The Interplay of Collagen IV, Tumor Necrosis Factor-α, Gelatinase B (Matrix Metalloprotease-9), and Tissue Inhibitor of Metalloproteases-1 in the Basal Lamina Regulates Sertoli Cell-Tight Junction Dynamics in the Rat Testis

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Abstract
During spermatogenesis, preleptotene and leptotene spermatocytes must translocate across the blood-testis barrier formed by inter-Sertoli cell-tight junctions (TJs) from the basal compartment of the seminiferous epithelium adjacent to the basement membrane to the adluminal compartment at stages VIII–IX for further development. Because of the close proximity between extracellular matrix (ECM) that constitutes the basement membrane and the blood-testis barrier, we sought to investigate the role of ECM in Sertoli cell TJ dynamics. When Sertoli cells were cultured in vitro to initiate the assembly of the Sertoli cell TJ-permeability barrier, the presence of an anticollagen IV antibody indeed perturbed the barrier. Because ECM is known to maintain a pool of cytokines and TNFα has been shown to regulate TJ dynamics in other epithelia, we investigated whether TNFα can regulate Sertoli cell TJ function via its effects on collagen α3(IV) and other proteins that maintain the homeostasis of ECM. As expected, recombinant TNFα perturbed the Sertoli cell TJ-barrier assembly in vitro dose dependently. TNFα also inhibited the timely induction of occludin, which is known to associate with the Sertoli cell TJ-barrier assembly. Furthermore, TNFα induced the expression of Sertoli cell collagen α3(IV), gelatinase B (matrix metalloprotease-9, MMP-9) and tissue inhibitor of metalloproteases-1 but not gelatinase A (matrix metalloprotease-2), and promoted the activation of pro-MMP-9. These results thus suggest that the activated MMP-9 induced by TNFα is used to cleave the existing collagen network in the ECM, thereby perturbing the TJ-barrier. This in turn creates a negative feedback that causes TNFα to induce collagen α3(IV) and tissue inhibitor of metalloproteases-1 expression so as to replenish the collagen network in the disrupted TJ-barrier and limit the activity of MMP-9. Taken collectively, these observations strengthen the notion that ECM is involved in the regulation of junction dynamics in addition to its structural role in the testis.