Effect of pravastatin, a HMG CoA reductase inhibitor, on blood lipids and aortic lipidosis in cholesterol-fed White Carneau pigeons.

The effect of pravastatin, an inhibitor of HMG CoA reductase, on blood lipids and aortic lipidosis was studied in young cholesterol-fed White Carneau pigeons. The birds were fed with normal ('N group', n = 20) or atherogenic diet (grains + 0.4% cholesterol + 4% lard) alone ('C group', n = 20) and in association with pravastatin ('P group', n = 20). Plasma lipids and aortic intima lipidosis were studied after 3-5 and 8-12 months of the diet. Compared to the N group, pigeons from C group exhibited hypercholesterolemia (TC = 1000 mg/dl) and hyperlipoproteinemia of which level was independent of the duration of the diet. Total VLDL (VLDL+LDL)-cholesterol and apolipoprotein-B levels rose significantly 15, 8 and 4 times, respectively, whereas HDL were increased two times (P < 0.01) in females only. Macroscopically visible intima lipidosis areas covered 40% and 80% of aortic surface after 3-5 and 8-12 months of the diet. In P group, the increase in plasma lipid values was significantly lower than in WC from C group: -40% for total cholesterol (600 mg/dl) (P < 0.01), -71% for VLDL (P < 0.001), -53% for (VLDL+LDL)-cholesterol (P < 0.01) and -54% for apo-B (P < 0.05). HDL remained as high as in C group. Consequently TC/HDL-C ratio was improved and atherogenic risk of cholesterol was reduced by 41% (P < 0.05). Intima lipidosis areas were lowered by 35% (P < 0.01). We conclude that pravastatin treatment involves (1) a decrease in hypercholesterolemia and hyperlipoproteinemia and (2) a lowering in extensiveness and severity of macroscopically visible aortic lipidosis in cholesterol-fed White Carneau pigeon.

Hypertensive arterial disease and atherogenesis. Part. 1. Intimal changes in the old, spontaneously hypertensive rat (SHR).

The histological, ultrastructural and permeability aspects of the intima in 60 70-week-old spontaneously hypertensive (SHR) and Wistar Kyoto normotensive (WK) rats were studied and compared. The intima of aorta, coronary and renal arteries was unequally thickened owing to the smooth muscle cell (SMC) migration and proliferation, blood cell immigration and endothelial cell activation. The present work describes intimal changes that may act as potential atherogenic factors, i.e. hyper-reactivity of endothelial cells; decreased thinness of endothelial cell periphery; reduced intercellular junction pathways; increased quantity of basement lamina and glycosaminoglycan subendothelial material; platelet and monocyte-macrophage infiltration; widened fenestrae in the internal elastic lamina, and smooth muscle cell migration and proliferation. These changes might possibly be able to reduce the atheroresistance of this species by reducing the barrier function, increasing the trapping effect and stimulating smooth muscle cell proliferation and fibrogenesis. They are believed to promote the development of arterial lipidosis when hyperlipemia is an added risk factor.

Tunica media changes in the spontaneously hypertensive rat (SHR).

The histological, ultrastructural, morphometrical and histochemical aspects of the arterial media were studied in young and aged SHR, and compared to normotensive Wistar Kyoto rats. The diffuse thickening was the most characteristic feature of the hypertensive media. It seems due to three processes: Early generalized hypertrophy of smooth muscle cells (smc); connective matrix neogenesis and smc proliferation, more evident in peripheral vasculature. The present paper discusses the following hypertensive tunica media changes in relation to the atherosclerotic process: the decrease in lipolytic esterase and cholinesterase activities; the activation of some lysosomal enzymes; the increase in collagen, glycosaminoglycan and elastin content; the increased media thickness; the modified smc behavior (migration, secretion, proliferation). These alterations might positively influence arterial susceptibility to atherosclerosis through reduced smc lipolytic activity; slowed transmural diffusion; perturbed efflux and aggravated media hypoxia.

[Aortic development and ageing in the rat aorta -- Comparative histochemical and histoenzymological study (author's transl)]. / Développement et sénescence de l'aorte de rat. Histochimie et histoenzymologie comparative.

A comparative study was made of 36 aortic enzyme activities (E.A.) and 6 macromelecular substances at different stages of ontogenesis in 49 male rats. In the foetal aorta (19th day) the E.Z. were moderate or weak and restricted to a few metabolic pathways: glycolysis, diaphorases, esterolysis of some nucleotides and glucosaminoglycan (GAG) metabolism. During the neonatal period (1st-3rd day), the pre-existing E.A. increased; some aerobic and lipolytic activities became histochemically detectable; longitudinal and radial gradients became established. These changes seemed to provide evidence of increased morphogenic activities and metabolic exchanges. During the prepuberty and puberty period (10-20th day-2nd month) all the E.A., as well as metachromasis and pyroniophilia increased, and new E.A. appeared (GluDH, GPDH, 5/Nase, Ac.Pase-Ca++, Mg++, pH 7.2). These changes appeared to be related to cell proliferation and connective tissue increase during the period of fast aortic growth. The increase of some E.A. (Est/ase, Ch. est, ATPase-Ca++, Mg++, 5/Nase, Alk. Pase) suggested a correlation between enzymatic differentiation and hormonal maturation. During adulthood (6-12th month), the E.A. were stable except for 5/Nase, lysosomal and lipolytic activities which increased. Some E.A. were found to be high (diaphorases, glycolytic and esterolytic enzymes), or moderate (aerobic oxidoreductases, lysosome, lipolysis and GAG-linked E.A.), while others were weak or absent (glycogen pathway E.A.). These observations seemed to correlate with synthetic processes and defence mechanisms. Ageing (17th month) was characterized by an increase of metachromatic GAG and acid lipids and by a decrease of pyroninophillia. Lysosome, glycolysis and phospholipogenesis-linked E.A. increased. In some animals (individual reactivity) kreb's cycle and lypolysis-E.A. decreased.

The calcium inhibitor SR33805 reduces intimal formation following injury of the porcine carotid artery.

We studied the effect of SR33805, a calcium channel blocker, in vitro on the proliferation of vascular smooth muscle cells (SMC) stimulated by foetal calf serum, basic fibroblast growth factor and platelet derived growth factor, and in vivo with regard to SMC migration and proliferation which occurred following injury of the porcine carotid artery. The intimal lesion was induced by a silasten collar surgically positioned around the carotid artery and by a stenosis reducing blood flow by 50% for 30 days. Animals received SR33805 (5 mg/kg/day) 8 days before the induction of the lesion and up to 30 days after. In vitro, SR33805 inhibited in a dose-dependent manner growth factor-induced proliferation of SMC (0.20

Development of macroangiopathy in sand rats (Psammomys obesus), an animal model of non-insulin-dependent diabetes mellitus: effect of gliclazide.

The risk of developing macroangiopathy associated with diabetes led us to study in sand rats the long-term consequences of non-insulin-dependent diabetes on the development of arterial lesions promoted by feeding a high-cholesterol diet. Gliclazide, an agent whose preventive effect has previously been suggested in other experimental models of atheroma, was also investigated in these diabetic and hypercholesterolemic animals. Sand rats were fed a natural diet (ND group), a standard laboratory feed (StD group), or a high-cholesterol feed (HCD group) for 15 months. Biologic parameters were monitored throughout the period of the study, and histologic and histochemical examinations were conducted when the animals were killed (month 15). One StD group and one HCD group were treated with gliclazide from month 3 to month 15. The StD group developed a syndrome of obesity, hyperglycemia, hyperinsulinemia, and triglyceridemia. The high cholesterol feed further increased hypercholesterolemia. These biologic abnormalities were accompanied by arterial lesions (thickening of the intima, deposition of glycosaminoglycans). Foam cells were seen in the intima, and microthrombi were present in the lumen of the arteries of animals in the HCD group. Long-term gliclazide medication at doses that normalized serum glucose levels also reduced the obesity, hyperinsulinemia, lipid disorders, and it prevented or retarded the appearance of arterial lesions.

Relation between pancreatic islet cellular infiltration and plasma fibrinogen or alpha 1-acid glycoprotein levels in spontaneously and streptozotocin-diabetic rats: an increase in these protein levels is not necessary for inducing microcirculatory erythrocyte velocity alteration.

Plasma levels of fibrinogen, alpha 1-acid glycoprotein (AG) and albumin, pancreatic insulitis quantitative scores, and erythrocyte velocity in the mesoappendix microvessels were measured in BB diabetic (BBD) and streptozotocin-diabetic rats (WSTZ) in order to answer the following questions: (a) Does hyperfibrinogenemia or increase in AG plasma level occur in BBD and WSTZ rats, and if so, are these alterations secondary to the hyperglycemia or to an inflammatory process such as insulitis? (b) Is there a decrease in microcirculatory flow in the BBD and WSTZ rats, and if so, is it secondary to the hyperfibrinogenemia and/or the hyperglycemia? Insulitis was present in the BBD rats after 5 weeks of disease (with a score of 2.9 +/- 0.1 vs. 1.4 +/- 0.6 in the normoglycemic controls), but absent in WSTZ rats after 5 months of disease (1.2 +/- 0.06 vs. 1.1 +/- 0.06). Increase in fibrinogen and AG plasma levels was observed in the BBD rats only and appears linked to the insulitis. The major acute phase protein AG level is increased in BBD rats already on the first day of appearance of glycosuria. In the WSTZ rats, without insulitis, chronic hyperglycemia alone did not induce an increase in fibrinogen and AG plasma levels. A decreased microcirculatory erythrocyte velocity has been found in both BBD and WSTZ rats. Thus an increase in fibrinogen or AG plasma levels is not necessary for inducing a decrease in erythrocyte velocity. Hyperglycemia is probably the main factor responsible for the decrease in microcirculatory flow in the BBD and WSTZ rats.

Effect of long-term treatment with a purified micronized flavonoid fraction on pancreatic mononuclear cell infiltration in diabetic BB rats.

Bio Breeding (BB) rats develop a genetically determined insulin-dependent diabetes, because of the early destruction of pancreatic beta cells of Langerhans islets, massively infiltrated by inflammatory mononuclear cells. S 5682, registered as Daflon, 500 mg, is a purified micronized flavonoid fraction (90% diosmin, 10% hesperidin), which has been shown to possess antiinflammatory properties, including anti-free radical activity, effects on vascular permeability, venous tone, and perivenous inflammation. We studied the effect of S 5682 on the course of pancreatic insulitis in diabetic BB rats. All the diabetic BB rats were hyperglycemic, with an increase of plasma levels of fructosamine, alpha-1 acid glycoprotein, and fibrinogen, and a dramatic decrease of C-peptide level. These parameters were not modified by S 5682. Pancreas histologic studies showed that in S 5682-treated diabetic BB rats, lymphocytic infiltration of Langerhans islets was less important and frequent than in untreated diabetic BB rats. By quantitative analysis, a highly significant difference was observed for insulitis, as well as perivasculitis, between S 5682-treated and untreated diabetic BB rats. This inhibitory effect of S 5682 on pancreatic mononuclear cell infiltration may be useful for a complementary treatment to decrease the development of insulitis in human insulin-dependent diabetes mellitus.

Metabolic and anti-atherogenic effects of long-term benfluorex in dyslipidemic insulin-resistant sand rats (Psammomys obesus).

Benfluorex is a clinical lipid-lowering agent with antihyperglycemic properties. The effect of long-term oral treatment (10 mg/kg/day for 7.5 months) on carbohydrate and lipid metabolism and aortic morphology was investigated in 24 insulin-resistant sand rats receiving a standard laboratory diet supplemented with cholesterol (2%). Untreated controls (n=34) developed impaired glucose tolerance, hyperinsulinemia, hypertriglyceridemia, hypercholesterolemia and elevated plasma LDL- and VLDL-cholesterol, positively correlated with the proportion of the thoracic aorta displaying oil red O-positive atherosclerosis; ultrastructural examination showed intimal lipid deposits, foam cells, polymorph infiltrates and fibrosis. Benfluorex-treated animals showed significant decreases in glucose intolerance, hyperinsulinemia, hypertriglyceridemia, hypercholesterolemia, and plasma LDL- and VLDL-cholesterol, with no evidence of aortic atheroma. The metabolic benefits of benfluorex may protect against the long-term development of atherosclerosis in the insulin-resistant dyslipidemic syndrome.

Morphometric and metabolic profile of the distal segment of the internal mammary artery: caution on its use for coronary anastomoses.

OBJECTIVE: Elastic arteries were found to be less prone to intimal hyperplasia than muscular arteries. The internal mammary artery (IMA), which is elastic in its proximal segment, presents a gradual decrease of media elastic fibers along its downstream course. Metabolic and morphometric studies of the distal end of the IMA with regard to its local susceptibility to develop intimal changes were undertaken in order to evaluate the reliability of its use as an anastomotic site for bypass grafting. METHODS: Twenty distal segments of IMA were harvested from patients who had undergone myocardial revascularization. Histologic, enzyme-histochemical and morphometric studies were undertaken on these arterial segments. RESULTS: Histologic examinations indicated an elastomuscular structure in 13 patients, a muscular structure in 6 and an elastic structure in 1. Of the 20 IMAs, none was found to have intimal thickening of greater than 25% of the diameter of the lumen. The enzyme-histochemical profile of the proliferating cells found in the intimal thickening differed from normal contractile smooth muscle medial cells in the loss of myosin and mitochondrial ATPase, plasma membrane 5' nucleotidase, moderately decreased aerobic dehydrogenase and increased lactate dehydrogenase activity and ribonucleoprotein-linked pyroninophilia. Lysosomal beta-glucuronidase and sulfatase were strongly active. This enzyme behavior is unfavorable to contractile function and favorable to cell proliferation and lipid accumulation, two events strongly involved in the atherogenic process. CONCLUSION: Intimal proliferative changes were observed in the distal segment of the IMA. Although there was no histologic evidence of atherosclerotic plaque, the enzyme-histochemical profile of this intimal thickening was favorable to cell proliferation and lipid accumulation. These findings suggest that it may be beneficial to avoid coronary anastomoses with the distal end of the IMA and to use a more proximal/elastic segment.

Enzyme cytochemical expression of aortic smooth muscle cell modulation in primary and secondary cultures.

"Contractile" arterial smooth muscle cells (SMC) return to a less differentiated "synthetic" state during adaptative and proliferative processes in vitro and in cell cultures. At present, the enzyme expression of the modulation of cultured SMC is partially unknown. In order to define metabolic events associated with cell modulation in vitro, we studied 16 enzyme activities in primary and secondary (P1-P3-P10) SMC cultures in comparison to in situ SMC in fetal and adult rat aorta. The "contractile" SMC in aorta of 2 months old rat showed very high nucleotide hydrolase activities (5'-nucleotidase, Mg-ATPase, Ca-ATPase), and naphthylesterase activities and weak lysosomal acid phosphatase activity; the glycolysis-linked dehydrogenases were expressed with higher activities than Krebs cycle-linked enzymes. In primary cultures, the SMC near the explant expressed a "contractile-like" enzyme behaviour, in opposite to cells in the peripheral part of growing area enzymatically similar to sub-cultured SMC. Proliferating SMC in secondary cultures were characterized by increased lysosomal activities and by the decrease or disappearance of Ca-ATPase, Mg-ATPase, 5'-nucleotidase, and butyrylcholinesterase activities like fetal SMC in vivo. These enzyme changes in subcultures might be related to a deficiency of nucleotide ester hydrolysis, abnormal adenosine and AMP levels, lowered lipolytic capability and increased lysosomal reactivity. In conclusion, subcultured "synthetic" SMC expressed enzyme cytochemical patterns different from those of "contractile" SMC and similar to those of fetal immature SMC. Their enzyme behaviour is unfavourable to contractile function and favourable to cell proliferation and lipid accumulation, two characteristic features of SMC in atherosclerotic plaques.

Evaluation of cryopreserved arteries as alternative small vessel prostheses.

Biologic or synthetic grafts have had limited success in small vessel applications. Studies were initiated to assess the potential use of cryopreserved (CP) arteries as coronary artery bypass conduits. Sheep carotid arteries (internal diameter: 4 mm; length: 10 cm) were cryopreserved in a nutrient media containing 10% DMSO and were stored in a nitrogen vapor at -150 degrees C. After thawing, histological, enzyme-histochemical and functional studies showed slight histological alterations, preservation of enzymal activities and an abolition of the contractile response. In a sheep model, arterial substitution of a 10 cm segment of carotid artery was realised by implantation of fresh autografts ( n = 4); fresh allografts (n = 9) and CP allografts (n = 9). After 3 months, all autografts were patent with slight histological alterations. Fresh and CP allografts showed similar modifications: patency rate was 7/9 in both groups. Intimal thickening with cell proliferation was seen in fresh (3/7) and CP (4/8) arteries; loss of smooth muscle medial cells was constant. Adventitia was always involved by a marked inflammatory reaction. One characteristic of CP allografts was the frequent presence of large dystrophic calcifications. In conclusion, morphologic and functional arterial changes occurred after freezing and thawing. In spite of vascular rejection, the patency rate of allografts after 3 months of implantation in arterial circulation remained high and does not seem influenced by cryopreservation.

[Hypertensive arteriopathy and atherogenesis: cellular and molecular interactions]. / Artériopathie hypertensive et athérogenèse: interactions cellulaires et moléculaires.

The spontaneously hypertensive rat (SHR)--animal model for human essential hypertension--develops a generalized arteriopathy. The present paper discusses the atherogenic influence of hypertensive arterial lesions. The following changes in the intima might influence its permeability and barrier function, increase the trapping effect and stimulate the smooth muscle cell proliferation: the hyper-reactivity of endothelial cells; the decreased thickness of endothelial cell periphery; the reduced intercellular junction pathways; the increase in basal lamina and glycosaminoglycan sub-endothelial material; the mononuclear cell infiltrations; the widened fenestrae in the internal elastic lamina. Some hypertensive changes of the tunica media may also interact with atherogenic process through reduced smooth muscle cell lipolytic capabilities, slowed transmural diffusion, perturbed efflux, aggravated media hypoxia, namely: the decrease in esterase and cholinesterase activities, the activations of some lysosomal enzymes, the increase in collagen, glycosaminoglycan and elastin content; the increased media thickness and transmural passage; the modified smooth muscle cell behavior.

[Experimental models of atherosclerosis. Contribution, limits and trends]. / Modèles expérimentaux d'athérosclérose. Apports, limites et perspectives.

Experimental approaches to the problem of atherosclerosis involve animal or cellular models and procedures of lesional induction. Relevant animal models are rare. The rat, the mouse and the dog are free of "natural" atherosclerosis and only develop diffuse lipidosis after high cholesterol diet and thyroid block. They are more appropriate models of experimental arteriosclerosis and intimal proliferation induced by different procedures. The rabbit, also free of spontaneous atherosclerosis, is extremely sensitive to lipid-rich diets, but the lesions induced resemble more a xanthomatosis than an atherosclerosis. Immunological procedures in this model result in a generalised immune arteriosclerotic arteriopathy. The monkey and pig, which are phylogenetically close to man, develop spontaneous atherosclerosis exacerbated by lipid-rich diets or other procedures: hormones, psychosocial stress. The cost and problems of upkeep make these two models inaccessible to most laboratories. Although the hen, turkey and pigeon are grain-eating, they develop natural atherosclerosis, are sensitive to atherogenic diets, and provide satisfactory replacement models, especially for research into the viral and tumoral theories of atherogenesis. The pigeon is particularly suitable for studying cellular, biochemical and genetic aspects of atherosclerosis: these spontaneous plaques, similar to those in man, are ontogenetically and topographically predictable. The species include genetic types both sensitive and resistant to the disease. Moderately lipid-rich diets induce lesions even in very young pigeons. They also lend themselves well to the study of the antiatherosclerotic effects of pharmacological agents. Endothelial, smooth muscle and macrophage cell cultures are widely used to study the factors influencing cellular modulation and proliferation, lipid metabolism and movement of cholesterol, cellular biosynthesis and cell-cell and cell-matrix interactions.

[Natural history of the regression of atherosclerosis: from animal models to men]. / Histoire naturelle de la régression de l'athérosclérose: des modèles animaux à l'homme.

Numerous studies carried out on animal models (apes excepted) have given encouraging results as regards the regression of experimental atherosclerosis after return to a normal or hypocaloric diet combined or not with various drugs. Regression is more obvious when lesions are recent and less severe: lipid striae disappear in less than 12 months, whereas more advanced and complicated lesions take years to regress. Intracellular lipids and cell alterations vanish more readily than extracellular lipids and alterations of connective and matrical tissues. Excessive accumulation of collagen accounts for the irreversibility of complicated plaques. Lesions of the intima are less stubborn than those of the media. Involution does not take place at the same time in coronary vessels and in the aorta. In non human primates, however, no noticeable regression is observed before several months if not years. In these animals, the degree and rapidity of involution after return to the normal vegetarian diet depend on the severity of the lesions induced, on the degree of fibrosis, on the level of residual hypercholesterolaemia and on the adjunction to the diet of certain drugs such as cholestyramine or alpha-alpha. The results of therapeutic trials conducted in man have not been so good because the patients treated had old and severe atherosclerosis: after a few years' treatment with low-cholesterol diet and appropriate drugs less than 10 p. 100 of them showed a clear-cut angiographic improvement. It is therefore illusory to rely on spontaneous regression when tackling a case of clinically detectable atherosclerosis. A preventive treatment is more promising, since infraclinical lesions may regress.

[Detection of cardiac graft rejection using proton nuclear magnetic resonance]. / Détection du rejet de greffe cardiaque par résonance magnétique nucléaire du proton.

Rejection of cardiac transplants can be detected by NMR imaging if it is associated with a change in myocardial T1 and/or T2 proton relaxation time. T1 and T2 were studied in 14 Lewis rats that underwent heterotopic cardiac transplantation. T1 and T2 were measured in vitro immediately after sacrifice 3, 4, 7 or 11 days after the graft using a Minispec BRUKER PC20. The myocardial water content was measured by dehydration in a vacuum for 24 hours. Histological analysis of sections classified the rejection process in 4 stages according to the degree of lymphocyte infiltration and percentage of myolysis. There was a significant difference between the ortho and heterotopic hearts: (Formula: see text). In particular, there was highly significant relationship between T2 and the stage of rejection (r = 0.90, p less than 0.005), and between T2 and % myolysis (r = 0.84, p less than 0.005). In addition, there was a close relationship between the T2 of the ortho and heterotopic hearts and their water content (r = 0.95, p less than 0.001). If these results are confirmed in man, is should be possible to detect rejection by NMR imaging using sequences of activation concentrating on changes of T2.

[Atheroma and calcium antagonists]. / Athérome et antagonistes calciques.

Calcic ions play a role in the initial and chronic development of atherosclerosis lesions. Demonstration of anti atherogenic properties of calcium antagonists has open a new therapeutic approach of atherosclerosis and of its complications. Mechanism of protection remains unclear but it is essentially a preventive effect requiring very early administration before onset of lesions. Several experimental studies have shown that calcium antagonists could reduce cholesterol content of arterial wall without any modification of lipid plasma or profile. Several other mechanisms have been proposed but none of them can actually explain the observed preventive effect of these drugs. The clinical relevance in human of such a preventive effect of calcium antagonists requires further investigations.