Search

Increased atherosclerosis risk in hyperlipidemic patients may be a result of the enhanced oxidizability of their plasma lipoproteins. We have previously shown that hypocholesterolemic drug therapy, including the 3-hydroxy-3-methyl-glutaryl CoenzymeA (HMG-CoA) reductase inhibitors, and the hypotriglyceridemic drug bezafibrate, significantly reduced the enhanced susceptibility to oxidation of low density lipoprotein (LDL) isolated from hyperlipidemic patients. Although this antioxidative effect could not be obtained in vitro with all of these drugs, the active drug metabolites, which are formed in vivo, could affect lipoprotein oxidizability. We thus sought to analyze the effect of atorvastatin and gemfibrozil, as well as specific hydroxylated metabolites, on the susceptibility of LDL, very low density lipoprotein (VLDL), and high density lipoprotein (HDL) to oxidation. LDL oxidation induced by either copper ions (10 microM CuSO4), by the free radical generator system 2'-2'-azobis 2-amidino propane hydrochloride (5 mM AAPH), or by the J-774A.1 macrophage-like cell line, was not inhibited by the parent forms of atorvastatin or gemfibrozil, but was substantially inhibited (57-97%), in a concentration-dependent manner, by pharmacological concentrations of the o-hydroxy and the p-hydroxy metabolites of atorvastatin, as well as by the p-hydroxy metabolite (metabolite I) of gemfibrozil. On using the atorvastatin o-hydroxy metabolite and gemfibrozil metabolite I in combination an additive inhibitory effect on LDL oxidizability was found. Similar inhibitory effects (37-96%) of the above metabolites were obtained for the susceptibility of VLDL and HDL to oxidation in the oxidation systems outlined above. The inhibitory effects of these metabolites on LDL, VLDL, and HDL oxidation could be related to their free radical scavenging activity, as well as (mainly for the gemfibrozil metabolite I) to their metal ion chelation capacities. In addition, inhibition of HDL oxidation was associated with the preservation of HDL-associated paraoxonase activity. We conclude that atorvastatin hydroxy metabolites, and gemfibrozil metabolite I possess potent antioxidative potential, and as a result protect LDL, VLDL, and HDL from oxidation. We hypothesize that in addition to their beneficial lipid regulating activity, specific metabolites of both drugs may also reduce the atherogenic potential of lipoproteins through their antioxidant properties.

In the current studies we describe the effects of PD 72953 and related compounds on lipoprotein levels in chow-fed male rats. After 2 weeks, 10 mg/kg of PD 72953 daily was as effective as 100 mg/kg gemfibrozil for elevating HDL-cholesterol. At 100 mg/kg, PD 72953 further elevated HDL-cholesterol to 232% of control levels, and was associated with increased HDL size and plasma apoE (169% of control), despite no change in hepatic apoE mRNA. ApoA-I rose transiently (at 1 week), but by 2 weeks only apoE remained elevated. PD 72953 dose-dependently reduced plasma apoB, VLDL-cholesterol, LDL-cholesterol, and triglyceride. Hepatic apoC-III mRNA reduction parallelled triglyceride lowering. After 1 week, 30 and 100 mg/kg per day PD 72953 reduced plasma apo-CIII levels by 30 and 34%, and triglycerides by 60 and 83%, respectively. PD 72953 treatment had no effect on triglyceride production rates; however, 125I-labeled VLDL apoB disappearance was enhanced. We compared PD 72953 to a structurally similar diacid, PD 69405, that also reduced VLDL and LDL, but had no effect on HDL elevation. Compared to PD 72953, PD 69405 further accelerated 125I-labeled VLDL apoB disappearance, decreased triglyceride production, and elevated the ratio of post-heparin hepatic to lipoprotein lipase activity. Whole animal studies, transient transfection studies in HepG2 cells, and chimeric receptor studies in kidney 293 cells suggest that PD 72953 is a ligand for the peroxisomal proliferation activated receptor alpha (PPARalpha), and PPARgamma. Overall, PD 72953 may act through a peroxisomal proliferation activated receptor and result in plasma triglycerides and apoB-containing lipoprotein reduction, while also raising HDL cholesterol. Reduced apoC-III may allow triglyceride-rich remnants to more efficiently bind and present substrate to peripheral tissue lipoprotein lipase, and therefore allow enhanced shedding of remnant phospholipid surface for HDL production.

Chow and sucrose-fed rats were used as animal models to study the dose-responses of bezafibrate and gemfibrozil in normolipidemic and hypertriglyceridemic states, respectively. Although both drugs lowered plasma triglycerides (TG) to about the same extent in chow-fed rats, gemfibrozil lowered liver TG as well as plasma total and LDL-cholesterol (LDL-C), but elevated HDL-cholesterol (HDL-C) and plasma apo E concentrations. Bezafibrate produced opposite effects, namely, decreased HDL-C, apo E and liver TG, and tended to increase LDL-C. TG lowering for both drugs in chow-fed rats was not due to changes in TG secretion (production) in normal rats but was associated with enhanced LPL activity. In hypertriglyceridemic rats both drugs modestly reduced TG secretion rates about 40% at a dose producing maximal TG lowering, but again, gemfibrozil elevated and bezafibrate lowered HDL-C and apo E. Unlike gemfibrozil, bezafibrate induced the appearance of LDL-C in hypertriglyceridemic rats which was not detected in control animals, and also tended to increase rather than decrease plasma apo B levels. Finally, changes in liver TG concentration (mg/g) in hypertriglyceridemic rats were opposite for these drugs, resulting in significant drug-related differences in liver TG content (mg/organ). From these data we postulate that, although similar with regard to TG lowering activity and mechanisms thereof, gemfibrozil and bezafibrate produce fundamentally different effects on LDL, HDL and apolipoprotein metabolism (apo B and apo E) in rats which may relate to potential differential effects on reverse cholesterol transport and atherogenesis.

For the last 30 years fibrates have been widely prescribed to treat human dyslipidemia. However, the primary mechanism by which they lower plasma lipid levels is still unknown. Studies with transgenic mice have suggested that changes in apoC-III expression levels have a dramatic influence on plasma triglyceride levels. These results suggested that fibrates could reduce lipid levels by lowering apoC-III gene expression. In the current studies, we sought to determine whether the selected fibrates, bezafibrate, clofibrate, fenofibrate, and gemfibrozil, could reduce hepatic apoC-III mRNA and plasma apoC-III levels. Chow-fed rats were orally gavaged daily with a dosing vehicle alone or with 100 mg/kg of each of the fibrates for 1 week and in addition with gemfibrozil for 2 weeks. Bezafibrate and fenofibrate lowered plasma triglyceride by approximately half and dramatically reduced hepatic apoC-III mRNA and plasma apoC-III levels. In contrast, clofibrate did not reduce plasma triglyceride levels and only partially reduced apoC-III mRNA and plasma protein levels. Gemfibrozil strongly reduced plasma triglyceride levels and had an intermediate but significant effect on apoC-III mRNA and plasma apoC-III levels. Some of the fibrates, especially gemfibrozil also reduced plasma apoC-II levels, an effect that could contribute to the observed triglyceride-lowering effect. In addition, the ratio of plasma apoE to plasma apoC-II plus apoC-III was strongly and inversely correlated with plasma triglyceride levels. As plasma apoE levels were not reduced in gemfibrozil-treated animals, this could also have contributed to the triglyceride-lowering effect of this fibrate. Fibrate-mediated triglyceride lowering was not the result of a decreased apoB or VLDL production and, therefore, suggested an enhanced VLDL remnant catabolism. Our results suggest that the mechanism by which fibrates lower plasma triglycerides is by reducing the level of hepatic apoC-III expression.

In vitro data suggests that lifibrol lowers plasma cholesterol by inhibiting cholesterol synthesis. We report that lifibrol is far less potent in vitro and in vivo than lovastatin for inhibiting 14C-acetate incorporation into sterols. Moreover, several major differences between lifobrol and lovastatin were noted in various animal models. In contrast, lifibrol exhibited several activities in common with gemfibrozil, another phenoxy-acid-type drug. Specifically, in normal rats lifibrol, like gemfibrozil, lowered plasma non-HDL-cholesterol and triglycerides, and increased liver weight and hepatic peroxisomal marker enzyme activities. Lovastatin only lowered plasma triglycerides. In cholesterol-fed rats lifibrol and gemfibrozil lowered non-HDL-cholesterol and elevated HDL-cholesterol while lovastatin was inactive. Finally, lovastatin but not lifibrol exhibited hypocholesterolemic activity in normal guinea pigs and resin-primed dogs. Our interpretation is that these data do not support the notion that lifibrol lowers plasma cholesterol in vivo by inhibiting cholesterol synthesis.

Due to the potential importance of acyl-coenzyme A:cholesterol O-acyltransferase (ACAT) in the generation of lipid-filled monocytes-macrophages, the ACAT inhibitor CI-976 (2,2-dimethyl-N-(2,4,6-trimethoxyphenyl)dodecanamide) was evaluated relative to selected lipid-lowering agents for their effect on atherosclerotic lesion regression and progression. Atherosclerotic lesions comparable in composition to human fatty streaks were induced by chronic endothelial denudation in the iliac-femoral artery of hypercholesterolemic New Zealand White rabbits before intervention, while naturally occurring fatty streaks developed in the thoracic aorta. CI-976 administered in a hypercholesterolemic diet at a dose that did not lower plasma cholesterol prevented the accumulation of monocytes-macrophages within the preestablished iliac-femoral lesion and reduced the foam cell area by 27-29% relative to the initiation of intervention. CI-976 also blunted the development of thoracic aortic fatty streak-like lesions and decreased the cholesteryl ester enrichment by 46%. CI-976 had no effect on plasma triglycerides and, more importantly, had no effect or decreased liver, iliac-femoral, and thoracic aortic free cholesterol content. Dietary intervention alone increased monocyte-macrophage involvement in the iliac-femoral lesion despite reductions in plasma, liver, and thoracic aortic cholesterol content. Conventional lipid-lowering therapy such as cholestyramine or cholestyramine/niacin required substantial decreases in plasma cholesterol levels to achieve comparable vascular changes. We conclude that inhibition of ACAT within the arterial wall by the potent and specific ACAT inhibitor CI-976, even in the absence of plasma cholesterol lowering, can result in the inhibition of atherosclerotic lesion progression and can enhance regression.

Gemfibrozil (Lopid) is a new plasma lipid-regulating drug that decreases very low and low density lipoprotein (VLD/LDL) and increases high density lipoprotein (HDL) concentrations in man. The present experiments tested the effects of gemfibrozil on plasma lipoproteins and apolipoproteins in rats fed high fat/high cholesterol diets. Compared to chow-fed rats, cholesterol feeding for 2 weeks (20% olive oil/2% cholesterol) produced the expected increases in VLDL and intermediate density lipoprotein (IDL) while lowering plasma HDL. This was documented by using three methods of lipoprotein isolation: sequential ultracentrifugation, density gradient ultracentrifugation, and agarose gel filtration. Gemfibrozil gavaged at 50 mg/kg per day for 2 weeks during cholesterol feeding prevented these changes such that lipoprotein patterns were similar to those in chow-fed animals. Whole plasma apoE and apoA-I concentrations were decreased and apoB increased due to cholesterol feeding as determined by electroimmunoassay, but again gemfibrozil treatment prevented these diet-induced alterations. Gradient polyacrylamide gel electrophoresis patterns of the total d less than 1.21 g/ml lipoprotein fractions reflected the changes in apolipoprotein concentrations and further demonstrated a greater increase of apoBl compared to apoBh in cholesterol-fed rats. Gemfibrozil lowered the concentration of both apoB variants and prevented the shift of apoE from HDL to lower density lipoproteins. Changes in the distribution of apoE were confirmed using agarose gel column chromatography followed by electroimmunoassay. These methods also revealed a shift of apoA-IV from HDL to the d greater than 1.21 g/ml, lipoprotein-free fraction with gemfibrozil treatment when blood was taken from fasted or postabsorptive animals. Since it was also noted that in chow-fed rats more apoA-IV was present in the d greater than 1.21 g/ml fraction in the postabsorptive or fed state compared to fasted animals, it could be postulated that the shift of apoA-IV into this fraction in gemfibrozil-treated rats is related to an accelerated clearance of chylomicrons. It is concluded that gemfibrozil largely prevents the accumulation of abnormal lipoproteins in this model of dyslipoproteinemia, and that apoE may play a critical role in this normalization process.