FUNCTIONAL, LONG-TERM HUMAN THECA AND GRANULOSA-CELL CULTURES FROM POLYCYSTIC OVARIES

Reproducible culture conditions for obtaining large numbers of functio nal PCOS theca interns and granulosalutein cells will be indispensable in studies focussing on the molecular basis for androgen overproducti on by ovarian cells of patients with polycystic ovarian syndrome (PCOS ). The objective of the present study was to determine if granulosa an d theca interna cells obtained from ovarian follicles of patients with PCOS could be passaged with maintenance of inducible steroidogenic ac tivity. PCOS theca interna and granulosa cells were obtained from indi vidual follicles of polycystic ovaries containing multiple cystic foll icles with characteristic hypertrophied theca interna. Utilizing condi tions for growing normal ovarian cells, both cell types were passaged successively and conditions for cell freezing, storing and thawing wer e established. In granulosa-lutein cultures grown and passed for succe ssive passages, and transferred into serum-free medium, forskolin stim ulated aromatase activity increased 3-10-fold over control non-stimula ted values. Concurrent treatment with ICF-I (50 ng/mL) enhanced forsko lin-stimulated aromatase activity in PCOS granulosa-lutein cultures. I n passaged PCOS theca interna cells, forskolin-stimulated 17 alpha-hyd roxyprogesterone production was increased 4-25-fold over control value s. Treatment of PCOS theca interna cells with insulin (50 ng/mL) enhan ced forskolin-stimulated 17 alpha-hydroxyprogesterone biosynthesis. Th e effects of various growth factors and phorbol esters on 17 alpha-hyd roxylase activity in cultured PCOS theca interna cells was also invest igated. Treatment of PCOS theca cells with EGF, FGF, TGF beta and TPA resulted in the inhibition of forskolin-stimulated 17 alpha-hydroxypro gesterone production. These data suggest that PCOS theca interna and g ranulosa cells respond to insulin and to the growth factors similarly to cells obtained from normal cycling ovaries.