针刺研究2024，Vol.49Issue(2)：135-144,10.DOI:10.13702/j.1000-0607.20221062
电头针调控RORγt促进IL-17A+Th17/FOXP3+Treg细胞平衡缓解MCAO大鼠脑缺血炎性损伤的研究
Electroacupuncture of scalp acupoint alleviates cerebral ischemic inflammatory injury by down-regulating RORγt and promoting balance of IL-17A+Th17/FOXP3+Treg in MCAO rats
摘要
Abstract
Objective To observe the effect of electroacupuncture(EA)of scalp acupoint(Dingnieqian-xiexian,MS6)on expression of retinoid-related orphan receptor γT(ROR γ t),interleukin(IL)-17A,IL-10,transfor-ming growth factor-β1(TGF-β1),IL-6,IL-21,and IL-17A+ T helper cells(Th)17 and forkhead transcription factor P3(FOXP3)+ regulatory T cells(Treg)differentiation of ischemic cortex in ischemic stroke rats,so as to explore its mo-lecular mechanisms underlying relief of inflammatory injury of ischemic stroke.Methods A total of 120 male SD rats were randomly assigned to sham operation,model,EA,inhibitor,agonist and EA+agonist groups,with 15 rats in each group.The ischemic stroke model was established by occlusion of the left middle cerebral artery according to Longa's methods.For rats of the EA group and EA+agonist group,EA(2 Hz/100 Hz,1 mA)was applied to bilateral MS6 for 30 min,once daily for 7 days.Rats of the inhibitor group received intraperitoneal injection of solution of SR1001(RORγt inhibitor)(2.5 mg/mL,10 mg/kg),once daily for 7 days.Rats of the agonist and EA+agonist groups received in-traperitoneal injection of solution of SR1078(RORγt agonist)(5 mg/mL,5 mg/kg)before EA,once daily for 7 days.Rats of the sham operation and model groups were grabbed and fixed in the same way with the other groups.The Zea-longa's score,modified neurological severity score(mNSS)and the neurobehavioral score were assessed before and after the intervention.At the end of experiments,the ischemic cortex tissue was collected.The 2,3,5-Triphenyltetrazo-lium chloride(TTC)staining was used to detect the volume of cerebral infarction.The expression of RORγt mRNA was detected by real-time quantitative PCR;the protein expression levels of RORγt,IL-17A,IL-10 and TGF-β1 were de-tected by Western blot;the immunoactivity of IL-6 and IL-21 were detected by immunohistochemistry;the fluorescence areas of IL-17A+Th17 and FOXP3+Treg cells were measured by immunofluorescence and their ratio was calculated in the tissue of ischemic cortex.Results Relevant to the sham operation group,the model group had a significant in-crease in the Zea-Longa's score,mNSS score,neurobehavioral score,cerebral infarct volume,expression levels of RORγt mRNA and protein,IL-17A protein,IL-6 and IL-21 immunoactivity,IL-17A+Th17 immunofluorescence intensity,and the ratio of IL-17A+Th17/FOXP3+Treg(P<0.01),and an obvious decrease in the expression levels of TGF-β1 and IL-10 proteins and FOXP3+Treg immunofluorescence intensity(P<0.01).In contrast to the model group,both EA and inhibitor groups had a significant decrease in the Zea-Longa's score,mNSS score,neurobehavioral score,cerebral infarct volume,expression levels of RORγt mRNA and protein,IL-17A protein,IL-6 and IL-21 immunoactivity,IL-17A+Th17 immunofluorescence intensity,and the ratio of IL-17A+Th17/FOXP3+Treg(P<0.01,P<0.05),and a marked increase in the expression levels of TGF-β1 and IL-10 proteins and FOXP3+Treg immunofluorescence intensity(P<0.05,P<0.01),while the above indicators of the agonist group were all reversed(P<0.01,P<0.05).Comparison between the agonist and EA+agonist groups showed that the Zea-Longa's score,mNSS score,neurobehavioral score,cerebral infarct volume,expression levels of RORγt mRNA and protein,IL-17A protein,IL-6 and IL-21 immunoactivity,IL-17A+Th17 immunofluorescence intensity,and the ratio of IL-17A+Th17/FOXP3+Treg were significantly lower(P<0.01,P<0.05),and the expression of TGF-β1 and IL-10 proteins and FOXP3+Treg immunofluorescence intensity were obviously higher(P<0.01,P<0.05)in the EA+agonist group than in the agonist group,suggesting that EA intervention can effectively weaken the effects of RORγt agonist.Conclusion EA of scalp acupoint MS6 can effectively improve the neurological function,behavior reaction and reduce cerebral infarct volume in ischemic stroke rats,which may be associated with its functions in down-regulating the expression of RORγt and promoting the balance of IL-17A+Th17/FOXP3+Treg to alleviate inflammatory injury after ischemic stroke.关键词
急性缺血性脑卒中/电头针/维甲酸相关孤儿受体γt/辅助性T细胞/调节性T细胞Key words
Acute ischemic stroke/Electroacupuncture of scalp acupoint/Retinoid-related orphan receptor γt/T helper cells/Regulatory T cells引用本文复制引用
袁博,王金海,方晓苓,杨尚伟,田甜,朱玲桂,李颖,杜小正,彭晓云,姚小强..电头针调控RORγt促进IL-17A+Th17/FOXP3+Treg细胞平衡缓解MCAO大鼠脑缺血炎性损伤的研究[J].针刺研究,2024,49(2):135-144,10.基金项目
国家自然科学基金项目(No.81960896) （No.81960896）
甘肃省科技计划项目(No.18JR2FA002) （No.18JR2FA002）
甘肃中医药大学附属医院院内科研及技术创新基金青年项目(No.gzfy-2019-13) （No.gzfy-2019-13）
