SHH基因转染BMMSC心肌移植对大鼠心肌梗死后血管新生的影响

Effects of BMMSC transfected with SHH gene transplantation on angiogenesis in rat myocardial infarcted area

目的探讨SHH（Sonic hedgehog）基因转染的骨髓间充质干细胞（BMMSCSHH）移植对大鼠心肌梗死区移植部位血管再生的影响。方法密度梯度离心法及贴壁培养获得BMMSC后，电转染法将SHH基因转入骨髓间充质干细胞。以结扎法制备大鼠急性心肌梗死模型，将实验动物200只大鼠按随机数字表法分5组，每组40只；分别在梗死与正常交界部位移植BMMSCSHH（转染组）、等量的BMMSC（细胞组）、pcDNA3．1-ShhDNA（基因组）、BMMSC和pcDNA3．1-ShhDNA的混合物（混合组）、等容积的低糖DMEM培养基（对照组）。于移植后第8周取正常心肌与心肌梗死交界区组织标本行免疫组化染色标记血管，通过血管密度分析法分析各组间交界区血管密度差异。结果经CD34免疫组化染色后行血管密度分析显示：转染组（36．48±5．22）较对照组（16．71±3．41）血管密度增加（P〈0．01）；较细胞组（29．46±2．27）、基因组（28．78±2．91）及混合组（29．55±4．55）血管密度增加（P〈0．05）。结论BMMSCSHH移植能有效表达目的基因、促进心肌梗死后血管新生，为缺血性心脏病的细胞转基因治疗提供理论依据。

Objective To investigate the angiogenesis at myocardial infarcted area of rats undergoing the transplantation of bone marrow mesenchymal stem cells （BMMSCs） transfected with SHH gene （ BMMSCSHH）. Methods After BMMSC was harvested from rats by density gradient centrifugation and adherent culture. The SHH gene was electrotransfected into BMMSC. Rat model of myocardial infarction that was constructed by ligation of the left anterior descending coronary artery were randomly divided into 5 groups （40 rats for each group）. BMMSCSHH was injected into rat model in the transfected group （group T） at the border of the myocardial infarction zone, and the same amount of BMMSC were injected into the cell group （group B）. Liposome-pcDNA3. 1-SHH I）NA compound was injected into the gene group （group G）. The same amount of BMMSC pulse plasmid was injected into the mix group （ group M）. Culture medium was injected into control group. At the 8th week after transplantation, the rats＇hearts were harvested and the border of the myocardial infarction zone was checked by immunohistochemical staining. Capillary density in the myocardial infarcted area of rats in each group was analyzed. Results The capillary densities analysis via CD34 immunohistochemistry showed the capillary densities in the group T （36.48 ± 5. 22） were significantly higher than that in the control group （ 16. 71 ±3.41 ） （ P 〈0. 01 ）, group B （29.46 ±2. 27）, group G （28. 78 ± 2. 91 ）, and group M （29. 55 ± 4. 55 ） （ P 〈 0. 05 ）. Conclusions BMMSC transfected with SHH gene was beneficial to its stable expression. Transplantation of BMMSC transfected with SHH gene was helpful for angiogenesis at myocardial infarcted area of rats. Its outcome was better than gene or cell therapy alone.