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The HSP70 co-chaperone BAG3 targets unfolded proteins to degradation via chaperone assisted selective autophagy (CASA), thereby playing pivotal roles in the proteostasis of adult cardiomyocytes (CMs). However, the complex functions of BAG3 for regulating autophagy in cardiac disease are not completely understood. Here, we demonstrate that conditional inactivation of Bag3 in murine CMs leads to age-dependent dysregulation of autophagy, associated with progressive cardiomyopathy. Surprisingly, Bag3-deficient CMs show increased canonical and non-canonical autophagic flux in the juvenile period when first signs of cardiac dysfunction appear, but reduced autophagy during later stages of the disease. Juvenile Bag3-deficient CMs are characterized by decreased levels of soluble proteins involved in synchronous contraction of the heart, including the gap junction protein Connexin 43 (CX43). Reiterative administration of chloroquine (CQ), an inhibitor of canonical and non-canonical autophagy, but not inactivation of Atg5, restores normal concentrations of soluble cardiac proteins in juvenile Bag3-deficient CMs without an increase of detergent-insoluble proteins, leading to complete recovery of early-stage cardiac dysfunction in Bag3-deficient mice. We conclude that loss of Bag3 in CMs leads to age-dependent differences in autophagy and cardiac dysfunction. Increased non-canonical autophagic flux in the juvenile period removes soluble proteins involved in cardiac contraction, leading to early-stage cardiomyopathy, which is prevented by reiterative CQ treatment., Competing Interests: Conflicts of interest None declared., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Using ferric chloride (FeCl3) to induce experimental superior sagittal sinus (SSS) thrombosis might interfere with magnetic resonance imaging (MRI)-assisted visualization and evaluation of the thrombus, the brain parenchyma, and the quality of the occlusion. The aim of this study was to investigate whether aluminum chloride (AlCl3)-induced thrombosis of the SSS has comparable properties to those of FeCl3 without causing artifacts in MRI. SSS thrombosis was induced in 14 male Wistar rats by exposure of the SSS and subsequent topical application of a filter paper strip soaked in AlCl3 (n = 7) or FeCl3 (n = 7) over a period of 15 min. The animals with AlCl3-induced SSS thrombosis showed a constant and complete occlusion with in histological analysis large thrombi. Blood flow measurements indicated a significant reduction on the first and seventh postoperative day compared to preoperative measurements. MRI enabled visualization and subsequent evaluation of the thrombus and the surrounding parenchyma. In comparison, FeCl3-induced SSS thrombosis could not be evaluated by MRI due to artifacts caused by the paramagnetic properties and increased susceptibility of FeCl3. The occluded sinus and the surrounding area appeared hypointense. The quality of SSS occlusion by AlCl3 was comparable to that of FeCl3. AlCl3 therefore represents a significant alternative substance in experimental SSS thrombosis ideally suited for studies using MRI., (© 2024. The Author(s).)

Postnatal maturation of cardiomyocytes is characterized by a metabolic switch from glycolysis to fatty acid oxidation, chromatin reconfiguration and exit from the cell cycle, instating a barrier for adult heart regeneration 1,2 . Here, to explore whether metabolic reprogramming can overcome this barrier and enable heart regeneration, we abrogate fatty acid oxidation in cardiomyocytes by inactivation of Cpt1b. We find that disablement of fatty acid oxidation in cardiomyocytes improves resistance to hypoxia and stimulates cardiomyocyte proliferation, allowing heart regeneration after ischaemia-reperfusion injury. Metabolic studies reveal profound changes in energy metabolism and accumulation of α-ketoglutarate in Cpt1b-mutant cardiomyocytes, leading to activation of the α-ketoglutarate-dependent lysine demethylase KDM5 (ref.  3 ). Activated KDM5 demethylates broad H3K4me3 domains in genes that drive cardiomyocyte maturation, lowering their transcription levels and shifting cardiomyocytes into a less mature state, thereby promoting proliferation. We conclude that metabolic maturation shapes the epigenetic landscape of cardiomyocytes, creating a roadblock for further cell divisions. Reversal of this process allows repair of damaged hearts., (© 2023. The Author(s).)

Background: The ability of the right ventricle (RV) to adapt to an increased pressure afterload determines survival in patients with pulmonary arterial hypertension. At present, there are no specific treatments available to prevent RV failure, except for heart/lung transplantation. The wingless/int-1 (Wnt) signaling pathway plays an important role in the development of the RV and may also be implicated in adult cardiac remodeling., Methods: Molecular, biochemical, and pharmacological approaches were used both in vitro and in vivo to investigate the role of Wnt signaling in RV remodeling., Results: Wnt/β-catenin signaling molecules are upregulated in RV of patients with pulmonary arterial hypertension and animal models of RV overload (pulmonary artery banding-induced and monocrotaline rat models). Activation of Wnt/β-catenin signaling leads to RV remodeling via transcriptional activation of FOSL1 and FOSL2 (FOS proto-oncogene [FOS] like 1/2, AP-1 [activator protein 1] transcription factor subunit). Immunohistochemical analysis of pulmonary artery banding -exposed BAT-Gal (β-catenin-activated transgene driving expression of nuclear β-galactosidase) reporter mice RVs exhibited an increase in β-catenin expression compared with their respective controls. Genetic inhibition of β-catenin, FOSL1/2, or WNT3A stimulation of RV fibroblasts significantly reduced collagen synthesis and other remodeling genes. Importantly, pharmacological inhibition of Wnt signaling using inhibitor of PORCN (porcupine O-acyltransferase), LGKK-974 attenuated fibrosis and cardiac hypertrophy leading to improvement in RV function in both, pulmonary artery banding - and monocrotaline-induced RV overload., Conclusions: Wnt- β-Catenin-FOSL signaling is centrally involved in the hypertrophic RV response to increased afterload, offering novel targets for therapeutic interference with RV failure in pulmonary hypertension., Competing Interests: Disclosures None.

Immune cell trafficking constitutes a fundamental component of immunological response to tissue injury, but the contribution of intrinsic RNA nucleotide modifications to this response remains elusive. We report that RNA editor ADAR2 exerts a tissue- and stress-specific regulation of endothelial responses to interleukin-6 (IL-6), which tightly controls leukocyte trafficking in IL-6-inflamed and ischemic tissues. Genetic ablation of ADAR2 from vascular endothelial cells diminished myeloid cell rolling and adhesion on vascular walls and reduced immune cell infiltration within ischemic tissues. ADAR2 was required in the endothelium for the expression of the IL-6 receptor subunit, IL-6 signal transducer (IL6ST; gp130), and subsequently, for IL-6 trans-signaling responses. ADAR2-induced adenosine-to-inosine RNA editing suppressed the Drosha-dependent primary microRNA processing, thereby overwriting the default endothelial transcriptional program to safeguard gp130 expression. This work demonstrates a role for ADAR2 epitranscriptional activity as a checkpoint in IL-6 trans-signaling and immune cell trafficking to sites of tissue injury., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Activation of endothelial YAP/TAZ signaling is crucial for physiological and pathological angiogenesis. The mechanisms of endothelial YAP/TAZ regulation are, however, incompletely understood. Here we report that the protocadherin FAT1 acts as a critical upstream regulator of endothelial YAP/TAZ which limits the activity of these transcriptional cofactors during developmental and tumor angiogenesis by promoting their degradation. We show that loss of endothelial FAT1 results in increased endothelial cell proliferation in vitro and in various angiogenesis models in vivo. This effect is due to perturbed YAP/TAZ protein degradation, leading to increased YAP/TAZ protein levels and expression of canonical YAP/TAZ target genes. We identify the E3 ubiquitin ligase Mind Bomb-2 (MIB2) as a FAT1-interacting protein mediating FAT1-induced YAP/TAZ ubiquitination and degradation. Loss of MIB2 expression in endothelial cells in vitro and in vivo recapitulates the effects of FAT1 depletion and causes decreased YAP/TAZ degradation and increased YAP/TAZ signaling. Our data identify a pivotal mechanism of YAP/TAZ regulation involving FAT1 and its associated E3 ligase MIB2, which is essential for YAP/TAZ-dependent angiogenesis., (© 2023. The Author(s).)

Aims: Mammalian models have been instrumental in investigating adult heart function and human disease. However, electrophysiological differences with human hearts and high costs motivate the need for non-mammalian models. The zebrafish is a well-established genetic model to study cardiovascular development and function; however, analysis of cardiovascular phenotypes in adult specimens is particularly challenging as they are opaque., Methods and Results: Here, we optimized and combined multiple imaging techniques including echocardiography, magnetic resonance imaging, and micro-computed tomography to identify and analyse cardiovascular phenotypes in adult zebrafish. Using alk5a/tgfbr1a mutants as a case study, we observed morphological and functional cardiovascular defects that were undetected with conventional approaches. Correlation analysis of multiple parameters revealed an association between haemodynamic defects and structural alterations of the heart, as observed clinically., Conclusion: We report a new, comprehensive, and sensitive platform to identify otherwise indiscernible cardiovascular phenotypes in adult zebrafish., Competing Interests: Conflict of interest: none declared., (© The Author(s) 2021. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Background: ADAR1 (adenosine deaminase acting on RNA-1)-mediated adenosine to inosine (A-to-I) RNA editing plays an essential role for distinguishing endogenous from exogenous RNAs, preventing autoinflammatory ADAR1 also regulates cellular processes by recoding specific mRNAs, thereby altering protein functions, but may also act in an editing-independent manner. The specific role of ADAR1 in cardiomyocytes and its mode of action in the heart is not fully understood. To determine the role of ADAR1 in the heart, we used different mutant mouse strains, which allows to distinguish immunogenic, editing-dependent, and editing-independent functions of ADAR1., Methods: Different Adar1 -mutant mouse strains were employed for gene deletion or specific inactivation of ADAR1 enzymatic activity in cardiomyocytes, either alone or in combination with Ifih1 (interferon induced with helicase C domain 1) or Irf7 (interferon regulatory factor 7) gene inactivation. Mutant mice were investigated by immunofluorescence, Western blot, RNAseq, proteomics, and functional MRI analysis., Results: Inactivation of Adar1 in cardiomyocytes resulted in late-onset autoinflammatory myocarditis progressing into dilated cardiomyopathy and heart failure at 6 months of age. Adar1 depletion activated interferon signaling genes but not NFκB (nuclear factor kappa B) signaling or apoptosis and reduced cardiac hypertrophy during pressure overload via induction of Irf7 . Additional inactivation of the cytosolic RNA sensor MDA5 (melanoma differentiation-associated gene 5; encoded by the Ifih1 gene) in Adar1 mutant mice prevented activation of interferon signaling gene and delayed heart failure but did not prevent lethality after 8.5 months. In contrast, compound mutants only expressing catalytically inactive ADAR1 in an Ifih1 -mutant background were completely normal. Inactivation of Irf7 attenuated the phenotype of Adar1 -deficient cardiomyocytes to a similar extent as Ifih1 depletion, identifying IRF7 as the main mediator of autoinflammatory responses caused by the absence of ADAR1 in cardiomyocytes., Conclusions: Enzymatically active ADAR1 prevents IRF7-mediated autoinflammatory reactions in the heart triggered by endogenous nonedited RNAs. In addition to RNA editing, ADAR1 also serves editing-independent roles in the heart required for long-term cardiac function and survival.

Oncostatin M (OSM) and leukemia inhibitory factor (LIF) signaling protects the heart after myocardial infarction (MI). In mice, oncostatin M receptor (OSMR) and leukemia inhibitory factor receptor (LIFR) are selectively activated by the respective cognate ligands while OSM activates both the OSMR and LIFR in humans, which prevents efficient translation of mouse data into potential clinical applications. We used an engineered human-like OSM (hlOSM) protein, capable to signal via both OSMR and LIFR, to evaluate beneficial effects on cardiomyocytes and hearts after MI in comparison to selective stimulation of either LIFR or OSMR. Cell viability assays, transcriptome and immunoblot analysis revealed increased survival of hypoxic cardiomyocytes by mLIF, mOSM and hlOSM stimulation, associated with increased activation of STAT3. Kinetic expression profiling of infarcted hearts further specified a transient increase of OSM and LIF during the early inflammatory phase of cardiac remodeling. A post-infarction delivery of hlOSM but not mOSM or mLIF within this time period combined with cardiac magnetic resonance imaging-based strain analysis uncovered a global cardioprotective effect on infarcted hearts. Our data conclusively suggest that a simultaneous and rapid activation of OSMR and LIFR after MI offers a therapeutic opportunity to preserve functional and structural integrity of the infarcted heart.

Background: Early recanalization of an occluded vessel is associated with a better clinical outcome in acute ischemic stroke. Intravenous thrombolysis using recombinant tissue plasminogen activator (rt-PA) is only available in a minority of patients and often fails to reopen the occluded vessel. Mechanical recanalization is more effective in this matter but only available for selected patients when a thrombectomy centre can be reached. Therefore, sonothrombolysis might represent an alternative or complementary approach. Here, we tested microbubble-mediated sonothrombolysis (mmSTL) in a thromboembolic stroke model for middle cerebral artery occlusion (MCAO) in rats., Methods: Sixty-seven male Wistar rats underwent MCAO using an autologous full blood thrombus and were randomly assigned to four groups receiving rt-PA, mmSTL, a combination of both, or a placebo. Diagnostic workup included neurological examination, assessment of infarct size, and presence of intracerebral haemorrhage by magnetic resonance imaging (MRI) and presence of microbleedings in histological staining., Results: Neurological examination revealed no differences between the treatment groups. In all treatment groups, there was a reduction in infarct size 24 hours after MCAO as compared to the placebo (P≤0.05), but there were no differences between the active treatment groups (P>0.05) (placebo 0.75±0.10 cm 3 ; mmSTL 0.43±0.07 cm 3 ; rt-PA 0.4±0.07 cm 3 ; mmSTL + rt-PA 0.27±0.08 cm 3 ). Histological staining displayed intracerebral microbleedings in all animals. The frequency of gross bleeding detected by MRI did not differ between the groups (placebo 3; mmSTL 4; rt-PA 2; mmSTL + rt-PA 2; P>0.05) and was not associated with worse performance in clinical testing (P>0.05). There were no statistical differences in the mortality between the groups (P>0.05)., Conclusions: Our study showed the efficacy and safety of mmSTL with or without rt-PA in an embolic rat stroke model using a continuous full blood thrombus. Sonothrombolysis might be useful for patients who need to be transported to a thrombectomy centre or for those with distal vessel occlusion., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/atm-21-75). TB reports grants from the University Medical Center Giessen and Marburg (UKGM), Germany; non-financial support from Bracco Suisse SA, Geneva, Switzerland, during the conduct of the study; non-financial support from Bayer Vital GmbH, outside the submitted work. The other authors have no conflicts of interest to declare., (2021 Annals of Translational Medicine. All rights reserved.)

Although NF-κB is known to play a pivotal role in lung cancer, contributing to tumor growth, microenvironmental changes, and metastasis, the epigenetic regulation of NF-κB in tumor context is largely unknown. Here we report that the IKK2/NF-κB signaling pathway modulates metastasis-associated protein 2 (MTA2), a component of the nucleosome remodeling and deacetylase complex (NuRD). In triple transgenic mice, downregulation of IKK2 (Sftpc-cRaf-IKK2DN) in cRaf-induced tumors in alveolar epithelial type II cells restricted tumor formation, whereas activation of IKK2 (Sftpc-cRaf-IKK2CA) supported tumor growth; both effects were accompanied by altered expression of MTA2. Further studies employing genetic inhibition of MTA2 suggested that in primary tumor growth, independent of IKK2, MTA2/NuRD corepressor complex negatively regulates NF-κB signaling and tumor growth, whereas later dissociation of MTA2/NuRD complex from the promoter of NF-κB target genes and IKK2-dependent positive regulation of MTA2 leads to activation of NF-κB signaling, epithelial-mesenchymal transition, and lung tumor metastasis. These findings reveal a previously unrecognized biphasic role of MTA2 in IKK2/NF-κB-driven primary-to-metastatic lung tumor progression. Addressing the interaction between MTA2 and NF-κB would provide potential targets for intervention of tumor growth and metastasis. SIGNIFICANCE: These findings strongly suggest a prominent role of MTA2 in primary tumor growth, lung metastasis, and NF-κB signaling modulatory functions., (©2020 American Association for Cancer Research.)

Long non-coding RNAs (lncRNAs) contribute to cardiac (patho)physiology. Aging is the major risk factor for cardiovascular disease with cardiomyocyte apoptosis as one underlying cause. Here, we report the identification of the aging-regulated lncRNA Sarrah (ENSMUST00000140003) that is anti-apoptotic in cardiomyocytes. Importantly, loss of SARRAH (OXCT1-AS1) in human engineered heart tissue results in impaired contractile force development. SARRAH directly binds to the promoters of genes downregulated after SARRAH silencing via RNA-DNA triple helix formation and cardiomyocytes lacking the triple helix forming domain of Sarrah show an increase in apoptosis. One of the direct SARRAH targets is NRF2, and restoration of NRF2 levels after SARRAH silencing partially rescues the reduction in cell viability. Overexpression of Sarrah in mice shows better recovery of cardiac contractile function after AMI compared to control mice. In summary, we identified the anti-apoptotic evolutionary conserved lncRNA Sarrah, which is downregulated by aging, as a regulator of cardiomyocyte survival.

Enlarged vestibular aqueduct (EVA) is one of the most commonly identified inner ear malformations in hearing loss patients including Pendred syndrome. While biallelic mutations of the SLC26A4 gene, encoding pendrin, causes non-syndromic hearing loss with EVA or Pendred syndrome, a considerable number of patients appear to carry mono-allelic mutation. This suggests faulty pendrin regulatory machinery results in hearing loss. Here we identify EPHA2 as another causative gene of Pendred syndrome with SLC26A4. EphA2 forms a protein complex with pendrin controlling pendrin localization, which is disrupted in some pathogenic forms of pendrin. Moreover, point mutations leading to amino acid substitution in the EPHA2 gene are identified from patients bearing mono-allelic mutation of SLC26A4. Ephrin-B2 binds to EphA2 triggering internalization with pendrin inducing EphA2 autophosphorylation weakly. The identified EphA2 mutants attenuate ephrin-B2- but not ephrin-A1-induced EphA2 internalization with pendrin. Our results uncover an unexpected role of the Eph/ephrin system in epithelial function.

Myogenic vasoconstriction is an autoregulatory function of small arteries. Recently, G-protein-coupled receptors have been involved in myogenic vasoconstriction, but the downstream signalling mechanisms and the in-vivo-function of this myogenic autoregulation are poorly understood. Here, we show that small arteries from mice with smooth muscle-specific loss of G 12 /G 13 or the Rho guanine nucleotide exchange factor ARHGEF12 have lost myogenic vasoconstriction. This defect was accompanied by loss of RhoA activation, while vessels showed normal increases in intracellular [Ca 2+ ]. In the absence of myogenic vasoconstriction, perfusion of peripheral organs was increased, systemic vascular resistance was reduced and cardiac output and left ventricular mass were increased. In addition, animals with defective myogenic vasoconstriction showed aggravated hypotension in response to endotoxin. We conclude that G 12 /G 13 - and Rho-mediated signaling plays a key role in myogenic vasoconstriction and that myogenic tone is required to maintain local and systemic vascular resistance under physiological and pathological condition., Competing Interests: RC, AW, JF, RL, RB, YJ, AW, FS, NW, DH, SO No competing interests declared, (© 2019, Chennupati et al.)

Constitutive expression of the chemokine Mcp1 in mouse cardiomyocytes creates a model of inflammatory cardiomyopathy, with death from heart failure at age 7-8 months. A critical pathogenic role has previously been proposed for induced oxidative stress, involving NADPH oxidase activation. To test this idea, we exposed the mice to elevated oxygen levels. Against expectation, this prevented, rather than accelerated, the ultrastructural and functional signs of heart failure. This result suggests that the immune signaling initiated by Mcp1 leads instead to the inhibition of cellular oxygen usage, for which mitochondrial respiration is an obvious target. To address this hypothesis, we combined the Mcp1 model with xenotopic expression of the alternative oxidase (AOX), which provides a sink for electrons blocked from passage to oxygen via respiratory complexes III and IV. Ubiquitous AOX expression provided only a minor delay to cardiac functional deterioration and did not prevent the induction of markers of cardiac and metabolic remodeling considered a hallmark of the model. Moreover, cardiomyocyte-specific AOX expression resulted in exacerbation of Mcp1-induced heart failure, and failed to rescue a second cardiomyopathy model directly involving loss of cIV. Our findings imply that mitochondrial involvement in the pathology of inflammatory cardiomyopathy is multifaceted and complex.

Hypoxia signaling plays a major role in non-malignant and malignant hyperproliferative diseases. Pulmonary hypertension (PH), a hypoxia-driven vascular disease, is characterized by a glycolytic switch similar to the Warburg effect in cancer. Ras association domain family 1A (RASSF1A) is a scaffold protein that acts as a tumour suppressor. Here we show that hypoxia promotes stabilization of RASSF1A through NOX-1- and protein kinase C- dependent phosphorylation. In parallel, hypoxia inducible factor-1 α (HIF-1α) activates RASSF1A transcription via HIF-binding sites in the RASSF1A promoter region. Vice versa, RASSF1A binds to HIF-1α, blocks its prolyl-hydroxylation and proteasomal degradation, and thus enhances the activation of the glycolytic switch. We find that this mechanism operates in experimental hypoxia-induced PH, which is blocked in RASSF1A knockout mice, in human primary PH vascular cells, and in a subset of human lung cancer cells. We conclude that RASSF1A-HIF-1α forms a feedforward loop driving hypoxia signaling in PH and cancer.

Pulmonary arterial hypertension (PAH) is a progressive disorder characterized by remodeling of the pulmonary vasculature and a rise in right ventricular (RV) afterload. The increased RV afterload leads to right ventricular failure (RVF) which is the reason for the high morbidity and mortality in PAH patients. The objective was to evaluate the therapeutic efficacy and antiremodeling potential of the phosphodiesterase type 5 (PDE5) inhibitor sildenafil and the soluble guanylate cyclase stimulator riociguat in a model of pressure overload RV hypertrophy induced by pulmonary artery banding (PAB). Mice subjected to PAB, one week after surgery, were treated with either sildenafil (100 mg/kg/d, n = 5), riociguat (30 mg/kg/d, n = 5), or vehicle ( n = 5) for 14 days. RV function and remodeling were assessed by right heart catheterization, magnetic resonance imaging (MRI), and histomorphometry. Both sildenafil and riociguat prevented the deterioration of RV function, as determined by a decrease in RV dilation and restoration of the RV ejection fraction (EF). Although both compounds did not decrease right heart mass and cellular hypertrophy, riociguat prevented RV fibrosis induced by PAB. Both compounds diminished TGF-beta1 induced collagen synthesis of RV cardiac fibroblasts in vitro . Treatment with either riociguat or sildenafil prevented the progression of pressure overload-induced RVF, representing a novel therapeutic approach.

Sirtuins (Sirt1-Sirt7) are NAD + -dependent protein deacetylases/ADP ribosyltransferases, which play decisive roles in chromatin silencing, cell cycle regulation, cellular differentiation, and metabolism. Different sirtuins control similar cellular processes, suggesting a coordinated mode of action but information about potential cross-regulatory interactions within the sirtuin family is still limited. Here, we demonstrate that Sirt1 requires autodeacetylation to efficiently deacetylate targets such as p53, H3K9, and H4K16. Sirt7 restricts Sirt1 activity by preventing Sirt1 autodeacetylation causing enhanced Sirt1 activity in Sirt7 -/- mice. Increased Sirt1 activity in Sirt7 -/- mice blocks PPARγ and adipocyte differentiation, thereby diminishing accumulation of white fat. Thus, reduction of Sirt1 activity restores adipogenesis in Sirt7 -/- adipocytes in vitro and in vivo. We disclosed a principle controlling Sirt1 activity and uncovered an unexpected complexity in the crosstalk between two different sirtuins. We propose that antagonistic interactions between Sirt1 and Sirt7 are pivotal in controlling the signaling network required for maintenance of adipose tissue., Competing Interests: The authors declare no conflict of interest.

GPR116 (ADGRF5) and ELTD1 (ADGRL4) belong to different subfamilies of the adhesion G-protein-coupled receptor group but are both expressed in endothelial cells. We therefore analyzed their functions in mice lacking these receptors. While loss of GPR116 or ELTD1 alone had no obvious effect on cardiovascular or kidney function, mice lacking both, GPR116 and ELTD1, showed malformations of the aortic arch arteries and the cardiac outflow tract leading to perinatal lethality in about 50% of the mutants. In addition to cardiovascular malformations, surviving mice developed renal thrombotic microangiopathy as well as hemolysis and splenomegaly, and their lifespan was significantly reduced. Loss of GPR116 and ELTD1 specifically in endothelial cells or neural crest-derived cells did not recapitulate any of the phenotypes observed in GPR116-ELTD1 double deficient mice, indicating that loss of GPR116 and ELTD1 expressed by other cells accounts for the observed cardiovascular and renal defects.

Control of energy homeostasis and metabolism is achieved by integrating numerous pathways, and miRNAs are involved in this process by regulating expression of multiple target genes. However, relatively little is known about the posttranscriptional processing of miRNAs and a potential role for the precursors they derive from. Here, we demonstrate that mature miRNA-22 is more abundant in muscle from male mice relative to females and that this enables sex-specific regulation of muscular lipid metabolism and body weight by repressing estrogen receptor alpha (ERα) expression. We found that the ERα adjusts its own activity by preventing processing of miR-22 via direct binding to a conserved ERα-binding element within the primary miR-22 precursor. Mutation of the ERα binding site within the pri-miR-22 in vivo eliminates sex-specific differences in miR-22 expression. We reason that the resulting tissue selective negative feedback regulation is essential to establish sex-specific differences in muscle metabolism and body weight development., (© 2017 The Authors.)

Right ventricular (RV) remodeling represents a complex set of functional and structural adaptations in response to chronic pressure or volume overload due to various inborn defects or acquired diseases and is an important determinant of patient outcome. However, the underlying molecular mechanisms remain elusive. We investigated the time course of structural and functional changes in the RV in the murine model of pressure overload-induced RV hypertrophy in C57Bl/6J mice. Using magnetic resonance imaging, we assessed the changes of RV structure and function at different time points for a period of 21 days. Pressure overload led to significant dilatation, cellular and chamber hypertrophy, myocardial fibrosis, and functional impairment of the RV Progressive remodeling of the RV after pulmonary artery banding (PAB) in mice was associated with upregulation of myocardial gene markers of hypertrophy and fibrosis. Furthermore, remodeling of the RV was associated with accumulation and activation of mast cells in the RV tissue of PAB mice. Our data suggest possible involvement of mast cells in the RV remodeling process in response to pressure overload. Mast cells may thus represent an interesting target for the development of new therapeutic approaches directed specifically at the RV., (© 2017 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.)

Plants and many lower organisms, but not mammals, express alternative oxidases (AOXs) that branch the mitochondrial respiratory chain, transferring electrons directly from ubiquinol to oxygen without proton pumping. Thus, they maintain electron flow under conditions when the classical respiratory chain is impaired, limiting excess production of oxygen radicals and supporting redox and metabolic homeostasis. AOX from Ciona intestinalis has been used to study and mitigate mitochondrial impairments in mammalian cell lines, Drosophila disease models and, most recently, in the mouse, where multiple lentivector-AOX transgenes conferred substantial expression in specific tissues. Here, we describe a genetically tractable mouse model in which Ciona AOX has been targeted to the Rosa26 locus for ubiquitous expression. The AOX Rosa26 mouse exhibited only subtle phenotypic effects on respiratory complex formation, oxygen consumption or the global metabolome, and showed an essentially normal physiology. AOX conferred robust resistance to inhibitors of the respiratory chain in organello; moreover, animals exposed to a systemically applied LD50 dose of cyanide did not succumb. The AOX Rosa26 mouse is a useful tool to investigate respiratory control mechanisms and to decipher mitochondrial disease aetiology in vivo., (© 2017. Published by The Company of Biologists Ltd.)

Rationale: Activated cardiac fibroblasts (CF) are crucial players in the cardiac damage response; excess fibrosis, however, may result in myocardial stiffening and heart failure development. Inhibition of activated CF has been suggested as a therapeutic strategy in cardiac disease, but whether this truly improves cardiac function is unclear., Objective: To study the effect of CF ablation on cardiac remodeling., Methods and Results: We characterized subgroups of murine CF by single-cell expression analysis and identified periostin as the marker showing the highest correlation to an activated CF phenotype. We generated bacterial artificial chromosome-transgenic mice allowing tamoxifen-inducible Cre expression in periostin-positive cells as well as their diphtheria toxin-mediated ablation. In the healthy heart, periostin expression was restricted to valvular fibroblasts; ablation of this population did not affect cardiac function. After chronic angiotensin II exposure, ablation of activated CF resulted in significantly reduced cardiac fibrosis and improved cardiac function. After myocardial infarction, ablation of periostin-expressing CF resulted in reduced fibrosis without compromising scar stability, and cardiac function was significantly improved. Single-cell transcriptional analysis revealed reduced CF activation but increased expression of prohypertrophic factors in cardiac macrophages and cardiomyocytes, resulting in localized cardiomyocyte hypertrophy., Conclusions: Modulation of the activated CF population is a promising approach to prevent adverse cardiac remodeling in response to angiotensin II and after myocardial infarction., (© 2016 American Heart Association, Inc.)

Obesity is an increasing health problem worldwide, and nonsurgical strategies to treat obesity have remained rather inefficient. We here show that acute loss of TGF-β-activated kinase 1 (TAK1) in adipocytes results in an increased rate of apoptotic adipocyte death and increased numbers of M2 macrophages in white adipose tissue. Mice with adipocyte-specific TAK1 deficiency have reduced adipocyte numbers and are resistant to obesity induced by a high-fat diet or leptin deficiency. In addition, adipocyte-specific TAK1-deficient mice under a high-fat diet showed increased energy expenditure, which was accompanied by enhanced expression of the uncoupling protein UCP1. Interestingly, acute induction of adipocyte-specific TAK1 deficiency in mice already under a high-fat diet was able to stop further weight gain and improved glucose tolerance. Thus, loss of TAK1 in adipocytes reduces the total number of adipocytes, increases browning of white adipose tissue, and may be an attractive strategy to treat obesity, obesity-dependent diabetes, and other associated complications.

Myostatin, a member of the TGF-β superfamily of secreted growth factors, is a negative regulator of skeletal muscle growth. In the heart, it is expressed at lower levels compared to skeletal muscle but up-regulated under disease conditions. Cre recombinase-mediated inactivation of myostatin in adult cardiomyocytes leads to heart failure and increased mortality but cardiac function of surviving mice is restored after several weeks probably due to compensatory expression in non-cardiomyocytes. To study long-term effects of increased myostatin expression in the heart and to analyze the putative crosstalk between cardiomyocytes and fibroblasts, we overexpressed myostatin in cardiomyocytes. Increased expression of myostatin in heart muscle cells caused interstitial fibrosis via activation of the TAK-1-MKK3/6-p38 signaling pathway, compromising cardiac function in older mice. Our results uncover a novel role of myostatin in the heart and highlight the necessity for tight regulation of myostatin to maintain normal heart function.

Cardiac healing after myocardial ischemia depends on the recruitment and local expansion of myeloid cells, particularly macrophages. Here we identify Reg3β as an essential regulator of macrophage trafficking to the damaged heart. Using mass spectrometry-based secretome analysis, we found that dedifferentiating cardiomyocytes release Reg3β in response to the cytokine OSM, which signals through Jak1 and Stat3. Loss of Reg3β led to a large decrease in the number of macrophages in the ischemic heart, accompanied by increased ventricular dilatation and insufficient removal of neutrophils. This defect in neutrophil removal in turn caused enhanced matrix degradation, delayed collagen deposition and increased susceptibility to cardiac rupture. Our data indicate that OSM, acting through distinct intracellular pathways, regulates both cardiomyocyte dedifferentiation and cardiomyocyte-dependent regulation of macrophage trafficking. Release of OSM from infiltrating neutrophils and macrophages initiates a positive feedback loop in which OSM-induced production of Reg3β in cardiomyocytes attracts additional OSM-secreting macrophages. The activity of the feedback loop controls the degree of macrophage accumulation in the heart, which is instrumental in myocardial healing.

Rationale: Recent studies indicate that tumor-associated macrophages (MΦ) with an M2 phenotype can influence cancer progression and metastasis, but the regulatory pathways remain poorly characterized., Objectives: This study investigated the role of tumor-associated MΦ in lung cancer., Methods: Coculturing of MΦ with mouse Lewis lung carcinoma (LLC1) and 10 different human lung cancer cell lines (adenocarcinoma, squamous cell carcinoma, and large cell carcinoma) caused up-regulation of CCR2/CCL2 and CX3CR1/CX3CL1 in both the cancer cells and the MΦ., Measurements and Main Results: In the MΦ-tumor cell system, IL-10 drove CCR2 and CX3CR1 up-regulation, whereas CCL1, granulocyte colony-stimulating factor, and MIP1α were required for the up-regulation of CCL2 and CX3CL1. Downstream phenotypic effects included enhanced LLC1 proliferation and migration and MΦ M2 polarization. In vivo, MΦ depletion (clodronate, MΦ Fas-induced apoptosis mice) and genetic ablation of CCR2 and CX3CR1 all inhibited LLC1 tumor growth and metastasis, shifted tumor-associated MΦ toward M1 polarization, suppressed tumor vessel growth, and enhanced survival (metastasis model). Furthermore, mice treated with CCR2 antagonist mimicked genetic ablation of CCR2, showing reduced tumor growth and metastasis. In human lung cancer samples, tumor MΦ infiltration and CCR2 expression correlated with tumor stage and metastasis., Conclusions: Tumor-associated MΦ play a central role in lung cancer growth and metastasis, with bidirectional cross-talk between MΦ and cancer cells via CCR2 and CX3CR1 signaling as a central underlying mechanism. These findings suggest that the therapeutic strategy of blocking CCR2 and CX3CR1 may prove beneficial for halting lung cancer progression.

Mutations in EMD, encoding emerin cause skeletal muscle and heart defects in patients with X-linked Emery-Dreifuss muscular dystrophy (X-EDMD) but the underlying mechanisms leading to cardiac defects are poorly understood. Here, we investigated the role of emerin in controlling cardiomyocyte proliferation and cardiac remodeling and explored its function in regulation of the Wnt/β-catenin pathway. We observed a remarkable increase of cardiomyocytes in emerin-null adult mice accompanied with decreased numbers of multinucleated cells. Depletion of emerin in mouse ES cell-derived cardiomyocytes by shRNA caused hyperactivation of Wnt/β-catenin signaling, increased proliferation and abrogated timely cardiac differentiation. Likewise, emerin-null mice exhibited increased Wnt/β-catenin signaling, cardiac dysfunction and perturbed hypertrophic remodeling following pressure overload. Pharmacological inhibition of β-catenin normalized proliferation and differentiation of ES cell-derived cardiomyocytes while inactivation of a single allele of β-catenin efficiently rescued cardiac dysfunction in emerin-null mice. We conclude that emerin constrains β-catenin signaling in the heart providing tight control of cardiomyocyte numbers. Enhanced Wnt/β-catenin signaling seems to contribute to cardiac defects observed in X-EDMD. Hence, therapeutic inhibition of Wnt/β-catenin signaling might be beneficial for X-EDMD patients., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objective: The serotonin (5-HT) pathway was shown to play a role in pulmonary hypertension (PH), but its functions in right ventricular failure (RVF) remain poorly understood. The aim of the current study was to investigate the effects of Terguride (5-HT2A and 2B receptor antagonist) or SB204741 (5-HT2B receptor antagonist) on right heart function and structure upon pulmonary artery banding (PAB) in mice., Methods: Seven days after PAB, mice were treated for 14 days with Terguride (0.2 mg/kg bid) or SB204741 (5 mg/kg day). Right heart function and remodeling were assessed by right heart catheterization, magnetic resonance imaging (MRI), and histomorphometric methods. Total secreted collagen content was determined in mouse cardiac fibroblasts isolated from RV tissues., Results: Chronic treatment with Terguride or SB204741 reduced right ventricular fibrosis and showed improved heart function in mice after PAB. Moreover, 5-HT2B receptor antagonists diminished TGF-beta1 induced collagen synthesis of RV cardiac fibroblasts in vitro., Conclusion: 5-HT2B receptor antagonists reduce collagen deposition, thereby inhibiting right ventricular fibrosis. Chronic treatment prevented the development and progression of pressure overload-induced RVF in mice. Thus, 5-HT2B receptor antagonists represent a valuable novel therapeutic approach for RVF.

The electrical properties of the heart are primarily determined by the activity of ion channels and the activity of these molecules is permanently modulated and adjusted to the physiological needs by adrenergic signaling. miRNAs are known to control the expression of many proteins and to fulfill distinct functions in the mammalian heart, though the in vivo effects of miRNAs on the electrical activity of the heart are poorly characterized. The miRNAs miR-1 and miR-133a are the most abundant miRNAs of the heart and are expressed from two miR-1/133a genomic clusters. Genetic modulation of miR-1/133a cluster expression without concomitant severe disturbance of general cardiomyocyte physiology revealed that these miRNA clusters govern cardiac muscle repolarization. Reduction of miR-1/133a dosage induced a longQT phenotype in mice especially at low heart rates. Longer action potentials in cardiomyocytes are caused by modulation of the impact of β-adrenergic signaling on the activity of the depolarizing L-type calcium channel. Pharmacological intervention to attenuate β-adrenergic signaling or L-type calcium channel activity in vivo abrogated the longQT phenotype that is caused by modulation of miR-1/133a activity. Thus, we identify the miR-1/133a miRNA clusters to be important to prevent a longQT-phenotype in the mammalian heart.

Rationale: Myostatin is a major negative regulator of skeletal muscle mass and initiates multiple metabolic changes, including enhanced insulin sensitivity. However, the function of myostatin in the heart is barely understood, although it is upregulated in the myocardium under several pathological conditions., Objective: Here, we aimed to decipher the role of myostatin and myostatin-dependent signaling pathways for cardiac function and cardiac metabolism in adult mice. To avoid potential counterregulatory mechanisms occurring in constitutive and germ-line-based myostatin mutants, we generated a mouse model that allows myostatin inactivation in adult cardiomyocytes., Methods and Results: Cardiac MRI revealed that genetic inactivation of myostatin signaling in the adult murine heart caused cardiac hypertrophy and heart failure, partially recapitulating effects of the age-dependent decline of the myostatin paralog growth and differentiation factor 11. We found that myostatin represses AMP-activated kinase activation in the heart via transforming growth factor-β-activated kinase 1, thereby preventing a metabolic switch toward glycolysis and glycogen accumulation. Furthermore, myostatin stimulated expression of regulator of G-protein signaling 2, a GTPase-activating protein that restricts Gaq and Gas signaling and thereby protects against cardiac failure. Inhibition of AMP-activated kinase in vivo rescued cardiac hypertrophy and prevented enhanced glycolytic flow and glycogen accumulation after inactivation of myostatin in cardiomyocytes., Conclusions: Our results uncover an important role of myostatin in the heart for maintaining cardiac energy homeostasis and preventing cardiac hypertrophy., (© 2014 American Heart Association, Inc.)

Aims: Mouse models of myocardial infarction (MI) are commonly used to explore the pathophysiological role of the monocytic response in myocardial injury and to develop translational strategies. However, no study thus far has examined the potential impact of inter-individual variability and sham surgical procedures on monocyte subset kinetics after experimental MI in mice. Our goal was to investigate determinants of systemic myeloid cell subset shifts in C57BL/6 mice following MI by developing a protocol for sequential extensive flow cytometry (FCM)., Methods and Results: Following cross-sectional multiplex FCM analysis we provide for the first time a detailed description of absolute quantities, relative subset composition, and biological variability of circulating classical, intermediate, and non-classical monocyte subsets in C57BL/6 mice. By using intra-individual longitudinal measurements after MI induction, a time course of classical and non-classical monocytosis was recorded. This approach disclosed a significant reduction of monocyte subset dispersion across all investigated time points following MI. We found that in the current invasive model of chronic MI the global pattern of systemic monocyte kinetics is mainly determined by a nonspecific inflammatory response to sham surgery and not by the extent of myocardial injury., Conclusions: Application of sequential multiplexed FCM may help to reduce the impact of biological variability in C57BL/6 mice. Furthermore, the confounding influence of sham surgical procedures should always be considered when measuring monocyte subset kinetics in a murine model of MI.

Heart failure (HF) is a common and potentially deadly condition, which frequently develops as a consequence of various diseases of the heart. The incidence of heart failure continuously increases in aging societies illustrating the need for new therapeutic approaches. We recently discovered that continuous activation of oncostatin M (OSM), a cytokine of the interleukin-6 family that induces dedifferentiation of cardiomyocytes, promotes progression of heart failure in dilative cardiomyopathy. To evaluate whether inhibition of OSM signaling represents a meaningful therapeutic approach to prevent heart failure we attenuated OSM-receptor (Oβ) signaling in a mouse model of inflammatory dilative cardiomyopathy. We found that administration of an antibody directed against the extracellular domain of Oβ or genetic inactivation of a single allele of the Oβ gene reduced cardiomyocyte remodeling and dedifferentiation resulting in improved cardiac performance and increased survival. We conclude that pharmacological attenuation of long-lasting Oβ signaling is a promising strategy to treat different types and stages of HF that go along with infiltration by OSM-releasing inflammatory cells.

Cardiac pressure overload-induced hypertrophy and pathological remodelling frequently leads to right ventricular dysfunction, which is the most frequent cause of death in patients with pulmonary arterial hypertension. Nowadays, accumulating reports support the concept that proinflammatory cytokines and growth factors play crucial roles in the failing heart. We recently identified Fn14 as an endogenous key regulator in cardiac fibrosis in the PAB (Pulmonary Artery Banding) pressure-overload model. Right ventricular overload after PAB is also characterized by hypertrophy. The aim of this study was to determine whether right ventricular (RV) cardiac hypertrophy induced by PAB is mediated by the TWEAK/Fn14 axis. After baseline MRI, Fn14(-/-) mice and wild-type (WT) littermates were randomly assigned to two groups: (1) SHAM-operated (n⩾4, per genotype) and (2) PAB (n⩾11, per genotype). The results of MRI and histological analysis demonstrated that Fn14(-/-) mice exhibit less PAB-induced cardiac hypertrophy compared to WT littermates. Moreover, Fn14 overexpression in cultured adult rat cardiomyocytes enhanced cardiomyocyte size. Collectively, our studies demonstrate that Fn14 ablation attenuates RV hypertrophy after PAB and that activation of TWEAK/Fn14 signaling promotes cardiomyocyte growth in vitro. These results nominate Fn14 as a potential novel target for the treatment of heart hypertrophy., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Pulmonary arterial hypertension (PAH) is a fatal disease for which no cure is yet available. The leading cause of death in PAH is right ventricular (RV) failure. Previously, the TNF receptor superfamily member fibroblast growth factor-inducible molecule 14 (Fn14) has been associated with different fibrotic diseases. However, so far there is no study demonstrating a causal role for endogenous Fn14 signaling in RV or LV heart disease. The purpose of this study was to determine whether global ablation of Fn14 prevents RV fibrosis and remodeling improving heart function. Here, we provide evidence for a causative role of Fn14 in pulmonary artery banding (PAB)-induced RV fibrosis and dysfunction in mice. Fn14 expression was increased in the RV after PAB. Mice lacking Fn14 (Fn14(-/-)) displayed substantially reduced RV fibrosis and dysfunction following PAB compared to wild-type littermates. Cell culture experiments demonstrated that activation of Fn14 induces collagen expression via RhoA-dependent nuclear translocation of myocardin-related transcription factor-A (MRTF-A)/MAL. Furthermore, activation of Fn14 in vitro caused fibroblast proliferation and myofibroblast differentiation, which corresponds to suppression of PAB-induced RV fibrosis in Fn14(-/-) mice. Moreover, our findings suggest that Fn14 expression is regulated by endothelin-1 (ET-1) in cardiac fibroblasts. We conclude that Fn14 is an endogenous key regulator in cardiac fibrosis and suggest this receptor as potential new target for therapeutic interventions in heart failure.

Inflammation is a major factor in heart disease. IκB kinase (IKK) and its downstream target NF-κB are regulators of inflammation and are activated in cardiac disorders, but their precise contributions and targets are unclear. We analyzed IKK/NF-κB function in the heart by a gain-of-function approach, generating an inducible transgenic mouse model with cardiomyocyte-specific expression of constitutively active IKK2. In adult animals, IKK2 activation led to inflammatory dilated cardiomyopathy and heart failure. Transgenic hearts showed infiltration with CD11b(+) cells, fibrosis, fetal reprogramming, and atrophy of myocytes with strong constitutively active IKK2 expression. Upon transgene inactivation, the disease was reversible even at an advanced stage. IKK-induced cardiomyopathy was dependent on NF-κB activation, as in vivo expression of IκBα superrepressor, an inhibitor of NF-κB, prevented the development of disease. Gene expression and proteomic analyses revealed enhanced expression of inflammatory cytokines, and an IFN type I signature with activation of the IFN-stimulated gene 15 (ISG15) pathway. In that respect, IKK-induced cardiomyopathy resembled Coxsackievirus-induced myocarditis, during which the NF-κB and ISG15 pathways were also activated. Vice versa, in cardiomyocytes lacking the regulatory subunit of IKK (IKKγ/NEMO), the induction of ISG15 was attenuated. We conclude that IKK/NF-κB activation in cardiomyocytes is sufficient to cause cardiomyopathy and heart failure by inducing an excessive inflammatory response and myocyte atrophy.

Cardiomyocyte remodeling, which includes partial dedifferentiation of cardiomyocytes, is a process that occurs during both acute and chronic disease processes. Here, we demonstrate that oncostatin M (OSM) is a major mediator of cardiomyocyte dedifferentiation and remodeling during acute myocardial infarction (MI) and in chronic dilated cardiomyopathy (DCM). Patients suffering from DCM show a strong and lasting increase of OSM expression and signaling. OSM treatment induces dedifferentiation of cardiomyocytes and upregulation of stem cell markers and improves cardiac function after MI. Conversely, inhibition of OSM signaling suppresses cardiomyocyte remodeling after MI and in a mouse model of DCM, resulting in deterioration of heart function after MI but improvement of cardiac performance in DCM. We postulate that dedifferentiation of cardiomyocytes initially protects stressed hearts but fails to support cardiac structure and function upon continued activation. Manipulation of OSM signaling provides a means to control the differentiation state of cardiomyocytes and cellular plasticity., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Aims: The transcription factor FoxO3 contributes to anti-hypertrophic signalling in the heart presumably by regulating autophagic-lysosomal and ubiquitin-proteasomal pathways. We wanted to study FoxO3 function in the adult heart in vivo by expressing a constitutively active mutant of FoxO3 in transgenic mice., Methods and Results: We generated transgenic mice in which a tetracycline-regulated constitutively active FoxO3 transgene (FoxO3-CA) is controlled by the heart-specific α-myosin heavy chain promoter. Cardiac-specific expression in adult mice resulted in a decrease in heart weight by 25% and a reduction in stroke volume and cardiac output. The decrease in heart size was due to a reduction in the size of individual cardiomyocytes, whereas there was no evidence for increased cell death. FoxO3 activation was accompanied by the initiation of a foetal gene programme with increased expression of β-myosin heavy chain and natriuretic peptides, and by the activation of AKT and mammalian target of rapamycin signalling. As shown by electron microscopy, FoxO3-CA massively stimulated destruction of sarcomeres and autophagy, and induced expression of LC3-II and BNIP3. When FoxO3-CA expression was shut off in affected mice, cardiac atrophy and dysfunction as well as molecular markers were normalized within 1 month. FoxO3-CA expression did not counteract hypertrophy induced by transverse aortic constriction., Conclusion: Heart-specific expression of constitutively active FoxO3 leads to reversible heart atrophy. The reversibility of the phenotype suggests a remarkable ability of the adult myocardium to respond to different regulatory cues.

Objectives: To determine whether adult newts (Notophthalmus viridescens) are able to repair experimentally-induced joint damage in order to generate a model system for the study of endogenous joint regeneration., Methods: Joint instability and articular cartilage lesions of the knee joint of adult newts (N viridescens) were induced by intra-articular injection of collagenase. The changes over time were analysed clinically, by MRI, histologically and by reverse transcription PCR to detect selected relevant markers., Results: After rapid onset of disease with joint luxation, loss of proteoglycans and cartilage volume, the signs ameliorated continuously by regeneration of the affected joint compartments. The majority of joints were morphologically intact and functionally operative after 10 weeks. Upregulation of chondrogenic key genes, homogenous expression levels of factors implicated in cartilage homeostasis and limb regeneration as well as the distribution of the blastemal marker 22/18 in both treated and untreated knees suggest that joint regeneration in adult newts only partially invokes pathways of embryological organogenesis., Conclusions: Newts are able to regenerate articular cartilage injuries and to restore tissue integrity and function after induction of damage using a procedure known to induce experimental osteoarthritis in murine models. Further analysis of the underlying molecular mechanisms may contribute to the development of novel treatment approaches in joint failure.

The right ventricle (RV) differs in several aspects from the left ventricle (LV) including its embryonic origin, physiological role and anatomical design. In contrast to LV hypertrophy, little is known about the molecular circuits, which are activated upon RV hypertrophy (RVH). We established a highly reproducible model of RVH in mice using pulmonary artery clipping (PAC), which avoids detrimental RV pressure overload and thus allows long-term survival of operated mice. Magnetic resonance imaging revealed pathognomonic changes with striking similarities to human congenital heart disease- or pulmonary arterial hypertension-patients. Comparative, microarray based transcriptome analysis of right- and left-ventricular remodeling identified distinct transcriptional responses to pressure-induced hypertrophy of either ventricle, which were mainly characterized by stronger transcriptional responses of the RV compared to the LV myocardium. Hierarchic cluster analysis revealed a RV- and LV-specific pattern of gene activity after induction of hypertrophy, however, we did not find evidence for qualitatively distinct regulatory pathways in RV compared to LV. Data mining of nearly three thousand RV-enriched genes under PAC disclosed novel potential (co)-regulators of long-term RV remodeling and hypertrophy. We reason that specific inhibitory mechanisms in RV restrict excessive myocardial hypertrophy and thereby contribute to its vulnerability to pressure overload., (Copyright 2010 Elsevier Ltd. All rights reserved.)

Chromatin modifying enzymes play a critical role in cardiac differentiation. Previously, it has been shown that the targeted deletion of the histone methyltransferase, Smyd1, the founding member of the SET and MYND domain containing (Smyd) family, interferes with cardiomyocyte maturation and proper formation of the right heart ventricle. The highly related paralogue, Smyd2 is a histone 3 lysine 4- and lysine 36-specific methyltransferase expressed in heart and brain. Here, we report that Smyd2 is differentially expressed during cardiac development with highest expression in the neonatal heart. To elucidate the functional role of Smyd2 in the heart, we generated conditional knockout (cKO) mice harboring a cardiomyocyte-specific deletion of Smyd2 and performed histological, functional and molecular analyses. Unexpectedly, cardiac deletion of Smyd2 was dispensable for proper morphological and functional development of the murine heart and had no effect on global histone 3 lysine 4 or 36 methylation. However, we provide evidence for a potential role of Smyd2 in the transcriptional regulation of genes associated with translation and reveal that Smyd2, similar to Smyd3, interacts with RNA Polymerase II as well as to the RNA helicase, HELZ.

Circulating blood-derived vasculogenic cells improve neovascularization of ischemic tissue by a broad repertoire of potential therapeutic actions. Whereas initial studies documented that the cells incorporate and differentiate to cardiovascular cells, other studies suggested that short-time paracrine mechanisms mediate the beneficial effects. The question remains to what extent a physical incorporation is contributing to the beneficial effects of cell therapy. By using the inducible suicide gene thymidine kinase to deplete transplanted cells, we determined the contribution of physical incorporation in 3 animal models. After acute myocardial infarction, depletion of cells 14 days after infusion resulted in a reduction of capillary density and a substantial deterioration of heart function. Likewise, neovascularization of Matrigel plugs and ischemic limbs was significantly suppressed when infused cells were depleted 7 days after infusion. Induction of cell death in the previously transplanted cells reduced perfusion and led to vascular leakage as evidenced by Evans blue extravasation. These results indicate that physical incorporation and persistence of cells contribute to cell-mediated improvement of neovascularization and cardiac function. Long-term paracrine activities and/or cell intrinsic mechanisms may have contributed to the maintenance of functional improvement.