The rate of de novo synthesis of cholesterol by human fetal adrenal tissue was computed from the rate of incorporation of tritium from [3H]water into cholesterol. Cholesterol biosynthesis in this tissue was stimulated by ACTH, as evidenced by 1) the incorporation of tritium from [3H]water into cholesterol, 2) the incorporation of [14C]acetate into cholesterol, and 3) the specific activity of 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase in microsome-enriched preparations from fetal adrenal tissue. ACTH also stimulated the incorporation of tritium from [3H]water into triglycerides but not the incorporation of [14C]oleic acid into triglycerides. When the specific activity of HMG CoA reductase was determined in microsomes prepared from tissue homogenized in the presence of NaF (50 HIM), the activity was 0.3–0.6 times the specific activity in microsomes prepared from tissue homogenized in the absence of NaF, regardless of whether the tissue was fresh or had been maintained in organ culture for 3 days in the presence or absence of ACTH. The specific activity of HMG CoA reductase that was most consistent with the estimated rate of cholesterol biosynthesis, determined using the technique of incorporation of tritium from [3H]water into cholesterol, was that observed in microsomes prepared from tissue homogenized in the presence of NaF. Based on these results, we suggest that much of the HMG CoA reductase in the human fetal adrenal gland may be in an inactive form, whether or not the tissue is stimulated by ACTH. Based on a comparison of the rate of de novo synthesis of cholesterol with the rates of secretion of the steroid hormones, dehydroisoandrosterone sulfate and cortisol, it is concluded that de novo cholesterol synthesis could account for the synthesis of some 30% of the daily secretion rate of these steroids.
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