A 22-amino acid synthetic peptide corresponding to the second extracellular loop of rat occludin perturbs the blood-testis barrier and disrupts spermatogenesis reversibly in vivo

When Sertoli cells were cultured in vitro on Matrigel-coated bicameral unit s, the assembly of the inter-Sertoli tight junction (TJ) permeability barri er correlated with an induction of occludin expression. Inclusion of a 22-a mino acid peptide, NH2-GSQIYTICSQFYTPGGTGLYVD-COOH, corresponding to residu es 209-230 in the second extracellular loop of rat occludin, at 0.2-4 muM i nto Sertoli cell cultures could perturb the assembly of Sertoli TJs dose-de pendently and reversibly. This peptide apparently exerts its effects by int erfering with the homotypic interactions of two occludin molecules between adjacent Sertoli cells at the sites of TJs, thereby disrupting TJs, which, in turn, causes a decline in transepithelial electrical resistance across t he Sertoli cell epithelium. When similar experiments were performed using a 22-amino acid myotubularin peptide, NH2-TKVNERYELCDTYPALLAVPAN-COOH (resid ues 156-177), no effects on the assembly of inter-Sertoli TJs in vitro were noted. When a single dose of this synthetic occludin peptide was administe red to adult rats intratesticularly at 1.5-10 mg/testis, germ cells began t o deplete from the seminiferous epithelium within 8-16 days. By 27 days, vi rtually all tubules were devoid of germ cells. This antispermatogenic effec t was reversible, because germ cells progressively repopulated the epitheli um thereafter. Treated testes were indistinguishable from normal or control testes by 68 days post-occludin peptide treatment when assessed using hist ological analysis. In contrast, control rats receiving either no treatment, vehicle alone, or a 22-amino acid synthetic peptide of myotubularin displa yed no changes in the testicular morphology at all time points. The occludi n peptide-induced germ cell depletion was also accompanied by a disruption of the blood-testis barrier (BTB) when assessed by micropuncture techniques quantifying [I-125]-BSA in rete testis fluid and seminiferous tubular flui d following i.v. administration of [I-125]-BSA through the jugular vein. Th ese results illustrate that the occludin peptide-induced disruption of the BTB may possibly affect the underlying adherens junctions, which causes pre mature release of germ cells from the epithelium and reversible infertility .