隔药饼灸对免疫抑制兔Tim-4及PD-1的泛素化的影响  

Effects of herbal cake separated moxibustion on Tim-4 and ubiquitination of PD-1 in rabbits with immunosuppression

作　　者：熊罗节 田岳凤 徐小珊 翟春涛 李玮 XIONG Luo-jie;TIAN Yue-feng;XU Xiao-shan;ZHAI Chun-tao;LI Wei(College of Acupuncture-Moxibustion,Tuina and Rehabilitation,Hunan University of CM,Changsha 410208,China;Second Clinical College,Shanxi University of CM,Jinzhong030619)

机构地区：[1]湖南中医药大学针灸推拿与康复学院,长沙410208 [2]山西中医药大学第二临床学院,晋中030619

出　　处：《中国针灸》2023年第5期529-536,共8页Chinese Acupuncture & Moxibustion

基　　金：国家自然科学基金资助项目:82174504、81674062;湖南中医药大学研究生创新课题项目:2022CX20。

摘　　要：目的:观察隔药饼灸对免疫抑制兔巨噬细胞效应分子T细胞免疫球蛋白黏蛋白分子-4(Tim-4)及细胞程序性死亡受体1(PD-1)泛素化的影响,探讨隔药饼灸改善免疫抑制的可能机制。方法:将32只大耳白兔随机分为空白组、模型组、艾条灸组及隔药饼灸组,每组8只。除空白组外,其余3组均以60 mg/kg剂量腹腔注射环磷酰胺制备免疫抑制模型。艾条灸组予艾条灸,每穴灸1壮;隔药饼灸组予隔药饼灸,每穴灸5壮。均选取“神阙”“肾俞”“足三里”等穴,隔日1次,共干预10次。采用血细胞分析仪检测外周血白细胞含量;流式细胞术检测外周血PD-1在CD_(4)^(+)、CD_(8)^(+)T淋巴细胞及CD_(68)^(+)巨噬细胞中的阳性表达;ELISA法检测血清白细胞介素2(IL-2)、CD8、CD68及Tim-4含量;免疫组化法检测肝脏、脾脏Tim-4及F-box蛋白38(FBXO38)表达。结果:与空白组比较,模型组外周血白细胞计数(WBC)及中性粒细胞百分比(NEU%)降低、淋巴细胞百分比(LYM%)升高(P<0.01);外周血PD-1在CD_(4)^(+)、CD_(8)^(+)T淋巴细胞及CD_(68)^(+)巨噬细胞中的阳性表达率升高(P<0.01);血清IL-2、CD68及Tim-4含量升高(P<0.01),CD8含量降低(P<0.01);肝脏Tim-4及肝脏、脾脏FBXO38平均吸光度值增高(P<0.01)。与模型组比较,艾条灸组与隔药饼灸组WBC及NEU%升高(P<0.01);外周血PD-1在CD_(4)^(+)、CD_(8)^(+)T淋巴细胞及CD_(68)^(+)巨噬细胞中的阳性表达率降低(P<0.01);血清IL-2、CD68与Tim-4含量降低(P<0.01),CD8含量升高(P<0.01);肝脏Tim-4、FBXO38及脾脏FBXO38平均吸光度值降低(P<0.01,P<0.05)。与艾条灸组比较,隔药饼灸组外周血PD-1在CD_(68)^(+)巨噬细胞中的阳性表达率升高(P<0.05);血清Tim-4含量增高(P<0.01);肝脏Tim-4平均吸光度值降低(P<0.05)。结论:隔药饼灸可通过调节巨噬细胞效应分子Tim-4表达及FBXO38介导的PD-1泛素化改善免疫抑制状态,Tim-4可能是其参与免疫调控的特异性指标之一。Objective To observe the effects of herbal cake separated moxibustion on macrophage effector molecule T-cell immunoglobulin and mucin-domain containing-4(Tim-4)and ubiquitination of programmed cell death protein 1(PD-1)in rabbits with immunosuppression,and to explore the possible mechanism on herbal cake separated moxibustion in improving immunosuppression.Methods Thirty-two big-ear white rabbits were randomly divided into a normal group,a model group,a moxa stick moxibustion group and a herbal cake separated moxibustion group,8 rabbits in each group.Except the normal group,the immunosuppression model was established by intraperitoneal injection of cyclophosphamide of 60 mg/kg in the other 3 groups."Shenque"(CV 8),"Shenshu"(BL 23),"Zusanli"(ST 36),etc.were selected in both the moxa stick moxibustion group and the herbal cake separated moxibustion group.Moxa stick moxibustion was applied in the moxa stick moxibustion group,one cone at each acupoint;herbal cake separated moxibustion was applied in the herbal cake separated moxibustion group,5 cones at each acupoint.The intervention was given once every other day for 10 times in both groups.Leukocyte content in peripheral blood was detected by blood cell analyzer;the positive expression of PD-1 in CD_(4)^(+)T lymphocytes,CD_(8)^(+)T lymphocytes and CD_(68)^(+)macrophages in peripheral blood was measured by flow cytometry,the serum levels of interleukin 2(IL-2),CD8,CD68 and Tim-4 were detected by ELISA,and the expression of Tim-4 and F-box only protein 38(FBXO38)in the liver and spleen tissues was measured by immunohistochemistry.Results Compared with the normal group,in the model group,white blood cell count(WBC)and percentage of neutrophils(NEU%)were decreased while percentage of lymphocyte(LYM%)was increased(P<0.01)in peripheral blood;the positive expression rates of PD-1 in CD_(4)^(+)T lymphocytes,CD_(8)^(+)T lymphocytes and CD_(68)^(+)macrophages in peripheral blood were increased(P<0.01);the serum levels of IL-2,CD68 and Tim-4 were increased(P<0.01),the serum l

关 键 词：免疫抑制 隔药饼灸 艾条灸 巨噬细胞 T淋巴细胞 F-box蛋白38 T细胞免疫球蛋白黏蛋白分子-4 泛素化 

分 类 号：R245.8[医药卫生—针灸推拿学]