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Title: [Effect of Kruppel-like factor 2 on the migration of human liver sinusoidal endothelial cells].
Author: Li N, Liu C, Pan DY, Tseng YZ, Zhou J, Zeng XQ, Luo TC, Chen SY.
Journal: Zhonghua Yi Xue Za Zhi; 2018 Feb 13; 98(7):527-532. PubMed ID: 29495223.
Abstract:
Objective: To investigate the effect and mechanism of Kruppel-like factor 2 (KLF2) on the migration of human liver sinusoidal endothelial cells (LSEC). Methods: Cultured human LSEC were infected with different lenti-viruses to overexpress or suppress KLF2 expression (LV5-KLF2 and LV3-shKLF2, respectively), the infection efficacies were examined by real-time PCR and Western blot analysis.Transwell migration assay was used to investigate the role of KLF2 on the migration of LSEC.The mRNA and protein expression of vascular endothelial growth factor receptor-2 (VEGFR-2) were detected by real-time PCR and Western blot analysis, respectively.The expression and phosphorylation of Src, P38 MAPK, and P44/42 MAPK were detected by Western blot. Results: The up-regulation of KLF2 expression dramatically inhibited migration of treated LSEC, compared with LV5-NC and WT control cells, fewer LV5-KLF2 cells migrated to the lower side of the filter after 12 h [ (35.6±1.4), (71.3±2.4) and (69.3±1.6), P<0.001 for all comparisons]. In contrast, the down-regulation of KLF2 expression promoted the migration of LSEC, more LV3-KLF2 cells migrated to the lower side of the filter compared with the LV3-NC and WT control cells [(189.5±5.4), (83.4±2.5) and (82.2±3.4), P<0.001 for all comparisons]. Furthermore, up-regulation of KLF2 reduced the mRNA and protein expression level of VEGFR2, while down-regulation of KLF2 significantly increased its expression in LSEC.Additionally, up-regulation of KLF2 inhibited the phosphorylation of Src, P38 MAPK, and P44/42 MAPK pathway in LSEC, whereas down-regulation of KLF2 promoted the phosphorylation of those signaling pathway proteins. Conclusions: KLF2 may inhibit the migration of human LSEC through the Src/ MAPK signaling pathway. 目的： 探索Kruppel样转录因子2(KLF2)对肝窦内皮细胞(LSEC)体外迁移的影响及其机制。 方法： 采用慢病毒感染的方式构建过表达和干扰KLF2表达的LSEC(分别为LV5－KLF2和LV3－shKLF2)，采用荧光定量PCR和蛋白印迹法检测病毒感染效率，未处理LSEC作为野生对照(WT)组。采用Transwell实验检测KLF2表达的改变对LSEC迁移能力的影响，荧光定量PCR和蛋白印迹法检测促迁移因子血管内皮细胞生长因子受体2(VEGFR2)的表达，蛋白印迹法检测迁移相关信号通路蛋白Src，P38 MAPK，P44/42 MAPK的表达。 结果： Transwell实验计数12 h细胞迁移数，过表达KLF2可抑制LSEC的体外迁移能力，LV5－KLF2组LSEC细胞迁移数目较LV5－NC和WT组少[(35.6±1.4)、(71.3±2.4)和(69.3±1.6)]，差异均有统计学意义(均P<0.001)。而下调KLF2表达后，LV3－shKLF2组LSEC的细胞迁移能力较LV5－NC和WT组增强，细胞迁移数目较对照组多[(189.5±5.4)、(83.4±2.5)和(82.2±3.4)]，差异均有统计学意义(均P<0.001)。荧光定量PCR和蛋白印迹法证实过表达KLF2可抑制LSEC中促迁移因子VEGFR2的mRNA和蛋白水平的表达，而干扰KLF2则上调VEGFR2的mRNA和蛋白表达水平。蛋白印迹结果提示过表达KLF2，抑制LSEC中信号通路蛋白p－Src、p－P38 MAPK及p－P44/42的表达，而干扰KLF2则促进LSEC中上述磷酸化蛋白的表达。 结论： KLF2可能通过Src/MAPK通路抑制LSEC的体外迁移。.

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