艾灸治疗慢性炎性内脏痛的机制研究:脊髓p38 MAPK/ELK1信号通路的调节  

Mechanism of moxibustion in treating chronic inflammatory visceral pain:regulation of the p38 MAPK/ELK1 signaling pathway in the spinal cord

作　　者：张丹 李志元 于华鹏 吴焕淦 吴丽洁 杨韵 杨光 谢晨 洪珏 杨延婷 马晓芃 ZHANG Dan;LI Zhiyuan;YU Huapeng;WU Huangan;WU Lijie;YANG Yun;YANG Guang;XIE Chen;HONG Jue;YANG Yanting;MA Xiaopeng(Shanghai Research Institute of Acupuncture and Meridian,Shanghai 200030,China;Shenzhen Hospital,Beijing University of Chinese Medicine,Shenzhen 518100,China;Eye&ENT Hospital of Fudan University,Shanghai 200031,China;The First Affiliated Hospital of Zhejiang Chinese Medical University,Hangzhou 310006,China;Yueyang Hospital of Integrated Traditional Chinese and Western Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200437,China)

机构地区：[1]Shanghai Research Institute of Acupuncture and Meridian,Shanghai 200030,China [2]Shenzhen Hospital,Beijing University of Chinese Medicine,Shenzhen 518100,China [3]Eye&ENT Hospital of Fudan University,Shanghai 200031,China [4]The First Affiliated Hospital of Zhejiang Chinese Medical University,Hangzhou 310006,China [5]Yueyang Hospital of Integrated Traditional Chinese and Western Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200437,China

出　　处：《Journal of Acupuncture and Tuina Science》2024年第4期263-272,共10页针灸推拿医学（英文版）

基　　金：国家自然科学基金项目,No.82104985,No.81273843,No.81674073,No.81202754;上海市自然科学基金项目,No.23ZR1460100,No.23ZR1460000;上海市领军人才项目,No.060.

摘　　要：目的:从调节脊髓p38丝裂原活化蛋白激酶(MAPK)/Ets样转录因子1(ELK1)信号通路角度,探讨艾灸治疗慢性炎性内脏痛(CIVP)的中枢机制及其镇痛效应。方法:清洁级雄性Sprague-Dawley大鼠随机分为正常组、模型组、隔药灸组、假隔药灸组、p38 MAPK抑制剂组和二甲基亚砜(DMSO)组。采用2,4,6-三硝基苯磺酸合50%乙醇灌肠制备CIVP大鼠模型。隔药灸组给予隔附子饼灸治疗;假隔药灸组处理同隔药灸组,但不点燃艾炷;p38 MAPK抑制剂组在大鼠L5-L6间鞘内注射SB203580;DMSO组在L5-L6间鞘内注射2%DMSO。通过腹壁撤回反射(AWR)评分、机械刺激缩足阈值(MWT)及热缩足潜伏期(TWL)观察各组大鼠痛行为变化;应用苏木素-伊红染色镜下观察各组大鼠结肠组织形态学变化;采用免疫印迹法和实时荧光定量聚合酶链反应检测各组大鼠脊髓凋亡信号调节激酶1(ASK1)、MAPK激酶(MKK)3/6、p38 MAPK、ELK1和丝裂原应激活化蛋白激酶1(MSK1)磷酸化蛋白和mRNA的表达。结果:与正常组比较,CIVP大鼠结肠炎性损伤严重,病理损伤评分明显升高;不同压力结直肠扩张(CRD)刺激下AWR评分均增加,MWT及TWL均降低;脊髓p38 MAPK、ELK1、MSK1、ASK1、MKK3/6磷酸化蛋白和mRNA表达显著增加(P<0.01)。经隔药灸治疗后,大鼠结肠损伤有所修复,病理损伤评分降低;不同压力CRD刺激下,AWR评分均减少,MWT、TWL升高;脊髓p38 MAPK、ELK1、ASK1、MKK3磷酸化蛋白和mRNA表达降低,与模型组、假隔药灸组比较,差异均有统计学意义(P<0.01)。隔药灸组与p38 MAPK抑制剂组相比、DMSO组与模型组相比,上述指标均无显著差异(P>0.05)。结论:隔药灸能通过下调CIVP大鼠脊髓ASK1、MKK3、p38 MAPK、ELK1磷酸化蛋白和mRNA的表达发挥镇痛效应。抑制脊髓p38 MAPK/ELK1信号通路活化可能是隔药灸缓解疼痛,治疗CIVP的机制之一。Objective:To investigate the central mechanism of moxibustion in treating chronic inflammatory visceral pain(CIVP)and its analgesic effect from the perspective of the p38 mitogen-activated protein kinase(MAPK)/Ets-like transcription factor 1(ELK1)signaling pathway in the spinal cord.Methods:Clean-grade male Sprague-Dawley rats were randomly divided into a normal group,a model group,a herb-partitioned moxibustion(HPM)group,a sham-HPM group,a p38 MAPK inhibitor group,and a dimethyl sulfoxide(DMSO)group.CIVP rat models were prepared using an enema mixture of 2,4,6-trinitrobenzene sulfonic acid solution and 50%ethanol.The HPM group was treated with HPM;the sham-HPM group was treated the same as the HPM group,but the moxa cones were not ignited;rats in the p38 MAPK inhibitor group received L5-L6 intrathecal injection of p38 MAPK inhibitor(SB203580);rats in the DMSO group received L5-L6 intrathecal injection of 2%DMSO.Abdominal withdrawal reflex(AWR),mechanical withdrawal threshold(MWT),and thermal withdrawal latency(TWL)were used to observe pain-related behaviors in each group.Hematoxylin-eosin staining was used to observe the morphological changes in rat colon tissue.Western blotting and real-time quantitative reverse-transcription polymerase chain reaction were used to detect the phosphorylated protein and mRNA expression of apoptosis signal-regulating kinase 1(ASK1),MAPK kinase(MKK)3/6,p38 MAPK,ELK1,and mitogen and stress-activated protein kinase 1(MSK1)in the spinal cord.Results:Compared with the normal group,CIVP rats had severe colonic inflammatory injuries,and the pathological injury scores increased significantly,along with increased AWR scores under different colorectal distension(CRD)stimulation pressures and decreased MWT and TWL;the mRNA and phosphorylated protein expression of p38 MAPK,ELK1,MSK1,ASK1,MKK3,and MKK6 all increased in the spinal cord(P<0.01).After HPM treatment,the colon injuries were repaired,and the pathological injury scores decreased;under different CRD stimulation pressures,the AWR score

关 键 词：灸法 药饼灸疗法 炎症性肠病 内脏痛 内脏高敏感 P38丝裂原活化蛋白激酶信号通路 大鼠 

分 类 号：R2-03[医药卫生—中医学]