Free Radical Biology & Medicine, Vol. 9, pp. 245-265, 1990
0891-5849/90 $3.00 + .00 Copyright© 1990PergamonPress plc
Printedin the USA. All rightsreserved.
Review Article BIOCHEMICAL BASIS OF OZONE TOXICITY MOHAMAD G. MUSTAFA Department of Environmental Health Sciences, School of Public Health, University of California, Los Angeles, CA 90024, U.S.A. (Received 18 December 1989; Revised and Accepted 18 May 1990)
Abstract--Ozone (03) is the major oxidant of photochemical smog. Its biological effect is attributed to its ability to cause oxidation or peroxidationof biomoleculesdirectly and/or via free radical reactions. A sequence of events may include lipid peroxidationand loss of functional groups of enzymes, alteration of membrane permeability, and cell injury or death. An acute exposure to 03 causes lung injury involving the ciliated cell in the airways and the type 1 epithelial cell in the alveolar region. The effects are particularly localized at the junction of terminal bronchioles and alveolar ducts, as evident from a loss of cells and accumulation of inflammatory cells. In a typical short-term exposure the lung tissue response is biphasic: an initial injury-phase characterized by cell damage and loss of enzyme activities, followed by a repair-phase associated with increased metabolic activities, which coincide with a proliferation of metabolically active cells, for example, the alveolar type 2 cells and the bronchiolar Clam cells. A chronic exposure to 03 can cause or exacerbate lung diseases, including perhaps an increased lung tumor incidence in susceptible animal models. Ozone exposure also causes extrapulmonary effects involving the blood, spleen, central nervous system, and other organs. A combination of 03 and NO2, both of which occur in photochemicalsmog, can produce effects which may be additive or synergistic. A synergisticlung injury occurs possibly due to a formation of more powerful radicals and chemical intermediates. Dietary antioxidants, for example, vitamin E, vitamin C, and selenium, can offer a protection against 03 effects. Keywords--Antioxidant protection, Extrapulmonary effects, Lung injury, Lung tumor, Nitrogen dioxide' toxicity, Ozone toxicity, Photochemical oxidants, Synergistic effects, Free radicals
INTRODUCTION
incidence, are thought to result from exposures to this oxidant. The toxic effects are attributed to its ability to generate free radical reactions in the biological system. The purpose o f this presentation is to review our understanding o f the toxicity and health effects o f O 3, including the possible involvement o f free radicals in oxidant injury and antioxidant protection.
Ozone is the major oxidant of photochemical smog, and generally occurs in association with other oxidant components, namely, nitrogen dioxide (NO2), peroxyacyl nitrates (PAN), hydrogen peroxide, alkyl peroxides, nitrous and nitric acids, formaldehyde and formic acid, and traces of other compounds. 1-3 Collectively, these chemicals are referred to as oxidants because they have the ability to remove electrons from other molecules. Of these oxidants, toxicity of O3 has been studied in relatively greater details, because of its abundance and earlier recognition in photochemical smog and the magnitude o f its potential effects on humans, plants, and ecosystem. The presence o f 0 3 in the air poses a serious health concern. Many short-term health effects and chronic lung diseases, including, perhaps, an increased tumor
OCCURRENCE OF OZONE
Ozone occurs as a natural component o f the atmosphere. However, there is a distinction in its occurrence in the upper atmosphere (stratosphere) and the lower atmosphere (e.g., smog in lower troposphere) as well as the biologic consequences. Ozone in smog is a common cause of urban air pollution, posing a threat to human health and property. 1-3 Stratospheric 03 is known to protect us from the harmful cosmic radiation, 4-7 although it may be a problem in high-altitude flights because the air-intake for airplane cabins includes O3 .s-12
Mohammad G. Mustafa (b. 1940, Bangladesh; Ph.D., 1969 in Biochemistry), is Professor of Enviommental Health Sciences, University of California, Los Angeles (UCLA). He worked with the Pulmonary (Air Pollution) Research Group at the U.C. Davis until his move to UCLA in 1975. His current researches focus on pulmonary effects of oxidant air pollutants (ozone, NO2) and other areas of inhalation
A. Formation in stratosphere
Atmospheric 03 concentration is the highest in stratosphere, a region extending from about 8 km at the poles and 17 km at the equator to about 50 km above the
toxicology. 245
246
M.G. MUSTAFA
earth's surface. 13 Ozone is thought to be generated according to the Chapman cycle of photochemical dissociation of molecular oxygen ( 0 2) by ultraviolet radiation (reaction 1.1) and then a chemical combination of atomic oxygen (O) with O 2 (reaction 1.2). 14-17 Ozone thus formed does not accumulate endlessly. The process of 03 formation is in a dynamic equilibrium with its natural destruction, for example, due to photochemical dissociation by ultraviolet radiation (reaction 1.3a) and also by infrared radiation (reaction 1.3b), collision with atomic oxygen (reaction 1.4), reaction with hydroxyl (HO) radical (reaction 1.5), and various other chemical and physical processes. 14-17 Some of the stratospheric 0 3 evidently reaches the lower troposphere providing the global O 3 background of up to 0.04 ppm. 1.2,18,19 However, it is the 0 3 produced in ambient air by photochemical reactions of primary (or precursor) pollutants that is of air pollution concern.
O2
hv (k
O+ O
(1.1)
M
O2 + O > 03 (1.2) (M = a third-body molecule absorbing excess energy of reaction) hv (k O2 + O (1.3a) hv (h < 1140 nm) O3 > O2 + O (1.3b) 03 + O > 02 + 02 (1.4) O3 + HO > HO2 + 02 (1.5)
Stratospheric 0 3 layer acts as a shield for the earth, protecting its surface, that is, preventing adverse effects on human health (skin cancer and suppression of immune response) and on terrestrial and aquatic ecosystems, from all UV-C (k
0 3 + 2NO 2
NO3 + NO2 N205 + 03
M
--'> M
>
N205 + 02
(1.19)
N205
(1.20)
NO 2 + 03
(1.21)
In addition, the reactions of 03, NO2, N205, and NO 3 in the moist environment of the bronchiolo-alveolar junctions (or with lung tissue) give rise to other chemical species, including HNO2, HNO 3, N O 2 - , N O 3 - , and HO radical (reactions 1.22-1.30). In lung tissue, NO 2 and NO 3 can be reduced to NO 2- and NO 3 - , respectively, in the presence of an electron donor (D), for example, a reducing substance, which then becomes a
254
M.G. MUSTAFA
radical cation, D + (reactions 1.22 and 1.30). Isolated lung perfusion studies have shown that the product of NO 2 absorption (or reaction) in lung tissue is N O 2 - , and it is not oxidized to NO 3 - (reaction 1.25) unless the red blood cells (RBC) are also present. 212'213 Oxidation of NO 2- to NO 3- and conversion of ferrohemoglobin (Hb) or oxyhemoglobin (HbO 2) to ferrihemoglobin (HB ÷ or metHb) represent a rather complex reaction system, 2~4-217 but the overall process can be shown as reaction 1.25. However, NO 3 radical can directly oxidize a biomolecule (D) and become reduced (reaction 1.30) without the presence of RBC,
NOz + D NO2 + NO + HzO HNO2 4NO 2 + 4HbO2 + 4H + N205 + H20 NO3 + H20 NO2 + HO HNO3 NO 3 + D
--> --> --> -->
--> -->
--> --> -->
NO2- + D ÷ 2HNO2
(1.22) (1.23)
NO2- + H + + 4Hb ÷ + Oz + 2H20 2HNO3 HNO 3 + HO HNO 3 NO 3- + H + NO3- + D +
(1.24) (1.25)
4NO 3 -
(1.26) (1.27) (1.28) (1.29) (1.30)
Formation of intermediate chemical species discussed above may explain the synergistic effects of 0 3 plus NO t system relative to 0 3 or NO 2 alone. The toxicity caused by 0 3, NO z, and their radical and nonradical reaction products must be understood apart from one another. Once the toxic potential of, at least, the major components are known, the phenomenon of additive or synergistic effects can be better explained.
D. Long-term effects Most experimental studies involving animals and controlled human subjects provide data on O3 toxicity that are derived from relatively short-term exposures, and often using O a concentrations higher than those encountered in the ambient atmosphere. Nonetheless, they are essential for regulatory processes and formulation of measures to prevent both short-term and longterm health effects.
a. Chronic diseases. Relatively long-term or chronic effects of 0 3 are not well documented. Experimental studies involving long-term exposures are limited. Epidemiological studies suggest that chronic oxidant exposures affect the baseline respiratory functions ,48 although the studies linking increased incidence of chronic disease with oxidants of photochemical smog are not very convincing. Based on relatively short-term animal
exposure studies, it is predicted that an acceleration of the deterioration of lung function with time (e.g., age of individuals) might result from chronic oxidant exposures. There is a depletion of lung cells and loss of lung reserves, which may not be obvious clinically but might produce a cumulative effect over a period of some years. 139 Destructive lung diseases, for example, emphysema, and other chronic obstructive lung diseases, for example, chronic bronchitis, and development of airway hypersensitivity or asthma might be the ultimate results of chronic oxidant exposures. 1-3,12,31,41,48.218-221 A continued (or cumulative effect of) oxidative and free radical reactions might be the underlying cause of lung cell depletion and functional deterioration. 222'223
b. Tumorigenic effects. A carcinogen (or tumorigen) is capable of causing several cellular reactions or processes that are thought to be associated with carcinogenesis (or tumorigenesis). These include induction of electron transfer or free radical mechanism; 224-227 chromosomal alteration or mutagenesis; 228'229 hyperplasia, irritation, and inflammation; 229'23° and omithine decarboxylase (ODC) activity. 231'232 Ozone exposure causes hyperplasia, irritation, and inflammation at the sites of lung injury. The hyperplasia can be repeatedly induced by recurrent or intermittent exposures. 42"149'150'154'156 Inflammatory cells release free radicals, for example, 0 2- and HO, which may cause further tissue injury, including DNA breaks.43"~38 Ozone causes chromosomal alterations, s~-55'233-236 which may be due to an attack on chromosomes by either 0 3 or free radicals generated by 03. Recently, a stimulation of lung ODC activity has been observed with 0 3 exposure. 237'238 Ozone, therefore, may be a potential carcinogen or promoter of carcinogenic process. Several experimental studies have been conducted to explore the carcinogenic potential of 0 3. Kotin et al.239,240 showed that mice after a year-long exposure to ozonized gasoline had a greater incidence of lung adenomas compared with control mice breathing a "washed atmosphere." Their synthetic atmosphere was intended to be qualitatively similar to Los Angeles smog, except the 0 3 concentration which was at 4 ppm (7840 ~g/m3). Stokinger 24~ reported 0 3 as a lung tumor accelerator on the basis of an increased tumor incidence in mice exposed to 1 ppm (1960 ixg/m3) 03 daily for 15 months. Gardner 242 exposed mice to ambient smog of Los Angeles for 7-11 months, and observed a trend toward increased incidence of pulmonary adenomas compared to that in age-matched clean-air control mice. The oxidant concentration in the smog ranged between 0.04 and 0.25 ppm (78 and 490 i~g/m3). A likelihood that ambient smog possesses a tumor-promoting activity in mice was suggested in this study. Two other studies in which O3-induced lung tumor incidence was significant
Biochemical basis of ozone toxicity
255
were carded out by Werthamer et al.243 and Nettesheim et al.244 In the latter study, mice were exposed to artificial smog (ozonized gasoline) containing 1 ppm (1960 p,g/m2) O 3 for up to 18 months. The incidence of lung tumors, which were adenomas and adenocarcinomas, was significantly greater in the smog-exposed mice than in the clean-air controls. Recently, Hassett et al. 245 examined the carcinogenic potential of 0 3 using A/J strain mice. A group of 45 mice was exposed to 0.50 ppm (980 ~g/m 3) 0 3 intermittently for 6 months; another group of 45 mice breathed clean air for the same time period. The lung tumor incidence was significantly higher in the O3-exposed mice. The result with a lower level of 0 3 (0.31 ppm) was not statistically significant. In another study, Last et al. 246 exposed AJJ strain and Swiss-Webster (SW) mice to 0.8 ppm 0 3 intermittently for approximately 4 months, and observed a significant increase in lung adenomas in m/J strain but not in SW mice relative to their corresponding controls. Since SW mice are relatively resistant to spontaneous or induced tumorigenesis, a 4-month long exposure may not have been chronic enough. In this study, exposure to a lower level of 03 (0.4 ppm) was not effective for A/J strain mice. Again, the exposure period may not have been chronic enough to make a difference between spontaneous and induced tumorigenesis. Although the major finding of Hassett et al. 245 and of Last et al. 246 that O 3 increases tumorigenesis in a susceptible mouse strain is in agreement, there remains an important point of difference. Hassett et al. 245 observed a synergism in lung adenoma formation when O 3 exposure was associated with a recurrent low-dose administration of urethane. Last et al. 246 observed an inhibition of lung adenoma formation when 0 3 exposure followed a onetime large-dose administration of urethane. This apparent contradiction may have to be resolved in a study specifically designed to address the issue of whether 0 3 is a cotumorigen/cocarcinogen. The subject of 0 3 tumorigenicity/carcinogenicity has recently been
defined. The effect of 03 on RBC and other hematologic parameters has been studied after both in vitro and in
r e v i e w e d . 48,247-249
Other extrapulmonary organs and tissues that have been examined for 0 3 effects include cardiovascular system, spleen, liver, endocrine system, and reproductive system. In animal studies, the heart rate and blood pressure were variably affected by O 3 exposure. 179.264 Spleen was found to be enlarged after a subchronic 0 3 exposure, 265 and this might be related to 0 3 effects on the blood. The O3 effects in the liver were variable. An evidence for alterations of zenobiotic metabolism, for example, an increase in pentobarbital-induced sleeping time, was reported, 266-268 but measurement of cytochrome P-450 concentration94"269 or benzopyrene hydroxylase activity89 showed no change. Ozone exposure was found to cause morphological changes in the parathyroid, 2°6 and a decrease in thyrotropin and thryoid hromones. 27°-273 Studies of O 3
EFFECTS IN OTHER ORGANS
Although the lung is the primary target for O 3 toxicity, the effects of 0 3 in other organs have also been documented. How 0 3 toxicity can reach the extrapulmonary tissues and organs is not clear, but it is possible that 0 3 reaction products cross the alveolar air-blood barrier, causing damage to various extrapulmonary sites. A. Hematopoietic system
Hematological parameters are frequently used to assess 0 3 toxicity, because blood chemistry is relatively simple and RBC morphology and metabolism are well
v i v o exposure. 52'54'69'71'159'160'171'250-257 T h e s e paranl-
eters include RBC morphology, survival, osmotic fragility, and oxyhemoglobin affinity; levels of GSH, ascorbate, hemoglobin, Heinz bodies, and methemoglobin; levels of serum lipids, cholesterol, albumin, and globulin; and RBC or plasma enzyme activities, namely, acetylcholine esterase, glucose-6-phosphate dehydrogenase, creatine phosphokinase, glutamate-pyruvate transaminase, pyruvate kinase, lactate dehydrogenase, and glutathione peroxidase. Ozone had either depressing or stimulating effects on these parameters, depending upon the 0 3 exposure conditions and nutritional status of the animals or controlled human subjects. For example, RBC morphology was altered, 253 survival was decreased, 252 and osmotic fragility increasedSZ'17a'251'254; GSH level was decreased251 or not changed159"255; serum cholesterol was increased256'257; acetylcholine esterase activity was decreased, 166 and the activities of glutathione peroxidase, pyruvate kinase, and lactate dehydrogenase were increased. 160
B. Central nervous system
Although CNS is adversely affected by O 3 exposure, and various physical and behavioral effects, for example, dizziness and visual impairment, have been noted, their biochemical basis is not clear. In animal studies, motor activity was found to be decreased by 0 3 exposure. 258 A change in behavior associated with a decreased running activity was reported by several laboratories. 259-262 A variability in enzyme activity, for example, a decrease in catechol methyl transferase and increase in monoamine oxidase, was also noted. 263
C. Other organs and tissues
256
M.G. MUSTAFA
effects on the reproductive system are limited. Some of the important effects noted were a decreased maternal weight gain and increased fetal resorption, 274 an increased neonatal mortality, 52 and slower development of offspring. 275 In addition, chromosomal alterations were observed in vitro in peripheral blood lymphocytes 54'234'235"276'277 and other mammalian cells. 51'236'278 The molecular mechanism for 0 3 effects on chromosomes is not clear, but single-strand DNA breaks may contribute to this effect. Zelac et al. 54 reported DNA strand breaks in lymphocytes of Chinese hamsters after in vivo 03 exposure, but others failed to observe such an in vivo effect in lymphocytes of Chinese hamsters 277 or humans. 235
ANTIOXIDANT PROTECTION
Since oxidative, peroxidative, and free radical reactions are important mechanisms of 0 3 toxicity, investigations have been carried out to discern antioxidant defense against oxidant damage. Living cells contain metabolic mechanisms (a variety of enzyme activities and metabolites) 42'8°'279-281 to combat oxidant stress, and these may be referred to as the intrinsic metabolic defense. Although the metabolic defense process prevents the formation of and/or eliminates most of the free radicals and peroxides, some free radicals (e.g., HO and HO 2) may not be destroyed by this means. Supplementation of ceils or organisms with antioxidant substances has been attempted as a means to contain or prevent free radical damage. External antioxidants often act in conjunction with metabolic defense and provide a strong antioxidant protection. 279-281
A. Metabolic defense Various antioxidant enzyme activities and metabolites (generated in situ by metabolic activites) contribute to metabolic defense mechanisms. 42'8°'279-281 One such mechanism, referred to as the GSH redox cycle, is thought to play a pivotal role in antioxidant defense against lipid peroxides and other toxic and related oxygenated intermediates. 42'44'45'279-281 In this cycle, GSH is oxidized to GSSG while reducing a peroxide in a GP-catalyzed reaction (1.31). GSH is then regenerated through a GR-catalyzed reaction (1.32) using the reducing equivalents of NADPH furnished by glucose oxidation via the pentose phosphate-shunt pathway (reactions 1.33ab). Activities of various enzymes, namely, glutathione peroxidase and associated thiol-metabolizing enzymes (glutathione reductase, disulfide reductase, thiol-disulfide transhydrogenase), peroxidase, catalase, superoxide dismutase, and key enzymes of the pentoseshunt pathway (glucose-6-phosphate and 6-phosphogluconate dehydrogenases), have been studied as a part of
the metabolic defense mechanism in the lung. 74'75'82'149' 150,151,154,157,158,162-165,167,168,279-282 Typical antioxidant metabolites studied are GSH and NADPH. 38"42' 84,279-281 In addition to antioxidant protection, metabolic defense mechanisms can promote the repair of injured cells and tissues, and enable the lung to withstand continuing injury. 42'84 2GSH +
GP
-> GSSG + ROH + H20
(1.31)
-> 2GSH + NADP ÷
(1.32)
ROOH
GSSG + NADPH + H+ NADP ÷ + G6P 6PG + NADP ÷
GR
G6PD 6PGD
-> NADPH + H ÷ 6PG
(1.33a)
•> R5P + NADPH + H
(1.33b)
(where, ROOH = a hydroperoxide; GP = glutathione peroxidase; GR = glutathione reductase; G6P --glucose-6-phosphate; 6PG = 6-phosphogluconate; G6PD and 6PGD = dehydrogenases pertaining to G6P and 6PG, respectively; and R5P = ribulose-5-phosphate). The enzyme activities and metabolites associated with metabolic defense are increased with O 3 exposure. 39'42'93'149'150 Whether this elevation is a protective mechanism against oxidant stress (i.e., the enzymes are synthesized de novo or metabolic pathways are activated for the purpose of defense) is not certain. It appears more likely that this is a response of lung tissue to oxidant damage in which the metabolically active cells (e.g., the alveolar type 2 and the bronchiolar Clara cells) through proliferation contribute to metabolic increases. This is corroborated by the fact that the time-course of metabolic increases coincides with that of morphological changes. 119'12° However, the issue is not entirely resolved since protection against oxidants is afforded by the development of metabolic defense mechanisms. Increased activities of antioxidant enzymes evidently prevent further damage against continued exposure as well as high-level e x p o s u r e . 74'75'149'154"156'159
B. Antioxidant supplements Various natural and synthetic antioxidants have been used as supplements (dietary or otherwise) to prevent and/or decrease O a damage. These are vitamin E (tocopherols), selenium (SE), vitamin C (ascorbate), vitamin A and beta-carotene, p-aminobenzoic acid, phenolic antioxidants (butylated hydroxytoluene, butylated hydroxyanisole), gluatathione and other sulfhydryl compounds. 42,74.75,77,78,84.149.150,158,160--164,169-171,255,279--297 These substances, usually administered as nutritional
Biochemical basis of ozone toxicity
supplements or as therapeutic agents, are known to act as free radical scavengers and/or as antioxidants against nonradical o x i d a t i o n . 3s'42'43'45,46'279-2sl'283'284'298-3°3 It is plausible that these compounds, particularly tocopherols (ArOH), donate a hydrogen to a free radical and stop or contain the chain reaction (reaction 1.34ab). 2s1'3°3'3°4 In that process they may become radicals (ArO) themselves, th,tt is, the antioxidants are oxidized, and then are regenerated or lost from the system. 37'28°'2s1'283 For tocopherols, an in vivo regeneration pathway has been proposed by Tappel, 283 and experimentally supported by others. 281'3°5-31°
HO + ArOH RO2 + ArOH
> >
HOH + ArO R O 2 H+ ArO
(1.34a) (1.34b)
Most typically, vitamin E is administered to experimental animals through dietary regimen. The effects of 0 3 can be elegantly demonstrated in vitamin E-deficient animals when compared with vitamin E-supplemented animals. Various degrees of protection, for example, prevention of massive lung injury or death due to high-level exposures and decrement of biochemical and morphological signs of injury due to low-level exposures, have been o b s e r v e d . 74'75'161'162A69-171'283-288 However, the protective role of vitamin E against oxidant (03) effects is not clearly demonstrated in humans, thus making any extrapolation from animals to man imprecise. There may be several explanations for this discrepancy. Experimental animals can be strictly controlled with dietary regimens, leading to vitamin E deficiency. Humans are not so restricted with diet and made deficient in vitamin E. Thus, a supplemented vitamin E dose in humans may not make any obvious difference in the protection already afforded by the basal dose. A lack of dietary vitamin E does not produce any obvious or acute signs of disease, probably because of the presence of other mechanisms of protection against lipid peroxidation and/or oxidant damage. Furthermore, the physiological and/or biochemical parameters examined in humans may not be sensitive enough to pick up the differences elicited by the basal (or average) dose and the supplemented dose. Also, the absence of any difference may not reflect a lack of vitamin E protective effects. In humans, vitamin E has found a clinical or pharmacologic usage against a variety of diseases, including aging, all of which may have a relationship with free radical r e a c t i o n s . 222'312-315 Vitamin E is thought to have no known human toxicity, although some clinical disorders are claimed to be associated with high doses. 311 It is not unreasonable to assume that a vitamin E supplementation has potential benefits against oxidant air
257
pollution (03), but the question remains on etablishing a judicial dose that can provide an optimal protection. Based on the premise that a vitamin E deficiency may be widespread in humans with symptoms of chronic lung disease, heart disease, or premature aging, and that an exposure to oxidant air pollution ( 0 3) may initiate and/or exacerbate these symptoms, a dietary supplementation of vitamin E may be a prudent means of prophylaxis against chronic free radical damage. In recent years, there is a concern that population exposed to photochemical smog or other environmental oxidants may require increased levels of dietary antioxidants, for example, vitamin E. Human requirements for vitamin E have been reviewed by Horwitt3~6 and Bled and Evarts. 317 The average human diet may contain 7 to 9 IU vitamin E per kg of food. The daily dosage received by the general population may appear marginal, considering the daily recommended dietary allowance of this vitamin which ranges form 4 to 15 IU for infants, children, and adults. 31s Therefore, some investigators have recommended large doses of vitamin E (as much as 1 g or roughly equivalent to 1000 IU daily) and other antioxidants for protection against oxidant effects and other diseases. 31°-312 However, the protection offeted by vitamin E or any of the other antioxidants is not absolute. Even with a high degree of supplementation lung injury occurs due to oxidant exposure. 42'2ss Similarly, Se, which is an integral part of glutathione peroxidase, has been found to influence 0 3 effects. Animals deficient in dietary Se were more susceptible to oxidants than those supplemented with this nutritional element, 163,164,288 and this susceptibility appeared to be due to a decreased function of glutathione peroxidase. 163 Se as selenocysteine and selenomethionine acts as potent water-soluble free radical scavengers. 37 The antioxidant role of Se is also evident from its ability to prevent the symptoms of vitamin E deficiency, such as caused by an oxidant (O 3) s t r e s s . 37'164"269 This is referred to as a sparing action between Se and vitamin E. However, Se and vitamin E do not fully replace each other in their antioxidant roles. 37 Also, Se is toxic in excess doses, and therefore cannot be widely used for protection against oxidant air pollution. 299 Vitamin C plays an antioxidant role against oxidant toxicity. 279-2sl'287'309 It is present in the normal lung, 29°'295 and its level is found to decrease with O 3 exposure. 167'296'297 Supplementation of animals with vitamin C offers protection against acute 03 injury. 29x293 Although vitamin C has the ability to break free radical chain reactions, 2sL3°s'3°9'314 it is not considered a major antioxidant because it can also act as a prooxidant. 2sL319-32° Ascorbate can reduce ferric iron while itself becoming an ascorbyl radical which would form superoxide and OH radicals in an aerobic environment. 283.321-324
258
M.G. MUSTAFA
Glutathione and other sulfhydryl compounds capable of furnishing -SH or -SS- groups have been found to offer a variable degree of protection against oxidant toxicity. 87,28t The exact mode of action is not known but thiyl radicals are thought to occur which may act as free radical scavengers.
2.
3. 4.
CONCLUSION
Ozone produces toxic effects in the lung through oxidation and peroxidation of biomolecules directly and/or by free radical chain reaction. Antioxidants, for example, vitamin E, possibly limit cellular injury by breaking the chain reactions, whereas cellular enzymatic mechanisms, for example, the GSH redox cycle, serve to eliminate the toxic reaction products. Antioxidant nutrition may have the virtue of retarding, if not entirely preventing, oxidant damage. There is abundant documentation of short-term 0 3 effects in the lung, which are characterized by an initial injury, and then a rebounding of metabolic activities, including the level of antioxidant metabolites, for example, GSH and NADPH. The long-term consequence of 0 3 exposure is not clear, and is surmised mostly based on short-term effects. As accelerated deterioration of lung structure and function with chronic exposure appears plausible. The incidence of chronic lung diseases may have multiple causes, but chronic 0 3 exposure remains a contributing factor. Inasmuch as free radical reactions are the underlying cause of 0 3 effects, a variety of diseases (viz., premature aging, chronic lung diseases, including lung tumor incidence) may be linked with chronic 0 3 exposures. Specific experimental findings, for example, chronic exposure causing lung tumor induction or enhancement, might shed light on the long-term risks of 0 3 exposures. There may be a good deal of similarity in the mode of biological reactions of 0 3 and NO2, but the synergistic effects produced by O3-NO 2 interaction is of concern because their combined exposure is the reality in a typical photochemical smog. The toxic chemical intermediates, for example, NO 3 and N20 5, along with the parent gases might expand or deepen cellular injury, resulting in the synergism. Further studies are needed to explain its molecular basis.
5.
6. 7.
8. 9.
10. 11. 12. 13.
14. 15. 16.
17. 18.
19.
20. 21. 22. 23. 24. 25.
Acknowledgements -- The support of U.S. EPA (R-812869) is gratefully appreciated. The author wishes to thank Mr. A.H. Saaduddin for technical assistance, and Mrs. Mary Hunter and Evalon Witt for preparation of the manuscript in a word processor.
26. 27.
REFERENCES 28. 1. National Research Council. Ozone and other photochemical oxidants. Vol. 19. Committee on Medical and Biologic Effects of Environmental Pollutants. Subcommittee on Ozone and Other
29.
Photochemical Oxidants. Washington, DC: National Academy of Sciences; 1977. U.S. Environmental Protection Agency. Air quality criteria for ozone and other photochemical oxidants. EPA Report No. EPA-600/8.79-O04. Research Triangle Park, NC: Environmental Criteria and Assessment Office, U.S. EPA; 1978. World Health Organization. Photochemical oxidants: Environmental health criteria 7. Geneva: WHO; 1979. National Academy of Sciences. Causes and effects of changes in stratospheric ozone: update 1983. Washington, DC: National Academy Press; 1984. U.S. Environmental Protection Agency. Ultraviolet radiation and melanoma with a special focus on assessing the risks of stratospheric ozone depletion. Washington, DC: Office of Air and Radiation, U.S. EPA; 1987. Jones, R.S. Ozone depletion and cancer risk. Lancet 2(No. 8556): 443-445; 1987. Redman, J.C. Stratospheric ozone depletion -- a proposed solution to the problem. Am. J. Dermatopathol. 9:457--458; 1987. Bennett, G. Ozone contamination of high altitude aircraft cabins. Aerosp. Med. 33:969-973; 1962. Jaffe, L.; Estes, H. Ozone toxicity in cabins of high altitude aircraft -- a review and current program. Aerosp. Med. 34: 633-643; 1963. Bischof, W. Ozone measurements on jet airline cabin air. Water Air Soil Pollut. 2:3-14; 1973. Reed, D.; Glaser, S.; Kaldor, J. Ozone toxicity symptoms among flight attendants. Am. J. lndust. Med. i:43-54; 1980. Stokinger, H.E. Ozone toxicology. Arch. Environ. Health 10: 719-731; 1965. Watson, R.T. Atmospheric ozone. In: Titus, J.G., ed. Effects of changes in stratospheric ozone and global climate. Washington, DC: U.S. Environmental Protection Agency; 1986:69-82. National Academy of Sciences. Environmental impact ofstratospheric flight. Washington, DC: National Academy Press; 1975. Heicklen, J. Atmospheric chemistry. New York: Academic Press; 1976. Perner, D. The consequences of increasing CFM concentrations for chemical reactions in the stratosphere and their impact on climate. In: Bach, W.; Pankrath, J.; Kellogg, W., eds. Man's impact on climate. Amsterdam: Elsevier; 1978:215-228. Kondrat'ev, K.Y. Changes in global climate (Russian Translations Series 26). Rotterdam: A.A. Balkema; 1986. Singh, H.B.; Viezee, W.; Johnson, W.B.; Ludwig, F.L. The impact of stratospheric ozone on tropospheric air quality. J. Air Pollut. Control Assoc. 311:1009-1017; 1980. Altshuller, A.P. Estimation of the natural background of ozone present at surface rural locations. J. Air Pollut. Control Assoc. 37:1409-1417; 1987. Haagen-Smit, A.J. Chemistry and physiology of Los Angeles smog. Ind. Eng. Chem. 44:1342-1346; 1952. Haagen-Smit, A.J. Photochemistry and smog. J. Air Pollut. Control Assoc. 13:446-454; 1963. Demerjian, K.L.; Kerr, J.A.; Calvert, J.G. The mechanism of photochemical smog formation. Adv. Environ. Sci. Technol. 4:1-262; 1974. Levy, H.H. Photochemistry of the troposphere. Adv. Photochem. 9:369-524; 1974. Finlayson, B.J.; Pitts, Jr., J.N. Photochemistry of the polluted troposphere. Science 192:111-119; 1976. Logan, J.A.; Prather, M.J.; Wofsy, S.C.; McElroy, M.B. Tropospheric chemistry: a global perspective. J. Geophys. Res. 86:7210-7254; 1981. Whitten, G.Z. The chemistry of smog formation: a review of current knowledge. Environ. Int. 9:447-463; 1983. Atkinson, R.; Lloyd, A.C. Evaluation of kinetic and mechanistic data for modeling of photochemical smog. J. Phys. Chem. Ref. Data. 13:315--444; 1984. Atkinson, R. Kinetic and mechanisms of the gas-phase reactions of the hydroxyl radical with organic compounds under atmospheric conditions. Chem. Rev. 86:69-201; 1986. Atkinson, R.; Carter, W.P.L. Kinetics and mechanisms of the
Biochemical basis of ozone toxicity
30. 31.
32. 33. 34. 35. 36. 37. 38.
39. 40.
41. 42. 43.
44.
45.
46. 47. 48. 49. 50.
51. 52.
gas-phase reactions of ozone with organic compounds under atmospheric conditions. Chem. Rev. 84:437-470; 1984. Nieto, LA.; Salvi, R.P.C.; Gutierrez C. A. Ozone hazard incurred in gamma-plant operation. Health Phys. 42:861-865; 1982. Borek, C.; Mehiman, M.A. Evaluation of health effects toxicity and biochemical mechanisms of ozone. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Publishers; 1983:325-361. Edwards, Jr., E.K. Ozone production by dermatologic phototherapy equipment. Curtis 28:433-434; 1981. South Coast Air Quality Management District. Seasonal and diurnal variation in air quality in California's South Coast Air Basin. E1 Monte, CA: SCAQMD; 1980. South Coast Air Quality Management District. 1982 summary of air quality in California's South Coast Air Basin. E1 Monte, CA: SCAQMD; 1983. Weast, R.C., ed. Handbook of chemistry and physics, 51st ed. Cleveland, OH: The Chemical Rubber Co.; 1970:D-67, D-68, D-112. Nasr, A.N.M. Biochemical aspects of ozone intoxication: a review. J. Occup. Med. 9:589-597; 1967. Menzel, D.B. Toxicity of ozone, oxygen and radiation. Annu. Rev. Pharmacol. 10:379-394; 1970. Menzel, D.B. The role of free radicals in the toxicity of air pollutants (nitrogen oxides and ozone). In: Pryor, W.A., ed. Free radicals in biology. Vol. II. New York: Academic Press; 1976:181-202. Cross, C.E.; Delucia, A.J.; Reddy, A.K.; Hussain, M.Z.; Chow, C.K.; Mustafa, M.G. Ozone interaction with lung tissue: biochemical approaches. Am. J. Med. 60"929-935; 1976. Mudd, J.B.; Freeman, B.A. Reaction of ozone with biological membranes. In: Lee, S.D, ed. Biochemical effects of environmental pollutants. Ann Arbor, MI: Ann Arbor Science Publishers; 1977:97-133. Coffin, D.L.; Stokinger, H.E. Biologic effects of air pollutants. In: Stern, A.C., ed Air pollution. Vol. II. New York: Academic Press; 1977:231-360. Mustafa, M.G.; Tierney, D. F. Biochemical and metabolic changes in the lung with oxygen, ozone, and nitrogen dioxide. Am. Rev. Respir. Dis. 118:1061-1090; 1978. Goldstein, B.D. The pulmonary and extrapulmonary effects of ozone. In: CIBA Foundation symposium: oxygen free radicals and tissue damage. Vol. 65. New York: Elsevier Press; 1979: 295-319. Pryor, W.A.; Dooley, M.M.; Church, D.F. Mechanisms for the reaction of ozone with biological molecules, the source of the toxic effects of ozone. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:7-19. Chow, C.K. Influence of dietary vitamin E on susceptibility to ozone exposure. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:75-93. Menzel, D.B. Ozone: an overview of its toxicity in man and animals. J. Toxicol. Environ. Health 13:183-204; 1984. Mehlman, M.A.; Borek, C. Toxicity and biochemical mechanisms of ozone. Environ. Res. 42:36-53; 1987. Lippmann, M. Health effects of ozone--a critical review. J. Air Waste Manage. Assoc. (JAPCA) 39:672-695; 1989. Brinkman, R.; Lamberts, H.B. Ozone as a possible radiomimetic gas. Nature (London) 181:1202-1203; 1958. Scheel, L.D.; Dobrogorski, D.J.; Mountain, LT.; Svirbely, J.L.; Stokinger, H.E. Physiologic, biochemical, immunologic and pathologic changes following ozone exposure. J. Appl. Physiol. 14:67-80; 1959. Fetner, R.H. Ozone induced chromosome breakage in human cell culture. Nature (London) 194:793-794; 1962. Brinkman, R.; Lamberts, H.B.; Veninga, T.S. Radiomimetic toxicity of ozonized air. Lancet 1(No. 7325):133-136; 1964.
259
53. Veninga, T.S. Toxicity of ozone in comparison with ionizing radiation. Strahlentherapi. 134:469-477; 1967. 54. Zelac, R.E.; Croruroy, H.L.; Bolch, Jr., W.E.; Dunavant, B.G.; Bevis, H.A. Inhaled ozone as a mutagen. I. Chromosome aberrations induced in Chinese hamster lymphocytes. Environ. Res. 4:262-282, 1971. 55. Zelac, R.E., Cromroy, H.L.; Bolch, Jr., W.E.; Dunavant, B.G., Bevis, H.A. Inhaled ozone as a mutagen. H. Effect on the frequency of chromosome aberrations observed in irradiated Chinese hamsters. Environ. Res. 4:325-342; 1971. 56. Koppenol, W.H. The reduction potential of the couple O3/03-, consequences for mechanisms of ozone toxicity. FEBS Lett. 140:169-172; 1982. 57. Pryor, W.A.; Ohto, N.; Church, D.F. Reaction of cumene with ozone to form cumyl hydrotr/oxide and the kinetics of decomposition of cumyl hydrotrioxide, l. Am. Chem. Soc. 105:36143622; 1983. 58. Joigne, J.; Bader, H. The role of hydroxyl radical reactions in ozonation process in aqueous solutions. Water Res. 10:377-386; 1979. 59. Carmichael, N.G.; Winder, C.; Borges, S.H.; Backhouse, B.L.; Lewis, P.D. The health implications of water treatment with ozone. Life Sci. 30:117-129; 1982. 60. Glaze, W.H. Reaction products of ozone: a review. Environ. Health Perspect. 69:151-157; 1986. 61. Goldstein, B.D.; Balchum, O.J.; Demopoulos, H.D.; Duke, P.S. Electron paramagnetic resonance spectroscopy. Free radical signals associated with ozonation of linoleic acid. Arch. Environ. Health 17:46-49; 1968. 62. Pyror, W.A.; Prier, D.G.; Church, D.F. Radical production from the interaction of ozone and PUFA as demonstrated by electron spin resonance spin-trapping techniques. Environ. Res. 24: 42-52, 1981. 63. Pyror, W.A. The role of free radical reactions in biological systems. In: Pryor, W.A., ed. Free radicals in biology. Vol. 1. New York: Academic Press; 1976:1-49. 64. Kurz, M.E.; Pryor, W.A. Radical production from the interaction of closed-shell molecules. 9. Reactions of ozone with tert-butyl hydroperoxide. J. Am. Chem. Soc. 100:7953-7959; 1978. 65. Pryor, W.A. Mechanisms and detection of pathology caused by free radicals. Tobacco smoke, nitrogen dioxide and ozone. In: McKinney, J.D., ed. Environmental health chemistry. Ann Arbor, MI: Ann Arbon Science Publishers; 1981:445-466. 66. Cortesi, R.; Privett, O.S. Toxicity of fatty ozonides and peroxides. Lipids 7:715-721; 1972. 67. Pryor, W.A. The toxicity of oxidized fats: lipid hydroperoxides, their decomposition products and their cooxidation products. In: Taylor, T.G.; Jenkins, N.K., eds. Proceedings of the XIII international congress of nutrition. London: John Libbey and Co.; 1986:589-593. 68. Wu, G.-S.; Milion, T.V.; Subramanian, C.S.; Mnstafa, M.G.; Afifi, A.A.; the late Mead, J.F. Effect of ozone and constituents of rat lung tissue and lavage fluid. Submitted for publication (1990). 69. Goldstein, B.D.; Balchum, O.J. Effect of ozone on lipid peroxidation in the red blood cell. Proc. Soc. Biol. Med. 126:356-358; 1967. 70. Balchum, O.J.; O'Brien, J.S.; Goldstein, B.D. Ozone and unsaturated fatty acids. Arch. Environ. Health 22: 32-34; 1971. 71. Goldstein, B.D. Hydrogen peroxide in erythrocytes. Detection in rats and mice inhaling ozone. Arch. Environ. Health 26:279280; 1973. 72. Goldstein, B.D.; Lodi, C.; Collinson, C.; Balchum, O.J. Ozone and lipid peroxidation. Arch. Environ. Heath 18:631-635; 1969. 73. Sevanian, A.; Mead, J.F.; Stein, R.A. Epoxide products of lipid peroxidation in rat lungs. Lipids 14:634--643; 1979. 74. Chow, C.K.; Tappel, A.L. An enzymatic protective mechanism against lipid peroxidatiun damage to lungs of ozone-exposed rats. Lipids 7: 518-524; 1972. 75. Chow, C.K.; Tappel, A.L. Activities of pentose shunt and glycolytic enzymes in lungs of ozone-exposed rats. Arch. Environ. Health 26:205-208; 1973.
260
M.G. MUSTAFA
76. Sagai, M.; Arakawa, K.; Ichinose, T.; Shimojo, N. Biochemical effects of combined gases of nitrogen dioxide and ozone. I. Species differences of lipid peroxides and phospholipids in lungs. Toxicol. 46:251-265; 1987. 77. Dumelin, E.E.; Dillard, C.J.; Tappel, A.L. Effect of vitamin E and ozone on pentane and ethane expired by rats. Arch. Environ. Health 33:129-135; 1978. 78. Dumelin, E.E.; Dillard, C.J.; Tappel, A.L. Breath ethane and pentane as measures of vitamin E protection of Macaca radiata against 90 days of exposure to ozone. Environ. Res. 15:38-43; 1978. 79. Gelmont, D.; Stein, R.A.; Mead, J.F. The bacterial origin of rat breath pentane. Biochem. Biophys. Res. Commun. 102:932-936; 1981. 80. Mustafa, J.G. General enzymology of the lung. In: Witschi, H.P.; Brain, J.D., eds. Toxicology of inhaled materials. Berlin: Springer-Verlag; 1985;369-419. 81. Menzel, D.B. Oxidation of biologically active reducing substances by ozone. Arch. Environ. Heath 23:149-153; 1971. 82. DeLucia, A.J.; Hoque, P.M.; Mustafa, M.G.; Cross, C.E. Ozone interaction with rodent lung. Effect on sulfhydryl and sulfhydryl-containing enzyme activities. J. Lab. Clin. Med. 80"559-566; 1972. 83. DeLucia, A.J.; Mustafa, M.G.; Hussaln, M.Z.; Cross, C.E. Ozone interaction with rodent lung. III. Oxidation of reduced glutathione and mixed disulfide formation between protein and nonprotein sulfhydryls. J. Clin. Invest. 55:794-802; 1975. 84. Fairchild, II, E.J.; Murphy, S.D.; Stokinger, H.E. Protection by sulfur compounds against the air pollutants ozone and nitrogen dioxide. Science 130:861-862; 1959. 85. Mountain, J.T. Detecting hypersensitivity to toxic substances. Arch. Environ. Health 6:357-365; 1963. 86. Freeman, B.A.; Mudd, J.B. Reaction of ozone with sulfhydryls of human erythrocytes. Arch. Biochem. Biophys. 208:212-220; 1981. 87. Mudd, J.B.; Leavitt, R.; Ongun, A.; McManus, T.T. Reaction of ozone with amino acids and proteins. Atmos. Environ. 3:669-681; 1969. 88. Hurst, D.J.; Coffin, D.L. Ozone effect on lysosomal hydrolases of alveolar macrophages in vitro. Arch. Intern. Med. 127" 1059-1063; 1971. 89. Palmer, M.S.; Swanson, D.H.; Coffin, D. L. Effect of ozone on benzpyrene hydroxylase activity in the Syrian golden hamster. Cancer Res. 31:730-733; 1971. 90. Palmer, M.S.; Exley, R.W.; Coffin, D.L. Influence of pollutant gases on benzpyrene hydroxylase activity. Arch. Environ. Health 25:439--442; 1972. 91. Mustafa, M.G.; DeLucia, A.J.; York, G.K.; Arth, C.; Cross, C.E. Ozone interaction with rodent lung. II. Effects on oxygen consumption of mitochondria. J. Lab. Clin. Med. 82:357-365; 1973. 92. Mustafa, M.G.; Cross, C.E. Effects of short-term ozone exposure on lung mitochondrial oxidative and energy metabolism. Arch. Biochem. Biophys. 162:585-594; 1974. 93. Mustafa, M.G. Augmentation of mitochondrial oxidative metabolism in lung tissue during recovery from acute ozone exposure. Arch. Biochem. Biophys. 165:531-538; 1974. 94. Goldstein, B.D.; Solomon, S.; Pasternack, B.S.; Bickers, D.R. Decrease in rabbit lung microsomal cytochrome P-450 levels following ozone exposure. Res. Commun. Chem. Pathol. Pharmacol. 10:759-762; 1975. 95. Montgomery, M.R.; Niewoehner, D.E. Oxidant-induced alterations in pulmonary microsomal mixed-function oxidation: acute effects of paraquat and ozone. J. Environ. Sci. Health C13: 205-219; 1979. 96. Menzel, D.B.; Anderson, W.G.; Abou-Donia, M.B. Ozone exposure modifies prostaglandin biosynthesis in perfused rat lungs. Res. Commun. Chem. Pathol, Pharmacol. 15:135-147; 1976. 97. Madden, M.C.; Eling, T.E.; Friedman, M. Ozone inhibits endothelial cell cyclooxigenase activity through formation of hydrogen peroxide. Prostagland. 34:445-463; 1987. 98. Gordon, T.; Taylor, B.F.; Amdur, M.O. Ozone inhibition of
tissue cholinesterase in guinea-pigs. Arch. Environ. Health 36:284-288; 1981. 99. Johnson, D.A. Ozone inactivation of human alpha-l-antiproo teinase inhibitor. Am. Rev. Respir. Dis. 121"1031-1038; 1980. 100. Koontz, A.E.; Heath, R.L. Ozone alteration of transport of cations and the Na/K ATPase in human erythrocytes. Arch. Biochem. Biophys. 198.493-500; 1979. 101. Witz, G.; Amoruso, M.A.; Goldstein, B.D. Effect of ozone on alveolar macrophage function: membrane dynamic properties. In: Lee S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Pub-
lishers; 1983:263-272. 102. Rietjens, I.M.C.M.; van Tilburg, C.A.M.; Coenen, T.M.M.; Alink, G.M.; Konings, A.W.T. Influence of polyunsaturated fatty acid supplementation and membrane fluidity on ozone and nitrogen dioxide sensitivity of rat alveolar macrophages. J. Toxicol. Environ. Health 21:45-56; 1987. 103. Alpert, S.M.; Schwartz, B.B.; Lee, S.D.; Lewis, T.R. Alveolar protein accumulation: a sensitive indicator of low level oxidant toxicity. Arch. Intern. Med. 128:69-73; 1971. 104. Reasor, M.J.; Adams, G.K.; Brooks, J.K.; Rubin, R.J. Enrichment of albumin and IgG in the airway secretions of dogs breathing ozone. J. Environ. Sci. Health C13:335-346; 1979. 105. Williams, S.J.; Charles, J.M.; Menzel, D.B. Ozone-induced alterations in phenol red absorption from the rat lung. Toxicol. Lett. 6:213-219; 1980. 106. Hu, P.C.; Miller, F.J.; Daniels, M.J.; Hatch, G.E.; Graham, J.A.; Gardner, D.E.; Selgrade, M.K. Protein accumulation in lung lavage fluid following ozone exposure. Environ. Res. 29:377-388; 1982. 107. Bhalla, D.K.; Crocker, T.T. Tracheal permeability in rats exposed to ozone. Am. Rev. Respir. Dis. 134:572-579; 1986. 108. Bhalla, D.K.; Mannix, R.C.; Kleinman, M.T.; Crocker, T.T. Relative permeability of nasal, tracheal, and bronchoalveolar mucosa to macromolecules in rats exposed to ozone. J. Toxicol. Environ. Health 17:269-283; 1986. 109. Kehrl, H.R.; Vincent, L.M.; Kowalsky, R.J.; Horstman,D.H.; O'Neil, J.J.; McCartney, W.H.; Bromberg, P.A. Ozone exposure increases respiratory epithelial permeability in humans. Am. Rev. Respir. Dis. 135:1124-1128; 1987. 110. Seltzer, J.; Bigby, B.G.; Stulbarg, M.; Holtzman, M.J.; Nadel, J.A.; Ueki, I.F.; Leikauf, G.D.; Goetzel, E.J.; Boushey, H.A. O3-induced change in bronchial reactivity to methacholine and airway inflammation in humans. J. Appl. Physiol. 60:13211326; 1986. 111. Koren, H.S.; Devlin, R.B.; Graham, D.E..; Mann, R.; McGee, M.P.; Horstman, D.H.; Kozumbo, W.J.; Becker, S.; House, D.E.; McDonnell, W.F.; Bromberg, P.A. Ozone-induced inflammation in the lower airways of human subjects. Am. Rev. Respir. Dis. 139:407-415; 1989. 112. Ford-Hutichinson, A.W.; Bray, M.A.; Doig, M.V.; Shipley, M.E.; Smith, M.J.H. Leukotriene b, a potent chemokinetic and aggregating substance released from polymorphonuclear leukocytes. Nature 286:264-265; 1980. 113. Dixon, J.R.; Mountain, J.T. Role of histamine and related substances in the development of tolerance to edemagenic gases. Toxicol Appl. Pharmacol. 7:756-766; 1965. 114. Fairchild, II, E.J. Neurohormonal factors in injury from inhaled irritants. Arch. Environ. Health 6:85-92; 1963. 115. Fairchild, II, E.J.; Graham, S.L. Thyroid influence on the toxicity of respiratory irritant gases, ozone and nitrogen dioxide. J. Pharmacol. Exp. Ther. 139:177-184; 1963. 116. Fairchild, II, E.J.; Graham, S.L.; Hite, M.; Killens, R.; Scheel, L.D. Changes in thyroid I TM activity in ozone-tolerant and ozone-susceptible rats. Toxicol Appl. Pharmacol. 6:607-613; 1964. 117. Yokoyama, E.; Frank, R. Respiratory uptake of ozone in dogs. Arch. Environ. Health 25:132-138; 1972. 118. Miller, F.J. Biomathematical modeling applications in the evaluation of ozone toxicity. In: Mudd, J.B.; Lee, S.D., eds. Assessing toxic effects of environmental pollutants. Ann Arbor, MI: Ann Arbor Science Publishers; 1979:263-286.
Biochemical basis of ozone toxicity 119. Dungworth, D.L.; Castleman, W.L.; Chow, C.K.; Me[lick, P.W.; Mustafa, M.G.; Tarkington, B.; Tyler, W.S. Effects of ambient levels of ozone on monkeys. Fed. Proc. 34:1670-1674; 1975. 120. Schwartz, W.L.; Dungworth, D.L.; Mustafa, M.G.; Tarkington, B.K.; Tyler, W.S. Pulmonary response of rats to ambient levels of ozones: effects of 7-day intermittent or continuous exposure. Lab. Invest. 34:565-578; 1976. 121. Cavender, F.L.; Steinhagen, W.H.; Ulrich, C.E.; Busey, W.M.; CockreU, B.Y.; Haseman, J.K.; Hogan, M.D.; Drew, R.T. Effects in rats and guinea-pigs of short-term exposures to sulfuric acid mist, ozone, and their combination. J. Toxicol. Environ. Health 3:521-533; 1977. 122. Castleman, W.L.; Tyler, W.S.; Dungworth, D.L. Lesions in respiratory bronchioles and conducting airways of monkeys exposed to ambient levels of ozone. Exp. Mol. Pathol. 26: 3~os. A.90; 1977. 123. Mellick, P.W.; Dungworth, D.L.; Schwartz, L.W.; Tyler, W.S. Short-term morphologic effects of high ambient levels of ozone on lungs of rhesus monkeys. Lab. Invest. 36:82-90; 1977. 124. Ibrahim, A.L.; Zee, Y.C.; Osebold, J.W. The effects of ozone on the respiratory epithelium of mice. II. Ultrastructural alterations. J. Environ. Pathol. Toxicol. 3:251-258; 1980. 125. Wilson, D.W.; Plopper, C.G.; Dungworth, D.L. The response of the macaque ffacheobronchial epithelium to acute ozone injury. A quantitative ultrastructural and autoradiographic study. Am. J. Pathol. 116:193-206; 1984. 126. Eustis, S.L.; Schwartz, L.W.; Kosch, P.C.; Dungworth, D.L. Chronic bronchiolitis in nonhuman primates after prolonged ozone exposure. Am. J. Pathol. 105:121-137; 1981. 127. Yokoyama, E.; Ichikawa, I.; Kawal, K. Does nitrogen dioxide modify the respiratory effects of ozone? In: Lee, S.D., ed. Nitrogen oxides and their effects on health. Ann Arbor, MI: Ann Arbor Science Publishers; 1980:217-229. 128. Stephens, R.J.; Sloan, M.F.; Evans, M.J.; Freeman, G. Early response of lung to low levels of ozone. Am. J. Pathol. 73:711-726; 1974. 129. Stephens, R.J.; Sloan, M.F.; Evans, M.J.; Freeman, G. Alveolar type 1 cell response to exposure to 0.5 ppm 03 for short periods. Exp. Mol. Pathol. 20:11-23; 1974. 130. Evans, M.J.; Johnson, L.V.; Stephens, R.J.; Freeman, G. Cell renewal in the lungs of rats exposed to low levels of ozone. Exp. Mol. Pathol. 24:70--83; 1976. 131. Lure, H.; Schwartz, L.W.; Dungworth, D.L.; Tyler, W.S. A comparative study of cell renewal after exposure to ozone or oxygen: response of terminal broncbiolar epithelium in the rat. Am. Rev. Respir. Dis. 118:355-345; 1978. 132. Boorman, G.A.; Schwartz, L. W.; Dungworth, D.L. Pulmonary effects of prolonged ozone insult in rats. Morphometric evaluation of the central acinus. Lab. Invest. 43:108-115; 1980. 133. Barry, B.E.; Miller, F.J.; Crapo, LD. Alveolar epithelial injury caused by inhalation of 0.25 ppm ozone. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:299-309. 134. Sherwin, R.P.; Richters, V.; Okimoto, D. Type 2 pneumocyte hyperplasia in the lungs of mice exposed to an ambient level (0.3 ppm) of ozone. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:289-297. 135. Castelman, W.L.; Dungworth, D.L.; Tyler, W.S. Cytochemically detected alterations of lung acid phosphatase reactivity following ozone exposure. Lab. Invest. 29:310-319; 1973. 136. Castelman, W.L.; Dungworth, D.L.; Tyler, W.S. Histochemically detected enzymatic alterations in rat lung exposed to ozone. Exp. Mol. Pathol. 19:402-421; 1973. 137. Brummer, M.E.G.; Schwartz, L.W.; McQuillen, N.K. A quan-. titative study of lung damage by scanning electron microscopy: inflammatory cell response to high-ambient levels of ozone. Scanning Electron Microsc. 2:513-518; 1977. 138. Brain, J.D. Macrophage damage in relation to the pathogenesis of lung diseases. Environ. Health Perspect. 35:21-28; 1980.
261
139. Oomichi, S.; Kits, H. Effect of air pollutants on ciliary activity of respiratory tract. Bull. Tokyo Med. Dent. Univ. 21:327-343; 1974. 140. Frager, N.B.; Phalen, R.F.; Kenoyer, J.L. Adaptations to ozone in reference to mucociliary clearance. Arch. Environ. Health 34:51-57; 1979. 141. Kenoyer, J.L.; Phalen, R.F.; Davis, J.R. Particle clearance from the respiratory tract as a test of toxicity: effect of ozone on short and long term clearance. Exp. Lung Res. 2:111-120; 1981. 142. Schlesinger, R.B.; Driscoll, K.E. Mucociliary clearance from the lungs of rabbits following single and intermittent exposure to ozone. J. Toxicol. Environ. Health 20:125-134; 1987. 143. Abraham, W.M.; Januskiewicz, A.J.; Mingle, M.; Wekler, M.; Wanner, A.; Sackner, M.A. Sensitivity of bronchoprovocatiun and tracheal mucous velocity in detecting airway responses to 03. J. Appl. Physiol. 48:789-793; 1980. 144. Phipps. R.J.; Denas, S.M.; Sielczak, M.W.; Wanner, A. Effects of 0.5 ppm ozone on glycoprotein secretion, ion and water fluxes in sheep trachea. J. Appl. Physiol. 60:918-927; 1986. 145. Foster, W.M.; Costa, D.L.; Langenback, E.G. Ozone exposure alters tracheobronchial mucociliary function in humans. J. Appl. Physiol. 63:996-1002; 1987. 146. Coffin, D.L.; Gardner, D.E. Interaction of biological agents and chemical air pollutants. Ann. Occup. Hyg. 15:219--234; 1972. 147. Miller, F.J.; Illing, J.W.; Gardner, D.E. Effects of urban ozone levels on lahoratory-induced respiratory infections. Toxicol. Len. 2:163-169; 1978. 148. Driscoll, K.E.; Vollmuth,T.A.; Schlocsinger, R.B. Early alveolar clearance of particles in rabbits undergoing acute and subchronic exposure to ozone. Fundam. Appl. Toxicol. 7: 264-271; 1986. 149. Mustafa, M.G.; Lee, S.D. Pulmonary biochemical alterations resulting from ozone exposure. Ann. Occup. Hyg. 19:17-26; 1976. 150. Mustafa, M.G.; Hacker, A.D.; Ospital, J.J.; Hnssain, M.Z.; Lee, S.D. Biochemical effects of environmental oxidant pollutants in animal lungs. In: Lee, S.D., ed. Biochemical effects of environmental pollutants. Ann Arbor, NII: Ann Arbor Science Publishers; 1977:59-96. 151. Mustafa, M.G.; Elsayed, N.M.; Quinn, C.L.; Postlethwalt, E.M.; Gardner, D.E.; Graham, LA. Comparison of pulmonary biochemical effects of low level ozone exposure on mice and rats. J. Toxicol. Environ. Health 9:857-865; 1982. 152. Elsayed, N.M.; Mustafa, M.G.; Postlethwalt, E.M. Age-dependent pulmonary response of rats to ozone exposure. J. Toxicol. Environ. Health 9:835-848; 1982. 153. Mustafa, M.G.; Elsayed, N.M.; Graham, J.A.; Gardner, D.E. Effects of ozone exposure on lung metabolism: influence of animal age, species, and exposure conditions. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A. eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983: 57-73. 154. Chow, C.K.; Hussaln, M.Z.; Cross, C.E.; Dungworth, D.L.; Mustafa, M.G. Effects of low levels of ozone on lung. I. Biochemical responses during recovery and reexposure. Exp. Mol. Pathol. 25:182-188; 1976. 155. Hussain, M.Z.; Cross, C.E.; Mustafa, M.G.; Bhatnagar, R.S. Hydroxyproline contents and prolyl hydroxylase activities in lungs of rats exposed to low levels of ozone. Life Sci. 15: 897-904; 1976. 156. Plopper, C.G.; Chow, C.K.; Dungworth, D.L.; Brummer, M.; Nemeth, T.J. Effect of low level of ozone on rat lungs. II. Morphological responses during recovery and reexposure. Exp. Mol. Pathol. 29:4(X)-411; 1978. 157. Plopper, C.G.; Chow, C.K.; Dungworth, D.L.; Tyler, W.S. Pulmonary alterations in rats exposed to 0.2 and 0.1 ppm ozone: a correlated morphological and biochemical study. Arch. Environ. Health 34:390-395; 1979. 158. Chow, C.K.; Dillard, C.J.; Tappel, A.L. Gluthione peroxidas¢ system and lysozyme in rats exposed to ozone or nitrogen dioxide. Environ. Res. 7:311-319; 1974. 159. Chow, C.K.; Mustafa, M.G.; Cross, C.E.; Tarkington, B.K.
262
160. 161.
162.
163.
164.
165.
166.
167. 168. 169. 170. 171. 172.
173. 174.
175.
176. 177. 178. 179.
M.G. MuSTAFA Effects of ozone exposure on the lungs and the erythrocytes of rats and monkeys: relative biochemical changes. Environ. Physiol. Biochem. 5:142-146; 1975. Chow, C.K.; Kaneko, J.J. Influence of dietary vitamin E on the red cells of ozone-exposed rats. Environ. Res. 19:49-55; 1979. Chow, C.K.; Plopper, C.G.; Dungworth, D.L. Influence of dietary vitamin E on the lungs of ozone-exposed rats: a correlated biochemical and morphological study. Environ. Res. 20:309317; 1979. Chow, C.K.; Plopper, C.G.; Chin, M.; Dungworth, D.L. Dietary vitamin E and pulmonary biochemical and morphological alterations of rats exposed to 0.1 ppm ozone. Environ. Res. 24:315-324; 1981. Elsayed, N.M.; Hacker, A.; Mustafa, M.; Kuehn, K.; Schrauzer, K. Effects of decreased glutathione peroxidase activity on the pentose phosphate cycle in mouse lung. Biochem, Biophys. Res. Commun. 104:564-569; 1982. Elsayed, N.M.; Hacker, A.D.; Kuehn, K.; Mustafa, M.G.; Schrauzer, G.N. Dietary antioxidants and the biochemical response to oxidant inhalation. II. Influence of dietary selenium on the biochemical effects of ozone exposure in mouse lung. Toxicol. Appl. Pharmacol. 71:398--406; 1983. Lunan, K.D.; Short, P.; Negi, D.; Stephens, R.J. Glucose6-phosphate dehydrogenase response of postnatal lungs to NO 2 and 03. In: Sanders, C.L.; Schneider, R.P.; Dagle, G.E.; Ragan, H.A., eds. Pulmonary macrophages and epithelial cells. Washington, DC: Energy Research and Development Administration (ERDA symposium series: 43): 1977;236-247. Moore, G.S.; Calabrese, E.J.; Grinberg-Funes, R.A. The C57L/J mouse strain as a model for extrapulmonary effects of ozone exposure. Bull. Environ. Contain. Toxicol. 25:578-585; 1980. Fukase, O.; Isomura, K.; Watanabe, H. Effect of ozone on glutathione in vivo. Taiki Osen Kenkyu 10:58-62; 1975. Tyson, C.A.; Lunan, K.D.; Stephens, R.J. Age-related differences in GSH-shuttle enzymes in NOz or O3 exposed rat lungs. Arch. Environ. Health 37:167-176; 1982. Roehm, J.N.; Hadley, J.G.; Menzel, D.B. Antioxidant vs. lung disease. Arch. Intern. Med. 128:88-93; 1971. Roehm, J.N.; Hadley, J.G.; Menzel, D.B. The influence of vitamin E on the lung fatty acids of rats exposed to ozone. Arch. Environ. Health 24:237-242; 1972. Menzel, D.B.; Roehm, J.N.; Lee, S.D. Vitamin E: the biological and environmental antioxidant. J. Agric. Food Chem. 20:481486; 1972. Shimasaki, H.T.; Takatori, W.R.; Anderson, H.L.; Shimasaki, H.; Takatori, T.; Anderson, W.R.; Horten, H.L.; Privett, O.S. Alteration of lung lipids in ozone exposed rats. Biochem. Biophys. Res. Commun. 68:1256-1262; 1976. Huber, G.L.; Mason, R.J.; LaForce, M.; Spencer, N.J.; Gardner, D.E.; Coffin, D.L. Alterations in the lung following the administration of ozone. Arch. Intern. Med. 128:81-87; 1971. Myers, A.B.; Dubick, M.A.; Reiser, K.M.; Gerriets, J.E.; Last, J.A.; Rucker, R.B. Ozone exposure, food restriction and protein deficiency: changes in collagen and elastin in rodent lung. Toxicol. Lett. 23: 43-49; 1984. Bhatnagar, R.S.; Hussain, M.Z.; Sorensen, K.R.; Mustafa, M.G.; yon Dohlen, F.M.; Lee, S.D. Effect of ozone on lung collagen biosynthesis. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:311-321. Last, J.A.; Kaizu, T. Mucous glycoprotein secretion by tracheal explants: effects of pollutants. Environ. Health Perspect. 35: 131-138; 1980. Last, J.A.; Greenberg, D.B.; Castleman, W.L. Ozone-induced alterations in collagen metabolism of rat lungs. Toxicol. Appl. Pharmacol. 51:247-258; 1979. Last, J.A.; Greenberg, D.B. Ozone-induced alterations in collagen metabolism of rat lungs. II. Long-term exposure. Toxicol. Appl. Pharmacol. 55:108-114; 1980. Costa, D.L.; Kutzman, R.S.; Lehrnann, J.R.; Popenoe, E.A.; Drew, R.T.A. Subchronic multi-dose ozone study in rats. In:
180. 181. 182. 183. 184.
185.
186. 187.
188. 189.
190.
191.
192. 193.
194.
195.
196, 197. 198.
Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:369-393. Last, J.A.; Reiser, K.M.; Tyler, W.S.; Rucker, R.B. Long-term consequences of exposure to ozone. I. Lung collagen content. Toxicol. Appl. Pharmacol. 72:111-118; 1984. Hacker, A.D.; Mustafa, M.G.; Ospital, J.J.; Elsayed, N.M.; Lee, S.D. Relationship of age to rat lung collagen synthesis with ozone exposure. Age 9:1-5; 1986. Hesterberg, T.W.; Last, J.A. Ozone-induced acute pulmonary fibrosis in rats: prevention of increased rates of collagen synthesis by methyl prednisolone. Am. Rev. Respir. Dis. 123:47-52; 1981. Last, J.A.; Jennings, M,D.; Schwartz, L.W.; Cross, C.E. Glycoprotein secretion by tracheal explants cultured from rats exposed to ozone. Am. Rev. Respir. Dis. 116:695-703; 1977. Last, J.A.; Cross, C.E. A new model for health effects of air pollutants: evidence for synergistic effects of mixtures of ozone and sulfuric acid aerosols on rat lungs. J, Lab. Clin. Med. 91:328-339; 1978. Dillard, C.J.; Urribarri, C.N.; Reddy, K.; Fletcher. B.; Taylor, S.; de Lumen, B.; Tappel, A.L. Increased lysosomal enzymes in lungs of ozone-exposed rats. Arch. Environ. Health 25:426--431; 1972. Evans, M.J.; Dekker, N.P.; Cabral-Anderson, L.J.; Shami, S.G. Morphological basis of tolerance to ozone. Exp. Mol. Pathol. 42: 366-376; 1985. Last, J.A.; Gerriets, J.E.; Hyde, D.M. Synergistic effects on rat lungs of mixtures of oxidant air pollutants (ozone or nitrogen dioxide) and respirable aerosols. Am. Rev. Respir. Dis. 128: 539-544; 1983. Last, J.A.; Hyde, D.M.; Chang, D.P.Y. A mechanism of synergistic lung damage by ozone and a respirable aerosol. Exp. Lung Res. 7:223-235; 1984. Hackney, J.D.; Linn, W.S.; Mohler, J.G.; Pederson, E.E,; Breisacher, P.; Russo, A. Experimental studies on human health effects of air pollutants. II. Four-hour exposure to ozone alone and in combination with other pollutant gases. Arch. Environ. Health 30:379-384; 1975. Hackney, J.D.; Linn, W.S.; Law, D.C.; Karuza, S.K.; Greenberg, H.; Buckley, R.D.; Pederson, E.E. Experimental studies on human health effects of air pollutants. III. Two-hour exposure to ozone alone and in combination with other pollutant gases. Arch. Environ. Health 30:385-390; 1975. von Neiding, G.; Wagner, H.M.; Krekeler, H.; LoUgen, H.; Fries, W.; Beuthan, A. Controlled studies of human exposure to single and combined action of NO 2, O3, and SO2. Int. Arch. Occup. Environ. Health 43:195-210; 1979. Folinsbee, L.J.; Bedi, J.F.; Horvath, S.M. Combined effects of ozone and nitrogen dioxide on respiratory function in man. Am. Ind. Hyg. Assoc. J. 42" 534-541; 1981. Toyama, T.; Tsunoda, T.; Nakaza, M.; Higashi, T.; Nakadate, T. Airway response to short-term inhalation of NO 2, 03, and their mixture in healthy men. Sangyo lgaku (Japan J. Ind. Hyg.) 23:285-293; 1981. Kagawa, J.; Tsuru, K. Respiratory effects of 2-hour exposure to ozone and nitrogen dioxide alone and in combination in normal subject performing intermittent exercise. Nippon Kyobu Shikkan Gakkai Zasshi (Japan J. Thorac. Dis.) 17:765-774; 1979. Freeman, G.; Jnhos, L.T.; Furiosi, N.J.; Mussenden, R.; Stephens, R.J.; Evans, M.J. Pathology of pulmonary disease from exposure to interdependent ambient gases (nitrogen dioxide and ozone). Arch. Environ. Health 29:203-210; 1974. Goldstein, E.; Warshauer, D.; Lippert, W.; Tarkington, B. Ozone and nitrogen dioxide exposure: murine pulmonary defense mechanisms. Arch. Environ. Health 28:85-90; 1974. Ehrlich, R.; Findlay, J.C,; Fenters, J.D.; Gardner, D.E. Health effects of short-term inhalation of nitrogen dioxide and ozone mixtures. Environ. Res. 14:223-231; 1977. Ehrlich, R.; Findlay, J.C.; Gardner, D.E. Effects of repeated exposures to peak concentrations of nitrogen dioxide and ozone on resistance to streptococcal pneumonia. J. Toxicol. Environ. Health 5:631--642; 1979.
Biochemical basis of ozone toxicity 199. Ehrlich, R. Interaction between environmental pollutants and respiratory infections. Environ. Health Perspect. 35:89-100; 1980. 200. Ehrlich, R. Changes in susceptibility to respiratory infection caused by exposures to photochemical oxidant pollutants. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biochemical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific PUbfishers; 1983:273-285. 201. Graham, J.A.; Gardner, D.E.; Blommer, E.J.; Honsem, D.E.; Menache, M.G.; Miller, F.J. Influence of exposure patterns of nitrogen dioxide and modifications by ozone on susceptibility to bacterial infectious disease in mice. J. Toxicol. Environ. Health 21:113-125; 1987. 202. Goldstein, B.D. Combined exposure to ozone and nitrogen dioxide. Environ. Health Perspect. 13:107-110; 1976. 203. Mnstafa, M.G.; Elsayed, N.M.; yon Dohien, F.M.; Hasset, C.M.; Postlethwait, E.M.; Quinn, C.L.; Graham, J.A.; Gardner, D.E. A comparison of biochemical effects of nitrogen dioxide, ozone, and their combination in mouse lung. I. Intermittant exposure. Toxicol. Appl. Pharmacol. 72:82-90; 1984. 204. Mnstafa, M.G.; Elsayed, N.M.; Lee, S.D. The combined effects of ozone and nitrogen dioxide on pulmonary biochemistry. In: Lee, S.D., ed. Evaluation of the scientific basis for ozone/ oxidants standards. Pittsburgh, PA: Air Pollution Control Association; 1985:337-347. 205. Mantz, W.J.; Kleinman, M.T.; Phalen, R.F.; Crocker, T.T. Effects of exercise I exposure on toxic interactions between inhaled oxidant and aldehyde air pollutants. J. Toxicol. Environ. Health 25:165-177; 1988. 206. Lee, J.-S.; Afifi, A.A.; Mustafa, M.G. Effects of short-term, single and combined exposure of rats to NO2 and 03 on lung tissue enzyme activities, lnhal. Toxicol. 1:21-35; 1989. 207. Lee, J.-S.; Mustafa, M.G.; Afifi, A.A. Effects of short-term, single and combined exposure to low-level NO2 and 03 on lung tissue enzyme activities in rats. J. Toxicol. Environ. Health 29:293-305; 1990. 208. Goldstein, B.D. Combined exposure to ozone and nitrogen dioxide. Environ. Health Perspect. 30:87-89; 1979. 209. Diggie, W.M.; Gage, J.C. The toxicity of ozone in the presence of oxides of nitrogen. Brit. J. lndustr. Med. 12:60-64; 1955. 210. Pins, Jr., J.N. Formation and fate of gaseous and particulate mutagens and carcinogens in real and simulated atmospheres. Environ. Health Perspect. 47:115-140; 1983. 211. piatt, U.F.; Wirier, A.M.; Blerman, H.W.; Atkinson, R.; Pitts, Jr., LN. Measurement of nitrate radical concentrations in continental air. Environ. Sci. Technol. 16:365-369; 1984. 212. Postlethwait, E.M.; Mustafa, M.G. The fate of inhaled nitrogen dioxide in the isolated perfused rat lung. J. Toxicol. Environ. Health 7:861-872; 1981. 213. Postlethwait, E.M.; Mustafa, M.G. Effect of altered dose rate on NO2 uptake and transformatiion in isolated lungs. J. Toxicol. Environ. Health 26:497-507; 1989. 214. Oda, H.; Nogami, H.; Nakajima, T. Reaction of hemoglobin with nitric oxide and nitrogen dioxide in mice. J. Toxicol. Environ. Health 6:673-678; 1980. 215. Chiodi, H.; Collier, C.R.; Mothler, J.G. In vitro methemogiobin formation in human blood exposed to NO 2. Environ. ICes. 30:9-15; 1983. 216. Kosaka, H.; Tyuma, I. Mechanism of autocatalytic oxidation of oxyhemoglobin by nitrite. Environ. Health Perspect. 7:147-151; 1987. 217. Klimmek, R.; Krettek, C.; Werner, H.W. Ferrihemoglobin formation by amyl nitrite and sodium nitrite in different species in vivo and in vitro. Arch. Toxicol. 62:152-160; 1988. 218. Stokinger, H.E.; Wagner, W.D.; Dobrogorski, O.J. Ozone toxicity studies. HI. Chronic effects. Arch. lndast. Health 16:514-522; 1957. 219. Melton, C.E. Effects of long-term exposure to low levels of ozone: a review. Aviat. Space Environ. Med. $3:105-111; 1982. 220. Filipowicz, C.; McCauley, R. The effect of chronic ozone exposure on lung benzo(a)pyrene oxidase and monoamine oxidase. Res. Commun. Chem. Pathol. Phormacol. $1:289-296;
263
1986. 221. Gross, K.B.; White, H.J. Functional and pathologic consequences of a 52-week exposure to 0.5 ppm ozone followed by a clean air recovery period. Lung 165:283-295; 1987. 222. Harman, D. Free radical theory of aging: effect of the amount and degree of unsaturation of dietary fat on mortality rate. J. Gerontol. 26:451-457; 1971. 223. Libovitz, B.E.; Siegel, B.V. Aspects of free radical reactions in biological systems and aging. J. GerontoL 35:45-56; 1980. 224. Ts'o, P.O.P.; Caspary, W.J.; Loventzen, R.J. The involvement of free radicals in chemical carcinogenesis. In: Pryor, W.A., ed. Free radicals in biology. Vol. 3. New York: Academic Press; 1977:251-303. 225. Demopoulos, H.B.; Peitronigro, E.S.; Flamm, E.S.; Seligman, M.L. The possible role of free radical reactions in carcinogenesis. In: Demopoulos, H.B.; Mehiman, M.A., eds. Cancer and the environment. Park Forest South, IL: Pathotox Publishers; 1980:273-303. 226. Totter, J.R. Spontaneous cancer and its possible relationship to oxygen metabolism. Proc. Natl. Acad. Sci. (USA) 77:17631767; 1980. 227. Troll, W.; Witz, G.; Goldstein, B.; Stone, D.; Sugimura, T. The role of free oxygen radical in tumor pormotion and carcinogenesis. In: Hecker, E.; Fusenig, N.E.; Kunz, W.; Marks, F.; Thielman, H.W., eds. Carcinogenesis and biological effects of tumor promoters. New York: Raven Press; 1982:593-597. 228. Chu, E.H.Y.; Trosko, J.E.; Chang, C. Mutational approaches to the study of carcinogenesis. J. Tox. Environ. Health 2:13171334; 1977. 229. Diamond, L.; O'Brien, T.G.; Ballrd, W.M. Tumor promoters and the mechanism of tumor promotion. Adv. Cancer Res. 32:1-74; 1980. 230. van Duuren, B.L. Tumor promoting and co-carcinogenic agents chemical carcinogenesis. In: Searle, C.E., ed. Chemical carcinogens. Washington, DC: Amercian Chemical Society; 1976: 24-51. 231. O'Brien, T.G.; Silmsiman, R.C.; Boutwell, R.K. Induction of the polyamine-biosynthetic enzymes in mouse epidermis and their specificity for tumor promotion. Cancer Res. 35:24262433; 1975. 232. Boutwell, R.K. Biochemical mechanism of tumor promotion. In: Slaga, T.J.; Sivak, A.; Boutwell, R.K., eds. Carcinogenesis, Vol 2. Mechanisms of tumor promotion and cocarcinogenesis. New York: Raven Press; 1978:49-58. 233. Hamelin, C.; Chung, Y.S. The effect of low concentrations of ozone on Escherichia coli chromosome. Murat. Res. 28:131132; 1975. 234. Marz, T.; Bender, M.; Kerr, H.; guile, T. Observations of aberrations in chromosomes of lymphocytes from human subjects exposed to ozone at concentrations of 0.5 ppm for 6 and 10 hours. Murat. Res. 31:299-302; 1975. 235. Guerrero, R.R.; Rounds, D.E.; Olson, R.S.; Hackney, J.D. Mutagenic effects of ozone on human cells exposed in vivo and in vitro based on sister chromatid exchange analysis. Environ. Res. 18:336--346; 1979. 236. Borek, C.; Ong, A.; Cleaver, J.E. DNA damage from ozone and radiation in human epithelial cells. Toxicol. lndast. Health 4:547-553; 1988. 237. Elsayed, N.M. Influence of vitamin E on polyamine metabolism in ozone-exposed rat lungs. Arch. Biochem. Biophys. 255: 392-399; 1987. 238. Elsayed, N.M.; Ellingson, A.S.; Tiemey, D.F.; Mustafa, M.G. Effects of ozone inhalation on polyamine metabolism and tritiated thymidine incorporation into DNA of rat lungs. Toxicol. Appl. Pharmacol. 102:1-8; 1990. 239. Kotin, P.; Falk, H.L. The experimental induction of pulmonary tumors in strain-A mice after their exposure to an atmosphere of ozonized gasoline. Cancer 9:910-917; 1956. 240. Kotin, P.; F a l l H.; McCammon, C J . The experimental induction of pulmonary tumors and changes in the respiratory epithelium in C57BL mice following their exposure to an atmosphere of ozoulzed gasoline. Cancer 11:473--481; 1958. 241. Stokinger, H.E. Effects of air pollution on animals. In: Stern,
264
M.G. MUSTAFA
A.C., ed. Air pollution. New York: Academic Press; 1962: 282-334. 242. Gardner, M.B. Biological effects of urban air pollution: lung tumors in mice. Arch. Environ. Health 12:305-313; 1966. 243, Werthamer, S.; Schwartz, L.H.; Soskind, L. Bronchial epithelial alterations and pulmonary neoplasia induced by ozone. Path. Microbiol. 35:224-230; 1970. 244. Nettesheim, P.; Hanna, Jr., M.G.; Doherty, D.G.; Newell, R.F.; Hellman, A. Effects of chronic exposure to artificial smog and chromium oxide dust on the incidence of lung tumors in mice. In: Hanna, M.G.; Nettesheim, P.; Gilbert, J.G., eds. Inhalation carcinogenesis, symposium series No. 18. Washington, DC: USAEC Division of Technical Information Extension; 1970: 305-317. 245. Hassett, C.; Mustafa, M.G.; Coulson, W.F.; Elashoff, R.M. Murine lung carcinogenesis following exposure to ambient ozone concentrations. J. Nat. Cancer Inst. 75:771-777; 1985. 246. Last, J.A.; Warren, D.L.; Pecquet-Goad, E.; Witschi, H.P. Modification of lung tumor development in mice by ozone. J. Nat. Cancer Inst. 78:149-154; 1986. 247. Maugh, T.H. New link between ozone and cancer. Science 216:396--397; 1982. 248. Mustafa, M.G.; Hassett, C.M.; Newell, G.W.; Schrauzer, G.N. Pulmonary carcinogenic effects of ozone. In: Maltoni, C.; Selikoff, I.J., eds. Living in a chemical world: occupational and environmental significance of industrial carcinogens. New York: Ann. N.Y. Acad. Sci.; 1988:714--723. 249. Witschi, H.P. Ozone, nitrogen dioxide and lung cancer: a review of some recent issues and problems. Toxicol. 48:1-20; 1988. 250. Menzel, D.B.; Slaughter, R.J.; Bryant, A.M.; Jauregui, H.O. Heinz bodies formed in erythrocytes by fatty acid ozonides and ozone. Arch. Environ. Health 3t1:296-301; 1975. 251. Clark, K.W.; Posin, C.I.; Buckley, R.D. Biochemical response of squirrel monkeys to ozone. J. Toxicol. Environ. Health 4"741-753; 1978. 252. Moore, G.S.; Calabrese, E.J.; Labato, F.J. Erythrocyte survival in sheep exposed to ozone. Bull. Environ. Contain. Toxicol. 27:126-138; 1981. 253. Larkin, E.C.; Goheen, S.C.; Rao, G.A. Morphology and fatty acid compositon of erythrocytes from monkeys exposed to ozone for one year. Environ. Res. 32:445-454; 1983. 254. Calabrese, E.J.; Moore, G.S.; Grunwald, E.L. Ozone-induced decrease of erythrocyte survival in adult rabbits. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983: 103-I 17. 255. Ballew, M.; Calabrese, E.J.; Moore, G.S. The effect of dietary vitamin C on ozone-induced oxidative changes in guinea pig erythrocytes. J. Environ. Sci. Health A18:597-610; 1983. 256. Vaughan, W.J.; Adamson, G.L.; Lindgren, F.T.; Schooley, J.C. Serum lipid and lipoprotein concentrations following exposure to ozone. J. Environ. Pathol. Toxicol. 0ncol.5:165-173; 1984. 257. Mole, Jr., M.L.; Stead, A.G.; Gardner, D.E.; Miller, F.J.; Graham, J.A. Effect of ozone on serum lipids and lipoproteins in the rat. Toxicol. Appl. Pharmacol. 80:367-376; 1985. 258. Konisberg, A.S.; Bachrnan, C.H. Ozonized atmosphere and gross motor activity or rats. Int. J. Biometeorol. 14:261-266; 1970. 259. Murphy, S.D.; Ulrich, C.E.; Frankowitz, S.H.; Xintaras, C. Altered function in animals inhaling low concentrations of ozone and nitrogen dioxide. Am. Ind. Hyg. Assoc. J. 2,5.246-253; 1964. 260. Weiss, B.; Ferin, J.; Merigan, W.; Stern, S.; Cox, C. Modification of rat operant behavior by ozone exposure. Toxicol. Appl. Pharmacol. 58:244-251; 1981. 261. Tepper, J.L.; Weiss, B.; Cox, C. Microanalysis of ozone depression of motor activity. Toxicol. Appl. Pharmacol. 64: 317-326; 1982. 262. Tepper, J.L.; Weiss, B.; Wood, R.W. Behavioral indices of ozone exposure. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.G., eds. International symposium on the biomedical effects of ozone-related photochemical oxidants. Princeton, NJ: Princeton
Scientific Publishers; 1983:515-526. 263. Trams, E.G.; Lauter, C.J.; Brown, E.A.B.; Young, O. Cerebral cortical metabolism after chronic exposure to ozone. Arch. Environ. Health 24:153-159; 1972. 264. Friedman, M.; Gallo, J.M.; Nichols, H.P.; Bromberg, P.A. Changes in inert gas rebreathing parameters after ozone exposure in dogs. Am. Rev. Respir. Dis. 128:851-856; 1983. 265. Hassett, C.; Mustafa, M.G.; Coulson, W.F.; Elashoff, R.M. Splenomegally in mice following exposure to ambient levels of ozone. Toxicol. Lett. 26:139-144; 1985. 266. Gardner, D.F.; Illing, J.W.; Miller, F.J.; Coffin, D.L. The effects of ozone on pentobarbital sleeping time in mice. Res. Commun. Chem. Pathol. Pharmacol. 9:689-700; 1974. 267. Graham, J.A.; Menzel, D.B.; Miller, F.J.; Illing, J.W.; Gardner, D.E. Influence of ozone on pentobarbital-induced sleeping time in mice, rats, and hamsters. Toxicol. Appl. Pharmacol. 61: 64-73; 1981. 268. Graham, J.A.; Menzel, D.B.; Miller, F.J.; Illing, J.W.; Gardner, D.E. Effect of ozone on drug-induced sleeping time in mice pretreated with mixed-function oxidase inducers and inhibitors. Toxicol. Appl. Pharmacol. 62:489--497; 1982. 269. Graham, J.A.; Miller, F.J.; Gardner, D.E.; Ward, R.; Menzel, D.B. Influence of ozone and nitrogen dioxide on hepatic microsomal enzymes in mice. J. Toxicol. Environ. Health 9: 849-856; 1982. 270. Atwal, O.S.; Wilson, T. Parathyroid gland changes following ozone inhalation: a morphologic study. Arch. Environ. Health 28:91-100; 1974. 271. Clemons, G.K.; Garcia, J.F. Changes in thyroid function after short-term ozone exposure in rats. J. Environ. Pathol. Toxicol. 4:359-369; 1980. 272. Clemons, G.K.; Garcia, J.F. Endocrine aspects of ozone exposure in rats. Arch. Toxicol. (Suppl.) 4:301-304; 1980. 273. Clemons, G.K.; Wei, D. Effect of short-term ozone exposure on exogenous thyroxine levels in thyroidectomized and hypophysectomized rats, Toxciol. Appl. Pharmacol. 74:86--90; 1984. 274. Kavlock, R.; Daston, G.; Grabowski, C.T. Studies on the development toxicity of ozone. I. Prenatal effects. Toxicol. Appl. Pharmacol. 48:19-28; 1979. 275. Kavlock, R.J.; Meyer, E.; Grabowski, C.T. Studies on the developmental toxicity of ozone: postnatal effects. Toxicol. Len. 5:3-9; 1980. 276. Gooch, P.C.; Creasia, D.A.; Brewen, J.G. The cytogenetic effect of ozone: inhalation and in vitro exposures. Environ. Res. 12:188-195; 1976. 277. Tice, R.R.; Bender, M.A.; Ivett, J.L.; Drew, R.T. Cytogenetic effects of inhaled ozone. Mutat. Res. 58:293-304; 1978. 278. Sweet, F.; Kao, M.-S.; Lee, S.D.; Hager, W.L.; Sweet, W.E. Ozone selectively inhibits growth of human cancer cells. Science 2119:931-933; 1980. 279. Chow, C.K. Nutritional influence on cellular antioxidant defense systems. Am. J. Clin. Nutr. 32:1066-1081; 1979. 280. Machlin, L.J.; Bendich, A. Free radical tissue damage: protective role of antioxidant nutrients. FASEB J. 1:441--445; 1987. 281. Heffner, J.E.; Repine, I.E. State of the art--pulmonary strategies of antioxidant defense. Am. Rev. Respir. Dis. 140:531-554; 1989. 282. Fukase, O.; Watanabe, H.; Isomura, K. Effects of exercise on mice exposed to ozone. Arch. Environ. Health 33:198-200; 1978. 283. Tappel, A.L. Will antioxidant nutrients slow aging process? Geriatrics 23:97-105; 1968. 284. Goldstein, B.D.; Buckley, R.D.; Cardenas, R.; Balchum, O.J. Ozone and vitamin E. Science 169:605--606; 1970. 285. Fletcher, B.L.; Tappel, A.L. Protective effects of alpha-tochopherol in rats exposed to toxic levels of ozone and nitrogen dioxide. Environ. Res. 6:165-175; 1973. 286. Mustafa, M.G. Influence of dietary vitamin E on lung cellular sensitivity to ozone in rats, Nutr. Rep. lnt'l. 11:473-476; 1975. 287. Donovan, D.H.; Williams, S.J.; Charles, J.M.; Menzel, D.B. Ozone toxicity: effect of dietary vitamin E and polyunsaturated fatty acids. Toxicol. Lett. 1:135-139; 1977. 288. Elsayed, N.M.; Kass, R.; Mustafa, M.G.; Hacker, A.D.; Ospi-
Biochemical basis of ozone toxicity
289.
290. 291. 292. 293. 294. 295. 296. 297. 298.
299. 300. 301.
302. 303.
304.
305. 306. 307.
308. 309. 310. 311.
tal, J.J.; Chow, C.K.; Cross, C.E. Effect of dietary vitamin E level on the biochemical response of rat lung ot ozone. DrugNutrient Interact. 5:373--386; 1989. EBayed, N.M.; Mustafa, M.G.; Hacker, A.D.; Kuehn, K.; Schrauzer, G.N. Dietary antioxidants and the biochemical response to oxidant inhalation. IIL Selenium influence on mouse lung response and tolerance to ozone. Biol. Trace Element Res. 6:249-261; 1984. Willis, R.I.; Kratzing, C.C. Ascorbic acid in rat lung. Biochem. Biophys. Res. Commun. 59:1250-1253; 1974. Stokinger, H.E. Evaluation of the hazards of ozone and oxides of nitrogen. Arch. Inudst. Health 15:181-190; 1957. Matzen, R.N. Effect of vitamin C and hydrocortisone on the pulmonary edema produced by ozone in mice. J. Appl. Physiol. 11:105-109; 1957. Freebairn, H.T. Reversal of inhibitory effects of ozone on oxygen uptake of mitochondria. Science 126:303-304; 1957. Matzen, R.N. Development of tolerance to ozone in reference to pulmonary edema. Am. J. Physiol. 190:84-88; 1957. Willis, R.J.; Kratzing, C.C. Extracellular ascorbic acid in lung. Biachim. Biophys. Acta 144:108-117; 1976. Kratziug, C.C.; Willis, R.J. Decreased levels of ascorbic acid in lung following exposure to ozone. Chem. Biol. Interact. 30: 53-56; 1980. Dubick, M.A.; Heng, H.; Rucker, R.B. Effects of protein deficiency and food restriction on lung ascorbic acid and glutathione in rats exposed to ozone. J. Nutr. 115:1050-1056; 1985. Cawthome, M.A.; Bunyan, J.; Sennitt, M.V.; Green, J. Vitamin E hepatotoxic agents. 3. Vitamin E, synthetic antioxidants and carbon tetrachloride toxicity in the rats. Br. J. Nutr. 24:357-384; 1970. Goyer, R.A.; Mehlman, M.A. Toxicology of trace elements. Washington, DC: Hemisphere Publishing Corp.; 1977:191-240. Chortyk, O.T.; Baker, J.T.; Chamberlain, W J . Selenium-mediated reduction in the mutagenicity of cigarette smoke. Environ. Molec. Mutage. 11:369-378; 1988. Johnson, G.J.; Vatassery, G.T.; Finkel, B.; Allen, D.W. Highdose vitamin E does not decrease the rate of chronic hemolysis in giucose-6-phosphate dehydrogenase deficiency. N. Eng. J. Med. 308:1014-1017; 1983. Diplock, A.T.; Lucy, J.A. The biochemical modes of action of vitamin E and selenium; A hypothesis. FEBS Lett. 29:205-210; 1973. Wilson, R.L. Free radical protection: Why vitamin E, not vitamin C, b-carotene or glutathione? In: Biology of vitamin E. Ciba Foundation Symposium 101. London: Pitman Books; 1983: 19--44. Burton, G.W.; Joyce, A.; Ingold, K.U. Is vitamin E the only lipid-soluble, chain-breaking antioxidant in human blood plasma and erythrocyte membranes? Arch. Biochem. Biophys. 221: 281-290; 1983. Packer, J.E.; Slater, T.F.; Wilson, R.W. Direct observation of a free radical interaction between vitamin E and vitamin C. Nature (London) 278:737-738; 1979. Chem, L.H.; Lee, M.S.; Hsing, W.F.; Chen, S.H. Effect of vitamin C on tissue antioxidant status of vitamin E deficient rats. Int. J. Nutr. Res. 50:156-162; 1980. Leung, H.-W.; Wang, M.J.; Mavis, R.D. The cooperative interaction between vitamin E and vitamin C in suppression of peroxidation of membrane phospholipids. Biochim Biophys. Acta 664:266-272; 1981. Slater, T.F. Free radical mechanisms of tissue injury. Biochem. J. 222:1-15; 1984. McCay, P.B. Vitamin E: interactions with free radicals and ascorhate. Annu. Rev. Nutr. 5:323-340; 1985. Lambelet, P.; Saucy, F.; Loliger, J. Chemical evidence for interactions between vitamins E and C. Experientia 41:13841388; 1985. Roberts, H.J. Perspective on vitamin E as therapy. J. Am. Med. Assoc. 246:129-131; 1981.
265
312. Fen'ell, P.M. Deficiency states, pharmacological effects, and nutrient requirements. In: Machlin, L.J., ed. Vitamin E: a comprehensive treatise. New York: Marcel Dekker; 1980:520620. 313. Witting, L.A. Vitamin E and lipid antioxidants in free-radical initiated reactions. In: Pryor, W.A., ed. Free radicals in biology, Vol. 4. New York: Academic Press; 1980:295-319. 314. Walton, J.R.; Packer, L. Free radical damage and protection: relatrionship to cellular aging and cancer. In: Machlin, LJ., ed. Vitamin E: a comprehensive treatise. New York: Marcel Dekker; 1980:495-517. 315. Bieri, J.G.; Corash, L.; Hubbard, V.S. Medical uses of vitamin E. N. Eng. J. Med. 308:1063-1071; 1983. 316. Horwitt, N.K. Status of human requirements for vitamin E. Am. J. Clin. Nutr. 27:1182-1193; 1974. 317. Bieri, J.G.; Evarts, R.P. Tocopherols and fatty acids in American diets. The recommended allowance for vitamin E. J. Am. Diet Assoc. 62:147-151; 1973. 318. National Academy of Sciences. Recommended dietary allowances. 9th ed. Washington, DC: National Academy Press; 1980:63--69. 319. Forman, H.J.; Fisher, A.B. Antioxidant enzymes of rat granular pneumocytes--constitutive levels and effect of hypoxia. Lab. Invest. 45:1-6; 1981. 320. Halliwell, B.; Wast, M.; Cn'ootveld, M. Biologically significant scavenging of the myeloperoxidase-derived oxidant hypochlorous acid by ascorbic acid. FEBS Left. 213:15-18; 1987. 321. Higson, F.K.; Keen, R.; Chevion, M. Iron enhancement of ascorbate toxicity. Free Rad. Res. Commun. 5:107-115; 1989. 322. Heys, A.D.; Dormandy, T.L. Lipid peroxidation in iron-overloaded spleens. Clin. Sci. 60:295-301; 1981. 323. Halliwell, B.; Gutteridge, J.M.C. Oxygen toxicity, oxygen radicals, transition metals and disease. Biochem. J. 219:1-14; 1977. 324. Rawley, D.A.; Halliwell, B. Formation of hydroxyl radicals from hydrogen peroxide and iron salts by superoxide- and ascorhate-dependent mechanisms: relevance to the pathology of rheumatoid disease. Clin. Sci. 64:649--653; 1983.
ABBREVIATIONS
VOC--volatile organic compounds NOx--oxides of nitrogen PUFA--polyunsaturated fatty acids SH--sulfhydryl group SS--disulfide group PSH--protein sulfhydryls NPSH--nonprotein sulfhydryls GSH--reduced glutathione GSSG--oxidized glutathione PSSG--mixed disulfide GP-- glutathione peroxidase GR--glutathione reductase MAO--monoamine oxidase RBC--red blood cells Hb-- ferrohemoglobin HbO:-- oxyhemoglobin Hb--ferdhemoglobin or methemoglobin butylated hydroxytoluene-- 3,5-tert-butyl-4-hydroxytoluene butylated hydroxyanisole--2- or 3-tert-butyl-4-hydroxyanisole
0891-5849/90 $3.00 + .00 Copyright© 1990PergamonPress plc
Printedin the USA. All rightsreserved.
Review Article BIOCHEMICAL BASIS OF OZONE TOXICITY MOHAMAD G. MUSTAFA Department of Environmental Health Sciences, School of Public Health, University of California, Los Angeles, CA 90024, U.S.A. (Received 18 December 1989; Revised and Accepted 18 May 1990)
Abstract--Ozone (03) is the major oxidant of photochemical smog. Its biological effect is attributed to its ability to cause oxidation or peroxidationof biomoleculesdirectly and/or via free radical reactions. A sequence of events may include lipid peroxidationand loss of functional groups of enzymes, alteration of membrane permeability, and cell injury or death. An acute exposure to 03 causes lung injury involving the ciliated cell in the airways and the type 1 epithelial cell in the alveolar region. The effects are particularly localized at the junction of terminal bronchioles and alveolar ducts, as evident from a loss of cells and accumulation of inflammatory cells. In a typical short-term exposure the lung tissue response is biphasic: an initial injury-phase characterized by cell damage and loss of enzyme activities, followed by a repair-phase associated with increased metabolic activities, which coincide with a proliferation of metabolically active cells, for example, the alveolar type 2 cells and the bronchiolar Clam cells. A chronic exposure to 03 can cause or exacerbate lung diseases, including perhaps an increased lung tumor incidence in susceptible animal models. Ozone exposure also causes extrapulmonary effects involving the blood, spleen, central nervous system, and other organs. A combination of 03 and NO2, both of which occur in photochemicalsmog, can produce effects which may be additive or synergistic. A synergisticlung injury occurs possibly due to a formation of more powerful radicals and chemical intermediates. Dietary antioxidants, for example, vitamin E, vitamin C, and selenium, can offer a protection against 03 effects. Keywords--Antioxidant protection, Extrapulmonary effects, Lung injury, Lung tumor, Nitrogen dioxide' toxicity, Ozone toxicity, Photochemical oxidants, Synergistic effects, Free radicals
INTRODUCTION
incidence, are thought to result from exposures to this oxidant. The toxic effects are attributed to its ability to generate free radical reactions in the biological system. The purpose o f this presentation is to review our understanding o f the toxicity and health effects o f O 3, including the possible involvement o f free radicals in oxidant injury and antioxidant protection.
Ozone is the major oxidant of photochemical smog, and generally occurs in association with other oxidant components, namely, nitrogen dioxide (NO2), peroxyacyl nitrates (PAN), hydrogen peroxide, alkyl peroxides, nitrous and nitric acids, formaldehyde and formic acid, and traces of other compounds. 1-3 Collectively, these chemicals are referred to as oxidants because they have the ability to remove electrons from other molecules. Of these oxidants, toxicity of O3 has been studied in relatively greater details, because of its abundance and earlier recognition in photochemical smog and the magnitude o f its potential effects on humans, plants, and ecosystem. The presence o f 0 3 in the air poses a serious health concern. Many short-term health effects and chronic lung diseases, including, perhaps, an increased tumor
OCCURRENCE OF OZONE
Ozone occurs as a natural component o f the atmosphere. However, there is a distinction in its occurrence in the upper atmosphere (stratosphere) and the lower atmosphere (e.g., smog in lower troposphere) as well as the biologic consequences. Ozone in smog is a common cause of urban air pollution, posing a threat to human health and property. 1-3 Stratospheric 03 is known to protect us from the harmful cosmic radiation, 4-7 although it may be a problem in high-altitude flights because the air-intake for airplane cabins includes O3 .s-12
Mohammad G. Mustafa (b. 1940, Bangladesh; Ph.D., 1969 in Biochemistry), is Professor of Enviommental Health Sciences, University of California, Los Angeles (UCLA). He worked with the Pulmonary (Air Pollution) Research Group at the U.C. Davis until his move to UCLA in 1975. His current researches focus on pulmonary effects of oxidant air pollutants (ozone, NO2) and other areas of inhalation
A. Formation in stratosphere
Atmospheric 03 concentration is the highest in stratosphere, a region extending from about 8 km at the poles and 17 km at the equator to about 50 km above the
toxicology. 245
246
M.G. MUSTAFA
earth's surface. 13 Ozone is thought to be generated according to the Chapman cycle of photochemical dissociation of molecular oxygen ( 0 2) by ultraviolet radiation (reaction 1.1) and then a chemical combination of atomic oxygen (O) with O 2 (reaction 1.2). 14-17 Ozone thus formed does not accumulate endlessly. The process of 03 formation is in a dynamic equilibrium with its natural destruction, for example, due to photochemical dissociation by ultraviolet radiation (reaction 1.3a) and also by infrared radiation (reaction 1.3b), collision with atomic oxygen (reaction 1.4), reaction with hydroxyl (HO) radical (reaction 1.5), and various other chemical and physical processes. 14-17 Some of the stratospheric 0 3 evidently reaches the lower troposphere providing the global O 3 background of up to 0.04 ppm. 1.2,18,19 However, it is the 0 3 produced in ambient air by photochemical reactions of primary (or precursor) pollutants that is of air pollution concern.
O2
hv (k
O+ O
(1.1)
M
O2 + O > 03 (1.2) (M = a third-body molecule absorbing excess energy of reaction) hv (k O2 + O (1.3a) hv (h < 1140 nm) O3 > O2 + O (1.3b) 03 + O > 02 + 02 (1.4) O3 + HO > HO2 + 02 (1.5)
Stratospheric 0 3 layer acts as a shield for the earth, protecting its surface, that is, preventing adverse effects on human health (skin cancer and suppression of immune response) and on terrestrial and aquatic ecosystems, from all UV-C (k
0 3 + 2NO 2
NO3 + NO2 N205 + 03
M
--'> M
>
N205 + 02
(1.19)
N205
(1.20)
NO 2 + 03
(1.21)
In addition, the reactions of 03, NO2, N205, and NO 3 in the moist environment of the bronchiolo-alveolar junctions (or with lung tissue) give rise to other chemical species, including HNO2, HNO 3, N O 2 - , N O 3 - , and HO radical (reactions 1.22-1.30). In lung tissue, NO 2 and NO 3 can be reduced to NO 2- and NO 3 - , respectively, in the presence of an electron donor (D), for example, a reducing substance, which then becomes a
254
M.G. MUSTAFA
radical cation, D + (reactions 1.22 and 1.30). Isolated lung perfusion studies have shown that the product of NO 2 absorption (or reaction) in lung tissue is N O 2 - , and it is not oxidized to NO 3 - (reaction 1.25) unless the red blood cells (RBC) are also present. 212'213 Oxidation of NO 2- to NO 3- and conversion of ferrohemoglobin (Hb) or oxyhemoglobin (HbO 2) to ferrihemoglobin (HB ÷ or metHb) represent a rather complex reaction system, 2~4-217 but the overall process can be shown as reaction 1.25. However, NO 3 radical can directly oxidize a biomolecule (D) and become reduced (reaction 1.30) without the presence of RBC,
NOz + D NO2 + NO + HzO HNO2 4NO 2 + 4HbO2 + 4H + N205 + H20 NO3 + H20 NO2 + HO HNO3 NO 3 + D
--> --> --> -->
--> -->
--> --> -->
NO2- + D ÷ 2HNO2
(1.22) (1.23)
NO2- + H + + 4Hb ÷ + Oz + 2H20 2HNO3 HNO 3 + HO HNO 3 NO 3- + H + NO3- + D +
(1.24) (1.25)
4NO 3 -
(1.26) (1.27) (1.28) (1.29) (1.30)
Formation of intermediate chemical species discussed above may explain the synergistic effects of 0 3 plus NO t system relative to 0 3 or NO 2 alone. The toxicity caused by 0 3, NO z, and their radical and nonradical reaction products must be understood apart from one another. Once the toxic potential of, at least, the major components are known, the phenomenon of additive or synergistic effects can be better explained.
D. Long-term effects Most experimental studies involving animals and controlled human subjects provide data on O3 toxicity that are derived from relatively short-term exposures, and often using O a concentrations higher than those encountered in the ambient atmosphere. Nonetheless, they are essential for regulatory processes and formulation of measures to prevent both short-term and longterm health effects.
a. Chronic diseases. Relatively long-term or chronic effects of 0 3 are not well documented. Experimental studies involving long-term exposures are limited. Epidemiological studies suggest that chronic oxidant exposures affect the baseline respiratory functions ,48 although the studies linking increased incidence of chronic disease with oxidants of photochemical smog are not very convincing. Based on relatively short-term animal
exposure studies, it is predicted that an acceleration of the deterioration of lung function with time (e.g., age of individuals) might result from chronic oxidant exposures. There is a depletion of lung cells and loss of lung reserves, which may not be obvious clinically but might produce a cumulative effect over a period of some years. 139 Destructive lung diseases, for example, emphysema, and other chronic obstructive lung diseases, for example, chronic bronchitis, and development of airway hypersensitivity or asthma might be the ultimate results of chronic oxidant exposures. 1-3,12,31,41,48.218-221 A continued (or cumulative effect of) oxidative and free radical reactions might be the underlying cause of lung cell depletion and functional deterioration. 222'223
b. Tumorigenic effects. A carcinogen (or tumorigen) is capable of causing several cellular reactions or processes that are thought to be associated with carcinogenesis (or tumorigenesis). These include induction of electron transfer or free radical mechanism; 224-227 chromosomal alteration or mutagenesis; 228'229 hyperplasia, irritation, and inflammation; 229'23° and omithine decarboxylase (ODC) activity. 231'232 Ozone exposure causes hyperplasia, irritation, and inflammation at the sites of lung injury. The hyperplasia can be repeatedly induced by recurrent or intermittent exposures. 42"149'150'154'156 Inflammatory cells release free radicals, for example, 0 2- and HO, which may cause further tissue injury, including DNA breaks.43"~38 Ozone causes chromosomal alterations, s~-55'233-236 which may be due to an attack on chromosomes by either 0 3 or free radicals generated by 03. Recently, a stimulation of lung ODC activity has been observed with 0 3 exposure. 237'238 Ozone, therefore, may be a potential carcinogen or promoter of carcinogenic process. Several experimental studies have been conducted to explore the carcinogenic potential of 0 3. Kotin et al.239,240 showed that mice after a year-long exposure to ozonized gasoline had a greater incidence of lung adenomas compared with control mice breathing a "washed atmosphere." Their synthetic atmosphere was intended to be qualitatively similar to Los Angeles smog, except the 0 3 concentration which was at 4 ppm (7840 ~g/m3). Stokinger 24~ reported 0 3 as a lung tumor accelerator on the basis of an increased tumor incidence in mice exposed to 1 ppm (1960 ixg/m3) 03 daily for 15 months. Gardner 242 exposed mice to ambient smog of Los Angeles for 7-11 months, and observed a trend toward increased incidence of pulmonary adenomas compared to that in age-matched clean-air control mice. The oxidant concentration in the smog ranged between 0.04 and 0.25 ppm (78 and 490 i~g/m3). A likelihood that ambient smog possesses a tumor-promoting activity in mice was suggested in this study. Two other studies in which O3-induced lung tumor incidence was significant
Biochemical basis of ozone toxicity
255
were carded out by Werthamer et al.243 and Nettesheim et al.244 In the latter study, mice were exposed to artificial smog (ozonized gasoline) containing 1 ppm (1960 p,g/m2) O 3 for up to 18 months. The incidence of lung tumors, which were adenomas and adenocarcinomas, was significantly greater in the smog-exposed mice than in the clean-air controls. Recently, Hassett et al. 245 examined the carcinogenic potential of 0 3 using A/J strain mice. A group of 45 mice was exposed to 0.50 ppm (980 ~g/m 3) 0 3 intermittently for 6 months; another group of 45 mice breathed clean air for the same time period. The lung tumor incidence was significantly higher in the O3-exposed mice. The result with a lower level of 0 3 (0.31 ppm) was not statistically significant. In another study, Last et al. 246 exposed AJJ strain and Swiss-Webster (SW) mice to 0.8 ppm 0 3 intermittently for approximately 4 months, and observed a significant increase in lung adenomas in m/J strain but not in SW mice relative to their corresponding controls. Since SW mice are relatively resistant to spontaneous or induced tumorigenesis, a 4-month long exposure may not have been chronic enough. In this study, exposure to a lower level of 03 (0.4 ppm) was not effective for A/J strain mice. Again, the exposure period may not have been chronic enough to make a difference between spontaneous and induced tumorigenesis. Although the major finding of Hassett et al. 245 and of Last et al. 246 that O 3 increases tumorigenesis in a susceptible mouse strain is in agreement, there remains an important point of difference. Hassett et al. 245 observed a synergism in lung adenoma formation when O 3 exposure was associated with a recurrent low-dose administration of urethane. Last et al. 246 observed an inhibition of lung adenoma formation when 0 3 exposure followed a onetime large-dose administration of urethane. This apparent contradiction may have to be resolved in a study specifically designed to address the issue of whether 0 3 is a cotumorigen/cocarcinogen. The subject of 0 3 tumorigenicity/carcinogenicity has recently been
defined. The effect of 03 on RBC and other hematologic parameters has been studied after both in vitro and in
r e v i e w e d . 48,247-249
Other extrapulmonary organs and tissues that have been examined for 0 3 effects include cardiovascular system, spleen, liver, endocrine system, and reproductive system. In animal studies, the heart rate and blood pressure were variably affected by O 3 exposure. 179.264 Spleen was found to be enlarged after a subchronic 0 3 exposure, 265 and this might be related to 0 3 effects on the blood. The O3 effects in the liver were variable. An evidence for alterations of zenobiotic metabolism, for example, an increase in pentobarbital-induced sleeping time, was reported, 266-268 but measurement of cytochrome P-450 concentration94"269 or benzopyrene hydroxylase activity89 showed no change. Ozone exposure was found to cause morphological changes in the parathyroid, 2°6 and a decrease in thyrotropin and thryoid hromones. 27°-273 Studies of O 3
EFFECTS IN OTHER ORGANS
Although the lung is the primary target for O 3 toxicity, the effects of 0 3 in other organs have also been documented. How 0 3 toxicity can reach the extrapulmonary tissues and organs is not clear, but it is possible that 0 3 reaction products cross the alveolar air-blood barrier, causing damage to various extrapulmonary sites. A. Hematopoietic system
Hematological parameters are frequently used to assess 0 3 toxicity, because blood chemistry is relatively simple and RBC morphology and metabolism are well
v i v o exposure. 52'54'69'71'159'160'171'250-257 T h e s e paranl-
eters include RBC morphology, survival, osmotic fragility, and oxyhemoglobin affinity; levels of GSH, ascorbate, hemoglobin, Heinz bodies, and methemoglobin; levels of serum lipids, cholesterol, albumin, and globulin; and RBC or plasma enzyme activities, namely, acetylcholine esterase, glucose-6-phosphate dehydrogenase, creatine phosphokinase, glutamate-pyruvate transaminase, pyruvate kinase, lactate dehydrogenase, and glutathione peroxidase. Ozone had either depressing or stimulating effects on these parameters, depending upon the 0 3 exposure conditions and nutritional status of the animals or controlled human subjects. For example, RBC morphology was altered, 253 survival was decreased, 252 and osmotic fragility increasedSZ'17a'251'254; GSH level was decreased251 or not changed159"255; serum cholesterol was increased256'257; acetylcholine esterase activity was decreased, 166 and the activities of glutathione peroxidase, pyruvate kinase, and lactate dehydrogenase were increased. 160
B. Central nervous system
Although CNS is adversely affected by O 3 exposure, and various physical and behavioral effects, for example, dizziness and visual impairment, have been noted, their biochemical basis is not clear. In animal studies, motor activity was found to be decreased by 0 3 exposure. 258 A change in behavior associated with a decreased running activity was reported by several laboratories. 259-262 A variability in enzyme activity, for example, a decrease in catechol methyl transferase and increase in monoamine oxidase, was also noted. 263
C. Other organs and tissues
256
M.G. MUSTAFA
effects on the reproductive system are limited. Some of the important effects noted were a decreased maternal weight gain and increased fetal resorption, 274 an increased neonatal mortality, 52 and slower development of offspring. 275 In addition, chromosomal alterations were observed in vitro in peripheral blood lymphocytes 54'234'235"276'277 and other mammalian cells. 51'236'278 The molecular mechanism for 0 3 effects on chromosomes is not clear, but single-strand DNA breaks may contribute to this effect. Zelac et al. 54 reported DNA strand breaks in lymphocytes of Chinese hamsters after in vivo 03 exposure, but others failed to observe such an in vivo effect in lymphocytes of Chinese hamsters 277 or humans. 235
ANTIOXIDANT PROTECTION
Since oxidative, peroxidative, and free radical reactions are important mechanisms of 0 3 toxicity, investigations have been carried out to discern antioxidant defense against oxidant damage. Living cells contain metabolic mechanisms (a variety of enzyme activities and metabolites) 42'8°'279-281 to combat oxidant stress, and these may be referred to as the intrinsic metabolic defense. Although the metabolic defense process prevents the formation of and/or eliminates most of the free radicals and peroxides, some free radicals (e.g., HO and HO 2) may not be destroyed by this means. Supplementation of ceils or organisms with antioxidant substances has been attempted as a means to contain or prevent free radical damage. External antioxidants often act in conjunction with metabolic defense and provide a strong antioxidant protection. 279-281
A. Metabolic defense Various antioxidant enzyme activities and metabolites (generated in situ by metabolic activites) contribute to metabolic defense mechanisms. 42'8°'279-281 One such mechanism, referred to as the GSH redox cycle, is thought to play a pivotal role in antioxidant defense against lipid peroxides and other toxic and related oxygenated intermediates. 42'44'45'279-281 In this cycle, GSH is oxidized to GSSG while reducing a peroxide in a GP-catalyzed reaction (1.31). GSH is then regenerated through a GR-catalyzed reaction (1.32) using the reducing equivalents of NADPH furnished by glucose oxidation via the pentose phosphate-shunt pathway (reactions 1.33ab). Activities of various enzymes, namely, glutathione peroxidase and associated thiol-metabolizing enzymes (glutathione reductase, disulfide reductase, thiol-disulfide transhydrogenase), peroxidase, catalase, superoxide dismutase, and key enzymes of the pentoseshunt pathway (glucose-6-phosphate and 6-phosphogluconate dehydrogenases), have been studied as a part of
the metabolic defense mechanism in the lung. 74'75'82'149' 150,151,154,157,158,162-165,167,168,279-282 Typical antioxidant metabolites studied are GSH and NADPH. 38"42' 84,279-281 In addition to antioxidant protection, metabolic defense mechanisms can promote the repair of injured cells and tissues, and enable the lung to withstand continuing injury. 42'84 2GSH +
GP
-> GSSG + ROH + H20
(1.31)
-> 2GSH + NADP ÷
(1.32)
ROOH
GSSG + NADPH + H+ NADP ÷ + G6P 6PG + NADP ÷
GR
G6PD 6PGD
-> NADPH + H ÷ 6PG
(1.33a)
•> R5P + NADPH + H
(1.33b)
(where, ROOH = a hydroperoxide; GP = glutathione peroxidase; GR = glutathione reductase; G6P --glucose-6-phosphate; 6PG = 6-phosphogluconate; G6PD and 6PGD = dehydrogenases pertaining to G6P and 6PG, respectively; and R5P = ribulose-5-phosphate). The enzyme activities and metabolites associated with metabolic defense are increased with O 3 exposure. 39'42'93'149'150 Whether this elevation is a protective mechanism against oxidant stress (i.e., the enzymes are synthesized de novo or metabolic pathways are activated for the purpose of defense) is not certain. It appears more likely that this is a response of lung tissue to oxidant damage in which the metabolically active cells (e.g., the alveolar type 2 and the bronchiolar Clara cells) through proliferation contribute to metabolic increases. This is corroborated by the fact that the time-course of metabolic increases coincides with that of morphological changes. 119'12° However, the issue is not entirely resolved since protection against oxidants is afforded by the development of metabolic defense mechanisms. Increased activities of antioxidant enzymes evidently prevent further damage against continued exposure as well as high-level e x p o s u r e . 74'75'149'154"156'159
B. Antioxidant supplements Various natural and synthetic antioxidants have been used as supplements (dietary or otherwise) to prevent and/or decrease O a damage. These are vitamin E (tocopherols), selenium (SE), vitamin C (ascorbate), vitamin A and beta-carotene, p-aminobenzoic acid, phenolic antioxidants (butylated hydroxytoluene, butylated hydroxyanisole), gluatathione and other sulfhydryl compounds. 42,74.75,77,78,84.149.150,158,160--164,169-171,255,279--297 These substances, usually administered as nutritional
Biochemical basis of ozone toxicity
supplements or as therapeutic agents, are known to act as free radical scavengers and/or as antioxidants against nonradical o x i d a t i o n . 3s'42'43'45,46'279-2sl'283'284'298-3°3 It is plausible that these compounds, particularly tocopherols (ArOH), donate a hydrogen to a free radical and stop or contain the chain reaction (reaction 1.34ab). 2s1'3°3'3°4 In that process they may become radicals (ArO) themselves, th,tt is, the antioxidants are oxidized, and then are regenerated or lost from the system. 37'28°'2s1'283 For tocopherols, an in vivo regeneration pathway has been proposed by Tappel, 283 and experimentally supported by others. 281'3°5-31°
HO + ArOH RO2 + ArOH
> >
HOH + ArO R O 2 H+ ArO
(1.34a) (1.34b)
Most typically, vitamin E is administered to experimental animals through dietary regimen. The effects of 0 3 can be elegantly demonstrated in vitamin E-deficient animals when compared with vitamin E-supplemented animals. Various degrees of protection, for example, prevention of massive lung injury or death due to high-level exposures and decrement of biochemical and morphological signs of injury due to low-level exposures, have been o b s e r v e d . 74'75'161'162A69-171'283-288 However, the protective role of vitamin E against oxidant (03) effects is not clearly demonstrated in humans, thus making any extrapolation from animals to man imprecise. There may be several explanations for this discrepancy. Experimental animals can be strictly controlled with dietary regimens, leading to vitamin E deficiency. Humans are not so restricted with diet and made deficient in vitamin E. Thus, a supplemented vitamin E dose in humans may not make any obvious difference in the protection already afforded by the basal dose. A lack of dietary vitamin E does not produce any obvious or acute signs of disease, probably because of the presence of other mechanisms of protection against lipid peroxidation and/or oxidant damage. Furthermore, the physiological and/or biochemical parameters examined in humans may not be sensitive enough to pick up the differences elicited by the basal (or average) dose and the supplemented dose. Also, the absence of any difference may not reflect a lack of vitamin E protective effects. In humans, vitamin E has found a clinical or pharmacologic usage against a variety of diseases, including aging, all of which may have a relationship with free radical r e a c t i o n s . 222'312-315 Vitamin E is thought to have no known human toxicity, although some clinical disorders are claimed to be associated with high doses. 311 It is not unreasonable to assume that a vitamin E supplementation has potential benefits against oxidant air
257
pollution (03), but the question remains on etablishing a judicial dose that can provide an optimal protection. Based on the premise that a vitamin E deficiency may be widespread in humans with symptoms of chronic lung disease, heart disease, or premature aging, and that an exposure to oxidant air pollution ( 0 3) may initiate and/or exacerbate these symptoms, a dietary supplementation of vitamin E may be a prudent means of prophylaxis against chronic free radical damage. In recent years, there is a concern that population exposed to photochemical smog or other environmental oxidants may require increased levels of dietary antioxidants, for example, vitamin E. Human requirements for vitamin E have been reviewed by Horwitt3~6 and Bled and Evarts. 317 The average human diet may contain 7 to 9 IU vitamin E per kg of food. The daily dosage received by the general population may appear marginal, considering the daily recommended dietary allowance of this vitamin which ranges form 4 to 15 IU for infants, children, and adults. 31s Therefore, some investigators have recommended large doses of vitamin E (as much as 1 g or roughly equivalent to 1000 IU daily) and other antioxidants for protection against oxidant effects and other diseases. 31°-312 However, the protection offeted by vitamin E or any of the other antioxidants is not absolute. Even with a high degree of supplementation lung injury occurs due to oxidant exposure. 42'2ss Similarly, Se, which is an integral part of glutathione peroxidase, has been found to influence 0 3 effects. Animals deficient in dietary Se were more susceptible to oxidants than those supplemented with this nutritional element, 163,164,288 and this susceptibility appeared to be due to a decreased function of glutathione peroxidase. 163 Se as selenocysteine and selenomethionine acts as potent water-soluble free radical scavengers. 37 The antioxidant role of Se is also evident from its ability to prevent the symptoms of vitamin E deficiency, such as caused by an oxidant (O 3) s t r e s s . 37'164"269 This is referred to as a sparing action between Se and vitamin E. However, Se and vitamin E do not fully replace each other in their antioxidant roles. 37 Also, Se is toxic in excess doses, and therefore cannot be widely used for protection against oxidant air pollution. 299 Vitamin C plays an antioxidant role against oxidant toxicity. 279-2sl'287'309 It is present in the normal lung, 29°'295 and its level is found to decrease with O 3 exposure. 167'296'297 Supplementation of animals with vitamin C offers protection against acute 03 injury. 29x293 Although vitamin C has the ability to break free radical chain reactions, 2sL3°s'3°9'314 it is not considered a major antioxidant because it can also act as a prooxidant. 2sL319-32° Ascorbate can reduce ferric iron while itself becoming an ascorbyl radical which would form superoxide and OH radicals in an aerobic environment. 283.321-324
258
M.G. MUSTAFA
Glutathione and other sulfhydryl compounds capable of furnishing -SH or -SS- groups have been found to offer a variable degree of protection against oxidant toxicity. 87,28t The exact mode of action is not known but thiyl radicals are thought to occur which may act as free radical scavengers.
2.
3. 4.
CONCLUSION
Ozone produces toxic effects in the lung through oxidation and peroxidation of biomolecules directly and/or by free radical chain reaction. Antioxidants, for example, vitamin E, possibly limit cellular injury by breaking the chain reactions, whereas cellular enzymatic mechanisms, for example, the GSH redox cycle, serve to eliminate the toxic reaction products. Antioxidant nutrition may have the virtue of retarding, if not entirely preventing, oxidant damage. There is abundant documentation of short-term 0 3 effects in the lung, which are characterized by an initial injury, and then a rebounding of metabolic activities, including the level of antioxidant metabolites, for example, GSH and NADPH. The long-term consequence of 0 3 exposure is not clear, and is surmised mostly based on short-term effects. As accelerated deterioration of lung structure and function with chronic exposure appears plausible. The incidence of chronic lung diseases may have multiple causes, but chronic 0 3 exposure remains a contributing factor. Inasmuch as free radical reactions are the underlying cause of 0 3 effects, a variety of diseases (viz., premature aging, chronic lung diseases, including lung tumor incidence) may be linked with chronic 0 3 exposures. Specific experimental findings, for example, chronic exposure causing lung tumor induction or enhancement, might shed light on the long-term risks of 0 3 exposures. There may be a good deal of similarity in the mode of biological reactions of 0 3 and NO2, but the synergistic effects produced by O3-NO 2 interaction is of concern because their combined exposure is the reality in a typical photochemical smog. The toxic chemical intermediates, for example, NO 3 and N20 5, along with the parent gases might expand or deepen cellular injury, resulting in the synergism. Further studies are needed to explain its molecular basis.
5.
6. 7.
8. 9.
10. 11. 12. 13.
14. 15. 16.
17. 18.
19.
20. 21. 22. 23. 24. 25.
Acknowledgements -- The support of U.S. EPA (R-812869) is gratefully appreciated. The author wishes to thank Mr. A.H. Saaduddin for technical assistance, and Mrs. Mary Hunter and Evalon Witt for preparation of the manuscript in a word processor.
26. 27.
REFERENCES 28. 1. National Research Council. Ozone and other photochemical oxidants. Vol. 19. Committee on Medical and Biologic Effects of Environmental Pollutants. Subcommittee on Ozone and Other
29.
Photochemical Oxidants. Washington, DC: National Academy of Sciences; 1977. U.S. Environmental Protection Agency. Air quality criteria for ozone and other photochemical oxidants. EPA Report No. EPA-600/8.79-O04. Research Triangle Park, NC: Environmental Criteria and Assessment Office, U.S. EPA; 1978. World Health Organization. Photochemical oxidants: Environmental health criteria 7. Geneva: WHO; 1979. National Academy of Sciences. Causes and effects of changes in stratospheric ozone: update 1983. Washington, DC: National Academy Press; 1984. U.S. Environmental Protection Agency. Ultraviolet radiation and melanoma with a special focus on assessing the risks of stratospheric ozone depletion. Washington, DC: Office of Air and Radiation, U.S. EPA; 1987. Jones, R.S. Ozone depletion and cancer risk. Lancet 2(No. 8556): 443-445; 1987. Redman, J.C. Stratospheric ozone depletion -- a proposed solution to the problem. Am. J. Dermatopathol. 9:457--458; 1987. Bennett, G. Ozone contamination of high altitude aircraft cabins. Aerosp. Med. 33:969-973; 1962. Jaffe, L.; Estes, H. Ozone toxicity in cabins of high altitude aircraft -- a review and current program. Aerosp. Med. 34: 633-643; 1963. Bischof, W. Ozone measurements on jet airline cabin air. Water Air Soil Pollut. 2:3-14; 1973. Reed, D.; Glaser, S.; Kaldor, J. Ozone toxicity symptoms among flight attendants. Am. J. lndust. Med. i:43-54; 1980. Stokinger, H.E. Ozone toxicology. Arch. Environ. Health 10: 719-731; 1965. Watson, R.T. Atmospheric ozone. In: Titus, J.G., ed. Effects of changes in stratospheric ozone and global climate. Washington, DC: U.S. Environmental Protection Agency; 1986:69-82. National Academy of Sciences. Environmental impact ofstratospheric flight. Washington, DC: National Academy Press; 1975. Heicklen, J. Atmospheric chemistry. New York: Academic Press; 1976. Perner, D. The consequences of increasing CFM concentrations for chemical reactions in the stratosphere and their impact on climate. In: Bach, W.; Pankrath, J.; Kellogg, W., eds. Man's impact on climate. Amsterdam: Elsevier; 1978:215-228. Kondrat'ev, K.Y. Changes in global climate (Russian Translations Series 26). Rotterdam: A.A. Balkema; 1986. Singh, H.B.; Viezee, W.; Johnson, W.B.; Ludwig, F.L. The impact of stratospheric ozone on tropospheric air quality. J. Air Pollut. Control Assoc. 311:1009-1017; 1980. Altshuller, A.P. Estimation of the natural background of ozone present at surface rural locations. J. Air Pollut. Control Assoc. 37:1409-1417; 1987. Haagen-Smit, A.J. Chemistry and physiology of Los Angeles smog. Ind. Eng. Chem. 44:1342-1346; 1952. Haagen-Smit, A.J. Photochemistry and smog. J. Air Pollut. Control Assoc. 13:446-454; 1963. Demerjian, K.L.; Kerr, J.A.; Calvert, J.G. The mechanism of photochemical smog formation. Adv. Environ. Sci. Technol. 4:1-262; 1974. Levy, H.H. Photochemistry of the troposphere. Adv. Photochem. 9:369-524; 1974. Finlayson, B.J.; Pitts, Jr., J.N. Photochemistry of the polluted troposphere. Science 192:111-119; 1976. Logan, J.A.; Prather, M.J.; Wofsy, S.C.; McElroy, M.B. Tropospheric chemistry: a global perspective. J. Geophys. Res. 86:7210-7254; 1981. Whitten, G.Z. The chemistry of smog formation: a review of current knowledge. Environ. Int. 9:447-463; 1983. Atkinson, R.; Lloyd, A.C. Evaluation of kinetic and mechanistic data for modeling of photochemical smog. J. Phys. Chem. Ref. Data. 13:315--444; 1984. Atkinson, R. Kinetic and mechanisms of the gas-phase reactions of the hydroxyl radical with organic compounds under atmospheric conditions. Chem. Rev. 86:69-201; 1986. Atkinson, R.; Carter, W.P.L. Kinetics and mechanisms of the
Biochemical basis of ozone toxicity
30. 31.
32. 33. 34. 35. 36. 37. 38.
39. 40.
41. 42. 43.
44.
45.
46. 47. 48. 49. 50.
51. 52.
gas-phase reactions of ozone with organic compounds under atmospheric conditions. Chem. Rev. 84:437-470; 1984. Nieto, LA.; Salvi, R.P.C.; Gutierrez C. A. Ozone hazard incurred in gamma-plant operation. Health Phys. 42:861-865; 1982. Borek, C.; Mehiman, M.A. Evaluation of health effects toxicity and biochemical mechanisms of ozone. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Publishers; 1983:325-361. Edwards, Jr., E.K. Ozone production by dermatologic phototherapy equipment. Curtis 28:433-434; 1981. South Coast Air Quality Management District. Seasonal and diurnal variation in air quality in California's South Coast Air Basin. E1 Monte, CA: SCAQMD; 1980. South Coast Air Quality Management District. 1982 summary of air quality in California's South Coast Air Basin. E1 Monte, CA: SCAQMD; 1983. Weast, R.C., ed. Handbook of chemistry and physics, 51st ed. Cleveland, OH: The Chemical Rubber Co.; 1970:D-67, D-68, D-112. Nasr, A.N.M. Biochemical aspects of ozone intoxication: a review. J. Occup. Med. 9:589-597; 1967. Menzel, D.B. Toxicity of ozone, oxygen and radiation. Annu. Rev. Pharmacol. 10:379-394; 1970. Menzel, D.B. The role of free radicals in the toxicity of air pollutants (nitrogen oxides and ozone). In: Pryor, W.A., ed. Free radicals in biology. Vol. II. New York: Academic Press; 1976:181-202. Cross, C.E.; Delucia, A.J.; Reddy, A.K.; Hussain, M.Z.; Chow, C.K.; Mustafa, M.G. Ozone interaction with lung tissue: biochemical approaches. Am. J. Med. 60"929-935; 1976. Mudd, J.B.; Freeman, B.A. Reaction of ozone with biological membranes. In: Lee, S.D, ed. Biochemical effects of environmental pollutants. Ann Arbor, MI: Ann Arbor Science Publishers; 1977:97-133. Coffin, D.L.; Stokinger, H.E. Biologic effects of air pollutants. In: Stern, A.C., ed Air pollution. Vol. II. New York: Academic Press; 1977:231-360. Mustafa, M.G.; Tierney, D. F. Biochemical and metabolic changes in the lung with oxygen, ozone, and nitrogen dioxide. Am. Rev. Respir. Dis. 118:1061-1090; 1978. Goldstein, B.D. The pulmonary and extrapulmonary effects of ozone. In: CIBA Foundation symposium: oxygen free radicals and tissue damage. Vol. 65. New York: Elsevier Press; 1979: 295-319. Pryor, W.A.; Dooley, M.M.; Church, D.F. Mechanisms for the reaction of ozone with biological molecules, the source of the toxic effects of ozone. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:7-19. Chow, C.K. Influence of dietary vitamin E on susceptibility to ozone exposure. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:75-93. Menzel, D.B. Ozone: an overview of its toxicity in man and animals. J. Toxicol. Environ. Health 13:183-204; 1984. Mehlman, M.A.; Borek, C. Toxicity and biochemical mechanisms of ozone. Environ. Res. 42:36-53; 1987. Lippmann, M. Health effects of ozone--a critical review. J. Air Waste Manage. Assoc. (JAPCA) 39:672-695; 1989. Brinkman, R.; Lamberts, H.B. Ozone as a possible radiomimetic gas. Nature (London) 181:1202-1203; 1958. Scheel, L.D.; Dobrogorski, D.J.; Mountain, LT.; Svirbely, J.L.; Stokinger, H.E. Physiologic, biochemical, immunologic and pathologic changes following ozone exposure. J. Appl. Physiol. 14:67-80; 1959. Fetner, R.H. Ozone induced chromosome breakage in human cell culture. Nature (London) 194:793-794; 1962. Brinkman, R.; Lamberts, H.B.; Veninga, T.S. Radiomimetic toxicity of ozonized air. Lancet 1(No. 7325):133-136; 1964.
259
53. Veninga, T.S. Toxicity of ozone in comparison with ionizing radiation. Strahlentherapi. 134:469-477; 1967. 54. Zelac, R.E.; Croruroy, H.L.; Bolch, Jr., W.E.; Dunavant, B.G.; Bevis, H.A. Inhaled ozone as a mutagen. I. Chromosome aberrations induced in Chinese hamster lymphocytes. Environ. Res. 4:262-282, 1971. 55. Zelac, R.E., Cromroy, H.L.; Bolch, Jr., W.E.; Dunavant, B.G., Bevis, H.A. Inhaled ozone as a mutagen. H. Effect on the frequency of chromosome aberrations observed in irradiated Chinese hamsters. Environ. Res. 4:325-342; 1971. 56. Koppenol, W.H. The reduction potential of the couple O3/03-, consequences for mechanisms of ozone toxicity. FEBS Lett. 140:169-172; 1982. 57. Pryor, W.A.; Ohto, N.; Church, D.F. Reaction of cumene with ozone to form cumyl hydrotr/oxide and the kinetics of decomposition of cumyl hydrotrioxide, l. Am. Chem. Soc. 105:36143622; 1983. 58. Joigne, J.; Bader, H. The role of hydroxyl radical reactions in ozonation process in aqueous solutions. Water Res. 10:377-386; 1979. 59. Carmichael, N.G.; Winder, C.; Borges, S.H.; Backhouse, B.L.; Lewis, P.D. The health implications of water treatment with ozone. Life Sci. 30:117-129; 1982. 60. Glaze, W.H. Reaction products of ozone: a review. Environ. Health Perspect. 69:151-157; 1986. 61. Goldstein, B.D.; Balchum, O.J.; Demopoulos, H.D.; Duke, P.S. Electron paramagnetic resonance spectroscopy. Free radical signals associated with ozonation of linoleic acid. Arch. Environ. Health 17:46-49; 1968. 62. Pyror, W.A.; Prier, D.G.; Church, D.F. Radical production from the interaction of ozone and PUFA as demonstrated by electron spin resonance spin-trapping techniques. Environ. Res. 24: 42-52, 1981. 63. Pyror, W.A. The role of free radical reactions in biological systems. In: Pryor, W.A., ed. Free radicals in biology. Vol. 1. New York: Academic Press; 1976:1-49. 64. Kurz, M.E.; Pryor, W.A. Radical production from the interaction of closed-shell molecules. 9. Reactions of ozone with tert-butyl hydroperoxide. J. Am. Chem. Soc. 100:7953-7959; 1978. 65. Pryor, W.A. Mechanisms and detection of pathology caused by free radicals. Tobacco smoke, nitrogen dioxide and ozone. In: McKinney, J.D., ed. Environmental health chemistry. Ann Arbor, MI: Ann Arbon Science Publishers; 1981:445-466. 66. Cortesi, R.; Privett, O.S. Toxicity of fatty ozonides and peroxides. Lipids 7:715-721; 1972. 67. Pryor, W.A. The toxicity of oxidized fats: lipid hydroperoxides, their decomposition products and their cooxidation products. In: Taylor, T.G.; Jenkins, N.K., eds. Proceedings of the XIII international congress of nutrition. London: John Libbey and Co.; 1986:589-593. 68. Wu, G.-S.; Milion, T.V.; Subramanian, C.S.; Mnstafa, M.G.; Afifi, A.A.; the late Mead, J.F. Effect of ozone and constituents of rat lung tissue and lavage fluid. Submitted for publication (1990). 69. Goldstein, B.D.; Balchum, O.J. Effect of ozone on lipid peroxidation in the red blood cell. Proc. Soc. Biol. Med. 126:356-358; 1967. 70. Balchum, O.J.; O'Brien, J.S.; Goldstein, B.D. Ozone and unsaturated fatty acids. Arch. Environ. Health 22: 32-34; 1971. 71. Goldstein, B.D. Hydrogen peroxide in erythrocytes. Detection in rats and mice inhaling ozone. Arch. Environ. Health 26:279280; 1973. 72. Goldstein, B.D.; Lodi, C.; Collinson, C.; Balchum, O.J. Ozone and lipid peroxidation. Arch. Environ. Heath 18:631-635; 1969. 73. Sevanian, A.; Mead, J.F.; Stein, R.A. Epoxide products of lipid peroxidation in rat lungs. Lipids 14:634--643; 1979. 74. Chow, C.K.; Tappel, A.L. An enzymatic protective mechanism against lipid peroxidatiun damage to lungs of ozone-exposed rats. Lipids 7: 518-524; 1972. 75. Chow, C.K.; Tappel, A.L. Activities of pentose shunt and glycolytic enzymes in lungs of ozone-exposed rats. Arch. Environ. Health 26:205-208; 1973.
260
M.G. MUSTAFA
76. Sagai, M.; Arakawa, K.; Ichinose, T.; Shimojo, N. Biochemical effects of combined gases of nitrogen dioxide and ozone. I. Species differences of lipid peroxides and phospholipids in lungs. Toxicol. 46:251-265; 1987. 77. Dumelin, E.E.; Dillard, C.J.; Tappel, A.L. Effect of vitamin E and ozone on pentane and ethane expired by rats. Arch. Environ. Health 33:129-135; 1978. 78. Dumelin, E.E.; Dillard, C.J.; Tappel, A.L. Breath ethane and pentane as measures of vitamin E protection of Macaca radiata against 90 days of exposure to ozone. Environ. Res. 15:38-43; 1978. 79. Gelmont, D.; Stein, R.A.; Mead, J.F. The bacterial origin of rat breath pentane. Biochem. Biophys. Res. Commun. 102:932-936; 1981. 80. Mustafa, J.G. General enzymology of the lung. In: Witschi, H.P.; Brain, J.D., eds. Toxicology of inhaled materials. Berlin: Springer-Verlag; 1985;369-419. 81. Menzel, D.B. Oxidation of biologically active reducing substances by ozone. Arch. Environ. Heath 23:149-153; 1971. 82. DeLucia, A.J.; Hoque, P.M.; Mustafa, M.G.; Cross, C.E. Ozone interaction with rodent lung. Effect on sulfhydryl and sulfhydryl-containing enzyme activities. J. Lab. Clin. Med. 80"559-566; 1972. 83. DeLucia, A.J.; Mustafa, M.G.; Hussaln, M.Z.; Cross, C.E. Ozone interaction with rodent lung. III. Oxidation of reduced glutathione and mixed disulfide formation between protein and nonprotein sulfhydryls. J. Clin. Invest. 55:794-802; 1975. 84. Fairchild, II, E.J.; Murphy, S.D.; Stokinger, H.E. Protection by sulfur compounds against the air pollutants ozone and nitrogen dioxide. Science 130:861-862; 1959. 85. Mountain, J.T. Detecting hypersensitivity to toxic substances. Arch. Environ. Health 6:357-365; 1963. 86. Freeman, B.A.; Mudd, J.B. Reaction of ozone with sulfhydryls of human erythrocytes. Arch. Biochem. Biophys. 208:212-220; 1981. 87. Mudd, J.B.; Leavitt, R.; Ongun, A.; McManus, T.T. Reaction of ozone with amino acids and proteins. Atmos. Environ. 3:669-681; 1969. 88. Hurst, D.J.; Coffin, D.L. Ozone effect on lysosomal hydrolases of alveolar macrophages in vitro. Arch. Intern. Med. 127" 1059-1063; 1971. 89. Palmer, M.S.; Swanson, D.H.; Coffin, D. L. Effect of ozone on benzpyrene hydroxylase activity in the Syrian golden hamster. Cancer Res. 31:730-733; 1971. 90. Palmer, M.S.; Exley, R.W.; Coffin, D.L. Influence of pollutant gases on benzpyrene hydroxylase activity. Arch. Environ. Health 25:439--442; 1972. 91. Mustafa, M.G.; DeLucia, A.J.; York, G.K.; Arth, C.; Cross, C.E. Ozone interaction with rodent lung. II. Effects on oxygen consumption of mitochondria. J. Lab. Clin. Med. 82:357-365; 1973. 92. Mustafa, M.G.; Cross, C.E. Effects of short-term ozone exposure on lung mitochondrial oxidative and energy metabolism. Arch. Biochem. Biophys. 162:585-594; 1974. 93. Mustafa, M.G. Augmentation of mitochondrial oxidative metabolism in lung tissue during recovery from acute ozone exposure. Arch. Biochem. Biophys. 165:531-538; 1974. 94. Goldstein, B.D.; Solomon, S.; Pasternack, B.S.; Bickers, D.R. Decrease in rabbit lung microsomal cytochrome P-450 levels following ozone exposure. Res. Commun. Chem. Pathol. Pharmacol. 10:759-762; 1975. 95. Montgomery, M.R.; Niewoehner, D.E. Oxidant-induced alterations in pulmonary microsomal mixed-function oxidation: acute effects of paraquat and ozone. J. Environ. Sci. Health C13: 205-219; 1979. 96. Menzel, D.B.; Anderson, W.G.; Abou-Donia, M.B. Ozone exposure modifies prostaglandin biosynthesis in perfused rat lungs. Res. Commun. Chem. Pathol, Pharmacol. 15:135-147; 1976. 97. Madden, M.C.; Eling, T.E.; Friedman, M. Ozone inhibits endothelial cell cyclooxigenase activity through formation of hydrogen peroxide. Prostagland. 34:445-463; 1987. 98. Gordon, T.; Taylor, B.F.; Amdur, M.O. Ozone inhibition of
tissue cholinesterase in guinea-pigs. Arch. Environ. Health 36:284-288; 1981. 99. Johnson, D.A. Ozone inactivation of human alpha-l-antiproo teinase inhibitor. Am. Rev. Respir. Dis. 121"1031-1038; 1980. 100. Koontz, A.E.; Heath, R.L. Ozone alteration of transport of cations and the Na/K ATPase in human erythrocytes. Arch. Biochem. Biophys. 198.493-500; 1979. 101. Witz, G.; Amoruso, M.A.; Goldstein, B.D. Effect of ozone on alveolar macrophage function: membrane dynamic properties. In: Lee S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Pub-
lishers; 1983:263-272. 102. Rietjens, I.M.C.M.; van Tilburg, C.A.M.; Coenen, T.M.M.; Alink, G.M.; Konings, A.W.T. Influence of polyunsaturated fatty acid supplementation and membrane fluidity on ozone and nitrogen dioxide sensitivity of rat alveolar macrophages. J. Toxicol. Environ. Health 21:45-56; 1987. 103. Alpert, S.M.; Schwartz, B.B.; Lee, S.D.; Lewis, T.R. Alveolar protein accumulation: a sensitive indicator of low level oxidant toxicity. Arch. Intern. Med. 128:69-73; 1971. 104. Reasor, M.J.; Adams, G.K.; Brooks, J.K.; Rubin, R.J. Enrichment of albumin and IgG in the airway secretions of dogs breathing ozone. J. Environ. Sci. Health C13:335-346; 1979. 105. Williams, S.J.; Charles, J.M.; Menzel, D.B. Ozone-induced alterations in phenol red absorption from the rat lung. Toxicol. Lett. 6:213-219; 1980. 106. Hu, P.C.; Miller, F.J.; Daniels, M.J.; Hatch, G.E.; Graham, J.A.; Gardner, D.E.; Selgrade, M.K. Protein accumulation in lung lavage fluid following ozone exposure. Environ. Res. 29:377-388; 1982. 107. Bhalla, D.K.; Crocker, T.T. Tracheal permeability in rats exposed to ozone. Am. Rev. Respir. Dis. 134:572-579; 1986. 108. Bhalla, D.K.; Mannix, R.C.; Kleinman, M.T.; Crocker, T.T. Relative permeability of nasal, tracheal, and bronchoalveolar mucosa to macromolecules in rats exposed to ozone. J. Toxicol. Environ. Health 17:269-283; 1986. 109. Kehrl, H.R.; Vincent, L.M.; Kowalsky, R.J.; Horstman,D.H.; O'Neil, J.J.; McCartney, W.H.; Bromberg, P.A. Ozone exposure increases respiratory epithelial permeability in humans. Am. Rev. Respir. Dis. 135:1124-1128; 1987. 110. Seltzer, J.; Bigby, B.G.; Stulbarg, M.; Holtzman, M.J.; Nadel, J.A.; Ueki, I.F.; Leikauf, G.D.; Goetzel, E.J.; Boushey, H.A. O3-induced change in bronchial reactivity to methacholine and airway inflammation in humans. J. Appl. Physiol. 60:13211326; 1986. 111. Koren, H.S.; Devlin, R.B.; Graham, D.E..; Mann, R.; McGee, M.P.; Horstman, D.H.; Kozumbo, W.J.; Becker, S.; House, D.E.; McDonnell, W.F.; Bromberg, P.A. Ozone-induced inflammation in the lower airways of human subjects. Am. Rev. Respir. Dis. 139:407-415; 1989. 112. Ford-Hutichinson, A.W.; Bray, M.A.; Doig, M.V.; Shipley, M.E.; Smith, M.J.H. Leukotriene b, a potent chemokinetic and aggregating substance released from polymorphonuclear leukocytes. Nature 286:264-265; 1980. 113. Dixon, J.R.; Mountain, J.T. Role of histamine and related substances in the development of tolerance to edemagenic gases. Toxicol Appl. Pharmacol. 7:756-766; 1965. 114. Fairchild, II, E.J. Neurohormonal factors in injury from inhaled irritants. Arch. Environ. Health 6:85-92; 1963. 115. Fairchild, II, E.J.; Graham, S.L. Thyroid influence on the toxicity of respiratory irritant gases, ozone and nitrogen dioxide. J. Pharmacol. Exp. Ther. 139:177-184; 1963. 116. Fairchild, II, E.J.; Graham, S.L.; Hite, M.; Killens, R.; Scheel, L.D. Changes in thyroid I TM activity in ozone-tolerant and ozone-susceptible rats. Toxicol Appl. Pharmacol. 6:607-613; 1964. 117. Yokoyama, E.; Frank, R. Respiratory uptake of ozone in dogs. Arch. Environ. Health 25:132-138; 1972. 118. Miller, F.J. Biomathematical modeling applications in the evaluation of ozone toxicity. In: Mudd, J.B.; Lee, S.D., eds. Assessing toxic effects of environmental pollutants. Ann Arbor, MI: Ann Arbor Science Publishers; 1979:263-286.
Biochemical basis of ozone toxicity 119. Dungworth, D.L.; Castleman, W.L.; Chow, C.K.; Me[lick, P.W.; Mustafa, M.G.; Tarkington, B.; Tyler, W.S. Effects of ambient levels of ozone on monkeys. Fed. Proc. 34:1670-1674; 1975. 120. Schwartz, W.L.; Dungworth, D.L.; Mustafa, M.G.; Tarkington, B.K.; Tyler, W.S. Pulmonary response of rats to ambient levels of ozones: effects of 7-day intermittent or continuous exposure. Lab. Invest. 34:565-578; 1976. 121. Cavender, F.L.; Steinhagen, W.H.; Ulrich, C.E.; Busey, W.M.; CockreU, B.Y.; Haseman, J.K.; Hogan, M.D.; Drew, R.T. Effects in rats and guinea-pigs of short-term exposures to sulfuric acid mist, ozone, and their combination. J. Toxicol. Environ. Health 3:521-533; 1977. 122. Castleman, W.L.; Tyler, W.S.; Dungworth, D.L. Lesions in respiratory bronchioles and conducting airways of monkeys exposed to ambient levels of ozone. Exp. Mol. Pathol. 26: 3~os. A.90; 1977. 123. Mellick, P.W.; Dungworth, D.L.; Schwartz, L.W.; Tyler, W.S. Short-term morphologic effects of high ambient levels of ozone on lungs of rhesus monkeys. Lab. Invest. 36:82-90; 1977. 124. Ibrahim, A.L.; Zee, Y.C.; Osebold, J.W. The effects of ozone on the respiratory epithelium of mice. II. Ultrastructural alterations. J. Environ. Pathol. Toxicol. 3:251-258; 1980. 125. Wilson, D.W.; Plopper, C.G.; Dungworth, D.L. The response of the macaque ffacheobronchial epithelium to acute ozone injury. A quantitative ultrastructural and autoradiographic study. Am. J. Pathol. 116:193-206; 1984. 126. Eustis, S.L.; Schwartz, L.W.; Kosch, P.C.; Dungworth, D.L. Chronic bronchiolitis in nonhuman primates after prolonged ozone exposure. Am. J. Pathol. 105:121-137; 1981. 127. Yokoyama, E.; Ichikawa, I.; Kawal, K. Does nitrogen dioxide modify the respiratory effects of ozone? In: Lee, S.D., ed. Nitrogen oxides and their effects on health. Ann Arbor, MI: Ann Arbor Science Publishers; 1980:217-229. 128. Stephens, R.J.; Sloan, M.F.; Evans, M.J.; Freeman, G. Early response of lung to low levels of ozone. Am. J. Pathol. 73:711-726; 1974. 129. Stephens, R.J.; Sloan, M.F.; Evans, M.J.; Freeman, G. Alveolar type 1 cell response to exposure to 0.5 ppm 03 for short periods. Exp. Mol. Pathol. 20:11-23; 1974. 130. Evans, M.J.; Johnson, L.V.; Stephens, R.J.; Freeman, G. Cell renewal in the lungs of rats exposed to low levels of ozone. Exp. Mol. Pathol. 24:70--83; 1976. 131. Lure, H.; Schwartz, L.W.; Dungworth, D.L.; Tyler, W.S. A comparative study of cell renewal after exposure to ozone or oxygen: response of terminal broncbiolar epithelium in the rat. Am. Rev. Respir. Dis. 118:355-345; 1978. 132. Boorman, G.A.; Schwartz, L. W.; Dungworth, D.L. Pulmonary effects of prolonged ozone insult in rats. Morphometric evaluation of the central acinus. Lab. Invest. 43:108-115; 1980. 133. Barry, B.E.; Miller, F.J.; Crapo, LD. Alveolar epithelial injury caused by inhalation of 0.25 ppm ozone. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:299-309. 134. Sherwin, R.P.; Richters, V.; Okimoto, D. Type 2 pneumocyte hyperplasia in the lungs of mice exposed to an ambient level (0.3 ppm) of ozone. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:289-297. 135. Castelman, W.L.; Dungworth, D.L.; Tyler, W.S. Cytochemically detected alterations of lung acid phosphatase reactivity following ozone exposure. Lab. Invest. 29:310-319; 1973. 136. Castelman, W.L.; Dungworth, D.L.; Tyler, W.S. Histochemically detected enzymatic alterations in rat lung exposed to ozone. Exp. Mol. Pathol. 19:402-421; 1973. 137. Brummer, M.E.G.; Schwartz, L.W.; McQuillen, N.K. A quan-. titative study of lung damage by scanning electron microscopy: inflammatory cell response to high-ambient levels of ozone. Scanning Electron Microsc. 2:513-518; 1977. 138. Brain, J.D. Macrophage damage in relation to the pathogenesis of lung diseases. Environ. Health Perspect. 35:21-28; 1980.
261
139. Oomichi, S.; Kits, H. Effect of air pollutants on ciliary activity of respiratory tract. Bull. Tokyo Med. Dent. Univ. 21:327-343; 1974. 140. Frager, N.B.; Phalen, R.F.; Kenoyer, J.L. Adaptations to ozone in reference to mucociliary clearance. Arch. Environ. Health 34:51-57; 1979. 141. Kenoyer, J.L.; Phalen, R.F.; Davis, J.R. Particle clearance from the respiratory tract as a test of toxicity: effect of ozone on short and long term clearance. Exp. Lung Res. 2:111-120; 1981. 142. Schlesinger, R.B.; Driscoll, K.E. Mucociliary clearance from the lungs of rabbits following single and intermittent exposure to ozone. J. Toxicol. Environ. Health 20:125-134; 1987. 143. Abraham, W.M.; Januskiewicz, A.J.; Mingle, M.; Wekler, M.; Wanner, A.; Sackner, M.A. Sensitivity of bronchoprovocatiun and tracheal mucous velocity in detecting airway responses to 03. J. Appl. Physiol. 48:789-793; 1980. 144. Phipps. R.J.; Denas, S.M.; Sielczak, M.W.; Wanner, A. Effects of 0.5 ppm ozone on glycoprotein secretion, ion and water fluxes in sheep trachea. J. Appl. Physiol. 60:918-927; 1986. 145. Foster, W.M.; Costa, D.L.; Langenback, E.G. Ozone exposure alters tracheobronchial mucociliary function in humans. J. Appl. Physiol. 63:996-1002; 1987. 146. Coffin, D.L.; Gardner, D.E. Interaction of biological agents and chemical air pollutants. Ann. Occup. Hyg. 15:219--234; 1972. 147. Miller, F.J.; Illing, J.W.; Gardner, D.E. Effects of urban ozone levels on lahoratory-induced respiratory infections. Toxicol. Len. 2:163-169; 1978. 148. Driscoll, K.E.; Vollmuth,T.A.; Schlocsinger, R.B. Early alveolar clearance of particles in rabbits undergoing acute and subchronic exposure to ozone. Fundam. Appl. Toxicol. 7: 264-271; 1986. 149. Mustafa, M.G.; Lee, S.D. Pulmonary biochemical alterations resulting from ozone exposure. Ann. Occup. Hyg. 19:17-26; 1976. 150. Mustafa, M.G.; Hacker, A.D.; Ospital, J.J.; Hnssain, M.Z.; Lee, S.D. Biochemical effects of environmental oxidant pollutants in animal lungs. In: Lee, S.D., ed. Biochemical effects of environmental pollutants. Ann Arbor, NII: Ann Arbor Science Publishers; 1977:59-96. 151. Mustafa, M.G.; Elsayed, N.M.; Quinn, C.L.; Postlethwalt, E.M.; Gardner, D.E.; Graham, LA. Comparison of pulmonary biochemical effects of low level ozone exposure on mice and rats. J. Toxicol. Environ. Health 9:857-865; 1982. 152. Elsayed, N.M.; Mustafa, M.G.; Postlethwalt, E.M. Age-dependent pulmonary response of rats to ozone exposure. J. Toxicol. Environ. Health 9:835-848; 1982. 153. Mustafa, M.G.; Elsayed, N.M.; Graham, J.A.; Gardner, D.E. Effects of ozone exposure on lung metabolism: influence of animal age, species, and exposure conditions. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A. eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983: 57-73. 154. Chow, C.K.; Hussaln, M.Z.; Cross, C.E.; Dungworth, D.L.; Mustafa, M.G. Effects of low levels of ozone on lung. I. Biochemical responses during recovery and reexposure. Exp. Mol. Pathol. 25:182-188; 1976. 155. Hussain, M.Z.; Cross, C.E.; Mustafa, M.G.; Bhatnagar, R.S. Hydroxyproline contents and prolyl hydroxylase activities in lungs of rats exposed to low levels of ozone. Life Sci. 15: 897-904; 1976. 156. Plopper, C.G.; Chow, C.K.; Dungworth, D.L.; Brummer, M.; Nemeth, T.J. Effect of low level of ozone on rat lungs. II. Morphological responses during recovery and reexposure. Exp. Mol. Pathol. 29:4(X)-411; 1978. 157. Plopper, C.G.; Chow, C.K.; Dungworth, D.L.; Tyler, W.S. Pulmonary alterations in rats exposed to 0.2 and 0.1 ppm ozone: a correlated morphological and biochemical study. Arch. Environ. Health 34:390-395; 1979. 158. Chow, C.K.; Dillard, C.J.; Tappel, A.L. Gluthione peroxidas¢ system and lysozyme in rats exposed to ozone or nitrogen dioxide. Environ. Res. 7:311-319; 1974. 159. Chow, C.K.; Mustafa, M.G.; Cross, C.E.; Tarkington, B.K.
262
160. 161.
162.
163.
164.
165.
166.
167. 168. 169. 170. 171. 172.
173. 174.
175.
176. 177. 178. 179.
M.G. MuSTAFA Effects of ozone exposure on the lungs and the erythrocytes of rats and monkeys: relative biochemical changes. Environ. Physiol. Biochem. 5:142-146; 1975. Chow, C.K.; Kaneko, J.J. Influence of dietary vitamin E on the red cells of ozone-exposed rats. Environ. Res. 19:49-55; 1979. Chow, C.K.; Plopper, C.G.; Dungworth, D.L. Influence of dietary vitamin E on the lungs of ozone-exposed rats: a correlated biochemical and morphological study. Environ. Res. 20:309317; 1979. Chow, C.K.; Plopper, C.G.; Chin, M.; Dungworth, D.L. Dietary vitamin E and pulmonary biochemical and morphological alterations of rats exposed to 0.1 ppm ozone. Environ. Res. 24:315-324; 1981. Elsayed, N.M.; Hacker, A.; Mustafa, M.; Kuehn, K.; Schrauzer, K. Effects of decreased glutathione peroxidase activity on the pentose phosphate cycle in mouse lung. Biochem, Biophys. Res. Commun. 104:564-569; 1982. Elsayed, N.M.; Hacker, A.D.; Kuehn, K.; Mustafa, M.G.; Schrauzer, G.N. Dietary antioxidants and the biochemical response to oxidant inhalation. II. Influence of dietary selenium on the biochemical effects of ozone exposure in mouse lung. Toxicol. Appl. Pharmacol. 71:398--406; 1983. Lunan, K.D.; Short, P.; Negi, D.; Stephens, R.J. Glucose6-phosphate dehydrogenase response of postnatal lungs to NO 2 and 03. In: Sanders, C.L.; Schneider, R.P.; Dagle, G.E.; Ragan, H.A., eds. Pulmonary macrophages and epithelial cells. Washington, DC: Energy Research and Development Administration (ERDA symposium series: 43): 1977;236-247. Moore, G.S.; Calabrese, E.J.; Grinberg-Funes, R.A. The C57L/J mouse strain as a model for extrapulmonary effects of ozone exposure. Bull. Environ. Contain. Toxicol. 25:578-585; 1980. Fukase, O.; Isomura, K.; Watanabe, H. Effect of ozone on glutathione in vivo. Taiki Osen Kenkyu 10:58-62; 1975. Tyson, C.A.; Lunan, K.D.; Stephens, R.J. Age-related differences in GSH-shuttle enzymes in NOz or O3 exposed rat lungs. Arch. Environ. Health 37:167-176; 1982. Roehm, J.N.; Hadley, J.G.; Menzel, D.B. Antioxidant vs. lung disease. Arch. Intern. Med. 128:88-93; 1971. Roehm, J.N.; Hadley, J.G.; Menzel, D.B. The influence of vitamin E on the lung fatty acids of rats exposed to ozone. Arch. Environ. Health 24:237-242; 1972. Menzel, D.B.; Roehm, J.N.; Lee, S.D. Vitamin E: the biological and environmental antioxidant. J. Agric. Food Chem. 20:481486; 1972. Shimasaki, H.T.; Takatori, W.R.; Anderson, H.L.; Shimasaki, H.; Takatori, T.; Anderson, W.R.; Horten, H.L.; Privett, O.S. Alteration of lung lipids in ozone exposed rats. Biochem. Biophys. Res. Commun. 68:1256-1262; 1976. Huber, G.L.; Mason, R.J.; LaForce, M.; Spencer, N.J.; Gardner, D.E.; Coffin, D.L. Alterations in the lung following the administration of ozone. Arch. Intern. Med. 128:81-87; 1971. Myers, A.B.; Dubick, M.A.; Reiser, K.M.; Gerriets, J.E.; Last, J.A.; Rucker, R.B. Ozone exposure, food restriction and protein deficiency: changes in collagen and elastin in rodent lung. Toxicol. Lett. 23: 43-49; 1984. Bhatnagar, R.S.; Hussain, M.Z.; Sorensen, K.R.; Mustafa, M.G.; yon Dohlen, F.M.; Lee, S.D. Effect of ozone on lung collagen biosynthesis. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:311-321. Last, J.A.; Kaizu, T. Mucous glycoprotein secretion by tracheal explants: effects of pollutants. Environ. Health Perspect. 35: 131-138; 1980. Last, J.A.; Greenberg, D.B.; Castleman, W.L. Ozone-induced alterations in collagen metabolism of rat lungs. Toxicol. Appl. Pharmacol. 51:247-258; 1979. Last, J.A.; Greenberg, D.B. Ozone-induced alterations in collagen metabolism of rat lungs. II. Long-term exposure. Toxicol. Appl. Pharmacol. 55:108-114; 1980. Costa, D.L.; Kutzman, R.S.; Lehrnann, J.R.; Popenoe, E.A.; Drew, R.T.A. Subchronic multi-dose ozone study in rats. In:
180. 181. 182. 183. 184.
185.
186. 187.
188. 189.
190.
191.
192. 193.
194.
195.
196, 197. 198.
Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983:369-393. Last, J.A.; Reiser, K.M.; Tyler, W.S.; Rucker, R.B. Long-term consequences of exposure to ozone. I. Lung collagen content. Toxicol. Appl. Pharmacol. 72:111-118; 1984. Hacker, A.D.; Mustafa, M.G.; Ospital, J.J.; Elsayed, N.M.; Lee, S.D. Relationship of age to rat lung collagen synthesis with ozone exposure. Age 9:1-5; 1986. Hesterberg, T.W.; Last, J.A. Ozone-induced acute pulmonary fibrosis in rats: prevention of increased rates of collagen synthesis by methyl prednisolone. Am. Rev. Respir. Dis. 123:47-52; 1981. Last, J.A.; Jennings, M,D.; Schwartz, L.W.; Cross, C.E. Glycoprotein secretion by tracheal explants cultured from rats exposed to ozone. Am. Rev. Respir. Dis. 116:695-703; 1977. Last, J.A.; Cross, C.E. A new model for health effects of air pollutants: evidence for synergistic effects of mixtures of ozone and sulfuric acid aerosols on rat lungs. J, Lab. Clin. Med. 91:328-339; 1978. Dillard, C.J.; Urribarri, C.N.; Reddy, K.; Fletcher. B.; Taylor, S.; de Lumen, B.; Tappel, A.L. Increased lysosomal enzymes in lungs of ozone-exposed rats. Arch. Environ. Health 25:426--431; 1972. Evans, M.J.; Dekker, N.P.; Cabral-Anderson, L.J.; Shami, S.G. Morphological basis of tolerance to ozone. Exp. Mol. Pathol. 42: 366-376; 1985. Last, J.A.; Gerriets, J.E.; Hyde, D.M. Synergistic effects on rat lungs of mixtures of oxidant air pollutants (ozone or nitrogen dioxide) and respirable aerosols. Am. Rev. Respir. Dis. 128: 539-544; 1983. Last, J.A.; Hyde, D.M.; Chang, D.P.Y. A mechanism of synergistic lung damage by ozone and a respirable aerosol. Exp. Lung Res. 7:223-235; 1984. Hackney, J.D.; Linn, W.S.; Mohler, J.G.; Pederson, E.E,; Breisacher, P.; Russo, A. Experimental studies on human health effects of air pollutants. II. Four-hour exposure to ozone alone and in combination with other pollutant gases. Arch. Environ. Health 30:379-384; 1975. Hackney, J.D.; Linn, W.S.; Law, D.C.; Karuza, S.K.; Greenberg, H.; Buckley, R.D.; Pederson, E.E. Experimental studies on human health effects of air pollutants. III. Two-hour exposure to ozone alone and in combination with other pollutant gases. Arch. Environ. Health 30:385-390; 1975. von Neiding, G.; Wagner, H.M.; Krekeler, H.; LoUgen, H.; Fries, W.; Beuthan, A. Controlled studies of human exposure to single and combined action of NO 2, O3, and SO2. Int. Arch. Occup. Environ. Health 43:195-210; 1979. Folinsbee, L.J.; Bedi, J.F.; Horvath, S.M. Combined effects of ozone and nitrogen dioxide on respiratory function in man. Am. Ind. Hyg. Assoc. J. 42" 534-541; 1981. Toyama, T.; Tsunoda, T.; Nakaza, M.; Higashi, T.; Nakadate, T. Airway response to short-term inhalation of NO 2, 03, and their mixture in healthy men. Sangyo lgaku (Japan J. Ind. Hyg.) 23:285-293; 1981. Kagawa, J.; Tsuru, K. Respiratory effects of 2-hour exposure to ozone and nitrogen dioxide alone and in combination in normal subject performing intermittent exercise. Nippon Kyobu Shikkan Gakkai Zasshi (Japan J. Thorac. Dis.) 17:765-774; 1979. Freeman, G.; Jnhos, L.T.; Furiosi, N.J.; Mussenden, R.; Stephens, R.J.; Evans, M.J. Pathology of pulmonary disease from exposure to interdependent ambient gases (nitrogen dioxide and ozone). Arch. Environ. Health 29:203-210; 1974. Goldstein, E.; Warshauer, D.; Lippert, W.; Tarkington, B. Ozone and nitrogen dioxide exposure: murine pulmonary defense mechanisms. Arch. Environ. Health 28:85-90; 1974. Ehrlich, R.; Findlay, J.C,; Fenters, J.D.; Gardner, D.E. Health effects of short-term inhalation of nitrogen dioxide and ozone mixtures. Environ. Res. 14:223-231; 1977. Ehrlich, R.; Findlay, J.C.; Gardner, D.E. Effects of repeated exposures to peak concentrations of nitrogen dioxide and ozone on resistance to streptococcal pneumonia. J. Toxicol. Environ. Health 5:631--642; 1979.
Biochemical basis of ozone toxicity 199. Ehrlich, R. Interaction between environmental pollutants and respiratory infections. Environ. Health Perspect. 35:89-100; 1980. 200. Ehrlich, R. Changes in susceptibility to respiratory infection caused by exposures to photochemical oxidant pollutants. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biochemical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific PUbfishers; 1983:273-285. 201. Graham, J.A.; Gardner, D.E.; Blommer, E.J.; Honsem, D.E.; Menache, M.G.; Miller, F.J. Influence of exposure patterns of nitrogen dioxide and modifications by ozone on susceptibility to bacterial infectious disease in mice. J. Toxicol. Environ. Health 21:113-125; 1987. 202. Goldstein, B.D. Combined exposure to ozone and nitrogen dioxide. Environ. Health Perspect. 13:107-110; 1976. 203. Mnstafa, M.G.; Elsayed, N.M.; yon Dohien, F.M.; Hasset, C.M.; Postlethwait, E.M.; Quinn, C.L.; Graham, J.A.; Gardner, D.E. A comparison of biochemical effects of nitrogen dioxide, ozone, and their combination in mouse lung. I. Intermittant exposure. Toxicol. Appl. Pharmacol. 72:82-90; 1984. 204. Mnstafa, M.G.; Elsayed, N.M.; Lee, S.D. The combined effects of ozone and nitrogen dioxide on pulmonary biochemistry. In: Lee, S.D., ed. Evaluation of the scientific basis for ozone/ oxidants standards. Pittsburgh, PA: Air Pollution Control Association; 1985:337-347. 205. Mantz, W.J.; Kleinman, M.T.; Phalen, R.F.; Crocker, T.T. Effects of exercise I exposure on toxic interactions between inhaled oxidant and aldehyde air pollutants. J. Toxicol. Environ. Health 25:165-177; 1988. 206. Lee, J.-S.; Afifi, A.A.; Mustafa, M.G. Effects of short-term, single and combined exposure of rats to NO2 and 03 on lung tissue enzyme activities, lnhal. Toxicol. 1:21-35; 1989. 207. Lee, J.-S.; Mustafa, M.G.; Afifi, A.A. Effects of short-term, single and combined exposure to low-level NO2 and 03 on lung tissue enzyme activities in rats. J. Toxicol. Environ. Health 29:293-305; 1990. 208. Goldstein, B.D. Combined exposure to ozone and nitrogen dioxide. Environ. Health Perspect. 30:87-89; 1979. 209. Diggie, W.M.; Gage, J.C. The toxicity of ozone in the presence of oxides of nitrogen. Brit. J. lndustr. Med. 12:60-64; 1955. 210. Pins, Jr., J.N. Formation and fate of gaseous and particulate mutagens and carcinogens in real and simulated atmospheres. Environ. Health Perspect. 47:115-140; 1983. 211. piatt, U.F.; Wirier, A.M.; Blerman, H.W.; Atkinson, R.; Pitts, Jr., LN. Measurement of nitrate radical concentrations in continental air. Environ. Sci. Technol. 16:365-369; 1984. 212. Postlethwait, E.M.; Mustafa, M.G. The fate of inhaled nitrogen dioxide in the isolated perfused rat lung. J. Toxicol. Environ. Health 7:861-872; 1981. 213. Postlethwait, E.M.; Mustafa, M.G. Effect of altered dose rate on NO2 uptake and transformatiion in isolated lungs. J. Toxicol. Environ. Health 26:497-507; 1989. 214. Oda, H.; Nogami, H.; Nakajima, T. Reaction of hemoglobin with nitric oxide and nitrogen dioxide in mice. J. Toxicol. Environ. Health 6:673-678; 1980. 215. Chiodi, H.; Collier, C.R.; Mothler, J.G. In vitro methemogiobin formation in human blood exposed to NO 2. Environ. ICes. 30:9-15; 1983. 216. Kosaka, H.; Tyuma, I. Mechanism of autocatalytic oxidation of oxyhemoglobin by nitrite. Environ. Health Perspect. 7:147-151; 1987. 217. Klimmek, R.; Krettek, C.; Werner, H.W. Ferrihemoglobin formation by amyl nitrite and sodium nitrite in different species in vivo and in vitro. Arch. Toxicol. 62:152-160; 1988. 218. Stokinger, H.E.; Wagner, W.D.; Dobrogorski, O.J. Ozone toxicity studies. HI. Chronic effects. Arch. lndast. Health 16:514-522; 1957. 219. Melton, C.E. Effects of long-term exposure to low levels of ozone: a review. Aviat. Space Environ. Med. $3:105-111; 1982. 220. Filipowicz, C.; McCauley, R. The effect of chronic ozone exposure on lung benzo(a)pyrene oxidase and monoamine oxidase. Res. Commun. Chem. Pathol. Phormacol. $1:289-296;
263
1986. 221. Gross, K.B.; White, H.J. Functional and pathologic consequences of a 52-week exposure to 0.5 ppm ozone followed by a clean air recovery period. Lung 165:283-295; 1987. 222. Harman, D. Free radical theory of aging: effect of the amount and degree of unsaturation of dietary fat on mortality rate. J. Gerontol. 26:451-457; 1971. 223. Libovitz, B.E.; Siegel, B.V. Aspects of free radical reactions in biological systems and aging. J. GerontoL 35:45-56; 1980. 224. Ts'o, P.O.P.; Caspary, W.J.; Loventzen, R.J. The involvement of free radicals in chemical carcinogenesis. In: Pryor, W.A., ed. Free radicals in biology. Vol. 3. New York: Academic Press; 1977:251-303. 225. Demopoulos, H.B.; Peitronigro, E.S.; Flamm, E.S.; Seligman, M.L. The possible role of free radical reactions in carcinogenesis. In: Demopoulos, H.B.; Mehiman, M.A., eds. Cancer and the environment. Park Forest South, IL: Pathotox Publishers; 1980:273-303. 226. Totter, J.R. Spontaneous cancer and its possible relationship to oxygen metabolism. Proc. Natl. Acad. Sci. (USA) 77:17631767; 1980. 227. Troll, W.; Witz, G.; Goldstein, B.; Stone, D.; Sugimura, T. The role of free oxygen radical in tumor pormotion and carcinogenesis. In: Hecker, E.; Fusenig, N.E.; Kunz, W.; Marks, F.; Thielman, H.W., eds. Carcinogenesis and biological effects of tumor promoters. New York: Raven Press; 1982:593-597. 228. Chu, E.H.Y.; Trosko, J.E.; Chang, C. Mutational approaches to the study of carcinogenesis. J. Tox. Environ. Health 2:13171334; 1977. 229. Diamond, L.; O'Brien, T.G.; Ballrd, W.M. Tumor promoters and the mechanism of tumor promotion. Adv. Cancer Res. 32:1-74; 1980. 230. van Duuren, B.L. Tumor promoting and co-carcinogenic agents chemical carcinogenesis. In: Searle, C.E., ed. Chemical carcinogens. Washington, DC: Amercian Chemical Society; 1976: 24-51. 231. O'Brien, T.G.; Silmsiman, R.C.; Boutwell, R.K. Induction of the polyamine-biosynthetic enzymes in mouse epidermis and their specificity for tumor promotion. Cancer Res. 35:24262433; 1975. 232. Boutwell, R.K. Biochemical mechanism of tumor promotion. In: Slaga, T.J.; Sivak, A.; Boutwell, R.K., eds. Carcinogenesis, Vol 2. Mechanisms of tumor promotion and cocarcinogenesis. New York: Raven Press; 1978:49-58. 233. Hamelin, C.; Chung, Y.S. The effect of low concentrations of ozone on Escherichia coli chromosome. Murat. Res. 28:131132; 1975. 234. Marz, T.; Bender, M.; Kerr, H.; guile, T. Observations of aberrations in chromosomes of lymphocytes from human subjects exposed to ozone at concentrations of 0.5 ppm for 6 and 10 hours. Murat. Res. 31:299-302; 1975. 235. Guerrero, R.R.; Rounds, D.E.; Olson, R.S.; Hackney, J.D. Mutagenic effects of ozone on human cells exposed in vivo and in vitro based on sister chromatid exchange analysis. Environ. Res. 18:336--346; 1979. 236. Borek, C.; Ong, A.; Cleaver, J.E. DNA damage from ozone and radiation in human epithelial cells. Toxicol. lndast. Health 4:547-553; 1988. 237. Elsayed, N.M. Influence of vitamin E on polyamine metabolism in ozone-exposed rat lungs. Arch. Biochem. Biophys. 255: 392-399; 1987. 238. Elsayed, N.M.; Ellingson, A.S.; Tiemey, D.F.; Mustafa, M.G. Effects of ozone inhalation on polyamine metabolism and tritiated thymidine incorporation into DNA of rat lungs. Toxicol. Appl. Pharmacol. 102:1-8; 1990. 239. Kotin, P.; Falk, H.L. The experimental induction of pulmonary tumors in strain-A mice after their exposure to an atmosphere of ozonized gasoline. Cancer 9:910-917; 1956. 240. Kotin, P.; F a l l H.; McCammon, C J . The experimental induction of pulmonary tumors and changes in the respiratory epithelium in C57BL mice following their exposure to an atmosphere of ozoulzed gasoline. Cancer 11:473--481; 1958. 241. Stokinger, H.E. Effects of air pollution on animals. In: Stern,
264
M.G. MUSTAFA
A.C., ed. Air pollution. New York: Academic Press; 1962: 282-334. 242. Gardner, M.B. Biological effects of urban air pollution: lung tumors in mice. Arch. Environ. Health 12:305-313; 1966. 243, Werthamer, S.; Schwartz, L.H.; Soskind, L. Bronchial epithelial alterations and pulmonary neoplasia induced by ozone. Path. Microbiol. 35:224-230; 1970. 244. Nettesheim, P.; Hanna, Jr., M.G.; Doherty, D.G.; Newell, R.F.; Hellman, A. Effects of chronic exposure to artificial smog and chromium oxide dust on the incidence of lung tumors in mice. In: Hanna, M.G.; Nettesheim, P.; Gilbert, J.G., eds. Inhalation carcinogenesis, symposium series No. 18. Washington, DC: USAEC Division of Technical Information Extension; 1970: 305-317. 245. Hassett, C.; Mustafa, M.G.; Coulson, W.F.; Elashoff, R.M. Murine lung carcinogenesis following exposure to ambient ozone concentrations. J. Nat. Cancer Inst. 75:771-777; 1985. 246. Last, J.A.; Warren, D.L.; Pecquet-Goad, E.; Witschi, H.P. Modification of lung tumor development in mice by ozone. J. Nat. Cancer Inst. 78:149-154; 1986. 247. Maugh, T.H. New link between ozone and cancer. Science 216:396--397; 1982. 248. Mustafa, M.G.; Hassett, C.M.; Newell, G.W.; Schrauzer, G.N. Pulmonary carcinogenic effects of ozone. In: Maltoni, C.; Selikoff, I.J., eds. Living in a chemical world: occupational and environmental significance of industrial carcinogens. New York: Ann. N.Y. Acad. Sci.; 1988:714--723. 249. Witschi, H.P. Ozone, nitrogen dioxide and lung cancer: a review of some recent issues and problems. Toxicol. 48:1-20; 1988. 250. Menzel, D.B.; Slaughter, R.J.; Bryant, A.M.; Jauregui, H.O. Heinz bodies formed in erythrocytes by fatty acid ozonides and ozone. Arch. Environ. Health 3t1:296-301; 1975. 251. Clark, K.W.; Posin, C.I.; Buckley, R.D. Biochemical response of squirrel monkeys to ozone. J. Toxicol. Environ. Health 4"741-753; 1978. 252. Moore, G.S.; Calabrese, E.J.; Labato, F.J. Erythrocyte survival in sheep exposed to ozone. Bull. Environ. Contain. Toxicol. 27:126-138; 1981. 253. Larkin, E.C.; Goheen, S.C.; Rao, G.A. Morphology and fatty acid compositon of erythrocytes from monkeys exposed to ozone for one year. Environ. Res. 32:445-454; 1983. 254. Calabrese, E.J.; Moore, G.S.; Grunwald, E.L. Ozone-induced decrease of erythrocyte survival in adult rabbits. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.A., eds. International symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, NJ: Princeton Scientific Publishers; 1983: 103-I 17. 255. Ballew, M.; Calabrese, E.J.; Moore, G.S. The effect of dietary vitamin C on ozone-induced oxidative changes in guinea pig erythrocytes. J. Environ. Sci. Health A18:597-610; 1983. 256. Vaughan, W.J.; Adamson, G.L.; Lindgren, F.T.; Schooley, J.C. Serum lipid and lipoprotein concentrations following exposure to ozone. J. Environ. Pathol. Toxicol. 0ncol.5:165-173; 1984. 257. Mole, Jr., M.L.; Stead, A.G.; Gardner, D.E.; Miller, F.J.; Graham, J.A. Effect of ozone on serum lipids and lipoproteins in the rat. Toxicol. Appl. Pharmacol. 80:367-376; 1985. 258. Konisberg, A.S.; Bachrnan, C.H. Ozonized atmosphere and gross motor activity or rats. Int. J. Biometeorol. 14:261-266; 1970. 259. Murphy, S.D.; Ulrich, C.E.; Frankowitz, S.H.; Xintaras, C. Altered function in animals inhaling low concentrations of ozone and nitrogen dioxide. Am. Ind. Hyg. Assoc. J. 2,5.246-253; 1964. 260. Weiss, B.; Ferin, J.; Merigan, W.; Stern, S.; Cox, C. Modification of rat operant behavior by ozone exposure. Toxicol. Appl. Pharmacol. 58:244-251; 1981. 261. Tepper, J.L.; Weiss, B.; Cox, C. Microanalysis of ozone depression of motor activity. Toxicol. Appl. Pharmacol. 64: 317-326; 1982. 262. Tepper, J.L.; Weiss, B.; Wood, R.W. Behavioral indices of ozone exposure. In: Lee, S.D.; Mustafa, M.G.; Mehlman, M.G., eds. International symposium on the biomedical effects of ozone-related photochemical oxidants. Princeton, NJ: Princeton
Scientific Publishers; 1983:515-526. 263. Trams, E.G.; Lauter, C.J.; Brown, E.A.B.; Young, O. Cerebral cortical metabolism after chronic exposure to ozone. Arch. Environ. Health 24:153-159; 1972. 264. Friedman, M.; Gallo, J.M.; Nichols, H.P.; Bromberg, P.A. Changes in inert gas rebreathing parameters after ozone exposure in dogs. Am. Rev. Respir. Dis. 128:851-856; 1983. 265. Hassett, C.; Mustafa, M.G.; Coulson, W.F.; Elashoff, R.M. Splenomegally in mice following exposure to ambient levels of ozone. Toxicol. Lett. 26:139-144; 1985. 266. Gardner, D.F.; Illing, J.W.; Miller, F.J.; Coffin, D.L. The effects of ozone on pentobarbital sleeping time in mice. Res. Commun. Chem. Pathol. Pharmacol. 9:689-700; 1974. 267. Graham, J.A.; Menzel, D.B.; Miller, F.J.; Illing, J.W.; Gardner, D.E. Influence of ozone on pentobarbital-induced sleeping time in mice, rats, and hamsters. Toxicol. Appl. Pharmacol. 61: 64-73; 1981. 268. Graham, J.A.; Menzel, D.B.; Miller, F.J.; Illing, J.W.; Gardner, D.E. Effect of ozone on drug-induced sleeping time in mice pretreated with mixed-function oxidase inducers and inhibitors. Toxicol. Appl. Pharmacol. 62:489--497; 1982. 269. Graham, J.A.; Miller, F.J.; Gardner, D.E.; Ward, R.; Menzel, D.B. Influence of ozone and nitrogen dioxide on hepatic microsomal enzymes in mice. J. Toxicol. Environ. Health 9: 849-856; 1982. 270. Atwal, O.S.; Wilson, T. Parathyroid gland changes following ozone inhalation: a morphologic study. Arch. Environ. Health 28:91-100; 1974. 271. Clemons, G.K.; Garcia, J.F. Changes in thyroid function after short-term ozone exposure in rats. J. Environ. Pathol. Toxicol. 4:359-369; 1980. 272. Clemons, G.K.; Garcia, J.F. Endocrine aspects of ozone exposure in rats. Arch. Toxicol. (Suppl.) 4:301-304; 1980. 273. Clemons, G.K.; Wei, D. Effect of short-term ozone exposure on exogenous thyroxine levels in thyroidectomized and hypophysectomized rats, Toxciol. Appl. Pharmacol. 74:86--90; 1984. 274. Kavlock, R.; Daston, G.; Grabowski, C.T. Studies on the development toxicity of ozone. I. Prenatal effects. Toxicol. Appl. Pharmacol. 48:19-28; 1979. 275. Kavlock, R.J.; Meyer, E.; Grabowski, C.T. Studies on the developmental toxicity of ozone: postnatal effects. Toxicol. Len. 5:3-9; 1980. 276. Gooch, P.C.; Creasia, D.A.; Brewen, J.G. The cytogenetic effect of ozone: inhalation and in vitro exposures. Environ. Res. 12:188-195; 1976. 277. Tice, R.R.; Bender, M.A.; Ivett, J.L.; Drew, R.T. Cytogenetic effects of inhaled ozone. Mutat. Res. 58:293-304; 1978. 278. Sweet, F.; Kao, M.-S.; Lee, S.D.; Hager, W.L.; Sweet, W.E. Ozone selectively inhibits growth of human cancer cells. Science 2119:931-933; 1980. 279. Chow, C.K. Nutritional influence on cellular antioxidant defense systems. Am. J. Clin. Nutr. 32:1066-1081; 1979. 280. Machlin, L.J.; Bendich, A. Free radical tissue damage: protective role of antioxidant nutrients. FASEB J. 1:441--445; 1987. 281. Heffner, J.E.; Repine, I.E. State of the art--pulmonary strategies of antioxidant defense. Am. Rev. Respir. Dis. 140:531-554; 1989. 282. Fukase, O.; Watanabe, H.; Isomura, K. Effects of exercise on mice exposed to ozone. Arch. Environ. Health 33:198-200; 1978. 283. Tappel, A.L. Will antioxidant nutrients slow aging process? Geriatrics 23:97-105; 1968. 284. Goldstein, B.D.; Buckley, R.D.; Cardenas, R.; Balchum, O.J. Ozone and vitamin E. Science 169:605--606; 1970. 285. Fletcher, B.L.; Tappel, A.L. Protective effects of alpha-tochopherol in rats exposed to toxic levels of ozone and nitrogen dioxide. Environ. Res. 6:165-175; 1973. 286. Mustafa, M.G. Influence of dietary vitamin E on lung cellular sensitivity to ozone in rats, Nutr. Rep. lnt'l. 11:473-476; 1975. 287. Donovan, D.H.; Williams, S.J.; Charles, J.M.; Menzel, D.B. Ozone toxicity: effect of dietary vitamin E and polyunsaturated fatty acids. Toxicol. Lett. 1:135-139; 1977. 288. Elsayed, N.M.; Kass, R.; Mustafa, M.G.; Hacker, A.D.; Ospi-
Biochemical basis of ozone toxicity
289.
290. 291. 292. 293. 294. 295. 296. 297. 298.
299. 300. 301.
302. 303.
304.
305. 306. 307.
308. 309. 310. 311.
tal, J.J.; Chow, C.K.; Cross, C.E. Effect of dietary vitamin E level on the biochemical response of rat lung ot ozone. DrugNutrient Interact. 5:373--386; 1989. EBayed, N.M.; Mustafa, M.G.; Hacker, A.D.; Kuehn, K.; Schrauzer, G.N. Dietary antioxidants and the biochemical response to oxidant inhalation. IIL Selenium influence on mouse lung response and tolerance to ozone. Biol. Trace Element Res. 6:249-261; 1984. Willis, R.I.; Kratzing, C.C. Ascorbic acid in rat lung. Biochem. Biophys. Res. Commun. 59:1250-1253; 1974. Stokinger, H.E. Evaluation of the hazards of ozone and oxides of nitrogen. Arch. Inudst. Health 15:181-190; 1957. Matzen, R.N. Effect of vitamin C and hydrocortisone on the pulmonary edema produced by ozone in mice. J. Appl. Physiol. 11:105-109; 1957. Freebairn, H.T. Reversal of inhibitory effects of ozone on oxygen uptake of mitochondria. Science 126:303-304; 1957. Matzen, R.N. Development of tolerance to ozone in reference to pulmonary edema. Am. J. Physiol. 190:84-88; 1957. Willis, R.J.; Kratzing, C.C. Extracellular ascorbic acid in lung. Biachim. Biophys. Acta 144:108-117; 1976. Kratziug, C.C.; Willis, R.J. Decreased levels of ascorbic acid in lung following exposure to ozone. Chem. Biol. Interact. 30: 53-56; 1980. Dubick, M.A.; Heng, H.; Rucker, R.B. Effects of protein deficiency and food restriction on lung ascorbic acid and glutathione in rats exposed to ozone. J. Nutr. 115:1050-1056; 1985. Cawthome, M.A.; Bunyan, J.; Sennitt, M.V.; Green, J. Vitamin E hepatotoxic agents. 3. Vitamin E, synthetic antioxidants and carbon tetrachloride toxicity in the rats. Br. J. Nutr. 24:357-384; 1970. Goyer, R.A.; Mehlman, M.A. Toxicology of trace elements. Washington, DC: Hemisphere Publishing Corp.; 1977:191-240. Chortyk, O.T.; Baker, J.T.; Chamberlain, W J . Selenium-mediated reduction in the mutagenicity of cigarette smoke. Environ. Molec. Mutage. 11:369-378; 1988. Johnson, G.J.; Vatassery, G.T.; Finkel, B.; Allen, D.W. Highdose vitamin E does not decrease the rate of chronic hemolysis in giucose-6-phosphate dehydrogenase deficiency. N. Eng. J. Med. 308:1014-1017; 1983. Diplock, A.T.; Lucy, J.A. The biochemical modes of action of vitamin E and selenium; A hypothesis. FEBS Lett. 29:205-210; 1973. Wilson, R.L. Free radical protection: Why vitamin E, not vitamin C, b-carotene or glutathione? In: Biology of vitamin E. Ciba Foundation Symposium 101. London: Pitman Books; 1983: 19--44. Burton, G.W.; Joyce, A.; Ingold, K.U. Is vitamin E the only lipid-soluble, chain-breaking antioxidant in human blood plasma and erythrocyte membranes? Arch. Biochem. Biophys. 221: 281-290; 1983. Packer, J.E.; Slater, T.F.; Wilson, R.W. Direct observation of a free radical interaction between vitamin E and vitamin C. Nature (London) 278:737-738; 1979. Chem, L.H.; Lee, M.S.; Hsing, W.F.; Chen, S.H. Effect of vitamin C on tissue antioxidant status of vitamin E deficient rats. Int. J. Nutr. Res. 50:156-162; 1980. Leung, H.-W.; Wang, M.J.; Mavis, R.D. The cooperative interaction between vitamin E and vitamin C in suppression of peroxidation of membrane phospholipids. Biochim Biophys. Acta 664:266-272; 1981. Slater, T.F. Free radical mechanisms of tissue injury. Biochem. J. 222:1-15; 1984. McCay, P.B. Vitamin E: interactions with free radicals and ascorhate. Annu. Rev. Nutr. 5:323-340; 1985. Lambelet, P.; Saucy, F.; Loliger, J. Chemical evidence for interactions between vitamins E and C. Experientia 41:13841388; 1985. Roberts, H.J. Perspective on vitamin E as therapy. J. Am. Med. Assoc. 246:129-131; 1981.
265
312. Fen'ell, P.M. Deficiency states, pharmacological effects, and nutrient requirements. In: Machlin, L.J., ed. Vitamin E: a comprehensive treatise. New York: Marcel Dekker; 1980:520620. 313. Witting, L.A. Vitamin E and lipid antioxidants in free-radical initiated reactions. In: Pryor, W.A., ed. Free radicals in biology, Vol. 4. New York: Academic Press; 1980:295-319. 314. Walton, J.R.; Packer, L. Free radical damage and protection: relatrionship to cellular aging and cancer. In: Machlin, LJ., ed. Vitamin E: a comprehensive treatise. New York: Marcel Dekker; 1980:495-517. 315. Bieri, J.G.; Corash, L.; Hubbard, V.S. Medical uses of vitamin E. N. Eng. J. Med. 308:1063-1071; 1983. 316. Horwitt, N.K. Status of human requirements for vitamin E. Am. J. Clin. Nutr. 27:1182-1193; 1974. 317. Bieri, J.G.; Evarts, R.P. Tocopherols and fatty acids in American diets. The recommended allowance for vitamin E. J. Am. Diet Assoc. 62:147-151; 1973. 318. National Academy of Sciences. Recommended dietary allowances. 9th ed. Washington, DC: National Academy Press; 1980:63--69. 319. Forman, H.J.; Fisher, A.B. Antioxidant enzymes of rat granular pneumocytes--constitutive levels and effect of hypoxia. Lab. Invest. 45:1-6; 1981. 320. Halliwell, B.; Wast, M.; Cn'ootveld, M. Biologically significant scavenging of the myeloperoxidase-derived oxidant hypochlorous acid by ascorbic acid. FEBS Left. 213:15-18; 1987. 321. Higson, F.K.; Keen, R.; Chevion, M. Iron enhancement of ascorbate toxicity. Free Rad. Res. Commun. 5:107-115; 1989. 322. Heys, A.D.; Dormandy, T.L. Lipid peroxidation in iron-overloaded spleens. Clin. Sci. 60:295-301; 1981. 323. Halliwell, B.; Gutteridge, J.M.C. Oxygen toxicity, oxygen radicals, transition metals and disease. Biochem. J. 219:1-14; 1977. 324. Rawley, D.A.; Halliwell, B. Formation of hydroxyl radicals from hydrogen peroxide and iron salts by superoxide- and ascorhate-dependent mechanisms: relevance to the pathology of rheumatoid disease. Clin. Sci. 64:649--653; 1983.
ABBREVIATIONS
VOC--volatile organic compounds NOx--oxides of nitrogen PUFA--polyunsaturated fatty acids SH--sulfhydryl group SS--disulfide group PSH--protein sulfhydryls NPSH--nonprotein sulfhydryls GSH--reduced glutathione GSSG--oxidized glutathione PSSG--mixed disulfide GP-- glutathione peroxidase GR--glutathione reductase MAO--monoamine oxidase RBC--red blood cells Hb-- ferrohemoglobin HbO:-- oxyhemoglobin Hb--ferdhemoglobin or methemoglobin butylated hydroxytoluene-- 3,5-tert-butyl-4-hydroxytoluene butylated hydroxyanisole--2- or 3-tert-butyl-4-hydroxyanisole
