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Background: Using a rat in vivo infarct model we tested the hypothesis that fetal cardiomyocyte (FCM) implantation would increase repolarization heterogeneity. Methods: Four groups of rats were studied: two groups served as controls and underwent injection with FCM or culture medium in a region of the left ventricle (LV) supplied by the left anterior descending artery (LAD), and two groups underwent LAD ligation followed 3 weeks later by the injection of either FCM or culture medium. Epicardial monophasic action potential (MAP) recordings were obtained 4 to 5 weeks after cell injection from the right ventricle (RV), LV infarction region and LV region remote from the LAD. The maximum difference in action potential duration (MAPD90) between the three sites was defined as repolarization heterogeneity. Results: LAD ligation (in the control media injection group) resulted in an increase in repolarization heterogeneity from 6.9 ± 0.9 to 13.8 ± 1.2 ms (p < 0.005). Similarly, injection of FCM without coronary ligation resulted in an increase in heterogeneity from 6.9 ± 3.9 to 20.7 ± 1.3 ms (p = 0.001). However, injection of FCM into regions of infarction did not result in an increase in heterogeneity when compared with the control media group (13.8 ± 1.9 vs 13.0 ± 2.6 ms, respectively, p = 0.752). Conclusions: Both fetal cardiomyocyte engraftment in the normal myocardium and coronary ligation increased repolarization heterogeneity. However, fetal cardiomyocyte engraftment in an infarcted region did not further increase repolarization heterogeneity. [Copyright &y& Elsevier]

Cardioprotective strategies are needed to prevent perioperative myocardial dysfunction in high-risk patients undergoing cardiac surgery. Despite accumulating evidence that statins exert lipid-independent cardioprotective effects, these have been ascribed primarily to improvements in endothelial function and neutrophil-endothelial interaction. The direct effects of statins on cardiomyocytes (independent of endothelial cells) remain unknown. Using a well-characterized model of low-volume hypoxia and reoxygenation, we studied the effects of pravastatin on human ventricular cardiomyocytes. Cardiomyocytes were subjected to 90 min of low-volume hypoxia and 30 min of reoxygenation in the presence and absence of pravastatin (1, 10, and 100 μm) (n = 10 per group). In some experiments, the effects of endothelin (ET) receptor blockade (with bosentan) and nitric oxide synthase (NOS) inhibition (with l-NAME) on pravastatin-mediated cardioprotection were evaluated. Cell survival, NO, and ET-1 production and protein kinase Akt activation were determined. Pravastatin treatment prevented cardiomyocyte cell death following simulated hypoxia and reoxygenation (P < 0.01). This effect was mediated via an increase in NO release, decrease in myocyte ET-1 production/action, and an increase in protein kinase Akt activation. We demonstrate, for the first time, novel protective effects of pravastatin in human ventricular cardiomyocytes independent of endothelial cells or other cell types. Statin therapy may restore ischemic hearts to full functional integrity during cardioplegic arrest through a direct effect on cardiomyocyte survival. [Copyright &y& Elsevier]

Presents a study that found that low-volume anoxia and reoxygenation increased nitric oxide synthase (NOS) activity and facilitated the conversion of L-arginine to nitric oxide (NO) to protect against heart cell injury. Measurement of NO production; Characterization of NOS isoforms in human heart cells after low-volume anoxia and reoxygenation; Effects of L-arginine on cellular injury and NO production after low-volume anoxia and reoxygenation.

The importance of the immune system for cardiac repair following myocardial infarction is undeniable; however, the complex nature of immune cell behavior has limited the ability to develop effective therapeutics. This limitation highlights the need for a better understanding of the function of each immune cell population during the inflammatory and resolution phases of cardiac repair. The development of reliable therapies is further complicated by aging, which is associated with a decline in cell and organ function and the onset of cardiovascular and immunological diseases. Aging of the immune system has important consequences on heart function as both chronic cardiac inflammation and an impaired immune response to cardiac injury are observed in older individuals. Several studies have suggested that rejuvenating the aged immune system may be a valid therapeutic candidate to prevent or treat heart disease. Here, we review the basic patterns of immune cell behavior after myocardial infarction and discuss the autonomous and nonautonomous manners of hematopoietic stem cell and immune cell aging. Lastly, we identify prospective therapies that may rejuvenate the aged immune system to improve heart function such as anti-inflammatory and senolytic therapies, bone marrow transplant, niche remodeling and regulation of immune cell differentiation. [ABSTRACT FROM AUTHOR]

Background—Efficient cardiac function requires synchronous ventricular contraction. After myocardial infarction, the nonconductive nature of scar tissue contributes to ventricular dysfunction by electrically uncoupling viable cardiomyocytes in the infarct region. Injection of a conductive biomaterial polymer that restores impulse propagation could synchronize contraction and restore ventricular function by electrically connecting isolated cardiomyocytes to intact tissue, allowing them to contribute to global heart function. Methods and Results—We created a conductive polymer by grafting pyrrole to the clinically tested biomaterial chitosan to create a polypyrrole (PPy)-chitosan hydrogel. Cyclic voltammetry showed that PPy-chitosan had semiconductive properties lacking in chitosan alone. PPy-chitosan did not reduce cell attachment, metabolism, or proliferation in vitro. Neonatal rat cardiomyocytes plated on PPy-chitosan showed enhanced Ca2+ signal conduction in comparison with chitosan alone. PPy-chitosan plating also improved electric coupling between skeletal muscles placed 25 mm apart in comparison with chitosan alone, demonstrating that PPy-chitosan can electrically connect contracting cells at a distance. In rats, injection of PPy-chitosan 1 week after myocardial infarction decreased the QRS interval and increased the transverse activation velocity in comparison with saline or chitosan, suggesting improved electric conduction. Optical mapping showed increased activation in the border zone of PPy-chitosan-treated rats. Echocardiography and pressure-volume analysis showed improvement in load-dependent (ejection fraction, fractional shortening) and load-independent (preload recruitable stroke work) indices of heart function 8 weeks after injection. Conclusions—We synthesized a biocompatible conductive biomaterial (PPy-chitosan) that enhances biological conduction in vitro and in vivo. Injection of PPy-chitosan better maintained heart function after myocardial infarction than a nonconductive polymer. [ABSTRACT FROM AUTHOR]

Abstract: The goal of cardiac tissue engineering is to restore function to the damaged myocardium with regenerative constructs. Human embryonic stem cell–derived cardiomyocytes (hESC-CMs) can produce viable, contractile, three-dimensional grafts that function in vivo. We sought to enhance the viability and functional maturation of cardiac tissue constructs by cyclical stretch. hESC-CMs seeded onto gelatin-based scaffolds underwent cyclical stretching. Histological analysis demonstrated a greater proportion of cardiac troponin T–expressing cells in stretched than non-stretched constructs, and flow sorting demonstrated a higher proportion of cardiomyocytes. Ultrastructural assessment showed that cells in stretched constructs had a more mature phenotype, characterized by greater cell elongation, increased gap junction expression, and better contractile elements. Real-time PCR revealed enhanced mRNA expression of genes associated with cardiac maturation as well as genes encoding cardiac ion channels. Calcium imaging confirmed that stretched constructs contracted more frequently, with shorter calcium cycle duration. Epicardial implantation of constructs onto ischemic rat hearts demonstrated the feasibility of this platform, with enhanced survival and engraftment of transplanted cells in the stretched constructs. This uniaxial stretching system may serve as a platform for the production of cardiac tissue-engineered constructs for translational applications. [Copyright &y& Elsevier]

Abstract: Following a myocardial infarction (MI), fibroblasts differentiate to myofibroblasts, which possess some of the characteristics of smooth muscle cells (SMCs) and contribute to wound healing. Previous studies suggested that the miR-143/-145 cluster plays a critical role in SMC differentiation. Therefore, we determined whether miR-145 promoted differentiation of cardiac fibroblasts to myofibroblasts. Following coronary occlusion in mice, myocardial miR-145 expression was downregulated at 3days but was restored at 7days. In vitro studies showed that hypoxia also downregulated miR-145 in cardiac fibroblasts. The number of α-smooth muscle actin (α-SMA) positive cells in fibroblast cultures was employed to determine their transdifferentiation to cardiac myofibroblasts and was increased by 73.5% after transient transfection with miR-145. Ultrastructural analysis of α-SMA stress fibers revealed that ~95% of the α-SMA+ cells treated with miR-145 organized their actin-filament bundles with a specific orientation compared to only 15% in the scrambled control group. This orientation of the SMA bundles and their integration with the filamentous actin fibers of the cytoskeleton permit infarct wound contraction. Structural and functional studies showed that miR-145 induced a myofibroblast phenotype, and miR-145 also potentiated the production of mature collagen by myofibroblasts. Repression of KLF5, a target of miR-145, was validated by a chimeric luciferase construct tagged with the full-length 3′-UTR of KLF5. A dramatic decrease in KLF5 and a corresponding increase in myocardin expression were observed after transfecting cultured fibroblasts with miR-145. Similar results were found in vivo: the transient decrease in miR-145 expression 3days post-MI was associated with an increase in KLF5 and a decrease in myocardin. In addition, in vivo delivery of a miR-145 antagomir 1day prior to and 2 and 6days after MI decreased myofibroblast formation and increased scar size. The antagomir also reversed the suppressed expression of KLF5 protein in the scar region at day 7 after MI. In summary, we describe a novel association between miR-145 and fibroblast differentiation toward myofibroblasts. These observations provide a new approach to promote endogenous scar healing and contracture by stimulating the transdifferentiation of cardiac fibroblasts to myofibroblasts. [Copyright &y& Elsevier]

Cardiac cell therapies offer distinct and exciting advantages over current treatments to prevent postinfarction heart failure because they can reverse ventricular remodeling and improve function, but only if the implanted stem cells contribute biological functions and achieve prolonged engraftment within the hostile environment of the damaged heart. Unfortunately, function is diminished in autologous stem cells isolated from older patients and those with comorbidities, and so clinical trials testing the implantation of healthy, allogeneic bone marrow–derived stromal cells (MSCs) isolated from young donors are currently underway. MSCs are unique because, in addition to exerting paracrine effects that restore blood flow and recruit endogenous stem cells to the infarct, they exhibit immune-modulating properties in culture that—if retained after allogeneic implantation—imply the cells may escape immune recognition within the heart. At present, the scope of MSC immune modulation after implantation is unclear. (J Card Surg 2010;25:359-366) [ABSTRACT FROM AUTHOR]

Introduction: Cardiovascular disease is the leading cause of death worldwide and its prevalence increases with age. Replacing the bone marrow (BM) stem cells of aged mice with young BM stem cell antigen-1+ (Y-Sca-1+) cells rejuvenates the response of the aged heart to injury. Autophagy, a critical cellular recycling process, decreases with age. We hypothesized that the high rate of autophagy in Y-Sca-1+ BM cells contributes to the rejuvenation of the aged heart and improves tissue repair. Methods: In vitro: Young (Y) and old (O) Sca-1+ BM cells were isolated from Y and O mice and cultured in normoxia and hypoxia conditions to examine the rate of autophagy. To determine the effect of secretory factors, conditioned medium (CM) was produced from Y-Sca-1+ and O-Sca-1+ cells and used to culture old primary cardiac fibroblasts (O-Fs). In vivo: Old mice were lethally irradiated and reconstituted with Y-Sca-1+ or O-Sca-1+ BM cells from GFP mice to generate YP and OP chimeras, respectively. After 3 months, mice were sacrificed and hearts collected for molecular and histological analyses. Results: In vitro: Cultured Y-Sca-1+ cells exhibited a higher rate of autophagy and were more responsive to hypoxia-induced autophagy than O-Sca-1+ cells. Thus, Y-Sca-1+ cells may pre-condition and rejuvenate the aged heat. In support of this theory, factors secreted from Y-Sca-1+, but not O-Sca-1+ BM cells, enhanced several functions of O-Fs, including autophagy, cell migration, proliferation and trans-differentiation. This phenotype was not observed after boiling the CM, suggesting that rejuvenation is mediated by a secreted factor. In vivo: We found that reconstitution of aged mice with Y-Sca-1+ BM cells improved autophagy of the aged heart. The autophagy rate was increased in YP relative to OP hearts as demonstrated by an increase in the LC3b II/I ratio. Immunostaining of the reconstituted heart showed that the number of LC3b+ cells was 3-fold higher in YP than OP hearts. These LC3b+ cells were both donor and recepiant cells. Conclusions: The increase in autophagy rate in the YP heart suggests that autophagy is one of the mechanisms involved in rejuvenation of the aged heart after BM reconstitution. Y-Sca-1+ BM cells secret factors that improve the autophagy of recepient cardiac cells. [ABSTRACT FROM AUTHOR]

Introduction: Clinical trials of cell therapy for post-injury conditioning of the myocardium have shown only modest improvements, perhaps because the majority of donor cells die in the harsh post-ischemic environment. Hypothesis: We hypothesize that pre-conditioning the myocardium by delivering human CD34+ (hCD34+) cells through bone marrow (BM) engraftment may prevent adverse remodeling and heart failure post myocardial infarct (MI). BM engraftment may be especially important in elderly patients because of their reduced stem cell function. Methods: BM aspirates from 40 patients undergoing open-heart surgery were collected. Mononuclear cells were isolated and stem cell subpopulations were quantified using flow cytometry and magnetically sorted. Function of hCD34+ cells was analyzed by the colony forming unit (CFU) assay and their ability to reconstitute irradiated immune-deficient NSG mice. Engraftment was quantified as hCD45 chimerism by flow cytometry. To evaluate the benefits of hCD34+ cells in cardiac repair, coronary artery ligation was performed 12 weeks post-reconstitution. Left ventricular (LV) function and histology were assessed by echocardiography and immunohistochemistry. Results: Older patients (>60 years old, n=24) had lower numbers (p=0.0008) and reduced function (p<0.0001) of hCD34+ cells. We found a significant difference in the homing ability of hCD34+ (n=7) vs. hCD34- (n=6) cells to the bone marrow, blood, spleen and heart (p<0.05, all groups) 12 weeks post-reconstitution. hCD34+ mice engrafted with young patients' cells had higher reconstitution in blood, spleen and heart (p<0.01, all groups). At 4 weeks post-ligation, hCD34+ mice (n=8) had improved LV function, with greater fractional shortening (p=0.003), reduced LV dilation (p=0.02) and reduced scar size (p=0.0002). Immunohistochemistry of the peri-infarct region of CD34+ mice showed increased hCD34-derived vessels, indicating their role in infarct healing. Conclusions: hCD34+ cells have reduced frequency and function in older patients, but engraft and protect the myocardium from functional decline post-MI. Cardiac pre-conditioning by restoration of aged hCD34+ cells is a promising avenue for preventing cardiac dysfunction post-MI. [ABSTRACT FROM AUTHOR]