Study On The Mechanism Of Proliferation And Energy Metabolism Of Renal Carcinoma Vascular Endothelial Cells And Lactic Acid And Mono Carboxylic Transporter

Objectives: As a common malignant tumor of the urinary system,renal cell carcinoma(RCC)is not sensitive to conventional radiotherapy and chemotherapy,and there is no effective treatment in the late stage.In recent years,targeted therapy to inhibit tumor angiogenesis has become a hot topic in the treatment of renal cancer.Based on the 3D co-culture model,this study aimed to explore the effects of endothelial cells on the biological behavior of human renal cell carcinoma cells,and the mechanism of lactic acid and mono carboxylic acid transporters in endothelial cell proliferation and energy metabolism,thus providing a new way of thinking for the targeting therapy of RCC.Methods: Transwell method was adopted to achieve the co-culture of renal carcinoma cells(786-O)and vascular endothelial cells(HUVEC).The proliferation ability of two cells was detected by Cell Counting Kit-8(CCK-8),the migration ability of the cells was detected by scratch test,and detection of cell invasion and metastasis by Transwell.To explore the effect of co-culture on cell biological behavior.Real-time PCR and Western blot were used to detect the expression of mono carboxylate transporter 1(MCT1)and MCT4 in two groups of cells in culture room.The lactate concentration test kit was used to detect the change of lactic acid concentration in each medium,so as to analyze the changes of lactate metabolism in two kinds of cells under co-culture.After targeted inhibition of MCT1 and MCT4,cell proliferation ability and lactate concentration in the two groups were detected.After targeted inhibition of MCT1 and MCT4,the proliferation rates of the two groups were significantly downregulated.The concentration of lactic acid in the cell culture medium of the inhibitor AZD3965 was significantly lower than that in the control group,the concentration of lactic acid in the cell culture medium with the inhibitor 7ACC1 decreased obviously.Results: CCK-8 assay showed that the proliferation rate of 786-O and HUVEC in the co-culture group was significantly higher than that in the control(the individual culture group).In the cell scratch test,the closure rate of 786-O and HUVEC cells in the co-culture group was significantly higher than control.In the Transwell assay,the number of 786-Ocells through the matrigel in the co-culture group was significantly higher than the control group.Real-time PCR results showed that the expression of MCT1 and MCT4 in the two cells in the co-culture group was up to up compared with the control group.The Western blot experiment also confirmed that the expression of MCT1 and MCT4 in 786-O and HUVEC cell lines was obviously up-regulated before and after co-culture.The lactic acid concentration test kit showed that the concentration of lactic acid in the co-culture group was significantly higher than control group.Conclusions: In co-culture,HUVEC can significantly enhance the proliferation,migration and invasion of 786-O,and 786-O can also promote the proliferation and migration of HUVEC.The expressions of MCT1 and MCT4 in HUVEC and 786-O were all up-regulated,and the concentration of lactic acid in the medium increased,indicating that HUVEC increased energy supply by glycolysis,and 786-O enhanced by lactic acid energy supply.This study provides a theoretical and practical basis for the exploration of a new and long time target therapy for inhibiting the formation of renal cell carcinoma,and provides a basis for more effective treatment or coordination in the treatment of renal cell carcinoma in the later period.