膜联蛋白A7表达抑制对HepG-2细胞中galectin-3及PKC表达的影响

Expression of galectin-3 and PKC in annexin A7 knockdown HepG-2 Cells

目的探讨抑制膜联蛋白A7(ANXA7)表达对人肝癌HepG-2细胞半乳糖凝集素3(galectin-3)的影响以及联合佛波酯(PMA)激活PKC后对ANXA7表达的影响。方法将细胞分为siRNA干扰组、阴性对照组和空白对照组。采用RNA干扰技术将靶向ANXA7的siRNA和阴性对照siRNA脂质体转染法分别转染肝癌HepG-2细胞,空白对照组不予任何处理。转染48h后,采用Western blot和RT-qPCR法进行抑制效果的鉴定。Western blot和RT-qPCR法分别检测ANXA7表达抑制的HepG-2细胞中galectin-3的表达。转染48h后用100ng/ml PMA处理24h,Western blot法检测ANXA7及PKC的表达。结果靶向ANXA7的siRNA可显著抑制ANXA7的表达;ANXA7表达受抑后的HepG-2细胞中galectin-3的表达下降,与阴性对照组和空白对照组相比,差异有显著性(P<0.05)。PMA处理24h后,ANXA7及PKC表达变化不显著。结论 ANXA7表达抑制的HepG-2细胞中galectin-3的表达显著下调,这可能是ANXA7表达受抑的细胞行为学发生改变的机制之一。

Obj ective To detect the expression of galectin-3 and PKC activated by PMA in annexin A7(ANXA7)knockdown HepG-2 cells.Methods The HepG-2 cells were grouped into experiment group, negative group and blank group.RNA interference technology was usedto transfect ANXA7-targeted siR-NA,and scrambled siRNA into HepG-2 cells with lipofectineTM2000,without any treatment to the blank group.About 48 h after transfection,western blot and RT-qPCR were used to assure the suppression of ANXA7.Then the expression of galectin-3 was detected respectively with western blot and RT-qPCR a-bout 48 h after transfection and about 24 h after 100ng/ml PMA treatment,western blot was used to de-tect the expression of PKC and ANXA7.Results The expression of ANXA7 was significantly suppressed in the cells transfected with ANXA7 siRNA.When ANXA7 expression was suppressed,the expression of galectin-3 decreased significantly compared with that of the negtive control group and blank control group (P<0.05).There was no obvious change in the expression of ANXA7 and PKC about 24 h after 100ng/mL PMA treatment.Conclusion The expression of galectin-3 was down-regulated in the ANXA7 sup-pressed HepG-2 cells,which may be one of the mechanisms for the abnormal behaviour of the ANXA7 knockdown carcinoma cells.