Myocyte Loss Research Articles

Hyperglycemia induces defective Ca2+ homeostasis in cardiomyocytes.

We have investigated the effects of hyperglycemia on cardiomyocyte physiology and ventricular function. Our results indicate that defective Ca2+ handling is a critical component of the progressive deterioration of cardiac performance of the diabetic heart.

Unilateral perivertebral fibrosis associated with lordosis, kyphosis and scoliosis (LKS) in farmed Chinook salmon in New Zealand.

Vertebral column lordosis, kyphosis and scoliosis (LKS) can result in downgrading of farmed Chinook salmon Oncorhynchus tshawytscha in New Zealand. No cause of LKS has been identified. Radiography and histology were used to quantify LKS and perivertebral fibrosis in 27 fish with LKS visible at harvest and 30 visually normal fish from 3 New Zealand farms. Radiographic LKS was present in all 27 fish with LKS and in 18 of 30 fish without visible LKS. Quantification of the radiographic severity revealed significantly higher radiographic severity scores in fish with visible LKS (mean ± SD = 5.89 ± 2.41) than in fish with no visible, but radiographic LKS (1.44 ± 0.86, p < 0.001). The most frequent histological finding was unilateral perivertebral fibrosis that often extended into the horizontal septum and adjacent myomeres resulting in separation or loss of myocytes. Fibrosis was visible in all fish with LKS and in 12 of 30 fish without visible LKS. Fibrosis scores were higher in fish with visible LKS (3.32 ± 1.71) than in fish without visible LKS (0.35 ± 0.57, p < 0.001). The radiographic LKS severity scores were significantly correlated to the fibrosis scores (R2 = 0.59 p < 0.001) in the fish. Histology of other tissues revealed multifocal inflammation within muscle, peripheral connective tissues and myocardium which were considered most likely incidental in these fish. In this study, LKS was consistently and significantly associated with perivertebral fibrosis, suggesting that perivertebral fibrosis is an important process in the development of LKS. Further research to determine the cause of the fibrosis is required.

The modest outcome of clinical trials with bone marrow cells for myocardial repair: is the autologous source of cells the prime culprit?

Re-entry of cardiomyocytes into cell cycle and participation of the resident cardiac stem cells (CSCs) as part of the intrinsic repair process have been evidenced in the ischemic heart (1-3). Nevertheless, the inadequacy of these mechanisms of intrinsic repair necessitates outside intervention especially when the damage inflicted is massive in terms of the loss of functional myocytes. The situation is further aggravated by the advanced age of the patients with ischemic heart disease, and the complexity of the disease process which renders the tissue environment in the diseased heart more intimidating for the transplanted cells besides impacting the stemness characteristics and reparability of the intrinsically available stem cells (4).

Improving Delivery of a Biomaterial Payload in Myocardial Infarction

Survival after an acute coronary syndrome has greatly improved because of advances in thrombolytic therapy, stabilization of the culprit lesion such as with coronary stents, and the use of pharmacological approaches to prevent recurrent thrombotic and arrhythmic events. Despite these approaches, however, myocardial injury culminating in myocardial infarction (MI) continues to be an all too common sequela. The MI region is a highly heterogeneous structure that contains several cell types, such as residual cardiac myocytes, inflammatory cells of different lineages, proliferating and transdifferentiating fibroblasts, and extracellular matrix proteins and signaling molecules. Although canonical thought was that the MI region was a rather static, fibrotic structure, it is now clear that this is a dynamic and ever-evolving entity with changes in geometry, biophysical properties, and cellular/extracellular composition. The time course and extent of these post-MI processes have been generically termed post-MI remodeling and is recognized as a milestone event for the initiation and progression to left ventricular (LV) dilation and dysfunction. One ubiquitous feature of post-MI remodeling is a progressive thinning and dyskinesia of the MI region, which is because of the summation of several factors: a loss of cardiac myocytes and thus functional myocardium, continuous turnover and instability of the extracellular matrix, and significant shifts in stress–strain patterns within this region throughout the cardiac cycle. Specifically, significant heterogeneity arises in the stress–strain patterns both within and surrounding the MI region and in turn can contribute to a feed-forward process of continuous activation of bioactive molecules and extracellular matrix instability. These biomechanical changes result in continued MI thinning and recruitment of border zone myocardium into the MI region, which is defined as MI expansion. As the MI expansion process continues in an inexorable fashion, the degree of heterogeneity in stress–strain relations are exacerbated, bulging of the MI region becomes evident by …

Exogenous spermine inhibits hypoxia/ischemia-induced myocardial apoptosis via regulation of mitochondrial permeability transition pore and associated pathways.

Myocardial infarction (MI) is associated with a high mortality rate, which is attributed to the effects of myocyte loss that occurs as a result of ischemia-induced cell death. Very few therapies can effectively prevent or delay the effects of ischemia. Polyamines (PAs) are polycations required for cell growth and division, and their use may prevent cell loss. The aim of this study was to investigate the relationship between hypoxia/ischemia (H/I)-induced cell apoptosis and PA metabolism and to investigate the ability of spermine to limit H/I injury in cardiomyocytes by blocking the mitochondrial apoptotic pathway. Neonatal rat cardiomyocytes were placed under hypoxic conditions for 24 h after being subjected to 5 μM of spermine as a pretreatment therapy. H/I induced PA catabolism, which was indicated by a 1.3-fold up-regulation of spermidine/spermine N(1)-acetyltransferase expression. Exogenous spermine significantly reduced H/I-induced cell death rate (60 ± 2 to 36 ± 2%) and apoptosis rate (42 ± 2 to 21 ± 2%); it also attenuated lactate dehyodrogenase and creatine kinase leakage (440 ± 13 and 336 ± 16 U/L to 275 ± 15 and 235 ± 13 U/L). Furthermore, it decreases calcium overload (3.8 ± 0.2 to 2.2 ± 0.1 a.u.). Moreover, spermine pretreatment remarkably decreased cytochrome c release from the mitochondria to the cytosol, lowering the expression of cleaved caspase-3 and -9. With spermine pretreatment, there was an increase in Bcl-2 levels and phosphorylation of ERK1/2, phosphoinositide 3-kinase, Akt, and GSK-3β, preserving mitochondrial membrane potential and inducing an mitochondrial permeability transition pore opening. In conclusion, H/I decreased endogenous spermine concentrations in cardiomyocytes, which ultimately induced apoptosis. The addition of exogenous spermine effectively prevented myocyte cell death.

Exercise, Nrf2 and Antioxidant Signaling in Cardiac Aging.

Aging is represented by a progressive decline in cellular functions. The age-related deformities in cardiac behaviors are the loss of cardiac myocytes through apoptosis or programmed cell death. Oxidative stress (OS) and its deleterious consequence contribute to age-related mechanical remodeling, reduced regenerative capacity, and apoptosis in cardiac tissue. The pathogenesis of OS in the elderly can predispose the heart to other cardiac complications such as atherosclerosis, hypertension, ischemic heart disease, cardiac myopathy, and so on. At the molecular level, oxidant-induced activation of Nrf2 (Nuclear erythroid-2-p45-related factor-2), a transcription factor, regulates several genes containing AREs (Antioxidant Response Element) and bring the respective translates to counteract the reactive radicals and establish homeostasis. Myriad of Nrf2 gene knockout studies in various organs such as lung, liver, kidney, brain, etc. have shown that dysregulation of Nrf2 severely affects the oxidant/ROS sensitivity and predispose the system to several pathological changes with aberrant cellular lesions. On the other hand, its gain of function chemical interventions exhibited oxidant stress resistance and cytoprotection. However, thus far, only a few investigations have shown the potential role of Nrf2 and its non-pharmacological induction in cardiac aging. Therefore, here we review the involvement of Nrf2 signaling along with its responses and ramifications on the cascade of OS under acute exercise stress (AES), moderate exercise training (MET), and endurance exercise stress (EES) conditions in the aging heart.

Cardiac MRI: a Translational Imaging Tool for Characterizing Anthracycline-Induced Myocardial Remodeling.

Cardiovascular side effects of cancer therapeutics are the leading causes of morbidity and mortality in cancer survivors. Anthracyclines (AC) serve as the backbone of many anti-cancer treatment strategies, but dose-dependent myocardial injury limits their use. Cumulative AC exposure can disrupt the dynamic equilibrium of the myocardial microarchitecture while repeated injury and repair leads to myocyte loss, interstitial myocardial fibrosis, and impaired contractility. Although children are assumed to have greater myocardial plasticity, AC exposure at a younger age portends worse prognosis. In older patients, there is lower overall survival once they develop cardiovascular disease. Because aberrations in the myocardial architecture predispose the heart to a decline in function, early detection with sensitive imaging tools is crucial and the implications for resource utilization are substantial. As a comprehensive imaging modality, cardiac magnetic resonance (CMR) imaging is able to go beyond quantification of ejection fraction and myocardial deformation to characterize adaptive microstructural and microvascular changes that are important to myocardial tissue health. Herein, we describe CMR as an established translational imaging tool that can be used clinically to characterize AC-associated myocardial remodeling.

Necroptotic cell death in failing heart: relevance and proposed mechanisms.

As cardiomyocytes have a limited capability for proliferation, renewal, and repair, the loss of heart cells followed by replacement with fibrous tissue is considered to result in the development of ventricular dysfunction and progression to heart failure (HF). The loss of cardiac myocytes in HF has been traditionally believed to occur mainly due to programmed apoptosis or unregulated necrosis. While extensive research work is being carried out to define the exact significance and contribution of both these cell death modalities in the development of HF, recent knowledge has indicated the existence and importance of a different form of cell death called necroptosis in the failing heart. This new cell damaging process, resembling some of the morphological features of passive necrosis as well as maladaptive autophagy, is a programmed process and is orchestrated by a complex set of proteins involving receptor-interacting protein kinase 1 and 3 (RIP1, RIP3) and mixed lineage kinase domain-like protein (MLKL). Activation of the RIP1-RIP3-MLKL signaling pathway leads to disruption of cation homeostasis, plasma membrane rupture, and finally cell death. It seems likely that inhibition of any site in this pathway may prove as an effective pharmacological intervention for preventing the necroptotic cell death in the failing heart. This review is intended to describe general aspects of the signaling pathway associated with necroptosis, to describe its relationship with cardiac dysfunction in some models of cardiac injury and discuss its potential relevance in various types of HF with respect to the underlying pathologic mechanisms.

Bioengineered epicardial patch regenerates the adult mammalian heart

Event Abstract Back to Event Bioengineered epicardial patch regenerates the adult mammalian heart Vahid Serpooshan1, Ke Wei2, 3, Mark Mercola2, 3 and Pilar Ruiz-Lozano1 1 Stanford University School of Medicine, Stanford Cardiovascular Institute and Department of Pediatrics, United States 2 University of California, San Diego, Department of Bioengineering, United States 3 Sanford-Burnham-Prebys Medical Discovery Institute, United States Adult mammals, unlike lower vertebrates, cannot effectively regenerate heart tissue, and myocyte loss following myocardial infarction (MI) is an important cause of human heart failure. Therefore, the elucidation of endogenous factors that control regeneration is of major scientific and therapeutic importance. Recent evidence suggests that the epicardium promotes myocardial regeneration; however, the mechanism remains largely unknown. In this study, we discovered that conditioned media from epicardial-like cultures enhance cardiomyogenesis in vitro[1]. We subsequently demonstrated that epicardial secreted factors embedded in a biomimetic type I collagen scaffold activate cardiomyocyte proliferation after ischemic heart injury (MI) in the mouse model (Figure1)[1]. An acellular type I collagen scaffold, endowed with specific physiomechanical properties approaching those of the embryonic epicardium, was generated utilizing plastic compression technique and used as cardiac patch (Figure 1a,b)[1]-[3]. Mass Spectrometry analysis identified Follistatin-like 1 (FSTL1) protein as a main component of the epicardial-derived cardiogenic activity. Fstl1 is expressed in the adult epicardium but declines strikingly following MI[1]. Application of FSTL1 to the epicardial surface of the heart by a compressed collagen patch recapitulated the activity of epicardial-conditioned media and restored cardiac function in the mouse model (Figure 2). The engineered patch-FSTL1 treatment diminished pathological remodeling (ventricular dilation and fibrosis), restored vascularization, and induced cell cycle entry of pre-existing αMHC+ cells after MI, consequently improving long-term cardiac function (Figure 2)[1]. The epicardial patch-FSTL1 function was also evaluated in the swine model of ischemia-reperfusion (I/R) injury[1]. As determined by delayed enhancement MRI, application of the patch to the epicardium over the injured tissue at week 1 post-I/R stimulated recovery of contractile function (%EF) in 2 weeks. The recovery remained stable for an additional two weeks and was in contrast to the steady decline seen without treatment or following treatment with patch alone (Figure 3b). FSTL1-treated pigs demonstrated the least scar size of all treatments (green lines in Figure 3c-d). Immunostaining 4 weeks after patch implantation demonstrated cardiomyocytes in the border zone and ischemic area with evident EdU labelling and midbody-localized aurora B kinase (indicative of cytokinesis) (Figure 3e-j)[1].

Assessment of Experimental Permanent Coronary Artery Ligation Using Echocardiography and Invasive Real-Time Pressure-Volume (PV): A Practical Tips to Evaluate Rat Hemodynamics

Background: Rat permanent coronary artery ligation is surgical model mimicking coronary artery ischemia and myocardial infarct (MI) injury, both sequels of coronary artery disease (CAD). The aim of this publication is to provide comprehensive, detailed description of rat load-dependent and independent hemodynamic assessment at baseline and at 28 days post-myocardial ischemia and remodeling. Materials and Methods: the detailed depiction of rat-MI model is followed by a thorough assessment of hemodynamics by two-dimensional (2D) echocardiography and invasive pressure-volume (PV) catheterization. Results: Quantification of post-MI using 2D M-mode showed a significant increase in end-systolic and end-diastolic dimensions with a decrease of fractional shortening. PV load-dependent hemodynamics at 28 days showed a rightward shift of PV loop on the volume axis, characterized by an increase of LVEDV; (319±73 vs. 215±79 µl; P<0.001) and the LVESV (157±39 vs. 57±28 µl; P<0.001). Both dpdtmax and ESP were significantly influenced by decreasing the number of myocytes, leading to a significant decrease of dpdtmax (5786±1443 vs. 9609±4126 mmHg; P<0.01) and ESP (91.5±12 vs. 108.2±13 mmHg; P<0.001). Loss of contractile myocytes had an effect on the cardiac output (CO) and ejection fraction (EF); (49±12 vs. 68±3.5 ml/min; P<0.05) and (50±8.5 vs. 76±4.9 % P<0.001). Diastolic dysfunction had a major influence on LV lusitropy at day 28 post-MI characterized by prolonged LV filling at higher LVEDP (9.1±2.9 vs. 5.18±2.5 % P<0.01), higher Tau values, time to peak filling and dpdt min (-4850±1062 vs. 5876±1443 mmHg; P<0.001). Using histopathology, calculated HW/BW ratio (g/mg) (3.1±0.22 vs. 3.82±0.39; P<0.001), reflected hypertrophy of post-remodeled myocardium. Conclusion: Heart failure (HF) post-permanent coronary artery ligation influences both systolic and diastolic hemodynamics. Comprehensive assessment of modeled HF using load-dependent and independent indices enables its clinical translation.

EFFECT OF L-CARNITINE ON OXYDATIVE STRESS AT CARDIOVASCULAR DISEASES

L-carnitine (LC) and its esters do not only participate in the metabolic exchange of the cardiac myocytes (they are responsible for transfer of long-chain fatty acids from the cytoplasm in mytochondria) but have other vital types of activity. They have antioxidant and anti- nflammatory effect at various levels. LC is proved to be a direct antioxidant and to remove already formed oxygen radicals as well as to suppress generation of oxygen radicals by cytoplasmic and membrane-connected ferments as a result of formation of complexes with iron and copper ions in their active centers. The leading reason of LC protective effects is change of activity of redox-sensitive signal paths targeted at nuclear receptors (they are factors of NF-kB; PPARs; Nrf2, AP1 etc. gene transcription). These factors control activity of a group of genes responsible for survival and stability of the cardiovascular system to various types of stress, including oxidative stress, and prevent loss of cardiac myocytes and endothelial cells at ischemia/reperfusion and other hypoxia-conditioned pathologic states. The review considers a number of mechanisms participating in protective effects of LC and its derivatives in the heart and other oxygen-dependent organs and tissues.

РЕГУЛИРОВАНИЕ ФУНКЦИОНАЛЬНО-ТЕХНОЛОГИЧЕСКИХ СВОЙСТВ МЯСНЫХ СИСТЕМ С ИСПОЛЬЗОВАНИЕМ ФЕРМЕНТАТИВНЫХ КОМПЛЕКСОВ

Use of nonstandard raw meat involves the use of technological additives and components with proteolytic activity to stabilize the physical and mechanical properties of raw meat. Fermentive complexes have significant impact on the functional characteristics and allow adjusting water binding capacity and rheological properties of meat systems. The results of the studies have found that the addition of fermentive complexes in the recipes of restructured products allows changing the functional and technological, structural and mechanical properties of meat systems; provides high yield, optimum organoleptic characteristics of the finished product, and its high nutritive value. In the analysis of the structural and mechanical properties the effectiveness of the fermentive complexes in the formation of a high density of the meat has been proven: the highest shear stress is typical for samples containing fermentive complex and is 1.5–1.7 times higher than the control ones. Histological examination of samples of the meat emulsion a high discomplexion of muscle fibers is revealed; with the loss of cross striation homogenization myocytes and fragmentation of muscle fibers prototypes is increased by 21,9–104%. In samples of sausages made with the help of fermentive complexes an increase in the mass fraction of protein, B vitamins, and energy value compared to the classical recipe was revealed. Fermentive complexes preventing the loss of low molecular weight substances and causing intense proteolysis biopolymers provide the formation of flavor and aroma of the finished product. Meat developed by innovative technology is characterized by uniform color, smooth surface, rich meat taste, and aroma. As a result of scientific studies the possibility of the use of substandard raw materials in the production of meat products by correcting the functional and technological characteristics of meat systems through fermentive complexes was proved.

Disturbed integrin expression in the vascular media in CADASIL.

CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy) is an inherited angiopathy characterized by degeneration and loss of vascular smooth muscle cells (VSMCs) of still unknown pathomechanism. Many functions of VSMCs, such as adhesion, apoptosis, contraction, differentiation, migration, and proliferation are determined by integrins - surface adhesion receptors involved in binding and interactions between cells and extracellular matrix (ECM). Since integrins play such an important role in VSMCs biology, disturbances in their expression may influence myocytes behavior and fate in CADASIL. In this study, we focused on the most important compounds of VSMCs integrins: subunits α₄, β1, and β₃ in an attempt to elucidate their immune expression in the arterial media of CADASIL patients. The immunohistochemistry revealed adecreased expression of integrin β1 subunit (p < 0.001) but similar to the control expression of integrin subunits α₄ and β₃. Decreased β1 immunoreactivity was observed in capillary vessels, arterioles, and small arteries. The abnormal immune expression of integrin β1 subunit was found even in microvessels without microscopically noted degenerative changes, which suggests that this is an early phenomenon in CADASIL. Since integrin β1 subunit is acompound of 10 heterodimer integrin receptors, its disturbed expression may significantly influence VSMCs biology leading to myocytes degeneration and loss via anoikis - atype of apoptotic cell death due to loss or inappropriate cell adhesion to ECM.

Molecular and structural transition mechanisms in long-term volume overload.

AimWe have previously reported that early phase (1 week) of experimental volume overload (VO) has an adaptive phenotype while wall stress‐matched pressure overload (PO) is maladaptive. Here we investigate the transition from adaptation to heart failure (HF) in long‐term VO.Methods and results FVB/N wild‐type mice were subjected to VO induced by aortocaval shunt, and were followed by serial echocardiography until in vivo left ventricular ejection fraction was below <50% (135 ± 35 days). Heart failure was evident from increased lung and liver weight and increased mortality compared with sham. Maladaptive remodelling resulted in significantly reduced sarcomeric titin phosphorylation (causing increased sarcomeric stiffness), whereas interstitial fibrosis was not increased. This was paralleled by re‐expression of the fetal gene program, activation of calcium/calmodulin‐dependent protein kinase II (CaMKII), decreased protein kinase B (Akt) phosphorylation, high oxidative stress, and increased apoptosis. Consistently, development of HF and mortality were significantly aggravated in Akt‐deficient mice.ConclusionTransition to HF in VO is associated with decreased Akt and increased CaMKII signalling pathways together with increased oxidative stress and apoptosis. Lack of interstitial fibrosis together with sarcomeric titin hypophosphorylation indicates an increased stiffness at the sarcomeric but not matrix level in VO‐induced HF (in contrast to PO). Transition to HF may result from myocyte loss and myocyte dysfunction owing to increased stiffness.

Abstract 11714: Allogeneic Cardiosphere-Derived Cells Administered at the Time of Reperfusion Only Improve Ejection Fraction When Infused to Ischemic as well as Remote Myocardial Regions in Swine With Acute Myocardial Infarction

Introduction: While intracoronary infusion of cardiosphere-derived cells (CDCs) to myocardial infarcts using the stop-flow technique reduces scar volume, ejection fraction (EF) remains unchanged. Hypothesis: We hypothesized that this may relate to ineffective treatment of myocyte loss in normally-perfused remote regions which could negatively impact EF. We therefore compared CDC infusion to the entire heart with conventional regional infusion to the infarct-related artery. Methods: Swine (n=24) chronically treated with cyclosporine (100 mg/day) underwent a 1-hour LAD occlusion and EF decreased from 60 ± 2% at baseline to 46 ± 2% after reperfusion (p&lt;0.05 vs. baseline). We infused either 10x10^6 intracoronary allogeneic CDCs via stop-flow infusion (regional LAD infusion), 20x10^6 intracoronary allogeneic CDCs to the entire heart under flow (global infusion) or vehicle 30 minutes after reperfusion. Four weeks later we assessed left ventricular function (echo), myocyte proliferation (Ki67), infarct size (TTC), myocyte nuclear density and cell size. Data were compared to vehicle. Results: Regional LAD infusion increased wall thickening and reduced scar volume but did not affect EF (Table). In contrast, global infusion increased EF despite an insignificant reduction in infarct volume. This was accompanied by 2-fold increases in myocyte proliferation (Ki67) in border and remote regions which did not change with regional LAD infusion. Global infusion increased myocyte nuclear density by 12% and decreased cell size by 9% while there were no significant changes after regional LAD infusion. Conclusion: Although regional LAD infusion reduces scar size and improves regional function, only global infusion improves EF since it effectively stimulates endogenous myocyte regeneration throughout the entire left ventricle. Thus, global infusion may more effectively prevent deleterious post-infarction remodeling than regional administration alone.

Hepatocyte growth factor (HGF) promotes cardiac stem cell differentiation after myocardial infarction by increasing mTOR activation in p27kip haploinsufficient mice

Myocardial infarction (MI) is a major cause of death worldwide. The treatment of MI has improved during the last decade; however, the loss of myocytes remains a problem because the cells cannot be renewed. Cardiac stem cells were recently discovered and were thought to represent a promising treatment for MI. However, the efficiency of cardiac stem cell differentiation into myocyte is not sufficient. Hepatocyte growth factor (HGF) is related to the differentiation and proliferation of cardiac stem cells. p27kip1 is a potent cell cycle inhibitor in most organs, especially the heart. In this study, we investigated the relationship among p27, HGF and cardiac stem cells in post-MI cardiac repair. We found that p27 haploinsufficient mice exhibited preserved cardiac function and improved cardiomyocyte renewal compared with wild-type mice. In addition, p27 haploinsufficiency may not increase cardiac stem cell proliferation but could improve their differentiation by increasing mammalian target of rapamycin expression.

Abstract 243: The E3 Ubiquitin Ligase Parkin Mediates Clearance of Damaged Mitochondria via Two Distinct Pathways

Damaged mitochondria release reactive oxygen species and pro-death factors which can lead to loss of cardiac myocytes. To protect against such damage, myocytes have developed several mechanisms of quality control that act both on the protein and organelle levels. We have previously identified the E3 ubiquitin ligase Parkin as an important regulator of mitochondrial clearance via autophagy in the myocardium. Here, we report that Parkin can also mediate clearance of mitochondria via the endosomal-lysosomal pathway. We found that Parkin promotes clearance of damaged mitochondria in both wild type (WT) and autophagy-deficient Atg5 knockout mouse embryonic fibroblasts (MEFs) treated with the mitochondria uncoupler FCCP. Mitochondrial damage leads to rapid activation of the endosomal-lysosomal pathway in both WT and Atg5-/- MEFs. We further observed increased activation of Rab5, a protein involved in early endosome formation, in both WT and Atg5-/- MEFs after treatment with FCCP. In addition, we observed sequestration of damaged mitochondria in Rab5+ and Rab7+ early and late endosomes, respectively. Mitochondria also colocalized with Lamp2+ vesicles in Atg5-/- MEFs indicating that the mitochondria are ultimately being delivered to the lysosomes for degradation. Overexpression of Rab5S34N, a dominant negative of Rab5, reduces FCCP-mediated clearance and increases cell death in Atg5-/- MEFs. Pharmacological inhibition of the endosomal-lysosomal pathway also results in increased FCCP-mediated cell death. Furthermore, we confirmed that FCCP treatment or simulated ischemia reperfusion exposure induces Rab5 activation with subsequent mitochondrial sequestration in early endosomes in neonatal myocytes. Interestingly, the activation of Rab5 is abrogated in the presence of the mitochondrial targeted antioxidant Mito-Tempo, suggesting that mitochondrial ROS is involved in the activation the endosomal pathway. Mitochondrial clearance via this pathway is also dependent on Parkin, as FCCP treatment fails to activate Rab5 and induce mitochondrial clearance in both WT and Atg5-/- MEFS in the absence of Parkin. Thus, our data suggest that Parkin can mediate clearance of damaged mitochondria via two distinct pathways in cells.

How to Improve the Safe and Effective Use of Doxorubicin in Children with Cancer.

Doxorubicin, one of the anthracyclines, is a chemotherapeutic agent widely used to treat several pediatric cancers such as acute lymphocytic leukemia, acute myeloid leukemia, Hodgkin lymphoma, non-Hodgkin lymphoma, softtissue sarcoma, bone sarcoma, Ewing sarcoma, neuroblastoma, and Wilms tumor. As such, this drug has contributed substantially to the dramatic improvement of the 5-year survival rates for many of these childhood cancers [1]. However, a major potential adverse event of the use of doxorubicin is its cardiotoxicity. More than 50 % of patients will develop asymptomatic cardiac dysfunction, whereas one in six will result in clinical heart failure due to cardiomyopathy [2]. The exact causative mechanism of this devastating adverse event is, despite the fact that doxorubicin has been used for decades in many patients, not fully elucidated. There is more or less consensus about the important role of reactive oxygen species causing cardiac cell damage, progressive myocyte loss that ultimately will result in a decreased cardiac contractility [3]. To date, several clinical risk factors have been linked to the development of doxorubicin-induced cardiotoxicity and these are a higher single as well as total cumulative dose, cardiac irradiation, short infusion time duration, younger age, longer time since the treatment, and female sex [4]. In addition, much more recently, the potentially important role of genetic factors as predictors of doxorubicin-induced cardiotoxicity has been brought to our attention [5]. Multiple genetic variants in several genes associated with this cardiotoxicity have been identified [6]. However, until now, the currently available knowledge about both the clinical as well as the genetic risk factors has never resulted in sufficient discriminatory power to divide patients into groups with a high or low risk of developing cardiotoxicity. In this issue of the journal, Voller and colleagues have described a population pharmacokinetic model of doxorubicin in children in which they have used a power function to describe the relation between doxorubicin clearance and age [7]. They were able to show, for the first time, that the pharmacokinetics (PK) of doxorubicin in infants and children with cancer are age dependent. Clearly, this finding might eventually assist in designing a more tailored, safe, and effective dosing regimen for the use of doxorubicin in children with cancer. However, before this demonstrated lower clearance of doxorubicin in infants and children younger than 3 years of age will be able to change the currently used dosing regimens in pediatric patients with cancer, there is an absolute need to not only consider developmental PK but also, and probably more importantly, developmental pharmacodynamics (PD). At first glance, based on the demonstrated lower total body clearance, it seems that we need less doxorubicin in this group of young patients to reach the same amount of doxorubicin exposure as compared with older children and adolescents. However, what do we really know about the dose-concentration-effect relationship in children with cancer between 1 and 18 years of age? In other words, there is an urgent need to better This commentary refers to the article available at doi:10.1007/s40262-015-0272-4.

Anthracycline-Induced Cardiomyopathy in Adults.

Anthracyclines are one of the most commonly used antineoplastic agent classes, and a core part of the treatment in breast cancers, hematological malignancies, and sarcomas. Their benefit is decreased by their well-recognized cardiotoxicity. The purpose of this review is to outline the presentation, mechanisms, diagnosis, and treatment of anthracyclines-induced cardiotoxicity. Symptomatic heart failure occurs in 2% to 5% of patients treated with anthrayclines and may be higher in older patients or patients with cardiovascular risk factors. The mechanisms involved in anthracycline-induced cardiotoxicity involve myocyte loss by apoptosis in the presence of a limited regenerative capacity. Once symptomatic, anthracycline-induced cardiotoxicity is associated with markedly decreased survival. Left ventricular ejection fraction (LVEF), mostly determined using echocardiography, is used to monitor patients treated with anthracyclines. As more than 1/3 of patients treated with anthracyclines do not recover their baseline LVEF once it is decreased, more sensitive echocardiographic indices of LV function such as myocardial deformation or biomarkers have been studied in patients monitoring. Cardioprotective treatments such as angiotensin-converting enzyme inhibitors, beta-blockers, iron chelators, statins, and metformin are also the topic of research efforts.

Can Cytoprotective Cobalt Protoporphyrin Protect Skeletal Muscle and Muscle-derived Stem Cells From Ischemic Injury?

Extremity trauma is the most common injury seen in combat hospitals as well as in civilian trauma centers. Major skeletal muscle injuries that are complicated by ischemia often result in substantial muscle loss, residual disability, or even amputation, yet few treatment options are available. A therapy that would increase skeletal muscle tolerance to hypoxic damage could reduce acute myocyte loss and enhance preservation of muscle mass in these situations. In these experiments, we investigated (1) whether cobalt protoporphyrin (CoPP), a pharmacologic inducer of cytoprotective heme oxygenase-1 (HO-1), would upregulate HO-1 expression and activity in skeletal muscle, tested in muscle-derived stem cells (MDSCs); and (2) whether CoPP exposure would protect MDSCs from cell death during in vitro hypoxia/reoxygenation. Then, using an in vivo mouse model of hindlimb ischemia/reperfusion injury, we examined (3) whether CoPP pharmacotherapy would reduce skeletal muscle damage when delivered after injury; and (4) whether it would alter the host inflammatory response to injury. MDSCs were exposed in vitro to a single dose of 25 μΜ CoPP and harvested over 24 to 96 hours, assessing HO-1 protein expression by Western blot densitometry and HO-1 enzyme activity by cGMP levels. To generate hypoxia/reoxygenation stress, MDSCs were treated in vitro with phosphate-buffered saline (vehicle), CoPP, or CoPP plus an HO-1 inhibitor, tin protoporphyrin (SnPP), and then subjected to 5 hours of hypoxia (< 0.5% O2) followed by 24 hours of reoxygenation and evaluated for apoptosis. In vivo, hindlimb ischemia/reperfusion injury was produced in mice by unilateral 2-hour tourniquet application followed by 24 hours of reperfusion. In three postinjury treatment groups (n = 7 mice/group), CoPP was administered intraperitoneally during ischemia, at the onset of reperfusion, or 1 hour later. Two control groups of mice with the same injury received phosphate-buffered saline (vehicle) or the HO-1 inhibitor, SnPP. Myocyte damage in the gastrocnemius and tibialis anterior muscles was determined by uptake of intraperitoneally delivered Evans blue dye (EBD), quantified by image analysis. On serial sections, inflammation was gauged by the mean myeloperoxidase staining intensity per unit area over the entirety of each muscle. In MDSCs, a single exposure to CoPP increased HO-1 protein expression and enzyme activity, both of which were sustained for 96 hours. CoPP treatment of MDSCs reduced apoptotic cell populations by 55% after in vitro hypoxia/reoxygenation injury (from a mean of 57.3% apoptotic cells in vehicle-treated controls to 25.7% in CoPP-treated cells, mean difference 31.6%; confidence interval [CI], 28.1-35.0; p < 0.001). In the hindlimb ischemia/reperfusion model, CoPP delivered during ischemia produced a 38% reduction in myocyte damage in the gastrocnemius muscle (from 86.4% ± 7% EBD(+) myofibers in vehicle-treated, injured controls to 53.2% EBD(+) in CoPP-treated muscle, mean difference 33.2%; 95% CI, 18.3, 48.4; p < 0.001). A 30% reduction in injury to the gastrocnemius was seen with drug delivery at the onset of reperfusion (to 60.6% ± 13% EBD(+) with CoPP treatment, mean difference 25.8%; CI, 12.2-39.4; p < 0.001). In the tibialis anterior, however, myocyte damage was decreased only when CoPP was given at the onset of reperfusion, resulting in a 27% reduction in injury (from 78.8% ± 8% EBD(+) myofibers in injured controls to 58.3% ± 14% with CoPP treatment, mean difference 20.5%; CI, 6.1-35.0; p = 0.004). Delaying CoPP delivery until 1 hour after tourniquet release obviated the protective effect in both muscles. Mean MPO staining intensity per unit area, indicating the host inflammatory response, decreased by 27-34% across both the gastrocnemius and tibialis anterior muscles when CoPP was given either during ischemia or at the time of reperfusion. Delaying drug delivery until 1 hour after the start of reperfusion abrogated this antiinflammatory effect. CoPP can decrease skeletal muscle damage when given early in the course of ischemia/reperfusion injury and also provide protection for regenerative stem cell populations. Pharmacotherapy with HO-1 inducers, delivered in the field, on hospital arrival, or during trauma surgery, may improve preservation of muscle mass and muscle-inherent stem cells after severe ischemic limb injury.