Research On The Effect Of Lactic Acid On Human CD4~+T Cell Function

Objective:Research on Effects and Mechanism of CD4~+T cells of lactic acid.Methods:Human peripheral blood mononuclear cells(PBMCs)were isolated by density gradient centrifugation from human peripheral venous anticoagulant blood,and CD4~+T cells were further sorted from PBMC for the following experiments.According to the different components of lactic acid in the solution,five groups were established:unstimulated group,stimulated control group,lactic acid group,sodium lactate group and hydrochloric acid group.Flow cytometry was used to detect the production of cytokines(IL-2 and IFN-γ)by CD4~+T cells with stimulation of PMA/ionomycin.ELISA was used to detect IFN-γlevels in the supernatant of cell culture following TCR(anti-human CD3/CD28)stimulation.The production of IL-2 and IFN-γat the m RNA level was detected by q PCR.The expression of surface antigens CD25 and CD69 was detected by flow cytometry.MCT1 expression at m RNA level and protein level was detected by RT-PCR and Western Blot,respectively.First,MCT1 inhibitor was used to examine the effect of MCT1 on CD4~+T cell effector function.Furthermore,MCT1 in CD4~+T cells was silenced by lentivirus-mediated sh RNA interference,and then MCT1-knockdown CD4~+T cells were sorted and the knockdown effect was determined by Western Blot.Thse cells were used to evaluate the inhibitory effects of lactic acid on CD4~+T cell function.Results:(1)Lactic acid inhibits the production of IL-2,IL-4 and IFN-γand activation of CD4~+T cells;(2)MCT1 is not required for lactic acid-mediated inhibition of IL-2 and IFN-γproduction;(3)MCT1 is required for the suppression of CD25 and CD69 expression by lactic acid.Conclusion:The inhibitory effects of lactic acid on IL-2 and IFN-γexpression in CD4~+T cells are independent of MCT1.