Heart disease is the leading cause of death in the United States. Heart transplantation is the only clinically effective therapy for end-stage heart failure, but it is available to only a few patients. Transplantation of immature cells, such as stem cells or myoblasts, in the infarct area of the heart, known as cell therapy, appears to be a promising alternative strategy. The myoblasts, derived from leg skeletal muscle tissue, can proliferate and differentiate into functional cells and thereby improve heart function. However, the probability of survival/proliferation of the transplanted myoblasts is often less than 1%. Severe hypoxia (low oxygen levels) is hypothesized as a factor responsible for the poor outcome of cell therapy. Thus, there is a pressing need to precisely evaluate the oxygen concentration, particularly intracellular oxygenation, in the transplanted myoblasts as a function of cardiac tissue regeneration. Therefore, the goal of the present study is to develop innovative probes and methods for noninvasive monitoring of the changes in the cellular oxygenation during myocardial regeneration. We have developed a new class of spin probes that can be directly detected by electron paramagnetic resonance (EPR) spectroscopy/imaging with markedly higher sensitivity for detection. The probes are composed of stacks of neutral radicals of lithiated naphthalocyanine macrocyclic ligands. In addition, the probes can report absolute values of tissue oxygen concentration with remarkable oxygen sensitivity (better than 0.1 Torr). They enable precise, accurate and repeated measurements of myocardial oxygenation over a period of several months in closed chests. The probes can be efficiently internalized into cells to study cell proliferation and migration. The objective of this proposal is to utilize these novel probes for noninvasive monitoring of the survival and proliferation of the transplanted cells in the infarct area of the myocardium. The proposal specifically seeks to study: 1. Establishment of optimal internalization procedures and characterization of the spin probe in myoblasts for cell-tracking and determination of intracellular oxygen concentration;2. Noninvasive monitoring of in situ oxygenation at the site of implant, migration, and engraftment of transplanted stem cells in infarcted hearts;3. Evaluation of the efficacy of myoblast stem cells subjected to hypoxic culture or intermittent hypoxia/reoxygenation on the survival, retention, integration, and in situ oxygenation after their transplantation in the infarcted heart. The availability of these innovative probes and technologies will enhance our ability to monitor cell therapy and should open new avenues in the treatment of heart disease in humans.
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