共价结合生长因子的胶原补片改善大鼠室壁瘤修补术后移植细胞生存率的实验研究  

Growth Factor-conjugated Collagen Patch Prolongs Survival Rate of Transplanted Cells after Ventricular Aneurysm Repair in Rats

作　　者：康凯[1] 曲辉[2] 汤继权[3] 谢宝栋[1] 贾智博[1] 张宇南[1] 吴华[1] 李仁科[1] 蒋树林[1] 

机构地区：[1]哈尔滨医科大学附属第二医院心血管外科,心肌缺血机理与诊疗技术省部共建教育部重点实验室,哈尔滨150086 [2]哈尔滨医科大学附属第二医院儿内科,哈尔滨150086 [3]鸡西市人民医院胸心血管外科,黑龙江鸡西158100

出　　处：《中国胸心血管外科临床杂志》2013年第4期451-456,共6页Chinese Journal of Clinical Thoracic and Cardiovascular Surgery

基　　金：黑龙江省自然科学基金项目(LC2012C03);黑龙江省卫生厅科学技术项目资助(2012-634)~~

摘　　要：目的观察用碳化二亚胺[1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride chemistry,EDC]法共价结合血管内皮细胞生长因子(vascular endothelial growth factor,VEGF)+碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)或VEGF的胶原补片对接种于其上的人骨髓间充质干细胞(human bone marrow mesen-chymal stem cells,hBM-MSCs)作用的体内外表现。方法将EDC法活化的胶原补片作对照组,将活化后与VEGF或VEGF+bFGF共价结合的胶原补片作实验组(VEGF组、VEGF+bFGF组),以hBM-MSCs作种子细胞(0.5×106/补片),构建工程化心肌片(engineered heart tissue,EHT)。利用MTT法评价hBM-MSCs在3种支架上的体外增殖情况;结扎雄性SD大鼠左冠状动脉前降支制作心肌梗死后室壁瘤模型,以构建的3种EHT分别行左心室重建术。术后4周取材,抗α平滑肌动蛋白(α-SMA)免疫荧光染色观察EHT功能血管再生情况,抗人线粒体抗体(anti-h-mitochondria)免疫荧光染色观察移植细胞生存情况。结果 (1)hBM-MSCs在VEGF组及VEGF+bFGF组的增殖在移植后2 d、4 d均明显优于对照组(P<0.05)。(2)EHT移植后4周,抗α-SMA免疫荧光染色证明VEGF组及VEGF+bFGF组新生小动脉密度高于对照组(P<0.05);(3)抗h-mitochondria抗体免疫荧光染色示VEGF组及VEGF+bFGF组移植细胞的生存率均高于对照组(P<0.05);(4)相关性分析显示,补片上存留的移植细胞数目与补片新生小动脉密度存在明显的正相关性(r2=0.99,P=0.02)。结论共价结合VEGF或VEGF+bFGF的胶原补片可促进hBM-MSCs的体外增殖;可通过改善局部功能血管的形成改善移植细胞的存活率。Objective To observe the impact of collagen patches using 1-ethyl-3- （3-dimethylaminopropyl） carbod- iimide hydrochloride chemistry （EDC） to conjugate vascular endothelial growth factor （VEGF）＋ basic fibroblast growth factor （bFGF） or VEGF alone on the survival rate of transplanted human bone morrow mesenchymal stem cells （hBM-MSCs） in vitro and in vivo. Methods Collagen patches which were activated by EDC were used as the control group, and EDC activated collagen patches that were conjugated with VEGF or VEGF ＋ bFGF were used as the experiment groups （VEGF 6 group and VEGF ＋ bFGF group）, hBM-MSCs （0.5 X 10/patch） were used as seeding cells to construct engineered heart tissue （EHT）. MTT assay was performed to assess in vitro proliferation of hBM-MSCs on 3 different collagen patches. Ventricular aneurysm model after myocardial infarction was created by left anterior descending artery （LAD） ligation in male SD rats, and EHT which were constructed with 3 different patches were used for ventricular plasty. Four weeks later, immunofluorescence staining was used to examine arteriole density （anti-α-SMA staining） and transplanted cell survival （anti-h-mitochondria staining）. Results （ 1 ） hMSCs proliferation in VEGF group and VEGF ＋ bFGF group was signifi- cantly better than that in the control group on the 2nd and 4th day after cell transplantation （P 〈 0.05 ） ; （2） Four weeks after EHT implantation, immunofluorescence staining for α-SMA revealed that arteriole density of VEGF group and VEGF ＋ bFGF group was significantly higher than that of the control group （P 〈 0.05） ; （3） Immunofluorescence staining for h-mitochondria showed that survival rates of transplanted hBM-MSCs of VEGF group and VEGF ＋ bFGF group were significantly higher than that of the control group （P 〈 0.05 ）; （4） There was a significantly positive correlation between survival rate of hBM-MSCs and arteriole density （r 2=0.99, P=0.02）. Conclusion VEGF or VE

关 键 词：血管内皮细胞生长因子 碱性成纤维细胞生长因子 组织工程 心室成形术 

分 类 号：R654.3[医药卫生—外科学]