Effect of 3D substrate obtained from decellularized tissue of rat testis on the differentiation of mouse spermatogonial cells

 In males, spermatogonial stem cells (SSCs) are the cause of germ cell production and therefore fertility. Reducing and damaging SSCs is one of the causes of infertility. Culture, proliferation, and differentiation of SSCs in vitro can be a solution to treat some cases of infertility. In the present study, performed in 2019 at the Iranian Biological Resource Center, we examined the growth and differentiation of SSCs on decellularized testicular matrix (DTM) in rats.

 In the current experimental study, after extraction of SSCs by enzymatic method from testicular tissue of newborn mice, these cells were propagated in a specific culture medium for three weeks. After confirming the identity of the colonies resulting from the growth of these cells, they were cultured in two groups, one on a layer of DTM and the other in two-dimensional conditions of conventional culture dishes with differential culture medium. In the fourth week, since the initiation of the differentiation culture, the expression of pre meiotic (OCT4 & PLZF) and meiotic (SCP3 & Protamine-2) genes were measured in both groups. To ensure the results, all steps were performed with three biological replications and the results were evaluated using one way ANOVA.

  Examination of pre-meiotic and meiotic gene expression after 4 weeks of differential culture of SSCs on two-dimensional substrate and DTM using real-time PCR showed that the expression of meiotic genes was significantly higher on DTM substrate (P ≤ 0.01). ).

 In DTM three-dimensional culture, due to the better communication of cells with each other and the presence of a natural extracellular matrix, more ideal conditions are created for the preservation, proliferation and differentiation of SSCs than in two-dimensional culture.