干扰Hmgb3表达对肺腺癌A549细胞增殖的影响

doi:10.3971/j.issn.1000-8578.2014.10.001

干扰Hmgb3表达对肺腺癌A549细胞增殖的影响

Effect of Interfered Hmgb3 Expression on Proliferation of Lung Carcinoma Cells A549 by Inducible shRNA Tet-on Lentiviral

摘要

摘要: 目的构建针对Hmgb3的shRNA诱导型慢病毒，检测诱导稳定感染细胞株干扰序列的干扰效率及其对A549细胞增殖的影响。方法设计Hmgb3 及对照GFP shRNA序列，装入Tet-pLKO-puro 质粒，进行慢病毒包装，感染A549细胞，嘌呤霉素筛选稳定株，筛选最佳诱导浓度并诱导干扰序列表达， Western blot检测干扰效率，MTT实验检测其对A549细胞增殖的影响。结果构建的Hmgb3 shRNA慢病毒稳定细胞株诱导后可显著抑制Hmgb3的表达，干扰效率达73%（P<0.05），而对照组无明显干扰效果（P=0.721），对照组的增殖率为94%，实验组的增殖率下降为46%（P<0.05）。结论成功构建了针对Hmgb3的shRNA诱导型慢病毒，并有效沉默A549细胞靶基因，干扰Hmgb3对肺癌细胞的增殖有一定的抑制作用。

Abstract: Objective To test the proliferation effect of Hmgb3 on lung cancer cell line A549 after doxycyline-induced Hmgb3 shRNA lentivirus was constructed and packaged. Methods Hmgb3 and GFP shRNA sequences were designed and inserted into Tet-pLKO-puro plasmid to package lentivirus to infect A549 cells. Stable infected cells were obtained by puromycin screening. The doxycyline concentration was optimized to obtain the best induction effect. Western blot was used to test the silencing effect and MTT was for proliferation. Results Hmgb3 shRNA lentivirus could significantly reduce Hmgb3 expression in A549 cells and the efficiency of silencing was up to 73% (P<0.05). In the meantime, the proliferation rate of A549 cells in experimental group(46%)was much lower than that in control group(94%)(P<0.05). Conclusion Inducible Hmgb3 shRNA lentivirus was successfully constructed and silenced effectively target gene in A549 cells. Interfered Hmgb3 expression could inhibit the growth of A549 cells.

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