Multipotential ability of primitive germ cells from neonatal pig testis cultured in vitro
Sandeep Goel A C , Mayako Fujihara A , Kazuo Tsuchiya B , Yuji Takagi B , Naojiro Minami A , Masayasu Yamada A and Hiroshi Imai A DA Laboratory of Reproductive Biology, Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan.
B Faculty of Agriculture, Shinshu University, Nagano 399-4598, Japan.
C Centre for Cellular and Molecular Biology, Council for Scientific and Industrial Research, Uppal Road, Hyderabad 500 007, India.
D Corresponding author. Email: imai@kais.kyoto-u.ac.jp
Reproduction, Fertility and Development 21(5) 696-708 https://doi.org/10.1071/RD08176
Submitted: 19 August 2008 Accepted: 8 March 2009 Published: 22 May 2009
Abstract
Gonocytes are progenitor-type germ cells that arise from primordial germ cells and differentiate further into spermatogonia, thereby initiating spermatogenesis. In the present study, freshly isolated gonocytes were found to have either weak or no expression of pluripotency determining transcription factors, such as POU5F1, SOX2 and C-MYC. Interestingly, the expression of these transcription factors, as well as other vital transcription factors, such as NANOG, KLF4 and DAZL, were markedly upregulated in cultured cells. Cells in primary cultures expressed specific germ cell and pluripotency markers, such as lectin Dolichos biflorus agglutinin (DBA), KIT, ZBTB16, stage-specific embryonic antigen (SSEA-1), NANOG and POU5F1. Using a monoclonal antibody to specifically identify porcine germ cells, the stem cell potential of fresh and cultured cells was determined with a testis xenotransplantation assay. Colonised porcine germ cells were detected only in mouse testes that were either transplanted with fresh testicular cells or with cells from primary cultures. Interestingly, testes transplanted with cells from primary cultures showed colonisation of germ cells in the interstitial space, reflecting their tumourigenic nature. The formation of teratomas with tissues originating from the three germinal layers following the subcutaneous injection of cells into nude mice from primary cultures confirmed their multipotency. The results of the present study may provide useful information for the establishment of multipotent germ stem cell lines from neonatal pig testis.
Acknowledgements
The authors thank Kazuo Kishimoto and Sachiko Kishimoto for providing testes samples and Dr Miki Sugimoto for critical discussion of this manuscript. This work was supported by a grant (no. 17658123) from the Ministry of Education, Science and Culture (to H.I.).
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