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Effect of gonadotrophin on cell and matrix retention and expression of metalloproteinases and their inhibitor in cultured human granulosa cells modelling corpus luteum function.

Granulosa cells were prepared from follicular aspirates obtained at oocyte collection for in-vitro fertilization (IVF) and maintained in culture. Substantial loss of cells from the culture surface occurred in the absence of gonadotrophin when cells were maintained on a thin layer of extracellular matrix (ECM) using a defined, serum-free medium. This cell loss was clearly and significantly reduced in the presence of human chorionic gonadotrophin (HCG) by days 4-6 of culture, and occurred in conjunction with loss of ECM. Analysis of culture medium by zymography using gelatin as substrate demonstrated the presence of metalloproteinases (MMP), MMP-9 (gelatinase B) appearing as the predominant band. Measurement of overall gelatinase activity in culture media revealed a progressive fall in gelatinase expressed on a per cell basis in media from HCG-treated cultures and this was less marked in controls. This suppression of gelatinase activity was consistent with an observed increase in production of tissue inhibitor of metalloproteinase-1 (TIMP-1) by HCG-treated cells, which was significant by days 6-8 of culture. We speculate that stabilization of the ECM may be an important aspect of HCG action in the corpus luteum.