The Effect And Mechanism Of LDHA In The Lipiogenesis Of Glioma

Objective:The aim of the study is to investigate the effect and mechanism of lactate dehydrogenase A(LDHA)in the lipiogenesis of glioma cells,it provides a theoretical basis for further clinical treatment strategies targeting tumor metabolism.Methods: Firstly,bioinformatics was applied to analyze the expression pattern and clinical significance of LDHA in GBM,as well as the correlation between LDHA and indexes related to lipogenesis.Then,the effects of LDHA on lipogenesis,mitophagy pathway and AMPK signaling pathway in glioma cells were analyzed by Western blot.Besides,mitochondrial ROS and mitochondrial membrane potential in glioma cells were detected by specific fluorescence staining.Results: Clinical data showed that mRNA expression level of LDHA in glioma was higher than normal brain tissue;patients with high LDHA expression had a lower overall survival.Meanwhile,the results of human protein database analysis showed that the expression level of LDHA protein in malignant glioma was significantly higher than that in low-grade glioma and normal brain tissue;the correlation analysis results showed that LDHA was positively correlated with ACLY expression,but not significantly correlated with ACSS2 expression.The protein expressions of LDHA and ME2 in PN GBM cells SW1783 and U251 MG under normoxic and hypoxic conditions were at low levels,while the protein expressions of LDHA and ME2 in MES GBM cells U87 MG and LN229 were at high levels,and compared with normoxic conditions,the lipiogenesis indexes FASN,SCD1,ACLY and ACSS2 in SW1783 and U251 MG cells increased significantly under hypoxic conditions,while FASN,SCD1,ACLY and ACSS2 in U87 MG and LN229 cells showed no significant changes and were at a relatively high level.The expression of LDHA,ME2 and p-ACLY in U87 MG cells increased with the increase of hypoxic time.After interfering with LDHA in U87 MG cells,the protein expression level of ME2 decreased and the phosphorylation level of ACLY decreased,indicating that LDHA can promote the activation of ACLY,that is,LDHA can promote the synthesis of acetyl-CoA from mitochondrial citric acid pathway.After interfering with LDHA under normoxic conditions,the levels of ME2 and p-ACLY increased with the increase of the concentration of exogenous lactate,indicating that lactate,the final product catalyzed by LDHA during glycolysis,could activate ACLY and promote the synthesis of acetyl-CoA from mitochondrial citric acid pathway.Next,the sh-LDHA or sh-EGFP plasmids were transfected into the glioma cells U87 MG and LN229,respectively.Compared to sh-EGFP group,the production of mitochondrial ROS increased,the mitochondrial membrane potential was significantly reduced,and the expression of mitophagy-related proteins PINK1,Parkin,p-ubiquitin,p62,Optirineurin,BNIP3,BNIP3 L,and LC3 B increased.Also,AMPK signaling pathway-related proteins p-AMPK and p-TSC2 were significantly increased,while p-Raptor and p-mTOR were decreased under these processing conditions in the sh-LDHA group,indicating that LDHA can affect the mitophagy of MES GBM cells through the AMPK signaling pathway.Conclusions: The expression of LDHA in glioma was obviously high,which was negatively correlated with the survival of patients,and the expression of LDHA was positively correlated with the expression of ACLY.Under hypoxia,the lipogenesis in PN GBM cells was active,while LDHA in MES type GBM cells can promote the lipogenesis of mitochondrial citric acid pathway.In addition,we demonstrated that LDHA inhibits mitophagy in human glioma cells through the AMPK signaling pathway.In this study,the effects of LDHA on lipogenesis of mitochondrial citric acid pathway in MES GBM cells and its effect on mitophagy in MES GBM cells by affecting AMPK signaling pathway were investigated,providing a theoretical basis for further clinical treatment targeting tumor metabolism.