Isolation, Culture and Identification of Cattle Liver Stem Cells
SHAO Qi1, ZHANG Cai1,2*, LI Pengfei1, SONG Xinru1, LI Yuanxiao2, WANG Yuqin2
The aim of this study was to explore the in vitro growth characteristics of live stem cells isolated from Holstein dairy cattle, which provided research tools for comparative medical science. The stem cells were isolated by the modified two-step collagenase perfusion method, then purified by percoll single density gradient centrifugation. The expression of liver stem cell surface markers was detected by immunofluorescence and PCR. The isolated cells were adherent and emerged active proliferation capacity after 48 h. Cells cultured in low growth density can differentiate into hepatocyte-like cells after cultured 504 h. Cells in normal growth density can be transmitted to the P3 generation, which had large cell cytoplasm ratio and was mononuclear, infantile and in low differentiation state, shown by HE staining. In addition, AFP (alpha-fetoprotein), EpCAM (epithelial cell adhesion protein), hematopoietic stem cell surface markers CD34, c-kit (stem cell factor receptor protein), cell keratin CK18, CK19 were expressed by the P3 generation cells. The results indicated that the isolated liver stem cells were well characterized and expressed the markers of mature hepatocytes and bile duct epithelial cells.
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