锦灯笼醇提取物通过lncRNA AK093987基因的表达对结肠癌细胞增殖、凋亡的影响及机制研究

Experimental study on the effects of Jingdenglong ethanol extract on proliferation and apoptosis of colon cancer cells through the expression of lncRNA AK093987 gene

目的探讨锦灯笼醇提取物对长链非编码RNA(lncRNA)AK093987的影响,以及对结肠癌细胞增殖、凋亡的调控作用与机制。方法结肠癌细胞HT29分为8组,即对照(NC组),2.5、5、10μg/ml锦灯笼醇提取物(低、中、高剂量组),转染si-NC(si-NC组),转染si-lncRNA AK093987(si-lncRNA AK093987组),10μg/ml锦灯笼醇提取物+转染pcDNA-NC(高剂量+pcDNA-NC组),10μg/ml锦灯笼醇提取物+转染pcDNA-lncRNA AK093987(高剂量+pcDNA-lncRNA AK093987组)。应用Western blot、细胞计数试剂盒8(CCK-8)、流式细胞仪和qRT-PCR检测细胞周期蛋白D1(CyclinD1)、活化-含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved-caspase-3)表达、细胞增殖能力、凋亡率和lncRNA AK093987表达。结果与NC组比较,低、中和高剂量组锦灯笼醇提取物减少结肠癌细胞HT29中CyclinD1蛋白、lncRNA AK093987的表达水平,降低细胞增殖能力,增加凋亡率、Cleaved-caspase-3蛋白表达水平,差异有统计学意义(P<0.05)。si-lncRNA AK093987组结肠癌细胞HT29中lncRNA AK093987、CyclinD1蛋白的表达水平和增殖能力低于si-NC组,Cleaved-caspase-3蛋白的表达水平和凋亡率高于si-NC组,差异均有统计学意义(P<0.05)。高剂量+pcDNA-lncRNA AK093987组结肠癌细胞HT29中lncRNA AK093987、CyclinD1蛋白的表达水平和增殖能力高于高剂量+pcDNA-NC组,Cleaved-caspase-3蛋白的表达水平和凋亡率低于高剂量+pcDNA-NC组,差异均有统计学意义(P<0.05)。结论锦灯笼醇提取物通过下调lncRNA AK093987表达,减轻结肠癌细胞的增殖和加速凋亡。

Objective To investigate the effects of Jingdenglong ethanol extract on the long non coding RNA(lncRNA)AK093987,and its regulation effects and mechanism on the proliferation and apoptosis of colon cancer cells.Methods Colon cancer cells(HT29)were divided into 8 groups:control group,Jingdenglong ethanol extract low,medium,high dose groups(2.5,5,10μg/ml),and si NC transfection group,si lncRNA AK093987 transfection group,high-dose Jingdenglong ethanol extract+pcDNA NC transfection group(group A),and high-dose Jingdenglong ethanol extract+pcDNA lncRNA AK093987 transfection group(group B).Western Blot,cell counting kit 8(CCK-8),flow cytometry and qRT PCR were used to detect the expression levels of cyclin D1(cyclin D1),cleaved cysteine containing aspartate proteolytic enzyme 3(cleaved caspase 3),cell proliferation capacity,apoptosis rate and lncRNA AK093987 expression.Results As compared with those in control group,the expression levels of cyclin D1 protein,lnRNA AK093987 in the low,medium,and high dose groups were significantly decreased,and the cell proliferation rate was decreased,however,the apoptosis rate and cleaved caspase 3 protein expression level were significantly increased in colon cancer cells HT29(P<0.05).The expression levels of lncRNA AK093987 and cyclin D1 protein,and the proliferation capacity in si lncRNA AK093987 group were significantly lower than those in si NC group,however the expression levels of cleaved caspase 3 protein and apoptosis rate were significantly higher than those in si NC group,(P<0.05).The expression levels of lncRNA AK093987 and cyclin D1 protein,and the proliferation ability in group B were significantly higher than those in group A,however the expression levels of cleaved caspase 3 protein and apoptosis rate were significantly lower than those in group A(P<0.05).Conclusion Jingdenglong ethanol extracton can reduce the proliferation of colon cancer cells and accelerate the apoptosis by down regulating the expression of lncRNA AK093987.