Background:Spermatogonial stem cells(SSCs)are capable of both self-renewal and differentiation to mature functional spermatozoa,being the only adult stem cells in the males that can transmit genetic information to the...Background:Spermatogonial stem cells(SSCs)are capable of both self-renewal and differentiation to mature functional spermatozoa,being the only adult stem cells in the males that can transmit genetic information to the next generation.Porcine SSCs hold great value in transgenic pig production and in establishment of porcine models for regenerative medicine.However,studies and applications of porcine SSCs have been greatly hampered by the low number of SSCs in the testis as well as the lack of an ideal stable long-term culture system to propagate porcine SSCs perpetually.Results:In the present study,by lentiviral transduction of plasmids expressing the simian virus 40(SV40)large T antigen into porcine primary SSCs,we developed two immortalized cell lines with porcine SSC attributes.The established cell lines,with the expression of porcine SSC and germ cell markers UCHL1,PLZF,THY1,VASA and DAZL,could respond to retinoic acid(RA),and could colonize the recipient mouse testis without tumor formation after transplantation.The cell lines displayed infinite proliferation potential,and have now been cultured for more than 7 months and passaged for over 35 times without morphological abnormalities.Conclusions:We have for the first time established porcine SSC lines that could provide abundant cell sources for mechanistic studies on porcine SSC self-renewal and differentiation,thereby facilitating development of an optimal long-term culture system for porcine primary SSCs and their application to animal husbandry and medicine.展开更多
Background:Spermatogenesis is the process by which male gametes are formed from spermatogonial stem cells and it is essential for the reliable transmission of genetic information between generations.To date,the dynami...Background:Spermatogenesis is the process by which male gametes are formed from spermatogonial stem cells and it is essential for the reliable transmission of genetic information between generations.To date,the dynamic transcriptional changes of defined populations of male germ cells in pigs have not been reported.Results:To characterize the atlas of porcine spermatogenesis,we profiled the transcriptomes of~16,966 testicular cells from a 150-day-old pig testis through single-cell RNA-sequencing(scRNA-seq).The scRNA-seq analysis identified spermatogonia,spermatocytes,spermatids and three somatic cell types in porcine testes.The functional enrichment analysis demonstrated that these cell types played diverse roles in porcine spermatogenesis.The accuracy of the defined porcine germ cell types was further validated by comparing the data from scRNA-seq with those from bulk RNA-seq.Since we delineated four distinct spermatogonial subsets,we further identified CD99 and PODXL2 as novel cell surface markers for undifferentiated and differentiating spermatogonia,respectively.Conclusions:The present study has for the first time analyzed the transcriptome of male germ cells and somatic cells in porcine testes through scRNA-seq.Four subsets of spermatogonia were identified and two novel cell surface markers were discovered,which would be helpful for studies on spermatogonial differentiation in pigs.The datasets offer valuable information on porcine spermatogenesis,and pave the way for identification of key molecular markers involved in development of male germ cells.展开更多
基金This study was supported by the National Natural Science Foundation of China(Grant No.31572401,31772605)to W.Z.the Open Fund of Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province(Grant No.SNDK-KF-201804)Young Talent fund of University Association for Science and Technology in Shaanxi,China(Grant No.20180204)and a startup fund from Northwest A&F University(Grant No.2452018037)to Y.Z.
文摘Background:Spermatogonial stem cells(SSCs)are capable of both self-renewal and differentiation to mature functional spermatozoa,being the only adult stem cells in the males that can transmit genetic information to the next generation.Porcine SSCs hold great value in transgenic pig production and in establishment of porcine models for regenerative medicine.However,studies and applications of porcine SSCs have been greatly hampered by the low number of SSCs in the testis as well as the lack of an ideal stable long-term culture system to propagate porcine SSCs perpetually.Results:In the present study,by lentiviral transduction of plasmids expressing the simian virus 40(SV40)large T antigen into porcine primary SSCs,we developed two immortalized cell lines with porcine SSC attributes.The established cell lines,with the expression of porcine SSC and germ cell markers UCHL1,PLZF,THY1,VASA and DAZL,could respond to retinoic acid(RA),and could colonize the recipient mouse testis without tumor formation after transplantation.The cell lines displayed infinite proliferation potential,and have now been cultured for more than 7 months and passaged for over 35 times without morphological abnormalities.Conclusions:We have for the first time established porcine SSC lines that could provide abundant cell sources for mechanistic studies on porcine SSC self-renewal and differentiation,thereby facilitating development of an optimal long-term culture system for porcine primary SSCs and their application to animal husbandry and medicine.
基金This study was supported in part by the National Natural Science Foundation of China(Grant No.31772605)to WXZResearch Project of Shaanxi Science and Technology Department(2020NY-003)to TZ.
文摘Background:Spermatogenesis is the process by which male gametes are formed from spermatogonial stem cells and it is essential for the reliable transmission of genetic information between generations.To date,the dynamic transcriptional changes of defined populations of male germ cells in pigs have not been reported.Results:To characterize the atlas of porcine spermatogenesis,we profiled the transcriptomes of~16,966 testicular cells from a 150-day-old pig testis through single-cell RNA-sequencing(scRNA-seq).The scRNA-seq analysis identified spermatogonia,spermatocytes,spermatids and three somatic cell types in porcine testes.The functional enrichment analysis demonstrated that these cell types played diverse roles in porcine spermatogenesis.The accuracy of the defined porcine germ cell types was further validated by comparing the data from scRNA-seq with those from bulk RNA-seq.Since we delineated four distinct spermatogonial subsets,we further identified CD99 and PODXL2 as novel cell surface markers for undifferentiated and differentiating spermatogonia,respectively.Conclusions:The present study has for the first time analyzed the transcriptome of male germ cells and somatic cells in porcine testes through scRNA-seq.Four subsets of spermatogonia were identified and two novel cell surface markers were discovered,which would be helpful for studies on spermatogonial differentiation in pigs.The datasets offer valuable information on porcine spermatogenesis,and pave the way for identification of key molecular markers involved in development of male germ cells.
