灌注法制备离体大鼠心脏脱细胞基质  

Preparation of heart decellularized matrix in rats by perfusing method

作　　者：彭蒙蒙[1] 林贤丰[1] 邵培刚[1] 王辉[1] 张建色[1,2] 王志斌[2] 梅劲[1,2] 

机构地区：[1]温州医学院人体解剖学教研室,浙江温州325035 [2]温州医学院生物支架移植与免疫研究所,浙江温州325035

出　　处：《解剖学报》2012年第4期559-563,共5页Acta Anatomica Sinica

基　　金：国家自然科学基金资助项目(81071576);浙江省教育厅科研资助项目(Y201016491);温州市科技局资助项目(Y20100012);浙江省大学生科技创新活动计划"新苗人才计划"资助项目(2011R413015)

摘　　要：目的探索制备离体大鼠心脏脱细胞生物支架材料的新方法,为心脏组织工程研究提供三维立体天然支架。方法取30只成年SD大鼠心脏,运用冻融加化学萃取的组织工程学方法(胰蛋白酶、十二烷基硫酸钠和曲拉通X-100)处理离体大鼠心脏,同时观察心脏大体形态及颜色变化,并对脱细胞支架进行基因组DNA分析;HE染色,免疫荧光法,扫描和透射电镜进一步检测鉴定脱细胞支架的生物学特征。结果心脏脱细胞支架外观透明,包膜完整,维持心脏三维立体结构,肉眼可见心脏内脉管系统;脱细胞支架DNA残留量不及对照组的3%;HE染色、扫描和透射电镜结果显示,心脏脱细胞生物支架去细胞彻底,细胞外基质网状结构保留完整;免疫荧光结果表明,胶原、弹性蛋白等细胞外支架成分保留较完整,未见明显细胞核成分残留。结论运用冻融加化学萃取法所制备的离体心脏脱细胞生物支架去细胞彻底,细胞外基质保留较完整,是较为理想的心脏三维立体生物支架材料。Objective The aim is to explore an effective method of preparing the whole heart decellularized bio- derived scaffolds （HDBS） in rats for the heart tissue engineering. Methods The hearts were harvested from 30 adult SD rats. HDBS were prepared by the treatment with freeze thawing, typsin, SDS and Triton X-100, and obeserved the deeellularization process at the same time. After decellularization, the scaffolds were examined through genomic DNA content analysis, HE staining, scanning electron microscope, transmission electron microscope and immunofluorescence. Results HDBSs maintained the three-dimensional structure with transparent appearance. Based on the DNA quantitative analysis, DNA content of the HDBSs was less than 3% compared to that of the control hearts. HE staining, scanning electron microscope and transmission electron microscope revealed that HDBS were deeellularized completely and preserved the full extraeellular matrix, lmmunofluorescenee results showed that collagen, elastin and other extracellular components were preserved completely without obvious nuclear components. Conclusion The HDBS prepared by the treatment with freeze thawing, typsin, SDS and Triton X-100 are deeellularized completely, with the full extracellular matrix preserved, which are the ideal three-dimensional bio-derived scaffold material of the heart.

关 键 词：脱细胞 心 细胞外基质 组织工程 免疫荧光 大鼠 

分 类 号：R322[医药卫生—人体解剖和组织胚胎学]