Isolation and Culture of Adult Rat Liver and Identification of The Markers for Hepatic Oval Cells_CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Authors：

JIN Shilong , LIU Baohua , LIU Hongming , GU Hongguang , YANG Juntao , SHEN Haijun , XIAO Jing

Department of Hepatobiliary Surgery, Daping Hospital, The Third Military Medical University, Chongqing 400042， China;

Corresponding author：

Keywords：

Hepatic oval cell; Cell isolation; Culture

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Objective 　To study a simple and practical method of isolation, culture and identification of hepatic oval cells from adult rat.
Methods 　Wistar adult rats were fed by 2-acetaminofluorere （AAF） and were stimulated by partial hepatectomy to activate the proliferation of hepatic oval cells. After operation 12 days, the livers were resected for isolating oval cells. Hepatic tissue was digested by 0.10% collagenase Ⅳ and the obtained heterogeneous liver cells were then isolated and purified by density gradient centrifugation. The expressions of albumin and CK19 mRNA in hepatic oval cells were analyzed by immuno-fluorescence and RT-PCR.
Results　 The survival rate of the newly isolated oval cells was more than 90%. The hepatic stem cells were shown by immuno-fluorescence of stem cell’s antigen c-kit. The expressions of mRNA CK19 and albumin of the oval cell were also detected by PCR. The proliferation activity of the newly isolated oval cells was significantly high and they could be induced to differentiate into both hepatic and bile ductal cells by some growth factors.
Conclusion 　The successful development of the simple and feasible isolation and purification procedure as well as the identification method for hepatic oval cells may provide a fundamental for further studies about bionomics of the hepatic stem cell and the relation between stem cells and hepatic carcinoma.

Citation： JIN Shilong,LIU Baohua,LIU Hongming,GU Hongguang,YANG Juntao,SHEN Haijun,XIAO Jing. Isolation and Culture of Adult Rat Liver and Identification of The Markers for Hepatic Oval Cells. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2007, 14(1): 68-71. doi: Copy

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3. 　He ZP, Tan WQ, Tang YF, et al. Activation, isolation, identification and in vitro proliferation of oval cells from adult rat livers [J]. Cell Prolif， 2004; 37(2)∶177.
4. 　王宇明，陈耀凯，郎松，等. 大鼠肝卵圆细胞的分离培养及脾内移植研究 [J]. 中华肝脏病杂志, 2003; 11(6)∶328.
5. 　Weiskirchen R, Gressner AM. Isolation and culture of hepatic stellate cells [J]. Methods Mol Med， 2005; 117（1）∶99.
6. 　Qin AL, Zhou XQ, Zhang W, et al. Characterization and enrichment of hepatic progenitor cells in adult rat liver [J].World J Gastroenterol， 2004; 10(10)∶1480.
7. 　马俊勋，方驰华，张伟，等. 大鼠肝干细胞定位及形态学特征研究 [J]. 中华实验外科杂志， 2004； 21(8)∶940.
8. 　何忠杰，方弛华，马俊勋，等. 大鼠肝卵圆细胞免疫组化定位及超微结构观察 [J]. 解放军医学杂志， 2005； 30(4)∶332.
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10. 　Sakamoto T, Ezure T, Lunz J, et al.Concanavalin A simultaneously primes liver hematopoietic and epithelial progenitor cells for parallel expansion during liver regeneration after partial hepatectomy in mice [J]. Hepatology， 2000; 32(2)∶256.
11. 　张淑莲，曾平鲁，肖荣，等. 肝干细胞分离培养条件的筛选 [J]. 肝脏， 2003； 8(1)∶31.
12. 　姚鹏，詹轶群，许望翔，等. 细胞生长因子体外对大鼠肝干细胞的影响 [J]. 中华肝脏病杂志， 2003; 11(1)∶33.

CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Isolation and Culture of Adult Rat Liver and Identification of The Markers for Hepatic Oval Cells

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