Myocardial regeneration therapy using nanotechnology enhances self-regeneration in damaged myocardium

1.片状成肌细胞植入扩张型心肌病仓鼠27周龄BIO-2(DCM)大鼠心脏中度重构作为受体。将分离自Bio F1B仓鼠的成肌细胞培养在涂有聚N-异丙基丙烯酰胺的培养皿中。采用3种不同的治疗方法：(1)片状成肌细胞移植(S组，n=29)；(2)成肌细胞注射(M组，n=28)；(3)假手术(C组，n=28)。S组在左室壁植入2枚片状成肌细胞，M组将成肌细胞分别注入右室壁和左室壁。植入片状成肌细胞移植物后，超声心动图显示左心室扩张的内径明显缩小，而其他组的心脏则表现为左室扩张的进展。短轴缩短率在M组和C组显著降低，而S组维持在术前…术后3个月达到较高水平。组织学检查显示，与其他组相比，S组左室壁增厚，成肌细胞存活，心肌纤维化减轻。免疫组织化学染色显示，S组大鼠心肌细胞基底膜有α-肌聚糖和β-肌聚糖表达，其他各组均未见表达。S组的预期寿命显著延长。片状成肌细胞移植改善了DCM仓鼠的心功能和延长了预期寿命，相关因素包括减少心肌纤维化和细胞骨架蛋白的重组。2.成肌片植入大鼠心肌梗死模型左冠状动脉前降支结扎的Lewis大鼠心脏(2周)通过成肌细胞注射或植入2张单层构建的成肌细胞片(每片5×10(6)个细胞)或通过介质注射的方法获得1×10(7)自体骨骼肌成肌细胞。超声心动图结果显示，在细胞移植后8周，成肌细胞片组的心功能改善程度高于其他组。组织学比较显示，只有成肌片组的心脏在明显的均匀增厚的壁内有更多的细胞和大量广泛的新生毛细血管。与骨骼肌成肌细胞注射相比，骨骼肌成肌片植入显著减少了纤维化。成肌细胞片组心肌梗死区的造血干细胞(c-kit、干细胞抗原1和CD34)数量明显增多。逆转录-聚合酶链式反应结果显示基质衍生因子1、肝细胞生长因子、血管内皮生长因子的表达情况如下：成肌细胞片&GT；成肌细胞注射&GT；对照组。成肌层通过释放基质衍生因子1和其他生长因子，修复受损的心肌，减少纤维化，并防止与基质衍生因子1和其他生长因子的募集相关的重塑。3.自杀基因系统调节梗死大鼠心脏血管生成的作用我们利用单纯疱疹病毒(TK)的胸苷激酶基因和更昔洛韦(GCV)系统开发了产生人肝细胞生长因子(HHGF)的细胞，它调节hHGF的产生。我们测试了这些细胞是否诱导和调节了梗死心肌中的血管生成效应。将hHGF基因表达载体(NIH/HGF)稳定地导入NIH3T3细胞。然后，将TK(NIH/HGF/TK)稳定地导入NIH/HGF细胞。在重度联合免疫缺陷大鼠心脏结扎左前降支，移植4种材料：1)NIH/HGF(n=10)，2)NIH/HGF/TK(n=10)，3)NIH3T3(n=10)，4)培养液(n=10)。在体外，GCV可抑制NIH/HGF/TK细胞的增殖。在体内，移植后4周，NIH/HGF组和NIH/HGF/TK组的心功能和血管生成显著增加。尽管NIH/HGF组有肿瘤病变，但NIH/HGF/TK组肿瘤生长完全受控。使用TK-GCV自杀基因系统的血管生成基因细胞治疗在细胞生长控制下诱导和调节血管生成。较少

1.Sheet-shaped myoblast implantation in dilated cardiomyopathic hamstersMale 27-week-old BIO TO-2 (DCM) hamsters that showed moderate cardiac remodeling were used as recipients. Myoblasts isolated from BIO F1B hamsters were cultured on dishes coated with poly(N-isopropylacrylamide). Three different therapies were conducted : (1)sheet-shaped myoblast graft implantation (S group, n=29) ; (2)myoblast injection (M group, n=28) ; and (3)sham operation (C group, n=28). In the S group, two sheet-shaped myoblast grafts were implanted on the left ventricle (LV) wall, and in the M group, myoblasts were injected into the right ventricle (RV) and LV walls. After the sheet-shaped myoblast grafts were implanted, echocardiography demonstrated that the dilated LV dimension was significantly reduced, whereas the hearts in other groups showed a progression of LV dilation. The fractional shortening in the M and C groups decreased significantly while that in the S group was maintained at the preoperative … More level for 3 months after the operation. Histological examination demonstrated that in the S group, the LV wall thickness was increased, with viable myoblasts, and myocardial fibrosis was decreased compared with the other groups. Immunohistochemical staining demonstrated alpha-sarcoglycan and beta-sarcoglycan expression on the basement membrane of the cardiomyocytes in the S group but not in the other groups. The life expectancy was significantly prolonged in the S group. Sheet-shaped myoblast graft implantation improved cardiac performance and prolonged life expectancy, associated with a reduction in myocardial fibrosis and re-organization of the cytoskeletal proteins in DCM hamsters.2.Myoblast sheet implantation in rat myocardial infarction modelLeft anterior descending coronary artery-ligated Lewis rat hearts (2 weeks) received 1 x 10(7) autologous skeletal myoblasts by means of cell transplantation either through myoblast injection or implantation of 2 monolayer-constructed myoblast sheets (5 x 10(6) cells per sheet) or through medium injection. Echocardiographic results indicated higher improvement of cardiac performance in the myoblast sheet group than among the other groups until 8 weeks after cell transplantation. Histologic comparison revealed greater cellularity and abundant widespread neocapillaries within the noticeable uniform thickened wall in myoblast sheet group hearts only. Fibrosis was substantially reduced with skeletal myoblast sheet implantation compared with skeletal myoblast cell injection. Obviously higher numbers of hematopoietic stem cells (c-kit, stem cell antigen 1, and CD34) were observed in the myoblast sheet group infarct heart region. Reverse transcription-polymerase chain reaction results showed expression of stromal-derived factor 1, hepatocyte growth factor, and vascular endothelial growth factor as follows : myoblast sheets > myoblast injection > control. Myoblast sheets repaired the impaired myocardium, reduced fibrosis, and prevented remodeling in association with recruitment of hematopoietic stem cells through the release of stromal-derived factor 1 and other growth factors.3.Suicide gene system regulates the effect of angiogenesis in infarcted rat heartWe developed human HGF (hHGF)-producing cells that regulated hHGF production using the thymidine kinase gene of Herpes Simplex Virus (TK) and the Ganciclovir (GCV) system. We tested whether these cells induced and regulated angiogenic effects in infarcted myocardium. NIH3T3 cells were stably transfected with an hHGF cDNA expression plasmid (NIH/HGF). Next, the NIH/HGF cells were stably transfected with TK (NIH/HGF/TK). The left anterior descending artery was ligated in the heart of severe combined immunodeficiency rats, and four materials were transplanted : 1)NIH/HGF (n=10), 2)NIH/HGF/TK, with orally administered GCV (n=10), 3)NIH3T3 (n=10), and 4)culture medium (n=10). In vitro, the proliferation of NIH/HGF/TK cells was suppressed by GCV. In vivo, significant increases in cardiac performance and angiogenesis were observed in the NIH/HGF and NIH/HGF/TK groups 4 weeks after transplantation. Although tumorous lesions were detected in the NIH/HGF group, their growth was completely controlled in the NIH/HGF/TK group. Angiogenic gene cell therapy using the TK-GCV suicide gene system induces and regulates angiogenesis under the control of cell growth. Less