Mechanism study of Yunpi Jiedu Tongluo Qushi Decoction on central sensitization of collagen-induced arthritis mice via suppressing CD4+T cell migration by targeting IRE1α/XBP1/PDK1
Objective:To investigate the mechanism of Yunpi Jiedu Tongluo Qushi Decoction on central sensitization of collagen-induced arthritis(CIA)mice via suppressing CD4+T cell migration by targeting IRE1 α/XBP1/PDK1.Methods:Thirty-two male DBA/1 mice were randomly divided into 4 groups after adaptive feeding,including blank group,CIA group,traditional Chinese medicine(TCM)group and western medicine group,with 8 mice in each group.The CIA model was constructed by collagen injection.After the initial immunization,each drug group received the corresponding drug treatment for 42 consecutive days.Pain threshold and arthritis scores in each group were assessed weekly from day 28.After the intervention,peripheral blood and spinal cord tissues in each group were separated and collected.In addition,spleen CD4+T cells in blank group,CIA group and Chinese medicine group were collected by magnetic activated cell sorting,and the blank group was randomly divided into blank group and IRE1 α agonist group.The agonist group was treated with Thapsigargin 1μmol/L for 24h.The serum levels of interleukin(IL)-1β and IL-6 were detected by ELISA.The fluorescence intensity of CD4,cFos,IBA1 and IRE1 α/XBPl/PDK1 were detected by immunofluorescence.Lactic acid kit was used to detect the content of lactic acid in spinal dorsal horn.The fluorescence intensity of IRE1 α/XBP1/PDK1 pathway related proteins was detected by immunofluorescence,the lactic acid content of cells in each group was detected by lactic acid kit,and the migration ability of CD4+T cells in each group was observed by Transwell assay.Results:Compared with CIA group,the paw withdrawal threshold and paw withdrawal latency were significantly increased(P＜0.05),the arthritis score(P＜0.01)and serum IL-1 βand IL-6 levels in TCM group were significantly decreased(P＜0.05),the fluorescence intensity of CD4,cFos and IBA1 in spinal dorsal horn of TCM group was significantly decreased(P＜0.05),the fluorescence intensity of IRE1 α,XBP1 and PDK1 were significantly decreased(P＜0.05),the lactate concentration was decreased significantly(P＜0.05).In addition,in vitro experiments showed that compared with the blank group,the expression levels of IRE1 α,XBP1 and PDK1 in CD4+T cells were significantly increased(P＜0.01,P＜0.05),the lactate concentration was significantly increased(P＜0.01,P＜0.05),the migration ability of CD4T cell was significantly enhanced(P＜0.05)in the CIA group and IRE1 α agonist group.Compared with the CIA group,the expression levels of IRE1 α,XBP1 and PDK1 were significantly decreased(P＜0.01,P＜0.05),the cellular lactate concentration decreased significantly(P＜0.05),and the migration ability of CD4+T cell was significantly decreased in the TCM group(P＜0.05).Conclusion:Yunpi Jiedu Tongluo Qushi Decoction can inhibit the reprogramming of CD4+T cells to glycolysis and the central migration of CD4+T cells by targeting IRE1 α/XBP1/PDK1,thereby improving the central sensitization of CIA mice.
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