Regulation Effect Of Acupuncture And Moxibustion Of Treg Cells In Mice With Ulcerative Colitis Based On CD39/CD73/A2a Adenosine Metabolic Pathway

Objective:To investigate whether electro-acupuncture and moxibustion have influenced on Treg cells through CD39/CD73/A2a adenosine metabolic pathway under the mice model Ulcerative Colitis(UC).Methods:Dextran Sulfate Sodium(DSS)induced UC mice were set up and electro-acupuncture and moxibustion were used to treat the mice.The disease activity index(DAI),the length of the colitis and the pathological structure of each group were evaluated.Immunofluorescence(IF)and Western Blot(WB)were used to detect the protein expression of CD39,CD73 and A2a in the colon.The number of CD4~+CD39~+T cells,CD4~+CD39~+Foxp3~+T cells,CD4~+CD73~+T cells and CD4~+CD73~+Foxp3~+T cells in peripheral blood,spleen,and local draining lymph nodes were measured by flow cytometry(FCM).Results:1.Therapeutic effect of electro-acupuncture and moxibustion on UC model mice:1.1 After mice drinking 2%DSS,the general status gradually deteriorated,bloody stools or pus appeared,and the DAI score was also increased.Both electro-acupuncture and moxibustion can improve the general conditions and the DAI scores of the mice.1.2 Compared with the control group,shorter colons,ulcers and inflammatory cells infiltration were seen in the model group.However,the length of the colon recovered and histopathology improved by the electro-acupuncture and moxibustion treatment.2.The effect of electro-acupuncture and moxibustion on adenosine metabolic pathway in UC mice2.1 Compared with the control group,the fluorescence intensity of CD39,CD73,and A2a,the number of positive cells as well as their protein expression in the colons were decreased in the model group.After treatment,the above protein were significantly improved.In addition,compared with the electro-acupuncture group,the fluorescence intensity of CD73 and A2a,the number of positive cells,and the expression of CD39 and CD73 protein in the moxibustion group were higher than that in the electro-acupuncture group(P<0.01).2.2 The expression of CD39 and CD73 in peripheral blood,local draining lymph nodes and spleen Treg cells:2.2.1 Compared with the control group,the numbers of CD4~+CD39~+T cells,CD4~+Foxp3~+CD39~+T cells,CD4~+CD73~+T cells,and CD4~+Foxp3~+CD73~+T cells in the peripheral blood,lymph nodes and spleen of the model group all decreased(P<0.01).The ratio of CD4~+Foxp3~+CD39~+T cells to CD4~+CD39~+T cells and the ratio of CD4~+Foxp3~+CD73~+T cells to CD4~+CD73~+T cells in the peripheral blood,lymph nodes,and spleen of the model group were all decreased(P<0.01).2.2.2 After intervention with electro-acupuncture and moxibustion,the number of CD4~+CD39~+T,CD4~+Foxp3~+CD39~+T,CD4~+CD73~+T,and CD4~+Foxp3~+CD73~+T cells were enhanced in the peripheral blood,lymph nodes,and spleen of moxibustion and electro-acupuncture groups(P<0.01).The ratio of CD4~+Foxp3~+CD39~+T cells to CD4~+CD39~+T cells and CD4~+Foxp3~+CD73~+T cells to CD4~+CD73~+T cells in peripheral blood,lymph nodes,and spleen of moxibustion group and electro-acupuncture group also showed a significant increase(except the ratio of CD39~+T cells in the moxibustion group of lymph nodes was P>0.05,the others P<0.01).2.2.3 Compared with the electro-acupuncture group,the number of CD4~+Foxp3~+CD73~+T cells in the spleen of the moxibustion group were increased(P<0.01),however,CD4~+Foxp3~+CD39~+T cells in the peripheral blood of the moxibustion group,and CD4~+CD73~+T cells in spleen and in the lymph nodes were decreased(P<0.01).The ratio of CD4~+Foxp3~+CD39~+T cells to CD4~+CD39~+T cells in peripheral blood and lymph nodes of the moxibustion group was decreased(P<0.01,P<0.05).The ratio of CD4~+Foxp3~+CD73~+T cells to CD4~+CD73~+T cells in the spleen of the moxibustion group increased(P<0.01).Conclusion:1.Both moxibustion and electro-acupuncture can alleviate symptoms,including blo ody stool and weight loss in DSS-induced UC mice.2.Both treatments can increase the expression of CD39,CD73 and A2a protein in colonic tissues.Meanwhile,the ratio of CD4~+Foxp3~+CD39~+T cells and the ratio of CD4~+Foxp3~+CD73~+T cells in peripheral blood,lymph nodes and spleen were all increased by electro-acupuncture and moxibustion.3.Additionally,the therapeutic effects of moxibustion and electroacupuncture on UC mice may be related to the intervention of CD39/CD73/A2a adenosine metabolic pathway and influence of the anti-inflammatory immune effects of Treg cells.4.The effection of moxibustion or electroacupuncture on CD39/CD73/A2a adenosine metabolic pathways in UC mice were strong and yet to be further studied.