三七总皂苷对川崎病模型小鼠血管炎性损伤的影响

Effects of Panax Notoginseng Saponins on Vasculitis Injury in Kawasaki Disease Model Mice

目的:探讨三七总皂苷(PNS)对川崎病模型小鼠血管炎性损伤和炎性细胞因子表达的影响。方法:BALB/C雄性小鼠经腹腔注射1 mg·ml^(-1)的干酪乳杆菌细胞壁(LCWE)提取物0.5 ml建立川崎病模型。造模成功后随机分为模型组、PNS低、中、高(100,200,400 mg·kg^(-1))剂量组、阿司匹林组(250 mg·kg^(-1)),每组10只;另取10只小鼠经腹腔注射0.5 ml生理盐水,作为正常对照组;各组连续给药7 d,1次/d。给药期间观察小鼠饮食、体质量等一般状况;采用酶联免疫吸附(ELISA)检测血清中血管内皮素(ET)、血管内皮生长因子(VEGF)、肿瘤坏死因子(TNF-α)、白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)含量;用苏木精-伊红(HE)染色观察冠状动脉病理变化;用蛋白免疫印迹法(Western blot)检测TNF-α、内皮细胞黏附分子(CAM-1)、单核细胞趋化蛋白-1(MCP-1)、血管内皮微粒(EMP)蛋白表达水平;免疫组化检测炎症通路核转录因子κB(NF-κB)磷酸化指标p-NF-κB在血管组织中的表达。结果:与正常对照组相比,模型组小鼠出现颤抖、饮食活动减少现象,冠状动脉血管膜增厚、炎性细胞浸润等病理损伤严重,小鼠体质量下降(P<0.05),血清TNF-α、IL-1β、IL-6、ET、VEGF水平及冠状动脉组织TNF-α、CAM-1、MCP-1、EMP蛋白、p-NF-κB阳性表达均显著升高(P<0.05)。与模型组相比,PNS低、中、高剂量组及阿司匹林组小鼠颤抖及冠状动脉血管膜增厚、炎性浸润等病理损伤缓解,血清TNF-α、IL-1β、IL-6、ET、VEGF水平及TNF-α、CAM-1、MCP-1、EMP蛋白、p-NF-κB阳性表达均显著下降(P<0.05),且PNS各组呈剂量依赖性。结论:PNS可降低NF-κB/TNF-α炎症通路活化,降低炎症刺激引起的血管损伤,从而缓解川崎病小鼠血管炎性损伤状态。

Objective:To investigate the effects of panax notoginseng saponins(PNS)on the vasculitis injury and expression of inflammatory cytokines in Kawasaki disease(KD)mice.Methods:The male BALB/c mice were intraperitoneally injected with 1 mg·ml^(-1)of L.Casei cell wall(LCWE)extract(0.5 ml)to build KD model.The model mice were randomly divided into model group,PNS low dose group(100 mg·kg^(-1)),medium dose group(200 mg·kg^(-1))and high dose group(400 mg·kg^(-1))and aspirin positive control group(250 mg·kg^(-1))with 10 in each group;another 10 mice were injected with 0.5 ml normal saline intraperitoneally as normal control group.Each group was continuously administered for 7 days,once a day.The diet and weight of mice were observed during the administration,the levels of tumor necrosis factor(TNF-α),interleukin-1β(IL-1β),interleukin-6(IL-6)and vascular endothelial injury markers of endothelin(ET)and vascular endothelial growth factor(VEGF)were detected by enzyme-linked immunosorbent assay(ELISA).The coronary arteries of mice were taken and the pathological changes were observed by hematoxylin eosin(HE)staining.Western blot was used to detect the relative expression levels of TNF-α,endothelial cell adhesion molecule(CAM-1),monocyte chemoattractant protein-1(MCP-1)and endothelial microparticle(EMP)in coronary artery;immunohistochemical detection was used to determine nuclear transcription factor(NF-κB)phosphorylation index p-NF-κB in vascular tissues.Results:Compared with those in the normal control group,mice in the model group showed tremor,decreased dietary activity and severe pathological damage such as coronary artery vascular membrane thickening and inflammatory cell infiltration;the body weight of the mice was decreased(P<0.05);serum TNF-α,IL-1β,IL-6,ET and VEGF levels and coronary artery tissue TNF-α,CAM-1,MCP-1,EMP protein and p-NF-κB positive expression increased significantly(P<0.05).Compared with those in the model group,mice in the PNS low,medium and high dose groups and the aspirin group alleviated pathological damage such as tremor,coronary artery vascular membrane thickening and inflammatory infiltration;serum TNF-α,IL-1β,IL-6,ET and VEGF levels and coronary artery tissue TNF-α,CAM-1,MCP-1,EMP protein and p-NF-κB positive expression decreased significantly(P<0.05).The effects of PNS groups were dose-dependent.Conclusion:PNS can reduce the activation of NF-κB/TNF-αinflammatory pathway,reduce the vascular damage caused by inflammation,and alleviate the vasculitis damage of Kawasaki disease mice.