In vitro differentiation of spermatogonial stem cells using testicular cells from Guangxi Bama mini-pig

Abstract

In this study, we attempted to establish a culture system for in vitro spermatogenesis from spermatogonial stem cells (SSCs) of Bama mini-pig. Dissociated testicular cells from 1-month-old pigs were co-cultured to mimic in vivo spermatogenesis. The testicular cells were seeded in minimum essential medium alpha (α-MEM) supplemented with Knockout serum replacement (KSR). Three-dimensional colonies formed after 10 days of culture. The colonies showed positive staining for SSC-associated markers such as UCHL1, PLZF, THY1, OCT4, Dolichos biflorus agglutinin, and alkaline phosphatase. Induction of SSCs was performed in α-MEM + KSR supplemented with retinoic acid, bone morphogenetic protein 4, activin A, follicle-stimulating hormone, or testosterone. The results showed that STRA8, DMC1, PRM1, and TNP1 were upregulated significantly in the colonies after induction compared to that in testis from 1-month-old pigs, while expression levels of those genes were significantly low compared to those in 2-month-old testis. However, upregulation of ACROSIN was not significant. Replacement of α-MEM and KSR with Iscove's modified Dulbecco's medium and fetal bovine serum did not upregulate expression of these genes significantly. These results indicate that SSCs of Bama mini-pig could undergo differentiation and develop to a post-meiotic stage in α-MEM supplemented with KSR and induction factors.

Distribution of extracellular matrix related proteins in normal and cryptorchid ziwuling black goat testes

“…Col IV mainly exists in basement membrane, Sertoli cells, spermatogenic cells and myoid cells in bovine testes ( Berkholtz et al, 2006 ). In vitro studies indicates that adding Col IV in mediums is beneficial to the growth and differentiation of porcine SSCS ( Zhao et al, 2018 ). Consistent with the mentioned result, Col IV is positively expressed in spermatogenic cells and peritubular myoid cells, and strong positively expressed in Sertoli cells, Leydig cells and capillary wall cells in normal testicular tissues of Ziwuling black goats.…”

High Temperature Inhibits The Differentiation of Spermatogonial Stem Cells

“…As a ligand of c-kit, SCF binds to it and plays an important role in regulating the balance between self-renewal and differentiation of SSCs [39]. Activin A plays an crucial role in germ cell maturation during the period when gonocytes resume mitosis to form the SSCs and differentiating germ cell populations [40], and Activin A is widely used as a cytokine in stem cell differentiation cultured in vitro [41][42][43]. Through this culture system, we are able to study SSC differentiation process.…”

The effects of growth factors on proliferation of spermatogonial stem cells from Guangxi Bama mini‐pig

“…Pigs are castrated at this age to improve meat quality. All procedures involving animals were performed in compliance with the Animal Care and Use Committee of the Germplasm Resource Center of Chinese Experimental Mini‐pig, Animal Care and Use Committee of Guangxi University (Zhao et al, ).…”

“…The cultured cells and testicular tissues from 1 month piglet were used for qRT‐PCR analysis, which was performed as our previous report (Zhao et al, ). Total RNA was extracted by using TRIzol Reagent (Cat no.15596–026; Invitrogen) according to the manufacturer's instructions.…”