Morphological and functional heterogeneity of the mouse intrahepatic biliary epithelium.
Morphological and functional heterogeneity of the mouse intrahepatic biliary epithelium.
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DOI：
10.1038/labinvest.2009.6
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发表时间：
2009-04
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--
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大鼠和人胆道上皮在形态和功能上是不同的。由于没有关于小鼠肝内胆道上皮异质性的信息，并且随着转基因小鼠模型研究肝脏疾病发病机制的增加，我们试图在正常和胆汁淤积小鼠模型中评估小胆管和大胆管以及纯化胆管细胞的形态学、分泌和增殖表型。形态学上，取正常和BDL小鼠（C57/BL6）肝脏，切成2 ~ 4 μm2块，石蜡包埋，切片，H&E染色。采用SigmaScan测量胆管和胆管细胞直径，评估胆管和胆管细胞的大小。在大小正常和BDL胆管细胞中，我们通过免疫荧光和western blot评估了胆管细胞特异性标志物角蛋白-19 （KRT19）、分泌素受体（SR）、囊性纤维化跨膜传导调节剂（CFTR）和碳酸氢盐阴离子交换剂2 （Cl-/HCO-3 AE2）的表达；细胞内cAMP水平和氯离子外排对分泌素的反应（100 nM）。为了评估胆管结扎（BDL）后胆管细胞的增殖反应，从BDL小鼠中分离出大小胆管细胞。通过流式细胞仪（FACS）分析细胞周期来确定增殖情况，通过肝切片中krt19阳性胆管的数量来确定胆管质量。原位形态测定证实小鼠胆道上皮在形态上存在异质性，较小的胆管细胞内衬较小的胆管，较大的胆管细胞内衬较大的胆管。小胆管细胞和大胆管细胞都表达KRT19，只有正常小鼠和BDL小鼠的大胆管细胞表达SR、CFTR和Cl-/HCO-3交换物，并对分泌素产生cAMP水平升高和氯离子外排的反应。BDL后，只有大鼠胆管细胞增生。与大鼠相似，小鼠肝内胆道上皮在形态和功能上都是异质的。小鼠是定义胆道树异质性的合适模型。
Rat and human biliary epithelium is morphologically and functionally heterogeneous. Since no information exists on the heterogeneity of the murine intrahepatic biliary epithelium, and with increased usage of transgenic mouse models to study liver disease pathogenesis, we sought to evaluate the morphological, secretory and proliferative phenotypes of small and large bile ducts and purified cholangiocytes in normal and cholestatic mouse models. For morphometry, normal and BDL mouse livers (C57/BL6) were dissected into blocks of 2-4 μm2, embedded in paraffin, sectioned, and stained with H&E. Sizes of bile ducts and cholangiocytes were evaluated by using SigmaScan to measure the diameters of bile ducts and cholangiocytes. In small and large normal and BDL cholangiocytes, we evaluated the expression of cholangiocyte specific markers, keratin-19 (KRT19), secretin receptor (SR), cystic fibrosis transmembrane conductance regulator (CFTR), and chloride bicarbonate anion exchanger 2 (Cl-/HCO-3 AE2) by immunofluorescence and western blot; and intracellular cAMP levels and chloride efflux in response to secretin (100 nM). To evaluate cholangiocyte proliferative responses after bile duct ligation (BDL), small and large cholangiocytes were isolated from BDL mice. The proliferation status was determined by analysis of the cell cycle by FACS and bile duct mass was determined by the number of KRT19-positive bile ducts in liver sections. In situ morphometry established that the biliary epithelium of mice is morphologically heterogeneous, which smaller cholangiocyte lining smaller bile ducts and larger cholangiocytes lining larger ducts. Both small and large cholangiocytes express KRT19 and only large cholangiocytes from normal and BDL mice express SR, CFTR, and Cl-/HCO-3 exchanger and respond to secretin with increased cAMP levels and chloride efflux. Following BDL, only large mouse cholangiocytes proliferate. Similar to rats, mouse intrahepatic biliary epithelium is morphologically, and functionally heterogeneous. The mouse is a suitable model for defining the heterogeneity of the biliary tree.
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