猪脱细胞主动脉瓣叶不同方法预处理后种植内皮细胞的比较研究

Effect of different pre-condition on decellular porcine aortic valve and seeding of endothelial cells: A comparative study

目的:猪脱细胞主动脉瓣叶经不同的方法预处理后,种植新生牛主动脉内皮细胞(BAECs),观察细胞的增殖状态.方法:猪脱细胞主动脉瓣叶分别经纤维连接蛋白(Fn,50μg/m1)预铺、胎牛血清(FCS)浸泡、无血清DMEM浸泡预处理12 h,采用间隔24 h、3次种植的方法种植BAECs,于种植后第4天、第7天进行细胞计数,并行MTT法和3H-TdR掺入量检测比较细胞的增殖状态.同时于第7天取瓣叶进行组织学检测,观察瓣叶表面内皮细胞覆盖情况.结果:种植后第4天和第7天,采用Fn预铺、FCS浸泡预处理的脱细胞瓣叶,其细胞计数、MTT的光密度和3H-TdR掺入值明显高于无血清DMEM预处理组(P＜0.01).瓣叶经H-E染色,可见Fn预铺、FCS浸泡预处理组内皮细胞的覆盖情况优于无血清DMEM浸泡组.结论:Fn或FCS预处理能明显增加内皮细胞在脱细胞瓣叶表面的黏附和增殖,有利于组织工程心脏瓣膜的体外构建.

Objective:To observe bovine aortic endothelial cells(BAECs) proliferation after they were seeded on decellu-lar porcine heart valve leaflets pre-conditioned by different methods. Methods: Decellular porcine aortic In-art valve leaflets were used as scaffold. They were pre-conditioned by fibronectin(Fn) >fetal calf serum(FCS) and Dulbecco minimum essential medium(DMEM) without FCS for 12 h.then BAECs were seeded onto them at 24 h interval for 3 times. On 4 and 7 d after seeding, the proliferations of BAECs were observed by cell counting,MTT,3H-thymidine (3H-TdR) incorporation. Results: There were significant differences in endothelial cell proliferation between group Fn,group FCS and group DMEM (P<0. 01) by cell counting, MTT, and 3H-TdR incorporation. As for histology,the endothelial cell proliferations of group Fn and FCS were better than group DMEM. Conculsion:Decellular porcine heart valve leaflets pre-conditioned by Fn and FCS can enhance the cell proliferation on the scaffold. These methods are helpful for creating tissue-engineered heart valve in vitro.