Effect Of Lactic Acid On Energy Metabolism Of U251 Glioma Cell

As tumor cells proliferate rapidly,tumors need more glucose uptake to provide energy compared with normal cells.However,duo to the less and abnormal distribution of blood vessels,the supply of glucose and oxygen is deficient in tumor,especially in tumor core area.And the tumor cells strengthen glycolysis and generate lots of lactic acid.Particularly,glinoma in brain which is isolated by blood-brain barrier is caught in lower glucose and worsen higher lactic acid condition.In such an adversity condition,the way tumor cells eliminate or utilize lactic acid is play great role in tumor survival and development.Presently,there have been reports about the utilization of lactic acid in tumor.However,the metabolic way of lactic acid in glioma is not clear.In this study,U251,glioma cell line,was treated with different concentration of lactic acid.And we found that 15mM lactic acid treatment was suitable concentration for U251 according to cell growth and ATP content status.In order to further explore on the metabolism way changes caused by lactic acid,U251 was cultured in normal culture medium with 15mM lactic acid for 24h,48h and 72h respectively.The results showed that when treated with lactic acid,more proliferous and higher ATP level were detected.While hydrochloric acid treated cell significantly decreased the cell growth and ATP content along with culture time.Those results indicated lactic acid can promote cell proliferation and ATP content independent on pH.Besides,the protein expression of lactate dehydrogenase(LDH),pyruvate dehydrogenase(PDH)and monocarboxylic acid transporter(MCT1,MCT4)were increased and glucose transporter GLUT1 was decreased.Metabolism regulated protein,hypoxia-inducible factor(HIF1-?)was down-regulated and proto-oncogene(C-myc)was up-regulated.Oxidative phosphorylation related proteins(NDUFB8,VDAC)increased in different degrees.Lactic acid could be transported and then utilized by the enhanced oxidative phosphorylation.In fact,glinoma is surrounded by the microenvironment including high lactic acid and low glucose.Furthermore,we analyzed the cell growth and ATP levels under low glucose medium with lactic acid(5mM G + 15mM Lac).We found that low glucose medium can only maintain cell survival for 48h,then a large number of cells were dead and ATP content was sharply declined.However,when lactic acid was added,U251 was able to continue growing and ATP content was kept in equal level to normal medium cultured cells.In low glucose condition,glucose could be preferentially used and then lactic acid could be role of fuel to keep cell growth and survival.Additionally,the protein expression of LDH,PDH,MCT1 and MCT4 were significantly increased.Otherwise,GLUT1 was decreased.The oxidative phosphorylation related proteins(NDUFB8,ND1,TFAM)were increased.Metabolism-related regulatory factor HIF-1? was decreased,C-myc was increased.Those results revealed that cells could enhance the oxidative phosphorylation pathway to produce sufficient ATP for cell survival in low glucose with high lactic acid.In order to verify the results of the above in vivo,we collected clinical glioma tissue and tested metabolic-related protein expression in different tumor area.The protein expression levels of LDH,PDH,MCT4 and HIF-la in the core region were significantly increased,while MCT1 and GLUT1 was decreased,which indicated that the lactic acid was mainly produced in core area through glycolysis.Then,through MCT4/MCT1 transporter,lactic acid could enter into adjacent tumor area and be utilized by oxidative phosphorylation as fuel.In this part,we found that different area of tumor tissue have different metabolic patterns according to surrounding microenvironment.And tumor cells can change metabolic way to adapt to glucose deficiency,lactic acid condition for survival and development.This study provides a new idea for further understanding the metabolic features of brain tumors.It may promote the development of new therapeutic strategies.