RNAi下调LOX表达对人肺癌细胞乏氧转移的影响及其分子机制

目的观察RNA干扰（RNAi）下调赖氨酸氧化酶（lysyl oxidase，LOX）表达对人肺癌细胞95D乏氧转移的影响及其分子机制。方法应用针对人LOX基因的小于扰RNA（LOX siRNA）转染常氧（19％O2）95D细胞，24h后将细胞蚩乏氧（0．5％O2）培养箱孵育，乏氧24h后采用实时PCR检测细胞LOX mRNA和Snail mRNA表达，Western blot技术评价Src、磷酸化Src（P-Src Y418）和Snail蛋白表达，Transwell小空评价细胞侵袭能力．结果与常氧95D细胞比较，乏氧引起细胞LOX mRNA表达和细胞侵袭能力分别增加14倍和2．12倍。与对照siRNA组比较，LOX siRNA转染使乏氧细胞LOX mRNA表达下降70％～75％，侵袭能力下降约30％，P—Src Y418和Snail蛋白表达降低约40％（P〈0．05），但不影响Src蛋白表达和Snail mRNA水平（P〉0．05）。结论LOX表达下调降低人肺癌细胞乏氧侵袭与减少Src活化和Snail蛋白表达有关，为LOX成为防治肺癌乏氧转移的潜在靶点提供了实验依据。

Influence of LOX downregulation by RNAi on hypoxic metastasis of human lung cancer cell and the underlying molecular mechanism

Objective To observe the influence of lysyl oxidase（LOX） downregulation via RNAi on hypoxic metastasis of human lung cancer cell 95D and stduy its molecular mechanism. Methods LOX siRNA was used to transfect 95D cell line in normoxia （19% O2）. After 24-hour incubation, the cells were cuhm＇ed in hypoxic incubator （0. 5% O2） for 24h. Real-time PCR assay was applied to detect LOX mRNA and Snail mRNA expression. Levels of Src, phosphoiTlation of Src （P-Src Y418） and Snail protein were determined by Western blot assay. Transwell chamber was used to evaluate the cellular invasion potential. Results Compared with 95D cells under normoxic conditions, hypoixa treatment increased LOX mRNA expression by 14 times and invasion ability by 2. 12 times respectively. Compared with siRNA control group, LOX siRNA transfection decreased LOX mRNA expression, the invasion ability of hypoxic ceils, and the protein expression of P-Src Y418 and Snail by 70% -75% , about 30% , and about 40% respectively （P 〈0. 05）. However, it didn＇t affect the expression level of Src protein or Snail mRNA （ P 〉 0. 05 ）. Conclusions hnpaired metastatic potential of hypoxic human lung cancer cell induced by LOX downregulation is associated with reduced expression level of Src activation and Snail protein. The present data provids experimental evidence for LOX as a potential target for prevention and treatment of lung cancer metastasis under hypoxia.