重组人血小板生成素对内毒素所致血小板减少症小鼠的影响  

Effect of recombinant human thrombopoietin on endotoxin-induced thrombocytopenia in mice

作　　者：蒋佳维 王兵[2] 余信 许华 王勇强[2] Jiang Jiawei;Wang Bing;Yu Xin;Xu Hua;Wang Yongqiang(Tianjin Medical University First Center Clinical College,Tianjin 300192,China;Department of Critical Care Medicine,Tianjin First Center Hospital,Tianjin Emergency Medicine Research Institute,Tianjin 300192,China)

机构地区：[1]天津医科大学一中心临床学院,300192 [2]天津市第一中心医院重症医学科,天津市急救医学研究所,300192

出　　处：《中华危重病急救医学》2019年第9期1108-1112,共5页Chinese Critical Care Medicine

基　　金：天津市卫生行业重点攻关项目(14KG101);国家临床重点专科建设项目(2011-873)。

摘　　要：目的探讨重组人血小板生成素(rhTPO)对内毒素脂多糖(LPS)所致血小板减少症(TCP)小鼠的影响。方法将60只雄性C57BL/6小鼠按随机数字表法分为生理盐水(NS)对照组(NS组)、脓毒症致TCP模型组(LPS组)和rhTPO处理组(LPS+rhTPO组),每组20只。给予小鼠单次腹腔注射LPS 30 mg/kg制备脓毒症致TCP模型;NS组给予等量NS。LPS+rhTPO组于腹腔注射LPS后即刻皮下注射rhTPO 2.7 kU/kg,每24 h注射1次;NS组和LPS组均皮下注射等量NS。各组均观察72 h。分别于制模前及制模后每24 h取小鼠目内眦血,应用动物血细胞计数仪检测血小板计数(PLT)。于制模后72 h取小鼠眼球血,应用流式细胞仪检测富血小板血浆中CD61+CD62p+细胞比例,反映血小板活化情况;处死小鼠取肺脏及脾脏组织,采用免疫组化法检测CD41的阳性表达,反映器官内血小板扣押情况;取小鼠单侧股骨的骨髓细胞,应用流式细胞仪检测CD41+CD61+细胞比例,反映骨髓巨核细胞增殖情况。结果制模前各组小鼠PLT比较差异无统计学意义。随制模后时间延长,LPS组小鼠PLT呈持续降低趋势,72 h略有回升,但仍明显低于NS组(×109/L:308.60±21.70比1152.72±50.27,P<0.05);LPS+rhTPO组PLT随制模后时间延长呈持续升高趋势,72 h时明显高于LPS组(×109/L:926.78±48.85比308.60±21.70,P<0.05)。制模后72 h,LPS组小鼠富血小板血浆中CD61+CD62p+细胞比例较NS组明显升高〔(25.07±2.55)%比(4.17±0.38)%,P<0.05〕,而LPS+rhTPO组CD61+CD62p+细胞比例较LPS组明显下降〔(15.92±1.26)%比(25.07±2.55)%,P<0.05〕;LPS组小鼠骨髓巨核细胞中CD41+CD61+细胞比例较NS组明显升高〔(11.84±0.80)%比(3.60±0.42)%,P<0.05〕,LPS+rhTPO组CD41+CD61+细胞比例较LPS组进一步升高〔(30.96±2.49)%比(11.84±0.80)%,P<0.05〕;免疫组化显示,LPS组肺脏和脾脏组织中CD41的阳性表达显著高于NS组〔A值:肺脏为828.94±119.30比447.09±16.19,脾脏为(280.15±16.71)×103比(0.65±0.26)×103,均P<0.05〕,而LPS+rhTPO�Objective To investigate the effect of recombinant human thrombopoietin(rhTPO)on thrombocytopenia(TCP)induced by endotoxin lipopolysaccharide(LPS)in mice.Methods Sixty male C57BL/6 mice were divided into normal saline(NS)control group(NS group),sepsis-induced TCP model group(LPS group)and rhTPO treatment group(LPS+rhTPO group)by random number table with 20 mice in each group.Sepsis-induced TCP model was reproduced by one intraperitoneal injection of LPS 30 mg/kg,and the mice in NS group were given the same amount of NS.In LPS+rhTPO group,2.7 kU/kg rhTPO was subcutaneously injected into mice immediately after intraperitoneal injection of LPS,once every 24 hours.The mice in NS group and LPS group were injected subcutaneously with the same amount of NS.The observation period of each group lasted for 72 hours.The inner canthus blood was harvested before and every 24 hours after modeling,and the platelet count(PLT)was measured by animal blood cell counter.The eyeball blood of mice was harvested at 72 hours after modeling,and the proportion of CD61+CD62p+cells in platelet-rich plasma was detected by flow cytometry,by which the platelet activation was reflected.Lung and spleen tissues of mice were harvested,and the positive expression of CD41 was determined by immunohistochemistry,by which the platelet sequestration in organs was reflected.Bone marrow cells from unilateral femur of mice were harvested,and the proportion of CD41+CD61+cells was determined by flow cytometry to reflect the proliferation of bone marrow megakaryocytes.Results There was no significant difference in PLT among the groups before modeling.With the extension of the time after modeling,PLT in LPS group was decreased continuously,and increased slightly at 72 hours,but it was still significantly lower than that in NS group(×109/L:308.60±21.70 vs.1152.72±50.27,P<0.05);PLT in LPS+rhTPO group was increased continuously with the extension of modeling time,and it was significantly higher at 72 hours than that in LPS group(×109/L:926.78±48.85 vs.308.60

关 键 词：骨髓巨核细胞 血小板减少症 重组人血小板生成素 血小板活化 富血小板血浆 CD41 C57BL/6小鼠 流式细胞仪检测 

分 类 号：R28[医药卫生—中药学]