人羊膜负载猪骨髓间充质干细胞体外生长的形态特点(英文)  

Morphological study on the growth of human amniotic membrane loaded with porcine bone marrow-derived mesenchymal stem cells

作　　者：闫国和[1] 艾国平[1] 汪代杰[2] 邹仲敏[1] 冉新泽[1] 王军平[1] 李蓉[1] 粟永萍[1] 程天民[1] 

机构地区：[1]解放军第三军医大学预防医学院全军复合伤研究所创伤,烧伤与复合伤国家重点实验室,重庆市400038 [2]泸州市人民医院肿瘤科,四川省泸州市646000

出　　处：《中国组织工程研究与临床康复》2007年第15期2985-2989,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：国家自然科学基金面上项目(30200142);国家重点基础研究发展规划项目("九七三"项目)(2005CB522605);军队"十一五"攻关项目(06G076)~~

摘　　要：背景:人羊膜含胶原、糖蛋白、蛋白多糖、整合素和板层体等多种成分,表达多种生长因子及其mRNA的相关蛋白,能为细胞的增殖、分化提供丰富的营养成分,有利于细胞的生长繁殖,其能否负载培养猪骨髓间充质干细胞良好生长值得研究。目的:建立人羊膜负载培养猪骨髓间充质干细胞生长的组织工程方法,观察骨髓间充质干细胞生长增殖的形态特点。设计:随机对照观察。单位:创伤、烧伤和复合伤国家重点实验室,解放军第三军医大学预防医学院全军复合伤研究所。材料:实验于2003-01/11在解放军第三军医大学预防医学院全军复合伤研究所,创伤、烧伤与复合伤国家重点实验室完成。选用3只出生二三个月的贵州小香猪,雌雄不拘,体质量6～8kg,由解放军第三军医大学实验动物中心提供。主要试剂:IMDM培养基(Hyclone,USA);优质胎牛血清PAA(Germany);Eosin B(Sigma,USA);OCT包埋剂(USA)。主要仪器:BX51立式荧光显微镜(Olympus,Japan);IX70型倒置荧光显微镜(Olympus,Japan);恒冷切片机(2700-Frigcut,德国);骨髓穿刺针(江苏);超净工作台(苏净集团安泰公司);CO2恒温培养箱(QUEUE,USA)。方法:按文献所述方法准备人羊膜,将按文献所述方法分离、培养的贵州小香猪骨髓间充质干细胞贵州小香猪的骨髓间充质干细胞原代培养、传代扩增后,以不同密度(0.84×105/cm2,1.54×105/cm2,2.75×105/cm2)接种于人羊膜基质面,倒置显微镜及扫描电镜下观察不同培养密度骨髓间充质干细胞的生长增殖变化情况;将第2、3代骨髓间充质干细胞以1.54×105/cm2的密度作为合适种植密度接种到由聚酯环支撑的人羊膜的基质面,最长培养至18d,逐日经卷膜、切片、苏木精-伊红染色光镜下观察人羊膜负载骨髓间充质干细胞生长情况,同时给予扫描及透射电镜。主要观察指标:不同培养密度及不同时间点骨髓间充质干细胞在人羊膜上的生长情�BACKGROUND： Human amniotic membrane （HAM） contains various ingredents such as collagen, glycoprotein, proteoglycan, integrin and laminated body, and so on, and expresses many kinds of growth factors and mRNA-associated proteins. And these ingredents can supply abundant nutriments for cellular proliferation and differentiation, and benefit cells to grow and propagate. Whether or not HAM can load porcine bone marrow-derived mesenchymal stem cells （BMSCs） to well grow on it deserves to be further investigated. OBJECTIVE： To set up a method of tissue engineering of human amniotic membrane loading porcine BMSCs and observe the morphological characteristics of growth and proliferation of BMSCs seeded on HAM. DESIGN ： Randomized controlled observation. SETTING： State Key Laboratory of Trauma, Burn and Combined Injury, General Institute of Combined Injuries, Academy of Preventive Medicine, Third Military Medical University of Chinese PLA. MATERIALS： This experiment was carried out in the State Key Laboratory of Trauma, Burn and Combined Injury, General Institute of Combined Injuries, Academy of Preventive Medicine, Third Military Medical University of Chinese PLA between January and November 2003. Three Guizhou minipigs of either gender, aged 2 to 3 months, weighing from 6 to 8 kg, were provided by the Experimental Animal Center, Third Military Medical University of Chinese PLA. Main reagent： ISCOVE＇S modified DULBECCO＇S medium （IMDM） culture medium （Hyclone, USA）; high-quality fetal bovine serum PAA （Germany）; haematoxylin （China）; Eosin B （Sigma, USA） and OCT embedding medium （USA）. Main instruments： BX51 stereoscopic fluorescence microscope （Olympus, JaPan）; IX70 inverted fluorescence microscope （Olympus, Japan）; cryostat （2700-Frigcut, Germany）; myeloid puncture needle （Jiangsu）; superclean bench （Sujing Bloc Antai Company）; CO2 constant-temperature incubator （QUEUE, USA）. METHODS ： HAM was prepared as previously descnbed. The BMSCs

关 键 词：人羊膜 猪骨髓间充质干细胞 体外生长 形态特点 

分 类 号：R394.2[医药卫生—医学遗传学]