Cholesterol biosynthesis in human fetal liver and adrenal.

Hepatic and adrenal cholesterol biosynthesis were investigated in the human fetus in vitro by measuring the incorporation of acetate2-14C into cholesterol by slices of liver and adrenals from the same fetus. To differentiate neutral lipids from cholesterol biosynthesis, the radioactivity was measured in the nonsaponifiable neutral lipid fraction and in cholesterol isolated and purified as its crystalline 5,6-dibromo-derivative. Expressed in m~noles acetate incorporated/hr/g tissue into neutral lipids (n.l.) and cholesterol, the values were: fetus I (ii wk. old), liver ( g4.4 ),, 49.1; adrenals (48.8), 9.3; fetus II (14 wk. old), liver (91.3), 47.3; adrenals (15.6), 5.5, respectively. Liver and adrenal cholesterogenesis was inhibited by a specific inhibitor Ltr~n$-l,4-b~(2-chlorobenzylaminomethyl)cyclohexane dihydrochloride = AY-9944] of 7-dehydrocholesterol A7-reductase. This finding provided (indirect) evidence that 7-dehydrocholesterol is an obligatory intermediate in the biosynthesis of cholesterol in both of these organs. The percent incorporation of acetate into cholesterol in the human fetal liver was greater than that in any other tissue previously reported. 2 S T E R O I D S 18:1 We have previously shown that cholesterol, perfused to the midterm (2) and term (3) human placenta in situ can be utilized in the synthesis of neutral steroids. Recently, Hellig et al (4) have demonstrated that circulating cholesterol (primarily of maternal origin) is utilized for most if not all of placental progesterone biosynthesis in pregnancies marked by anencephalic fetuses. However, the de novo synthesis of cholesterol by the placenta is minimal at best (5). In contrast, we and others have shown that the human midterm fetal testis has the capacity for de novo synthesis of free steroids; whereas, the fetal adrenal synthesizes both free and conjugated steroids (6-8). It seemed of interest to report our studies on the relative capacities of the human fetal adrenal and liver to synthesize cholesterol. Studies were also performed to assess indirectly whether 7-dehydrocholesterol is an obligatory intermediate in the biosynthesis of cholesterol in these organs. MATERIALS AND METHODS Human male fetal livers and adrenals were obtained following hysterotomy for therapeutic termination of pregnancy. The adrenals and livers were freed of extraneous tissue, blotted, and slices prepared within 15 min. of operation with a Stadie-Riggs microtome. Both adrenal and liver slices were preincubated (30 min.) and incubated (3 hr.) in Krebs-Ringer bicarbonate buffer (with added glucose), based on earlier studies on cholesterogenesis in bovine adrenals (9). Cholesterol was isolated (9) by the addition of I00 mg carrier, brominated (i0,ii) and the resulting 5,6-dibromocholestan-3~-ol recrystallized (4 times from ethyl acetate-methanol) to radiochemical purity. To assess whether 7-dehydrocholesterol is an obligatory intermediate in cholesterol biosynthesis in these organs, identical studies, from the same fetal specimens, were carried out in the presence of trans-l,4-bis (2-chlorobenzylaminomethyl) cyclohexane dihydrochloride (AY-9944), ixl0-4M final concentration. July 1971 s T E R O I D S 3