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The morphology of the exocrine secretory unit of the pancreas, i.e. the pancreatic acinus, is reviewed. The histological features of the acini and their relation with the duct system are described. The acinar three-dimensional architecture was studied by means of different ultrastructural techniques, some of which are complementary. The fine structure and morphodynamics of the acinar cells are also described. In addition, the location of the organelles in specific cytoplasmic domains and their close morphofunctional relationship with the sequential stages of secretion of the digestive enzymes are specially emphasized. Finally, morphological approaches are suggested to achieve a better comprehension of the physiological and pathological pancreatic activities whose morphodynamics need to be further elucidated or are almost totally unknown.

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.

The fine structure of the zona pellucida (ZP) covering the oocyte and of the mucus covering the surface of the intestinal villi was investigated by using a new method employing ruthenium red (RR), saponin, and osmium-thiocarbohydrazide impregnation. The glycoproteic matrices both appeared constituted by thin filaments (ranging from 22 to 50 nm in thickness) anastomosed to form a very fine network. RR prevented the dissolution and/or alteration of glycoproteins and polyanionic carbohydrates induced by acqueous fixatives. Saponin was a detergent of the soluble proteins. Osmium-thiocarbohydrazide preserved the glycoproteic matrix filaments from the mechanical stress induced by dehydration and critical point drying and reduced filaments packing and shrinkage. The technical improvement was demonstrated by the following results: 1) a regular arrangement of the filaments network; 2) a thickness of mucus filaments smaller than that obtained with other methods of preparation; 3) a homogeneous thickness of ZP filaments. This method allowed a very detailed study of the fine structural organization of the ZP and intestinal mucus. Therefore, this technique can be useful for a better evaluation of the morphodynamic of these and other glycoproteic matrices.

The human zona pellucida (ZP) and its changes during in vitro fertilization in oocytes at different maturational stages and polypronuclear ova at one- to four-cells stages were studied by transmission electron microscopy (TEM) and correlative scanning electron microscopy (SEM). To define the microstructure of the ZP, its amorphous masking material was removed using a detergent (saponin), and its structural glycoproteins were stabilized with a cationic dye, ruthenium red, followed by osmium-thiocarbohydrazide treatment. These methods allowed in all samples the clear visualization of variously arranged networks of filaments composing the outer and inner surfaces of the ZP. These filaments were straight or curved, 0.1-0.4 microns in length and 10-14 nm thick as seen via TEM or 22-28 nm thick as seen via SEM (the difference in thickness was due to the presence of the metal coating for SEM). The filament arrangement was remarkably different between the inner and outer surfaces of the ZP and among the various stages studied. The filaments of the outer surface of the ZP were basically arranged in "large" and "tight" meshed networks. Mature oocytes and fertilized (polypronuclear) ova had a regular alternating pattern of wide and tight meshed networks of filaments. On the other hand, immature and atretic oocytes displayed almost exclusively a tight meshed network of filaments. The inner surface filaments of the ZP of unfertilized oocytes at any stage were arranged in repetitive structures characterized by numerous short and straight filaments anastomosing with each other and sometimes forming at the intersections small, rounded structures. After fertilization, the inner surface of the ZP displayed numerous areas where filaments fused together. Collectively, these data clearly reveal that oocyte maturation and fertilization in humans are accompanied by changes of ZP filaments arrangement, which may be relevant in the processes of binding, penetration, and selection of spermatozoa.