Portal fibroblasts regulate the proliferation of bile duct epithelia via expression of NTPDase2
Portal fibroblasts regulate the proliferation of bile duct epithelia via expression of NTPDase2
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DOI：
10.1074/jbc.m412371200
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发表时间：
2005-06-17
影响因子：
4.8
通讯作者：
Dranoff, JA
中科院分区：
文献类型：
--
作者：
Jhandier, MN;Kruglov, EA;Dranoff, JA
胆管上皮是许多以胆管增生紊乱为特征的“胆管疾病”的靶点。尽管胆管增生的机制尚不清楚，但最近的证据表明，细胞外核苷酸通过激活P2Y受体调节细胞增殖。门脉成纤维细胞可能通过外核苷酸酶NTPDase2的表达调节胆管上皮P2Y受体。因此，我们验证了门脉成纤维细胞通过表达NTPDase2调节胆管上皮增殖这一假设。我们建立了一种新的MzChA - 1人胆管癌细胞和原代门脉成纤维细胞的共培养模型。通过溴脱氧尿苷摄取来测量细胞增殖。通过免疫荧光和实时定量逆转录聚合酶链反应评估NTPDase2的表达。使用小干扰RNA阻断门脉成纤维细胞中NTPDase2的表达。通过cDNA转染实现从胆管结扎大鼠分离的门脉肌成纤维细胞中NTPDase2的过表达。Mz - ChA - 1细胞与门脉成纤维细胞共培养使其增殖降低至对照的26%。可溶性外核苷酸酶三磷酸腺苷双磷酸酶和P2受体抑制剂苏拉明也诱导了Mz - ChA - 1增殖的类似降低。当使用NTPDase2特异性小干扰RNA抑制门脉成纤维细胞中NTPDase2的表达时，Mz - ChA - 1细胞的增殖恢复到基线水平。未转染且缺乏NTPDase2的门脉肌成纤维细胞对Mz - ChA - 1增殖没有影响，然而转染了NTPDase2 cDNA的门脉肌成纤维细胞抑制了Mz - ChA - 1增殖。我们得出结论，门脉成纤维细胞通过表达NTPDase2和阻断P2Y激活来抑制胆管增生。NTPDase2的缺失可能介导梗阻性胆汁淤积所特有的胆管增生。这种新的交互信号通路可能介导其他胆管疾病状况下胆管增生的病理改变。
Bile duct epithelia are the target of a number of "cholangiopathies" characterized by disordered bile ductular proliferation. Although mechanisms for bile ductular proliferation are unknown, recent evidence suggests that extracellular nucleotides regulate cell proliferation via activation of P2Y receptors. Portal fibroblasts may regulate bile duct epithelial P2Y receptors via expression of the ecto-nucleotidase NTPDase2. Thus, we tested the hypothesis that portal fibroblasts regulate bile duct epithelial proliferation via expression of NTPDase2. We generated a novel co-culture model of MzChA-1 human cholangiocarcinoma cells and primary portal fibroblasts. Cell proliferation was measured by bromodeoxyuridine uptake. NTPDase2 expression was assessed by immunofluorescence and quantitative real-time reverse transcription PCR. NTPDase2 expression in portal fibroblasts was blocked using short interfering RNA. NTPDase2 overexpression in portal myofibroblasts isolated from bile duct-ligated rats was achieved by cDNA transfection. Co-culture of Mz-ChA-1 cells with portal fibroblasts decreased their proliferation to 26% of control. Similar decreases in Mz-ChA-1 proliferation were induced by the soluble ecto-nucleotidase apyrase and the P2 receptor inhibitor suramin. The proliferation of Mz-ChA-1 cells returned to baseline when NTPDase2 expression in portal fibroblasts was inhibited using NT-PDase2-specific short interfering RNA. Untransfected portal myofibroblasts lacking NTPDase2 had no effect on Mz-ChA-1 proliferation, yet portal myofibroblasts transfected with NTPDase2 cDNA inhibited Mz-ChA-1 proliferation. We conclude that portal fibroblasts inhibit bile ductular proliferation via expression of NTPDase2 and blockade of P2Y activation. Loss of NTPDase2 may mediate the bile ductular proliferation typical of obstructive cholestasis. This novel cross-talk signaling pathway may mediate pathologic alterations in bile ductular proliferation in other cholangiopathic conditions.