Type VI Collagen in the Rat Testis: Monoclonal Antibody, Isolation, and Localization during Development1

Abstract

To investigate the components of the seminiferous tubule extracellular matrix (ST-ECM), monoclonal antibodies were raised against ST-ECM. One of these (15B6) recognized the nonreduced form of type VI collagen. With this antibody as a probe, type VI collagen was isolated from an SDS-urea extract of ST-ECM. It took a tetrameric form, and upon reduction dissociated into a major 140-kDa band and two bands at 190 and 210 kDa. Developmental changes in the distribution of type VI collagen were investigated by immunofluorescence microscopy. In 16-day rat embryos when myoid cells began to differentiate, type VI, collagen was seen in the urogenital mesentery, and it had begun to surround the gonad and, in some embryos, the seminiferous cords. In newborn rats, it was found throughout the extracellular spaces between the cords, being localized in the interstitium. In adults, however, it was confined to two parallel lines around the seminiferous tubules and to filamentous structures in the adventitia of vascular systems. Immunoelectron microscopy showed that this collagen was distributed in the interstitium independently from collagenous fibrils, but was excluded from the basal lamina of Sertoli cells. These observations indicate that type VI collagen is a major component of the ST-ECM and that it may be important in the organization of the matrix and in the differentiation of peritubular cells.

Developmental Changes in the Basement Membrane of the Normal and Hypothyroid Postnatal Rat Testis: Segmental Localization of Fibulin-2 and Fibronectin1

“…These changes demonstrate that the seminiferous tubule BM is undergoing subtle changes in its composition during the postnatal period before the onset of germ cell meiotic divisions. Previous studies have shown a change in the distribution of collagen VI within the postnatal rat testis, from an exclusive interstitial localization in the fetal and neonatal testis to a close association with the seminiferous tubule BM by Day 14 [18]. Our observations confirm these studies and demonstrate that the association of collagen VI with BMs occurs progressively from Day 10 to Day 20.…”

“…Collagen VI is initially restricted to the interstitial matrix between seminiferous cords of the fetal and early postnatal rat testis, and subsequently it is observed to be associated with the BM around Sertoli cells and peritubular cells by Day 14, when these cell types are beginning to differentiate [18]. In the mouse, the appearance of the ao3(IV) isoform chain of collagen IV in the BMs around seminiferous tubules coincides with the period when spermatogonia begin to proliferate and then migrate to the BM, suggesting that collagen ao3(IV) chains may have a role in these processes [19].…”

Passive Immunization with Anti-Laminin Immunoglobulin G Modifies the Integrity of the Seminiferous Epithelium and Induces Arrest of Spermatogenesis in the Guinea Pig1

“…In the normal developing rat testis, type VI collagen has been reported to fill the interstitial spaces between fetal cords, but in the adult, the same type of collagen was shown confined to the LIMM of the tubules and of the adventitia of the vessels [33,34]. Furthermore, it was concluded that type VI collagen was distributed independently from type I collagen fibrils in the interstitial spaces and that it was excluded from the basal lamina of the Sertoli cells [34], suggesting that peritubular myoid cells may be, at least in part, responsible for the deposit of type VI collagen.…”

27 kDa extracellular matrix protein revealed by a monoclonal antibody raised against rat testis

“…The ST-ECM was homogenized and used for immunization (Sawada and Yazama, 1994). Hybridomas were produced by fusing splenocytes from immunized BALB/c mice with X63-Ag8-6.5.3 myelomas with polyethylene glycol 1530 (Sigma Chemical Co.), according to the standard procedure (Galfre and Milstein, 1981).…”