Infarct Healing Research Articles

Effect of remote ischaemic conditioning on left ventricular function in ST-segment elevation myocardial infarction patients: The CONDI-2 echocardiographic sub-study.

Remote ischaemic conditioning (RIC) applied to the arm by inflation and deflation of a pneumatic cuff has been shown to reduce myocardial infarct size in patients with ST-elevation myocardial infarction undergoing primary percutaneous coronary intervention (PPCI). However, the effect of RIC on left ventricular ejection fraction (LVEF) following infarct healing remains unknown. To investigate whether RIC applied in the ambulance before PPCI can improve left ventricular (LV) function in STEMI patients 3 months following infarction. Echocardiography was performed in a total of 694 patients from the CONDI-2 study a median of 112 days (IQR 63) after the initial admission. LVEF and LV end-diastolic and end-systolic volumes were calculated using the modified Simpsons biplane method of disks. LV global longitudinal strain (GLS) was estimated using 2-dimensional cine-loops with a frame rate > 55 frames/second, measured in the three standard apical views. There was no difference in the measured echocardiographic parameters in the RIC group as compared to the control group, including LV EF, LV GLS, tricuspid annular plane systolic excursion or left ventricular volumes. In the control group, 32% had an ejection fraction < 50% compared to 37% in the RIC group (p = 0.129). In this largest to date randomized imaging study of RIC, RIC as an adjunct to PPCI was not associated with a change in echocardiographic measures of cardiac function compared to standard PPCI alone.

Extracellular Vesicles from NMN Preconditioned Mesenchymal Stem Cells Ameliorated Myocardial Infarction via miR-210-3p Promoted Angiogenesis

Mesenchymal stem cell-derived extracellular vesicles (MSCs-EVs) possess cardioprotection in acute myocardial infarction. Nevertheless, the therapeutic intervention potential and the molecular mechanism of EVs from NMN (Nicotinamide mononucleotide) preconditioned hUCMSCs (N-EVs) in acute myocardial infarction remains unknown. In the present study, EVs from hUCMSCs (M-EVs) and N-EVs were identified by electron microscopy, immunoblotting and nanoparticle tracking analysis. Compared with M-EVs, N-EVs significantly increased the proliferation, migration, and angiogenesis of HUVECs. Meanwhile, N-EVs markedly reduced apoptosis and cardiac fibrosis and promoted angiogenesis in the peri-infarct region in the MI rats. A high-throughput miRNA sequencing and qPCR methods analysis revealed that miR-210-3p was abundant in N-EVs and the expression of miR-210-3p was obviously upregulated in HUVECs after N-EVs treated. Overexpression of miR-210-3p in HUVECs significantly enhanced the tube formation, migration and proliferative capacities of HUVECs. However, downregulation of miR-210-3p in HUVECs markedly decreased the tube formation, migration and proliferative capacities of HUVECs. Furthermore, bioinformatics analysis and luciferase assays revealed that EphrinA3 (EFNA3) was a direct target of miR-210-3p. Knockdown of miR-210-3p in N-EVs significantly impaired its ability to protect the heart after myocardial infarction. Altogether, these results indicated that N-EVs promoted the infarct healing through improvement of angiogenesis by miR-210-3p via targeting the EFNA3. Graphical Created with Biorender.com.

Electrophysiological and histological characterization of atrial scarring in a model of isolated atrial myocardial infarction.

Background: Characterization of atrial myocardial infarction is hampered by the frequent concurrence of ventricular infarction. Theoretically, atrial infarct scarring could be recognized by multifrequency tissue impedance, like in ventricular infarction, but this remains to be proven. Objective: This study aimed at developing a model of atrial infarction to assess the potential of multifrequency impedance to recognize areas of atrial infarct scar. Methods: Seven anesthetized pigs were submitted to transcatheter occlusion of atrial coronary branches arising from the left coronary circumflex artery. Six weeks later the animals were anesthetized and underwent atrial voltage mapping and multifrequency impedance recordings. The hearts were thereafter extracted for anatomopathological study. Two additional pigs not submitted to atrial branch occlusion were used as controls. Results: Selective occlusion of the atrial branches induced areas of healed infarction in the left atrium in 6 of the 7 cases. Endocardial mapping of the left atrium showed reduced multi-frequency impedance (Phase angle at 307kHz: from -17.1° ± 5.0° to -8.9° ± 2.6°, p < .01) and low-voltage of bipolar electrograms (.2 ± 0.1mV vs. 1.9 ± 1.5mV vs., p < .01) in areas affected by the infarction. Data variability of the impedance phase angle was lower than that of bipolar voltage (coefficient of variability of phase angle at307kHz vs. bipolar voltage: .30 vs. .77). Histological analysis excluded the presence of ventricular infarction. Conclusion: Selective occlusion of atrial coronary branches permits to set up a model of selective atrial infarction. Atrial multifrequency impedance mapping allowed recognition of atrial infarct scarring with lesser data variability than local bipolar voltage mapping. Our model may have potential applicability on the study of atrial arrhythmia mechanisms.

UpStreAm doxycycline in ST-eLeVation myocArdial infarction: targetinG infarct hEaling and ModulatIon (SALVAGE-MI trial).

Experimental studies demonstrate protective effects of doxycycline on myocardial ischaemia-reperfusion injury. The trial investigated whether doxycycline administered prior to reperfusion in patients presenting with ST-elevation myocardial infarction (STEMI) reduces infarct size (IS) and ameliorates adverse left ventricular (LV) remodelling. In this randomized, double-blind, placebo-controlled trial, patients presenting with STEMI undergoing primary percutaneous coronary intervention (PPCI) were randomized to either intravenous doxycycline or placebo prior to reperfusion followed by 7 days of oral doxycycline or placebo. The primary outcome was final IS adjusted for area-at-risk (fIS/AAR) measured on two cardiac magnetic resonance scans ∼6 months apart. Of 103 participants, 50 were randomized to doxycycline and 53 to placebo and were matched for age (59 ± 12 vs. 60 ± 10 years), male sex (92% vs. 79%), diabetes mellitus (26% vs. 11%) and left anterior descending artery occlusion (50% vs. 49%), all P > 0.05. Patients treated with doxycycline had a trend for larger fIS/AAR [0.79 (0.5-0.9) vs. 0.61 (0.47-0.76), P = 0.06], larger fIS at 6 months [18.8% (12-26) vs. 13.6% (11-21), P = 0.08], but similar acute IS [21.7% (17-34) vs. 19.4% (14-27), P = 0.19] and AAR [26% (20-36) vs. 24.7% (16-31), P = 0.22] compared with placebo. Doxycycline did not ameliorate adverse LV remodelling [%Δend-diastolic volume index, 1.1% (-3.8-8.4) vs. -1.34% (-6.1-5.8), P = 0.42] and was independently associated with larger fIS (regression coefficient = 0.175, P = 0.03). Doxycycline prior to PPCI neither reduced IS acutely or at six months nor attenuated adverse LV remodelling. These data raise safety concerns regarding doxycycline use in STEMI for infarct modulation and healing.

Gasotransmitter-Induced Therapeutic Angiogenesis: A Biomaterial Prospective.

Gasotransmitters such as NO, H2S, and CO have emerged as key players in the regulation of various pathophysiological functions, prompting the development of gas therapy for various pathogeneses. Deficient production of gasotransmitters has been linked to various diseases such as hypertension, endothelial dysfunction, myocardial infarction, ischemia, and impaired wound healing, as they are involved in the regulatory action of angiogenesis. A better understanding of the regulatory mechanisms has given new hope to address the vascular impairment caused by the breakthroughs in gasotransmitters as therapeutics. However, the unstable nature and poor target specificity of gas donors limit the full efficacy of drugs. In this regard, biomaterials that possess excellent biocompatibility and porosity are ideal drug carriers to deliver the gas transmitters in a tunable manner for therapeutic angiogenesis. This review article provides a comprehensive discussion of biomaterial-based gasotransmitter delivery approaches for therapeutic angiogenesis. The critical role of gasotransmitters in modulating angiogenesis during tissue repair as well as their challenges and future directions are demonstrated.

Intramyocardial hemorrhage drives fatty degeneration of infarcted myocardium

Sudden blockage of arteries supplying the heart muscle contributes to millions of heart attacks (myocardial infarction, MI) around the world. Although re-opening these arteries (reperfusion) saves MI patients from immediate death, approximately 50% of these patients go on to develop chronic heart failure (CHF) and die within a 5-year period; however, why some patients accelerate towards CHF while others do not remains unclear. Here we show, using large animal models of reperfused MI, that intramyocardial hemorrhage - the most damaging form of reperfusion injury (evident in nearly 40% of reperfused ST-elevation MI patients) - drives delayed infarct healing and is centrally responsible for continuous fatty degeneration of the infarcted myocardium contributing to adverse remodeling of the heart. Specifically, we show that the fatty degeneration of the hemorrhagic MI zone stems from iron-induced macrophage activation, lipid peroxidation, foam cell formation, ceroid production, foam cell apoptosis and iron recycling. We also demonstrate that timely reduction of iron within the hemorrhagic MI zone reduces fatty infiltration and directs the heart towards favorable remodeling. Collectively, our findings elucidate why some, but not all, MIs are destined to CHF and help define a potential therapeutic strategy to mitigate post-MI CHF independent of MI size.

Agomir of miRNA-30d – a potential new therapeutic target for prevention of ischemic cardiomyopathy after myocardial infarction?

Abstract Background MicroRNA (miR)-30d is not only a valuable biomarker for assessing the extent of cardiac remodeling after myocardial infarction (MI), but also an important predictor of clinical outcome in heart failure. Overexpression of miRNA-30d appears to have a cardioprotective effect by preventing cardiomyocyte apoptosis as well as averting cardiac fibroblast proliferation via multiple molecular pathways. The aim of the present in vivo and in vitro study was to demonstrate whether an miR-30d can be a potential therapeutic target to reduce the risk of ischemic cardiomyopathy (iCMP) after MI. Methods First of all, miRNA profiling was performed by next generation sequencing (NGS) to assess differences in miRNA expression in ischemic vs. healthy myocardium in a rat model of MI using coronary artery ligation (ischemia/reperfusion injury, IR). MiR-30d was selected as the most promising target as it was significantly downregulated in ischemic myocardium and can be upregulated by cardioprotective agents. Therefore, an agomir of miR-30d was administered in the respective treatment group intraperitoneally, whereas non-functional, scrambled miRNA was administered in the control group. To analyze the ratio between phosphorylated p53 (pp53) and total p53, apoptosis was evaluated in human cardiomyocytes using a p53 and pp53 ELISA kit. To gain indirect insight into infarct healing, scratch assays were used to obtain information on cell migration in human umbilical vein endothelial cells (HUVEC) in vitro. Six weeks after the in vivo induction of acute MI/IR with consequential iCMP in a rat model, the extent of MI was evaluated by planimetry. Results The majority of miRNAs studied here showed significant up-regulation in the MI-induced heart tissue in comparison to the sham operated controls. In contrast, miRNA-30d was highly significantly reduced (p&amp;lt;0.001). Based on these investigations and the already repeatedly documented cardioprotective effect of miR-30d overexpression, an agomir was selected as a potential therapy target. Human cardiomyocytes under the influence of an agomir of miR-30d showed a decreased pp53/total p53 ratio (0.66±0.09 vs. 0.81±0.19) and thus a distinct tendency (p=0.055) for a reduction in apoptotic rate compared to the control group. In HUVECs, gap closure was significantly faster in the agomir treated cells 20h and 26h post-scratching (19.1% more than scrambled control after 20h; p=0.0028 and 18.7% more than scrambled control after 26h; p=0.0081). In the in vivo model, infarct size of left ventricle was significantly reduced by using the agomir (7.43±4.13% vs. 12.76±4.76%; p=0.0172). Conclusion Using an agomir of miR-30d underlines the cardioprotective effects of miR-30d in MIR/IR and could reduce the risk for iCMP development. Further investigations regarding its therapeutic potential in the human should be considered, as microRNA treatments are gaining more and more clinical applicability today. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Paracelsus Medical University, PMU-FFF

Effect of Pulsed-Field and Radiofrequency Ablation on Heterogeneous Ventricular Scar in a Swine Model of Healed Myocardial Infarction.

Pulsed-field ablation (PFA) is a nonthermal energy with higher selectivity to myocardial tissue in comparison to radiofrequency ablation (RFA). We compared the effects of PFA and RFA on heterogeneous ventricular scar in a swine model of healed infarction. In 9 swine, myocardial infarction was created by balloon occlusion of the left anterior descending artery. After a survival period of 8 to 10 weeks, ablation with PFA or RFA was performed at infarct border zones identified by abnormal electrograms. In the PFA group (4 swine), ablation was performed with a lattice catheter (Sphere-9, Affera, Inc). In the RFA group (5 swine), ablation was performed using a 3.5-mm tip catheter (Thermocool ST-SF; Biosense Webster). To further investigate the effect of RFA on temperature development in scar tissue, intramyocardial temperature was measured in healthy and infarcted myocardium using an ex vivo bath model. A total of 11 PFA and 15 RFA lesions were created at infarct border zones with heterogeneous scar. PFA produced uniform and well-demarcated lesions exhibiting irreversible injury characterized by cardiomyocyte death, contraction bands, and lymphocytic infiltration. This effect of PFA extended from the subendocardium through collagen and fat to the epicardial layers. In contrast, the effect of RFA is less uniform and largely limited to the subendocardium with minimal effect on viable myocardium deeper to separating layers of collagen and fat. PFA produced deeper and more transmural lesions (6.4 [interquartile range, 5.5-7.5) versus 5.4 [interquartile range, 4.8-5.9]), 72% versus 30%, respectively; P≤0.02 for each comparison). The limited effect of RFA on viable myocardium at deeper infarct layers was related to a lower intramyocardial maximal temperature compared with healthy myocardium (P=0.01). PFA may be advantageous for ablation in ventricular scar, producing lesions that unlike RFA are not limited to the subendocardium, but also eliminate viable myocardium separated from the catheter by collagen and fat.

Macrophages in myocardial infarction.

The heart contains a population of resident macrophages that markedly expands following injury through recruitment of monocytes and through proliferation of macrophages. In myocardial infarction, macrophages have been implicated in both injurious and reparative responses. In coronary atherosclerotic lesions, macrophages have been implicated in disease progression and in the pathogenesis of plaque rupture. Following myocardial infarction, resident macrophages contribute to initiation and regulation of the inflammatory response. Phagocytosis and efferocytosis are major functions of macrophages during the inflammatory phase of infarct healing, and mediate phenotypic changes, leading to acquisition of an anti-inflammatory macrophage phenotype. Infarct macrophages respond to changes in the cytokine content and extracellular matrix composition of their environment and secrete fibrogenic and angiogenic mediators, playing a central role in repair of the infarcted heart. Macrophages may also play a role in scar maturation and may contribute to chronic adverse remodeling of noninfarcted segments. Single cell studies have revealed a remarkable heterogeneity of macrophage populations in infarcted hearts; however, the relations between transcriptomic profiles and functional properties remain poorly defined. This review manuscript discusses the fate, mechanisms of expansion and activation, and role of macrophages in the infarcted heart. Considering their critical role in injury, repair, and remodeling, macrophages are important, but challenging, targets for therapeutic interventions in myocardial infarction.

Limb-Bud and Heart (LBH) Upregulation in Cardiomyocytes under Hypoxia Promotes the Activation of Cardiac Fibroblasts via Exosome Secretion.

The activation of cardiac fibroblasts (CFs) after myocardial infarction (MI) is essential for post-MI infarct healing, during which the regulation of transforming growth factor beta1 (TGF-β1) signaling is predominant. We have demonstrated that TGF-β1-mediated upregulation of LBH contributes to post-MI CF activation via modulating αB-crystallin (CRYAB), after being upregulated by TGF-β1. In this study, the effect of LBH-CRYAB signaling on the cardiac microenvironment via exosome communication and the corresponding mechanisms were investigated. The upregulation of LBH and CRYAB was verified in both cardiomyocytes (CMs) and CFs in hypoxic, post-MI peri-infarct tissues. CM-derived exosomes were isolated and identified, and LBH distribution was elevated in exosomes derived from LBH-upregulated CMs under hypoxia. Treatment with LBH+ exosomes promoted cellular proliferation, differentiation, and epithelial-mesenchymal transition-like processes in CFs. Additionally, in primary LBHKO CFs, western blotting showed that LBH knockout partially inhibited TGF-β1-induced CF activation, while LBH-CRYAB signaling affected TGF-β1 expression and secretion through a positive feedback loop. The administration of a Smad3 phosphorylation inhibitor to LBHKO CFs under TGF-β1 stimulation indicated that Smad3 phosphorylation partially accounted for TGF-β1-induced LBH upregulation. In conclusion, LBH upregulation in CMs in post-MI peri-infarct areas correlated with a hypoxic cardiac microenvironment and led to elevated exosomal LBH levels, promoting the activation of recipient CFs, which brings new insights into the studies and therapeutic strategies of post-MI cardiac repair.

Insulin-Like Growth Factor 1 Attenuates the Pro-Inflammatory Phenotype of Neutrophils in Myocardial Infarction.

Acute myocardial infarction (MI) induces an extensive sterile inflammation, which is dominated in the early phase by invading neutrophils and monocytes/macrophages. The inflammatory response after MI critically affects infarct healing and cardiac remodeling. Therefore, modulation of cardiac inflammation may improve outcome post MI. Insulin-like growth factor 1 (IGF1) treatment reduces infarct size and improves cardiac function after MI via IGF1 receptor mediated signaling in myeloid cells. Our study aimed to investigate the effect of IGF1 on neutrophil phenotype both in vitro and in vivo after MI. We show that IGF1 induces an anti-inflammatory phenotype in bone marrow derived neutrophils. On the molecular and functional level IGF1 treated neutrophils were indistinguishable from those induced by IL4. Surprisingly, insulin, even though it is highly similar to IGF1 did not create anti-inflammatory neutrophils. Notably, the IGF1 effect was independent of the canonical Ras/Raf/ERK or PI3K/AKT pathway, but depended on activation of the JAK2/STAT6 pathway, which was not activated by insulin treatment. Single cell sequencing analysis 3 days after MI also showed that 3 day IGF1 treatment caused a downregulation of pro-inflammatory genes and upstream regulators in most neutrophil and many macrophage cell clusters whereas anti-inflammatory genes and upstream regulators were upregulated. Thus, IGF1 acts like an anti-inflammatory cytokine on myeloid cells in vitro and attenuates the pro-inflammatory phenotype of neutrophils and macrophages in vivo after MI. IGF1 treatment might therefore represent an effective immune modulatory therapy to improve the outcome after MI.

Evolution of Myocardial Tissue Injury: A CMR Study Over a Decade After STEMI

BackgroundIn patients with a first ST-segment elevation myocardial infarction (STEMI), the multi-annual evolution of myocardial tissue injury parameters, as assessed by cardiac magnetic resonance (CMR), has not yet been described. ObjectivesThis study examined myocardial tissue injury dynamics over a decade after STEMI. MethodsSequential CMR examinations (within the first week after STEMI, and at 4, 12, months, and 9 years thereafter) were conducted in 74 patients with STEMI treated with primary percutaneous coronary intervention. Left ventricular function, infarct size (IS), and microvascular obstruction (MVO) were assessed at all time points. T2∗, T2, and T1 mapping (n = 59) were added at 9-year scan to evaluate the presence of iron and edema within the infarct core, respectively. ResultsIS decreased progressively and significantly between all CMR time points (all P < 0.001), with an average reduction rate of 5.8% per year (IQR: 3.5%-8.8%) and a relative reduction of 49% (IQR: 39%-76%) over a decade. MVO was present in 61% of patients at baseline, but was not present at the follow-up examinations. At 9-year CMR, 17 of 59 (29%) patients showed iron deposition within the infarct core, whereas 82% had persistent edema. Persistent iron and edema were associated with greater IS on any occasion (all P < 0.001), as well as the presence of MVO (P < 0.001). Patients with persistent iron and edema showed a lower relative regression of IS (P = 0.005 and P = 0.032, respectively) and greater end-systolic volumes over a decade (all P < 0.012 and P > 0.023, respectively). A T1 hypointense infarct core without evidence of T2∗ iron deposition (14 of 59 [24%] patients) was attributed to lipomatous metaplasia of the infarct. ConclusionsThe evolution of IS is a dynamic process that extends well beyond the first few months after STEMI. Persistence of iron and edema within the infarct core occurs up to a decade after STEMI and is associated with initial infarct severity and poor infarct healing.

Properties and Functions of Fibroblasts and Myofibroblasts in Myocardial Infarction

The adult mammalian heart contains abundant interstitial and perivascular fibroblasts that expand following injury and play a reparative role but also contribute to maladaptive fibrotic remodeling. Following myocardial infarction, cardiac fibroblasts undergo dynamic phenotypic transitions, contributing to the regulation of inflammatory, reparative, and angiogenic responses. This review manuscript discusses the mechanisms of regulation, roles and fate of fibroblasts in the infarcted heart. During the inflammatory phase of infarct healing, the release of alarmins by necrotic cells promotes a pro-inflammatory and matrix-degrading fibroblast phenotype that may contribute to leukocyte recruitment. The clearance of dead cells and matrix debris from the infarct stimulates anti-inflammatory pathways and activates transforming growth factor (TGF)-β cascades, resulting in the conversion of fibroblasts to α-smooth muscle actin (α-SMA)-expressing myofibroblasts. Activated myofibroblasts secrete large amounts of matrix proteins and form a collagen-based scar that protects the infarcted ventricle from catastrophic complications, such as cardiac rupture. Moreover, infarct fibroblasts may also contribute to cardiac repair by stimulating angiogenesis. During scar maturation, fibroblasts disassemble α-SMA+ stress fibers and convert to specialized cells that may serve in scar maintenance. The prolonged activation of fibroblasts and myofibroblasts in the infarct border zone and in the remote remodeling myocardium may contribute to adverse remodeling and to the pathogenesis of heart failure. In addition to their phenotypic plasticity, fibroblasts exhibit remarkable heterogeneity. Subsets with distinct phenotypic profiles may be responsible for the wide range of functions of fibroblast populations in infarcted and remodeling hearts.

Glucosamine regulates macrophage function in heart failure.

Glucosamine regulates macrophage function in heart failure.

Small vessels in the brain and heart: How similar are they?

AbstractBackgroundIn the brain and heart alike blood vessels are end arteries and their occlusion results in anaemic infarct with high mortality and severe morbidity in a high proportion of survivors. The role of collateral circulation and early recanalization is crucial in both contexts. Ischaemic preconditioning, reperfusion injury and haemorrhagic transformation are three key pathophysiological processes in both organs and the morphological and molecular processes at the level of microcirculation are fundamental to understand and therapeutically modulate the effects of ischaemia.MethodThe morphology of small vessels in brain and heart are compared based on published literature and pathologically assessed cases. The similarities and differences are related to the morphological and molecular consequences of hypoperfusion and vascular occlusion at the level of small vessels and microcirculation. In a case series, we retrospectively analysed the clinical data of 158 patients in a regional centre for acute stroke care. For clinico‐pathological analysis 98 cases (age 62±6.5y) were suitable.ResultAlthough small vessels and the microcirculation share morphological and molecular similarities in health and in response to ischaemic challenge, the specific and unique features of the blood‐brain barrier and the neurovascular unit in the brain are major differences, with implications for drug transfer to the ischaemic parenchyma. Haemorrhagic transformation accelerates the secondary progression of stroke and indicates higher risk of adverse outcome. Neurodegenerative changes impair the microcirculation and, vice versa, microcirculatory damage and dysfunction may trigger and accelerate the neurodegenerative cascade. Vascular and neurodegenerative mechanisms in dementia usually coexist. The morphological patterns of infarct evolution and healing are rather different in the brain and the heart, which also applies to the characteristics of the inflammatory cell composition.ConclusionOur findings underline the importance of post‐mortem pathological examination in the era of advanced imaging and laboratory diagnostic techniques. The fundamental differences in the histomorphology and in the sequence of cellular and molecular events during ischaemia and reperfusion in brain and heart indicate that adaptations of findings in cardiology and clinical neuroscience should be made with caution.

Abstract 12861: Adjoining Myofibroblasts Disrupt Impulse Propagation of Myocardial Tissue via Connexin 43-Mediated Heterocellular Gap Junctional Coupling

Introduction: The composite population of myofibroblasts (MFBs) and cardiomyocytes (CMCs) is known to alter impulse conduction in the heart. A previous study reported that structural disruption between cardiomyocytes by fibrosis or myofibroblast proliferation causes conduction delay due to insulating or reducing the intercellular current. However, it is unknown whether and how the conduction is altered when the myocardial tissue faces adjoining with MFB-rich granulation tissue, like, e.g. , the border zone of myocardial infarcts during infarct healing. Objectives: To clarify the influence of heterocellular gap-junctional coupling between CMCs and MFBs by calcium imaging in an injured myocardial tissue-mimicking model in which CMCs connect with MFBs via microporous membranes. Methods: Fluo-8 fluorescence patterns of impulse propagation were spatiotemporally imaged (27 x 18.9 mm, 333 frames/s, 32 °C) in neonatal rat CMC monolayers cultured on the upper sides of the Boyden chamber (pore diameter, 8 μm) of which MFB monolayer was co-cultured on the reverse sides (CMC-MFB group). For comparison, CMC monolayers were cultured on both membrane sides (CMC-CMC group). Results: During consecutive pacing at 1 - 4 Hz, CMC monolayers showed concentric propagation from the pacing site with slower conduction velocity (CV) and more irregular wavefronts in a frequency-dependent manner. The conduction slowing and its non-uniformity were more remarkable in the CMC-MFB group than in the CMC-CMC group. We confirmed the gap junction coupling between the upper CMC and lower MFB layers using co-immunostaining and calcein staining. Heptanol, gap junction inhibiter, inhibited calcein dye transfer from the upper CMC layer to the lower MFB layer. The knockdown of Cx43 in MFBs on the reverse layer improved the delay of CV and non-uniformity in the CMC-MFB group. Conclusions: In this study, we clarified the electric association between adjoining MFBs and CMCs. Our observations suggest that adjoining MFBs slow down conduction velocity and disrupt conduction uniformity on CMCs via Cx43-mediated heterocellular gap-junctional coupling between CMCs and MFBs.

The role of IL-6 receptor trans-signalling in ischemia-reperfusion injury, infarct healing and future adverse events in patients with ST-Elevation Myocardial Infarction

Abstract Background Inflammation has emerged as a new treatment target in patients with coronary artery disease, and inflammation seems to play an important role in the ischemia/reperfusion injury in ST-elevation myocardial infarction (STEMI). The pro-inflammatory cytokine interleukin-6 (IL-6) has been shown to be associated with myocardial injury and poor prognosis in patients with STEMI. Purpose The aim of the study was to further elucidate possible associations between the IL-6 trans-signalling system and final infarct size, myocardial function, adverse remodelling, and future cardiovascular events in patients with STEMI. Methods A total of 272 patients with first-time STEMI included in the POSTEMI study on ischaemic postconditioning, with symptom duration &amp;lt;6 hours and treated with percutaneous coronary intervention (PCI), were included. Blood samples for analysis of IL-6 and IL-6 receptor (IL-6R) were collected before PCI, immediately after PCI, at day 1 (median 18.3 hours after PCI), and at 4 months follow-up. Cardiac magnetic resonance imaging (CMR) was performed in the acute phase, median 2 days after admission, and repeated after 4 months. Clinical events and all-cause mortality were registered during 12 months' and 70 months' follow-up, respectively. Results There was a significant increase in IL-6 levels from admission to day 1 with a subsequent decline from day 1 to 4 months (Figure 1A). No significant change in IL-6R levels were found from admission to day 1 (Figure 1B). There was no difference between patients treated by postconditioning compared to routine PCI. High levels of IL-6 (&amp;gt; median) at all sampling points were significantly associated with increased infarct size and reduced left ventricular ejection fraction (LVEF) measured by CMR. Additionally, high levels of IL-6 (&amp;gt; median) at day 1 were associated with lower myocardial salvage, more presence of microvascular obstruction and larger increase in indexed LV end diastolic volume (LVEDVi). IL-6R measured during hospitalisation was significantly associated with change in LVEDVi, but did not associate with infarct size, LVEF or myocardial salvage. High levels of IL-6 (&amp;gt;75th percentile) at all sampling points were associated with an increased risk of having an adverse clinical event during the first year and with long-term all-cause mortality (Figure 2), whereas there was no association between IL-6R and adverse clinical events. Conclusion Patients with high IL-6 levels during the acute phase of STEMI had larger infarct size, reduced myocardial salvage, reduced LV function and worse clinical outcome than patients with lower levels of IL-6. High levels of IL-6 measured after 4 months were associated with larger infarct size, reduced LVEF and increased all-cause mortality. IL-6R was significantly associated with increase in LVEDVi. The results add important information to the role of IL-6 in myocardial injury in acute STEMI and the IL-6 pathway as a potential treatment target. Funding Acknowledgement Type of funding sources: Private grant(s) and/or Sponsorship. Main funding source(s): Stein Erik Hagen Foundation for Clinical Heart Research, Oslo, Norway. Figure 1Figure 2

Evolution of myocardial tissue injury over a decade after ST-elevation myocardial infarction: a cardiac magnetic resonance study

Abstract Background In patients with first ST-elevation myocardial infarction (STEMI), the evolution of myocardial tissue injury parameters over a decade as assessed by cardiac magnetic resonance (CMR) has not yet been described. Purpose This study examined long-term myocardial tissue injury dynamics in STEMI patients treated with primary percutaneous coronary intervention (PCI), as well as its association with patient characteristics. Methods A total of 104 patients with STEMI were included in this observational study. Sequential late gadolinium enhanced CMR studies (after 3 days [interquartile ranges (IQR) 2–4], 4 months [IQR 4–5] and 9 years [IQR 8–10]) were conducted to assess left ventricular (LV) dimensions and function, infarct size and microvascular obstruction (MVO). T2* mapping was added at 9 year scan to assess the presence of persistent iron within the infarct core. Results Infarct size decreased progressively from 13% of LV myocardial mass [IQR 7–21] to 10.2% [IQR 5.2–16.1] to 8% [IQR 2.4–12.3] (p&amp;lt;0.001), with an average reduction rate of 6.4% ± 3.4 per year. Relative reduction of infarct size from baseline to 9y follow-up was 43% [IQR 18–66], 21% [IQR 3–42] during the first 4m and 33% [IQR 8–54] between 4m and 9y after STEMI. Decrease of infarct size was associated with greater baseline infarct size (p&amp;lt;0.004) and extent of MVO (p=0.01). MVO was present in 60% (60/104) of patients at baseline, but in none of the follow-up examinations. Sixteen patients had persistent iron within the infarct core at 9 year CMR. Clinical and imaging associates of persistent iron included younger age at study inclusion (p=0.036), higher peak hs troponin T (p&amp;lt;0.001), higher peak creatine kinase (p&amp;lt;0.001) and higher peak CRP (p=0.036) as well as greater infarct size at any occasion (all p&amp;lt;0.001) and greater MVO (p&amp;lt;0.001). Patients with persistent iron showed less relative infarct size regression (51% [IQR 41–79] versus 46% [IQR 32–54], p=0.009). Conclusion In patients with STEMI, the evolution of infarct size is a dynamic process that extends well beyond the first few months after the acute event. MVO vanishes in the first few weeks after the index event in all patients. However, persistence of iron within the infarct core occurs up to a decade after reperfused STEMI, reflecting its irreversibility and is associated with the initial infarct severity and worse infarct healing. Funding Acknowledgement Type of funding sources: None. Central Illustration.Evolution of STEMI over a decade.

Abstract MP255: Intercellular Model Predicts Macrophage Signaling As A Driver Of Cardiac Fibroblast Response Post Myocardial Infarction

Introduction: Post-myocardial infarction (MI), cardiac fibroblasts and macrophages work together to regulate tissue homeostasis and infarct repair. Macrophage-fibroblast interactions in healthy tissue are stable and resistant to perturbations. However, this robustness post-MI has not been assessed. This study designs and implements an intercellular communication model of macrophage-fibroblast crosstalk to determine drivers of infarct repair. Methods: An ordinary differential equation model of post-MI cellular dynamics was developed ( Figure 1A ). Model inputs are time courses of cardiomyocytes, neutrophils, and monocytes. These cells, along with simulated macrophages and fibroblasts, secrete chemokines and cytokines which directed cell proliferation, removal, and chemical secretion. The outputs are macrophage and fibroblast densities, secreted factor dynamics, and produced collagen. Model validation was done using published data in post-MI mice. A sensitivity analysis was conducted by knocking down individual parameters to identify key drivers of fibroblast collagen production. Results: The simulated trends matched the validation time courses. Of the 28 validation relationships, 12 were input-output, 7 were knockouts, and 9 were inhibitor relationships. The validation passed at 78.5%; the 6 failed validations were due to the independent nature of the input curves. Sensitivity analysis ( Figure 1B ) identified macrophage differentiation rate, removal rate, and transforming growth factor-beta (TGFB) secretion rate as pro-fibrotic. Prolonged exposure to TGFB and granulocyte-macrophage colony stimulating factor was anti-fibrotic. Several clustered parameters differentially regulated macrophages and collagen production. Conclusions: The multicellular model identified macrophage density as a pro-fibrotic driver in the healing infarct. Differential regulation of macrophages and collagen production was predicted by the model.

Abstract 120: Pericytes Contribute To Myofibroblast Expansion And Vascular Maturation In The Infarcted Heart

Infarct healing is dependent on recruitment of inflammatory leukocytes and subsequent activation of myofibroblasts (MF) and neovessel formation, ultimately resulting in formation of a highly vascularized collagen-enriched scar. Though the heart has an abundant population of periendothelial pericytes, its role in wound healing upon myocardial infarction (MI) has not been studied. We hypothesized that in the infarcted myocardium, pericytes may become activated, contributing to inflammatory, fibrotic and angiogenic responses. We used pericyte/fibroblast reporter mice (NG2 DsRed ;PDGFRα GFP ), lineage tracing studies and in vitro approaches to study the fate and role of pericytes in the infarcted myocardium. In normal hearts, NG2+/PDGFRα- pericytes and PDGFRα+/NG2- fibroblasts had distinct transcriptomic profiles. Pericytes expressed mural genes like Acta2 , Pdgfrb and low amounts of extracellular matrix (ECM) genes, whereas fibroblasts synthesized collagens, Timp2/3 and matricellular genes. 7 days post-MI, expansion of the NG2+ population in the infarct zone was associated with emergence of non-mural NG2+/αSMA+ cells with MF characteristics. FACS-sorted NG2+/PDGFRα- cells from 7-day infarcts expressed higher levels of collagens when compared to NG2+/PDGFRα- cells from normal hearts. Infarct pericytes had high integrin and MMP14 expression, reflecting an activated migratory phenotype. Lineage tracing using NG2CreER TM ;Rosa tdTomato ;PDGFRα GFP mice showed that 5.7%±1.04 of PDGFRα+ fibroblasts and 10.49%±2.73 of infarct MFs were derived from NG2+ lineage. Pericyte-derived fibroblasts exhibited higher ECM gene synthesis, in comparison to fibroblasts from non-pericyte origin, while pericyte-derived mural cells showed accentuated inflammatory cytokine gene expression. Immunostaining showed pericytes actively contribute to vascular maturation, forming a mural cell coat enwrapping infarct neovessels. In vitro, TGFβ induced integrins, collagens and MMPs in human pericytes, similar to the changes observed in infarct pericytes. Taken together, our evidences show that after MI, pericytes become activated and contribute to repair by undergoing conversion to a subset of myofibroblasts and by coating infarct neovessels.