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目的明确小鼠肝脏中的细胞角蛋白19(cytokeratin 19,CK19)阳性(CK19~+)细胞是否可分化为成熟肝细胞。方法利用CK19~(CreERT)小鼠和Rosa26-GFP小鼠杂交得到CK19~(CreERT)/Rosa26-GFP双转基因小鼠,注射他莫昔芬(tamoxifen,TM)后检测小鼠肝脏中CK19~+细胞的GFP标记情况。在此基础上分别构建3,5-二乙氧基羰基-1,4-二氢-2,4,6-三甲基吡啶和四氯化碳(carbon tetrachloride,CCl4)肝脏损伤模型,采用肝脏组织冰冻切片结合免疫荧光染色检测肝脏中GFP标记的CK19~+细胞的分化情况。结果获得CK19~(CreERT)/Rosa26-GFP双转基因小鼠,免疫荧光染色结果显示CK19~+细胞可被GFP标记。在DDC肝脏损伤模型小鼠中检测到增生性小胆管内有GFP~+细胞,这些GFP~+细胞表达胆管上皮细胞标志物CK19,且DDC肝损伤模型小鼠中GFP~+胆管细胞比例高于未损伤对照组[(63.5±6.3)%vs(53.6±4.8)%,P<0.05];在CCl4肝损伤模型组小鼠中检测到肝脏实质细胞中有GFP~+细胞,这些GFP~+细胞表达成熟肝细胞标志物白蛋白(ALB),且CCl4肝损伤模型组小鼠肝脏中GFP~+细胞比例高于未损伤对照组[(0.15±0.02)%vs(0.008±0.003)%,P<0.01]。结论小鼠肝脏内的CK19~+细胞群体中存在具有肝向分化潜能的前体细胞,可能为真正的肝干细胞的识别鉴定提供一个新线索。
Objective To determine whether cytokeratin 19 (CK19) positive (CK19 ~ +) cells in mouse liver can differentiate into mature hepatocytes. Methods CK19 Creer / Rosa26-GFP double transgenic mice were crossed with CK19 Creer and Rosa26-GFP mice. After injection of tamoxifen (TM), CK19 ~ GFP labeling of cells. On the basis of these, a model of hepatic injury induced by 3,5-diethoxycarbonyl-1,4-dihydro-2,4,6-trimethylpyridine and carbon tetrachloride (CCl4) Tissue frozen section and immunofluorescence staining were used to detect the differentiation of GFP-labeled CK19 ~ + cells in the liver. Results The CK19 ~ (CreERT) / Rosa26-GFP double transgenic mice were obtained. Immunofluorescence staining showed that CK19 ~ + cells could be labeled by GFP. GFP ~ + cells were detected in proliferating small bile duct in DDC liver injury model mice. These GFP ~ + cells expressed bile duct epithelial cell marker CK19, and the proportion of GFP ~ + cholangiocytes in DDC liver injury model mice was higher than (63.5 ± 6.3)% vs (53.6 ± 4.8)%, P <0.05]. In the CCl4 liver injury model group, GFP ~ + cells were detected in the liver parenchymal cells. These GFP ~ + cells (ALB), and the percentage of GFP ~ + cells in the CCl4 liver injury model group was significantly higher than that in the untreated control group [(0.15 ± 0.02)% vs (0.008 ± 0.003)%, P < 0.01]. Conclusion The presence of hepatocyte differentiation potential in CK19 ~ + cell population in mouse liver may provide a new clue for identification and identification of real hepatic stem cells.