IMMUNOCYTOCHEMISTRY OF EXTRACELLULAR-MATRIX COMPONENTS IN THE RAT SEMINIFEROUS TUBULE - ELECTRON-MICROSCOPIC LOCALIZATION WITH IMPROVED METHODOLOGY

Background: Cell-cell interactions between Sertoli, myoid, Leydig, and germ cells are thought to be essential for spermatogenesis. These cel ls interact with each other through the extracellular matrices (ECMs) of the testicular lamina propria, which thus may have an important fun ction in spermatogenesis. For an understanding of the role of ECMs in spermatogenesis, it is important to investigate the molecular constitu tion of the testicular ECM. We examined the distribution of type IV, V , and VI collagens (Cols), laminin (LM), fibronectin (FN), and heparan sulfate proteoglycan (HSPG) in the rat testicular lamina propria. Met hods: Adult rat testes were fixed with 4% paraformaldehyde and 0.2% gl utaraldehyde. Ultrathin frozen sections were made and immunolabeling w as performed according to the method of Tokuyasu (1986 J. Microsc., 14 3:139-149). Alternatively, for preembedding immunocytochemistry, we us ed labeling with nanogold, followed by silver enhancement and gold ton ing. The tissues were then fixed with osmium and embedded in Epon (Saw ada and Esaki 1994 J. Electron Microsc., 43:361-366). These specimens were examined in a JEOL JEM-100C transmission electron microscope oper ated at 80 kV. Results: Col IV, LM, and HSPG were localized in the bas ement membranes of the seminiferous tubule (ST-BM) and in the BM of my oid cells. Cols V and VI, and FN were localized both in the connective tissue between the seminiferous tubule and the myoid cells (tubule-my oid connective tissue) and in the connective tissue between the myoid cells and the lymphatic endothelial cells (myoid-endothelium connectiv e tissue). Furthermore, statistical analysis of the micrographs of imm unogold labeling for Col IV, LM, and FN around the STEM suggested that the Col IV molecule is located uniformly in the ST-BM, whereas the LM molecule is distributed mainly in the lamina rara of ST-BM, and the F N molecule appears to be present predominantly in the tubule-myoid con nective tissue. Conclusions: Distinct distribution patterns were obser ved for each antigen within the testicular lamina propria at the ultra structural level. However, localization of some components was not con sistent with localizations reported by others. (C) 1997 Wiley-Liss, In c.