INFLUENCE OF FOLLICULAR MATURATION ON THE DEPOSITION OF FIBRONECTIN BY CHICKEN GRANULOSA-CELLS IN-VITRO - EFFECT OF 8-BROMO-CYCLIC AMP

Fibronectin deposition by avian granulosa cells and the effect of 8-bromo cAMP (8-br-cAMP) on this process was investigated in vitro. Granulosa cells obtained from the first (F1) and third (F3) largest preovulatory follicles and from small yellow follicles (SYF) were incubated in serum free medium 199 for 12 or 24 hr with and without 8-br-cAMP. Fibronectin deposited in incubation wells, secreted in medium or associated with cells was measured by ELISA. Basal fibronectin deposition increased with time and was greatest for F1 cells. Within 12 hr, 8-br-cAMP enhanced fibronectin deposition dose-dependently by F3 and SYF cells, but not by F1 cells. The amount of fibronectin deposition caused by 8-br-cAMP (which was greater after 24 hr incubation) was 0.10- to 0. 16-fold for F1 cells; 0.20- to 0.81-fold for F3 cells and more than 30-fold for SYF cells. Fibronectin secreted in the medium by unstimulated cells was also greatest in F1 cells. 8-bromo cAMP stimulated fibronectin secretion in medium dose-dependently by F3 and SYF cells, however, it had only a marginal stimulatory effect on this process in F1 cells, Cell-associated fibronectin was not increased significantly by 8-br-cAMP in F1, F3 or SYF cells. Total (deposited plus medium plus cell associated) fibronectin production was elevated in dose- and time-dependent manner by 8-br-cAMP in F3 and SYF cells, but only at high concentrations in F1 cells. Like deposited fibronectin, the relative effect of 8-br-cAMP on fibronectin secreted in the medium and on total fibronectin production was greatest in immature SYF cells. These results demonstrate that fibronectin synthesized and deposited by unstimulated chicken granulosa cells in vitro increase concomitantly with follicular maturation and cellular differentiation, and the relative change in fibronectin production caused by 8-br-cAMP was greatest in immature SYF cells.