电针刺激大肠俞对炎症性肠病小鼠内脏痛的影响及机制研究

Electroacupuncture on Dachangshu can ameliorate visceral pain in IBD mice

目的:观察电针刺激炎症性肠病(IBD)模型小鼠双侧大肠俞治疗内脏痛的效果并初步探讨其机制。方法:利用直肠注射2,4,6三硝基苯磺酸(TNBS)诱导炎症性肠病模型。32只8周龄C57BL/6J小鼠随机分为对照组、造模组、电针组和假针刺组,根据分组给予各组小鼠相应治疗,一共治疗7d。造模24h后每天采用腹泻指数评分评估小鼠肠道炎症的严重程度,共7d。治疗第7天,行结直肠扩张实验记录腹壁撤退反射评分;行旷场实验分别评估小鼠内脏痛觉过敏程度和焦虑情绪严重程度。完成行为学试验后,利用免疫印迹法(WB)检测小鼠结肠组织和背根神经节(DRGs)中诱导性一氧化氮合酶(iNOS)的表达;利用免疫荧光双标法检测小鼠胸11~腰1节段DRGs内脏痛敏感神经细胞中iNOS表达。结果:治疗第7天,电针组小鼠腹泻指数评分较造模组降低(P<0.01),电针组小鼠的腹壁撤退反射评分较造模组和假针刺组显著降低(均P<0.01);电针组小鼠的中心区域停留时间较造模组和假针刺组小鼠显著增加(均P<0.05)。WB结果显示,与造模组和假针刺组相比,电针组小鼠的结肠组织和DRGs中iNOS蛋白含量显著减少(均P<0.05);免疫荧光结果提示电针组小鼠DRGs中iNOS表达较造模组和假针刺组均减少(均P<0.05)。结论:电针刺激大肠俞能够改善IBD模型小鼠的炎性腹泻与内脏痛,并且能够改善内脏痛所致的焦虑,其机制可能与减少结肠和DRGs组织内炎性因子iNOS的生成,从而缓解了iNOS诱发的肠道炎症疼痛有关。

Objective:To observe the effects of electroacupuncture(EA)on Dachangshu to ameliorate visceral pain in inflammatory bowel diseases(IBD)mice and to discuss the potential mechanism tentatively.Methods:A total of 32 C57BL/6J mice of 8 weeks old were randomly divided into 4 groups:control(CTRL)group,IBD(TNBS)group,TNBS+EA group and sham electroacupuncture(TNBS+SHAM)group.IBD model was established by TNBS injection into the colon of mice and after that each mouse received corresponding treatment for 7 days.The diarrhea index scores during 7 days of EA,and the abdominal withdraw reflex score(AWRs)and open field test(OFT)at the 7th day of EA were recorded to assess the severity of colonic inflammation,visceral pain and the anxiety.Western blotting was used to evaluate the production of iNOS in colon and dorsal root ganglions(DRGs)of thoracic 11 to lumber 1 tissue of mice,and the double-labeling immunofluorescence was used to test the iNOS in neuronal cells related to visceral hyperalgesia in DRGs.Results:The diarrhea index scores in TNBS+EA group were decreased on the 7th day of EA treatment(P<0.01),meanwhile the AWRs in TNBS+EA group were improved as compared with those in the TNBS group(P<0.01)and the TNBS+SHAM group(P<0.01).The center zone time in TNBS+EA group was prolonged as compared with that in TNBS and TNBS+SHAM groups(P<0.05).The production of iNOS was declined in TNBS+EA group as compared with that in TNBS group and TN BS+SHAM group(P<0.05).The results of the double-labeling immunofluorescence indicated that the number of DRGs with co-location of iNOS and IB4 was reduced in TNBS+EA group as compared with that in TNBS group and TNBS+SHAM group(P<0.05).Conclusion:EA on Dachangshu could ameliorate inflammatory diarrhea,visceral pain and anxiety in IBD mice induced by TNBS through decreasing the expression of iNOS and its inflammatory stimulation on colon and DRGs.