Abstract-Heart failure remains a leading cause of morbidity and mortality. The cellular mechanism underlying the development of cardiac dysfunction is a decrease in the number of viable cardiomyocytes. Recent observations have suggested that the adult heart may contain a progenitor cell population. Side population (SP) cells, characterized by a distinct Hoechst dye efflux pattern, have been shown to exist in multiple tissues and are capable of tissue-specific differentiation. In this report, we confirm the existence of a cardiac SP cell population, immunophenotypically distinct from bone marrow SP cells. Moreover, we demonstrate that among cardiac SP cells, the greatest potential for cardiomyogenic differentiation is restricted to cells negative for CD31 expression and positive for stem cell antigen 1 (Sca1) expression (CD31 Ϫ /Sca1 ϩ ). Furthermore, we determine that CD31 Ϫ /Sca1 ϩ cardiac SP cells are capable of both biochemical and functional cardiomyogenic differentiation into mature cardiomyocytes, with expression of cardiomyocyte-specific transcription factors and contractile proteins, as well as stimulated cellular contraction and intracellular calcium transients indistinguishable from adult cardiomyocytes. We also determine the necessity of cell-extrinsic signaling through coupling, although not fusion, with adult cardiomyocytes in regulating cardiomyogenic differentiation of cardiac SP cells. We, therefore, conclude that CD31 Ϫ /Sca1 ϩ cardiac SP cells represent a distinct cardiac progenitor cell population, capable of cardiomyogenic differentiation into mature cardiomyocytes through a process mediated by cellular coupling with adult cardiomyocytes. (Circ Res. 2005;97:52-61.)

Abstract

Cardiac hypertrophy is a common response to injury and hemodynamic stress and an important harbinger of heart failure and death. Herein, we identify the Kruppel-like factor 15 (KLF15) as an inhibitor of cardiac hypertrophy. Myocardial expression of KLF15 is reduced in rodent models of hypertrophy and in biopsy samples from patients with pressure-overload induced by chronic valvular aortic stenosis. Overexpression of KLF15 in neonatal rat ventricular cardiomyocytes inhibits cell size, protein synthesis and hypertrophic gene expression. KLF15-null mice are viable but, in response to pressure overload, develop an eccentric form of cardiac hypertrophy characterized by increased heart weight, exaggerated expression of hypertrophic genes, left ventricular cavity dilatation with increased myocyte size, and reduced left ventricular systolic function. Mechanistically, a combination of promoter analyses and gel-shift studies suggest that KLF15 can inhibit GATA4 and myocyte enhancer factor 2 function. These studies identify KLF15 as part of a heretofore unrecognized pathway regulating the cardiac response to hemodynamic stress.

Antibodies that stimulate the immune system by targeting inhibitory T cell receptors were successfully introduced into oncological practice and are capable to overcome tumor-induced immune evasion. In particular, targeting of the inhibitory receptors CTLA-4 and PD-1 or its ligand PD-L1 have been shown to be beneficial for patients with melanoma, renal cell cancer, non-small cell lung cancer and a growing list of other cancers with impressive response rates. Here, we report a severe, potentially life-threatening side effect of anti-PD-1 immunotherapy with pembrolizumab, which has not been previously described in the literature. A 73-year-old woman with metastatic uveal melanoma treated with pembrolizumab in third line developed severe heart failure due to pembrolizumab-mediated autoimmune myocarditis. Echocardiographic studies revealed a severely impaired left ventricular function with dyssynchrony. All tests for cardiotropic viruses were negative and histological analysis of a myocardial biopsy showed lymphocytic infiltration with a predominance of CD8 positive cells and a reduction of FOXP3 positive regulatory T cells. After initiation of corticosteroids and guideline-conform heart failure therapy, the symptoms rapidly improved and the left ventricular function recovered. While autoimmune myocarditis is a documented side effect of other checkpoint inhibitors, as for example ipilimumab and in one case with anti-PD-L1 antibody, it is not described for anti-PD-1-antibodies like pembrolizumab or nivolumab. As the FDA recently approved both pembrolizumab and nivolumab for melanoma progressing after anti-CTLA-4 treatment with ipilimumab, more patients will soon receive anti-PD-1 therapy. Thus, it is important to be aware of such rare, but severe immune-related adverse events.Electronic supplementary materialThe online version of this article (doi:10.1186/s40425-015-0057-1) contains supplementary material, which is available to authorized users.

Tissue-specific progenitor cells contribute to local cellular regeneration and maintain organ function. Recently, we have determined that cardiac side-population (CSP) cells represent a distinct cardiac progenitor cell population, capable of in vitro differentiation into functional cardiomyocytes. The response of endogenous CSP to myocardial injury, however, and the cellular mechanisms that maintain this cardiac progenitor cell pool in vivo remain unknown. In this report we demonstrate that local progenitor cell proliferation maintains CSP under physiologic conditions, with little contribution from extracardiac stem cell sources. Following myocardial infarction in adult mice, however, CSP cells are acutely depleted, both within the infarct and noninfarct areas. CSP pools are subsequently reconstituted to baseline levels within 7 days after myocardial infarction, through both proliferation of resident CSP cells, as well as through homing of bone marrow-derived stem cells (BMC) to specific areas of myocardial injury and immunophenotypic conversion of BMC to adopt a CSP phenotype. We, therefore, conclude that following myocardial injury, cardiac progenitor cell populations are acutely depleted and are reconstituted to normal levels by both self-proliferation and selective homing of BMC. Understanding and enhancing such processes hold enormous potential for therapeutic myocardial regeneration.T issue-specific progenitor cell populations maintain the regenerative capacity of terminally differentiated organs, both under basal conditions and following local tissue injury. Side population (SP) cells, characterized by their intrinsic capacity to efflux Hoechst dye through ATP-binding cassette transporters, contribute to the long-term regenerative potential of hematopoietic and extrahematopoietic tissues. 1 Recently, we have demonstrated that cardiac SP (CSP) cells, immunophenotypically distinct from bone marrow (BM)-derived stem cells (BMC), are present in the adult heart and are capable of both biochemical and functional cardiomyogenic differentiation into mature cardiomyocytes, thereby identifying CSP as a distinct cardiac progenitor cell population. 2 Supplementation of cardiac progenitor cell pools after myocardial infarction (MI) with exogenous cells has been shown to improve ventricular function by regenerating myocardium and cardiac vasculature. [3][4][5] The response of endogenous CSP cells to myocardial injury, however, and the cellular mechanisms that maintain this endogenous cardiac progenitor cell pool under basal and after injury conditions remain unknown. We, therefore, serially assessed CSP pools in hearts following MI and determined the role of selfproliferation and BMC in reconstituting cardiac progenitor pools. Methods and MaterialsCSP were isolated from mouse hearts as described previously. 2 MI was performed in mice via permanent coronary ligation. 6 BM transplantation was performed in lethally irradiated mice using marrow isolated from C57bl/6-Tg(ACTbEGFP) mice. 7 All animals were obtained from The ...

Although the 'intermediate' clinical profile of HFmrEF between HFrEF and HFpEF would support the conclusion that HFmrEF is a distinct clinical entity, we hypothesize that HFmrEF has to be categorized as HFrEF because of the high prevalence of coronary artery disease and the similar benefit of NT-proBNP-guided therapy in HFrEF and HFmrEF, in contrast to HFpEF.

Abstract-Recently, the side population (SP) phenotype has been introduced as a reliable marker to identify subpopulations of cells with stem/progenitor cell properties in various tissues. We and others have identified SP cells from postmitotic tissues, including adult myocardium, in which they have been suggested to contribute to cellular regeneration following injury. SP cells are identified and characterized by a unique efflux of Hoechst 33342 dye. Key Words: Abcg2 Ⅲ Mdr1 Ⅲ progenitor cells Ⅲ proliferation Ⅲ SP cells R ecently, the side population (SP) phenotype has been introduced as a reliable marker to identify subpopulations of cells with stem/progenitor cell properties in various tissues including the heart. 1 On the molecular level, the SP phenotype is linked to the presence of ATP-binding cassette (ABC) transporters with the ability to efficiently efflux the DNA binding dye Hoechst 33342. 2 This ABC transporterdependent Hoechst efflux phenomenon confers the characteristic fluorescent-activated cell sorting (FACS) profile of SP cells as a Hoechst-low "side population" located to the periphery of the Hoechst-high main population. 2 Among the various members of the ABC transporter superfamily, Abcg2 (also referred to as breast cancer resistance protein 1 [Bcrp1]) and Mdr1 (also referred to as P-glycoprotein [p-gp] or Abcb1) have been shown to efficiently efflux Hoechst 33342 and thereby confer the SP phenotype. 3 Although both transporters are highly expressed in bone marrow (BM)SP cells, studies performed in mice with targeted disruption of the Mdr1a and Mdr1b genes, the murine homologs of the human Abcb1/Mdr1 gene, demonstrated that Abcg2 is the sole molecular determinant of the SP phenotype in hematopoietic stem cells. 4 Moreover, Abcg2 expression is conserved in SP cells from a wide range of tissues including blood, gonad, lung, skeletal muscle and the retina, suggesting an important role of Abcg2 in stem cells. 4 -7 We and others have characterized SP cells isolated from adult myocardium. 8 -11 These cardiac (c)SP cells are phenotypically distinct from BMSP cells, in that they are not hematopoietic but exhibit the potential to differentiate into functional cardiomyocytes. 10 As in SP cells from the bone marrow, Abcg2 is expressed in SP cells from the heart. 9 The contribution of Abcg2 to the cSP phenotype and its biological significance in cSP progenitor cells, however, remain unknown. In this study, we find that the contribution of Abcg2 to the SP phenotype in the heart exists in an age-dependent manner, with Abcg2 as the molecular determinant of the SP phenotype in the neonatal heart and Mdr1 as the basis for the SP phenotype in the adult heart. In addition, we demonstrate Original

Myocardial failure is associated with increased oxidative stress and abnormal excitation-contraction coupling characterized by depletion of sarcoplasmic reticulum (SR) Ca 2+ -stores and a reduction in Ca 2+ -transient amplitude. Little is known about the mechanisms whereby oxidative stress affects Ca 2+ -handling and contractile function; however, reactive thiols may be involved. We used an in vitro cardiomyocyte system to test the hypothesis that short-term oxidative stress induces SR Ca 2+ -depletion via redox-mediated regulation of sarco-endoplasmic reticulum Ca 2+ -ATPase (SERCA) and the sodium-Ca 2+ -exchanger (NCX) and that this is associated with thiol oxidation. Adult rat ventricular myocytes paced at 5 Hz were superfused with H 2 O 2 (100 μM, 15 min). H 2 O 2 caused a progressive decrease in cell shortening followed by diastolic arrest, which was associated with decreases in SR Ca 2+ -content, systolic [Ca 2+ ] i and Ca 2+ -transient amplitude, but no change in diastolic [Ca 2+ ] i . H 2 O 2 caused reciprocal effects on the activities of SERCA (decreased) and NCX (increased). Pretreatment with the NCX inhibitor KB-R7943 prior to H 2 O 2 increased diastolic [Ca 2+ ] i , and mimicked the effect of SERCA inhibition with thapsigargin. These functional effects were associated with oxidative modification of thiols on both SERCA and NCX. In conclusion, redox-mediated SR Ca 2+ -depletion involves reciprocal regulation of SERCA and NCX, possibly via direct oxidative modification of both proteins.