earticle

영어

Spermatogonial stem cells (SSCs) are the basis of spermatogenesis in male. Among the testicular germ cells, SSCs are considered as an important population because of their capability to multiply in numbers which is also known as self-renewal, and to differentiate into spermatocytes by subsequent meiotic processes finalized with the production of spermatozoa. As SSCs are present in a very small proportion among the total testis cells, in vitro proliferation of SSCs in undifferentiated state is very important in the field of experiments related to germ cell culture. The objective of the study was to establish an effective feeder cell for murine germ cell development in vitro. We used three types of feeder cells to grow murine germ cells, such as STO, tissue specific feeder layer, and C166 feeder cells and green fluorescence protein (GFP) rat SSCs were used to grow on them. A continuous culture was performed for each feeder groups, but C166 feeder cells found unable to proliferate germ cells even after several weeks. We observed a comparatively better proliferation rate of rat germ cells grown on tissue specific feeder layer where cultured germ cell colony and cell count were higher than STO feeder. Immunocytochemistry of undifferentiated SSC marker showed significantly higher expression for germ cell cultured on tissue specific feeder layer. Similarly, mRNA level of undifferentiated spermatogonia were found higher in case of tissue specific feeder layer and STO, although the expression level of differentiated spermatogonia was similar. Finally, the functional characteristics of SSCs were studied by cultured germ cell transplantation into recipient mice. We used donor rat germ cells cultured on both feeder groups for 4 months, where SSCs from tissue specific feeder layer showed higher functional and stemness activity. Transplantation of SSCs cultured for 8 months from tissue specific feeder layer group were performed to recipient rat and we could produce GFP offspring from those recipients. Thus the experiment conclude that tissue specific cells could be an effective feeder layer for murine SSCs culture based on their survivability and proliferation along with retaining the perfect functional characteristics and stemness properties.