CDMP1转基因间充质干细胞片修复兔软骨缺损

Repairing larynx cartilage defects with MSCs cell sheets transfected by CDMP1

目的 探讨软骨源性形态发生蛋白-1（CDMP-1）转基因细胞片修复兔软骨缺损的能力.方法 腺病毒转染CDMP1基因至第4代兔骨髓间充质干细胞（BMSCs）中,连续培养7～14 d后接种至温度敏感性培养皿中制备细胞片,real time PCR和Western blot法检测转基因细胞片的CDMP1及Ⅱ型胶原蛋白表达;用细胞片工程技术构建组织工程化细胞片并移植入兔甲状软骨缺损处修复软骨缺损.实验分：1）BMSCs细胞片组;2）转染As-CMV-eGFP细胞片组;3）转染As-CMV-hCDMP1-IRES-eGFP细胞片组.分别于术后4和8周检测工程化软骨的Ⅱ型胶原蛋白及蛋白多糖（GAG）表达.结果 检测到转基因细胞片中CDMP1的表达;所获得的工程化软骨免疫组化和阿利新蓝染色显示：A组Ⅱ型胶原蛋白和GAG表达阳性,B组和C组Ⅱ型胶原蛋白和GAG表达阴性（P＜0.05）.结论 CDMP转基因细胞片具有较好的软骨分化活性,可有效地修复兔喉软骨缺损.

Objective To detect the repairing capacity in rabbit cartilage defects by BMSCs cell sheets transfected by CDMP. Methods Rabbit bone marrow mesenchymal stem ceils were Isolated and cultured; the passage 4 cells were tansfected with exogenous recombinant human cartilage-derived morphogenetic protein 1 by adenovirus vector; the transgenosis cell sheets were harvested with temperature-responsive culture dish after 7-14 days. After detecting the expression of CDMP1 in transgenie cell sheet, the sheets were implanted into the rabbit thyroid cartilage defects. After 4 or 8 weeks, the culturing systems were analyzed at the gross level as well as at the histological level to observe the repair ability. The samples were divide into three groups ： 1 ） transfected by As-CMV-hCDMP1 -IRES-eGFP BMSCs cell sheets;2） transfected by Ad-CMV-eGFP BMSCs cell sheets;3）BMSCs cell sheets. Results The expression of CDMP1 in transgenosis cell sheets can be detected by real time PCR and Western blot; the detection of immunohistochemistry and Alcian Blue staining showed the expression of collagen Ⅱ and GAG in group A, while negative expression in group B and group C. There was statistical difference between group A, group B and C （P 〈 0. 05）. Conclusions CDMP transgenosis cell sheets with superior chondrogenic capacity can effectively repair rabbit thyroid cartilage defects.