The serine-threonine kinases Pim-1 and Akt regulate cellular proliferation and survival. Although Akt is known to be a crucial signaling protein in the myocardium, the role of Pim-1 has been overlooked. Pim-1 expression in the myocardium of mice decreased during postnatal development, re-emerged after acute pathological injury in mice and was increased in failing hearts of both mice and humans. Cardioprotective stimuli associated with Akt activation induced Pim-1 expression, but compensatory increases in Akt abundance and phosphorylation after pathological injury by infarction or pressure overload did not protect the myocardium in Pim-1-deficient mice. Transgenic expression of Pim-1 in the myocardium protected mice from infarction injury, and Pim-1 expression inhibited cardiomyocyte apoptosis with concomitant increases in Bcl-2 and Bcl-X(L) protein levels, as well as in Bad phosphorylation levels. Relative to nontransgenic controls, calcium dynamics were significantly enhanced in Pim-1-overexpressing transgenic hearts, associated with increased expression of SERCA2a, and were depressed in Pim-1-deficient hearts. Collectively, these data suggest that Pim-1 is a crucial facet of cardioprotection downstream of Akt.

Background Despite numerous studies demonstrating efficacy of cellular adoptive transfer for therapeutic myocardial regeneration, problems remain for donated cells with regard to survival, persistence, engraftment, and long-term benefits. This study redresses these concerns by enhancing the regenerative potential of adoptively transferred cardiac progenitor cells (CPCs) via genetic engineering to overexpress Pim-1, a cardioprotective kinase that enhances cell survival and proliferation. Methods and Results Intramyocardial injections of CPCs overexpressing Pim-1 were given to infarcted female mice. Animals were monitored over 4, 12, and 32-weeks to assess cardiac function and engraftment of Pim-1 CPCs using echocardiography, in vivo hemodynamics, and confocal imagery. CPCs overexpressing Pim-1 show increased proliferation and expression of markers consistent with cardiogenic lineage commitment following dexamethasone exposure in vitro. Animals that received CPCs overexpressing Pim-1 also produce greater levels of cellular engraftment, persistence, and functional improvement relative to control CPCs up to 32-weeks post-delivery. Salutary effects include reduction of infarct size, greater number of c-kit+ cells, and increased vasculature in the damaged region. Conclusions Myocardial repair is significantly enhanced by genetic engineering of CPCs using Pim-1 kinase. Ex vivo gene delivery to enhance cellular survival, proliferation, and regeneration may overcome current limitations of stem cell-based therapeutic approaches.

One of the greatest examples of integrated signal transduction is revealed by examination of effects mediated by AKT kinase in myocardial biology. Positioned at the intersection of multiple afferent and efferent signals, AKT exemplifies a molecular sensing node that coordinates dynamic responses of the cell in literally every aspect of biological responses. The balanced and nuanced nature of homeostatic signaling is particularly essential within the myocardial context, where regulation of survival, energy production, contractility, and response to pathological stress all flow through the nexus of AKT activation or repression. Equally important, the loss of regulated AKT activity is primarily the cause or consequence of pathological conditions leading to remodeling of the heart and eventual decompensation. This review presents an overview compendium of the complex world of myocardial AKT biology gleaned from more than a decade of research. Summarization of the widespread influence that AKT exerts upon myocardial responses leaves no doubt that the participation of AKT in molecular signaling will need to be reckoned with as a seemingly omnipresent regulator of myocardial molecular biological responses.

Abstract-The Notch network regulates multiple cellular processes, including cell fate determination, development, differentiation, proliferation, apoptosis, and regeneration. These processes are regulated via Notch-mediated activity that involves hepatocyte growth factor (HGF)/c-Met receptor and phosphatidylinositol 3-kinase/Akt signaling cascades.

Objective Enhancement of human cardiac progenitor cell (hCPC) reparative and regenerative potential by genetic modification for treatment of myocardial infarction. Background Regenerative potential of stem cells to repair acute infarction is limited. Improved hCPC survival, proliferation and differentiation into functional myocardium will increase efficacy and advance translational implementation of cardiac regeneration. Methods hCPCs isolated from myocardium of heart failure patients undergoing left ventricular assist device (LVAD) implantation are engineered to express green fluorescent protein (GFP; hCPCe) or Pim-1-GFP (hCPCeP). Functional tests of hCPC regenerative potential are performed with immunocompromised mice by intramyocardial adoptive transfer injection after infarction. Myocardial structure and function is monitored by echocardiographic and hemodynamic assessment for 20 weeks following delivery. hCPCe and hCPCeP expressing luciferase are followed by bioluminesence imaging (BLI) to non-invasively track persistence. Results hCPCeP exhibit augmentation of reparative potential relative to hCPCe control cells as demonstrated by significantly increased proliferation coupled with amelioration of infarction injury and increased hemodynamic performance at 20 weeks post-transplantation. Concurrent with enhanced cardiac structure and function, hCPCeP demonstrate increased cellular engraftment and differentiation with improved vasculature and reduced infarct size. Enhanced persistence of hCPCeP versus hCPCe is revealed by BLI at up to 8 weeks post delivery. Conclusion Genetic engineering of hCPCs with Pim-1 enhances repair of damaged myocardium. Ex vivo gene delivery to modify stem cells has emerged as a viable option addressing current limitations in the field. This study demonstrates that efficacy of human CPCs from the failing myocardium can be safely and significantly enhanced through expression of Pim-1 kinase, setting the stage for use of engineered cells in preclinical settings.

STEM CELLS 2008;26:1315-1324 Disclosure of potential conflicts of interest is found at the end of this article.

Mitochondrial morphological dynamics affect the outcome of ischemic heart damage and pathogenesis. Recently, mitochondrial fission protein dynamin-related protein 1 (Drp1) has been identified as a mediator of mitochondrial morphological changes and cell death during cardiac ischemic injury. In this study, we report a unique relationship between Pim-1 activity and Drp1 regulation of mitochondrial morphology in cardiomyocytes challenged by ischemic stress. Transgenic hearts overexpressing cardiac Pim-1 display reduction of total Drp1 protein levels, increased phosphorylation of Drp1-S637 , and inhibition of Drp1 localization to the mitochondria. Consistent with these findings, adenoviral-induced Pim-1 neonatal rat cardiomyocytes (NRCMs) retain a reticular mitochondrial phenotype after simulated ischemia (sI) and decreased Drp1 mitochondrial sequestration. Interestingly, adenovirus Pimdominant negative NRCMs show increased expression of Bcl-2 homology 3 (BH3)-only protein p53 up-regulated modulator of apoptosis (PUMA), which has been previously shown to induce Drp1 accumulation at mitochondria and increase sensitivity to apoptotic stimuli. Overexpression of the p53 up-regulated modulator of apoptosis-dominant negative adenovirus attenuates localization of Drp1 to mitochondria in adenovirus Pim-dominant negative NRCMs promotes reticular mitochondrial morphology and inhibits cell death during sI. Therefore, Pim-1 activity prevents Drp1 compartmentalization to the mitochondria and preserves reticular mitochondrial morphology in response to sI.

Key Words: Pim-1 Ⅲ mitochondria Ⅲ cardiomyocyte Ⅲ apoptosis C ardiovascular disease is the leading cause of death among men and women and affects approximately 33% of the US population. 1 A direct correlation between the decline in heart function and loss of cardiomyocytes via apoptosis involving the mitochondria occurs in cardiomyopathy, myocardial ischemia/reperfusion (I/R), and congestive heart failure. 2-9 Specifically, myocardial I/R injury generates calcium overload and oxidative stress, which initiate the intrinsic apoptotic pathway through activation of the mitochondrial permeability transition pore (mPTP). The ensuing chain of events result in dramatic changes to mitochondrial morphology associated with uncoupling of the electron transport chain, depolarization of the inner membrane, matrix swelling, unfolding of the cristae, and ultimately outer membrane rupture, with release of proapoptotic cytochrome c. 10 -15 Release of cytochrome c into the cytosol consequently activates apoptotic protease-activating factor, which mediates caspase cascade programmed cell death. 16 Thus, preservation of mitochondrial integrity is essential in designing molecular strategies to enhance cardiomyocyte cell survival by blunting injury attributed to cardiomyopathic insult.Cardioprotection mediated by survival kinase signal transduction acts through multiple mechanisms including preservation of mitochondrial integrity. 17 Numerous studies have documented antiapoptotic actions of the serine/threonine kinase AKT, which acts in part through protecting mitochon-