MiR-30a-LDHA-HIF-1α Axis Regulates The Proliferation Of Tumor Cells In Lung Adenocarcinoma

Objective: The effect of miR-30a-LDHA-HIF-1α signaling axis on the growth of tumor cells was investigated in lung adenocarcinoma,and the mechanism that miR-30a-LDHA-HIF-1α signaling axis affecting the tumor cell stemness was explored.The investigation suggested that miR-30 a might be a biomarker for diagnosis and a potential target for the treatment of lung cancer.Methods:miR-30a overexpression was performed by transfection of miR-30a mimics,and knockdowns of both LDHA and HIF-1αby transfection of siRNA.miR-30a,LDHA,and HIF-1αwere detected at RNA level by qRT-PCR.The proteins of LDHA,Oct4 and Sox2 were examined by Western blot.The expressions of LDHA and HIF-1αin lung adenocarcinoma were analyzed by TCGA database.The correlation analysis between LDHA and HIF-1αwas performed using TCGA data,and the interaction was analyzed by software GeneMANIA.To investigate the effect of miR-30a on glycolysis,lactic acid production was detected using lactic acid analyzing kit.To explore the effect of miR-30a on the proliferation,CCK-8 method and cell colony formation assay were employed.With overexpression of miR-30a,knockdowns of LDHA and HIF-1α,the apoptosis was analyzed by flow cytometry.Protein chip was hired to analyze the stemness of tumor cells affected by miR-30a-LDHA-HIF-1αaxis,and the effect of miR-30a on transcriptome in A549 cells was explored by high-throughput sequencing.Results: LDHA was a direct target of miR-30 a,and miR-30 a inhibited LDHA expression at both RNA and protein levels in A549 cell line.Using TCGA database,we found that LDHA expression in normal lung tissues was lower than that in lung adenocarcinoma tissues,and so did HIF-1α.Meanwhile,the LDHA expression was significantly correlated with HIF-1α.miR-30 a overexpression suppressed lactic acid production,which was according with the knockdowns of LDHA and HIF-1α.miR-30 a overexpression,knockdowns of LDHA and HIF-1α were able to inhibit tumor cell proliferation and induce the apoptosis of tumor cells in vitro.The effect of miR-30 a on the stemness of tumor cell was examined,and the expressions of both Oct4 and Sox2 were suppressed by miR-30 a overexpression by comparison of the Mock and the Negative control,which was according with the knockdowns of LDHA and HIF-1α.Using transcriptome sequencing,high expression of miR-30 a significantly down-regulated the expressions of ACOT11,KDELC2,CT62,F10,MADCAM1,FGF12 and BTBD8 and up-regulated the levels of EGR4,CYP3A7,ARG1,CFAP73,MYLK2,TVP23 A.Conclusion:The down-regulation of miR-30a promotes LDHA expression and lactic acid production,then induces HIF-1αexpression,resulting in the high expression of Oct4 and Sox2,which are the markers of tumor stem cells.The decreased miR-30a was able to promote tumor cell proliferation and suppress the apoptosis of tumor cells by regulating LDHA-HIF-1αaxis,leading to the development of lung adenocarcinoma.