乳酸脱氢酶A在肾细胞癌中的表达及意义

Expression and Significance of Lactate Dehydrogenase A in Renal Cell Carcinoma

【目的】分析乳酸脱氢酶A(LDHA)在肾细胞癌组织及细胞系中的表达情况并探讨其影响肾细胞癌细胞进展的方式。【方法】收集自2018年6月至2022年6月在我院通过手术方式获取的肾细胞癌组织标本52例及癌旁组织标本49例,免疫组织化学法比较LDHA的表达差异。通过qRT-PCR及Western blot实验检测LDHA在正常人近端肾小管上皮细胞系(HK-2)及肾细胞癌细胞系(A498,Caki-2,ACHN,786-O)中的表达水平。构建LDHAshRNA重组质粒,转染至786-O以敲低LDHA表达,利用CCK-8、克隆形成实验及EdU染色法检测敲低LDHA表达对癌细胞增殖活性的影响。利用细胞葡萄糖摄取试验及乳酸分泌试验检测细胞糖摄取水平和乳酸分泌水平的变化。利用糖酵解压力测试实验检测细胞糖酵解能力的变化。【结果】免疫组化结果可见LDHA在肾细胞癌组织中表达明显高于癌旁组织,并且临床TNM分期越高的肾癌组织,LDHA的表达水平越高(P<0.05)。qRT-PCR和Western blot结果可见,LDHA在各肾细胞癌细胞系中的表达均较HK-2有明显升高(P<0.05)。在786-O敲低LDHA表达后,细胞增殖活性及糖酵解能力均显著下调(P<0.05)。【结论】LDHA在肾细胞癌组织及细胞中的表达均明显升高,肿瘤分期越晚期,LDHA的表达越高。抑制LDHA表达能显著抑制肾细胞癌细胞786-O的增殖活性及有氧糖酵解活性。

【Objective】To analyze the expression of Lactate dehydrogenase A(LDHA)in both renal cell carcinoma(RCC)tissue and RCC cell lines,and to investigate the impact of LDHA expression on the progression of RCC.【Methods】From June 2018 to June 2022,totally 52 cases of RCC tissue samples and 49 cases of para-cancerous tissue samples were collected through surgical procedures from our hospital.LDHA expression was detected using immunohistochemistry(IHC).The expression levels of LDHA in vitro were also detected in the normal human proximal tubule epithelial cell line HK-2 and renal cell carcinoma cell lines A498,Caki-2,ACHN,and 786-O by using qRT-PCR and Western blot.A re⁃combinant plasmid carrying LDHA-shRNA was constructed and then transfected into 786-O cells to down-regulate the ex⁃pression of LDHA.Tumor proliferative capacity was monitored using CCK-8 assay,clonal formation assay and EdU assess⁃ments.Additionally,cell glycolytic activity was assessed through glucose uptake assay,lactate secretion assay,and ECAR analysis.【Results】IHC analysis revealed significantly higher expression of LDHA in RCC tissue compared to adjacent tis⁃sues(P<0.05).Furthermore,RCC tissues with higher TNM stage exhibited greater expression of LDHA than those with low⁃er TNM stage(P<0.05).The results of qRT-PCR and Western blot demonstrated that the expression of LDHA in each RCC cell line was significantly higher than that in HK-2(P<0.05).After blocking the expression of LDHA in 786-O,there was a significant down-regulation of cell proliferation and glycolysis capacity(P<0.05).【Conclusions】The expression of LDHA in RCC tissue and RCC cell lines is significantly overexpressed compared with normal one,particularly in those with high⁃er TNM stage.Knockdown of the expression of LDHA significantly suppresses cell proliferation and aerobic glycolysis ca⁃pacity in 786-O.