Endocrine Abstracts

Placenta is an endocrine organ, secreting steroids (progesterone [P4], estrogens) and hCG, thanks to villous tissu (cytotrophoblasts [VCT] and syncytiotrophoblast [ST]). P4 is required for implantation and maintenance of pregnancy. P4 is synthetized from maternal cholesterol thanks to mitochondrial transporter MLN64 and P450SCC and HSD3B1 enzymes. We aimed to characterize P4 production in human placenta during VCTs differentiation into ST in vitro. VCTs isolated from term placenta (n=6) were cultured for 72h to allow trophoblast differentiation. hCG, P4 and Pregnenolone secretions were measured in supernatants from 24h to 72h by immuno-assay (IA) and/or mass spectrometry (GC-MS/MS). Intracellular expression of transporter (MLN64) and enzymes (P450SCC and HSD3B1) were studied by western-blot and RT-qPCR. The same experiments were performed with 10 μM of Forskolin (FSK, a cAMP/PKA pathway activator) to stimulate trophoblast differentiation. hCG and P4 secretions increase during trophoblast differentiation (respectively 25 and 8 folds). An increase of P4 (12 folds) and pregnenolone (6 folds) is observed using GC-MS/MS during VCTs differentiation. However, 17-OH-Progesterone is not detected, confirming the absence of CYP17 enzyme in placenta. HSD3B1 and MLN64 expression increase (respectively 4 and 1,5 folds) during trophoblast differentiation at mRNA and protein levels whereas P450SCC expression remains constant. Using IA and/or CG-MS/MS, after FSK-induced differentiation, hCG level increases at 24h and 72h whereas P4 and pregnenolone levels increase (respectively 1,3 and 1,5-folds) only at 72h. Furthermore, HSD3B1 and P450SCC protein expressions increase (respectively 1,3 and 2,1 folds) in FSK-incubated trophoblasts at 72h. Our results highlight that steroidogenesis is already effective in VCT. Each steps of the synthesis increase during trophoblast differentiation leading to increase pregnenolone and progesterone secretions. Interestingly, FSK-induced trophoblast differentiation involves a rise in progesterone production. These results suggest that steroid production is linked to VCT differentiation and may involve hCG.