Luteinized human granulosa cells are associated with endogenous basement membrane-like components in culture

Human granulosa cells (GC), prepared from follicular aspirates using a non- enzymic method, were maintained in culture on chamber slides in a defined m edium without additional attachment factors or extracellular matrix (ECM), In this system, GC clustered to a limited extent and attached only loosely to the substratum necessitating medium replacement through repeated partial changes to avoid cell loss. Using this new culture system, cell size and p rogesterone production per cell increased, consistent with continuing lutei nization. These processes were associated with maintenance and deposition o f endogenous ECM components. Thus, pericellular heparan sulphate proteoglyc an (HSPG) was clearly visible by immunocytochemistry around the luteinized GC after culture. Also progressive accumulation of laminin (particularly al pha(2)-, beta(1)- and gamma(1)-subunits) during culture was shown by Wester n blotting of GC extracts. Small patches of collagen IV, shown to be alread y present between freshly prepared GC, were maintained in culture. A clear effect of gonadotrophin on the maintenance of progesterone production in cu lture was paralleled by an apparent increased pericellular deposition of HS PG. To conclude, luteinization and maintenance of the GC-derived layer of t he corpus luteum is likely to involve deposition and conservation of perice llular ECM components.