Effects Of Overexpression And Knockdown Of Lactate Dehydrogenase C4 On The Biological Characteristics Of Breast Cancer Cells

Objective:1.To evaluate the role of lactate dehydrogenase C4(LDH-C4)in breast cancer tissues and its possible significance in use.2.To detect the change of biological function with LDHC gene alteration in breast cancer cell HCC1937.3. To understand the underlying molecular mechanism as modifying the LDHC gene expression.Methods:1.Immunohistochemical staining technology was applied to evaluate the LDH-C4 protein expression and to roughly indicate its role in survival analysis.2. The LDHC gene overexpression and low expression lentiviral vector were constructed to infect breast cancer cells HCC1937 and the puromycin was used to screen stable cell lines.The RT-q PCR was conducted to evaluate the LDHC gene alteration.Besides,the changes in cell biological function of breast cancer cell HCC1937 were studied through the assays including CCK-8 colorimetry,scratch repair,and chamber analysis.3.The subsequent events occured in glycolytic pathways including lactic acid,pyruvate,and glucose were detected by microplate method while the AKT / mTOR-related pathways were shown by Western blot.Results:1.LDH-C4 protein was expressed by all 145 breast cancer tissues included and the breast cancer harbored higher expression level than the paracancer tissues.The outcome results showed that the LDH-C4 protein was positively correlated with the pathological grade,lymph node metastasis and N stage of the patients(P<0.05).Moreover,the results of Kaplan-Meier survival analysis showed patients with higher LDH-C4 had poorer quality of life(P<0.05).It was also indicated by Cox model results that the LDH-C4 protein was an factor for breast cancer poor prognosis(P<0.05).2.The breast cancer cells with over-expressed and under-expressed LDHC gene were successfully obtained.The results of CCK-8colorimetry,scratch repair,and transwell invasion and migration suggested that higher LDH-C4 could accelerate the migration and invasion ability of HCC1937 cells,while the lower LDH-C4 can inhibit the ability.However,the change of LDH-C4 expression has no effects on the growth and proliferation of breast cancer cell HCC1937.3.Up-regulated LDHC gene expression increased the glucose substrate consumption and lactic acid production while reduced the pyruvate production.On the other hand,down-regulated LDHC gene expression reduced the glucose substrate consumption and lactic acid product production while increased the pyruvate production.At the same time,western blot results showed that over-expression of LDH-C4 in breast cancer cells HCC1937 contributed to the higher expression of AKT and mTOR protein,while low expression of LDH-C4 resulted in the decreased level of AKT and mTOR protein.Conclusions:1.LDH-C4 protein is largely expressed by the breast cancer tissue,which also gives rise to poor prognosis.2.Higher LDH-C4 can accelerate the migration and invasion ability of HCC1937 cells while lower LDH-C4 can inhibit the ability.3.LDH-C4 maybe involved in breast cancer genesis through regulating the glycolysis pathway and AKT / mTOR signaling pathway.