Myocardial Infarction In Pigs Research Articles

Characterizing the immune response to myocardial infarction in pigs.

Though myocardial infarction (MI) in pigs is a well-established translational large animal model, it has not yet been widely used for immunotherapy studies, and a comprehensive description of the immune response to MI in this species is lacking. We induced MI in Landrace pigs by balloon occlusion of the left anterior descending artery over 90min. Within 14days, the necrotic myocardium was progressively replaced by scar tissue with involvement of myofibroblasts. We characterized the immune response in the heart ex vivo by (immuno)histology, flow cytometry, and RNA sequencing of myocardial tissue on days 3, 7, and 14 after MI. Besides a clear predominance of myeloid cells among heart-infiltrating leukocytes, we detected activated T cells and an increasing proportion of CD4+ Foxp3+ regulatory T cells (Treg), especially in the infarct core-findings that closely mirror what has been observed in mice and humans after MI. Transcriptome data indicated inflammatory activity that was persistent but markedly changing in character over time and linked to extracellular matrix biology. Analysis of lymphocytes in heart-draining lymph nodes revealed significantly higher proliferation rates of T helper cell subsets, including Treg on day 7 after MI, compared to sham controls. Elevated frequencies of myeloid progenitors in the spleen suggest that it might be a site of emergency myelopoiesis after MI in pigs, as previously shown in mice. We thus provide a first description of the immune response to MI in pigs, and our results can aid future research using the species for preclinical immunotherapy studies.

Abstract 17394: Minimally Invasive Delivery of Synthetic Cardiac Stromal Cells Improves Heart Repair in Pigs

Introduction: Myocardial infarction (MI) is the primary manifestation of cardiovascular events. Synthetic cardiac stromal cells (synCSCs) were shown to promote cardiac repair after MI. Minimally invasive delivery of synCSCs embedded in hyaluronic acid (HA) hydrogel via intrapericardial cavity (iPC) injections in a porcine model may provide a unique dataset for cardiac regeneration therapeutics. Hypothesis: Minimally invasive delivery of synCSCs via iPC injection will improve pig cardiac function and cardiomyocyte (CM) proliferation. Methods: A pig MI model was created using coronary balloon occlusion, and Yorkshire male pigs underwent video-assisted thoracoscopic surgery for iPC injections. Cardiac function studies were conducted using MRI to measure left ventricle ejection fraction (LVEF) and the infarct area. Histology and immunohistochemistry (IHC) analyses were performed to investigate CM proliferation. Single nucleus RNA-sequencing (snRNA-sq) analysis was performed to identify key gene features and phenotype changes that may be affected by the synCSC treatment. Results: The iPC injection of synCSCs embedded in HA hydrogel improves cardiac function, enhances cardiomyocyte proliferation, and induces phenotype alteration in the pig MI model. MRI analysis depicts significantly attenuated infarct size and improved LVEF at Day 28 in the synCSC group. Histology shows reduced fibrotic percentage and improved infarct wall thickness. IHC analysis shows a significant increase in cardiomyocyte proliferation. SnRNA-seq elucidates immature cardiomyocytes with higher proliferation gene features. Conclusion: The results show significant improvements in pig cardiac function and CM proliferation. Interestingly, immature cardiomyocytes of a different phenotype emerged in the therapeutic group. These findings provide an unprecedented dataset to facilitate the clinical translation of cardiac tissue regeneration therapeutics.

Abstract 12266: CCND2 Modified mRNA Activates the Cell Cycle in Cardiomyocytes and Improves Recovery From Myocardial Infarction in Small and Large Mammals

Background: Experiments in mammalian models of cardiac injury suggest that the cardiomyocyte (CM)-specific overexpression of cyclin D2 (CCND2 in humans) improves recovery from myocardial infarction (MI). The primary objective of this investigation was to demonstrate that our Specific Modified mRNA Translation System (SMRTs) can induce CCND2 expression in CMs (CCND2-CMs) and replicate the benefits observed in other studies of CM-specific CCND2 overexpression for myocardial repair. Methods: The CCND2-CM SMRTs consists of two modRNA constructs: one codes for CCND2 and contains a binding site for L7Ae, and the other codes for L7Ae and contains recognition elements for the CM-specific microRNAs miR-1 and miR-208. Thus, L7Ae suppresses CCND2 translation in noncardiomyocytes but is itself suppressed by endogenous miR-1 and -208 in CMs, thereby facilitating CM-specific CCND2 expression. Experiments were conducted in both mouse and pig models of MI, and control assessments were performed in animals treated with an SMRTs coding for the CM-specific expression of luciferase or GFP, in animals treated with L7Ae modRNA alone or with the delivery vehicle, and in Sham-operated animals. Results: CCND2 was abundantly expressed in cultured, post-mitotic CMs two days after transfection with CCND2-CM SMRTs, and the increase was accompanied by the upregulation of markers for cell-cycle activation and proliferation (e.g., Ki67 and Aurora B kinase). When the GFP-CM SMRTs was intramyocardially injected into infarcted mouse hearts, the GFP signal was observed in CMs but no other cell type. In both the mouse and pig MI models, CM proliferation (on Day 7 and Day 3 after treatment administration in mice and pigs, respectively) was significantly greater, left-ventricular ejection fractions (Days 7 and 28 in mice, Days 10 and 28 in pigs) were significantly higher, and infarcts (Day 28 in both species) were significantly smaller in animals treated with CCND2-CM SMRTs than in any other group that underwent MI induction. Conclusion: Intramyocardial injections of the CCND2-CM SMRTs promoted CM proliferation, reduced infarct size, and improved cardiac performance in small and large mammalian hearts with AMI.

Early Changes in Acute Myocardial Infarction in Pigs: Achieving Early Detection with Wearable Devices

We examined the changes in variables that could be recorded on wearable devices during the early stages of acute myocardial infarction (AMI) in an animal model. Early diagnosis of AMI is important for prognosis; however, delayed diagnosis is common because of patient hesitation and lack of timely evaluations. Wearable devices are becoming increasingly sophisticated in the ability to track indicators. In this study, we retrospectively reviewed the changes in four variables during AMI in a pig model to assess their ability to help predict AMI onset. AMI was created in 33 pigs by 90-min balloon occlusion of the left anterior descending artery. Blood pressure, EKG, and lactate and cardiac troponin I levels were recorded during the occlusion period. Blood pressure declined significantly within 15 min after balloon inflation (mean arterial pressure, from 61 ± 8 to 50 ± 8 mmHg) and remained at this low level. Within 5 min of balloon inflation, the EKG showed ST-elevation in precordial leads V1-V3. Blood lactate levels increased gradually after occlusion and peaked at 60 min (from 1.48 to 2.53 mmol/L). The continuous transdermal troponin sensor demonstrated a gradual increase in troponin levels over time. Our data suggest that significant changes in key indicators (blood pressure, EKG leads V1-V3, and lactate and troponin levels) occurred at the onset of AMI. Monitoring of these variables could be used to develop an algorithm and alert patients early at the onset of AMI with the help of a wearable device.

NLRP3-Inflammasome Inhibition with IZD334 Does Not Reduce Cardiac Damage in a Pig Model of Myocardial Infarction.

NLRP3-inflammasome-mediated signaling is thought to significantly contribute to the extent of myocardial damage after myocardial infarction (MI). The purpose of this study was to investigate the effects of the NLRP3-inflammasome inhibitor IZD334 on cardiac damage in a pig model of myocardial infarction. Prior to in vivo testing, in vitro, porcine peripheral blood mononuclear cells and whole blood were treated with increasing dosages of IZD334, a novel NLRP3-inflammasome inhibitor, and were stimulated with lipopolysaccharide (LPS) and adenosine triphosphate (ATP). After determination of the pharmacological profile in healthy pigs, thirty female Landrace pigs were subjected to 75 min of transluminal balloon occlusion of the LAD coronary artery and treated with placebo or IZD334 (1 mg/kg, 3 mg/kg, or 10 mg/kg once daily) in a blinded randomized fashion. In vitro, NLRP3-inflammasome stimulation showed the pronounced release of interleukin (IL)-1β that was attenuated by IZD334 (p < 0.001). In vivo, no differences were observed between groups in serological markers of inflammation nor myocardial IL-1β expression. After 7 days, the ejection fraction did not differ between groups, as assessed with MRI (placebo: 45.1 ± 8.7%, 1 mg/kg: 49.9 ± 6.1%, 3 mg/kg: 42.7 ± 3.8%, 10 mg/kg: 44.9 ± 6.4%, p = 0.26). Infarct size as a percentage of the area at risk was not reduced (placebo: 73.1 ± 3.0%, 1 mg/kg: 75.5 ± 7.3%, 3 mg/kg: 80.3 ± 3.9%, 10 mg/kg: 78.2 ± 8.0%, p = 0.21). In this pig MI model, we did not observe attenuation of the inflammatory response after NLRP3-inflammasome inhibition in vivo. Consecutively, no difference was observed in IS and cardiac function, while in vitro inhibition successfully reduced IL-1β release from stimulated porcine blood cells.

Hyperpolarized Metabolic and Parametric CMR Imaging of Longitudinal Metabolic-Structural Changes in Experimental Chronic Infarction

BackgroundProlonged ischemia and myocardial infarction are followed by a series of dynamic processes that determine the fate of the affected myocardium toward recovery or necrosis. Metabolic adaptions are considered to play a vital role in the recovery of salvageable myocardium in the context of stunned and hibernating myocardium. ObjectivesThe potential of hyperpolarized pyruvate cardiac magnetic resonance (CMR) alongside functional and parametric CMR as a tool to study the complex metabolic-structural interplay in a longitudinal study of chronic myocardial infarction in an experimental pig model is investigated. MethodsMetabolic imaging using hyperpolarized [1-13C] pyruvate and proton-based CMR including cine, T1/T2 relaxometry, dynamic contrast-enhanced, and late gadolinium enhanced imaging were performed on clinical 3.0-T and 1.5-T MR systems before infarction and at 6 days and 5 and 9 weeks postinfarction in a longitudinal study design. Chronic myocardial infarction in pigs was induced using catheter-based occlusion and compared with healthy controls. ResultsMetabolic image data revealed temporarily elevated lactate-to-bicarbonate ratios at day 6 in the infarcted relative to remote myocardium. The temporal changes of lactate-to-bicarbonate ratios were found to correlate with changes in T2 and impaired local contractility. Assessment of pyruvate dehydrogenase flux via the hyperpolarized [13C] bicarbonate signal revealed recovery of aerobic cellular respiration in the hibernating myocardium, which correlated with recovery of local radial strain. ConclusionsThis study demonstrates the potential of hyperpolarized CMR to longitudinally detect metabolic changes after cardiac infarction over days to weeks. Viable myocardium in the area at risk was identified based on restored pyruvate dehydrogenase flux.

CineCT platform for in vivo and ex vivo measurement of 3D high resolution Lagrangian strains in the left ventricle following myocardial infarction and intramyocardial delivery of theranostic hydrogel

Myocardial infarction (MI) produces acute changes in strain and stiffness within the infarct that can affect remote areas of the left ventricle (LV) and drive pathological remodeling. We hypothesized that intramyocardial delivery of a hydrogel within the MI region would lower wall stress and reduce adverse remodeling in Yorkshire pigs (n = 5). 99mTc-Tetrofosmin SPECT imaging defined the location and geometry of induced MI and border regions in pigs, and in vivo and ex vivo contrast cine computed tomography (cineCT) quantified deformations of the LV myocardium. Serial in vivo cineCT imaging provided data in hearts from control pigs (n = 3) and data from pigs (n = 5) under baseline conditions before MI induction, post-MI day 3, post-MI day 7, and one hour after intramyocardial delivery of a hyaluronic acid (HA)-based hydrogel with shear-thinning and self-healing properties to the central infarct area. Isolated, excised hearts underwent similar cineCT imaging using an ex vivo perfused heart preparation with cyclic LV pressurization. Deformations were evaluated using nonlinear image registration of cineCT volumes between end-diastole (ED) and end-systole (ES), and 3D Lagrangian strains were calculated from the displacement gradients. Post-MI day 3, radial, circumferential, maximum principal, and shear strains were reduced within the MI region (p < 0.04) but were unchanged in normal regions (p > 0.6), and LV end diastolic volume (LV EDV) increased (p = 0.004), while ejection fraction (EF) and stroke volume (SV) decreased (p < 0.02). Post-MI day 7, radial strains in MI border zones increased (p = 0.04) and dilation of LV EDV continued (p = 0.052). There was a significant negative linear correlation between regional radial and maximum principal/shear strains and percent infarcted tissue in all hearts (R2 > 0.47, p < 0.004), indicating that cineCT strain measures could predict MI location and degree of injury. Post-hydrogel day 7 post-MI, LV EDV was significantly reduced (p = 0.009), EF increased (p = 0.048), and radial (p = 0.021), maximum principal (p = 0.051), and shear strain (p = 0.047) increased within regions bordering the infarct. A smaller strain improvement within the infarct and normal regions was also noted on average along with an improvement in SV in 4 out of 5 hearts. CineCT provides a reliable method to assess regional changes in strains post-MI and the therapeutic effects of intramyocardial hydrogel delivery.

"Evaluation of Myocardial Infarction in Pigs by Coronary Spectral CT Angiography"

"Evaluation of Myocardial Infarction in Pigs by Coronary Spectral CT Angiography"

An optimal non-viral gene transfer method for genetically modifying porcine bone marrow-derived endothelial progenitor cells for experimental therapeutics.

No currently available treatment is able to generate new contractile tissue or significantly improve cardiac function after myocardial infarction (MI), a leading cause of morbidity and mortality worldwide. Although gene transfer-enhanced endothelial progenitor cells (GTE-EPCs) show effectiveness in MI treatment in small animal models, no clinical trials using GTE-EPCs have been documented. Before the introduction of GTE-EPCs into human trials, gene-transfer-mediated augmentation of EPC function in animal models that reflect the human MI scenario should be tested. In this regard, a porcine model is the best choice since pigs have cardiac size, hemodynamics and coronary anatomy similar to that of humans. To examine GTE-EPC therapeutic efficacy in pig MI models, an efficient method for gene transfer into pig EPCs is required, which however, has been poorly documented. Pig bone marrow mononuclear cells were isolated and cultured in EGM-2 medium to obtain bone marrow-derived EPCs (BM-EPCs) that were characterized by immunostaining and the tube formation assay. Gene transfer was optimized in 6-well plates using a GFP and a VEGF plasmid, and scaled up in T75 flasks. Gene transfer efficiency was determined by fluorescence microscopy and flow cytometry. VEGF levels were measured by ELISA. Cell proliferation was assayed by the CCK-8 kit. (1) BM-EPCs expressed VEGFR2 and eNOS but not CD45 protein, and formed tube structures on Matrigel; (2) several chemical compounds were explored with the highest transfection efficiency of 41.4% ± 5.8% achieved using Lipofectamine 3000; (3) the VEGF level in culture medium after VEGF transfection was 378 ± 48 ng/106 cells; and (4) BM-EPCs overexpressing VEGF had significantly enhanced proliferation than GFP-transfected EPCs. A simple, easy and cheap method that can be applied to produce a large number of genetically-modified BM-EPCs was established, which will facilitate the study of GTE-EPC therapeutic efficacy in pig MI model.

Cardiovascular magnetic resonance imaging of functional and microstructural changes of the heart in a longitudinal pig model of acute to chronic myocardial infarction

BackgroundWe examined the dynamic response of the myocardium to infarction in a longitudinal porcine study using relaxometry, functional as well as diffusion cardiovascular magnetic resonance (CMR). We sought to compare non contrast CMR methods like relaxometry and in-vivo diffusion to contrast enhanced imaging and investigate the link of microstructural and functional changes in the acute and chronically infarcted heart.MethodsCMR was performed on five myocardial infarction pigs and four healthy controls. In the infarction group, measurements were obtained 2 weeks before 90 min occlusion of the left circumflex artery, 6 days after ischemia and at 5 as well as 9 weeks as chronic follow-up. The timing of measurements was replicated in the control cohort. Imaging consisted of functional cine imaging, 3D tagging, T2 mapping, native as well as gadolinium enhanced T1 mapping, cardiac diffusion tensor imaging, and late gadolinium enhancement imaging.ResultsNative T1, extracellular volume (ECV) and mean diffusivity (MD) were significantly elevated in the infarcted region while fractional anisotropy (FA) was significantly reduced. During the transition from acute to chronic stages, native T1 presented minor changes (< 3%). ECV as well as MD increased from acute to the chronic stages compared to baseline: ECV: 125 ± 24% (day 6) 157 ± 24% (week 5) 146 ± 60% (week 9), MD: 17 ± 7% (day 6) 33 ± 14% (week 5) 29 ± 15% (week 9) and FA was further reduced: − 31 ± 10% (day 6) − 38 ± 8% (week 5) − 36 ± 14% (week 9). T2 as marker for myocardial edema was significantly increased in the ischemic area only during the acute stage (83 ± 3 ms infarction vs. 58 ± 2 ms control p < 0.001 and 61 ± 2 ms in the remote area p < 0.001). The analysis of functional imaging revealed reduced left ventricular ejection fraction, global longitudinal strain and torsion in the infarct group. At the same time the transmural helix angle (HA) gradient was steeper in the chronic follow-up and a correlation between longitudinal strain and transmural HA gradient was detected (r = 0.59 with p < 0.05). Comparing non-gadolinium enhanced data T2 mapping showed the largest relative change between infarct and remote during the acute stage (+ 33 ± 4% day 6, with p = 0.013 T2 vs. MD, p = 0.009 T2 vs. FA and p = 0.01 T2 vs. T1) while FA exhibited the largest relative change between infarct and remote during the chronic follow-up (+ 31 ± 2% week 5, with p = N.S. FA vs. MD, p = 0.03 FA vs. T2 and p = 0.003 FA vs. T1). Overall, diffusion parameters provided a higher contrast (> 23% for MD and > 27% for FA) during follow-up compared to relaxometry (T1 17–18%/T2 10–20%).ConclusionDuring chronic follow-up after myocardial infarction, cardiac diffusion tensor imaging provides a higher sensitivity for mapping microstructural alterations when compared to non-contrast enhanced relaxometry with the added benefit of providing directional tensor information to assess remodelling of myocyte aggregate orientations, which cannot be otherwise assessed.

Exosomes secreted by hiPSC-derived cardiac cells improve recovery from myocardial infarction in swine.

Cell therapy treatment of myocardial infarction (MI) is mediated, in part, by exosomes secreted from transplanted cells. Thus, we compared the efficacy of treatment with a mixture of cardiomyocytes (CMs; 10 million), endothelial cells (ECs; 5 million), and smooth muscle cells (SMCs; 5 million) derived from human induced pluripotent stem cells (hiPSCs), or with exosomes extracted from the three cell types, in pigs after MI. Female pigs received sham surgery; infarction without treatment (MI group); or infarction and treatment with hiPSC-CMs, hiPSC-ECs, and hiPSC-SMCs (MI + Cell group); with homogenized fragments from the same dose of cells administered to the MI + Cell group (MI + Fra group); or with exosomes (7.5 mg) extracted from a 2:1:1 mixture of hiPSC-CMs:hiPSC-ECs:hiPSC-SMCs (MI + Exo group). Cells and exosomes were injected into the injured myocardium. In vitro, exosomes promoted EC tube formation and microvessel sprouting from mouse aortic rings and protected hiPSC-CMs by reducing apoptosis, maintaining intracellular calcium homeostasis, and increasing adenosine 5'-triphosphate. In vivo, measurements of left ventricular ejection fraction, wall stress, myocardial bioenergetics, cardiac hypertrophy, scar size, cell apoptosis, and angiogenesis in the infarcted region were better in the MI + Cell, MI + Fra, and MI + Exo groups than in the MI group 4 weeks after infarction. The frequencies of arrhythmic events in animals from the MI, MI + Cell, and MI + Exo groups were similar. Thus, exosomes secreted by hiPSC-derived cardiac cells improved myocardial recovery without increasing the frequency of arrhythmogenic complications and may provide an acellular therapeutic option for myocardial injury.

MicroRNA-directed cardiac repair after myocardial infarction in pigs.

Viral vector delivery of microRNA-199a after myocardial infarction in pigs can stimulate endogenous myocardial repair mechanisms and improve cardiac function; however, persistent expression of the miRNA leads to sudden arrhythmic death.

MicroRNA therapy stimulates uncontrolled cardiac repair after myocardial infarction in pigs.

Prompt coronary catheterization and revascularization have dramatically improved the outcome of myocardial infarction, but also have resulted in a growing number of survived patients with permanent structural damage of the heart, which frequently leads to heart failure. Finding new treatments for this condition is a largely unmet clinical need 1, especially because of the incapacity of cardiomyocytes to replicate after birth and thus achieve regeneration of the lost contractile tissue 2. Here we show that expression of human microRNA-199a in infarcted pig hearts is capable of stimulating cardiac repair. One month after myocardial infarction and delivery of this microRNA through an adeno-associated viral vector, the treated animals showed marked improvements in both global and regional contractility, increased muscle mass and reduced scar size. These functional and morphological findings correlated with cardiomyocyte de-differentiation and proliferation. At longer follow-up, however, persistent and uncontrolled expression of the microRNA resulted in sudden arrhythmic death of most of the treated pigs. Such events were concurrent with myocardial infiltration of proliferating cells displaying a poorly differentiated myoblastic phenotype. These results show that achieving cardiac repair through the stimulation of endogenous cardiomyocyte proliferation is attainable in large mammals, however this therapy needs to be tightly dosed.

Cardioprotective Effects of Curcumin-Nisin Based Poly Lactic Acid Nanoparticle on Myocardial Infarction in Guinea Pigs

Myocardial infarction (MI) is the most prevalent cause of cardiovascular death. A possible way of preventing MI maybe by dietary supplements. The present study was thus designed to ascertain the cardio-protective effect of a formulated curcumin and nisin based poly lactic acid nanoparticle (CurNisNp) on isoproterenol (ISO) induced MI in guinea pigs. Animals were pretreated for 7 days as follows; Groups A and B animals were given 0.5 mL/kg of normal saline, group C metoprolol (2 mg/kg), groups D and E CurNisNp 10 and 21 mg/kg respectively (n = 5). MI was induced on the 7th day in groups B-E animals. On the 9th day electrocardiogram (ECG) was recorded, blood samples and tissue biopsies were collected for analyses. Toxicity studies on CurNisNp were carried out. MI induction caused atrial fibrillation which was prevented by pretreatment of metoprolol or CurNisNp. MI induction was also associated with increased expressions of cardiac troponin I (CTnI) and kidney injury molecule-1 (KIM-1) which were significantly reduced in guinea pig’s pretreated with metoprolol or CurNisNp (P < 0.05). The LC50 of CurNisNp was 3258.2 μg/mL. This study demonstrated that the formulated curcumin-nisin based nanoparticle confers a significant level of cardio-protection in the guinea pig and is nontoxic.

Abstract 336: Human Bone Contains Primitive Cells With Angiogenic and Immunomodulatory Properties

Previously, we have shown that treatments of cortical bone derived stem cells (CBSCs) reduce ventricular remodeling and improve cardiac functions in both mouse and pig myocardial infarction (MI) models. The goals of the present study were to isolate and characterize human CBSCs (hCBSCs). Using bone fragments obtained at the time surgeries, we isolated three morphologically distinctive hCBSC phenotypes; 1) sphere forming (SF); 2) epithelial like monolayer (ELM); and 3) larger and spindle shaped (LSS) phenotype. The hCBSC phenotypes grew without senescence in culture. Mean doubling time (hour) for SF, ELM, and LSS at the passage number 20 (P20) were 19.54, 21.21, and 20.3 respectively and at the P40 were 18.01, 17.13, and 20.56 respectively. All of the phenotypes were found to have surface expression of CD44; however, other markers detected (CD105, CD90, CD73, CD106, CD271, and CD133) on mouse CBSCs (mCBSCs) were not detected along with the markers not detected on the surface of mCBSCs (CD45, CD11b, CD31, CD34, CD117, and CD325). Erythrocytes (CD235a) and immune cell markers (CD2, CD3, CD14, CD16, CD19, and CD56) were also not detected. The hCBSCs were found without histocompatibility antigen Ia (MHC-Ia: HLA-A, HLA-B, and HLA-C) and MHC-II (HLA-DR); instead, they expressed soluble forms of MHC-Ib (HLA-G5, HLA-G6, and HLA-G7), potent inhibitor of natural killer cells and other immune cells. These features are commonly found in immune privileged cells. Further, the hCBSCs produced the immunomodulatory and anti-inflammatory factors (IL-4, IL-1RA, TGF-b and IL-10). Additionally, the hCBSCs secreted angiogenic factors that organize endothelial cells into tube like structures. From these results, the hCBSCs we isolated have totally different surface marker characteristics compared to any other known cells including mCBSCs. In addition to their angiogenic effects, the hCBSCs have immunomodulatory properties that could make them suitable for cell therapy targeting inflammatory diseases such post myocardial infarction remodeling.

3006Chemical exchange saturation transfer for the measure of myocardium metabolism status: preliminary validation in myocardial infarction pigs

3006Chemical exchange saturation transfer for the measure of myocardium metabolism status: preliminary validation in myocardial infarction pigs

Role of Gated Myocardial Glucose Metabolic Imaging in Assessing Left Ventricular Systolic Dyssynchrony after Myocardial Infarction and the Influential Factors

In this study, we investigated the role of gated myocardial glucose metabolic imaging in assessing left ventricular (LV) systolic dyssynchrony after myocardial infarction (MI) and explored the influencing factors. Bama mini-pigs were divided into normal group and MI group and subjected to gated myocardial metabolic imaging (GMMI) and gated myocardial perfusion imaging (GMPI). The phase bandwidth (BW), standard deviation (SD) and the latest activation site of left ventricle were obtained using program Cedars QGS. The results showed that (1) BW and SD obtained in GMMI and GMPI showed significant correlation in pigs with MI, but not in the normal pigs, (2) BW and SD obtained in GMMI and GMPI had good consistency in both normal pigs and MI pigs, (3) GMMI and GMPI had a 66.7% identity in determining the latest activation site of left ventricle in the normal pigs and 77.8% identity in determining the latest activation site of left ventricle in pigs with MI. Multivariate stepwise regression analysis showed that total perfusion deficit and summed motion score were independent factors affecting BW and SD in GMMI. In conclusion, phase analysis of GMMI images could objectively reflect LV systolic dyssynchrony resulted from interactions of multiple factors.

Hyperactive ryanodine receptors in human heart failure and ischaemic cardiomyopathy reside outside of couplons.

AimsIn ventricular myocytes from humans and large mammals, the transverse and axial tubular system (TATS) network is less extensive than in rodents with consequently a greater proportion of ryanodine receptors (RyRs) not coupled to this membrane system. TATS remodelling in heart failure (HF) and after myocardial infarction (MI) increases the fraction of non-coupled RyRs. Here we investigate whether this remodelling alters the activity of coupled and non-coupled RyR sub-populations through changes in local signalling. We study myocytes from patients with end-stage HF, compared with non-failing (non-HF), and myocytes from pigs with MI and reduced left ventricular (LV) function, compared with sham intervention (SHAM).Methods and resultsSingle LV myocytes for functional studies were isolated according to standard protocols. Immunofluorescent staining visualized organization of TATS and RyRs. Ca2+ was measured by confocal imaging (fluo-4 as indicator) and using whole-cell patch-clamp (37°C). Spontaneous Ca2+ release events, Ca2+ sparks, as a readout for RyR activity were recorded during a 15 s period following conditioning stimulation at 2 Hz. Sparks were assigned to cell regions categorized as coupled or non-coupled sites according to a previously developed method. Human HF myocytes had more non-coupled sites and these had more spontaneous activity than in non-HF. Hyperactivity of these non-coupled RyRs was reduced by Ca2+/calmodulin-dependent kinase II (CaMKII) inhibition. Myocytes from MI pigs had similar changes compared with SHAM controls as seen in human HF myocytes. As well as by CaMKII inhibition, in MI, the increased activity of non-coupled sites was inhibited by mitochondrial reactive oxygen species (mito-ROS) scavenging. Under adrenergic stimulation, Ca2+ waves were more frequent and originated at non-coupled sites, generating larger Na+/Ca2+ exchange currents in MI than in SHAM. Inhibition of CaMKII or mito-ROS scavenging reduced spontaneous Ca2+ waves, and improved excitation–contraction coupling.ConclusionsIn HF and after MI, RyR microdomain re-organization enhances spontaneous Ca2+ release at non-coupled sites in a manner dependent on CaMKII activation and mito-ROS production. This specific modulation generates a substrate for arrhythmia that appears to be responsive to selective pharmacologic modulation.

Noninvasive Immunometabolic Cardiac Inflammation Imaging Using Hyperpolarized Magnetic Resonance.

Current cardiovascular clinical imaging techniques offer only limited assessment of innate immune cell-driven inflammation, which is a potential therapeutic target in myocardial infarction (MI) and other diseases. Hyperpolarized magnetic resonance (MR) is an emerging imaging technology that generates contrast agents with 10- to 20 000-fold improvements in MR signal, enabling cardiac metabolite mapping. To determine whether hyperpolarized MR using [1-13C]pyruvate can assess the local cardiac inflammatory response after MI. We performed hyperpolarized [1-13C]pyruvate MR studies in small and large animal models of MI and in macrophage-like cell lines and measured the resulting [1-13C]lactate signals. MI caused intense [1-13C]lactate signal in healing myocardial segments at both day 3 and 7 after rodent MI, which was normalized at both time points after monocyte/macrophage depletion. A near-identical [1-13C]lactate signature was also seen at day 7 after experimental MI in pigs. Hyperpolarized [1-13C]pyruvate MR spectroscopy in macrophage-like cell suspensions demonstrated that macrophage activation and polarization with lipopolysaccharide almost doubled hyperpolarized lactate label flux rates in vitro; blockade of glycolysis with 2-deoxyglucose in activated cells normalized lactate label flux rates and markedly inhibited the production of key proinflammatory cytokines. Systemic administration of 2-deoxyglucose after rodent MI normalized the hyperpolarized [1-13C]lactate signal in healing myocardial segments at day 3 and also caused dose-dependent improvement in IL (interleukin)-1β expression in infarct tissue without impairing the production of key reparative cytokines. Cine MRI demonstrated improvements in systolic function in 2-DG (2-deoxyglucose)-treated rats at 3 months. Hyperpolarized MR using [1-13C]pyruvate provides a novel method for the assessment of innate immune cell-driven inflammation in the heart after MI, with broad potential applicability across other cardiovascular disease states and suitability for early clinical translation.

Augmentation of M‐Current and Increased Synaptic Efficacy at Intrinsic Cardiac Neurons Contributes to an Enhanced Cardiac Responsiveness to Vagal Nerve Stimulation in the Infarcted Porcine Heart

Imbalances to autonomic regulation of cardiac function contribute to risk for arrhythmia after myocardial infarction (MI). We tested the hearts responsiveness to vagal nerve stimulation (VNS), 8 wks after MI, to determine if parasympathetic control of the heart is altered in MI pigs. A significantly greater bradycardic response was elicited in MI animals during 20Hz stimulation. Intracellular voltage recordings from whole‐mount preparations of intrinsic cardiac neurons (ICNs) were subsequently used to assess whether alterations in neuronal membrane properties or synaptic transmission underlie the observed increase in responsiveness to VNS in MI animals. General membrane properties (resting membrane potential, input resistance, action potential amplitude/duration, afterhyperpolarization amplitude/duration) of ICNs from control and MI pigs were not different, but ICN excitability was significantly depressed in MI pigs. Testing synaptic efficacy revealed a greater proportion of strong (high AP probability) vs weak synapses (low AP probability) in MI animals. Tetanic stimulation (20Hz) of interganglionic nerve bundles elicited slow excitatory postsynaptic potentials (sEPSPs) in a significantly greater subset of MI ICNs (10/22) vs controls (4/26). The sEPSP persisted in hexamethonium (500uM) but was blocked by atropine (1uM), indicating mediation by cholinergic inhibition of M‐current. Application of 1mM Ba2+ increased excitability of ICNs from both control and MI animals further supporting a role for M‐current in the regulation of porcine ICN excitability. In hexamethonium, the amplitude of the sEPSP was significantly greater in MI ICNs. In conclusion, we identify a important role of M‐current in the regulation of the excitability of porcine ICNs, and demonstrate enhanced membrane depolarization of ICNs from MI animals after presynaptic stimulation due, in part, to greater inhibition of IM during tetanic nerve stimulation which may underlie the enhanced sensitivity to VNS in MI pigs.Support or Funding InformationThis work was supported in part by NIH grant OT2OD023848.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.