摘要
目的探讨体外干扰乳酸脱氢酶A(LDHA)表达对人类表皮生长因子受体2(ErbB2)高表达乳腺癌SK-BR-3和MDA-MB-453细胞迁移和侵袭的影响及其分子机制。方法用si LDHA转染(以转染scramble siRNA为阴性对照)SK-BR-3和MDA-MB-453细胞干扰LDHA的表达;Western blot法检测LDHA蛋白表达水平;Transwell小室检测细胞的迁移和侵袭能力;采用乳酸脱氢酶(LDH)试剂盒测定细胞的LDH活性;采用葡萄糖和乳酸检测试剂盒分别测定培养基中的葡萄糖浓度和乳酸浓度,计算细胞对葡萄糖的摄取量和乳酸生成量。结果与阴性对照组比较,si LDHA下调SK-BR-3和MDA-MB-453细胞LDHA的蛋白水平(P<0.01);降低细胞的迁移和侵袭能力(P<0.001);下调SK-BR-3细胞的LDH活性,减少两种细胞葡萄糖摄取量和乳酸生成量,差异均有统计学意义(P<0.05)。结论干扰LDHA表达通过降低糖酵解从而抑制ErbB2高表达乳腺癌细胞的迁移和侵袭。
Objective To investigate the effect of small interfering RNA of lactate dehydrogenase A(si LDHA)on migration and invasion of epidermal growth factor receptor 2(ErbB2)over expressing breast cancer cell line SK-BR-3,MDA-MB-453 and its molecular mechanism.Methods SK-BR-3and MDA-MB-453 cells were transfected with si LDHAto interfere with the expression of LDHA.The transfection of scramble siRNA was used as negative control.The LDHA protein levels were detected by Western blot(P<0.01).Cell migration and invasion was detected by Transwell assays.Lactate dehydrogenase(LDH)activity was measured by LDH assay kit.The glucose and lactate concentration in the culture media was determined by glucose and lactate assay kit,respectively,and then glucose uptake and lactate production by the cells were calculated.Results si LDHA downregulated LDHA protein levels in SK-BR-3and MDA-MB-453cells(P<0.01).Compared with negative control group,si LDHAsignificantly decreased migration and invasion of SK-BR-3and MDA-MB-453cells(P<0.001).si LDHAreduced LDH activity in SK-BR-3cells,glucose uptake and lactate production in SK-BR-3and MDA-MB-453 cells,the difference was significant(P<0.05).Conclusion Knockdown of LDHA by siRNA inhibits the migration and invasion via downregulation of glycolysis in ErbB2 over expressing breast cancer cell line.
引文
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