Recent studies show that bone marrow stem cells (BMSC), mobilized or transplanted, can colonize the infarcted myocardium and participate in de novo regeneration of the myocardium. Several difficulties, however, afflict heart cell therapy approach due to lack of optimized methodology for directed differentiation of donor cells to adopt cardiac phenotype and to enhance their survival following transplantation. Ischemic preconditioning can impart cytoprotective influence against subsequent lethal ischemic injury. The present application is built on the premise that preconditioning of stem cells evokes multiple signaling pathways leading to their enhanced survival and engraftment in the ischemic myocardium. Hence, the central hypothesis of this proposal is that preconditioning of donor cells with preconditioning mimetics and cytokines enhances their survival, proliferation and differentiation potential in the ischemic myocardium after transplantation. To address this issue, we propose to determine whether 1) preconditioning renders the BMSC resistant to subsequent ischemic injury and the preconditioned BMSC overexpress growth factors or other protective proteins which reduce cell death and ultimately lead to successful engraftment and proliferation in the ischemic myocardium 2) transfection of BMSC with cytoprotective proteins enhance their survival and transdifferentiation potentials 3) mobilized stem cells by preconditioning stress release paracrine factors which protect myocytes from ischemic injury 4) preconditioning induced upregulation of protective proteins and growth factors involve protein kinase C signaling pathways 5) preconditioning BMSC with different cytokines/ growth factors promote angiogenic or myogenic response or commitment. Both in vitro and in vivo models will be used to support the proposed specific aims. A wide range of multidisciplinary techniques including biochemistry, cell biology, molecular biology, electron microscopy, immunocytochemistry and pharmacological approaches will be used to integrate the information at the cellular and molecular level with the function at the organ level. RT-PCR and Western blotting will be used to identify potential protein candidates and their importance in stem cell survival, homing, proliferation and differentiation in the ischemic heart. Thus signaling cascade of preconditioning will be exploited to promote provide both targeted commitment of stem cells as well as their enhanced survival rate. Knowledge gained with the novel approaches listed will have solid impact on the success of cell based therapies to treat cardiovascular diseases.
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