title: Sources of cholesterol for testosterone biosynthesis in murine Leydig cells.
abstract: The sources of cholesterol for testosterone production were investigated in freshly isolated murine Leydig cells. In vitro stimulation with human CG (hCG) (0.2 IU/ml) caused a 75-fold increase in testosterone production. Leydig cells contained approximately equal amounts of free and esterified cholesterol (7.8 vs. 8.7 micrograms/mg protein). The total cholesterol content of cells stimulated for 4 h with hCG was significantly decreased compared with unstimulated cells (8.4 vs. 17.6 micrograms/mg protein); both free and esterified cholesterol decreased by about 50%. In unstimulated Leydig cells incubated with [14C]acetate for 12 h, the majority of incorporated [14C] was found in free and esterified cholesterol, whereas, in the hCG-stimulated cells, 80% of incorporated 14C was in testosterone. The activity of hydroxymethylglutaryl-coenzyme A (HMG-CoA) reductase after 4 h in hCG-stimulated cells was 20% higher than in unstimulated cells (115.5 vs. 84.4 pmol/mg protein.min). However, by 6 h, HMG-CoA reductase activity doubled in the hCG-stimulated compared with unstimulated cells. By 12 h, HMG-CoA reductase activity in hCG-stimulated cells was 4 times the preincubation level and 8 times the 12-h level in unstimulated cells. HMG-CoA reductase activity induced by hCG was blocked by aminoglutethimide, an inhibitor of the cholesterol side-chain cleavage enzyme. Lovastatin, a potent inhibitor of HMG-CoA reductase, had no effect on unstimulated or hCG-stimulated testosterone production during a 12-h incubation. Murine high density lipoproteins (mHDL) increased HMG-CoA reductase activity in both unstimulated (29%) and hCG-stimulated (20%) cells. During a 6 h incubation, mHDL increased hCG-stimulated testosterone production by 20%, but had no effect on unstimulated testosterone production. These results suggest that murine Leydig cells store enough cholesterol and cholesteryl esters to support testosterone production for at least 12 h in vitro. Although mHDL does not have a major stimulatory effect on testosterone biosynthesis, it may be involved in the regulation of de novo cholesterol synthesis.
authors.affiliation: Department of Biochemistry, Emory University, Decatur, Georgia.
authors.forename: J W
authors.initials: JW
authors.lastname: Hou
authors.forename: D C
authors.initials: DC
authors.lastname: Collins
authors.forename: R L
authors.initials: RL
authors.lastname: Schleicher
mesh_terms: D000616:Aminoglutethimide / Q000494:pharmacology; D000818:Animals; D002784:Cholesterol / Q000378:metabolism*; D008076:Cholesterol, HDL / Q000494:pharmacology; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism; D007985:Leydig Cells / Q000378:metabolism*; D008148:Lovastatin / Q000494:pharmacology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D013261:Sterols / Q000378:metabolism; D013739:Testosterone / Q000096:biosynthesis*
chemical_list: D008076:Cholesterol, HDL; D013261:Sterols; D000616:Aminoglutethimide; D013739:Testosterone; D002784:Cholesterol; D008148:Lovastatin; D006903:Hydroxymethylglutaryl CoA Reductases