CYTOCHROME P-450(SCC) ACTIVITY AND SUBSTRATE SUPPLY IN HUMAN PLACENTAL TROPHOBLASTS

The degree of saturation of cytochrome P-450(scc) with cholesterol and the substrate turnover number of the cytochrome in cultured trophobla sts and mitochondria from the human placenta were investigated. Choles terol sulfate was found to be a suitable substrate for probing the deg ree of saturation of cytochrome P-450(scc) with substrate during cultu re and in isolated mitochondria, since it enabled the maximum velocity of the cholesterol side-chain cleavage reaction to be estimated. In c ontrast, 25-hydroxycholesterol and low density lipoprotein supported t rophoblast progesterone production at lower rates than that measured w ith saturating cholesterol sulfate. In the absence of exogenous substr ate, the highest rate of progesterone synthesis by trophoblasts was ob served at the beginning of the culture. With cholesterol sulfate as su bstrate, the turnover number of cytochrome P-450(scc) in cultured cell s was 2.8 min(-1) and was not significantly different to the turnover number of the cytochrome for placental mitochondria, where cholesterol is known to be saturating. Results indicate that cholesterol is limit ing for progesterone synthesis in cultured trophoblasts even in the pr esence of lipoprotein rich medium and 8-bromo-cAMP. The concentration of cytochrome P-450(scc) in trophoblasts was only 20% of that measured for placental homogenate suggesting an induction of the cytochrome oc curs when the trophoblasts fuse in vivo to form syncytiotrophoblasts.