WASF3反义寡核苷酸对非小细胞肺癌细胞增殖和迁移的影响及其机制

目的探讨Wiskott-Aldrich综合征蛋白家族成员3的反义寡核苷酸（WASF3-AS）对非小细胞肺癌（NSCLC）细胞增殖和迁移的影响及其作用机制。方法RT-qPCR法检测人NSCLC细胞株A549细胞和人正常肺细胞MRC-5中WASF3的表达水平。将A549细胞分为空白对照组、WASF3-NC组（阴性对照组）、WASF3-AS组（反义寡核苷酸WASF3组），应用RT-qPCR法检测WASF3-AS对A549细胞中WASF3表达的影响；应用CCK-8法检测细胞增殖能力，克隆实验检测细胞克隆能力，Transwell法检测细胞迁移能力；RT-qPCR和Western blotting法分别检测WASF3-AS对A549细胞中磷酸酶和张力蛋白同源基因（PTEN）、磷脂酰肌醇-3-激酶（PI3K）、蛋白激酶B（p-Akt）mRNA和蛋白表达水平。结果与人正常肺细胞MRC-5比较，A549细胞各组中WASF3的mRNA相对表达量均增加（P < 0.05~P < 0.01）；空白对照组和WASF3-NC组的WASF3的mRNA相对表达量差异无统计学意义（P>0.05），但均高于WASF3-AS组（P < 0.05和P < 0.01）。与WASF3-NC组比较，转染后48、72、96 h WASF3-AS组细胞增殖水平明显降低（P < 0.05）。细胞转染14 d后，与空白对照组和WASF3-NC组比较，WASF3-AS组细胞克隆形成率和细胞迁移数目均明显降低（P < 0.05）。与WASF3-NC组比较，WASF3-AS组细胞的PTEN mRNA和蛋白相对表达量明显增加，PI3K、p-Akt mRNA和蛋白相对表达量明显减少（P < 0.05）。结论人NSCLC细胞的WASF3表达水平明显高于正常肺细胞；WASF3-AS可能通过调控PTEN-PI3K/Akt信号通路，抑制NSCLC的增殖、克隆和迁移。

Study on the effects of WASF3 antisense oligonucleotides on the proliferation and migration of non-small cell lung cancer and its mechanism

ObjectiveTo investigate the effects of antisense oligonucleotides of Wiskott-Aldrich syndrome protein family member 3(WASF3-AS) on the proliferation and migration of non-small cell lung cancer(NSCLC) cells and its mechanism.MethodsThe expression levels of WASF3 in human NSCLC cell lung cancer cell line A549 and human normal lung cell MRC-5 were detected using RT-qPCR. The A549 cells were divide into the blank control group, WASF3-NC group(negative control group) and WASF3-AS group(antisense oligonucleotide WASF3 group). The effects of WASF3-AS on the WASF3 expression in A549 cells were detected using RT-qPCR. The CCK-8 method was used to detect the cell proliferation ability, the cloning experiment was used to detect the cell cloning ability, and the Transwell method was used to detect the cell migration ability. The mRNA and protein levels of the phosphatase and tensin homolog(PTEN), phosphatidylinositol-3-kinase(PI3K) and protein kinase B(p-Akt) in A549 cells of three groups were detected using RT-qPCR and Western blotting, respectively.ResultsCompared with the human normal lung cells MRC-5, the mRNA expression levels of WASF3 in three group increased(P < 0.05 to P < 0.01). There was no statistical significance in the mRNA expression level of WASF3 between the blank control group and WASF3-NC group(P>0.05), but it was higher than that of WASF3-AS group(P < 0.05 and P < 0.01). Compared with the WASF3-NC group, the proliferation levels of cells in WASF3-AS group significantly decreased after 48, 72 and 96 h of transfection(P < 0.05). After 14 days of cell transfection, the clone formation rate and number of cell migration in WASF3-AS group significantly decreased compared with the blank control group and WASF3-NC group(P < 0.05). Compared with the WASF3-NC group, the expression levels of PTEN mRNA and protein in WASF3-AS group significantly increased, while the expression levels of PI3K and p-Akt mRNA and protein significantly decreased(P < 0.05).ConclusionsThe expression level of WASF3 in human NSCLC cells is significantly higher than that in normal lung cells. The WASF3-AS may inhibit the proliferation, cloning and migration of NSCLC by regulating the PTEN-PI3K/Akt signaling pathway.