蛇伤胶囊对竹叶青蛇伤兔血小板活化功能影响的研究

Study on the effect of Sheshang Capsule on platelet activation function in rabbits injured by Trimeresurus stejnegeri

目的研究蛇伤胶囊对竹叶青蛇伤兔血小板活化功能的影响。方法选取24只新西兰大白兔随机分为正常对照组、模型组、蛇伤胶囊组,每组各8只,雌雄各半,根据前期研究方法模型组和蛇伤胶囊组以兔耳缘静脉注射0.75 mg/kg竹叶青蛇毒液建立竹叶青蛇伤兔模型,正常对照组注射等量生理盐水。注射6 h后予正常对照组和模型组10 ml/(kg·d)生理盐水灌胃,蛇伤胶囊组予10 ml/(kg·d)蛇伤胶囊所配药液灌胃。以上3组连续灌胃1周后兔耳缘静脉采血5 ml,血小板分离提取后应用实时荧光定量聚合酶链式反应技术(qPCR)检测各组血小板活化指标αⅡb整合素(CD41)、整合素β链Ⅲ(CD61)、血小板P选择素(CD62P)、溶酶体颗粒膜糖蛋白(CD63)信使核糖核酸(mRNA)表达,酶联免疫吸附试验(ELISA)检测膜糖蛋白Ⅱb/Ⅲa(GPⅡb/Ⅲa)水平。分析比较各组血小板活化指标CD41、CD61、CD62P、CD63 mRNA表达及GPⅡb/Ⅲa水平差异。结果各组血小板细胞CD41、CD61、CD62P、CD63 mRNA表达及GPⅡb/Ⅲa水平组间比较,差异均有显著统计学意义(P<0.01)。两两组间比较,与正常对照组比较,模型组CD41、CD61、CD62P、CD63 mRNA表达及GPⅡb/Ⅲa水平显著降低,差异有显著统计学意义(P<0.01),蛇伤胶囊组CD41、CD61、CD62P、CD63 mRNA表达显著降低,差异有显著统计学意义(P<0.01),GPⅡb/Ⅲa水平显著降低,差异有统计学意义(P<0.05);与模型组比较,蛇伤胶囊组CD41、CD61、CD62P、CD63 mRNA表达显著增加,差异有显著统计学意义(P<0.01),GPⅡb/Ⅲa水平显著增加,差异有统计学意义(P<0.05)。结论竹叶青蛇伤时血小板活化功能受抑制,蛇伤胶囊可通过提高CD41、CD61、CD62P、CD63、GPⅡb/Ⅲa水平,减轻血小板活化功能抑制,改善血小板活化功能。

Objective To study the effect of Sheshang Capsule on platelet activation function in rabbits injured by Trimeresurus stejnegeri.Methods A total of 24 New Zealand white rabbits were randomly divided into a normal control group,a model group,and a Sheshang Capsule group,with 8 rabbits in each group,half male and half female.According to the previous research method,the model group and the Sheshang Capsule group were injected with 0.75 mg/kg Trimeresurus stejnegeri’s venom into the rabbit ear vein to establish a rabbit injured by Trimeresurus stejnegeri model,while the normal control group was injected with an equal amount of physiological saline.After 6 hours of injection,the normal control group and the model group were gavaged with 10 ml/(kg·d)physiological saline,while the Sheshang Capsule group was gavaged with 10 ml/(kg·d)solution with Sheshang Capsule content.After continuous gavage of the above three groups for one week,5 ml of blood was collected from the rabbit ear vein respectively.After platelet separation and extraction,real-time fluorescence quantitative polymerase chain reaction(qPCR)was used to detect platelet activation indicators,i.e.,integrin α Ⅱb(CD41),integrin β Ⅲ chain(CD61),platelet P-selectin(CD62P),and lysosomal membrane glycoprotein(CD63)messenger RNA(mRNA)expression,and enzyme-linked immunosorbent assay(ELISA)to detect membrane glycoprotein Ⅱb/Ⅲa(GPⅡb/Ⅲa)levels in each group.The differences in platelet activation indicators CD41,CD61,CD62P,CD63mRNA expression,and GPⅡb/Ⅲa levels was analyzed and compared among different groups.Results There were significant differences in CD41,CD61,CD62P,CD63 mRNA expression and GPⅡb/Ⅲa levels between groups(P<0.01).Compared with the normal control group,the expression of CD41,CD61,CD62P,CD63 mRNA,and GPⅡb/Ⅲa levels in the model group were significantly reduced,with the differences were statistically significant(P<0.01),while the expression of CD41,CD61,CD62P and CD63 mRNA in the Sheshang Capsule group was significantly reduced,with the differences were statistically significant(P<0.01),and GPⅡb/Ⅲa levels in the Sheshang Capsule group were significantly reduced,with the differences were statistically significant(P<0.05).Compared with the model group,the expression of CD41,CD61,CD62P,and CD63 mRNA in the Sheshang Capsule group was significantly increased,with the differences were statistically significant(P<0.01),and the level of GPⅡb/Ⅲa in the Sheshang Capsule group was significantly increased,with the differences were statistically significant(P<0.05).Conclusion When injured by Trimeresurus stejnegeri,the platelet activation function is inhibited.The Sheshang Capsule can reduce the inhibition of platelet activation function and improve platelet activation function by increasing the levels of CD41,CD61,CD62P,CD63,and GPⅡb/Ⅲa.