Heterogeneous distribution of fibronectin, tenascin-C, and laminin immunoreactive material in the cumulus-corona cells surrounding mature human oocytes from IVF-ET protocols--evidence that they are composed of different subpopulations: an immunohistochemical study using scanning confocal laser and fluorescence microscopy.

Monoclonal antibodies and immunofluorescence microscopy, including laser confocal microscopy, were used in this study to point out the production of fibronectin, tenascin-c, and laminin in the cumulus-corona (CC) cells surrounding mature human oocytes from IVF-ET protocols in view of their presumptive importance in the coordination of the processes leading to fertilization and early embryo cleavage, including the final maturation of the ovum, the sperm-egg interaction, and the "complex biochemical dialogue" between the gamete and the oviduct through the tubal luminal environment. One hundred fifty mature oocyte-CC complexes were obtained from IVF-ET protocols and fixed in 4.0% buffered paraformaldehyde. Specimens were incubated with a panel of primary monoclonal antibodies (mabs) recognizing different epitopes of fibronectin, tenascin-c, and laminin and then with fluorescein isothiocyanate-conjugated goat anti-mouse IgG. Observations were made by a scanning confocal microscope (Sarastro 2000) and a photomicroscope (Polyvar, Reichert-Jung) equipped with epifluorescence optics. The immunohistochemical data demonstrated that human CC cells are capable of producing fibronectin and tenascin-c but that their production is not homogeneous in the CC population. In fact, fibronectin immunoreactivity was shown mostly by inner CC cells (mainly corona cells), whereas tenascin was produced by some cells scattered in the entire cumulus mass. Moreover, fibronectin and tenascin-c immunoreactive material was observed in the intracytoplasmic areas, at the plasma membrane level as well as in the extracellular matrix. On the contrary, laminin immunofluorescent material was found around plasma membranes of almost all CC cells, but a clear intracytoplasmic reaction was never observed. This leads us to assume that laminin in the extracellular matrix remains entrapped once produced by granulosa follicular cells and that in the postovulatory period no active secretion occurs in CC cells. Even though the functional role of these extracellular matrix proteins remains still unclear, it is reasonable to suggest that they are necessary in various steps of the reproductive process, i.e., from the pick-up of the oocyte, its transport through the oviduct, and fertilization, up until the early cleavage of the embryo. Finally, functional differences between "corona radiata" and "cumulus" cells during the oocyte denudation may be accounted for particular distribution of these adhesive proteins.