Coronary artery hyalinosis in rats fed allylamine

EXPERIMENTAL

AND

MOLECULAR

Coronary

PATHOLOGY

Artery

14-26

10,

Hyalinosis JOSEPH

Department

of Pathology,

University

’

in Rats Fed Allylamine’

J. LALICH

of Wisconsin

Received

(1969)

Medical

September

School,

Madison,

Wisconsin

53706

15,1968

Although a considerable number of chemicals when administered parenterally are able to injure blood vessels, relatively few are capable of doing so when taken by mouth (fed). Ingested vasculotoxins must resist inactivation during intestinal absorption, somehow circumvent hepatic detoxication, and promote selective cytotoxicity for some component of the arterial wall. Only cholesterol, vitamin D, ethionine, lathyrogenic chemicals, and the alkaloid of Crotuluria spectabilis are known to do so at the present time. While “thrombogenic” diets have produced spectacular and widespread alterations in the arteries of rats, multiple chemicals are necessary to attain such results (Thomas and Hartroft, 1959; Wilgram, 1959). Interestingly, each model can be readily identified by the type of cytologic alterations produced in the vessel. Vitamin D promoted cellular degeneration and calcification of arteries and soft tissues (Gillman, 1957; Kent et uZ., 1958). Cholesterol tends to accumulate in the subintimal space and in the smooth muscles of arteries in rabbits (Imai et al., 1966), or in intimal foam cells in monkeys (Taylor et al., 1962). Ethionine induces an accumulation of periodic acid Schiff-positive material, acid mucopolysaccharide, and iron in arteries (Klavins et al., 1964). Lathyrogens cause dissecting hemorrhages and perforation of the aorta in addition to skeletal deformities (Lalich, 1967; Walker and Wirtshafter, 1956). C. spectubilis or its alkaloid monocrotaline predisposes to necrosis, inflammation, and thrombosis in the pulmonary arteries of rats (Kay et al., 1967; Lalich and Ehrhart, 1962). The unique cytologic affinity and anatomic localization in arteries manifested by each of these chemicals suggests that a variety of mechanisms may play a role in the production of vascular degeneration or necrosis in animals. In view of the different kinds of cytologic arterial injury described after ingestion of various dietary angiotoxins, one wonders whether other vasculotoxic chemicals will mimic what has already been documented or wilI further widen the spectrum of arterial response. In an attempt to find a new dietary angiotoxin, it appeared reasonable to test a chemical such as allylamine, known to be vasculotoxic following parenteral administration (Conrad et al., 1956; Guzman et al., 1961; Horst et al., 1960; and Waters, 1948). We were able to demonstrate that allylamine, when ingested, manifests vasculotoxic effects which differ appreciably from those induced by parenteral administration. In view of this observation, the following factors appeared worthy of study: ‘This investigation was supported by grant Research Grants, Public Health Services.

AM-06444-05 14

and

HE

12162-06

from

the Division

of

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IN

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15

(1) To determine the optimum concentration of drug in the diet and the duration of feeding required to consistently produce coronary artery hyalinosis and myocardial infarction; (2) to observe whether medial hyalinosis of equivalent magnitude develops in arteries of similar caliber in other organs; and (3) to describe sequentially the cellular alterations which eventuate in intimal plaque formation in the aortic arch. METHOD It was necessary to employ varying concentrations of drug to establish the quantity of allylamine in the diet which was compatible with sustained weight gains in immature rats. Some Sprague-Dawley rats received from 20 to 40 ml of 5% allylamine in corn oil/kg of ground diet. Allylamine mixed with corn oil is volatile and produces an offensive odor. For this reason, an allylamine fumarate sah was prepared for feeding in later assays. Thirty-four milliliters of allylamine was added to 200 ml of 95% ethanol. Twenty-five grams of fumaric acid was gradually added and dissolved with constant stirring in this solution. After filtration through glass wool, excess allylamine and ethanol were evaporated to l/3 the original volume under water-tap vacuum at room temperature. Thereafter, the solution was kept at 4-6°C for 24-36 hours. After precipitation, the crystals of allylamine was collected in a Biichner funnel, washed with 50 ml of cold ethanol ether (1: 1) and set aside to air dry. The salt of allylamine fumarate was pulverized with mortar and pestle before mixing with the ground diet.’ Concentrations of 1.5-2.5 gm of salt/kg of diet were fed to another group of rats. Due to the prevalence of murine pneumonia in studies which require 3 or more months, only two rats were housed in a medium-sized, open-bottom mesh cage. Food and water were fed ad l&turn. An excess of allylamine was assumed to be present in the diet when the rats stopped eating or failed to gain weight. At such times the concentration of drug in the diet was reduced. It was necessary to weight the rats at weekly intervals until it was established that the growth was sustained. Tabulation of time factor is complicated by the fact that death occurred in some test rats while the others survived. In those assays in which rats died of cardiac failure, it was necessary to add a second column to show when death occurred. After the rats began to consume their food each day and manifest a steady growth, their weights are recorded at monthly intervals. Several control rats maintained on a ground commercial diet were included in each assay which utilized weanling rats. In order that space requirements could be reduced in the table, the controls in these six assays are shown as one group. The weight tended to attain a plateau in the control and test rats after the fourth month. When pneumonia was presumed to be present, due to weight loss and coughing, the rat was killed. Control and test rats which maintained their weight and did not develop snuffles were fed up to 300 days before the animals were killed with ether anesthesia. At autopsy the organs were inspected for gross abnormalities, and samples for microscopy were put into neu2 Lab-Blox,

Allied

Mills

Inc.,

Chicago,

Illinois.

16

LALICH

TABLE RESPONSE

I

OF RATS TO ALLYLAMINE

FEEDING

Microscopic Allylamine fedperk of diet

Feedhg duration of surviVOIS in hrs

Deaths due to myocardial failure

DW when death owurred

Rats Ave. wt which insurvivors suvived(initial/linal)

GX-OSS aortic intimal plaque

alterations

Myocardium

NlXXosis

C0DIlaly hyalinoSk

Fibrosis L.V.

Sep.

R.V.

Control

189-300

“lm

2Oml

213-222

77’

3

25 ml

191-197

‘/8

5

25ml

267-284

127-143

“t5

4

30 ml

132-199

104

75

4

1

30 ml 25 ml

23 206-268

5

26-59

3

3

1

35 ml 25 ml

14 208-210

3

110-118

7

6

4

1

7

4Oml 30 ml 25ml

14 28 107-162

8

Z-56

2

4

4

1

1

3

1.75 gmb 1.50 g m

77 162-174

2

62-69

4

1

5

6

1

1

1.15 g m

152-151

2

32-62

3

1

2.5 g m 2.0 g m 1.75gm

28 28 118-156

8

47-64

2

2

% ‘“/ass

1

%

4

Lwk!

1 2

Pneumonia

10

Edema 5 Hemorrhage

5

Edema 3 Hemorrhage

4

Hemorrhage

1

4 5 4

1

5

3

2

2

Pneumonia

2

2

Hemorrhage

1

4

’ Fed as a 5% allylamine corn-oil mixture. b Salt of allylamine fumarate was added to the diet. ’ One of seven rat aortas examined had an intimal plaque.

tral formalin. The aorta was freed of surrounding tissue and opened longitudinally to the iliac bifurcation. Tissues from the myocardium, lung, kidney, mesentery, tongue, aorta, and thyroid were included for microscopy. After fixation of the heart, it was cut transversely into three segments. After trimming, all tissues were impregnated with paraffin, then sectioned and stained with hematoxylin, saffron, and eosin. Following observations of arterial edema or hyalinosis in some sections of myocardium, selected specimens of formalin-fixed tissue were cut while frozen and stained with oil-red-O. In order that comparisons might be drawn of the response to the feeding of different concentrations of drug, the data have been assembled in tabular form. Shown in Table I are: The concentration and duration of allylamine feeding, whether the rats died or survived, days when death occurred from myocardial failure, the initial and final average weight in rats which survived and the numbers of abnormalities which were seen in the heart, lungs, and aorta.

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FIG. 1. Rat fed 40 ml for 14 days, 30 ml for 28 days, and 25 ml corn-oil mixture/kg for 3 days when it died. A white infarct was seen on the lateral aspect of the left ventricle near the apex. This section of the heart illustrates the area of ischemic necrosis in which leukocytic infiltration is minimal. H, S, and E. X5. FIG. 2. Rat fed 30 ml corn-oil mixture/kg for 28 days then 25 ml for 7 days when it died of acute myocardial infarction. The lateral, posterior, and septal aspects of the left ventricle are involved. In this instance, there has been extensive leukocytic infiltration of the three areas of necrosis which accounts for the darkening of hematoxylin. H, S, and E. X5. FIG. 3. Rat fed 30 ml of corn-oil mixture/kg for 199 days when an autopsy was performed. There was a white fibrous scar on the posterior aspect of the apical portion of the left ventricle. One can observe an appreciable loss of myocardial fibers in the area of fibrosis. H, S, and E. X 5. FIG. 4. Rat fed 30 ml of corn-oil mixture/kg when because of weakness the rat was sacrificed on day 132 of feeding. At autopsy, a flabby heart and an extensive bronchopneumonia were encountered. A significant loss of myocardial fibers is apparent in the posterior left ventricle and the septum. H, S, and E. X 5.

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LALICH

RESULTS Neither necrosis nor fibrosis of the heart were seen in 25 control rats. The media of the coronary arteries was not swollen and the smooth muscle nuclei were uniformly distributed. Moderate to severe bronchopneumonia was seen in 10 of these rats. The intima in the aorta was found to be smooth in 20 control rats. When immature rats were fed 20 ml of corn oil mixture/kg of diet for 213-222 days, the rats appeared healthy and grew almost as well as the controls. Focal hyalinosis of the coronaries was seen in four animals. Three of eight had myocardial infarcts. In one rat, multiple areas of infarction were seen in the right and left ventricles. Intimal thickening in the aortic arch was seen in two while cartilagenous dysplasia occurred once. When 25 ml corn-oil mixture was fed for 191-197 days to weanling rats, they grew reasonably well. Again, coronary artery edema and hyalinosis with myocardial infarction were frequent. In two of eight aortas, gross intimal plaques were noted in the aortic arch. In 83- to 88-gm rats fed 2.5 ml/kg of diet, two of six died before 143 days while the others were killed after 225 days. Again, typical alterations in the coronary arteries, the myocardium and aortic arch were seen. In five adult rats, the concentration was increased to 30 ml/kg of diet. One rat which died of myocardial failure was found to have myocardial fibrosis and a thrombus in the left ventricle. A second rat which was killed after 132 days due to weakness and weight loss, pneumonia and extensive myocardial fibrosis (Fig. 4). Three rats tolerated this concentration for periods of 197-199 days with a slight depression in their weight. Again, as before, the coronaries, the myocardium, or the aortic intima were found to be involved in two of three rats. In eight immature rats, a concentration of 30 ml for 4 weeks was followed by 25 ml thereafter. At this concentration, five of eight died before 60 days. Acute myocardial necrosis was seen in only one of these five rats (Fig. 2). Acute pulmonary edema or alveolar hemorrhage on the other hand, were present in all five rats which died. In three which survived beyond 208 days, the alterations in the coronary arteries, the myocardium and aortic intima were typical. In 10 immature rats, a mixture of 35 ml for 14 days was followed by 25 ml/kg of diet. Three of 10 died between 110 and 118 days. Seven of these rats were fed for 208-210 days. Despite variations in time, the coronary artery hyalinosis and edema with myocardial fibrosis were approximately similar. In 10 other immature rats, corn-oil mixtures of 40 ml for 14 days were followed by 30 ml for 4 weeks before reducing to a concentration of 25 ml/kg of diet. Again, 8 rats died before 57 days. In this assay, acute myocardial necrosis and fibrosis were accompanied by pulmonary edema and alveolar hemorrhages. In two rats which survived beyond 107 days, ventricular fibrosis and a thrombus of the left auricle were observed. When allylamine fumarate was fed, similar myocardial and coronary alterations were produced. A concentration of 1.75 gm for 77 days was followed by 1.50 gm/kg of diet thereafter in 88 to 91-gm rats. Two of six died between 62 and 69 days, while four others fed a reduced concentration were autopsied between 162174 days. Alterations in the myocardium and the coronaries were typical in rats which survived beyond 162 days. When 1.75 gm/kg was fed to five adult

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19

rata, it caused a slight loss of weight. Two rats died after 34 and 62 days while the others were fed for periods of 152-157 days. Either myocardial necrosis or fibrosis was seen in three rats. Weight-depressing concentrations of allylamine fumarate were fed to ten immature rata. They received 2.5 gm for 28 days, followed by 2.0 gm for 28 days and 1.75 gm/kg of diet thereafter. At these levels,

FIG. 5. Rat fed 35 ml for 2 weeks and 25 ml corn-oil mixture thereafter for 195 days. Loss of nuclei in the media is appreciable yet the intima appears to be normal. H, S, and E. X 150. FIG. 6. Rat was fed corn-oil mixture 35 ml/kg for 14 days followed by 25 ml for 96 days when an autopsy was performed. In addition to a loss of nuclei, hyalinosis of the media and proliferation of adventitial collagen, there is prominent intimal hyperplasia. Intimal hypertrophy of this magnitude was seen in only two other rats. H, S, and E. X 150. FIG. 7. Rat was fed 40 ml for 2 weeks followed by 30 ml for 26 days, then 25 ml corn-oil mixture/ kg for 10 days when it died. Section taken from an area of septal fibrosis illustrates six branches of the coronary artery with edema and hypertrophy of the media. H, S, and E. X 150. FIG. 8. Section of heart from the same rat as shown in Fig. 3. Infarcts undergoing fibrosis frequently contained foci of calcification and cartilagenous dysplasia. H, S, and E. X60.

20

LALICH

eight of ten died between 47 and 64 days. Myocardial necrosis or fibrosis was again accompanied by alveolar hemorrhage on four occasions. Two rats which survived beyond 118 days failed to grow properly; however, one of these had myocardial fibrosis. Despite repeated attempts by feeding weight-depressing concentrations of allylamine in corn oil, or as allylamine fumarate for short periods of time, it was not possible to produce, consistently, arterial hyalinosis of small arteries in other organs. At such higher concentrations, arterial nephrosclerosis was observed in only two rats. Hyalinosis of the mesenteric, the pancreatic, and the testicular arteries was seen in four other test rats. While there was a suggestion of pulmonary artery hypertrophy in a few rata, this was also of unusual occurrence . Focal hyalinosis induced by allylamine feeding was prominent in the proximal rather than the distal coronary tributaries (Fig. 5). Slight intimal fibrosis of the coronary arteries occurred in the proximal portions in less than 25% of the specimens. Following extensive hyalinosis, adventitial collagen tended to extend into the media in a majority of the rats. Extensive intimal hyperplasia, on the other hand, was rarely seen (Fig. 6). While medial hyalinosis was conspicuous in foci near the origin of the arteries, myocardial infarction occurred more frequently in the apical half of the ventricles. Coronary artery hyalinosis and myocardial fibrosis usually did not occur in the same transverse level of the myocardium. On the other hand, medial edema of small arteries was invariably seen in and around the areas of myocardial fibrosis (Fig. 7). Myocardial fibrosis, when present, was more common in the left ventricle (Figs. 3 and 4). Those portions of the left ventricle adjacent to the septum appeared to be particularly susceptible to injury. In areas of necrosis, calcification, and cartilagenous dysplasia were fairly common (Fig. 8). While necrosis or fibrosis occurred in 45 of 75 test rats, massive myocardial involvement was manifest in about one-third of these animals (Figs, 1, 2, and 4). In 14 animals which died suddenly with acute pulmonary edema and alveolar hemorrhage, myocardial necrosis, and fibrosis were absent in most instances. Nevertheless, myocardial failure was presumed to be the cause of death in such rats. Gross intimal plaque formation of the aortic arch with cartilagenous dysplasia was common in test rats fed for more than 150 days (Fig. 10). These intimal lesions appeared to evolve in the following sequence. Endothelial cells became less prominent while increased numbers of longitudinally oriented cells with hypertrophic fusiform nuclei and an eosinophilic cytoplasm increased the width of the intima (Fig. 11). Such intimal alterations appeared to be secondary to fragmentation and lysis of the innermost elastic membrane. Thereafter, some of these intimal cells assumed a cartilagenous appearance (Fig. 12). Whereas, intimal plaques which one could identify by gross inspection were limited to the lesser curvature of the aortic arch; microscopic examination of the intima frequently revealed multiple foci of intimal thickening at other levels of the aorta. In addition to the intimal alterations, smooth muscle fibers in the aortic

CORONARY

ARTERY

HYALINOSIS

IN RATS

21

FIG. 9. R.at was fed 25 ml corn-oil mixture/kg for 225 days when an autopsy was performed. The intima is smooth, however, there is a suggestion of probable intimal thickening along the lesser curvature of the aortic arch for a distance of 4 mm. Formalin-fixed aorta. X 2.4 FIG. 10. Rat was fed 25 ml corn-oil mixture/kg for 264 days when an autopsy was performed. A well-defined triangular intimal plaque 3 mm long is seen on the lesser aortic arch. Well-defined plaques of this type are seldom seen in aortas of control rats. Formalin-fixed aorta. X 2.4. FIG. 11. Rat fed 25 ml corn-oil mixture/kg for 191 days when an autopsy was performed. An area of questionable intimal hyperplasia on gross examination is found to be abnormal on microscopic inspection. The hyperplasia is due to proliferation of longitudinally oriented cells with hypertrophic nuclei and a prominent eosinophilic cytoplasm. The elastic membrane which usually lies immediately beneath the endothelial cells is absent in the focus of intimal thickening. H, S, and E. x 130. FIG. 12. Rat fed 25 ml corn-oil mixture/kg for 284 days when an autopsy was performed. About l/5 of the width of the aortic arch is hyperplastic intima in which cartilagenous dysplasia is occurring. This type of lesion can be identified on gross inspection. H, S, and E. X 130.

22

LALICH

media particularly around arterial ostia and along the adventitial border also assumed a longitudinal orientation. In such foci, one observed prominent nuclear and cytoplasmic hypertrophy of the smooth muscle cells. Of the microscopic alterations observed in the aorta, intimal hyperplasia appeared to be the most striking. Whereas, gross intimal plaques of the aortic arch were seen in 13 test rats, on microscopic examination intimal cartilagenous dysplasia occurred in 18 aortas (Fig. 12). Samples of formalin-fixed hearts from 19 test rats were cut while frozen and then stained with oil-red-O. Medial hyalinosis on routine examination was seen in 16 while the vessels in three were considered to be normal. Oil-red-O affinity for coronaries was found in three rats with hyalinosis and in one rat with a normal coronary. Areas of myocardial fibrosis stained negatively for fat whereas the phagocytes within these infarcts usually contained cytoplasmic vacuoles with an aflinity for this dye. Coronary artery hyalinosis therefore was not always associated with fatty metamorphosis. However, in view of the focal nature of this lesion, it will be necessary to examine more hearts by this method before a proper correlation can be drawn between hyalinosis and medial oilred-0 affinity. DISCUSSION It has been shown that the concentration of allylamine administered parenterally will influence the character of arterial injury. A minimal injury was produced with intravenous injections of allylamine into dogs (Conrad et al., 1956). They observed accumulations of glycogen and PAS positive material in the media of coronary arteries and the aorta. When Mellon et al. (1935) applied allylamine topically or injected the drug intradermally into rabbits, they produced a hemorrhagic proliferative endarteritis in the dermal arterioles. After multiple injections of intravenous allylamine of sufficient concentration, Waters (1948 and 1957) produced hemorrhagic medial necrosis, inflammatory cell infiltration, and eventually a conspicuous intimal fibrosis in the coronary arteries of dogs. Similar results were obtained by Bloor and Lowman (1963) who injected equivalent concentrations of allylamine into dogs. In addition, these workers pointed out that the medium and small coronary tributaries were particularly susceptible to allylamine injections. They also recorded that myocardial infarction occurred in four of their dogs. Intravenous injection of allylamine produces alveolar hemorrhage, renal infarcts, and necrosis of hepatic cells in calves (Will, 1967). It was necessary to resort to intracoronary injection of allylamine for the production of myocardial infarction. Parenteral administration of allylamine has been undertaken in rats on several occasions with different results. Intravenous injections of 15 mg of allylamine per kg of body weight failed to produce myocardial lesions (Guzman et al., 1961). Since this concentration is effective in dogs, these workers felt that rats were more resistant. Even the inhalation of lethal doses of allylamine failed to produce consistent coronary arteritis and myocardial infarction. Others have injected greater quantities of allylamine with more success (Horst et al., 1960). They recorded medial edema and fragmentation of elastic membranes of the aorta. When

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combined intravenous injections of allylamine and egg yolk were employed, medial necrosis and leukocytic infiltration were accompanied by focal intimal thickening and aortic lipid accumulation. In none of the previous studies, however, were correlations drawn between alterations in the coronary arteries and the location or severity of myocardial necrosis or fibrosis. Spontaneous myocardial fibrosis occurs rarely in normal rats less than 2 years old (Fox, 1933; Wilgram, 1959; Wilens and Sproul, 1938). In accord with these reports, we did not see any fibrosis in 25 control rats. Myocardial infarcts were seen in only 50% of the rats when lipomatous coronary occlusions were produced with “thrombogenic” diets (Wilgram, 1959). In addition to lipid infiltration of the itima, some of the rats also developed thrombosis in the coronary arteries (Thomas and Ha&oft, 1959; Wilgram, 1959). In contrast to the observations reported on rats fed “thrombogenic” diets, myocardial fibrosis was encountered without thrombosis or complete occlusion of the coronaries in this study. Since necrosis of myocardial fibers invariably preceeded fibrosis, it is assumed that infarction had occurred. Production of myocardial necrosis or fibrosis by allylamine without arterial occlusion may be due to the property of this drug either to injure the coronary arteries more extensively or initiate additional deleterious physiological and biochemical effects in the myocardium. Age of the rats did not appear to modify the tissue response appreciably. The most suitable concentration of drug for the production of coronary hyalinosis was one which permits moderate growth in immature rats. To induce consistent coronary artery hyalinosis and myocardial infarct, however, it was necessary to extend the feeding period beyond 3 months. Concentrations of drug which caused weight loss in adult rats were also responsible for sudden death when fed to immature animals. Pulmonary edema and alveolar hemorrhage were present in the majority of these rats at autopsy. Despite the elimination of overcrowding, the maintenance of constant surveillance for infection and scrupulous cleanliness, it was not possible to control murine pneumonia. Inspection of the hearts and arteries of control rats with pneumonia, however, excluded the possibility that infection induced either coronary artery hyalinosis or myocardial fibrosis in rats of this age in this study. Attempts which were made to produce equivalent arterial hyalinosis in other organs were invariably unsuccessful. The cause for the unusual and striking susceptibility of the coronary arteries and the intima in the aortic arch to allylamine feeding remains obscure. Neither is there an adequate explanation for the focal localization of edema or hyalinosis within the coronary arteries. If the development of arterial hyalinosis were merely a matter of plasma concentration of drug plus the time of exposure, then arteries of similar size in other organs would be equally involved. Future studies may resolve whether the increased susceptibility of coronary arteries to allylamine feeding is engendered by arterial torsion (Boucek, 1964) or hemodynamic stress associated with myocardial contraction (Texon, 1963). It has been suggested that arteriolar hyalinosis may be caused by accumulations of subendothelial fibrin or alterations of smooth muscle fibers (Dustin, 1962). Arterial injury produced by parenteral administration of allylamine has

24

LALICH

been attributed principally to alterations of endothelial permeability (Waters, 1948 and 1957). The production of hemorrhagic medial necrosis, leukocytic infiltration, thrombosis, and intimal fibrosis of the coronary arteries would certainly favor this point of view. Neither consistent intimal fibrosis nor thrombosis occur in the coronary arteries when allylamine is fed in the diet. Significant medial edema of the small coronary tributaries, however, is observed both after injection or feeding of the drug. Following ingestion of allylamine, edema, and degeneration of medial smooth muscle fibers appear to be more prominent than alterations in the endothelial cells of the intima. Adventitial proliferation of collagen, on the other hand, was common. It has not been possible to resolve whether altered endothelial permeability or a selective cytotoxicity of allylamine for smooth muscle exerts a greater influence on the production of medial coronary artery hyalinosis in this study. It would appear that a selective cytotoxicity of smooth muscle fibers may play a greater role that has been suspected. It is difficult to account for the focal intimal thickening and cartilagenous dysplasia which occurs in the aortic arch. Gross intimal alterations have rarely been seen in aortas of control rats of this age. Even ,&aminopropionitrile fed rats which develop obvious fusiform or saccular aortic aneurysms do not respond with gross intimal thicking (Lalich, 1967). Focal aortic intimal thickening when observed in allylamine-fed rats could be due to fibroblastic proliferation, hyperplasia of smooth muscle fibers, or a combination of both. On the basis of cytologic appearance of these cells in the aortic intima, and documented observations concerning myointimal cellular response following arterial injury (Buck, 1963) or insertion of aortic grafts (Florey et al., 1961), proliferation of smooth muscle fibers appears more likely. While focal intimal proliferation, presumably of smooth muscle fibers was observed microscopically at various levels, intimal plaques which could be identified on gross inspection developed only along the lesser curvature of the aortic arch. There is a suggestion that the consumption of allylamine promotes a cytoplasmic edema and necrosis of muscle cells particularly in the coronary arteries. The histologic alterations which are produced by allylamine ingestion, however, mimic spontaneous degeneration of the coronary arteries which is seen in aging rats more closely than any other type of experimental arterial injury (Wilens and Sproul, 1938). SUMMARY Different concentrations of allylamine in corn oil or as a fumarate salt was fed to 75 rats for periods up to 284 days. A majority of these rats developed focal hyalinosis of the proximal coronary arteries and conspicuous medial edema of the smaller branches together with myocardial infarction. Repeated attempts to produce comparable arterial hyalinosis in other parenchymatous organs were unsuccessful. Intimal cartilagenous plaques in the aortic arch were seen in 18 test rats. Coronary arteritis and intimal fibrosis usually produced by parenteral administration of allylamine to dogs did not develop when the drug was fed in the diet to rata. On the basis of a limited and focal arterial involvement and the character of cytologic alterations observed in the media of the coronary arteries, there is a suggestion that the smooth muscle fibers are selectively injured. Prolonged ingestion of allylamme therefore promotes a focal degeneration of the coronary arteries which closely resembles the morphologic alterations seen in aging rats.

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REFERENCES BOUCEK, R. J. (1964). Regional coronary artery motion and vascular fibrosis. In “Biological Aspects of Occlusive Vascular Disease.” (D. G. Chalmers and G. A. Gresham, eds.), pp. 136-155. Cambridge University Press, New York. BLOOR, C. M., and LOWMAN, R. M. (1963). Experimental coronary arteriography. The distribution and extent of allylamine-induced vascular lesions in the dog. Radiology 81, ‘770-777. BUCK, R. C. (1963). Histogenesis and morphology of arterial tissue. In, “Atherosclerosis and Its Origin.” (M. Sandler and G. H. Boume, eds.). pp. l-37, Academic Press, New York. CONRAD. L. L., GONZALEZ, I. E., JOEL, W., and FURMAN, R. H. (1956). Histochemical evaluation of canine coronary artery and aortic lesions induced by intravenous allylamine. Circulation Res. 4,263-267. DUSTIN, P., JR., (1962). Arteriolar hyalinosis. Intern. Reu. Exptl. Pathol. (G. W. Richter and M. A. Epstein, eds.). 1,73-130, Academic Press, New York. (1961). The pseudointima FLOREY, H. W., GREEN, S. J., POOLE, J. C. F., and WERTHESSEN, N. T. lining fabric grafts of the aorta. &it. J. Exptl. Puthol. 42, 236-246. FOX, H. (1933). Arteriosclerosis in lower mammals and birds: Its relation to the disease in man. In “Arteriosclerosis.” (E. V. Cowdry, ed.). pp. 153-193, Macmillan, New York. GILLMAN, T. (1957). Coronary artery disease-A possible pathogenesis. Lancet,lI, 1117-1119. GUZMAN, R. J., LOCUVAM, G. S., KODAMA, J. K., and HINE, C. H. (1961). Myocarditis produced by allylamines. Arch. Enuiron. Health 2, 62-73. HORST, A., ROZYNKOWA, D., and ZAGORSKA, I. (1960). Changes in the aorta of the rat after injection of allylamines and egg yolk. Acta Med. Polona. 1, 1-S. IMAI, H., LEE, K. T., PASTORI, S., PANLILIO, E., FLORENTIN, R., and THOMAS, W. A. (1966). Atherosclerosis in rabbits. Architectural and subcellular alterations of smooth muscle cells of aortas in response to hyperlipemia. Exptl. Mol. Pathol. 5, 273-310. KAY, J. M., HARRIS, P., and HEATH, D. (1967). Pulmonary hypertension produced in rats by ingestion of Crotalaria spectabilis seeds. Thorax. 22,176179. KENT, S. P., VAWTER, G. F., DOWBEN, R. M., and BENSON, R. E. (1958). Hypervitaminosis D in monkeys. A clinical and pathologic study. Am. J. Puthol. 34,37-59. KLAVINS, J. V., KINNEY, T. D., and KAUFMAN, N. (1964). Cardiovascular changes induced by ethionine. A. M. A. Arch. Pathol. 78, 200-208. LALICH, J. J. (1967). Aortic aneurysms in experimental lathyrism. A. M. A. Arch. Puthol. 84, 528-535. LALICH, J. J., and EHRHART, L. A. (1962). Monocrotaline induced pulmonary arteritis in rats. J. Atherosclerosis. 2,482-492. MELLON, R. R., BAKER, M. R., and M&ROY, A. P. (1935). Experimental necrotizing arteriolitis induced by a protein cleavage product. Proc. Sot. Exptl. Biol. Med. 33,92-95. TAYLOR, C. B., Cox, G. E., MANALO-ESTRELLA, P., SOUTHWORTH, J., PATTON, D. E., and CATHCART, C. (1962). Atherosclerosis in rhesus monkeys: II. Arterial lesions associat,ed with hypercholesteremia induced by dietary fat and cholesterol. A. M. A. Arch. Pathol. 74,16-34. TEXON, M. (1963). The role of vascular dynamics in the development of atherosclerosis. In “Atherosclerosis and its Origin.” (M. Sandler and G. H. Bourne, eds.). pp. 167-195, Academic Press, New York. THOMAS, W. A., and HARTROFT, W. S. (1959). Myocardial infarction in rats fed diets containing high fat cholesterol, thiouracil, and sodium cholate. Circulation 19, 65-72. WALTER, D., and WIRTSHAFTER, S. (1956). Histopathogenesis of aortic aneurysms in the Lathyrus odomtus fed rats. A. M. A. Arch. Puthol. 61,125-135. WATERS, L. L. (1948). Changes in the coronary arteries of the dog following injections of allylamine. Am. Heart J. 35,212-220. WATERS, L. L. (1957). Studies on the pathogenesis of vascular disease. The effect of intravenously injected human plasma and lipid rich human plasma globulins on inflammatory lesions in the coronary arteries of dogs. Yale J. Biol. Med. 30,57-71.

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