EXPERIMENTAL

AND

Arterial

MOLECULAR

PATHOLOGY

(1975)

Enzymes and Their Relation to Atherosclerosis in Pigeons l Tmox ZEMFWSNYI

Department

22, 225-241

of Medicine,

AND ALAN

J. ROSENSTEIN

Cardiology Section, University of Southern California Medicine, Los Angeles, California

School

of

Received July 3, 1974 Comparison of metabolic processes between the atherosclerosis-resistant Show Racer (SR) and susceptible White Carneau (WC) pigeon strains offers an opportunity to study factors which may predispose to atherosclerosis. The activity of several glycolytic enzymes, Krebs cycle enzymes, enzymes of the ATP cycle and of the glycerol phosphate shuttle was studied in SR and WC arteries. In 4-6 yr old pigeons the activity of lipoamide dehydrogenase and malate dehydrogenase is significantly lower in WC than SR arteries. The differences are not the result of aging or atherosclerosis, because they were also detected in arteries of very young 5-8 wk old pigeons. Furthermore, the arteries of the young pigeons revealed a significantly higher activity of two glycolytic enzymes, namely phosphofructokinase and aldolase, in the WC arteries as compared with SR arteries. The results in the young birds indicate that the differences between the two pigeon strains are of an inherited (genetic?) nature. It is suggested that low activity of lipoamide and malate dehydrogenases slows down the Krebs cycle and leads to low citrate and ATP production. The latter factor is an essential part of the feedback control adjustments that regulate the effici;?ncy of glycolysis via phosphofructokinase. Increased dependence of the WC arteries on glycolysis appears to facilitate the development of atherosclerosis in these birds, and the mechanism may be similar to the mechanism by which tissue hypoxia induces lipid accumulation and connective tissue alterations in the arterial wall. An additional Ending in these studies is the higher activity in female than male arteries of phosphofructokinase, aldolase, isocitrate dehydrogenase, glycerokinase, ATPase and creatine phosphokinase.

INTRODUCTION During the last decade it became evident that the pigeon is a useful laboratory animal for the study of atherosclerosis. Although birds are phylogenetically remote from man, some pigeon strains develop atherosclerotic lesions which quite closely resemble the human disease, and so the spontaneous pigeon disease became a useful model for studying not only the morphological but also the biochemical characteristics of atherosclerosis (Prichard et al., 1962, 1964, 1966; Lofland et al., 1965, 1967; Kottke et al., 1966; St. Clair et al., 1968; Lindsay and Nichols, 1971; Santerre et al., 1972; Olson and Young, 1973; Clarkson et al., 1973; Wagner et al., 1973; Thomas et al., 1973). 1 This investigation was supported USPHS Grant HL-14138.

by a Specialized

225 Copyright Q 1975 by Academic Press, Inc. All rights of reproduction in any form reserved.

Center of Research in Atherosclerosis,

226

ZEMPLENYI

AND

ROSENSTEIN

An important characteristic of pigeon atherosclerosis is the different susceptibility of some pigeon strains to spontaneous, as well as experimental atherosclerosis. The cause of this difference between the highly susceptible White Carneau (WC) and the resistant Show Racer (SR) strains has been an enigma. A genetic basis for the difference appears to emerge from cross breeding experiments (Wagner et al., 1973), but the biochemical background of the difference in susceptibility to atherosclerosis is unknown. Variations in blood lipid values appear to be insignificant (Lofland and Clarkson, 1959; Wagner et al., 1973). Our previous observations dealing with human and animal arteries revealed clear-cut enzymatic differences between arteries differing in susceptibility to atherosclerosis (Zemplenyi et al., 1966; Zemplenyi, 1968, 1974). Study of enzyme activities in the SR and WC 2 arteries could disclose additional facts. MATERIALS

AND

METHODS

White Carneau and Show Racer pigeons, 5-6 yr old (for experimental groups 1A and 1B) were obtained from the Bowman Gray School of Medicine in WinstonSalem, North Carolina. Breeding pairs of SR, WC, and WC-2 3 pigeons, 12 of each strain, were also obtained from the Bowman Gray School of Medicine. The progeny of these breeders, 5-8 wk old, were used for experimental group II. The birds were housed in pigeon fly pens, usually 15-30 in each pen. They had free access to water and food (Leach Feed and Grain Company, Downey, California) containing whole corn, Canadian peas, Austrian peas, vetch, mile, kafhr, wheat, and vitamin additive. The pigeons were killed by decapitation. The aortas, and adjacent small segments of the brachiocephalic arteries, were removed immediately and cleaned of perivascular and adventitial tissue. The arteries were rinsed several times in ice-cold saline to remove all adherent blood. In experimental group II the arteries were quickfrozen using a mixture of dry-ice and 2-methoxy-ethanol and 2 The abbreviations and “trivial enzyme names” used in this publication (systemic enzyme names and code numbers in parenthesis) are as follows: ACONIT: aconitase (citrate [isocitrate] hydro-lyase; E.C.4.2.1.3); ADP: adenosine 5’diphosphate; ALD: aldolase (fructose-l, 6-diphosphate n-glyceraIdehyde-3-phosphate-lyase; E.C.4.1.2.13); AMP: adenosine 5’-phosphate; ATP: adenosine 5’-triphosphate; ATPase: adenosinetriphosphatase, ( ATP phosphohydrolase; E.C.3.6.1.3); CPK: creatine phosphokinase (ATP: creatine phosphotransferase; E.C.2.7.3.2); DNA: deoxyribonucleic acid; EDTA: ethylenediamine tetraacetic acid; FFDW: fat-free dry weight; GK: glycerokinase ( ATP: glycerol phosphotransferase; E.C.2.7.1.30); GPDH: a-glycerolphosphate dehydrogenase (Lglycerol-3-phosphate: NAD oxidoreductase; E.C.1.1.1.8), ICDH: isocitrate dehydrogenase (threo-D.-isocitrate: NADP oxidoreductase decarboxylating; E.C.1.1.1.42); LDH: lactate dehydrogenase (L-lactate: NAD oxidoreductase; E.C.1.1.1.27 ); LIPOAM.DH: lipoamide dehydrogenase ( NADHz: lipoamide oxidoreductase; E.C.1.6.4.3); MDH: malate dehydrogenase (L-malate: NAD oxidoreductase; E.C.1.1.1.37); NAD: nicotinamide-adenine dinucleotide; NADH2: reduced NAD; NADP: nicotinamide-adenine dinucleotide phosphate; NADPHa: reduced NADP; PFK: phosphofructokinase ( ATP: n-fructose-6-phosphate l-phosphotransferase; E.C.2.7.1.11); SEM: standard error of the mean; SR: Show Racer; WC: White Carneau. s The WC-2 is an inbred strain (developed in the Bowman Gray School of Medicine) and is characterized by a greatly increased susceptibility to experimental atherosclerosis induced by a diet containing 0.5% cholesterol. The mean aortic cholesterol concentration in this strain was found by Wagner et al. (1973) to be more than twofold higher than the usual concentration in the WC strain,

ARTERIAL ENZYMES IN PIGEONS

227

stored overnight at -26°C. (Previous studies in our laboratory demonstrated that the activity of the enzymes investigated is not altered by this treatment.) In the other experimental series (group 1A and 1B) fresh arteries were chopped into small pieces and an approximate 6% homogenate (w/vol) in ice-cold saline (adjusted to pH 7.0) was prepared using a Polytron homogenizer with a PT-10 generator (Kinematica GMBH, Luzern). The homogenates were extracted by rotation for 10 min at 4°C and centrifuged at 145Og for 20 min at 4°C. The pellet was reextracted in an identical manner and supernatants combined. The enzymatic assays were as follows. Frwtose-6-phosphate kinuse (Phosphofructokinuse) was measured by a modification of the method described by Ling et al. (1966). The assay is based on the conversion of fructose-1,6diphosphate (formed by the primary reaction) to dihydroxyacetone phosphate in the presence of excess aldolase and triosephosphate isomerase. The latter reaction is coupled with an indicator reaction catalyzed by (xglycerolphosphate dehydrogenase, in which DHAP is reduced. The concomitant oxidation of NADHz is measured at 340 nm. The composition of the reaction mixture, in a total vol of 3 ml, was as follows: ATP( 0.73 mM ) ; KCl( 6.67 mM ) ; MgS04 ( 1.4 mM ) ; NADHz (0.08 mM ) ; Cleland’s reagent ( 1.7 mM ); auxiliary enzyme mixture 0.2 ml (consisting of 0.9 U aldolase, 2.43 U ar-glycerolphosphate dehydrogenase, and 13.7 U triosephosphate isomerase prepared according to Ling et al., 1966; Tris-HCl buffer, 0.2 M, pH 8.0; and tissue extract (usually 0.5 ml). The reaction was started with fructose-6-phosphate (1.4 mM), and the oxidation of NADHz was recorded with a Beckman DB-GT spectrophotometer. A&&e was measured by a method basically identical with that mentioned above, except that fructose-1,6diphosphate served as substrate for the primary aldolase-catalyzed reaction. The reaction mixture (3 ml) consisted of NADH2 ( 0.14 mM ) ; auxiliary enzyme mixture (prepared as for PFK assay but without aldolase) 0.2 ml; triethanolamine buffer, 0.05 M, pH 7.6; and the tissue extract (usually 0.5 ml). The reaction was started with fructose-l ,6-diphosphate tetracyclohexylammonium salt (0.6 mM ), and ,the oxidation of NADH2 was recorded at 340 nm. Lactate dehydrogenase was assayed by recording spectrophotometrically the oxidation of NADHz during the LDH-catalyzed reduction of pyruvate to lactate. The assay mixture (3 ml) consisted of sodium pyruvate (0.73 mM); NADHz (0.14 mM); tissue extract (usually 0.1 ml); and phosphate buffer, 0.1 M, pH 7.5. Malute dehydrogenase was assayed essentially in the same way as LDH but using oxalacetic acid (0.25 mM, prepared immediately before the assay) instead of pyruvate. Lipoamide dehydrogenase was measured by a modification of the methods devised by Reed and Willms (1966) and Massey ( 1966). The oxidation of the reduced NAD during the enzyme-catalyzed reduction of lipoamide was measured spectrophotometrically at 340 nm. The assay mixture consisted of DL-6, 8-thioctic acid amide (0.9-3.1 mM, dissolved in equal volumes of acetone and water, or in 95% ethanol); NADH2 (0.23 mM); NAD (0.24 mM); EDTA (1.1 mM); bovine serum albumin (2%) 0.2 ml; tissue extract (usually 0.2-0.5 ml); and phosphate buffer, 0.1 M, pH 5.9 or 6.5, to make up a total v01 of 3.1 ml. The reaction was started with the tissue extract.

228

ZEMPLENYI

AND ROSENSTEIN

Aconitase was assayed by linking the primary aconitase-catalyzed reaction to an indicator reaction in which isocitrate was oxidized to a-oxoglutarate and COZ, and NADP reduced by the catalytic action of isocitrate dehydrogenase. The reaction mixture (3 ml) consisted of sodium citrate (0.2 mM); NADP (0.35 mM); M&O4 (1.33 mM); Cleland’s reagent (1.7 mM); isocitrate dehydrogenase 10 Units; tissue extract (usually 1 ml); and Tris buffer, pH 7.4, 0.1 M. lsocitrate dehydrogenuse was assayed by measuring spectrophotometrically the reduction of NADP in a way analogous to the indicator reaction of the aconitase assay (see above). The reaction mixture (3.1 ml) consisted of isocitric acid, trisodium salt (1.2 mM); NADP, monosodium salt (0.13 mM); MnCl? (1.0 mM); tissue extract (usually 1 ml); and Tris-HC1 buffer, 0.1 M, pH 7.5. ATPase was assayed by linking the primary reaction to an auxiliary step in which ADP was phosphorylated by phosphoenolpyruvate in a reaction catalyzed by pyruvate kinase. The pyruvate that was formed was in turn determined by an indicator reaction catalyzed by lactate dehydrogenase. To inhibit adenylate kinase (myokinase) activity, an excess of AMP was used. The reaction mixture consisted of phosphoenolpyruvate tricyclohexylammonium salt (0.81 mA4); ATP (0.91 mM); AMP (1.82 mM); NADH2 (0.14 mM); MgS04 (0.70 m&f); KC1 (1.3 mM); pyruvate kinase (44.4 units); lactate dehydrogenase (Sigma, 17.0 units); tissue extract (usually 0.5 ml); and Tris-HCl buffer, O.lM, pH 8.0. Creative phosphokinase was assayed by a slight modification of the method described by Oliver ( 1955). It is based on the formation of ATP by the primary reaction. In an auxiliary reaction, catalyzed by hexokinase, glucose is phosphorylated; and in the indicator reaction, glucose-6-phosphate is oxidized and NADP reduced by the catalytic action of glucose-6-phosphate dehydrogenase. The assay mixture (2.8 ml) consisted of glucose (19.5 mM); Mg acetate (7.88 mM); ADP (0.83 mM); NADP (0.56 mM); AMP ( 17.8 m&i’); glutathione (0.98 mM), tissue extract (0.1-0.2 ml) and triethanolamine buffer, 0.05 M, pH 7.0 to make up the desired total volume. The reaction was started with the tissue extract and the reduction ,of NADP recorded at 340 nm. Glycerol-3-phosphate dehydrogenuse (a-glycerol phosphate dehydrogenuse) was assayed by recording spectrophotometrically the reduction of NAD during the enzyme-catalyzed oxidation of glycerol-3-phosphate to dihydroxyacetone phosphate. The assay mixture consisted of NAD (1.0 MM); glycerol-3-phosphate disodium salt (15.4 mM); tissue extract (usually 1 ml); and glycine hydrazine buffer, 0.2 M, pH 9.5 (prepared according to Hohorst (1965), or purchased from Sigma) to make up a total volume of 3 ml. The reaction was started with the tissue extract. Glycerokinase was assayed by linking the primary glycerokinase-catalyzed reaction to an indicator reaction in which glycerol-3-phosphate was oxidized to dihydroxyacetone phosphate and NAD reduced by the catalytic action of glycerol-3-phosphate dehydrogenase. The assay mixture consisted of NAD (0.5 mM); ATP (0.6 mM); glycerol (3.3 mM); MgC12 (1.0 mM); auxiliary enzyme mixture (4.86 U of a-glycerolphosphate dehydrogenase prepared as for the PFK assay but without aldolase) 0.2 ml; tissue extract (usually I ml); and glycine hydrazine buffer pH 9.5 to make up a total volume of 3 ml. The reaction was started with the tissue extract.

ARTERIAL

ENZYMES IN PIGEONS

229

Pig Artery .----A

FIG. 1. &ogenase change in described

Pigeon Artery

Proportionality between arterial phospbofructokinase, aldolase,and lactate dehyconcentration and observedactivity. (Abscissa:extract volume; ordinate: here optical density ( X10’) resulting from oxidation of NADH,. ) Details of methods are in text.

All the above enzyme activities were measured spectrophotometrically using cuvettes of 1 cm light path in a Beckman DB-GT Spectrophotometer equipped with a thermocirculator water bath for temperature regulation, recorder and automatic sampling accessory. This arrangement makes it possible to follow changes of optical densities in six samplessimultaneously. In the NAD or NADP-linked enzyme assays, the amount of substrate consumed was calculated from the amount of the coenzyme oxidized or reduced. The activities were expressed in milliunits (mu), i.e., in nanomoles of substrate converted per minute per ml of extract and finally calculated on three different bases: the protein content (Lowry et al., 1951) of extracts, the DNA content (Burton, 1962) of the artery and the fat-free dry weight. (Since in normal and atherosclerotic arteries the concentrations of metabolically inert components may differ, we consider more meaningful calculations on the FFDW basis than on the “wet” arterial weight basis.) Optimal conditions for enzyme activity measurement were established for pigeon as well as for arteries of other species. Figures 1, 2, 3 show that under the assay conditions. used the relationship between the enzyme concentrations and changes in optica densities was linear. RESULTS 1. Experiments with Arteries of Old Pigeons 4 In the fir& group (group 1A) arterial enzyme activities of nine Show Racer and nine White Carneau pigeons were compared. The ages of birds of both breeds were between 4 and 6 yr. In this group the activity of lactate dehydrogenase, malate dehydrogenase, isocitrate dehydrogenase, aconitase and lipoamide dehydrogenase were measured. Table I summarizes the results. Lipoamide dehydrogenase has a significantly lower activity in the WC than SR arteries, whether 4 A preliminaryabstracton experimentswith old pigeonswas publishedbefore (Zemplenyi et al., 1072).

ZEMPLENYI

230

Lipoamide

AND

HOSENSTEIN

DH

*

cl20

.poo

,i‘ t

\80 g60

,?' 40

'

420

f

i

OF

.2 .4 .6 .8 1.0

I

.2

A

.8 IO 12

6

I

Ejjy/Jj? ml. ext

.I

.02 .05

ml.

ext.

.2

-

FIG. 2. Proportionality between arterial lipoamide dehydrogenase, aconitase, isocitrate dehydrogenase, and malate dehydrogenase concentration and observed activity (Abscissa: exresulting from oxidation or retract volume; ordinate: here changes in optical density ( X10:') duction of NADHZ or NADP, respectively.) Details of methods are described in text.

expressed on the DNA or FFDW basis. The activity of malate dehydrogenase is also lower in the WC arteries but only when calculated on the fat-free dry weight basis. Other differences were not detected in this group of birds. In the second group (group 1B) more enzymes were studied than in the previous group. The ages of the birds were again between 4 and 6 yr. The activity of phosphofructokinase, aldolase, lactate dehydrogenase, lipoamide dehydrogenase, isocitrate dehydrogenase, aconitase, malate dehydrogenase, and Mg dependent ATPase was measured. Again, as shown in Table II, we observed a significantly lower activity of lipoamide dehydrogenase and malate dehydrogenase in the WC arteries than in the SR arteries, when calculated

IO GK 8J

2

.4 .6 .8 1.0 I2

.2 .4 .6 .8 I.0 l2

.2

A .6 .8 I.0

ml. ext. FIG. 3. Proportionality between arterial a-glycerol phosphate dehydrogenase, glycerokinase, creatine phosphokinase, and adenosine triphosphatase concentration and observed activity. (Abscissu: extract volume: ordinate: here change in optical density (X10’) resulting from oxidation or reduction of NAD or NADP.) Details of methods are described in text.

For

abbreviations

see footnote,

p. 226.

0.270 0.290

4.177 12.18 9.944

WC SR WC

Lipoam.

SR WC

171.0 3.658

WC SR

ICDH

Acon.

211.8 167.8

WC SR

MDH

DH

198.4

Act/100 (Mean

SR

Strain

LDH

Enzyme

ARTERIAL

f f

f h f

f f

i f

0.050 0.037

0.250 1.233 1.300

8.667 0.173

11.00 10.67

h 10.67

fig p/min f SEM)

ENZYME

N.S.

N.S.

N.S.

N.S.

N.S.

SigIl.

ACTIVITIES

IN 44

TABLE OLD

PIGEONS

3.513 3.099

45.97 145.8 107.4

1873.0 44.65

2321.7 2038.7

2427.9

139.0

f f

f f f

f f

0.667 0.367

3.567 12.00 12.90

107.7 2.039

& 133.7 f 95.00

f

Act/100 pg DNA/min (Mean i SEM)

YR

I (GROUP

N.S.

p < .05

N.S.

N.S.

N.S.

Sign.

1A)

0.369 0.277

4.071 15.24 9.735

166.3 4.633

206.7 213.9

251.4

i f

f f f

i f

f f

f

0.073 0.033

0.263 1.40 1.27

8.667 0.270

12.00 17.33

15.33

Act/mg FFDW/min (Mean f SEM)

N.S.

p < .Ol

N.S.

p < .05

p < .05

Sign.

M c

! UY

52 5¶ ij

f5 CA

? 2

is

rl E

?-

For

see footnote,

p. 226.

SR WC

ATPase

abbreviations

SR WC

MDH

SR WC

Lipoamide

SR WC

SR WC

LDH

ICDH

SR WC

Aldolase

DH

SR WC

Strain

PFK

Enzyme

31.13 31.78

177.1 155.7

3.511 4.129

13.53 11.85

173.6 186.2

5.137 5.513

1.040 1.026

Act/100 (Mean

ARTERIAL

f i

f f

f f

f f

f f

f f

f f

1.202 0.867

7.071 6.667

0.240 0.177

0.788 0.500

5.480 8.733

0.453 0.507

0.120 0.160

pg p/min f SEM)

ENZYME

N.S.

p < .05

N.S.

N.S.

N.S.

N.S.

N.S.

Sign.

ACTIVITIES

YR

II OLD

PIGEONS

383.1 372.5

2160.7 1817.9

43.33 48.10

164.6 137.3

2122.0 2159.6

62.17 63.51

12.930 12.044

86.26 94.67

3.500 2.367

8.591 4.633

80.72 94.17

4.667 4.933

1.754 1.980

f 23.33 zk 20.00

f f

f f

f f

f f

f f

f f

Act/100 pg DNA/min (Mean f SEM)

IN 4-6

TABLE

N.S.

N.S.

N.S.

Sign.

1B)

N.S.

p < .05

N.S.

p < .Ol

(GROUP

35.37 32.23

199.3 158.0

4.070 4.201

15.10 11.94

197.0 187.3

5.668 5.511

1.192 1.046

Act/mg (Mean

f xt

f f

f f

f f

f f

zk f

f f

2.651 1.533

11.31 8.333

0.424 0.267

0.813 0.500

13.19 7.767

0.368 0.413

0.148 0.174

FFDW/min f SEM)

N.S.

p < .Ol

N.S.

p < .Ol

N.S.

N.S.

N.S.

Sign.

ARTERIAL

ENZYMES

IN PIGEONS

233

on the DNA or FFDW basis. (In the case of malate dehydrogenase also on the extract protein basis.) The activity of the other enzymes did not reveal ,any significant difference between the two pigeon breeds. 2. Experiments with Arteries of Young Pigeons In these experiments, the arteries of 11 Show Racer and 18 White Carneau pigeons (10 WC and eight WC-2) were used. The birds were onIy 5-8 wk old. The activities of phosphofructokinase, aldolase, lactate dehydrogenase, lipoamide dehydrogenase, isocitrate dehydrogenase, malate dehydrogenase, ATPase, creatine phosphokinase, glycerokinase, and glycerol-3-phosphate dehydrogenase were measured. The results are summarized in Table III and Figs. 4, 5, and 6. Similarly, as in the experiments with the arteries of the older birds, the comparison of lipoamide dehydrogenase activity reveals a significant difference between SR and WC or WC-2 arteries when calculated on the DNA or FFDW basis. The activity of malate dehydrogenase also is significantly lower in the WC arteries, in accordance with the findings in the old birds, but fails to show this difference in the WC-2 arteries. In addition, the young bird arteries show a lower activity of creatine phosphokinase, but only when calculated on the FFDW basis. In sharp contrast, phosphofructokinase reveals an unequivocally higher activity in the WC and WC-2 arteries than in the SR arteries, whether calculated on the basis of extract protein, DNA content or fat-free dry weight. Another glycolytic enzyme, aldolase, tends to be higher in the WC-2 arteries. 3. Sex-related Differences in Arterial Enzyme Activities In the group of the young pigeons peculiar sex differences in enzyme activities could be detected in spite of the small number of animals. The significant differences are summarized in Table IV. The activity of .phosphofructokinase is significantly higher in the female than male WC and WC-2 arteries whether calculated on the extract protein or DNA content or FFDW basis. A similar tendency (i.e., significant difference when calculated at least on one of those bases) was manifested by the activity of aldolase, isocitrate dehydrogenase, glycerokinase, ATPase and creatine phosphokinase. In the SR pigeons the only sex difference was the slightly higher activity in the male arteries of ATPase, calculated on the extract protein basis, DISCUSSION As outlined in the introduction, the difference in susceptibility to atherosclerosis between the SR and WC pigeons seemsto be a well established and generally accepted fact. However, the findings of some investigators are equivocal in this regard (Lindsay and Nichols, 1971; Santerre et al., 1972). One possible explanation of this discrepancy is the fact that the strains presently available from the Palmetto Pigeon Plant (the source of the pigeons for most studies) ,are not identical with the original s,trains investigated in the laboratories of the Rowman 1 Gray ‘School of Medicine. According to the information obtained from the Palmetto Plant, their White Carneau pigeons have been cross mated with White

f & h

f f f

f f f

f f f

f f f

f f f

f f f f f f

i f f

f f f

SR and other

0.812 1.411 1.003

10.92 11.33 9.573

4.239 5.159 3.943

0.251 0.143 0.141

0.242 0.287 0.162

5.977 7.918 6.600

0.062 0.065 0.070 0.032 0.025 0.035

1.132 0.151 0.120

0.030 0.220 0.115

pg p/min f SEM)

between

32.43 32.32 32.96

162.8 142.2 174.7

117.5 103.0 106.5

3.945 3.769 4.030

4.888 4.133 4.038

148.2 159.7 144.3

0.764 0.758 0.758 0.497 0.522 0.480

2.954 3.105 3.255

0.59 1.260 0.976

Act/100 (Mean

significant differences see footnote, p. 226.

SR WC WC-2

ATPsse

p values indicate For abbreviations

SR WC WC-2

CPK

SR WC WC-2

Lipoam.

SR WC WC-2

SR WC WC-2

LDH

MDH

SR WC WC-2

GK

SR WC WC-2

SR WC WC-2

a-GPDH

ICDH

SR WC WC-2

Aldolase

DH

SR WC WC-2

Strain

PFK

Enzyme

III

strain

N.S.

N.S.

p < .05 N.S.

N.S.

N.S. p c.02

N.S.

N.S.

N.S.

N.S.

p < .Ol p c.002

Sign.

arteries.

501.3 486.5 536.2

2482.5 2094.2 2840.6

60.15 56.63 65.27 1802.0 1508.9 1720.0

74.49 63.76 65.42

2270.3 2337.1 2337.0

7.699 7.842 7.780

11.95 11.66 12.10

9.192 20.59 15.85 45.31 45.70 53.01

0.577 0.661 0.415

1.221 1.416 0.848

0.650 4.377 1.995 1.977 2.697 2.745

f f f

f f f

f i f f f f

f f f

23.36 38.37 24.59

136.0 180.9 176.0

3.048 4.142 2.078 64.06 71.53 41.26

2.606 2.310 2.675

I& 83.50 f 110.7 f 105.0

f f f

f f f

f f f f z!z f

Act/100 pg DNA/min (Mean f SEM)

N.S.

N.S.

p < .Ol N.S.

N.S.

p < .05

p < .Ol

N.S.

N.S.

N.S.

N.S. p

Arterial enzymes and their relation to atherosclerosis in pigeons.

EXPERIMENTAL AND Arterial MOLECULAR PATHOLOGY (1975) Enzymes and Their Relation to Atherosclerosis in Pigeons l Tmox ZEMFWSNYI Department 22,...
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