人胎盘来源的造血干细胞和间充质干细胞的分离及鉴定  

Isolation and identification of human placenta-derived hematopoietic stem cells and mesenchymal stem cells

作　　者：李欣 赵中利[1] 罗晓彤[1] 李珈莹 曹阳[1] 于永生[1] LI Xin;ZHAO Zhong-li;LUO Xiao-tong;LI Jia-ying;CAO Yang;YU Yong-sheng(Branch of Animal Husbandry,Jilin Academy of Agricultural Sciences,Gongzhuling 136100,Jilin Province,China)

机构地区：[1]吉林省农业科学院畜牧科学分院,吉林公主岭136100 [2]吉林省银丰生物工程技术有限公司,吉林长春130012

出　　处：《中国生物制品学杂志》2019年第1期95-99,105,共6页Chinese Journal of Biologicals

摘　　要：目的建立人胎盘来源的造血干细胞(placenta hematopoietic stem cells,hP-HSCs)和间充质干细胞(placentaderived mesenchymal stem cells,hP-MSCs)的分离方法,并进行鉴定和组分分析。方法选取10份新生健康婴儿胎盘组织,采用机械破碎法联合磁珠分选法分离h P-HSCs,胎盘绒毛膜组织块贴壁法分离hP-MSCs,利用形态学观察、集落培养、流式细胞术等进行鉴定。结果 hP-HSCs:分离后有核细胞数(total nucleated cell number,TNC)为(11. 82±2. 46)×10~8个,TNC回收率≥80%;细胞活性为(99. 7±0. 3)%;细胞表面抗体CD34^+CD45dim表达率为(8. 69±0. 36)%,CD34^+总数为(108. 0±6. 48)×10~6个;集落形成总数为(1. 88±1. 07)×10~6。hP-MSCs:冻存细胞总数为(40. 78±9. 35)×10~7个;细胞活性为(99. 0±1. 5)%;细胞表面抗体CD34^+CD45^+表达率为(0. 1±0. 1)%,CD44^+CD105^+为(99. 6±0. 2)%,CD14^+CD19^+为(0. 1±0. 1)%,CD90^+CD73+为(98. 9±0. 2)%;且具有良好成脂、成骨分化潜能。结论成功建立了hP-HSCs和hP-MSCs体外分离培养方法,为胎盘的临床应用奠定了基础,并提供了细胞种子资源。Objective To develop a method for isolation of human placenta-derived hematopoietic stem cells and mesenchymal stem cells,identify the isolated cells and analyze their components.Methods Ten placental tissues of healthy newborns were collected,from which placenta hematopoietic stem cells(hP-HSCs) were isolated by mechanical disruption combined with magnetic bead sorting,while placenta-derived mesenchymal stem cells(hP-MSCs)by placental chorionic membrane tissue block adherence method,observed for morphology,and identified by colony culture and flow cytometry.Results The total nucleated cell(TNC) number of hP-HSCs was(11.82 ± 2.46)×10^8,while the TNC recovery was not less than 80%,the cell viability was(99.7 ± 0.3)%,the proportion of CD34^+CD45 dim cells was(8.69 ± 0.36)%,the total CD34^+ cell number was(108.0 ± 6.48)× 10^6,and the total number of colonies formed was(1.88 ± 1.07) × 10^6.However,the total number of frozen hP-MSCs was(40.78 ± 9.35) × 10^7,while the cell viability was(99.0 ± 1.5)%,and the proportions of CD34^+CD45^+,CD45^+CD105^+,CD14^+CD19^+ and CD90^+CD73^+ cells were(0.1 ± 0.1)%,(99.6 ± 0.2)%,(0.1 ± 0.1)% and(98.9 ± 0.2)%,respectively.The hP-MSCs showed potential in osteogenic and adipogenic differentiations.Conclusion The methods were isolation and culture of h P-hSCs and hP-MSCs were successfully developed,which laid a foundation of clinical application of placenta and provided a resource of cell seeds.

关 键 词：胎盘 造血干细胞 间充质干细胞 组织块法 磁珠分选 

分 类 号：Q28[生物学—细胞生物学]