Objective: To investigate the effect of aquaporin 5(AQP5) on proliferation and migration of ectopic endometrial epithelial cells.
Methods: AQP5 shRNA interference fragments were designed and transfected into ectopic endometrial epithelial cells stably by lentivirus technology. Fluorescence quantitative RT-PCR and Western blotting were used to detect the AQP5 mRNA and protein expression, respectively. The cell proliferation and migration were determined by using MTT method and Transwell system, respectively. Levels of phosphorylated AKT(p-AKT) and total AKT were examined by Western blotting. The nude mice model of endometriosis was constructed and the endometrial cell nodule formation was observed.
Results: AQP5 shRNA transfection inhibited cell proliferation and migration compared with control group (both P<0.05). The activation of AKT in AQP5 shRNA transfected cells was lower than that in control cells (P<0.01). Compared to control group, the endometrial cells nodule formation was suppressed in mice inoculated with AQP5 shRNA-silencing ectopic endometrial epithelial cells.
Conclusion: Down-regulation of AQP5 expression can suppress the proliferation and migration of ectopic endometrial epithelial cells and endometrial cell nodule formation in nude mice, in which AKT pathway may be involved.
目的: 探讨水通道蛋白(AQP)5基因对异位子宫内膜腺上皮细胞增殖及迁移的影响。
方法: 构建可靶向沉默AQP5基因的质粒, 在293T细胞中包装形成病毒颗粒后, 感染原代培养的异位子宫内膜腺上皮细胞, 分别通过逆转录PCR及蛋白质印迹法鉴定异位子宫内膜腺上皮细胞中AQP5 mRNA和蛋白表达。MTT法测定细胞增殖; Transwell技术检测细胞的体外迁移能力; 蛋白质印迹法检测丝氨酸/苏氨酸蛋白酶(AKT)活化情况。构建裸鼠腹腔内子宫内膜异位症体内模型, 考察阴性对照组和AQP5沉默组子宫内膜腺上皮细胞在裸鼠腹腔内瘤体生长情况及腹膜转移情况。
结果: 体外实验结果显示, AQP5沉默后异位子宫内膜腺上皮细胞的增殖能力在第7天时明显抑制( P < 0.05);AQP5沉默组异位子宫内膜腺上皮细胞磷酸化(p-AKT)表达减少, 而AKT表达无改变, p-AKT/AKT减少( P < 0.05);AQP5沉默组比阴性对照组迁移细胞数减少( P < 0.05)。体内实验结果显示, 与阴性对照组比较, AQP5沉默组瘤体结节数相对较少, 体积较小, 其中腹膜瘤体结节数少于阴性对照组( P < 0.05)。
结论: AQP5基因沉默可抑制异位子宫内膜腺上皮细胞的增殖及迁移能力, 其机制可能与AKT活化有关。