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Background Bezafibrate reportedly lowers serum lipids and plasma fibrinogen in humans and rats, but the mechanism of this process has still to be clarified. We have addressed this problem by investigating the effects of bezafibrate on fibrinogen in the human hepatoma HepG2 cell line. Methods In cells cultured with and without bezafibrate, intra- and extracellular fibrinogen was quantified using an enzyme-linked immunosorbent assay (ELISA), and changes in the biosynthesis and secretion of bezafibrate were monitored by [35S]-methionine labelling. In addition, expression of the corresponding mRNA was investigated by Northern blotting. As fibrinogen is an acute-phase protein, HepG2 cells exposed to interleukin 6 were also analysed. Results Bezafibrate decreased fibrinogen biosynthesis by about 25%, regardless of whether or not the cultures were also exposed to interleukin 6. mRNA was reduced in proportion to the concentration of interleukin 6 to which the cells were exposed, but intracellular transport and release of the protein were not affected. Conclusion Apparently, bezafibrate interferes with the biosynthesis of fibrinogen at the pretranslational level but does not affect the export of the protein.

The effect of the piperazine derivative BM 15.766 (4-(2-[1-(4-chlorocinnamyl)piperazin-4-yl]ethyl]-benzoic acid) on the biosynthesis of sterols was investigated in adult rat hepatocytes in primary monolayer culture. The substance led to a dose-dependent reduction of cholesterol in the serum of various species of animals such as rat, dog and marmoset. BM 15.766 showed a dose-dependent action on the incorporation of 14C-acetate in neutral, nonsaponifiable lipids. The inhibition of the overall incorporation was 10-12% (10(-5) M). No inhibition was observed in the hepatocytes over the entire dose range of 10(-8) M to 2 X 10(-5) M, while the release of the neutral lipids from the hepatocytes into the culture medium was reduced by up to 40%. The biosynthesis of cholesterol could be reduced by more than 90%. Simultaneously, 7-dehydrocholesterol levels rose in the cells and, to a less marked extent, in the medium. This can be interpreted as an indication that 7-dehydrocholesterol is incorporated into the cell membrane, which results in a lower release of 7-dehydrocholesterol into the medium in comparison with controls. The site of attack is the inhibition of the delta 5.7-sterol delta 7-reductase. The formation of desmosterol and cholestatrienol as well as other 7-dehydrocholesterol precursors could also be demonstrated. After longer incubation, there was an additional accumulation of squalene and lanosterol with simultaneous reduction of 7-dehydrocholesterol by BM 15.766, whereas the total 14C-acetate incorporation in neutral lipids was increased.

Rat liver cells derived from male and female animals in primary monolayer cultures were investigated for suitability as a test system for xenobiotics affecting the cholesterol pathway. An appropriate mode of extraction and separation of newly formed cholesterol and precursors is described. This system can be widely applied. Rat liver cells from females in oestrus cycle had a higher synthesis rate of cholesterol than those from males. The disadvantages related to the cycle phases make male rats more appropriate donor animals for the test system developed. The altered in vitro cholesterol synthesis is relevant to that in vivo. The extraction of newly synthesized cholesterol and its precursors by means of columns packed with large-pore kieselgur is precise and time saving. The modified separation by thin-layer chromatography on silica gel layers impregnated with silver nitrate enables direct separation from the extract and is sufficient to recognize cholesterol and its precursors. The method in this form is suitable for processing a large number of specimens e.g. for screening.

Bezafibrate is a potent hypolipidemic agent, which causes marked proliferation of peroxisomes in rat liver. At the same dosage, bezafibrate is more effective in male than in female rats. This is probably related to divergent pharmacokinetics, which cause differences in drug level in serum and liver. The volume density of peroxisomes and several of their enzymes such as carnitine acetyl transferase and acyl-COA oxidase increase in a dose-related fashion. The hypolipidemic effect of bezafibrate, however, does not correlate with the used dosage. This implies that peroxisomal proliferation may play only a minor role in the hypolipidemic action of bezafibrate. In animals treated for 26 months with 300, 750, or 1500 ppm bezafibrate, the relative liver weight and serum triglycerides did not differ significantly from controls. Peroxisomal proliferation varied in different cells, being most prominent in single hepatocytes. The liver catalase activity was significantly reduced, but carnitine acetyl transferase was increased. Abnormal peroxisomes and mitochondria with longitudinal cristae were quite frequent. In one focus, catalase activity was severely diminished ahd peroxisomes were markedly reduced. The incidence of liver tumors was the same (1-3%) in treated animals as in controls.

The pharmacokinetics and pharmacodynamics of BM 13.177 were investigated in eight healthy men who received a single oral dose of 800 mg on the first day and seven equal doses in 8-hour intervals on the second to fourth days. Pharmacodynamic effects were measured ex vivo by the testing of platelet functions such as shape change, aggregation, and [3H] serotonin release. The maximum serum concentration of 6.6 or 6.7 mg/L was achieved within 1.6 hours after the first dose and within 1.5 hours after multiple doses, respectively. Afterwards, BM 13.177 was eliminated in urine with a terminal elimination t½of 0.84 or 1.0 hours after single and multiple dosing, respectively. The inhibition of platelet function showed the same close correlation with the serum concentrations of BM 13.177 after single and after multiple doses. Apparently, BM 13.177 induces neither refractoriness to BM 13.177 nor desensitization of the platelet thromboxane receptor. Because BM 13.177 was also well tolerated without subjective or objective side effects, this drug appears to be useful in evaluating the clinical benefit of thromboxane receptor blockade. Clinical Pharmacology and Therapeutics (1986) 39, 145–150; doi:10.1038/clpt.1986.25