多联袋无菌制备脐带血间质干细胞的方法及效果  

Sterile preparation of umbilical cord derived mesenchymal stem cells with multiple bags:Method and effect

作　　者：李建斌[1] 满勇[2] 单泓[1] 赵林娜[1] 段艳丽[1] 

机构地区：[1]河南省红十字血液中心,河南省郑州市450012 [2]郑州市第二人民医院,河南省郑州市450000

出　　处：《中国组织工程研究与临床康复》2007年第24期4781-4784,共4页Journal of Clinical Rehabilitative Tissue Engineering Research

摘　　要：目的:分析人脐带血间质干细胞分离培养方法及临床应用的安全性。方法:实验于2005-04/12在河南省红十字血液中心和郑州市第二人民医院完成。实验方法:①无菌条件下将脐带血液采集到无菌采血袋内,产妇及其家属均知情同意,并经医院伦理委员会批准。随机抽查62份样本,单份采集量80～160mL,计数有核细胞。②以1:1体积比将Ficoll-Hypaque混合溶液加入四联袋主袋内,将四联袋与脐带血收集袋相连,再将样本血液沿袋壁缓慢加入四联袋主袋内的液面上。离心后将四联袋主袋放在分浆夹上,袋内液体自上而下分为血浆血小板层、单个核细胞层、淋巴细胞分离液层、多核细胞层、红细胞层5层。将血浆血小板层上2/3液体挤压入四联袋一空袋内,将剩余的血浆血小板层、单个核细胞层、淋巴细胞分离液层上部挤压入四联袋另一空袋内,即为富含有核细胞液体。③随机选择本院2005-04/12收治脑卒中后遗症患者10例。男9例,女1例,35～75岁,病程3个月～7年。诊断均由CT、MRI检查证实,符合第四届全国脑血管病学术会议修订的脑卒中诊断标准,患者均对治疗方案知情同意。将脐带血间充质干细胞经静脉输注入患者体内,每次输注1份,每例患者平均输注6份,每份间隔时间平均4d。于治疗前和治疗3个月后评价患者神经功能状态、肢体运动功能及日常生活活动能力。采用脑卒中患者临床神经功能缺损程度评分标准评估神经功能,分数越低,神经功能状态越好;采用Fugl-Meyer评估患侧肢体运动功能,分数越高,肢体运动功能越好;采用Barthel指数评估日常生活活动能力,分数越高,日常生活活动能力越高。结果:①分离的62份脐血间质干细胞,每份含有有核细胞(3.62±2.39)×109,红细胞残余量为(1.64±1.25)×109,CD34+细胞的比率为(2.31±0.93)%,CD133+的比率为(1.36±1.23)%。培养7d后贴壁细胞为(3.27±1.85)×107,CD133+�AIM： To analyze the isolated culture method of human umbilical cord derived mesenchymal stem ceils and the safety of clinical application. METHODS： The experiment was performed at the Henan Red Cress Blood Center and Zhengzhou Second People＇s Hospital from April to December 2005. ①Cord blood was collected sterilely in sterile blood bag. Puerperant and their family members knew and agreed to participate the trial that was approved by ethics committee. Totally 62 samples were randomly checked with the single collection volume of 80-160 mL. Karyocytes were counted. ②Ficoll-Hypaque mixed solution at the ratio of 1：1 was put in major bag of quadriad bags that was connected with the collection bag of cord blood, and then sample blood was added slowly in major bag of quadriad bags along bag walt. After centrifugalization, major bag of quadriad bags was put on plasma isolation clamp, so the liquor was isolated into platelet layer, mononuclear cell layer, lymphocyte isolation liquor layer, polykaryocyte layer and erythrecyte layer from upper to lower. Two-thirds liquor on platelet layer was pressed into a empty quadriad bags, and vestigial upper platelet layer, mononuclear cell layer and lymphocyte isolation liquor layer were pressed in another empty quadriad bags, namely abundant karyocyte liquor. ③Totally 10 patients with stroke residual were enrolled randomly from April to December 2005, including 9 males and 1 female, aged 35-75 years, with the disease course of 3 months to 7 years. The diagnosis was verified by CT and MRI, which met the diagnostic standards revised by Fourth National Cerebrevascular Disease Academic Conference. The patients knew the therapeutic regimen and agreed. Umbilical cord derived mesenchymal stem cells were injected into the patients by intravenous infusion, once a portion, 6 portions in each patient averagely, with the interval of mean 4 days between two portions. Neural Function, limb motor function and ability of activities of daily living were evaluated before treatment and 3

关 键 词：胎盘脐带血 间质干细胞 脑卒中后遗症 

分 类 号：R394.2[医药卫生—医学遗传学]