• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2003년도 학술발표대회 발표논문초록집
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• pp.82-82
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• 2003

The objective of the present study was to investigate the survival effect after transplantation of pig spermatogonia cells into mouse testis. Donor cells were collected from porcine testis and the isolated spermatogonial stem cells were labeled with a fluorescent marker before transplantation and transplanted into testes of busulfan-treated recipient mice. Testes were examined for the presence and localization of labeled donor cells immediately after transplantation or every week for 4 wk. Transplanted germ cells were present in the seminiferous epithelium at 4 weeks after the transplantation, but any differentiating porcine-derived cells were not detected in mouse testis. These results indicate that porcine-derived spermatogonial stem cells can be survived in the recipient, but suggest that porcine-derived male stem cells can not proceed to further differentiating step without helping of immunosuppressor agents.

• Reproductive and Developmental Biology
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• 제32권3호
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• pp.193-198
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• 2008

본 연구는 돼지에 있어서 정원줄기세포를 포함하는 정소세포를 recipient 돼지의 정소 내로 이식할 수 있는 기법을 개발하기 위하여 시행되었다. 공여세포는 $10{\sim}14$일령의 돼지로부터 채취된 정소에서 효소처리법을 이용하여 회수하였고, recipient의 정소 내로 이식하기 전 형광 마커(PHK26)로 표지하였다. 외과적 수술을 통하여 recipient 돼지부터 정소를 꺼낸 후 초음파 기기와 이식 장치를 이용하여 형광표지된 공여세포를 recipient 정소의 세정관 내로 이식하였다. 14주령의 recipient 정소에 $5{\sim}7ml$의 공여 세포부유액을 주입하여 정소 내 50% 이상의 세정관 내로 새포부유액의 주입이 가능하였고, 세포부유액이 주입된 세정관 내에서 형광표지된 정소세포들이 고루 이식되어짐이 관찰되었다. 본 연구에서 개발한 이식 기법을 이용하여 효율적인 정소세포의 이식이 가능함에 따라 향후 돼지 정원줄기세포의 연구 및 활용법 개발에 획기적인 돌파구가 마련될 것으로 기대된다.

• Reproductive and Developmental Biology
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• 제38권4호
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• pp.137-142
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• 2014

A tremendous increase in the human population has put poultry industry under an increased pressure to meet steep increase in the demand. Poultry is contributing 25% of the total world's meat production and lesser cost of investment per bird makes it more suitable for the further breeding programmes. Major poultry diseases frequently lead to cardiac damage and cause huge economic losses to poultry industry due to mortality. The in vitro embryonic stem cell (ESC) technology has a futuristic approach for homogeneous populace of differentiated cells, for their further transplantations. During in vitro conditions the differentiated cell populace can be used in grafting and transplantation processes to regenerate damaged tissues. Therefore, the current study targeted the use of spermatogonial stem cells (SSCs) in the poultry production system through cardiac regeneration. The current study will also open new boulevard for the similar kind of research in other livestock species for the management of heart diseases.

• Asian-Australasian Journal of Animal Sciences
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• 제20권7호
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• pp.1015-1022
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• 2007

The spermatogonial stem cell (SSCs) is unique in that it is the only cell in the adult male that can contribute genes to a subsequent generation. Permanent modification of the germ cell line may be realized if stem cells could be cultured, transfected with unique genes, and then transplanted into recipient testes. We developed a culture system that supported long-term viability of SSCs. We used a retrovirus vector (pMSCV including ${\beta}$-galactosidase) to stably transfect spermatogonia following long-term culture using the system developed. Expression of the reporter gene ${\beta}$-galactosidase controlled by the retroviral vector was stable in long-term cultured SSCs. We confirmed the retroviral-mediated ${\beta}$-galactsidase gene could be expressed in germ cells in recipient mice following SSCs transplantation.

• 한국가금학회:학술대회논문집
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• 한국가금학회 2006년도 제23차 정기총회 및 학술발표회
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• pp.43-58
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• 2006

As a bioreactor, bird has proved to be most efficient system for producing useful therapeutic proteins. More than half of the egg white protein content derives from the ovalbumin gene with four other proteins(lysozyme, ovomucoid, ovomucin and conalbumin) present at levels of 50 milligrams or greater. And the naturally sterile egg also contains egg white protein at high concentration allowing for a long shelf life of recombinant protein without loss in activity. In spite of these advantages, transgenic procedures for the bird have lagged far behind because of its complex process of fertilized egg and developmental differences. Recently, a system to transplant mouse testis cells from a fertile donor male to the seminiferous tubules of an infertile recipient male has been developed. Spermatogenesis is generated from transplanted cells, and recipients are capable of transmitting the donor haplotype to progeny. After transplantation, primitive donor spermatogonia migrate to the basement membrane of recipient seminiferous tubules and begin proliferating. Eventually, these cells establish stable colonies with a characteristic appearance, which expands and produces differentiating germ cells, including mature spermatozoa. Thus, the transplanted cells self-renew and produce progeny that differentiate into fully functional spermatozoa. In this study, to develop an alternative system of germline chimera production that operates via the testes rather than through developing embryos, the spermatogonial stem cell techniques were applied. This system consisted of isolation and in vitro-culture of chicken testicular cells, transfer of in vitro-maintained cells into heterologous testes, production of germline chimeras and confirmation of germline transmission for evaluating production of heterologous, functional spermatozoa.

• Molecules and Cells
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• 제37권6호
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• pp.473-479
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• 2014

Spermatogonial stem cells (SSCs, also called germline stem cells) are self-renewing unipotent stem cells that produce differentiating germ cells in the testis. SSCs can be isolated from the testis and cultured in vitro for long-term periods in the presence of feeder cells (often mouse embryonic fibroblasts). However, the maintenance of SSC feeder culture systems is tedious because preparation of feeder cells is needed at each subculture. In this study, we developed a Matrigel-based feeder-free culture system for long-term propagation of SSCs. Although several in vitro SSC culture systems without feeder cells have been previously described, our Matrigel-based feeder-free culture system is time- and cost-effective, and preserves self-renewability of SSCs. In addition, the growth rate of SSCs cultured using our newly developed system is equivalent to that in feeder cultures. We confirmed that the feeder-free cultured SSCs expressed germ cell markers both at the mRNA and protein levels. Furthermore, the functionality of feeder-free cultured SSCs was confirmed by their transplantation into germ cell-depleted mice. These results suggest that our newly developed feeder-free culture system provides a simple approach to maintaining SSCs in vitro and studying the basic biology of SSCs, including determination of their fate.

• Journal of Animal Science and Technology
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• 제63권5호
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• pp.1194-1203
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• 2021

Preparation of recipient stallions is critical step to produce donor spermatogonial stem cell (SSC) derived sperm using transplantation technique. This study was conducted to evaluate the effects of intravenous busulfan infusion on germ cell depletion, semen production, and libido in stallions. Six Thoroughbred stallions were separated into two treatment groups: 1) a multiple low-dose (2.5 mg/kg bw for the first 4 weeks and 5 mg/kg bw for the 5th week); and 2) control group treated with PBS. Testicular samples were obtained at 11 weeks and classified into three different patterns of spermatogenesis, such as normal, Sertoli cell only, and destroyed. Semen collection and libido experiments were performed 1 week before treatment, and 4 and 8 weeks after treatment. For the sperm analysis, total spermatozoa and motility were measured using a light microscope with a motility analyzing system. In the multiple low-dose group, the numbers of tubules categorized as Sertoli cell only were significantly higher than those in the control as well as the total population and total/progressive motility of sperm were significantly decreased 8 weeks after the start of the treatment. The sperm production and motility in the multiple low-dose group appears to be reduced, while libido was maintained. In conclusion, multiple administration of 2.5 mg/kg bw busulfan depletes endogenous germ cells in the stallion recipients for SSC transplantation.

• 한국가금학회:학술대회논문집
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• 한국가금학회 2003년도 제20차 정기총회 및 학술발표회
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• pp.71-72
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• 2003

최근 생쥐에서 정원세포를 이용한 생식선 카이메라의 생산이 보고되었다. 정원세포의 경우 성축으로부터 세포를 다량으로 얻기가 쉬우며, 수용체 정소 내로 이식될 경우 생식선 카이메라의 생산능력이 있어서 이전의 배아줄기 세포를 이용할 때의 문제점을 효율적으로 해결할 수 있다. 또한 유전자가 도입된 정원세포의 이식에 의한 수용체 정소 내에서의 정자형성의 보고는 정원세포를 이용한 형질전환 동물의 생산 시스템으로의 개발 가능성을 보여준다. 본 실험에서는 닭에서 기존에 이용되어 왔던 형질전환 동물 생산 시스템의 문제점을 극복하고자 주령별 정원세포의 분리 및 이식을 통하여 조류에서 정원세포의 이식방법을 확립하고 생식선 카이메라 생산효율을 증진시키기 위하여 불임제인 부설판 등을 이용한 불임화 기술을 확립하여, 결국 조류에서의 형질전환 조류 생산 시스템으로서의 개발가능성을 제시하고자 한다.