S.C. Nam

Rapid production of advanced atherosclerosis in swine by a combination of endothelial injury and cholesterol feeding

Abstract The aim of the current study was to find out whether a combination of a balloon-endothelial-cell-denudation procedure and cholesterol feeding would result in more rapid growth of atherosclerotic lesions in the abdominal aorta of swine than if either were used alone. The results far exceeded our expectation. The two procedures appear to act synergistically. In the first 2 or 3 mo lesions are patchy and scattered but by 6 mo they become confluent so that practically the entire abdominal aorta is covered with thick lipid-rich atherosclerotic lesions. The lesions produced by 6 mo have many of the characteristics of advanced human lesions.

Experimental atherosclerosis in rhesus monkeys: I. Gross and light microscopy features and lipid values in serum and aorta☆

Abstract Forty-five rhesus monkeys were divided into five groups fed four different high-fat diets and a stock diet. The duration of feeding varied from 33 to 70 weeks. All monkeys fed the high-fat diets developed aortic proliferative lesions composed predominantly of spindle-shaped cells, most of them laden with lipid. The thickest of these proliferative lesions measured 0.85 mm, and most were thinner, suggesting that in the monkey receiving these high-fat diets some proliferative lesions can reach this range of thickness before undergoing necrosis. Atheromatous aortic lesions characterized by necrosis and the accumulation of lipid debris were found only in monkeys receiving high-fat diets for 70 weeks. In monkeys receiving a diet in which the lipid component was 30% peanut oil and 5% cholesterol, the fatty acid composition of the aorta was similar tothat in humans developing coronary artery atherosclerosis, in that the fatty acids showing the greatest accumulation were oleic and linoleic. The largest number of atheromatous lesions, however, was associated with a diet otherwise identical to the peanut oil-cholesterol diet but containing as its lipid component 30% butter and 5% cholesterol. This latter diet also resulted in higher serum lipid levels and higher aortic lipid levels per milligram of DNA than did peanut oil diet. High-fat diets fed to rhesus monkeys appear to produce both proliferative and atheromatous aortic lesions similar to those seen in the human as regards both light microscopy features and lipid composition. Work is now in progress assessing some of the metabolic features of these lesions.

Short-term feeding of unsaturated versus saturated fat in the production of atherosclerosis in the rat☆

Abstract Complex diets containing either 40% peanut oil, corn oil, or butter were fed to weanling rats for 3 months. The rats fed the peanut oil-containing diets developed intimal spindle cell lesions similar in many respects to early human atherosclerotic lesions. Rats receiving the butter-containing diets developed foam cell lesions which do not resemble human atherosclerosis. Under electron microscopy the spindle cells were of the smooth muscle type. They appeared more undifferentiated than the smooth muscle cells seen in aortic lesions in rats fed butter-containing diets for 12 months in a previous experiment. The primitive nature of the cells in the present experiment may have been due to their age, their unsaturated fatty acid environment, or other factors. The difference in serum fatty acids was possibly an important factor determining the type of lesion produced, since both the peanut oil- and butter-fed rats had much the same degree of hyperlipemia as shown by similar serum cholesterol and triglyceride levels. Both the peanut oil- and butter-fed rats showed a sharp drop in the relative percentage of serum arachidonic acid, even though the peanut oil-fed group had excessively high levels of serum linoleates. The absolute level of serum arachidonic acid, however, changed very little.

Increased 3H-thymidine incorporation into endothelial cells of swine fed cholesterol for 3 days☆

Segments of abdominal aortas from swine fed either a chlesterol or a control diet for 1 or 3 days were incubated for 20 minutes in a medium containing 3H-thymidine. En face preparations of endothelium were then made and labeling indices determined. At 3 days the labeling indices were 2- to 3-fold greater in cholesterol-fed than in control swine.

Alterations in population dynamics of arterial smooth muscle cells during atherogenesis: I. Activation of interphase cells in cholesterol-fed swine prior to gross atherosclerosis demonstrated by “postpulse salvage labeling”

Abstract The effect of a high-cholesterol diet on the cell dynamics of swine aortic smooth muscle was investigated by pulse labeling in vivo with 8 H-thymidine autoradiography, and the determination of nuclear grain-count distributions. The shift in grain-count distributions with time after labeling was analyzed with the aid of a mathematical model. The conclusions inferred were that: (1) the initially labeled cells virtually all completed their partial cell cycle from S phase through mitosis within 2 days, and gave no evidence of further division up to 30 days; (2) control and cholesterol-fed animals did not differ in this respect; and (3) the known effect of cholesterol diet on labeling index is accounted for by newly recruited G 0 cells or by acceleration of slowly moving G 1 cells. This was demonstrated by postpulse labeling, presumably resulting from reutilization of label salvaged from dead cells in the intestine and elsewhere which continued for at least 7 days. These conclusions were supported by estimation of the G 2 population withthe aid of microspectrophotometry, by a detailed analysis of grain counts and mitoses during the first 2 days with the help of successive 5-hour colchicine collections, and by a comparison of labeling in carotid artery cells in which one artery was excluded from the systemic circulation during the initial pulsing period and subsequently readmitted to the circulation. Since no observations were made beyond 30 days, it was not possible to decide whether the diet ultimately did affect the generation time of the pulse-labeled cells, nor whether the cells newly recruited by the diet resulted from activation of G 0 cells or from acceleration of G 1 cells. However, it was possible with the help of preliminary data on the S period and some further mathematical analysis to estimate a range of values for the generation time and for the fraction of cells in G 0 .

Population dynamics of arterial smooth muscle cells: III. Inhibition by aortic tissue extracts of proliferative response to intimal injury in hypercholesterolemic swine☆

Abstract Previously we demonstrated in swine that a single ip injection of an aqueous extract of swine aortic tissue (containing less than a mg of protein) would inhibit entry of arterial smooth muscle cells into mitosis for at least 2 hr. In the current study we investigated the effect of repeated injections of aortic extract for periods up to 104 days on the development of intimal hyperplasia resulting from a combination of hypercholesterolemia and trauma producd by an intraarterial balloon. Quantitatively both the extent and the thickness of intimal lesions were found to be significantly less in aortic extract treated swine than in paired controls given only saline. Qualitative features did not appear to differ and no general or local toxic effects were observed.

Geographic Pathology of Myocardial Infarction

Abstract Myocardial infarction is common in the United States and rare in certain parts of the Orient and Africa. Such geographic variations in occurrence rates of myocardial infarction could be due either to genetic factors or to environmental factors such as diet, climate and socioeconomic factors. If the genetic factor is the main factor responsible for the apparent geographic differences in the occurrence rates of myocardial infarction, the Orientals and Negroes who immigrated to the United States should maintain their low occurrence rates at least to some extent. On the other hand, if environmental factors are mainly responsible for variations, the occurrence rates of myocardial infarcts in those people should have changed considerably when they moved into an entirely different environment. To investigate the importance of genetic compared to environmental factors, we have analyzed all death certificates, autopsy data from 17 general hospitals and from coroners in San Francisco, and from 3 general hospitals in Los Angeles for the period 1956 to 1960. Detailed information was obtained on all Orientals, including Chinese, Japanese and Filipinos. All the autopsied Oriental cases were compared with age-sex matched cases from the white populations. Similar information was obtained from Japan, Korea, Uganda and Nigeria, and from Negro and white groups in New Orleans and Albany. The average length of residence of the Orientals in San Francisco and Los Angeles was approximately 30 years, and the occurrence rates of myocardial infarction in those Orientals were much higher in all age groups than in the Orientals in their home lands, and approached those of the whites. In the male 50 to 59 age group which included substantial numbers of cases for meaningful comparisons, the occurrence rates of myocardial infarction were similar in both racial groups. This suggests that environmental factors seem to play a major role among the Orientals in the etiology of myocardial infarction and coronary thrombosis, and the genetic factor plays a relatively minor role, if any. In the Uganda and Nigeria autopsy series, myocardial infarction was almost nonexistent. Negroes in the United States have lived in this country for many generations and the occurrence rates of myocardial infarction among them are significantly greater than among Negroes in Africa and approach the occurrence rates of whites. As with the Orientals, this suggests that environmental factors are more important in the etiology of myocardial infarction than genetic factors in Negroes in the United States. However, definitive conclusions cannot be drawn for many reasons including the fact that whites, Orientals and Negroes do not represent pure genetic strains since all three groups have immigrated to the United States only a few generations ago from diverse regions and have not remained as sharply defined races.

Geographic pathology of arteriosclerosis: A study of the “critical level” of dietary fat as related to myocardial infarction in Koreans☆

Abstract It is well established that people whose diet contains little fat (10% or less by calories) have low serum lipid levels and a very low incidence of myocardial infarction. By contrast, people whose diet contains much fat (40% or more), such as most Americans, have higher serum lipid levels and a relatively high occurrence rate of myocardial infarction. Whether or not the relationships between dietary fat and myocardial infarction is one of cause and effect, it seems probable that at some as yet-to-be determined critical level of dietary fat above 10%, infarcts begin to appear, possibly to increase in frequency in proportion to an increase in fat, or possibly to reach a point beyond which no further increase in frequency of infarcts occurs, regardless of diet. Although it is not difficult to find peoples at the dietary extremes, with diets containing less than 10% or more than 40% fat, it is difficult to establish clear-cut dietary groups that lie in between and from whom information as to a possible critical level of dietary fat might be obtained. Our studies in three socio-economic groups of Koreans have shown that dietary fat intake is 7% of calories in farmers, 13% in middle class and 17% in upper class people. Myocardial infarcts are practically nonexistent in lower economic classes but do occur in small numbers in higher economic classes suggesting the possibility that a “critical level” of dietary fat at or near 17% may exist beyond which myocardial infarcts become increasingly common. All the individuals studied were well nourished and in apparent good health. Although this study allows for tentative opinions or hypotheses regarding critical levels of fat, it is apparent that more information is needed before definitive conclusions are made. Among other things, we would like to have true cross sections of the population groups in which we are interested with selection based on census rolls; dietary histories checked for accuracy by sample “weigh-backs”; information regarding chemical content of foods based on actual analyses instead of on standard tables; autopsy data on status of coronary arteries on at least some of the subjects of this study. The implications of the information already obtained would appear to warrant further investigation along these lines.

DIETARY LIPID AND THROMBOSIS: STUDY OF "TRIGGER" MECHANISM FOR THROMBOSIS IN STOCK FED RATS.

Abstract Wessler and his associates induced stasis thrombosis and shortening of both glass and silicone clotting times in rabbits by systemic infusion of serum, and concluded that these changes might be caused by activation of factors XII, XI, and IX. Serum has been known for many years to produce platelet aggregation and fusion in vitro . Recently, adenosine diphosphate was demonstrated to induce aggregation and fusion of platelets in vitro in the native plasma. In order to obtain further information on the initial phase of coagulation, two experiments were carried out using rats. In the first experiment we attempted to investigate the effect of serum infusion on in vitro and in vivo clotting, and in the second we investigated the effect of serum and ADP infusion on in vitro and in vivo platelet aggregation and clotting. In rats, intravenous serum infusion resulted in stasis thrombi and shortening of silicone and glass clotting times, as in rabbits, but dose requirements per unit body weight were much greater. With a reduced dose, stasis thrombi could be produced without shortened silicone or glass clotting times. This suggests that something other than, or in addition to, factors XII, XI, and IX may be responsible for the production of the serum-induced hypercoagulable state in rats. When serum was infused, a significant reduction in platelet counts and platelet aggregation in the native plasma was demonstrated. Infusion of ADP solution alone also produced not only significant reduction in platelet counts but also a state of hypercoagulability manifested by marked reduction of both glass and silicone whole-blood clotting times and production of stasis thrombi. Furthermore, the native plasma clotted in most instances during the centrifugation at 4°C and not after infusion of serum in doses currently used. The mechanism involved in the production of such hypercoagulable state after infusion of ADP is not clear, but we believe massive aggregation of platelets is an important factor. Electron micrographs taken from a stasis thrombus after serum and ADP infusion were similar and demonstrated many areas of platelet aggregation and fusion amidst red cells; scattered strands of fibrin were also seen. The effects of infusion of serum and ADP solution on platelet aggregation and fusion, whole-blood clotting times, and development of stasis thrombi were similar in the current experiment, suggesting that at least platelets were similarly involved in the two conditions.

Geographic Pathology of Myocardial Infarction: Part I. Myocardial infarction in orientals and whites in the United States; Part II. Myocardial infarction in orientals in Korea and Japan; Part III. Myocardial infarction in Africans in Africa and negroes and whites in the United States; Part IV. Measurement of amount of coronary arteriosclerosis in Africans, Koreans, Japanese and New Yorkers☆

Abstract Myocardial infarction is common in the United States and rare in certain parts of the Orient and Africa. Such geographic variations in occurrence rates of myocardial infarction could be due either to genetic factors or to environmental factors such as diet, climate and socioeconomic factors. If the genetic factor is the main factor responsible for the apparent geographic differences in the occurrence rates of myocardial infarction, the Orientals and Negroes who immigrated to the United States should maintain their low occurrence rates at least to some extent. On the other hand, if environmental factors are mainly responsible for variations, the occurrence rates of myocardial infarcts in those people should have changed considerably when they moved into an entirely different environment. To investigate the importance of genetic compared to environmental factors, we have analyzed all death certificates, autopsy data from 17 general hospitals and from coroners in San Francisco, and from 3 general hospitals in Los Angeles for the period 1956 to 1960. Detailed information was obtained on all Orientals, including Chinese, Japanese and Filipinos. All the autopsied Oriental cases were compared with age-sex matched cases from the white populations. Similar information was obtained from Japan, Korea, Uganda and Nigeria, and from Negro and white groups in New Orleans and Albany. The average length of residence of the Orientals in San Francisco and Los Angeles was approximately 30 years, and the occurrence rates of myocardial infarction in those Orientals were much higher in all age groups than in the Orientals in their home lands, and approached those of the whites. In the male 50 to 59 age group which included substantial numbers of cases for meaningful comparisons, the occurrence rates of myocardial infarction were similar in both racial groups. This suggests that environmental factors seem to play a major role among the Orientals in the etiology of myocardial infarction and coronary thrombosis, and the genetic factor plays a relatively minor role, if any. In the Uganda and Nigeria autopsy series, myocardial infarction was almost nonexistent. Negroes in the United States have lived in this country for many generations and the occurrence rates of myocardial infarction among them are significantly greater than among Negroes in Africa and approach the occurrence rates of whites. As with the Orientals, this suggests that environmental factors are more important in the etiology of myocardial infarction than genetic factors in Negroes in the United States. However, definitive conclusions cannot be drawn for many reasons including the fact that whites, Orientals and Negroes do not represent pure genetic strains since all three groups have immigrated to the United States only a few generations ago from diverse regions and have not remained as sharply defined races.