The Study On The Therapy Of Autotransplantation With Mesenchymal Stem Cells For End-stage Congestive Heart Failure

Objective: Cellular cardiomyoplasty has been proposed as an alternative strategy for augmenting the function of diseased myocardium. Experimental data have suggested that mesenchymal stem cells (MSC) may contribute to the healing of myocardial infarction(MI). We will investigate whether autotransplantation with MSC can treatend-stage heart failure and its mechanism of via intracoronary autologous mesenchymalstem cell transplantation.Methods:1. To establish a controllable swine congestive heart failure model, ten pigs were randomly divided into 2 groups, namely control group and CHF group. The control group received sham operation, while the CHF received rapid right ventricular pacing at the rate of 230 beats per minute for 4 weeks, and 190 beats per minute for another 4 weeks, Clinical symptoms and hemodynamic parameters were used to evaluate the severity of heart failure .2. Bone marrow nspirate was taken from the iliac crest of healthy swines, and MSCs were isolated with Ficoll lymophocyte, then co-cultured with 5-aza at different concentration for 24hours. After cultured for additional 2 weeks, MSCs were identified by means of immunohistochemistry technique.3. The coding sequence of VEGF was cloned from human brain by reverse transcription PCR (RT-PCR) and was subcloned into pENTR eukaryotic expression vector. The recombinant VEGF vector was then transfected into cardiomyocyte-like cells mediated by transfection reagent. Analysis by restriction enzyme digestion was carried out to demonstrate the sequence of the Ad-VEGF and its activity was thendetermined using adenovirus transfected cardiomyocyte-like cells .4. Heart failure was created in 25 swines by right ventricular pacing-induced. All the swines were divided into five groups: DMEM implantation group(DMEM); MSC implantation group (MSC); cardiomyocyte-like cell implantation group(CSC); VEGF implantation group (VEGF) and VEGF-cardiomyocyte-like cell implantation group CSC-VE). The stem cells were injected into coronary artery of the porcines with heart failure under DSA. The ejection fraction (EF)> the thickness of left ventricular wall, the diastolic diameter of left ventricle (LVDd) and CO were assessed by echocardiography before and 4 week after implantation.The TNF ^ TGF were detected with ELISA in the serum;the percentage of apotosis^ the concentrtion of collagen and the ratio of collagen I /III were detected in the heart tissues5. Dialated cardiomyopathy patients with heart failure were divided into two groups:the control group(A) ; MSC implantation group (MSC).The thickness of left ventricular wall, the diasiolic diameter of left ventricle (LVDd) and EF were assessed by echocardiography before and 4 week after implantation.Results1. As pacing prolonged, the pigs showed variant symptoms and the hemodynamic parameters implied that the severity of heart failure increased progressively.2. A part of the cells induced by 5-aza had morphological changes and 29 % stained positively Troponin I.3. The results of restriction enzyme and DNA sequencing revealed that VEGF cloning was successful. The adenovirus vector with VEGF could express active VEGF protein in the cardiomyocyte-like cells.4. Compared with that of the control group, LVDd of implantation groups was smaller 4 weeks after stem cells implantation (P<0.05), whereas EF was increased (P<0.05):. The TNF . the concentrtion of collagen and the percentage of apotosis decreased, whereas TGF. the ratio of collagen I /III increased in implantation groups vs control group.5. The MSC transplantation had significantly sustained improvements in left ventricular ejection fraction than that before transplant. The improvement in EF was similar in the MSC group and the induced MSC group.Conclusion:1. The congestive heart failure model induced by rapid ventricular pacing has the advantage of controllability and is a useful animal model for clinical research of heart failure.2. The MSCs from adult bone marrow can differentia...