Endocrine Abstracts

In human granulosa-lutein (hGL) cells, cortisol (F) is inter-converted with cortisone (E) by 11BHSD. To date two isoforms of 11BHSD have been cloned. Human granulosa cells have been shown to express 11BHSD2 mRNA during the follicular phase of the ovarian cycle, switching to 11BHSD1 mRNA expression as luteinization occurs. However, it is not known whether (1) the protein expression, and (2) the 11BHSD enzyme activities reflect this pattern of mRNA expression. Hence, the aims of the current study were to investigate the expression and activities of 11BHSD proteins in luteinzing human granulosa-lutein (hGL) cells. After isolation from the follicular aspirates of patients undergoing gonadotrophin-stimulated IVF-ET, hGL cells were cultured for up to 3 days with the assessment of enzyme activities (11B-dehydrogenase and 11-ketosteroid reductase) and the expression of 11BHSD1 and 11BHSD2 proteins on each day of culture. In Western blots, an immunopurified 11BHSD1 antibody recognised a 38kDa protein in hGL cells and in human embryonic kidney (HEK) cells stably transfected with human 11BHSD1. The 11BHSD2 antibody recognised a 48kDa protein in HEK cells transfected with human 11BHSD2 cDNA but 11BHSD2 protein was not expressed in hGL cells throughout the 3 days of culture. While the expression of 11BHSD1 protein increased progressively by 3.5-fold over 3 days as hGL cells luteinized, both 11B-dehydrogenase and reductase activities declined (by 52.9% and 34.2%; P<0.05) over this same period. Changes in enzyme expression and activity were unaffected by the suppression of endogenous steroidogenesis in the hGL cells using 0.1mM aminoglutethimide. We conclude that luteinizing hGL cells express 11BHSD1 protein, and may also secrete a paracrine/autocrine inhibitor of 11BHSD1 which does not appear to be a locally-synthesised steroid. We have recently identified the presence of such a compound in the hydrophobic fractions of follicular fluid.

[ Supported by a BBSRC project grant ].