Figure 4.

Effect of cortisol on steroidogenic enzyme activities in nonadrenal cells engineered to express HSD3B2 or CYP17A1. Representative TLC and quantification of metabolic products of [³H]-DHEA in cells transiently transfected with HSD3B2 (A and B) or of [³H]-pregnenolone in cells transiently transfected with CYP17A1 (C). In the absence of transfection of either gene, no metabolism of the labeled steroid was observed. Substrate-velocity (A) and Lineweaver-Burk (B) plots were used to obtain Michaelis-Menten kinetic constants for 3βHSD2 metabolism of DHEA in the absence and presence of cortisol (50 μm). There was little change in relative maximal velocity, V_max (P = 0.28), and a 3-fold increase in the apparent Michaelis constant, K_m (P < 0.001), with the addition of cortisol, indicating that cortisol is a competitive inhibitor of 3βHSD2. In the representative TLC chromatogram in A, the DHEA substrate concentration was 0.5 μm. C, Increasing amounts of cortisol were associated with small but significant increases in 17αOH-pregnenolone and decreases in DHEA (P = 0.006 and P = 0.003, respectively for effect of cortisol), indicating a modest decrease in 17, 20 lyase activity.