Improve Infarct Healing Research Articles

CRISPR-Cas9 editing of TLR4 to improve the outcome of cardiac cell therapy

Inflammation and fibrosis limit the reparative properties of human mesenchymal stromal cells (hMSCs). We hypothesized that disrupting the toll-like receptor 4 (TLR4) gene would switch hMSCs toward a reparative phenotype and improve the outcome of cell therapy for infarct repair. We developed and optimized an improved electroporation protocol for CRISPR-Cas9 gene editing. This protocol achieved a 68% success rate when applied to isolated hMSCs from the heart and epicardial fat of patients with ischemic heart disease. While cell editing lowered TLR4 expression in hMSCs, it did not affect classical markers of hMSCs, proliferation, and migration rate. Protein mass spectrometry analysis revealed that edited cells secreted fewer proteins involved in inflammation. Analysis of biological processes revealed that TLR4 editing reduced processes linked to inflammation and extracellular organization. Furthermore, edited cells expressed less NF-ƙB and secreted lower amounts of extracellular vesicles and pro-inflammatory and pro-fibrotic cytokines than unedited hMSCs. Cell therapy with both edited and unedited hMSCs improved survival, left ventricular remodeling, and cardiac function after myocardial infarction (MI) in mice. Postmortem histologic analysis revealed clusters of edited cells that survived in the scar tissue 28 days after MI. Morphometric analysis showed that implantation of edited cells increased the area of myocardial islands in the scar tissue, reduced the occurrence of transmural scar, increased scar thickness, and decreased expansion index. We show, for the first time, that CRISPR-Cas9-based disruption of the TLR4-gene reduces pro-inflammatory polarization of hMSCs and improves infarct healing and remodeling in mice. Our results provide a new approach to improving the outcomes of cell therapy for cardiovascular diseases.

Endothelial progenitor cells overexpressing endothelial no-synthase may improve infarct healing: Results from the enhanced angiogenic cell therapy -acute myocardial infarction (ENACT-AMI) trial

Endothelial progenitor cells overexpressing endothelial no-synthase may improve infarct healing: Results from the enhanced angiogenic cell therapy -acute myocardial infarction (ENACT-AMI) trial

YAP/TAZ deficiency reprograms macrophage phenotype and improves infarct healing and cardiac function after myocardial infarction.

Adverse cardiac remodeling after myocardial infarction (MI) causes structural and functional changes in the heart leading to heart failure. The initial post-MI pro-inflammatory response followed by reparative or anti-inflammatory response is essential for minimizing the myocardial damage, healing, and scar formation. Bone marrow-derived macrophages (BMDMs) are recruited to the injured myocardium and are essential for cardiac repair as they can adopt both pro-inflammatory or reparative phenotypes to modulate inflammatory and reparative responses, respectively. Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) are the key mediators of the Hippo signaling pathway and are essential for cardiac regeneration and repair. However, their functions in macrophage polarization and post-MI inflammation, remodeling, and healing are not well established. Here, we demonstrate that expression of YAP and TAZ is increased in macrophages undergoing pro-inflammatory or reparative phenotype changes. Genetic deletion of YAP/TAZ leads to impaired pro-inflammatory and enhanced reparative response. Consistently, YAP activation enhanced pro-inflammatory and impaired reparative response. We show that YAP/TAZ promote pro-inflammatory response by increasing interleukin 6 (IL6) expression and impede reparative response by decreasing Arginase-I (Arg1) expression through interaction with the histone deacetylase 3 (HDAC3)-nuclear receptor corepressor 1 (NCoR1) repressor complex. These changes in macrophages polarization due to YAP/TAZ deletion results in reduced fibrosis, hypertrophy, and increased angiogenesis, leading to improved cardiac function after MI. Also, YAP activation augmented MI-induced cardiac fibrosis and remodeling. In summary, we identify YAP/TAZ as important regulators of macrophage-mediated pro-inflammatory or reparative responses post-MI.

Immunotherapy for Infarcts: In Vivo Postinfarction Macrophage Modulation Using Intramyocardial Microparticle Delivery of Map4k4 Small Interfering RNA.

The myeloid cells infiltrating the heart early after acute myocardial infarction elaborate a secretome that largely orchestrates subsequent ventricular wall repair. Regulating this innate immune response could be a means to improve infarct healing. To pilot this concept, we utilized (β1,3-d-) glucan-encapsulated small interfering RNA (siRNA)-containing particles (GeRPs), targeting mononuclear phagocytes, delivered to mice as a one-time intramyocardial injection immediately after acute infarction. Findings demonstrated that cardiac macrophages phagocytosed GeRPs in vivo and had little systemic dissemination, thus providing a means to deliver local therapeutics. Acute infarcts were then injected in vivo with phosphate-buffered saline (PBS; vehicle) or GeRPs loaded with siRNA to Map4k4, and excised hearts were examined at 3 and 7 days by quantitative polymerase chain reaction, flow cytometry, and histology. Compared with infarcted PBS-treated hearts, hearts with intrainfarct injections of siRNA-loaded GeRPs exhibited 69–89% reductions in transcripts for Map4k4 (mitogen-activated protein kinase kinase kinase kinase 4), interleukin (IL)-1β, and tumor necrosis factor α at 3 days. Expression of other factors relevant to matrix remodeling—monocyte chemoattractant protein-1 (MCP-1), matrix metalloproteinases, hyaluronan synthases, matricellular proteins, and profibrotic factors transforming growth factor beta (TGF-β), and connective tissue growth factor (CTGF)—were also decreased. Most effects peaked at 3 days, but, in some instances (Map4k4, IL-1β, TGF-β, CTGF, versican, and periostin), suppression persisted to 7 days. Thus, direct intramyocardial GeRP injection could serve as a novel and clinically translatable platform for in vivo RNA delivery to intracardiac macrophages for local and selective immunomodulation of the infarct microenvironment.

Nuclear Imaging of Post-infarction Inflammation in Ischemic Cardiac Diseases - New Radiotracers for Potential Clinical Applications.

Acute myocardial infarction is one of the leading causes of death in the western world. Despite major improvements in myocardial reperfusion with sophisticated percutaneous coronary intervention technologies and new antithrombotic agents, there is still no effective therapy for preventing post- infarction myocardial injury and remodeling. Death of cardiomyocytes following ischemia results in "danger signals" that elicit an inflammatory reaction to remove cell debris and form scar tissue. Optimal healing of the damaged myocardial tissue requires a coordinated cellular response for sufficient wound healing and scar formation. However, if this inflammatory reaction is overactive or incompletely resolved, adverse left ventricular remodeling and heart failure may occur. Treatment aimed at the modulation of the post-MI inflammatory response has been widely pursued and investigated. Although improved infarct healing was shown in many experimental preclinical studies, to date, clinical trials using anti-inflammatory treatment strategies have been far less successful. Clearly, a need exists for predicting and selecting patients at risk and selecting the most appropriate therapy for individual patients. To this end, imaging of the post-MI response has been a topic of significant interest. In this review, we first discuss the clinical complications resulting from myocardial inflammation following AMI and the need for non-invasive imaging techniques using radiolabeled tracers. We then discuss the inflammatory reaction cascade following acute myocardial infarction, the inflammatory reaction cascade following acute myocardial infarction focusing on inflammatory cell types involved herein, and potential imaging targets for identifying these cells during the inflammatory process. In addition, we discuss specific characteristics and limitations of various preclinical animal models for ischemic heart disease since they are crucial in the development and evaluation of the imaging techniques. Finally, we discuss the need for non-invasive imaging approaches using radiolabeled tracers.

CD226 deletion improves post-infarction healing via modulating macrophage polarization in mice.

Macrophages are essential for wound repair after myocardial infarction (MI). CD226, a member of immunoglobulin superfamily, is expressed on inflammatory monocytes, however, the role of CD226 in infarct healing and the effect of CD226 on macrophage remain unknown.Methods: Wild type and CD226 knockout (CD226 KO) mice were subjected to permanent coronary ligation. CD226 expression, cardiac function and ventricular remodeling were evaluated. Profile of macrophages, myofibroblasts, angiogenesis and monocytes mobilization were determined.Results: CD226 expression increased in the infarcted heart, with a peak on day 7 after MI. CD226 KO attenuated infarct expansion and improved infarct healing after MI. CD226 deletion resulted in increased F4/80+ CD206+ M2 macrophages and diminished Mac-3+ iNOS+ M1 macrophages accumulation in the infarcted heart, as well as enrichment of α-smooth muscle actin positive myofibroblasts and Ki67+ CD31+ endothelial cells, leading to increased reparative collagen deposition and angiogenesis. Furthermore, CD226 deletion restrained inflammatory monocytes mobilization, as revealed by enhanced retention of Ly6Chi monocytes in the spleen associated with a decrease of Ly6Chi monocytes in the peripheral blood, whereas local proliferation of macrophage in the ischemic heart was not affected by CD226 deficiency. In vitro studies using bone marrow-derived macrophages showed that CD226 deletion potentiated M2 polarization and suppressed M1 polarization.Conclusion: CD226 expression is dramatically increased in the infarcted heart, and CD226 deletion improves post-infarction healing and cardiac function by favoring macrophage polarization towards reparative phenotype. Thus, inhibition of CD226 may represent a novel therapeutic approach to improve wound healing and cardiac function after MI.

EGF-like repeats and discoidin I-like domain 3 (EDIL3) deficiency improves post-myocardial infarction healing via neutrophil extracellular traps (NETs) mediated pro-inflammatory macrophage polarization

Abstract After myocardial infarction (MI), injured cardiomyocytes recruit neutrophils, leading to extravasated monocytes polarized to inflammatory and reparative macrophages sequentially, which contribute to the healing and regenerative process. As an endogenous leukocyte-endothelial adhesion inhibitor, the role of EDIL3 in MI remains obscure. We found that serum EDIL3 level of MI patients was negatively associated with MI biomarkers, including CK-MB, cTnI and MGB. EDIL3−/− mice showed improved infarct healing with lower mortality, better cardiac function and decreased infarcted size compared with wild type (WT). Enhanced neutrophil infiltration resulted in the increase of Ly6Chi MHClo Mertk− macrophages, which we identified to be pro-inflammatory, protecting EDIL3−/− mice hearts from excessive fibrosis. A neutrophil infiltration antagonist not only eliminated the difference in cardiac function between WT and EDIL3−/− mice, but also reduced Ly6Chi MHClo Mertk− macrophage polarization. These data implied that improved cardiac healing was associated with increased infiltration of neutrophil and it mediated macrophage polarization. Notably, we confirmed that this polarization depended on NETs, which were more abundant in hearts and serum in EDIL3−/− mice than in WT. Indeed, self-DNA released from NETs primed the pro-inflammatory polarization of macrophages. The degradation of NETs by DNase I reduced this polarization and thus affected post-MI outcome, indicating the pivotal role of NETs in MI healing. Meanwhile, the pro-inflammatory macrophages induced apoptosis of neutrophils and restrained excessive inflammation. Our findings reveal a novel neutrophil–macrophage crosstalk essential to cardiac healing after MI.

Sufentanil-medetomidine anaesthesia compared with fentanyl/fluanisone-midazolam is associated with fewer ventricular arrhythmias and death during experimental myocardial infarction in rats and limits infarct size following reperfusion.

To improve infarct healing following myocardial infarction in humans, therapeutic interventions can be applied during the inflammatory response. Animal models are widely used to study this process. However, induction of MI in rodents is associated with high mortality due to ventricular fibrillation (VF) during coronary artery ligation. The anaesthetic agent used during the procedure appears to influence the frequency of this complication. In this retrospective study, the effect on ventricular arrhythmia incidence during ligation and infarct size following in vivo reperfusion of two anaesthetic regimens, sufentanil–medetomidine (SM) and fentanyl/fluanisone–midazolam (FFM) was evaluated in rats. Anaesthetics were administered subcutaneously using fentanyl/fluanisone (0.5 mL/kg) with midazolam (5 mg/kg) (FFM group, n = 48) or sufentanil (0.05 mg/kg) with medetomidine (0.15 mg/kg) (SM group, n = 47). The coronary artery was ligated for 40 min to induce MI. Heart rate and ventricular arrhythmias were recorded during ligation, and infarct size was measured via histochemistry after three days of reperfusion. In the SM group, heart rate and VF incidence were lower throughout the experiment compared with the FFM group (6% versus 30%) (P < 0.01). Fatal VF did not occur in the SM group whereas this occurred in 25% of the animals in the FFM group. Additionally, after three days of reperfusion, the infarcted area following SM anaesthesia was less than half as large as that following FFM anaesthesia (8.5 ± 6.4% versus 20.7 ± 5.6%) (P < 0.01). Therefore, to minimize the possibility of complications related to VF and acute death arising during ligation, SM anaesthesia is recommended for experimental MI in rats.

Loss of Macrophage Wnt Secretion Improves Remodeling and Function After Myocardial Infarction in Mice.

BackgroundMacrophages and Wnt proteins (Wnts) are independently involved in cardiac development, response to cardiac injury, and repair. However, the role of macrophage‐derived Wnts in the healing and repair of myocardial infarction (MI) is unknown. We sought to determine the role of macrophage Wnts in infarct repair.Methods and ResultsWe show that the Wnt pathway is activated after MI in mice. Furthermore, we demonstrate that isolated infarct macrophages express distinct Wnt pathway components and are a source of noncanonical Wnts after MI. To determine the effect of macrophage Wnts on cardiac repair, we evaluated mice lacking the essential Wnt transporter Wntless (Wls) in myeloid cells. Significantly, Wntless‐deficient macrophages presented a unique subset of M2‐like macrophages with anti‐inflammatory, reparative, and angiogenic properties. Serial echocardiography studies revealed that mice lacking macrophage Wnt secretion showed improved function and less remodeling 30 days after MI. Finally, mice lacking macrophage‐Wntless had increased vascularization near the infarct site compared with controls.ConclusionsMacrophage‐derived Wnts are implicated in adverse cardiac remodeling and dysfunction after MI. Together, macrophage Wnts could be a new therapeutic target to improve infarct healing and repair.

Targeting and modulating infarct macrophages with hemin formulated in designed lipid-based particles improves cardiac remodeling and function

Uncontrolled activation of pro-inflammatory macrophages after myocardial infarction (MI) accelerates adverse left ventricular (LV) remodeling and dysfunction. Hemin, an iron-containing porphyrin, activates heme oxygenase-1 (HO-1), an enzyme with anti-inflammatory and cytoprotective properties. We sought to determine the effects of hemin formulated in a macrophage-targeted lipid-based carrier (denoted HA-LP) on LV remodeling and function after MI.Hemin encapsulation efficiency was ~100% at therapeutic dose levels. In vitro, hemin/HA-LP abolished TNF-α secretion from macrophages, whereas the same doses of free hemin and drug free HA-LP had no effect. Hemin/HA-LP polarized peritoneal and splenic macrophages toward M2 anti-inflammatory phenotype. We next induced MI in mice and allocated them to IV treatment with hemin/HA-LP (10mg/kg), drug free HA-LP, free hemin (10mg/kg) or saline, one day after MI. Active in vivo targeting to infarct macrophages was confirmed with HA-LP doped with PE-rhodamine. LV remodeling and function were assessed by echocardiography before, 7, and 30days after treatment. Significantly, hemin/HA-LP effectively and specifically targets infarct macrophages, switches infarct macrophages toward M2 anti-inflammatory phenotype, improves angiogenesis, reduces scar expansion and improves infarct-related regional function.In conclusion, macrophage-targeted lipid-based drug carriers with hemin switch macrophages into an anti-inflammatory phenotype, and improve infarct healing and repair. Our approach presents a novel strategy to modulate inflammation and improve infarct repair.

Microstructural characterization of myocardial infarction with optical coherence tractography and two-photon microscopy.

Myocardial infarction leads to complex changes in the fiber architecture of the heart. Here, we present a novel optical approach to characterize these changes in intact hearts in three dimensions. Optical coherence tomography (OCT) was used to derive a depth‐resolved field of orientation on which tractography was performed. Tractography of healthy myocardium revealed a smooth linear transition in fiber inclination or helix angle from the epicardium to endocardium. Conversely, in infarcted hearts, no coherent microstructure could be identified in the infarct with OCT. Additional characterization of the infarct was performed by the measurement of light attenuation and with two‐photon microscopy. Myofibers were imaged using autofluorescence and collagen fibers using second harmonic generation. This revealed the presence of two distinct microstructural patterns in areas of the infarct with high light attenuation. In the presence of residual myofibers, the surrounding collagen fibers were aligned in a coherent manner parallel to the myofibers. In the absence of residual myofibers, the collagen fibers were randomly oriented and lacked any microstructural coherence. The presence of residual myofibers thus exerts a profound effect on the microstructural properties of the infarct scar and consequently the risk of aneurysm formation and arrhythmias. Catheter‐based approaches to segment and image myocardial microstructure in humans are feasible and could play a valuable role in guiding the development of strategies to improve infarct healing.

UM206, a selective Frizzled antagonist, attenuates adverse remodeling after myocardial infarction in swine

Modulation of Wnt/Frizzled signaling with UM206 reduced infarct expansion and prevented heart failure development in mice, an effect that was accompanied by increased myofibroblast presence in the infarct, suggesting that Wnt/Frizzled signaling has a key role in cardiac remodeling following myocardial infarction (MI). This study investigated the effects of modulation of Wnt/Frizzled signaling with UM206 in a swine model of reperfused MI. For this purpose, seven swine with MI were treated with continuous infusion of UM206 for 5 weeks. Six control swine were treated with vehicle. Another eight swine were sham-operated. Cardiac function was determined by echo in awake swine. Infarct mass was estimated at baseline by heart-specific fatty acid-binding protein ELISA and at follow-up using planimetry. Components of Wnt/Frizzled signaling, myofibroblast presence, and extracellular matrix were measured at follow-up with qPCR and/or histology. Results show that UM206 treatment resulted in a significant decrease in infarct mass compared with baseline (−41±10%), whereas infarct mass remained stable in the Control-MI group (+3±17%). Progressive dilation of the left ventricle occurred in the Control-MI group between 3 and 5 weeks after MI, while adverse remodeling was halted in the UM206-treated group. mRNA expression for Frizzled-4 and the Frizzled co-receptor LRP5 was increased in UM206-treated swine as compared with Control-MI swine. Myofibroblast presence was significantly lower in infarcted tissue of the UM206-treated animals (1.53±0.43% vs 3.38±0.61%) at 5 weeks follow-up. This study demonstrates that UM206 treatment attenuates adverse remodeling in a swine model of reperfused MI, indicating that Wnt/Frizzled signaling is a promising target to improve infarct healing and limit post-MI remodeling.

Silencing collapsin response mediator protein-2 reprograms macrophage phenotype and improves infarct healing in experimental myocardial infarction model.

BackgroundDelayed M1 toward M2 macrophage phenotype transition is considered one of the major causes for the impaired healing after myocardial infarction (MI). While searching for molecules that modulate M1 and M2 macrophage polarization, we identified collapsin response mediator protein-2 (CRMP2) as a novel molecule involved in macrophage polarization to M1. In this study, we evaluated the effect of silencing CRMP2 on macrophage polarization, inflammation and fibrosis post myocardial infarction.MethodsCRMP2 expression was assessed with Western blotting or immunohistochemistry. Macrophage phenotypes were measured with flow cytometry, quantitative real-time PCR (qPCR), Western blotting or immunohistochemistry. CRMP2 siRNA was delivered into the macrophages infiltrated in the wound of ApoE−/− mice through lipidoid nanoparticle, and fibrosis, leukocyte infiltration and inflammation parameters were measured with qPCR. Infarct size was measured with Masson’s trichrome staining. Echocardiography was performed to assess ventricular systolic dimension, left ventricular diastolic dimension, anterior wall thickness and posterior wall thickness. Student’s t-test (for 2 groups) and ANOVA (for > 2 groups) were used for statistical analyses.ResultsCRMP2 was expressed in a higher level in M1 macrophages than M2 subsets, and CRMP2 RNA interference (RNAi) resulted in a switch of bone marrow-derived macrophages from M1 to M2 phenotype. High level of CRMP2 was also observed in the macrophages infiltrated in the infarct area 3 days post MI in both wildtype (WT) and ApoE−/− mice, and the expression of CRMP2 retained in the infiltrated macrophages of ApoE−/− mice but not in that of WT mice 10 days after MI. Nanoparticle-mediated delivery of CRMP2 siRNA to ApoE−/− mice with MI resulted in dramatic switch of wound macrophages from M1 to M2 phenotype, marked decrease in inflammation and fibrosis, and significant attenuation of post-MI heart failure and mortality.ConclusionCRMP2 is highly expressed in M1 macrophages and silencing CRMP2 reprograms macrophage phenotype and improves infarct healing in atherosclerotic mice.

Positive effect of intravenous iron-oxide administration on left ventricular remodelling in patients with acute ST-elevation myocardial infarction – A cardiovascular magnetic resonance (CMR) study

ObjectivesThis study investigated the safety profile and potential “therapeutic” effect of intravenous ultrasmall superparamagnetic iron-oxide (USPIO)-based iron administration regarding infarct healing in patients with ST-elevation myocardial infarction (STEMI). USPIO-administration was recently shown to enable an improved characterization of myocardial infarct pathology in acute STEMI patients. Materials and MethodsSeventeen study patients (IRON, 54±9yrs, 88% male) and 22 matched controls (CONTROL, 57±9yrs, 77% male) both with primary reperfused STEMI underwent multi-parametric CMR studies in the first week and three months after acute MI. Only IRON patients received a single intravenous bolus of 510mg elemental iron as ferumoxytol (FerahemeTM) within four days following acute MI. ResultsThree months later, all patients were alive and there were no adverse cardiac events. Significant improvement in left ventricular (LV) ejection fraction (IRON: 53±10% to 59±9%, p=0.002; CONTROL: 54±6% to 57±10%, p=0.005) as well as shrinkage of infarct size were seen in both groups at follow-up. There was a more pronounced decrease in infarct size in the IRON group (IRON: −10.3±5.4% vs. CONTROL: −7.0±8.4%, p=0.050) in addition to a significant decrease in both endocardial extent and prevalence of transmural infarctions in IRON but not in CONTROL patients. A significant decrease in LV end systolic volume was only seen in the IRON group (71±25mL to 59±25mL, p=0.002). ConclusionsIntravenous iron administration in acute STEMI patients seems to be associated with an improved infarct healing and a beneficial global left ventricular remodelling. These findings together with the good safety profile make USPIO-based iron administration a promising future candidate as a “diagnostic” and “therapeutic” adjunctive solution in acute MI management.

In Vivo Silencing of the Transcription Factor IRF5 Reprograms the Macrophage Phenotype and Improves Infarct Healing

The aim of this study was to test whether silencing of the transcription factor interferon regulatory factor 5 (IRF5) in cardiac macrophages improves infarct healing and attenuates post-myocardial infarction (MI) remodeling. In healing wounds, the M1 toward M2 macrophage phenotype transition supports resolution of inflammation and tissue repair. Persistence of inflammatory M1 macrophages may derail healing and compromise organ functions. The transcription factor IRF5 up-regulates genes associated with M1 macrophages. Here we used nanoparticle-delivered small interfering ribonucleic acid (siRNA) to silence IRF5 in macrophages residing in MIs and in surgically-induced skin wounds in mice. Infarct macrophages expressed high levels of IRF5 during the early inflammatory wound-healing stages (day 4 after coronary ligation), whereas expression of the transcription factor decreased during the resolution of inflammation (day 8). Following in vitro screening, we identified an siRNA sequence that, when delivered by nanoparticles to wound macrophages, efficiently suppressed expression of IRF5 in vivo. Reduction of IRF5 expression, a factor that regulates macrophage polarization, reduced expression of inflammatory M1 macrophage markers, supported resolution of inflammation, accelerated cutaneous and infarct healing, and attenuated development of post-MI heart failure after coronary ligation as measured by protease targeted fluorescence molecular tomography-computed tomography imaging and cardiac magnetic resonance imaging (p < 0.05). This work identified a new therapeutic avenue to augment resolution of inflammation in healing infarcts by macrophage phenotype manipulation. This therapeutic concept may be used to attenuate post-MI remodeling and heart failure.

Macrophage Subpopulations Are Essential for Infarct Repair With and Without Stem Cell Therapy

This study sought to investigate the hypothesis that the favorable effects of mesenchymal stromal cells (MSCs) on infarct repair are mediated by macrophages. The favorable effects of MSC therapy in myocardial infarction (MI) are complex and not fully understood. We induced MI in mice and allocated them to bone marrow MSCs, mononuclear cells, or saline injection into the infarct, with and without early (4 h before MI) and late (3 days after MI) macrophage depletion. We then analyzed macrophage phenotype in the infarcted heart by flow cytometry and macrophage secretome in vitro. Left ventricular remodeling and global and regional function were assessed by echocardiography and speckle-tracking based strain imaging. The MSC therapy significantly increased the percentage of reparative M2 macrophages (F4/80(+)CD206(+)) in the infarcted myocardium, compared with mononuclear- and saline-treated hearts, 3 and 4 days after MI. Macrophage cytokine secretion, relevant to infarct healing and repair, was significantly increased after MSC therapy, or incubation with MSCs or MSC supernatant. Significantly, with and without MSC therapy, transient macrophage depletion increased mortality 30 days after MI. Furthermore, early macrophage depletion produced the greatest negative effect on infarct size and left ventricular remodeling and function, as well as a significant incidence of left ventricular thrombus formation. These deleterious effects were attenuated with macrophage restoration and MSC therapy. Some of the protective effects of MSCs on infarct repair are mediated by macrophages, which are essential for early healing and repair. Thus, targeting macrophages could be a novel strategy to improve infarct healing and repair.

Selective Homocysteine Lowering Gene Transfer Improves Infarct Healing, Attenuates Remodelling, and Enhances Diastolic Function after Myocardial Infarction in Mice

Background and aimsHomocysteine levels predict heart failure incidence in prospective epidemiological studies and correlate with severity of heart failure in cross-sectional surveys. The objective of this study was to evaluate whether a selective homocysteine lowering intervention beneficially affects cardiac remodelling and cardiac function after myocardial infarction (MI) in a murine model of combined hypercholesterolemia and hyperhomocysteinemia.Methodology and principal findingsA selective homocysteine lowering gene transfer strategy was evaluated in female C57BL/6 low density lipoprotein receptor (Ldlr)−/− cystathionine-ß-synthase (Cbs)+/− deficient mice fed a hyperhomocysteinemic and high saturated fat/high cholesterol diet using an E1E3E4-deleted hepatocyte-specific adenoviral vector expressing Cbs (AdCBS). MI was induced by permanent ligation of the left anterior descending coronary artery 14 days after saline injection or gene transfer. AdCBS gene transfer resulted in a persistent more than 5-fold (p<0.01) decrease of plasma homocysteine levels and significantly improved endothelial progenitor cell function. Selective homocysteine lowering enhanced infarct healing as indicated by a 21% (p<0.01) reduction of infarct length at day 28 after MI and by an increased number of capillaries and increased collagen content in the infarct zone. Adverse remodelling was attenuated in AdCBS MI mice as evidenced by a 29% (p<0.05) reduction of left ventricular cavity area at day 28, by an increased capillary density in the remote myocardium, and by reduced interstitial collagen. The peak rate of isovolumetric relaxation was increased by 19% (p<0.05) and the time constant of left ventricular relaxation was reduced by 21% (p<0.05) in AdCBS MI mice compared to control MI mice, indicating improved diastolic function.Conclusion/significanceSelective homocysteine lowering gene transfer improves infarct healing, attenuates remodelling, and significantly enhances diastolic function post-MI in female C57BL/6 Ldlr −/− Cbs +/− mice. The current study corroborates the view that hyperhomocysteinemia exerts direct effects on the myocardium and may potentiate the development of heart failure.

931Beneficial effect of intravenous iron administration on left ventricular remodelling in patients with acute ST-elevation myocardial infarction – a cardiovascular magnetic resonance (CMR) study

Background: The infusion of iron has been associated with improved symptoms, functional capacity, and quality of life in patients with heart failure and chronic myocardial infarction – even in the absence of anemia. Objectives: We aimed to investigate the safety profile and effect of intravenous iron administration on short-term ventricular remodeling in patients with myocardial infarction in the acute setting. Material and methods: Seventeen study patients (IRON, 54 ± 9 yrs, 88% male) and 22 matched controls (CONTROL, 57 ± 9 yrs, 77% male) both with primary reperfused ST-elevation myocardial infarctions (STEMI) underwent cine and late gadolinium enhancement cardiovascular magnetic resonance (CMR) imaging in the first week and three months after the acute event. IRON patients received a unique intravenous bolus of 510 mg elemental iron as ferumoxytol (Feraheme®) in the 24 hours following the first CMR exam. Results: Ferumoxytol administration was well tolerated by all patients, with no adverse events. No significant differences regarding baseline infarct characteristics or ventricular function were present between groups. At 3 months follow-up, all patients were alive, with no new adverse cardiac events. In both groups significant changes in left ventricular ejection fraction (IRON: 53 ± 10 to 59 ± 9%, p = 0.002; CONTROL: 54 ± 6 to 57 ± 10, p = 0.005), LV mass index (IRON: 65, 59-76 gr/m2 to 54, 38-114 gr/m2, p = 0.0006; CONTROL: 65, 58-92 gr/m2 to 62, 49-74 gr/m2, p<0.0001) and infarct size (IRON: 27 ± 8 to 17 ± 9%, p<0.0001; CONTROL: 27 ± 12 to 20 ± 11, p<0.0001) were seen at follow-up with a clear (but non-significant) trend towards a higher increase in ejection fraction and smaller scars in the IRON group. In addition, in IRON but not in CONTROL patients, significant decreases in LV end systolic volume (71 ± 25 to 59 ± 25 mL, p = 0.002) and transmural infarctions (17, 100% to 7, 41%, p = 0.0003) were encountered. Conclusion: Intravenous iron administration in acute STEMI patients is associated with both improved infarct healing and beneficial global left ventricular remodeling compared to controls. Moreover, a good safety profile for iron administration was observed.

Monocyte-Directed RNAi Targeting CCR2 Improves Infarct Healing in Atherosclerosis-Prone Mice

Exaggerated and prolonged inflammation after myocardial infarction (MI) accelerates left ventricular remodeling. Inflammatory pathways may present a therapeutic target to prevent post-MI heart failure. However, the appropriate magnitude and timing of interventions are largely unknown, in part because noninvasive monitoring tools are lacking. Here, we used nanoparticle-facilitated silencing of CCR2, the chemokine receptor that governs inflammatory Ly-6C(high) monocyte subset traffic, to reduce infarct inflammation in apolipoprotein E-deficient (apoE(-/-)) mice after MI. We used dual-target positron emission tomography/magnetic resonance imaging of transglutaminase factor XIII (FXIII) and myeloperoxidase (MPO) activity to monitor how monocyte subset-targeted RNAi altered infarct inflammation and healing. Flow cytometry, gene expression analysis, and histology revealed reduced monocyte numbers and enhanced resolution of inflammation in infarcted hearts of apoE(-/-) mice that were treated with nanoparticle-encapsulated siRNA. To follow extracellular matrix cross-linking noninvasively, we developed a fluorine-18-labeled positron emission tomography agent ((18)F-FXIII). Recruitment of MPO-rich inflammatory leukocytes was imaged with a molecular magnetic resonance imaging sensor of MPO activity (MPO-Gd). Positron emission tomography/magnetic resonance imaging detected anti-inflammatory effects of intravenous nanoparticle-facilitated siRNA therapy (75% decrease of MPO-Gd signal; P<0.05), whereas (18)F-FXIII positron emission tomography reflected unimpeded matrix cross-linking in the infarct. Silencing of CCR2 during the first week after MI improved ejection fraction on day 21 after MI from 29% to 35% (P<0.05). CCR2-targeted RNAi reduced recruitment of Ly-6C(high) monocytes, attenuated infarct inflammation, and curbed post-MI left ventricular remodeling.

Pre-clinical data on involvement of mineralocorticoid receptor activation in healing and remodelling post-myocardial infarction

Myocardial infarction (MI) is one of the leading causes of death. Therapeutic strategies that target pathophysiological mechanisms of infarct healing and that improve cardiac remodeling after MI appear to be useful tools to prevent progressive myocardial dysfunction, heart failure and sudden death. Recent preclinical data highlight that activation of the mineralocorticoid receptor (MR) plays a pivotal role in post-infarct healing and remodelling. Immediate MR blockade after MI enhanced infarct neovascularization and reduced early myocardial dilatation and dysfunction. Moreover, cardiomyocyte-specific MR deficiency improved infarct healing and prevented progressive adverse cardiac remodeling, contractile dysfunction, and molecular alterations in ischemic heart failure. This review summarizes new insights into the involvement of MR activation in wound healing and remodelling after myocardial infarction.