Wuzi-Yanzong-Wan Prevents the Defect of Cell-Cell Junctions between Sertoli-Germ Cells by Up-Regulating the Expression of TAp73

The TAp73 gene is an anti-cancer gene that also affects the junction between Sertoli and germ cells. Inhibition of this gene causes infertility in male mice. Our previous research proved that Wuzi-Yanzong-Wan (WZYZW) can protect spermatogenesis and maturation by preventing TAp73 inhibition.

This study aimed to investigate the effect of drug-containing serum of WZYZW on the defect of cell-cell junctions in the Sertoli-germ cells co-culture system in vitro.

LC-HRMS was used to analyze the content of active ingredients in WZYZW-medicated serum. Then, primary extraction and co-culture of germ cells and Sertoli cells were carried out. Co-cultured cells were added with PFT-α to induce the TAp73 inhibition model, with WZYZW-medicated serum at 2.5%, 5%, and 10% treated in parallel. Sloughing of germ cells from Sertoli cells was calculated. Transmission electron microscopy (TEM), Immunofluorescence, qRT-PCR, and western blot methods were employed.

The drug-containing serum of WZYZW contained schisandrin, hyperoside, geniposidic acid, ellagic acid, and quercetin. Using TEM assay, we observed restoration of the desmosome-like (Des), tight junctions (TJ), and basal ectoplasmic specialization (ES) structure following WZYZW treatment. WZYZW caused inhibition of peptidase and protease inhibitors (tissue inhibitor of metalloproteinase-1 (TIMP1), Serpina3n) by immunofluorescence analysis. Western blot and qRT-PCR analysis revealed that WZYZW was able to ameliorate the expressions of peptidase and protease inhibitors and cell adhesion factors, such as TAp73, TIMP1, Serpina3n, Desmocollin-3, N-cadherin, and Nectin-2.

WZYZW-medicated serum could prevent the defect of cell-cell junctions between Sertoli-germ cells co-culture system in vitro by up-regulating the expression of TAp73.