Treatment Of Myocardial Ischemia With Bone Marrow-derived Mesenchymal Stem Cells Bearing Hepatocyte Growth Factor And Studies On Its Correlative Mechanisms

Coronary heart disease is characterized by the occlusion of a coronary vessel that results in local myocardial ischemia, death of some cardiomyocytes and local collagen deposition. To cure the disease, both restoration of the local blood flow and regeneration of the lost cardiomyocytes must be achieved. In view of the powerful angiogenic activity and anti-fibrosis function of hepatocyte growth factor (HGF), and the ability of mesenchymal stem cells (MSC) to differentiate to cardiomyocytes, we treat the cardiac ischemia with a strategy of combination MSC and HGF in a rat's myocardial ischemia model. On the base of this, we further investigated the cellular or molecular mechanisms of HGF-regulated angiogenesis. The aims of the present study were to: (1) find out the feasibility and effect of MSC modified with HGF gene in the treatment of myocardial ischemia, and provide experimental foundations for clinical treatment of organ or tissue injury by stem cells modified with certain genes; (2) investigate the mechanisms of HGF-regulated angiogenesis, including the mobilization effect of HGF on bone marrow stem cells and biological functions of HGF on the endothelial cells and related molecular events. Thus provide experimental data for using HGF gene modify stem cells to treat ischemia-related diseases.Human or rat's bone marrow-derived MSC were isolated using Percoll density gradient method and according to the adherent property of MSC. Immunohistochemical staining showed that these cells expressed c-Met, receptor for HGF. Then the cells were treated with recombinant human HGF and the mitosis, differentiation and migration of the cells were observed. Results showed that HGF had no obvious effect on proliferation and differentiation of MSC, but could induce migration of MSC markedly and could protect MSC from detachment-induced apoptosis (anoikis). Based on these results, we infected rat's MSC with adenovirus vector encoding GFP or HGF gene (Ad-HGF or Ad-GFP). The efficiency of infection and expression of HGF were detected by FACS and ELISA assay respectively. It showed that transfection efficiency was increasing along with the increase of MOI. Anearly 100% cells were transfected when the 400 MOI was administrated. MSC expressed HGF at a high level when being transfected with the Ad-HGF. The HGF expression could maintain for at least two weeks. Then the female rats' myocardial ischemia model was established by ligation of left anterior descending (LAD) coronary artery and male's MSC overexpressing HGF was injected into risk areas. Results showed that cell implantation improved the function of impaired myocardium, including diminishing the area of ischemia, increasing the number of. capillaries and reduction of fibrosis. Hemodynamics assay showed that left ventricular end-diastolic pressure (LVEDP) decreased but left ventricular systolic pressure (LVSP) and LV peak velocities of contraction and relaxation (dP/dtmax) increased remarkably compared to the ligation control group and Ad-GFP treatment group, indicating that both systolic and diastolic functions were best preserved in the mesenchymal stem cells expressing HGF group after myocardial infarction. By using sry gene as a marker, we also identified that the engrafted cells or their progenies incorporated into ischemic cardiac muscle. These results showed that treatment of myocardial ischemia with bone marrow-derived MSC overexpressing HGF could be a novel strategy that can both restore of the local blood flow and regenerate the lost cardiomyocytes.Then we investigated the cellular or molecular mechanisms of HGF in promotion of angiogenesis. Firstly, in view of following facts: (1) HGF, also known as scatter factor (SF), is a multifunctional factor that can promote mobility or migration of a variety of cell type. (2) Recent studies showed that bone marrow-derived stem cells played important roles in angiogenesis. We assessed whether HGF recruited bone marrow-derived stem cells into blood circulation to participate in postnatal angiogenesis and endothelium repair.