人胎肝干细胞在重症联合免疫缺陷小鼠损伤肝脏中的植入  

Engrafting human fetal liver stem cells into injured livers of mice with severe combined immunodeficiency disorder

作　　者：苏娟[1] 李文林[1] 陶欣荣[1] 朱海英[1] 王新民[1] 李建秀[1] 田棣[1] 余宏宇[2] Joseph T Lau 胡以平[1] 

机构地区：[1]第二军医大学基础医学部细胞生物学教研室 [2]第二军医大学长征医院病理科 [3]The Department of Molecular and Cellular Biology,Roswell Park Cancer Institute,Elm and Carlton Streets,Buffalo,NY 14263,USA

出　　处：《第二军医大学学报》2005年第3期243-246,共4页Academic Journal of Second Military Medical University

基　　金：国家自然科学基金( 30470876; 30270668;30270603;30200138);上海市科委重大项目(03DJ14020)

摘　　要：目的:从孕4.5～6周的人胚胎肝脏中分离培养肝干细胞,并观察其在重症联合免疫缺陷(SCID)小鼠损伤肝脏中的植入.方法:从孕4.5～6周的人胚胎肝脏中分离干细胞并进行体外长期培养,用RT PCR方法检测肝干细胞相关分子标志以及八聚物结合蛋白4(Oct-4)的表达,然后植入经四氯化碳(CCl4)造成急性肝损伤的SCID小鼠体内,在移植后2、l0、17d分别对受体鼠肝脏冰冻切片进行荧光观察和H-E染色.结果:所建立的胎肝细胞系不仅表达二肽水解酶Ⅳ(DPPⅣ)、白蛋白(Alb)、细胞角蛋白(CK)19、γ-谷氨酰转肽酶(γ-GT)、CK18、肝细胞生长因子受体(c-met)、干细胞因子受体(c-kit)等肝干细胞相关标志,还表达干细胞转录因子Oct-4.在细胞移植后2 d,肝脏冰冻切片未见绿色荧光;移植后10 d和17d肝脏冰冻切片见散在的绿色荧光.对同一切片做H-E染色,发现绿色荧光部位为肝板内的多个成熟肝细胞.结论:从人胚胎肝脏中分离的干细胞能够植入SCID小鼠的损伤肝脏.Objective:To separate hepatic stem cells from human fetal livers of 4.5-6-week gestation and observe their engraftment into injured livers of mice with severe combined immunodeficiency (SCID). Methods: Stem cells isolated from human fetal livers were long-term cultured in vitro. RT-PCR was used to detect expression of hepatic stem cell related markers and octamer-binding protein 4 (Oct-4) in those cells. Then they were transplanted into SCID mouse liver with acute injury caused by carbon tetrachloride (CCl 4 ). Liver cryostat sections of recipient mice were observed by a fluorescence microscope at 2 d, 10 d and 17 d post-transplantation,then hematoxylin and eosin (H-E) dying was performed on the same liver sections. Results: The fetal liver cell lines not only expressed hepatic stem cell related markers such as dipeptidylpeptidase Ⅳ (DPPⅣ), albumin (Alb), cytokeratin (CK) 19, gamma-glutamyl transpeptidase (γ-GT), CK18, hepatocyte growth factor receptor (c-met), stem cell factor receptor (c-kit), but also expressed stem cell transcription factor Oct-4. At 2 d after transplantation no green fluorescence was observed in the recipient livers. At 10 d and 17 d several cell clusters with green fluorescence in different sizes were observed. H-E staining confirmed that there were clusters of hepatocytes in hepatic cord. Conclusion: The stem cells isolated from human fetal liver can be engrafted into injured livers of SCID mice.

关 键 词：肝干细胞 胚胎 重症联合免疫缺陷 细胞移植 

分 类 号：R593.3[医药卫生—内科学]