穴位埋线对哮喘大鼠肺组织中p38MAPK信号通路及细胞间黏附分子-1、白细胞介素-4和嗜酸性粒细胞的影响

Acupoint catgut embedment may reduce airway inflammation reaction by down-regulating ICAM-1 and EOS by suppressing p38MAPK signaling in lung tissue of asthmatic rats

目的:观察穴位埋线对哮喘大鼠肺组织中p38丝裂原活化蛋白激酶(p38MAPK)、细胞间黏附分子-1(ICAM-1)、白细胞介素-4(IL-4)和嗜酸性粒细胞(EOS)的影响,探讨其通过调控EOS减轻哮喘的作用机制。方法:40只SPF级雄性Wistar大鼠随机分为对照组、模型组、穴位埋线组和地塞米松组,每组10只。采用腹腔注射卵蛋白(OVA)和氢氧化铝混悬液法制备哮喘大鼠模型。穴位埋线组于"肺俞""定喘""膻中"处行埋线治疗1次;地塞米松组予腹腔注射地塞米松,每日1次,连续2周。HE染色法观察大鼠肺组织形态结构的改变,瑞氏-姬姆萨染色法计数肺泡灌洗液(BALF)中EOS的相对含量,TUNEL法检测大鼠肺组织EOS的凋亡水平,透射电镜观察大鼠肺组织EOS超微结构的变化,荧光定量PCR法检测大鼠肺组织中p38MAPK、ICAM-1和IL-4 mRNA的相对表达量。结果:(1)与对照组比较,模型组可见明显的支气管变形和炎性细胞浸润;与模型组比较,穴位埋线组和地塞米松组支气管变形和周围炎性细胞浸润程度缓解。(2)与对照组比较,模型组BALF中EOS含量显著增加(P<0.01);与模型组比较,穴位埋线组和地塞米松组BALF中EOS含量显著下降(P<0.01);与穴位埋线组比较,地塞米松组BALF中EOS含量下降(P<0.05)。(3)与对照组比较,模型组肺组织EOS的凋亡指数显著下降(P<0.01);与模型组比较,穴位埋线组和地塞米松组肺组织EOS的凋亡指数显著升高(P<0.01)。(4)与对照组比较,模型组肺组织EOS细胞膜缺损,细胞质不均匀,嗜酸性颗粒肿胀、中心键偏移;与模型组比较,穴位埋线组和地塞米松组肺组织EOS包膜完整,细胞质皱缩,且细胞内可见自噬小体或自噬泡结构。(5)与对照组比较,模型组肺组织中p38MAPK、ICAM-1和IL-4 mRNA相对表达量显著增加(P<0.01);与模型组比较,穴位埋线组和地塞米松组肺组织中p38MAPK、ICAM-1和IL-4 mRNA相对表达量显著下降(P<0.01,P<0.05);与穴位埋线组比较,地塞米松组肺组织中ICAM-1 mRNA相对表达量下降(P<0.05)。结论:穴位埋线可能是通过抑制p38MAPK、ICAM-1和IL-4 mRNA的表达,减少肺组织EOS的聚集,促进EOS的凋亡,从而缓解哮喘气道炎性反应。

Objective To observe the effect of acupoint catgut embedment(ACE)on expression of p38 mitogen activated protein kinase(p38 MAPK),intercellular adhesion molecule-1(ICAM-1),interleukin-4(IL-4)and eosinophils(EOS)in lung tissue of asthmatic rats,so as to explore its mechanism underlying improvement of asthma.Methods Forty male Wistar rats were randomly divided into control,model,ACE and dexamethasone(DEX)groups,with 10 rats in each group.The asthmatic model was established by intraperitoneal injection of mixture suspension(1 mL)of ovalbumin(OVA,10%)and 10%Al(OH)_(3)+normal saline,followed by inhalation of atomized 1%OVA solution for 30 min,once daily for 2 weeks to trigger occurrence of asthmatic symptoms.The ACE was applied once to“Feishu”(BL13),“Dingchuan”(EX-B1)and“Danzhong”(CV17).Rats of the DEX group were given intraperitoneal injection of DEX once a day for 2 weeks.H.E.staining was used to evaluate histopathological changes of the lung tissue.The relative number of EOS in the bronchoalveolar lavage fluid(BALF)was detected by Wright Giemsa staining.The apoptosis level of EOS in the lung tissue was detected by TUNEL staining.The ultrastructural changes of EOS in the lung tissues were observed by transmission electron microscope(TEM).The expression of p38 MAPK,ICAM-1 and IL-4 mRNAs in the lung tissue was detected by quantitative real-time PCR.Results Findings of optical microscope and TEM showed obvious bronchial deformation and inflammatory cell infiltration,rupture of EOS cell membrane,uneven cytoplasm with swelling and uneven density of eosinophilic granules in EOS of the model group,which was relatively milder in the ACE and DEX groups.Compared with the control group,the EOS number in BALF and the expressions of p38 MAPK,ICAM-1 and IL-4 mRNAs in the lung tissue were significantly increased(P<0.01),and the apoptosis index of EOS in the lung tissue was significantly decreased(P<0.01)in the model group.After intervention,the EOS number in BALF and expression levels of p38 MAPK,ICAM-1 and IL-4 mRNAs in the lung tissue of ACE and DEX groups were significantly decreased(P<0.01,P<0.05),as well as the apoptosis index of EOS in the lung tissue was significantly increased in both ACE and DEX groups(P<0.01)in comparison with the model group.The EOS number in BALF and expression of ICAM-1 mRNA were significantly lower in the DEX group than those in the ACE group(P<0.05).Conclusion Catgut embedding at acupoints may alleviate the airway inflammatory response in asthma rats,which may be related with its effects in down-regulating p38 MAPK signaling,ICAM-1 and IL-4 mRNA expression,reducing the aggregation of EOS,and promoting the apoptosis of EOS.