Clinical and Experimental Allergy, 1992, Volume 22, pages 667-672
REVIEW
The effects of air pollution on allergic bronchial responsiveness N. A. MOLFINO*, A. S. SLUTSKYf and N. ZAMELJ * Medical Research Council of Canada, ^Respiratory Division, University of Toronto and %Tri-Hospital Pulmonary Function Laboratories, University of Toronto, Toronto, Ontario, Canada
Introduction
Asthma morbidity and mortality appear to be increasing in most countries [1]. A number of investigators have postulated that this disturbing observation may be related to increased air pollution [2-5], changes in meteorologic parameters [6] as well as seasonal variations [7]. In addition, allergens are well known triggers for acute severe asthmatic exacerbations [8,9]. Since airborne allergens and air pollution are often both increased during the same season [10], potentiation, either in the degree of acquired sensitization [11] or in the degree of the response to allergens [12], should be considered as another important factor that might help to explain the increasing asthma morbidity and mortality rates in some developed countries [1]. Unfortunately, there have been few human studies addressing this potential interaction. In this article, we review the available evidence from animal and human studies regarding the possible interaction between pollutants and allergens. Since indoor air pollution levels can be as high as those outdoors [13] and since most of the population spend the majority of the time indoors, exposure to pollutants from either source is for the purpose of simplification, used interchangeably in this review and a similar rationale will be applied to allergens.
Studies in animals
A number of studies have been performed in animal models to investigate the interaction between air pollutants and allergens. In general, these studies have been useful because it has been possible to use higher doses of pollutants than would be possible in human studies, and hence examine potentially toxic effects over a range of concentrations. However, because of the higher doses usually used and possible inter-species differences [14],
Correspondence: Dr N. Zamel, Mount Sinai Hospital, 600 University Avenue, Room 656, Toronto, Ontario, Canada M5G 1X5.
care has to be taken in extrapolating these results to humans. Table 1 summarizes the most important reports that have investigated a relationship between pollutants and antigens in different animal species in the last 20 years. In the majority of these studies the mechanisms for such a potentiation have been related to an increment in the allergen absorption [15] with greater sensitization [16-22] and greater response [16,18,23-28]. From these reports it is evident that a deleterious interaction of pollutants and antigens exists at least in animal models. However, the mechanisms for this potentiation might be different depending on the pollutant, antigen and animal model used. Furthermore, it has been shown that particles present in diesel exhausts also act as allergen carriers [16,19,20]. Matsumura [21] reported that most of the animals exposed to ozone at 5 p.p.m. and to nitrogen dioxide at 70 p.p.m. developed anaphylatic shock after nebulization of antigen aerosol (albumin). This was in striking contrast to the control group in which none or only one developed anaphylaxis. Moreover, eight of the 11 animals exposed to ozone at 5 p.p.m. and five of the 10 animals exposed to sulphur dioxide at 330 p.p.m. showed increments in haemagglutination tests suggesting that these gases might have induced sensitization via the airways of the animals. In addition, to provoke cutaneous anaphylaxis in the exposed animals, the mean minimal dose of antigen required was significantly lower than in the control group. Thus, in guinea pigs, repeated 30-min periods of exposure via the respiratory tract enhanced sensitization to albumin. Matsumura suggested that ozone, nitrogen dioxide and sulphur dioxide stimulated the immune system promoting activation of the alveolar macrophages with greater absorption or prolonged retention of antigen in the lung. Matsumura further investigated this possibility [15] by infusing I'-^' labelled albumin after a 30-min exposure to 8 p.p.m. of ozone. Radioactivity in the blood of exposed animals increased much more rapidly and was significantly higher than in the control group and lung radioactivity was twice as high in the exposed than in the non-exposed group. This lead Matsumura to conclude 667
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N. A. Molfino, A. S. Slutsky and N. Zamel
that ozone for 30 min at a concentration of 8 p.p.m. facilitated the absorption of antigen and prolonged its retention in the lung of guinea pigs. Finally, Matsumura [26] reported that guinea pigs could also be sensitized via the intraperitoneal route with egg albumin with the induction of anaphylaxis by nebulized antigen following exposure to more than 2 p.p.m. ozone and to 40 p.p.m. nitrogen dioxide but not to 400 p.p.m. sulphur dioxide. This was one of the first reports suggesting different mechanisms of action among pollutants based on the different solubility of sulphur dioxide in water and thus, different degrees of absorption by the respiratory system. Osebold et ai. [17] showed that exposure of mice to 0-5 or 0-8 p.p.m. of ozone was accompanied by immunological changes in the lungs including increases in secretory igA as well as increases in IgA containing cells in the bronchus lymphoid associated tissue and also along mucous membranes. Since serum albumin levels increased in respiratory secretions, Osebold et ai. hypothesized that there was a loss of membrane integrity after the exposure to ozone leading to an increase in immunologic responses. Osebold et ai. extended their previous findings [24] by nebulizing antigen (albumin) to mice previously exposed to ozone or sulphur dioxide. They found not only an enhancement ofthe immune response in terms of anaphylaxis but also synergism between ozone and sulphur dioxide. They used their previous findings [17], namely disruption of membranes, to explain this observation and those of Matsumura [15,21,26] and concluded that pollutants would increase the number of sensitized individuals. Following these reports, Gershwin et al. [22] found that mice exposed to ozone (0-5 or 0 8 p.p.m.) had a 34 2-fold increment of IgE-containing cells following the inhalation of antigen while the control group (no exposure to ozone) had only a 9 4-fold increment. This enhancement correlated well with anaphylactic sensitivity to intravenous ovalbumin; however, threshold ozone levels were not determined in the reports by Osebold et al. [17,24] or Gershwin et al. [22]. Biagini et al. [28] lent support to these reports by showing that primates exposed to both ozone at 1 p.p.m. and platinum (a specific airway sensitizer) had an increased incidence of positive skin tests to platinum and of bronchial responsiveness to inhaled methacholine as well as inhaled platinum. These increments were more than those obtained when any ofthe two exposures were performed separately. They concluded that some of their findings could explain the respiratory symptoms experienced by workers exposed to platinum and suggested that ozone could accelerate the production of specific IgE in this circumstance. Another group of investigators [16,19,20] conducted
studies, in which they exposed mice to diesel-exhausts. They found an overall increment in the IgE production when the animals were immunized with diesel exhausts and ovalbumin [19] or red cedar [16] intraperitoneally [19] as well as by the intranasal route [20]. Riedel et al. [23] reported a sulphur dioxide (at 01 p.p.m.) facilitation of allergic sensitization to ovalbumin in guinea pigs. This was one of the first reports utilizing levels of a pollutant lower than those suggested by the United States Clean Air Act (for sulphur dioxide = 014 p.p.m.). Exposure to these levels during 8 hr on 5 consecutive days induced allergic bronchial obstruction and increased concentrations of specific antibodies to ovalbumin in bronchoalveolar lavage. When concentrations of sulphur dioxide were increased, a greater enhancement was detected plateauing at a concentration of 16 6 p.p.m. There have been a few conflicting reports regarding the effects of exposure to ozone on the acute and late allergic response to Ascaris suum in dogs. Kleeberger et al. [29] reported an attenuation ofthe acute response by exposing the animals to ozone. This was associated with a significant increase of neutrophils and degranulated mast cells. Subsequently, Turner et al. [30] found no influence of a previous inhalation of ozone on the acute response, but they did observe blunting of the late response to antigen despite a significant increase in the number of mast cells and neutrophils in the airway of the dogs 3 hr after exposure to 1 p.p.m. of ozone for only 5 min. These differences in the acute response in the two studies may be related to the different dose of antigen used in both studies [29,30]. Although several hypotheses were proposed [29] to explain the findings, the blunting ofthe late response in the presence of an increase in inflammatory cells remains speculative. More recently, Yanai et al. [18] found that previous administration of inhaled ozone at 3 p.p.m. increased airway responsiveness to acethylcholine and to Ascaris suum only in sensitized dogs. OKY-046, a thromboxane synthesis-inhibitor, blocked the ozone-induced increments to acethylcholine bronchial responsiveness but it failed to blunt the specific response to antigen. Based on these results they concluded that ozone increased the responsiveness to inhaled antigen through a different mechanism from that of ozone-induced muscarinic hypcrresponsiveness. Although there are some contradictory data in the literature related to the animal studies, the weight of evidence supports the concept that there is a potentiation between different pollutants and various types of antigen. The following section will focus on the human data— epidemiologic and experimental—addressing the interaction between pollutants and allergens.
Air pollution and allergic bronchial responsiveness
Studies in humans Epidemiological evidence A body of epidemiological literature exists relating air pollution with asthma [4,5,31-36]. Epidemiological studies allow investigation of a large number of people exposed to pollution under real life conditions. However, they lack adequate control of the many variables present in the environment [37] (weather, allergens, smoking, etc.) and of those factors which can influence the patients' behaviour (age, housing, medical care, socioeconomic status, etc). Further, epidemiological studies fail to measure all potentially relevant substances in the air. Moreover, a potentiation between pollutants can occur in the environment [38] making the conclusions from epidemiological studies even weaker when they are intended to determine specific causative factors. Because of all these limitations, the epidemiological evidence attempting to link pollution with aggravation of allergic conditions [2] is in general controversial. Nevertheless, these studies are very important in identifying potential factors affecting asthmatic exacerbations [7]. Weill et al. [39] as well as Salvaggio and Klein [40] suggested that only atopic invidivuals were affected by air pollution during periodic exacerbations of obstructive pulmonary disease in New Orleans in the late 1950s.
669
However, it has also been proposed [39] that the presence of a grain elevator on the Mississippi river coast and not air pollution per se was responsible for the increments in asthma crises registered among atopic individuals. A similar episode was reported in 1964 from the province of Saskatchewan in Canada [41]. Furthermore, the increased incidence of asthmatic episodes in military personnel that started in 1954 in the highly industrialized Tokyo-Yokohama area [35,36] initially seemed to have had a greater efl"ect on individuals with a family history of allergies [35,36]. Yet, the same author of the original description [36] subsequently changed the term Tokyo-Yokohama asthma' to 'Tokyo-Yokohama respiratory disease' probably to emphasize that emphysematous and chronic bronchitis patients had been affected more severely. Similarly, Brown and Ipsen [33] reported that atopic asthmatics followed for several years, suffered from exacerbations as pollen counts increased; however, the relationship with 10 indices of air pollution and humidity showed inconsistencies. Thus, the number and degree to which atopic patients were affected during these episodes in the 1950s and 1960s remains unclear. In 1981, Richards et al. [31] studied the relationship between asthma emergency visits, metereological conditions as well as levels of different pollutants and aeroaller-
Table 1 . Studies showing a potentiation between pollutants and antigens in different animal species
Reference
Year
Species
Pollutant
Antigen
25 21 15 26
1967 1970 1970 1970
Mice Guinea pigs
Ozone Ozone, nitrogen and sulphur dioxides
Streptococcus Albumins
Increased susceptibility to infection Increased sensitization, absorption, lung retention and anaphylaxis
Result
17
1979
Mice
Ozone
Ovalbumin
Increased IgA in secretions and lymphoid tissue
24
1980
Mice
Ozone, sulphur dioxide
Ovalbumin
Potentiation of the allergic response and synergism between pollutants
22
1981
Mice
Ozone
Ovalbumin
Increased IgE-containing cells and enhancement of sensitization
27
1983
Sheep
Ozone
Ascaris suum
Potentiation of the response to antigen
28
1986
Primates
Ozone
Platinum
Increased dermal sensitivity and bronchial response to methacholine and platinum
19
1986
Mice
Diesel exhausts
Increased production of IgE
Increased bronchial response to methacholine and antigen
20
1987
Mice
Diesel exhausts
23
1988
Guinea pigs
Sulphur dioxide
16
1989
Mice
Diesel exhausts
Ovalbumin Ovalbumin Ovalbumin Cedar pollen
18
1990
Dogs
Ozone
Ascaris suum
Increased production of IgE Potentiation of the allergic bronchial response Increased immune response
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N. A. Molfino, A. S. Slutsky and N. Zamel
gens in Los Angeles. The authors found a positive correlation between worsening of asthma and nitric oxide, haze, hydrocarbons, wind conditions and total allergen counts. Negative correlations were found with levels of ozone, sulphur dioxide, temperature and relative humidity. In this context, the negative correlation with ozone and sulphur dioxide is not surprising since it might have been influenced by the low temperatures that were registered simultaneously during the conduct of the study. It is generally known that low temperatures and humidity [6,40,42] may not only worsen asthma per se but also reduce the levels of ozone and sulphur dioxide [4]. This is an example of the difficulty in interpreting data from the epidemiological studies. Subsequently, in 1983 [43], Bates and Sizto reported an increase in hospital admissions 24 hr after a rise in ozone concentration in one region of Canada. This lag time suggested that pulmonary function in sensitized asthmatic subjects deteriorated, not only because of exposure to ozone but also perhaps because of an additional exposure to some other agent (e.g. allergens). Ishizaki et al. [44] examined the incidence of hay fever in a district in Japan. They found that the incidence depended on the levels of pollutants. The average value was 9 7%; however, if the towns or villages had high allergen counts and motor vehicle traffic then the incidence was 13 2%, while high pollen concentration and low automobile exhausts yielded a value of 8 8%. In forest regions with high pollen counts and no vehicles the value was 5-1% and in mountain regions without both factors the incidence was as low as 1-7%; however, in towns and villages without pollen but with pollutants the incidence was back up to 9 6%. Recently, Popp et al. [45] have shown also that sensitization to aeroallergens can occur in polluted areas more frequently than in rural non-polluted regions. In this study, allergen specific IgE was significantly higher in patients from highly polluted areas than in the patients from rural areas, despite the lack of a significant increase in reported symptoms. Finally, more recently in Canada, Bates et al. have reported a relationship between markers of asthma morbidity and air pollutants [4,5]. Unfortunately, none of these last two studies differentiated between atopic and non-atopic asthmatics. The southern Ontario study [5] showed a positive correlation only with levels of ozone. The population affected were subjects who were older than 14 years, suggesting that patients with chronic airflow limitation and not only asthmatics were affected. The Vancouver study [4] indicated that there was a positive relationship between temperature and emergency visits for asthma in nine hospitals during three consecutive years in all age groups (with hotter days more visits occurred). In the 15-60 age group, asthma and respira-
tory visits were significantly correlated with SO2 and SO4 levels. Although the epidemiological evidence suggests a causal relationship between pollutants and worsening of atopic conditions as well as seasonal trends [7], this evidence remains inconclusive. In addition, by the nature of these studies they have rarely arrived at conclusions regarding specific mechanisms linking pollutants and atopy. Laboratory evidence Laboratory studies relating pollutants and allergens in humans are few in number, despite the fact that their potential value was recognized many years ago [32]. Holtzman et al. [46] reported one of the first investigations suggesting that non-smoking atopic subjects had an increased susceptibility to inhaled pollutants compared with healthy non-smokers. They found that histamine challenge performed following a 2 hr-exposure to 0 6 p.p.m. of ozone doubled the specific airway resistance in non-atopic subjects but almost tripled it in atopies. Recently, Molfino et al. [12] demonstrated that inhalation of 012 p.p.m. of ozone increased the bronchial response to allergen in resting asthmatic subjects without changes in baseline airway function. Starting from the same baseline FEVi, the mean dose of allergen required after breathing ozone for 1 hr to produce a similar early response was approximately half the dose required when air preceded the allergen challenge. Furthermore, when inhalation of ozone was followed by placebo instead of allergen there were no changes in FEVi during the six following hours. This was the first study to demonstrate that levels of ozone commonly observed in large urban centres can increase the allergic bronchial responsiveness in asthmatic subjects. However, before the results of this study are used to affect public policy, they must be replicated in a larger number of patients and in other laboratories. These results are different from those of Bascom et al. [47] obtained in subjects with allergic rhinitis, suggesting that mechanisms at the level of the nose may differ substantially from those in the lower airways in response to both pollutants and antigens. Using even lower levels of ozone (0 08 and 0-10 p.p.m.), but for a longer period of time (6 hr), and using exercise to increase minute ventilation in normal subjects, Devlin et al. [48] reported both biochemical and cellular inflammatory changes in bronchoalveolar lavage performed 18 hr after the ozone exposure in healthy subjects. This study suggests that lower levels than those currently accepted as safe for environmental ozone (012 p.p.m.) may be deleterious for normal airways and suggests that patients with airway diseases (e.g. asthma) may suffer from even greater impairment after such an exposure.
Air poilution and allergic bronchial responsiveness
Clearly more laboratory studies are needed to elucidate the possible mechanisms underlying the interaction between pollutants and allergens. Hypotheses to explain possible interactions include that of Leskowitz et al. [49] who proposed an increased absorption of allergens across the mucosal surface and a more efficient contact between antigen and antigen presenting cells that could lead to an increased sensitization to allergen. As mentioned previously this was also proposed by Matsumura [15,21,26] and Osebold et al. [17,24] based on results from animal studies. More recently, Ruffin et al. [50] have suggested that pollens can increase their antigenicity if a concomitant exposure to air pollutants (SO2, NO2 and CO) exists. This would also be the human counterpart of the studies done in mice by Muranaka and colleagues [19,20]. Finally, the oxidant effects of ozone may play a critical role in its deleterious effect [10,48]. Ozone contains two unpaired electrons and can oxidize target molecules or generate free radicals as well as act on the sulphydril group of glutathione, reacting with compounds such as NADH and NADPH. Indeed, the study of Devlin et al. [48] strongly supports the hypothesis that the inflammatory effect on the airway is the result ofthe oxidant effects on biological membranes described above. This would damage the epithelium with production of inflammatory mediators and neutrophil infiltration that would potentiate inflammation [10,48]. Conclusions
During the same period in which the prevalence of atopie asthma appears to be increasing, there is a concomitant increase in the concentrations of atmospheric pollutants (oxides of nitrogen, ozone, sulphur dioxide, aerosol particles and vehicle exhausts) [10]. Evidence for the seasonal nature of asthma morbidity and mortality has been reported [7] and data from a number of investigators from different parts of the world suggests that air pollution contributes to a potentiation of the airway allergic response in susceptible populations. It is evident by examining the current state of the knowledge [2] that further studies in humans identifying the specific agents and the concentrations of these agents that can affect pulmonary function are required. These studies may help to explain the increasing problem of asthma morbidity and mortality and may offer a plausible explanation for the putative beneficial effects of a geographic move for patients with allergic respiratory disease. References 1 Sears MR. Epidemiology of asthma. In: O'Byrne PM, ed. Asthma as an inflammatory disease. New York: Dekker, 1990:15-48. 2 Lippert FW, Morris SC. Air pollution benefit-cost assessment. Science 1991; 253:606-9.
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3 Imai M, Yoshida K, Kitabake M. Mortality from asthma and chronic bronchitis associated with changes in sulfur oxides air pollution. Arch Environ Health 1986; 41:29-35. 4 Bates DV, Baker-Anderson M, Sizto R. Asthma attack periodicity: a study of hospital emergency visits in Vancouver. Environ Res 1990; 51:51-70. 5 Bates DV, Sizto R. The Ontario Air Pollution Study: identification of the causative agent. Environ Health Perspect 1989; 79:69-72. 6 Piccolo MC, Perillo GME, Ramon CG, DiDio V. Outbreaks of asthma attacks and meteorologic parameters in Bahia Blanca, Argentina. Ann Allergy 1988; 60:107-10. 7 Weiss KB. Seasonal trends in US asthma hospitalizations and mortality. J Am Med Ass 1991; 263:2323-8. 8 O'Hollaren MT, Yunginger JW, Offord KP, Somers MJ, O'Connell EJ, Ballard DJ, Sachs MI. Exposure to an aeroallergen as a possible precipitating factor in respiratory arrest in young patients with asthma. N Engl J Med 1991; 324:359-63. 9 Pollart SM, Chapman MD, Fiocco GP, Rose G, Platts-Mills AE. Epidemiology of acute asthma: IgE antibodies to common inhalant allergens as a risk factor for emergency from visits. J Allergy Clin Immunol 1989; 83:875-82. 10 Editorial. Ozone: too much in the wrong place. Lancet 1991; 330:221-2. 11 Pauli G, Bessot JC, Quoix E. Effect of the environment on the development of respiratory allergies. Rev Pneumol Clin 1989; 45:231-6. 12 Molfino NA, Wright SC, Katz I, Tarlo S, Silverman F, McClean PA, Szalai JP, Raizenne M, Slutsky AS, Zamel N. Effect of low concentrations of ozone on inhaled allergen responses in asthmatic subjects. Lancet 1991; 338:199-203. 13 Weschler CJ, Shields HC, Naik DV. Indoor ozone exposures. J Air Pollut Control Assoc 1989; 39:1562-8. 14 Wanner A, Abraham WM, Douglas JS, Drazen JM, Richerson HB, Ram JS. Models of airway hyperresponsiveness. Am Rev Respir Dis 1990; 141:253-7. 15 Matsumura Y. The effects of ozone, nitrogen dioxide and sulfur dioxide on the experimentally induced allergic respiratory disorder in guinea pigs. II. The effects of ozone on the absorption and the retention of antigen in the lung. Am Rev Respir Dis 1970; 102:438-43. 16 Suzuki S, Takafuji S, Miyamoto T. Particulate air pollutants as enhancers of IgE production. J Allergy Clin Immunol 1989; 1:76-8. 17 Osebold JW, Owens SL, Zee YC, Dotson WM, LaBarre DD. Immunological alterations in the lungs of mice following ozone exposure: changes in immunoglobulin levels and antibody containing cells. Arch Environ Health 1979; 34:258-65. 18 Yanai M, Ohrui T, Aikawa T, Okayama H, Sekizawa K, Maeyama K, Sasaki H, Takishima T. Ozone increases susceptibility to antigen inhalation in allergic dogs. J Appl Physiol 1990; 68:2267-73. 19 Muranaka M, Suzuki S, Koizumi K, Takafuji S, Miyamoto T, Ikemori R, Tokiwa H. Adjuvant activity of diesel-exhaust particulates for the production of IgE antibody in mice. J Allergy Clin Immunol 1986; 77:616-23.
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20 Takafuji S, Suzuki S, Koizumi K, Tadokoro K, Miyamoto T, Ikemori R, Muranaka M. Diesel-exhaust particulates inoculated by the intranasal route have an adjuvant activity for IgE production in mice. J Allergy Clin Immunol 1987; 79:639-45. 21 Matsumura Y. The effects of ozone, nitrogen dioxide, and sulfur dioxide on the experimentally induced allergic respiratory disorder in guinea pigs I. The effect on sensitization with albumin through the airway. Am Rev Respir Dis 1970; 102:430-7. 22 Gershwin LJ, Osebold JW, Zee YC. Immunoglobulin Econtaining cells in mouse lung following allergen inhalation and ozone exposure. Int Arch Allergy Appl Immunol 1981; 65:266-77. 23 Riedel F, Kraemer M, Scheibenbogen C, Rieger CHL. Effects of SO2 exposure on allergic sensitization in the guinea pig. J Allergy Clin Immunol 1988; 82:527-34, 24 Osebold JW, Gershwin LJ, Zee YC. Studies on the enhancement of allergic lung sensitization by inhalation of ozone and sulphuric acid aerosol. J Environ Pathol Toxicol 1980; 3:221-34. 25 Coffin DL, Blommer EJ. Enhancement of mortality in mice from Streptococcal pneumoniae by irradiated auto exhaust. Arch Environ Health 1967; 15:36-8. 26 Matsumura Y. The effects of ozone, nitrogen dioxide, and sulfur dioxide on the experimentally induced allergic respiratory disorder in guinea pigs III. The effect on the occurrence of dyspneic attacks. Am Rev Respir Dis 1970; 102:444-7. 27 Abraham WM, Yerger L, Marchette B, Wanner A. The efTcct of ozone on antigen-induced bronchospasm in allergic sheep. In: Lee SD, Mustafa MG, Mehlman MA, eds. International Symposium on the biomedical effects of ozone and related photochemical oxidants. Princeton, New Jersey: Princeton Scientific Publishers, 1983:193-203. 28 Biagini RE, Moorman WJ, Lewis TR, Bernstein IL. Ozone enhancement of platinum asthma in a primate model. Am Rev Respir Dis 1986; 134:719-25. 29 Kleeberger SR, Kolbe J, Turner CR, Spannhake EW. Exposure to 1 ppm ozone attenuates the immediate antigenic response of canine peripheral airways. J Toxicol Environ Health 1989; 23:349-62. 30 Turner CR, Kleeberger SR, Spannhake EW. Pre-exposure to ozone blocks the antigen-induced late asthmatic response of the canine peripheral airways. J Toxicol Environ Health 1989; 28:363-71. 31 Richards W, Azen SP, Weiss J, Stocking S, Church J. Los Angeles air pollution and asthma in children. Ann Allergy 1981; 47:348-54. 32 Zweiman B, Slavin RG, Feinberg RJ, Falliers CJ, Aaron TH. Effects of air pollution on asthma: a review. J Allergy Clin Immunol 1972; 50:305-14. 33 Brown EB, Ipsen J. Changes in severity of symptoms of asthma and allergic rhinitis due to air pollutants. J Allergy 1968; 41:254-68. 34 Wuthrich B. Epidemiology of the allergic diseases: are they really on the increase? Int Arch Allergy Appl Immunol 1989; 90:3-10.
35 Phelps HW, Sabel GW, Fischer NE. Air pollution asthma among military personnel in Japan. J Am Med Ass 1961; 175:990-3. 36 Phelps HW. Follow-up studies in Tokyo-Yokohama respiratory disease. Arch Environ Health 1965; 10:143-6. 37 Boushey HA, Sheppard D. Air Pollution. In: Murray JF, Nadel JA, eds. Textbook of respiratory medicine. Philadelphia: Saunders, 1988:1617-30. 38 Koenig JQ, Covert DS, Hanley QS, van Belle G, Pierson WE. Prior exposure to ozone potentiates subsequent response to sulphur dioxide in adolescent asthmatic subjects. Am Rev Respir Dis 1990; 141:377-80. 39 Weill H, Ziskind MM, Dickerson RC, Derbes VJ. Allergenic air pollutants in New Orleans. J Air PoUut Control Assoc 1965; 15:467-71. 40 Salvaggio J, Klein R. New Orleans asthma I. J Allergy 1967; 39:227-9. 41 Skoulas A, Williams N, Merriman JE. Exposure to grain dust. J Occ Med 1964; 6:319-29. 42 Greenburg L, Field F, Reed J, Erhardt C. Asthma and temperature change: an epidemiologic study of emergency clinic visits for asthma in three large New York hospitals. Arch Environ Health 1964; 8:642-55. 43 Bates DV, Sizto R. Relationship between air pollutant levels and hospital admissions in southern Ontario. Can J Public Health 1983; 74:117-22. 44 Ishizaki T, Koizumi K, Ikemori R, Ishiyama Y, Kushibiki E. Studies of prevalence of Japanese cedar pollinosis among the residents in a densely cultivated area. Ann Allergy 1987; 58:265-70. 45 Popp W, Zwick H, Steyrer K, Rauscher H, Wanke T. Sensitization to aeroallergens depends on environmental factors. Allergy 1989; 44:572-5. 46 Holtzman MJ, Cunningham JH, Sheller JR, Irsigler GB, Nadel JA, Boushey HA. Effect of ozone on bronchial reactivity in atopie and nonatopic subjects. Am Rev Respir Dis 1979; 120:1059-67. 47 Bascom R, Nacleiro RM, Fitzgerald TK, Kagey-Sobotka A, Proud D. Effect of ozone inhalation on the response to nasal challenge with antigen of allergic subjects. Am Rev Respir Dis 1990; 142:594-601. 48 Devlin RB, McDonnell WF, Mann R, Becker S, House DE, Schreinemachers D, Koren HS. Exposure of humans to ambient levels of ozone for 6 6 hours causes cellular and biochemical changes in the lung. Am J Respir Cell Mol Biol 1991; 4:72-81. 49 Leskowitz S, Salvaggio JE, Schwartz HJ. An hypothesis for the development of atopie allergy in man. Clin Allergy 1972; 2:237-46. 50 Ruffin J, Liu MYG, Sessoms R, Banerjee S, Banerjee UC. Effects of certain atmospheric pollutants (SO2, NO2 and CO) on the soluble amino acids, molecular weight and antigenicity of some airborne pollen grains. Cytobios 1986; 46:11929.
REVIEW
The effects of air pollution on allergic bronchial responsiveness N. A. MOLFINO*, A. S. SLUTSKYf and N. ZAMELJ * Medical Research Council of Canada, ^Respiratory Division, University of Toronto and %Tri-Hospital Pulmonary Function Laboratories, University of Toronto, Toronto, Ontario, Canada
Introduction
Asthma morbidity and mortality appear to be increasing in most countries [1]. A number of investigators have postulated that this disturbing observation may be related to increased air pollution [2-5], changes in meteorologic parameters [6] as well as seasonal variations [7]. In addition, allergens are well known triggers for acute severe asthmatic exacerbations [8,9]. Since airborne allergens and air pollution are often both increased during the same season [10], potentiation, either in the degree of acquired sensitization [11] or in the degree of the response to allergens [12], should be considered as another important factor that might help to explain the increasing asthma morbidity and mortality rates in some developed countries [1]. Unfortunately, there have been few human studies addressing this potential interaction. In this article, we review the available evidence from animal and human studies regarding the possible interaction between pollutants and allergens. Since indoor air pollution levels can be as high as those outdoors [13] and since most of the population spend the majority of the time indoors, exposure to pollutants from either source is for the purpose of simplification, used interchangeably in this review and a similar rationale will be applied to allergens.
Studies in animals
A number of studies have been performed in animal models to investigate the interaction between air pollutants and allergens. In general, these studies have been useful because it has been possible to use higher doses of pollutants than would be possible in human studies, and hence examine potentially toxic effects over a range of concentrations. However, because of the higher doses usually used and possible inter-species differences [14],
Correspondence: Dr N. Zamel, Mount Sinai Hospital, 600 University Avenue, Room 656, Toronto, Ontario, Canada M5G 1X5.
care has to be taken in extrapolating these results to humans. Table 1 summarizes the most important reports that have investigated a relationship between pollutants and antigens in different animal species in the last 20 years. In the majority of these studies the mechanisms for such a potentiation have been related to an increment in the allergen absorption [15] with greater sensitization [16-22] and greater response [16,18,23-28]. From these reports it is evident that a deleterious interaction of pollutants and antigens exists at least in animal models. However, the mechanisms for this potentiation might be different depending on the pollutant, antigen and animal model used. Furthermore, it has been shown that particles present in diesel exhausts also act as allergen carriers [16,19,20]. Matsumura [21] reported that most of the animals exposed to ozone at 5 p.p.m. and to nitrogen dioxide at 70 p.p.m. developed anaphylatic shock after nebulization of antigen aerosol (albumin). This was in striking contrast to the control group in which none or only one developed anaphylaxis. Moreover, eight of the 11 animals exposed to ozone at 5 p.p.m. and five of the 10 animals exposed to sulphur dioxide at 330 p.p.m. showed increments in haemagglutination tests suggesting that these gases might have induced sensitization via the airways of the animals. In addition, to provoke cutaneous anaphylaxis in the exposed animals, the mean minimal dose of antigen required was significantly lower than in the control group. Thus, in guinea pigs, repeated 30-min periods of exposure via the respiratory tract enhanced sensitization to albumin. Matsumura suggested that ozone, nitrogen dioxide and sulphur dioxide stimulated the immune system promoting activation of the alveolar macrophages with greater absorption or prolonged retention of antigen in the lung. Matsumura further investigated this possibility [15] by infusing I'-^' labelled albumin after a 30-min exposure to 8 p.p.m. of ozone. Radioactivity in the blood of exposed animals increased much more rapidly and was significantly higher than in the control group and lung radioactivity was twice as high in the exposed than in the non-exposed group. This lead Matsumura to conclude 667
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N. A. Molfino, A. S. Slutsky and N. Zamel
that ozone for 30 min at a concentration of 8 p.p.m. facilitated the absorption of antigen and prolonged its retention in the lung of guinea pigs. Finally, Matsumura [26] reported that guinea pigs could also be sensitized via the intraperitoneal route with egg albumin with the induction of anaphylaxis by nebulized antigen following exposure to more than 2 p.p.m. ozone and to 40 p.p.m. nitrogen dioxide but not to 400 p.p.m. sulphur dioxide. This was one of the first reports suggesting different mechanisms of action among pollutants based on the different solubility of sulphur dioxide in water and thus, different degrees of absorption by the respiratory system. Osebold et ai. [17] showed that exposure of mice to 0-5 or 0-8 p.p.m. of ozone was accompanied by immunological changes in the lungs including increases in secretory igA as well as increases in IgA containing cells in the bronchus lymphoid associated tissue and also along mucous membranes. Since serum albumin levels increased in respiratory secretions, Osebold et ai. hypothesized that there was a loss of membrane integrity after the exposure to ozone leading to an increase in immunologic responses. Osebold et ai. extended their previous findings [24] by nebulizing antigen (albumin) to mice previously exposed to ozone or sulphur dioxide. They found not only an enhancement ofthe immune response in terms of anaphylaxis but also synergism between ozone and sulphur dioxide. They used their previous findings [17], namely disruption of membranes, to explain this observation and those of Matsumura [15,21,26] and concluded that pollutants would increase the number of sensitized individuals. Following these reports, Gershwin et al. [22] found that mice exposed to ozone (0-5 or 0 8 p.p.m.) had a 34 2-fold increment of IgE-containing cells following the inhalation of antigen while the control group (no exposure to ozone) had only a 9 4-fold increment. This enhancement correlated well with anaphylactic sensitivity to intravenous ovalbumin; however, threshold ozone levels were not determined in the reports by Osebold et al. [17,24] or Gershwin et al. [22]. Biagini et al. [28] lent support to these reports by showing that primates exposed to both ozone at 1 p.p.m. and platinum (a specific airway sensitizer) had an increased incidence of positive skin tests to platinum and of bronchial responsiveness to inhaled methacholine as well as inhaled platinum. These increments were more than those obtained when any ofthe two exposures were performed separately. They concluded that some of their findings could explain the respiratory symptoms experienced by workers exposed to platinum and suggested that ozone could accelerate the production of specific IgE in this circumstance. Another group of investigators [16,19,20] conducted
studies, in which they exposed mice to diesel-exhausts. They found an overall increment in the IgE production when the animals were immunized with diesel exhausts and ovalbumin [19] or red cedar [16] intraperitoneally [19] as well as by the intranasal route [20]. Riedel et al. [23] reported a sulphur dioxide (at 01 p.p.m.) facilitation of allergic sensitization to ovalbumin in guinea pigs. This was one of the first reports utilizing levels of a pollutant lower than those suggested by the United States Clean Air Act (for sulphur dioxide = 014 p.p.m.). Exposure to these levels during 8 hr on 5 consecutive days induced allergic bronchial obstruction and increased concentrations of specific antibodies to ovalbumin in bronchoalveolar lavage. When concentrations of sulphur dioxide were increased, a greater enhancement was detected plateauing at a concentration of 16 6 p.p.m. There have been a few conflicting reports regarding the effects of exposure to ozone on the acute and late allergic response to Ascaris suum in dogs. Kleeberger et al. [29] reported an attenuation ofthe acute response by exposing the animals to ozone. This was associated with a significant increase of neutrophils and degranulated mast cells. Subsequently, Turner et al. [30] found no influence of a previous inhalation of ozone on the acute response, but they did observe blunting of the late response to antigen despite a significant increase in the number of mast cells and neutrophils in the airway of the dogs 3 hr after exposure to 1 p.p.m. of ozone for only 5 min. These differences in the acute response in the two studies may be related to the different dose of antigen used in both studies [29,30]. Although several hypotheses were proposed [29] to explain the findings, the blunting ofthe late response in the presence of an increase in inflammatory cells remains speculative. More recently, Yanai et al. [18] found that previous administration of inhaled ozone at 3 p.p.m. increased airway responsiveness to acethylcholine and to Ascaris suum only in sensitized dogs. OKY-046, a thromboxane synthesis-inhibitor, blocked the ozone-induced increments to acethylcholine bronchial responsiveness but it failed to blunt the specific response to antigen. Based on these results they concluded that ozone increased the responsiveness to inhaled antigen through a different mechanism from that of ozone-induced muscarinic hypcrresponsiveness. Although there are some contradictory data in the literature related to the animal studies, the weight of evidence supports the concept that there is a potentiation between different pollutants and various types of antigen. The following section will focus on the human data— epidemiologic and experimental—addressing the interaction between pollutants and allergens.
Air pollution and allergic bronchial responsiveness
Studies in humans Epidemiological evidence A body of epidemiological literature exists relating air pollution with asthma [4,5,31-36]. Epidemiological studies allow investigation of a large number of people exposed to pollution under real life conditions. However, they lack adequate control of the many variables present in the environment [37] (weather, allergens, smoking, etc.) and of those factors which can influence the patients' behaviour (age, housing, medical care, socioeconomic status, etc). Further, epidemiological studies fail to measure all potentially relevant substances in the air. Moreover, a potentiation between pollutants can occur in the environment [38] making the conclusions from epidemiological studies even weaker when they are intended to determine specific causative factors. Because of all these limitations, the epidemiological evidence attempting to link pollution with aggravation of allergic conditions [2] is in general controversial. Nevertheless, these studies are very important in identifying potential factors affecting asthmatic exacerbations [7]. Weill et al. [39] as well as Salvaggio and Klein [40] suggested that only atopic invidivuals were affected by air pollution during periodic exacerbations of obstructive pulmonary disease in New Orleans in the late 1950s.
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However, it has also been proposed [39] that the presence of a grain elevator on the Mississippi river coast and not air pollution per se was responsible for the increments in asthma crises registered among atopic individuals. A similar episode was reported in 1964 from the province of Saskatchewan in Canada [41]. Furthermore, the increased incidence of asthmatic episodes in military personnel that started in 1954 in the highly industrialized Tokyo-Yokohama area [35,36] initially seemed to have had a greater efl"ect on individuals with a family history of allergies [35,36]. Yet, the same author of the original description [36] subsequently changed the term Tokyo-Yokohama asthma' to 'Tokyo-Yokohama respiratory disease' probably to emphasize that emphysematous and chronic bronchitis patients had been affected more severely. Similarly, Brown and Ipsen [33] reported that atopic asthmatics followed for several years, suffered from exacerbations as pollen counts increased; however, the relationship with 10 indices of air pollution and humidity showed inconsistencies. Thus, the number and degree to which atopic patients were affected during these episodes in the 1950s and 1960s remains unclear. In 1981, Richards et al. [31] studied the relationship between asthma emergency visits, metereological conditions as well as levels of different pollutants and aeroaller-
Table 1 . Studies showing a potentiation between pollutants and antigens in different animal species
Reference
Year
Species
Pollutant
Antigen
25 21 15 26
1967 1970 1970 1970
Mice Guinea pigs
Ozone Ozone, nitrogen and sulphur dioxides
Streptococcus Albumins
Increased susceptibility to infection Increased sensitization, absorption, lung retention and anaphylaxis
Result
17
1979
Mice
Ozone
Ovalbumin
Increased IgA in secretions and lymphoid tissue
24
1980
Mice
Ozone, sulphur dioxide
Ovalbumin
Potentiation of the allergic response and synergism between pollutants
22
1981
Mice
Ozone
Ovalbumin
Increased IgE-containing cells and enhancement of sensitization
27
1983
Sheep
Ozone
Ascaris suum
Potentiation of the response to antigen
28
1986
Primates
Ozone
Platinum
Increased dermal sensitivity and bronchial response to methacholine and platinum
19
1986
Mice
Diesel exhausts
Increased production of IgE
Increased bronchial response to methacholine and antigen
20
1987
Mice
Diesel exhausts
23
1988
Guinea pigs
Sulphur dioxide
16
1989
Mice
Diesel exhausts
Ovalbumin Ovalbumin Ovalbumin Cedar pollen
18
1990
Dogs
Ozone
Ascaris suum
Increased production of IgE Potentiation of the allergic bronchial response Increased immune response
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N. A. Molfino, A. S. Slutsky and N. Zamel
gens in Los Angeles. The authors found a positive correlation between worsening of asthma and nitric oxide, haze, hydrocarbons, wind conditions and total allergen counts. Negative correlations were found with levels of ozone, sulphur dioxide, temperature and relative humidity. In this context, the negative correlation with ozone and sulphur dioxide is not surprising since it might have been influenced by the low temperatures that were registered simultaneously during the conduct of the study. It is generally known that low temperatures and humidity [6,40,42] may not only worsen asthma per se but also reduce the levels of ozone and sulphur dioxide [4]. This is an example of the difficulty in interpreting data from the epidemiological studies. Subsequently, in 1983 [43], Bates and Sizto reported an increase in hospital admissions 24 hr after a rise in ozone concentration in one region of Canada. This lag time suggested that pulmonary function in sensitized asthmatic subjects deteriorated, not only because of exposure to ozone but also perhaps because of an additional exposure to some other agent (e.g. allergens). Ishizaki et al. [44] examined the incidence of hay fever in a district in Japan. They found that the incidence depended on the levels of pollutants. The average value was 9 7%; however, if the towns or villages had high allergen counts and motor vehicle traffic then the incidence was 13 2%, while high pollen concentration and low automobile exhausts yielded a value of 8 8%. In forest regions with high pollen counts and no vehicles the value was 5-1% and in mountain regions without both factors the incidence was as low as 1-7%; however, in towns and villages without pollen but with pollutants the incidence was back up to 9 6%. Recently, Popp et al. [45] have shown also that sensitization to aeroallergens can occur in polluted areas more frequently than in rural non-polluted regions. In this study, allergen specific IgE was significantly higher in patients from highly polluted areas than in the patients from rural areas, despite the lack of a significant increase in reported symptoms. Finally, more recently in Canada, Bates et al. have reported a relationship between markers of asthma morbidity and air pollutants [4,5]. Unfortunately, none of these last two studies differentiated between atopic and non-atopic asthmatics. The southern Ontario study [5] showed a positive correlation only with levels of ozone. The population affected were subjects who were older than 14 years, suggesting that patients with chronic airflow limitation and not only asthmatics were affected. The Vancouver study [4] indicated that there was a positive relationship between temperature and emergency visits for asthma in nine hospitals during three consecutive years in all age groups (with hotter days more visits occurred). In the 15-60 age group, asthma and respira-
tory visits were significantly correlated with SO2 and SO4 levels. Although the epidemiological evidence suggests a causal relationship between pollutants and worsening of atopic conditions as well as seasonal trends [7], this evidence remains inconclusive. In addition, by the nature of these studies they have rarely arrived at conclusions regarding specific mechanisms linking pollutants and atopy. Laboratory evidence Laboratory studies relating pollutants and allergens in humans are few in number, despite the fact that their potential value was recognized many years ago [32]. Holtzman et al. [46] reported one of the first investigations suggesting that non-smoking atopic subjects had an increased susceptibility to inhaled pollutants compared with healthy non-smokers. They found that histamine challenge performed following a 2 hr-exposure to 0 6 p.p.m. of ozone doubled the specific airway resistance in non-atopic subjects but almost tripled it in atopies. Recently, Molfino et al. [12] demonstrated that inhalation of 012 p.p.m. of ozone increased the bronchial response to allergen in resting asthmatic subjects without changes in baseline airway function. Starting from the same baseline FEVi, the mean dose of allergen required after breathing ozone for 1 hr to produce a similar early response was approximately half the dose required when air preceded the allergen challenge. Furthermore, when inhalation of ozone was followed by placebo instead of allergen there were no changes in FEVi during the six following hours. This was the first study to demonstrate that levels of ozone commonly observed in large urban centres can increase the allergic bronchial responsiveness in asthmatic subjects. However, before the results of this study are used to affect public policy, they must be replicated in a larger number of patients and in other laboratories. These results are different from those of Bascom et al. [47] obtained in subjects with allergic rhinitis, suggesting that mechanisms at the level of the nose may differ substantially from those in the lower airways in response to both pollutants and antigens. Using even lower levels of ozone (0 08 and 0-10 p.p.m.), but for a longer period of time (6 hr), and using exercise to increase minute ventilation in normal subjects, Devlin et al. [48] reported both biochemical and cellular inflammatory changes in bronchoalveolar lavage performed 18 hr after the ozone exposure in healthy subjects. This study suggests that lower levels than those currently accepted as safe for environmental ozone (012 p.p.m.) may be deleterious for normal airways and suggests that patients with airway diseases (e.g. asthma) may suffer from even greater impairment after such an exposure.
Air poilution and allergic bronchial responsiveness
Clearly more laboratory studies are needed to elucidate the possible mechanisms underlying the interaction between pollutants and allergens. Hypotheses to explain possible interactions include that of Leskowitz et al. [49] who proposed an increased absorption of allergens across the mucosal surface and a more efficient contact between antigen and antigen presenting cells that could lead to an increased sensitization to allergen. As mentioned previously this was also proposed by Matsumura [15,21,26] and Osebold et al. [17,24] based on results from animal studies. More recently, Ruffin et al. [50] have suggested that pollens can increase their antigenicity if a concomitant exposure to air pollutants (SO2, NO2 and CO) exists. This would also be the human counterpart of the studies done in mice by Muranaka and colleagues [19,20]. Finally, the oxidant effects of ozone may play a critical role in its deleterious effect [10,48]. Ozone contains two unpaired electrons and can oxidize target molecules or generate free radicals as well as act on the sulphydril group of glutathione, reacting with compounds such as NADH and NADPH. Indeed, the study of Devlin et al. [48] strongly supports the hypothesis that the inflammatory effect on the airway is the result ofthe oxidant effects on biological membranes described above. This would damage the epithelium with production of inflammatory mediators and neutrophil infiltration that would potentiate inflammation [10,48]. Conclusions
During the same period in which the prevalence of atopie asthma appears to be increasing, there is a concomitant increase in the concentrations of atmospheric pollutants (oxides of nitrogen, ozone, sulphur dioxide, aerosol particles and vehicle exhausts) [10]. Evidence for the seasonal nature of asthma morbidity and mortality has been reported [7] and data from a number of investigators from different parts of the world suggests that air pollution contributes to a potentiation of the airway allergic response in susceptible populations. It is evident by examining the current state of the knowledge [2] that further studies in humans identifying the specific agents and the concentrations of these agents that can affect pulmonary function are required. These studies may help to explain the increasing problem of asthma morbidity and mortality and may offer a plausible explanation for the putative beneficial effects of a geographic move for patients with allergic respiratory disease. References 1 Sears MR. Epidemiology of asthma. In: O'Byrne PM, ed. Asthma as an inflammatory disease. New York: Dekker, 1990:15-48. 2 Lippert FW, Morris SC. Air pollution benefit-cost assessment. Science 1991; 253:606-9.
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3 Imai M, Yoshida K, Kitabake M. Mortality from asthma and chronic bronchitis associated with changes in sulfur oxides air pollution. Arch Environ Health 1986; 41:29-35. 4 Bates DV, Baker-Anderson M, Sizto R. Asthma attack periodicity: a study of hospital emergency visits in Vancouver. Environ Res 1990; 51:51-70. 5 Bates DV, Sizto R. The Ontario Air Pollution Study: identification of the causative agent. Environ Health Perspect 1989; 79:69-72. 6 Piccolo MC, Perillo GME, Ramon CG, DiDio V. Outbreaks of asthma attacks and meteorologic parameters in Bahia Blanca, Argentina. Ann Allergy 1988; 60:107-10. 7 Weiss KB. Seasonal trends in US asthma hospitalizations and mortality. J Am Med Ass 1991; 263:2323-8. 8 O'Hollaren MT, Yunginger JW, Offord KP, Somers MJ, O'Connell EJ, Ballard DJ, Sachs MI. Exposure to an aeroallergen as a possible precipitating factor in respiratory arrest in young patients with asthma. N Engl J Med 1991; 324:359-63. 9 Pollart SM, Chapman MD, Fiocco GP, Rose G, Platts-Mills AE. Epidemiology of acute asthma: IgE antibodies to common inhalant allergens as a risk factor for emergency from visits. J Allergy Clin Immunol 1989; 83:875-82. 10 Editorial. Ozone: too much in the wrong place. Lancet 1991; 330:221-2. 11 Pauli G, Bessot JC, Quoix E. Effect of the environment on the development of respiratory allergies. Rev Pneumol Clin 1989; 45:231-6. 12 Molfino NA, Wright SC, Katz I, Tarlo S, Silverman F, McClean PA, Szalai JP, Raizenne M, Slutsky AS, Zamel N. Effect of low concentrations of ozone on inhaled allergen responses in asthmatic subjects. Lancet 1991; 338:199-203. 13 Weschler CJ, Shields HC, Naik DV. Indoor ozone exposures. J Air Pollut Control Assoc 1989; 39:1562-8. 14 Wanner A, Abraham WM, Douglas JS, Drazen JM, Richerson HB, Ram JS. Models of airway hyperresponsiveness. Am Rev Respir Dis 1990; 141:253-7. 15 Matsumura Y. The effects of ozone, nitrogen dioxide and sulfur dioxide on the experimentally induced allergic respiratory disorder in guinea pigs. II. The effects of ozone on the absorption and the retention of antigen in the lung. Am Rev Respir Dis 1970; 102:438-43. 16 Suzuki S, Takafuji S, Miyamoto T. Particulate air pollutants as enhancers of IgE production. J Allergy Clin Immunol 1989; 1:76-8. 17 Osebold JW, Owens SL, Zee YC, Dotson WM, LaBarre DD. Immunological alterations in the lungs of mice following ozone exposure: changes in immunoglobulin levels and antibody containing cells. Arch Environ Health 1979; 34:258-65. 18 Yanai M, Ohrui T, Aikawa T, Okayama H, Sekizawa K, Maeyama K, Sasaki H, Takishima T. Ozone increases susceptibility to antigen inhalation in allergic dogs. J Appl Physiol 1990; 68:2267-73. 19 Muranaka M, Suzuki S, Koizumi K, Takafuji S, Miyamoto T, Ikemori R, Tokiwa H. Adjuvant activity of diesel-exhaust particulates for the production of IgE antibody in mice. J Allergy Clin Immunol 1986; 77:616-23.
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35 Phelps HW, Sabel GW, Fischer NE. Air pollution asthma among military personnel in Japan. J Am Med Ass 1961; 175:990-3. 36 Phelps HW. Follow-up studies in Tokyo-Yokohama respiratory disease. Arch Environ Health 1965; 10:143-6. 37 Boushey HA, Sheppard D. Air Pollution. In: Murray JF, Nadel JA, eds. Textbook of respiratory medicine. Philadelphia: Saunders, 1988:1617-30. 38 Koenig JQ, Covert DS, Hanley QS, van Belle G, Pierson WE. Prior exposure to ozone potentiates subsequent response to sulphur dioxide in adolescent asthmatic subjects. Am Rev Respir Dis 1990; 141:377-80. 39 Weill H, Ziskind MM, Dickerson RC, Derbes VJ. Allergenic air pollutants in New Orleans. J Air PoUut Control Assoc 1965; 15:467-71. 40 Salvaggio J, Klein R. New Orleans asthma I. J Allergy 1967; 39:227-9. 41 Skoulas A, Williams N, Merriman JE. Exposure to grain dust. J Occ Med 1964; 6:319-29. 42 Greenburg L, Field F, Reed J, Erhardt C. Asthma and temperature change: an epidemiologic study of emergency clinic visits for asthma in three large New York hospitals. Arch Environ Health 1964; 8:642-55. 43 Bates DV, Sizto R. Relationship between air pollutant levels and hospital admissions in southern Ontario. Can J Public Health 1983; 74:117-22. 44 Ishizaki T, Koizumi K, Ikemori R, Ishiyama Y, Kushibiki E. Studies of prevalence of Japanese cedar pollinosis among the residents in a densely cultivated area. Ann Allergy 1987; 58:265-70. 45 Popp W, Zwick H, Steyrer K, Rauscher H, Wanke T. Sensitization to aeroallergens depends on environmental factors. Allergy 1989; 44:572-5. 46 Holtzman MJ, Cunningham JH, Sheller JR, Irsigler GB, Nadel JA, Boushey HA. Effect of ozone on bronchial reactivity in atopie and nonatopic subjects. Am Rev Respir Dis 1979; 120:1059-67. 47 Bascom R, Nacleiro RM, Fitzgerald TK, Kagey-Sobotka A, Proud D. Effect of ozone inhalation on the response to nasal challenge with antigen of allergic subjects. Am Rev Respir Dis 1990; 142:594-601. 48 Devlin RB, McDonnell WF, Mann R, Becker S, House DE, Schreinemachers D, Koren HS. Exposure of humans to ambient levels of ozone for 6 6 hours causes cellular and biochemical changes in the lung. Am J Respir Cell Mol Biol 1991; 4:72-81. 49 Leskowitz S, Salvaggio JE, Schwartz HJ. An hypothesis for the development of atopie allergy in man. Clin Allergy 1972; 2:237-46. 50 Ruffin J, Liu MYG, Sessoms R, Banerjee S, Banerjee UC. Effects of certain atmospheric pollutants (SO2, NO2 and CO) on the soluble amino acids, molecular weight and antigenicity of some airborne pollen grains. Cytobios 1986; 46:11929.
