Morphological effects of moderate diet and clofibrate on swine atherosclerosis

Early proliferative coronary atherosclerosis was produced in swine by feeding them a high-fat, high-cholesterol diet for 17 months, at which time one group of animals was killed (reference group), while the remainder was transferred for 12 months to a moderate diet that resulted in serum cholesterol levels of about 190 mg/100 ml. The moderate diet only did not decrease the size of coronary lesions, but prevented their progression. The addition of clofibrate therapy caused regression that involved a significant decrease in size, gross sudanophilia, and extent of calcification and the disappearance of foam-cell lesions. Resultant serum cholesterol levels appear to be more important than the amount of dietary cholesterol in the progression, prevention, and regression of swine coronary atherosclerosis.     

Some effects of a grain-based mash diet on cholesterol metabolism in swine

The aim of the first part of this study was to find out in swine how much fat could be included in a cholesterol-free semi-purified diet without resulting in changes in serum cholesterol levels or hepatic microsomal HMG-CoA reductase activities from those in swine fed a grain-based mash diet. Graded levels of peanut oil from 5 to 40% by calories were included in a casein-carbohydrate-cellulose based diet and effects compared with those in swine fed mash (10% fat) for 2 weeks. Serum cholesterol levels were essentially unaffected by any of the diets. However, in the semipurified diet-fed swine at all levels of dietary fat hepatic microsomal HMG-CoA reductase activities were reduced to values circa 25% of those in the mash-fed swine. This is as great as the reduction from mash levels of activity produced by high fat, high-cholesterol hyperlipidemic diets.The aim of the second part of the study was to find out in terms of whole body cholesterol balance difference in the effects of a cholesterol-free, low-fat (9%) semi-purified diet and a mash diet that might account for the differences in hepatic microsomal HMG-CoA reductase activities. The principal difference proved to be in the fecal output of bile acids (908 ± 86 mg/day on mash vs 292 ± 11 mg/day on a low-fat semi-purified diet). Thus the higher hepatic microsomal HMG-CoA reductase activities in the mash fed swine are presumably in response to the greater loss of steroids. The mixed natural fibers in the grain-based mash diet, as compared to cellulose only in the other, probably accounts for the increased fecal output of bile acids.     

Effect of dietary protein and cholesterol on atherosclerosis in swine

Two groups of pigs, each consisting of six animals, were fed on isocaloric amounts of experimental diet with a high cholesterol content but no added fat and with varying levels of protein (5% vs 25% by weight of the diet) for 16 mo.Animals of the low-protein group were confirmed to have developed a protein-deficiency state by the characteristic microscopic changes in the viscera and hypoproteinemia due to reduced albumin fraction. They had a larger surface area of the aorta involved with atherosclerosis, and the lesions had a higher lipid and cholesterol and lower phospholipid content. The serum cholesterol was significantly higher, and the serum cholesterol esters contained larger proportion of oleic acid at the expense of linoleic acid than the animals of the high protein group.The results indicate that very low levels of dietary protein have a promotive effect on the induction of hypercholesterolemia and atherosclerotic lesions in the presence of cholesterol alone and in the absence of additional fats. The precise mechanism of this variation is not understood.     

Ultrastructural study of hepatic mitochondrial abnormality in swine treated with clofibrate

Administration of clofibrate frequently reduces serum cholesterol levels in hypercholesterolemic swine to near the base level within a few weeks even when the hypercholesterolemic diet is continued. The current study was designed to investigate hepatic ultrastructural changes that might be associated with the hypocholesterolemic effect of clofibrate. Characteristic ultrastructural changes were observed in 17% of hepatocellular mitochondria after 6 wk of clofibrate treatment. The most frequent mitochondrial alteration was the presence of discrete bundles of tubular or spiral structures which appeared at random in orientation. The number of microbodies increased moderately but no structural changes were noted.     

HSP22在高脂致动脉粥样硬化病变中的作用及对他汀干预的影响

目的:研究热休克蛋白22(heat shock protein 22,HSP22)在高脂饮食诱导的小鼠动脉粥样硬化(atherosclerosis,AS)病变中的作用,及其对阿托伐他汀(atorvastatin,Ator)干预的影响。方法:8~9周龄Apo E~(-/-)和HSP22~(-/-)Apo E~(-/-)和HSP22+Apo E~(-/-)雄性小鼠各18只,每种小鼠分为2个亚组,分别为对照组与他汀干预组(Ator组),HSP22~(-/-)组(KO组)与HSP22~(-/-)他汀干预组(KO+Ator组)及HSP22~+组(Tg组)与HSP22~+他汀干预组(Tg+Ator组),各干预组从第5周开始给予Ator(10 mg·kg~(-1)·d~(-1))干预,各对照组给予等量生理盐水,共饲养13周。采用油红O及苏木精-伊红(hematoxylin-eosin,HE)染色法观察AS病变程度,免疫组化法、Western blot法和ELISA法检测主动脉和血清中HSP22、NF-κB、eNOS、ICAM-1及IL-6的表达。结果:油红O及HE染色示Tg组主动脉斑块相对面积低于KO组(P0.05)。KO组的血清和主动脉中HSP22的蛋白表达显著低于对照组和Tg组,对照组显著低于Tg组。Tg组及对照组的主动脉eNOS蛋白的表达显著高于KO组。对照组的主动脉NF-κB及ICAM-1蛋白表达较KO组显著降低,较Tg组显著升高。对照组、KO组及Tg组血清IL-6水平的差异无统计学显著性。结论:HSP22基因缺失可上调NF-κB和ICAM-1的表达,降低eNOS的表达,加速AS的进展;HSP22基因过表达可降低NF-κB和ICAM-1的表达,从而改善AS。HSP22基因缺失部分限制了他汀下调NF-κB和ICAM-1及上调eNOS表达的作用;其过表达可促进他汀下调ICAM-1表达,进一步改善AS。     

The effects of clofibrate ingestion on alveolar macrophage peroxisome content and oxygen metabolism

Respiratory burst activity in alveolar macrophages in response to particulate and soluble challenges, such as zymosan particles and phorbol myristate acetate (PMA), is not nearly as dependent upon membrane stimulation as in neutrophils. Microperoxisomes are subcellular organelles containing catalase and are present in lung macrophages and cells of other organs. Evidence from liver cells indicates that peroxisomes are intimately involved with hydrogen peroxide and lipid metabolism. Clofibrate (2-(p-chlorophenoxy)-2-methylpropionic acid ethyl ester, Atromid-S-^®), a hypolipidemic drug known to cause peroxisomal proliferation in liver cells, was studied with respect to its ability to cause increases in the microperoxisome content and to alter the cellular metabolism of alveolar macrophages. Liver weight increased over a 2-week drug treatment period while lung weight remained unchanged. Plasma triglyceride levels were decreased by the treatment, indicating the effectiveness of the drug. Unlike the effect on liver cells, however, clofibrate did not cause a proliferation of microperoxisomes, as determined by morphometric analysis. Oxygen concumption and hydrogen peroxide generation by alveolar macrophages in response to either stimulant (zymosan or PMA) was no greater in clofibrate-treated rats than in controls. Superoxide release, when expressed as the change in response to PMA, appeared elevated in the drug group; statistical significance, however, was not demonstrated. The hexose monophosphate shunt (HMP), which produces reducing equivalents for lipid biosynthesis, was elevated in macrophages from clofibrate-treated rats when expressed similarly. The significance of these results is discussed in relation to the known effects of the drug on liver cells.     

Dietary magnesium effect on swine coronary atherosclerosis induced by hypervitaminosis D

The effects of magnesium (Mg) supplement on coronary arteries of 61 swine, fed various levels of vitamin D3 (VD3), were studied by light and electron microscopy. High frequencies of smooth muscle cell degeneration were observed in groups of swine fed various levels of excess VD3. Swine fed moderately excessive level of VD3 with a basal level of Mg displayed great incidence and magnitude of intimal thickening, while swine fed the same level of VD3 with supplementary Mg sustained only mild intimal thickening. In groups of swine fed high levels of VD3, the prevention of calcification and smooth muscle cell degeneration was observed as a supplementary Mg effect. Plasma analyses indicated that supplementary Mg with excess VD3 increased cholesterol levels but decreased arterial damage. It is concluded that the dietary Mg supplement prevents coronary atherosclerosis induced by hypervitaminosis D.     

Ultrastructural and elemental analysis of calcification of advanced swine aortic atherosclerosis

During progression and in the early phase on a regression regimen, calcification of the necrotic portion of the atheroma of swine abdominal aorta occurred primarily in degenerated cells or in membranous, vesicular cellular degradation products which varied in size, shape, and the amount of mineral deposit. Calcium appeared to be deposited in amorphous granular or needle-like crystalline forms. Energy dispersive X-ray and line profile analysis showed that the major elements in the heavily calcified portions of the plaques were calcium and phosphorus. There was a direct relationship between the distribution and concentration of these elements indicating that the mineral deposit was a calcium phosphate. Select area electron diffraction analysis of grossly calcified portions of the plaque gave a diffraction pattern identical to that of calcium hydroxyapatite. Calcification was not observed to occur on elastic tissue or collagen fibers.     

Effect of clofibrate,cholestyramine, cholesterol and feeding pattern on the diurnal variation of cholesterol 7 alpha-hydroxylation in swine

A rhythmic change in hepatic microsomal cholesterol 7 alpha-hydroxylase activity in young male Yorkshire swine fed either low-fat, low-cholesterol mash or high-fat, high-cholesterol diets in a single daily meal at 9:00 am has been established. In mash-fed swine, dual peaks were observed; the first peak with 2-fold increase in the enzyme activity over that of 9:00 am at 2:00 pm, and another peak appeared at midnight. In swine fed a high-fat, high-cholesterol diet, the enzyme activity peaked only at 2:00 pm, then the enzyme activity gradually returned to the base line.The total daily activities were not significantly different between mash (one meal) and high-fat, high-cholesterol (one meal) groups. When swine were fed mash in three divided meals daily, the peak seemed virtually to disappear. But the daily total activity integrated for 24 hr was the same as that of swine fed single mash meals.Addition of cholestyramine in mash (one meal) increased amplitude of the diurnal peak, but the rhythmic pattern was not altered. Addition of clofibrate in mash (one meal) swine did not affect the amplitude nor pattern of enzyme activities. Both cholestyramine and clofibrate treatments reduced serum cholesterol concentrations after 2 weeks.     

Lack of effect of clofibrate on hepatic HMG-CoA reductase activity in young swine in the postabsorptive state.

Effects of clofibrate treatment of 7 days duration were studied on hepatic HMG-CoA (β-hydroxy-β-methylglutaryl Coenzyme A) reductase (EC 1.1.1.34) activity and on acetate incorporation into cholesterol by tissue slices from liver and ileum in 29 young male mash-fed swine in the postabsorptive state. Swine treated with either 2 or 10 gm of clofibrate daily showed marked reductions in serum cholesterol levels. Unlike the large decrease observed in rats, hepatic HMG-CoA reductase activity was not significantly reduced in either treated group. Acetate incorporation into cholesterol by liver and ileal slices was similarly unaffected by clofibrate. These results and our previous cholesterol balance studies of clofibrate-treated swine suggest that the drug does not inhibit cholesterol synthesis in these animals. The mechanism of action of clofibrate in swine may be different from that in rats.     

Time-dependent effects of the hypolipidemic agent clofibrate on peroxisomes and mitochondria in mouse hepatocytes

Mice of the NMRI strain were fed a diet containing 0.5% clofibrate up to 25 days to study its effect on peroxisomes and mitochondria in the hepatocytes. Liver samples were analyzed by electron microscopy after cytochemical staining for catalase using diaminobenzidine in order to detect all peroxisomes for a quantitative analysis. The liver weights increased significantly after two days of treatment and reached a maximal level of 40% over the control animals. The number of peroxisomal profiles increased 3- to 4-fold at maximal proliferation and their average area nearly doubled, hence the number of peroxisomes with large diameters was increased making a 5-fold increase within the total cytoplasmic area. The size distribution of the peroxisomes was changed at day two of treatment to a more spread and even distribution going up in a higher size range. The mitochondrial profiles were less affected but there was a slight increase in their number, a small decrease in average area, a transient decrease in axial ratio but no significant effects on their cytoplasmic fractional area. The frequency distribution of mitochondrial area was shifted somewhat to lower values by clofibrate. At the end of the treatment the hepatocytes contained about the same number of peroxisomal and mitochondrial profiles but the latter were still occupying a two-fold higher fractional volume of the cytoplasm. Mouse hepatocytes thus seem to be a good system for studies on peroxisomal biogenesis as is their counterpart in rats, although they differ in their levels of various peroxisomal enzymes.     

Biochemical interaction between effects of beclomethasone dipropionate and salbutamol or formoterol in sputum cells from mild to moderate asthmatics

BACKGROUND: Several in vitro studies demonstrate that corticosteroids and long-acting beta(2) agonists may have a complementary and synergistic mode of action on the inflammatory processes in asthma. METHODS: Sputum was induced in 20 mild to moderate asthmatic patients and the induced sputum cells (ISC) were cultured with beclomethasone dipropionate (BDP) 10(-7) M, salbutamol 10(-8) M and formoterol 10(-8) M either alone or in combination, BDP plus salbutamol and BDP plus formoterol, for 24 h. We measured the levels of growth macrophages-colony stimulating factor (GM-CSF), released on activation normal T cells expressed and activated (RANTES) and interleukin-8 (IL-8), in the supernatant of stimulated cells by ELISA. Furthermore, we assessed nuclear translocation of glucocorticoid receptor (GR) and the expression of beta(2) receptor in ISC by immunofluorescence and RT-PCR, respectively. RESULTS: The release of GM-CSF, RANTES and IL-8 in ISC was significantly reduced by BDP plus salbutamol or formoterol as compared with either drug alone (P < 0.0001). beta(2) receptor expression was increased after 30 min of incubation with BDP and continued to increase over a time period of 4 h (P < 0.0001). Furthermore after 30 min of incubation, nuclear translocation of GR was greater with BDP plus salbutamol or formoterol than with any of the drugs alone (P < 0.0001). CONCLUSION: The present ex vivo study demonstrates a complementary mode of action between BDP and salbutamol or formoterol leading to an enhanced anti-inflammatory activity.     

Experimental atherosclerosis in swine: effect of dietary protein and high fat

Two groups of pigs each consisting of 6 animals were fed for 18 months on isocaloric amounts of an experimental diet with a high fat content and cholesterol but with widely different levels of protein (5% vs. 25% by weight of the diet). In addition, a third group consisting of 4 animals was maintained on normal stock diet to serve as control.Animals of the low protein group showed the maximal intimal surface area involvement with atherosclerotic lesions in the aorta and coronary arteries, and also the most severe among the three groups. No significant differences were noted in the extent and severity of lesions between the high protein-high fat fed animals as compared with the high protein-low fat fed controls. Lesions of the low protein group had a higher cholesterol content and a raised cholesterol:phospholipid ratio than those in the other two groups.Extremely low levels of dietary proteins seem to have had a promotive effect on the induction of atherosclerotic lesions by an atherogenic diet, whereas adequate levels of dietary proteins have had a protective influence. The precise mechanism by which varying levels of dietary proteins have such effects is not understood. It may possibly be related to the aberrations of lipid metabolism induced by extremely low levels of dietary proteins.     

Biochemical effects of corticosteroids on neural tissues

Corticosteroids appear capable of exerting an impressive array of effects on the metabolism of neural tissues. The diversity of these effects is perhaps not surprising given the wide variety of biochemically and morphologically distinguishable cell types present in the combined central and peripheral nervous systems. In conclusion, it seems useful to summarize the state of knowledge in some of the most critical research areas discussed in this review and to predict what major advances are probably forthcoming in the next few years.