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

使用纳米技术的心肌再生疗法可增强受损心肌的自我再生

基本信息

批准号：
15209046
负责人：
MATSUDA Hikaru
金额：
$ 29.37万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2005
项目状态：
已结题

项目摘要

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

1.在扩张的心肌病仓鼠中的片状成肌细胞植入显示中度心脏重塑的27周龄雄性BIO TO-2（DCM）仓鼠用作受体。将从BIO F1 B仓鼠分离的成肌细胞在用聚（N-异丙基丙烯酰胺）包被的培养皿上培养。实验分为三组：（1）成肌细胞片状移植组（S组，n=29）;（2）成肌细胞注射组（M组，n=28）;（3）假手术组（C组，n=28）。在S组中，将两个片状成肌细胞移植物植入左心室（LV）壁，在M组中，将成肌细胞注入右心室（RV）和LV壁。植入片状成肌细胞移植物后，超声心动图显示扩张的LV尺寸显著减小，而其他组的心脏显示LV扩张进展。M组和C组的短轴缩短率显著降低，而S组的短轴缩短率维持在术前水平。 ...更多信息手术后3个月。组织学检查显示，S组左室壁厚度增加，成肌细胞存活，心肌纤维化程度减轻。免疫组化染色显示S组心肌细胞基底膜上有α-肌聚糖和β-肌聚糖表达，而其他组无表达。S组平均寿命明显延长。片状成肌细胞移植物植入改善了心脏性能并延长了预期寿命，与DCM仓鼠心肌纤维化和细胞骨架蛋白重组的减少相关。2.大鼠心肌梗死模型中的成肌细胞片植入左前降支冠状动脉结扎的刘易斯大鼠心脏（2周）接受1 × 10（7）通过成肌细胞注射或植入2个单层构建的成肌细胞片（每张5 x 10（6）个细胞）或通过培养基注射进行细胞移植的自体骨骼成肌细胞。超声心动图结果表明，直到细胞移植后8周，成肌细胞片组的心脏性能改善程度高于其他组。组织学比较显示，仅在成肌细胞片组心脏中，在明显均匀增厚的壁内存在更大的细胞结构和丰富的广泛的新包膜。与骨骼肌成肌细胞注射相比，骨骼肌成肌细胞片植入显著减少了纤维化。在成肌细胞片组梗死心脏区域观察到明显较高数量的造血干细胞（c-kit、干细胞抗原1和CD 34）。逆转录-聚合酶链反应结果显示基质衍生因子1、肝细胞生长因子和血管内皮生长因子的表达如下：成肌细胞片>成肌细胞注射液>对照。成肌细胞片修复受损的心肌，减少纤维化，通过释放基质衍生因子1和其他生长因子来防止与造血干细胞募集相关的重塑。3.自杀基因系统调节梗死大鼠心脏中血管生成的作用我们开发了产生人HGF（hHGF）的细胞，其利用单纯疱疹病毒（TK）的胸苷激酶基因来调节hHGF的产生。和更昔洛韦（GCV）系统。我们测试了这些细胞是否诱导和调节梗死心肌的血管生成作用。用hHGF cDNA表达质粒（NIH/HGF）稳定转染NIH 3 T3细胞。接着，用TK稳定转染NIH/HGF细胞（NIH/HGF/TK）。结扎重症联合免疫缺陷大鼠心脏左前降支，移植NIH/HGF（n=10）、NIH/HGF/TK（n=10）+GCV（n=10）、NIH 3 T3（n=10）和4）培养基（n=10）。GCV可抑制NIH/HGF/TK细胞的增殖。在体内，移植后4周，NIH/HGF和NIH/HGF/TK组中观察到心脏性能和血管生成显著增加。尽管在NIH/HGF组中检测到肿瘤病变，但在NIH/HGF/TK组中它们的生长被完全控制。使用TK-GCV自杀基因系统的血管生成基因细胞疗法在细胞生长的控制下诱导和调节血管生成。少