摘要
的 研究泻火解毒法对竹叶青蛇伤血管内皮细胞炎性因子的影响.方法 ①动物实验：选择50只健康新西兰大白兔,根据计算机统计软件产生的随机数分为5组,每组10只.采用经兔右后腿皮下注射0.75 mL/kg竹叶青蛇毒液的方法复制竹叶青蛇伤动物模型;对照组经兔右后腿皮下注射等量生理盐水.制模后6h,低、中、高剂量蛇伤胶囊组给予蛇伤胶囊,剂量分别为174、348和522 mg· kg-1·d^-1,使用时以生理盐水稀释为17.4、34.8和52.2 g/L的药液,灌胃量则均为10 mL· kg-1·d^-1;对照组和模型组灌胃等量生理盐水;各组均每日灌胃1次,连续1周.于末次灌胃后24 h经耳缘静脉取血,分离血清,用五分类血细胞计数仪检测血中白细胞计数（WBC）和单核细胞（MO）数;用酶联免疫吸附试验（ELISA）测定血清白细胞介素（IL-1、IL-2、IL-6）和肿瘤坏死因子-α（TNF-α）水平.②细胞实验：用MEM培养液培养人脐静脉内皮细胞（HUVEC） 24 h后更换培养液并按随机数字表法分为5组,每组10个样本.采用细胞中加入5 mg/L竹叶青蛇毒液的方法复制竹叶青蛇毒细胞模型.培养6h后,空白对照组和模型组给予10％正常兔血清培养液,低、中、高剂量蛇伤胶囊组分别加5％、10％和15％含中药兔血清继续培养.培养72 h后,收集细胞培养液,用ELISA试验检测HUVEC培养液中IL-1、IL-2、IL-6、TNF-α水平.结果 模型组血WBC、MO、IL-1、IL-2、IL-6、TNF-α和HUVEC培养液中IL-1、IL-2、IL-6、TNF-α水平均较对照组明显升高,随着药物剂量增加,低、中、高剂量蛇伤胶囊组上述指标均较模型组下降,以中剂量蛇伤胶囊组降低更显著[血清：WBC（×10^9/L）为9.03±2.16比12.11±4.38,MO（×10^9/L）为0.44±0.18比0.63±0.16,IL-1 （ng/L）为92.46±9.46比110.80±29.78,IL-2（ng/L）为92.11±17.55比112.0±18.83,IL-6 （ng/L）为481.73±147.12比667.26±226.41,TNF-α（ng/L）为7.12±2.96比9.41±1.76;HUVEC培养液：IL-1 （ng/L）为56.76±10.37比75.80±22.49,IL-2 （ng/L）为76.77±13.73比92.80±17.82,IL-6（ng/L）为231.70±107.91比413.25±178.65,TNF-α（ng/L）为223.98±30.31比252.86±30.75,均P＜ 0.05].结论 竹叶青蛇毒可造成血管内皮细胞炎性损伤;泻火解毒法可治疗竹叶青蛇伤血管内皮炎性损伤.
Objective To investigate the influence of traditional Chinese medicine （TCM） purging fire and removing toxin therapy on levels of vascular endothelial cell inflammatory factors in trimeresurus stejnegeri bites. Methods① Animal experiment： fifty healthy New Zealand white rabbits were chosen. According to random numbers generated by software, they were divided into five groups with 10 animals in each group. Snake venom 0.75 mL/kg was injected into the subcutaneous tissues of rabbits＇ right hind leg to reproduce the models of trimeresurus stejnegeri bites. And the same volume of normal saline was injected into the same site of rabbits in the control group. 6 hours after model establishment, the rabbits in low, middle and high dose Sheshang capsule groups received 174, 348 and 522 mg·kg^-1·d^^-1 of the capsule respectively; capsules were dissolved in normal saline to make liquids with 17.4, 34.8 and 52.2 g/L Sheshang capsule and the volume of gavage was 10 mL·kg^-1·d^^-1 in the three groups. The same volume of normal saline was given to the control and model groups; the gavage was given once daily for a week. Twenty-four hours after the last gavage, the blood of the rabbits was collected through auricular vein and the serum was separated. The numbers of white blood cells （WBC） and mononuclear cells （MO） were measured by the blood cell counter. The levels of interleukins （IL-1, IL-2, IL-6） and tumor necrosis factor -α （TNF-α） of rabbit serum were measured by enzyme linked immunosorbent assay （ELISA）. ② Cell experiment： the human umbilical venous endothelial cell （HUVEC） was cultured with MEM. After culture for 24 hours, the medium was replaced and the cells were randomly divided by using random number table into five groups with 10 samples in each group. The trimeresurus stejnegeri bite models were reproduced by culturing the cells with 5 mg/L snake venom. After culture for 6 hours, the cells of blank and model groups received 10% blank normal rabbit serum cultural media for filrther culture, while the cells of low, middle and high dose Sheshang capsule groups received rabbit serum containing 5%, 10% and 15% traditional Chinese medicine respectively for further culture. After culture for 72 hours, the medium was collected for measurements of the levels of IL-1, IL-2, IL-6 and TNF-α by ELISA. Results The levels of WBC, MO in blood and IL-1, IL-2, IL-6, TNF- ot in the serum and HUVEC medium in the model group were all higher than those in the control group. Along with the increase of dosage of the capsule, the contents of the above indexes in low, middle and high dose Sheshang capsule groups were all decreased compared with those in the model group. And the degrees of descent in the middle dose Sheshang capsule group were the most significant [in blood： WBC （ ×10^9/L） was 9.03±2.16 vs. 12.11±4.38, MO （×10^9/L） was 0.44±0.18 vs. 0.63±0.16; in serum： IL-1 （ng/L） was 92.46±9.46 vs. 110.80±29.78, IL-2 （ng/L） was 92.11±17.55 vs. 112.00±8.83, IL-6 （ng/L） was 481.73±147.12 vs. 667.26±226.41, TNF-α （ng/L） was 7.12±2.96 vs. 9.41±1.76, in the HUVEC medium： IL-1 （ng/L） was 56.76 ±10.37 vs. 75.80±22.49, IL-2 （ng/L） was 76.77±13.73 vs. 92.80±17.82, IL-6 （ng/L） was 231.70±107.91 vs. 413.25±178.65, TNF-α （ng/L） was 223.98±30.31 vs. 252.86±30.75, all P 〈 0.05]. Conclusions Trimeresurus stejnegeri venom may damage vascular endothelial cells by causing inflammatory response; the TCM therapy of purging fire and removing toxin can treat inflammatory injury of vascular endothelial ceils induced by Trimeresurus stejnegeri snakebites.
出处
《中国中西医结合急救杂志》
CAS
北大核心
2016年第6期632-635,共4页
Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care
基金
国家自然科学基金（81302978）
福建省自然科学基金（2012J01379）
福建中医药大学校管课题重点学科项目（X2014033-学科,X2014034-学科,X2014035-学科）
关键词
泻火解毒法
蛇伤胶囊
竹叶青蛇伤
血管内皮细胞
炎性因子
The therapy of purging fire and removing toxin
Sheshang capsule
Trimeresurus stejnegeri bite
Vascular endothelial cells
Inflammatory factors
