肝动脉断流促进癌旁正常肝细胞上皮-间质转化

Hepatic arterial occlusion promotes epithelial-mesenchymal transition of peritumoral liver cells in hepatocellular carcinoma

目的观察肝动脉断流对癌旁正常肝细胞生物学特性的影响。方法肝动脉结扎（HAL）阻断裸鼠肝脏原位移植瘤（高转移性MHCC97H肝癌细胞）血供（nHAL=10，n假手术=10）。免疫组织化学（SP）法分别检测移植瘤内或癌旁缺氧诱导因子-1α（HIF-1α）、Pimonidazole及上皮-间质转化（EMT）标记分子如E-cadhefin和N-cadherin分子表达；体外用CoCl2模拟缺氧环境，免疫荧光和Westernblot检测缺氧处理后L-02肝细胞内上述分子的表达变化。结果HAL增加肝癌组织和癌旁肝组织内HIF-1α表达和Pimonidazole阳性细胞数（HAL组：9±3比假手术组：4±2，P〈0．05），减少癌旁肝细胞内E-cadhefin水平，增加其N-cadherin蛋白表达。100Ixmol／L CoCl2有效模拟体外缺氧环境，增加L-02肝细胞内HIF-1α表达，上调N-cadhefin同时降低E-cadhefin表达。结论HAL增加肝癌及癌旁组织缺氧，诱导癌旁正常肝细胞发生EMT。

Objective To investigate the effects of the hepatic arterial occlusion on biological characteristics of peritumoral liver cells in hepatocellular carcinoma （HCC）. Methods Using a metastatic human HCC orthotopic nude mice model （MHCC97H cells） , the effects of hepatic artery ligation （HAL） on the expression of hypoxia-inducible factor-1α （HIF-1α）, E-cadherin or N-cadherin, and pimonidazole positive staining were evaluated. To clarify the molecular background, the changes consistent with epithe- lial-mesenchymal transition （EMT） in normal liver cells L-02 were explored by using Western blotting and immunofluorescence staining under hypoxic conditions. Results HAL not only induced intratumoral hypoxia, but also led to peritumoral hypoxia as evidenced by higher HIF-1α level and augment number of pimonidazole positive cells （ HAL ： 9 ± 3vs. sham-operation ： 4 ± 2, P 〈 0. 05）. Besides the changes asso ciated with EMT were observed in HCC cells as demonstrated previously, and the molecular changes of EMT were also found in peritumoral liver cells, which were characterized by down-regulation of E-cadherin and up-regulation of N-cadherin. We introduced 100μmol/L CoC12 to mimic hypoxic conditions in vitro and found the HIF-1α expression was increased in L-02 cells. Moreover, the changes of EMT as described in vivo were also observed in these hypoxic cells by Western blotting or immunofluorescence staining. Conclusion HAL triggers the changes associated with EMT in tumor and peritumor tissues through inducing hypoxia.