In vitro differentiation of spermatogonial stem cells using testicular cells from Guangxi Bama mini-pig
2018
Huimin Zhao, Guangxi University, Nanning, China | Junyu Nie, Guangxi University, Nanning, China | Xiangxing Zhu, Guangxi University, Nanning, China | Yangqing Lu, Guangxi University, Nanning, China | Xingwei Liang, Guangxi University, Nanning, China | Huiyan Xu, Guangxi University, Nanning, China | Xiaogan Yang, Guangxi University, Nanning, China | Yunkai Zhang, Guangxi University, Nanning, China | Kehuan Lu, Guangxi University, Nanning, China | Shengsheng Lu, Guangxi University, Nanning, China
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.
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