Surgical Porcine Model of Chronic Myocardial Ischemia Treated by Exosome-laden Collagen Patch and Off-pump Coronary Artery Bypass Graft.

Chronic myocardial ischemia resulting from progressive coronary artery stenosis leads to hibernating myocardium (HIB), defined as myocardium that adapts to reduced oxygen availability by reducing metabolic activity, thereby preventing irreversible cardiomyocyte injury and infarction. This is distinct from myocardial infarction, as HIB has the potential for recovery with revascularization. Patients with significant coronary artery disease (CAD) experience chronic ischemia, which puts them at risk for heart failure and sudden death. The standard surgical intervention for severe CAD is coronary artery bypass graft surgery (CABG), but it has been shown to be an imperfect therapy, yet no adjunctive therapies exist to recover myocytes adapted to chronic ischemia. To address this gap, a surgical model of HIB using porcine that is amenable to CABG and mimics the clinical scenario was used. The model involves two surgeries. The first operation involves implanting a 1.5 mm rigid constrictor on the left anterior descending (LAD) artery. As the animal grows, the constrictor gradually causes significant stenosis resulting in reduced regional systolic function. Once the stenosis reaches 80%, the myocardial flow and function are impaired, creating HIB. An off-pump CABG is then performed with the left internal mammary artery (LIMA) to revascularize the ischemic region. The animal recovers for one month to allow for optimal myocardial improvement prior to sacrifice. This allows for physiologic and tissue studies of different treatment groups. This animal model demonstrates that cardiac function remains impaired despite CABG, suggesting the need for novel adjunctive interventions. In this study, a collagen patch embedded with mesenchymal stem cell (MSC)-derived exosomes was developed, which can be surgically applied to the epicardial surface distal to LIMA anastomosis. The material conforms to the epicardium, is absorbable, and provides the scaffold for the sustained release of signaling factors. This regenerative therapy can stimulate myocardial recovery that does not respond to revascularization alone. This model translates to the clinical arena by providing means of physiological and mechanistic explorations regarding recovery in HIB.

Stem cell-derived exosome patch with coronary artery bypass graft restores cardiac function in chronically ischemic porcine myocardium.

OBJECTIVE: This study aimed to investigate whether or not the application of a stem cell-derived exosome-laden collagen patch (EXP) during coronary artery bypass grafting (CABG) can recover cardiac function by modulating mitochondrial bioenergetics and myocardial inflammation in hibernating myocardium (HIB), which is defined as myocardium with reduced blood flow and function that retains viability and variable contractile reserve. METHODS: In vitro methods involved exposing H9C2 cardiomyocytes to hypoxia followed by normoxic coculture with porcine mesenchymal stem cells. Mitochondrial respiration was measured using Seahorse assay. GW4869, an exosomal release antagonist, was used to determine the effect of mesenchymal stem cells-derived exosomal signaling on cardiomyocyte recovery. Total exosomal RNA was isolated and differential micro RNA expression determined by sequencing. In vivo studies comprised 48 Yorkshire-Landrace juvenile swine (6 normal controls, 17 HIB, 19 CABG, and 6 CABG + EXP), which were compared for physiologic and metabolic changes. HIB was created by placing a constrictor on the proximal left anterior descending artery, causing significant stenosis but preserved viability by 12 weeks. CABG was performed with or without mesenchymal stem cells-derived EXP application and animals recovered for 4 weeks. Before terminal procedure, cardiac magnetic resonance imaging at rest, and with low-dose dobutamine, assessed diastolic relaxation, systolic function, graft patency, and myocardial viability. Tissue studies of inflammation, fibrosis, and mitochondrial morphology were performed posttermination. RESULTS: In vitro data demonstrated improved cardiomyocyte mitochondrial respiration upon coculture with MSCs that was blunted when adding the exosomal antagonist GW4869. RNA sequencing identified 8 differentially expressed micro RNAs in normoxia vs hypoxia-induced exosomes that may modulate the expression of key mitochondrial (peroxisome proliferator-activator receptor gamma coactivator 1-alpha and adenosine triphosphate synthase) and inflammatory mediators (nuclear factor kappa-light-chain enhancer of activated B cells, interferon gamma, and interleukin 1ß). In vivo animal magnetic resonance imaging studies demonstrated regional systolic function and diastolic relaxation to be improved with CABG + EXP compared with HIB (P = .02 and P = .02, respectively). Histologic analysis showed increased interstitial fibrosis and inflammation in HIB compared with CABG + EXP. Electron microscopy demonstrated increased mitochondrial area, perimeter, and aspect ratio in CABG + EXP compared with HIB or CABG alone (P < .0001). CONCLUSIONS: Exosomes recovered cardiomyocyte mitochondrial respiration and reduced myocardial inflammation through paracrine signaling, resulting in improved cardiac function.

An Adjuvant Stem Cell Patch with Coronary Artery Bypass Graft Surgery Improves Diastolic Recovery in Porcine Hibernating Myocardium.

Diastolic dysfunction persists despite coronary artery bypass graft surgery (CABG) in patients with hibernating myocardium (HIB). We studied whether the adjunctive use of a mesenchymal stem cells (MSCs) patch during CABG improves diastolic function by reducing inflammation and fibrosis. HIB was induced in juvenile swine by placing a constrictor on the left anterior descending (LAD) artery, causing myocardial ischemia without infarction. At 12 weeks, CABG was performed using the left-internal-mammary-artery (LIMA)-to-LAD graft with or without placement of an epicardial vicryl patch embedded with MSCs, followed by four weeks of recovery. The animals underwent cardiac magnetic resonance imaging (MRI) prior to sacrifice, and tissue from septal and LAD regions were collected to assess for fibrosis and analyze mitochondrial and nuclear isolates. During low-dose dobutamine infusion, diastolic function was significantly reduced in HIB compared to the control, with significant improvement after CABG + MSC treatment. In HIB, we observed increased inflammation and fibrosis without transmural scarring, along with decreased peroxisome proliferator-activated receptor-gamma coactivator (PGC1α), which could be a possible mechanism underlying diastolic dysfunction. Improvement in PGC1α and diastolic function was noted with revascularization and MSCs, along with decreased inflammatory signaling and fibrosis. These findings suggest that adjuvant cell-based therapy during CABG may recover diastolic function by reducing oxidant stress-inflammatory signaling and myofibroblast presence in the myocardial tissue.

Persistent diastolic dysfunction in chronically ischemic hearts following coronary artery bypass graft.

OBJECTIVE: A porcine model was used to study diastolic dysfunction in hibernating myocardium (HM) and recovery with coronary artery bypass surgery (CABG). METHODS: HM was induced in Yorkshire-Landrace juvenile swine (n = 30) by placing a c-constrictor on left anterior descending artery causing chronic myocardial ischemia without infarction. At 12 weeks, animals developed the HM phenotype and were either killed humanely (HIB group; n = 11) or revascularized with CABG and allowed 4 weeks of recovery (HIB+CABG group; n = 19). Control pigs were matched for weight, age, and sex to the HIB group. Before the animals were killed humanely, cardiac magnetic resonance imaging (MRI) was done at rest and during a low-dose dobutamine infusion. Tissue was obtained for histologic and proinflammatory biomarker analyses. RESULTS: Diastolic peak filling rate was lower in HIB compared with control (5.4 ± 0.7 vs 6.7 ± 1.4 respectively, P = .002), with near recovery with CABG (6.3 ± 0.8, P = .06). Cardiac MRI confirmed preserved global systolic function in all groups. Histology confirmed there was no transmural infarction but showed interstitial fibrosis in the endomysium in both the HIB and HIB+CABG groups compared with normal myocardium. Alpha-smooth muscle actin stain identified increased myofibroblasts in HM that were less apparent post-CABG. Cytokine and proteomic studies in HM showed decreased peroxisome proliferator-activator receptor gamma coactivator 1-alpha (PGC1-α) expression but increased expression of granulocyte-macrophage colony-stimulating factor and nuclear factor kappa-light-chain enhancer of activated B cells (NFκB). Following CABG, PGC1-α and NFκB expression returned to control whereas granulocyte-macrophage colony-stimulating factor, tumor necrosis factor-α, and interferon gamma remained increased. CONCLUSIONS: In porcine model of HM, increased NFκB expression, enhanced myofibroblasts, and collagen deposition along with decreased PGC1-α expression were observed, all of which tended toward normal with CABG. Estimates of impaired relaxation with MRI within HM during increased workload persisted despite CABG, suggesting a need for adjuvant therapies during revascularization.

Recovery of hibernating myocardium using stem cell patch with coronary bypass surgery.

OBJECTIVE: This study aims to investigate the utility of mesenchymal stem cells (MSCs) applied as an epicardial patch during coronary artery bypass graft (CABG) to target hibernating myocardium; that is, tissue with persistently decreased myocardial function, in a large animal model. METHODS: Hibernating myocardium was induced in juvenile swine (n = 12) using a surgically placed constrictor on the left anterior descending artery, causing stenosis without infarction. After 12 weeks, single-vessel CABG was performed using left internal thoracic artery to left anterior descending artery graft. During CABG, an epicardial patch was applied to the hibernating myocardium region consisting either of MSCs grown onto a polyglactin mesh (n = 6), or sham polyglactin mesh without MSCs (n = 6). Four weeks after CABG and patch placement, cardiac magnetic resonance imaging was performed and cardiac tissue was examined by gross inspection, including coronary dilators for vessel stenosis and patency, electron microscopy, protein assays, and proteomic analysis. RESULTS: CABG + MSC myocardium showed improvement in contractile function (78.24% ± 19.6%) compared with sham patch (39.17% ± 5.57%) during inotropic stimulation (P < .05). Compared with sham patch control, electron microscopy of CABG + MSC myocardium showed improvement in mitochondrial size, number, and morphology; protein analysis similarly showed increases in expression of the mitochondrial biogenesis marker peroxisome proliferator-activated receptor gamma coactivator 1-alpha (0.0022 ± 0.0009 vs 0.023 ± 0.009) (P < .01) along with key components of the electron transport chain, including succinate dehydrogenase (complex II) (0.06 ± 0.02 vs 0.14 ± 0.03) (P < .05) and adenosine triphosphate synthase (complex V) (2.7 ± 0.4 vs 4.2 ± 0.26) (P < .05). CONCLUSIONS: In hibernating myocardium, placement of a stem cell patch during CABG shows promise in improving myocardial function by improving mitochondrial morphology and function.

NHLBI-Sponsored Randomized Trial of Postconditioning During Primary Percutaneous Coronary Intervention for ST-Elevation Myocardial Infarction.

RATIONALE: Postconditioning at the time of primary percutaneous coronary intervention (PCI) for ST-segment-elevation myocardial infarction may reduce infarct size and improve myocardial salvage. However, clinical trials have shown inconsistent benefit. OBJECTIVE: We performed the first National Heart, Lung, and Blood Institute-sponsored trial of postconditioning in the United States using strict enrollment criteria to optimize the early benefits of postconditioning and assess its long-term effects on left ventricular (LV) function. METHODS AND RESULTS: We randomized 122 ST-segment-elevation myocardial infarction patients to postconditioning (4, 30 seconds PTCA [percutaneous transluminal coronary angioplasty] inflations/deflations)+PCI (n=65) versus routine PCI (n=57). All subjects had an occluded major epicardial artery (thrombolysis in myocardial infarction=0) with ischemic times between 1 and 6 hours with no evidence of preinfarction angina or collateral blood flow. Cardiac magnetic resonance imaging measured at 2 days post-PCI showed no difference between the postconditioning group and control in regards to infarct size (22.5±14.5 versus 24.0±18.5 g), myocardial salvage index (30.3±15.6% versus 31.5±23.6%), or mean LV ejection fraction. Magnetic resonance imaging at 12 months showed a significant recovery of LV ejection fraction in both groups (61.0±11.4% and 61.4±9.1%; P<0.01). Subjects randomized to postconditioning experienced more favorable remodeling over 1 year (LV end-diastolic volume =157±34 to 150±38 mL) compared with the control group (157±40 to 165±45 mL; P<0.03) and reduced microvascular obstruction ( P=0.05) on baseline magnetic resonance imaging and significantly less adverse LV remodeling compared with control subjects with microvascular obstruction ( P<0.05). No significant adverse events were associated with the postconditioning protocol and all patients but one (hemorrhagic stroke) survived through 1 year of follow-up. CONCLUSIONS: We found no early benefit of postconditioning on infarct size, myocardial salvage index, and LV function compared with routine PCI. However, postconditioning was associated with improved LV remodeling at 1 year of follow-up, especially in subjects with microvascular obstruction. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov . Unique identifier: NCT01324453.

Magnetic resonance imaging assessment of cardiac function in a swine model of hibernating myocardium 3 months following bypass surgery.

OBJECTIVE: Clinical studies demonstrate delayed recovery of hibernating myocardium (HM) following coronary artery bypass graft (CABG) surgery. Cardiac magnetic resonance (CMR) imaging is effective in identifying HM in clinical settings. Our animal model of HM shows partial but incomplete functional recovery 1 month following CABG using echocardiography. This study uses CMR imaging to determine completeness of recovery 3 months post-CABG. METHODS: Swine (N = 12) underwent left anterior descending artery (LAD) 1.5-cm constrictor placement creating a territory of HM over 12 weeks. CMR at 12 weeks confirmed hibernation without infarction (N = 12). Off-pump left internal thoracic artery (LITA) to the LAD was performed in 9 animals. Three animals were killed as HM controls. CMR imaging was repeated in revascularized animals before death at 1 (n = 4) or 3 months (n = 5). CMR imaging was performed at baseline and with dobutamine infusion (5 µg/kg/min). RESULTS: Twelve weeks after constrictor placement, CMR imaging confirmed viability in LAD region and LAD stenosis in all animals. In HM, wall thickening is reduced at baseline but with contractile reserve present during dobutamine infusion. Following revascularization, CMR imaging confirmed patent LITA graft (n = 9). Analysis of baseline regional function shows incomplete recovery of HM following CABG, with reduced contractile reserve at both 1 and 3 months post-CABG. CONCLUSIONS: CMR imaging provides accurate spatial resolution of regional contractile function and confirms the presence of HM at 12 weeks following instrumentation of the LAD. Three months following CABG, partial recovery of HM with contractile reserve is present in the single LAD territory.

Myocardial ATP hydrolysis rates in vivo: a porcine model of pressure overload-induced hypertrophy.

Left ventricular (LV) hypertrophy (LVH) and congestive heart failure are accompanied by changes in myocardial ATP metabolism. However, the rate of ATP hydrolysis cannot be measured in the in vivo heart with the conventional techniques. Here, we used a double-saturation phosphorous-31 magnetic resonance spectroscopy-magnetization saturation transfer protocol to monitor ATP hydrolysis rate in swine hearts as the hearts became hypertrophic in response to aortic banding (AOB). Animals that underwent AOB (n = 22) were compared with animals that underwent sham surgery (n = 8). AOB induced severe LVH (cardiac MRI). LV function (ejection fraction and systolic thickening fraction) declined significantly, accompanied by deferent levels of pericardial effusion, and wall stress increased in aorta banded animals at week 1 after AOB, suggesting acute heart failure, which recovered by week 8 when concentric LVH restored LV wall stresses. Severe LV dysfunction was accompanied by corresponding declines in myocardial bioenergetics (phosphocreatine-to-ATP ratio) and in the rate of ATP production via creatine kinase at week 1. For the first time, the same linear relationships of the rate increase of the constants of the ATP hydrolysis rate (kATP→Pi) vs. the LV rate-pressure product increase during catecholamine stimulation were observed in vivo in both normal and LVH hearts. Collectively, these observations demonstrate that the double-saturation, phosphorous-31 magnetic resonance spectroscopy-magnetization saturation transfer protocol can accurately monitor myocardial ATP hydrolysis rate in the hearts of living animals. The severe reduction of LV chamber function during the acute phase of AOB is accompanied by the decrease of myocardial bioenergetic efficiency, which recovers as the compensated LVH restores the LV wall stresses.

Cardiac repair in a porcine model of acute myocardial infarction with human induced pluripotent stem cell-derived cardiovascular cells.

Human induced pluripotent stem cells (hiPSCs) hold promise for myocardial repair following injury, but preclinical studies in large animal models are required to determine optimal cell preparation and delivery strategies to maximize functional benefits and to evaluate safety. Here, we utilized a porcine model of acute myocardial infarction (MI) to investigate the functional impact of intramyocardial transplantation of hiPSC-derived cardiomyocytes, endothelial cells, and smooth muscle cells, in combination with a 3D fibrin patch loaded with insulin growth factor (IGF)-encapsulated microspheres. hiPSC-derived cardiomyocytes integrated into host myocardium and generated organized sarcomeric structures, and endothelial and smooth muscle cells contributed to host vasculature. Trilineage cell transplantation significantly improved left ventricular function, myocardial metabolism, and arteriole density, while reducing infarct size, ventricular wall stress, and apoptosis without inducing ventricular arrhythmias. These findings in a large animal MI model highlight the potential of utilizing hiPSC-derived cells for cardiac repair.

Functional consequences of human induced pluripotent stem cell therapy: myocardial ATP turnover rate in the in vivo swine heart with postinfarction remodeling.

BACKGROUND: The use of cells derived from human induced pluripotent stem cells as cellular therapy for myocardial injury has yet to be examined in a large-animal model. METHODS AND RESULTS: Immunosuppressed Yorkshire pigs were assigned to 1 of 3 groups: A myocardial infarction group (MI group; distal left anterior descending coronary artery ligation and reperfusion; n=13); a cell-treatment group (MI with 4×10(6) vascular cells derived from human induced pluripotent stem cells administered via a fibrin patch; n=14); and a normal group (n=15). At 4 weeks, left ventricular structural and functional abnormalities were less pronounced in hearts in the cell-treated group than in MI hearts (P<0.05), and these improvements were accompanied by declines in scar size (10.4±1.6% versus 8.3±1.1%, MI versus cell-treatment group, P<0.05). The cell-treated group displayed a significant increase in vascular density and blood flow (0.83±0.11 and 1.05±0.13 mL·min(-1)·g(-1), MI versus cell-treatment group, P<0.05) in the periscar border zone (BZ), which was accompanied by improvements in systolic thickening fractions (infarct zone, -10±7% versus 5±5%; BZ, 7±4% versus 23±6%; P<0.05). Transplantation of vascular cells derived from human induced pluripotent stem cells stimulated c-kit(+) cell recruitment to BZ and the rate of bromodeoxyuridine incorporation in both c-kit(+) cells and cardiomyocytes (P<0.05). Using a magnetic resonance spectroscopic saturation transfer technique, we found that the rate of ATP hydrolysis in BZ of MI hearts was severely reduced, and the severity of this reduction was linearly related to the severity of the elevations of wall stresses (r=0.82, P<0.05). This decline in BZ ATP utilization was markedly attenuated in the cell-treatment group. CONCLUSIONS: Transplantation of vascular cells derived from human induced pluripotent stem cells mobilized endogenous progenitor cells into the BZ, attenuated regional wall stress, stimulated neovascularization, and improved BZ perfusion, which in turn resulted in marked increases in BZ contractile function and ATP turnover rate.

Novel imaging of atrial septal defects in isolated human hearts.

Within the adult population living with congenital heart defects, approximately 20 % have an atrial septal defect, which suggests that only 0.4-0.05 % of the entire adult population has an atrial septal defect (ASD). In patients with a left to right atrial shunt, treatments may include closure of the defect with a transcatheter device and/or surgical repair. From the perspective of a physician or engineer, it is vitally important to understand the anatomical nuances of such defects, not only to offer the most optimal treatment for the patient but also to call attention to the potential anatomy of ASDs which may go undetected. To do so, we reanimated two human hearts deemed not viable for transplant from 56-year-old and 68-year-old males. Neither patient history reported any heart conditions that would suggest an atrial defect, yet an ASD was found in each heart. Here, we present sets of images, videos, and 3D reconstructions that provide a clearer view of the anatomy of ASDs in functional human hearts. With an enhanced understanding of 3D functional aspects of ASDs, physicians can make improved decisions regarding treatment options and engineers can optimize device designs.

Induced pluripotent stem cells and their potential for basic and clinical sciences.

Induced pluripotent stem (iPS) cells, are a type of pluripotent stem cell derived from adult somatic cells. They have been reprogrammed through inducing genes and factors to be pluripotent. iPS cells are similar to embryonic stem (ES) cells in many aspects. This review summarizes the recent progresses in iPS cell reprogramming and iPS cell based therapy, and describe patient specific iPS cells as a disease model at length in the light of the literature. This review also analyzes and discusses the problems and considerations of iPS cell therapy in the clinical perspective for the treatment of disease.

Noninvasive mapping of transmural potentials during activation in swine hearts from body surface electrocardiograms.

The three-dimensional cardiac electrical imaging (3DCEI) technique was previously developed to estimate the initiation site(s) of cardiac activation and activation sequence from the noninvasively measured body surface potential maps (BSPMs). The aim of this study was to develop and evaluate the capability of 3DCEI in mapping the transmural distribution of extracellular potentials and localizing initiation sites of ventricular activation in an in vivo animal model. A control swine model (n = 10) was employed in this study. The heart-torso volume conductor model and the excitable heart model were constructed based on each animal's preoperative MR images and a priori known physiological knowledge. Body surface potential mapping and intracavitary noncontact mapping (NCM) were simultaneously conducted during acute ventricular pacing. The 3DCEI analysis was then applied on the recorded BSPMs. The estimated initiation sites were compared to the precise pacing sites; as a subset of the mapped transmural potentials by 3DCEI, the electrograms on the left ventricular endocardium were compared to the corresponding output of the NCM system. Over the 16 LV and 48 RV pacing studies, the averaged localization error was 6.1±2.3 mm, and the averaged correlation coefficient between the estimated endocardial electrograms by 3DCEI and from the NCM system was 0.62±0.09. The results demonstrate that the 3DCEI approach can well localize the sites of initiation of ectopic beats and can obtain physiologically reasonable transmural potentials in an in vivo setting during focal ectopic beats. This study suggests the feasibility of tomographic mapping of 3D ventricular electrograms from the body surface recordings.

Bioenergetic and functional consequences of cellular therapy: activation of endogenous cardiovascular progenitor cells.

RATIONALE: The mechanism by which endogenous progenitor cells contribute to functional and beneficial effects in stem cell therapy remains unknown. OBJECTIVE: Utilizing a novel (31)P magnetic resonance spectroscopy-2-dimensional chemical shift imaging method, this study examined the heterogeneity and bioenergetic consequences of postinfarction left ventricular (LV) remodeling and the mechanisms of endogenous progenitor cell contribution to the cellular therapy. METHODS AND RESULTS: Human embryonic stem cell-derived vascular cells (hESC-VCs) that stably express green fluorescent protein and firefly luciferase (GFP(+)/Luc(+)) were used for the transplantation. hESC-VCs may release various cytokines to promote angiogenesis, prosurvival, and antiapoptotic effects. Both in vitro and in vivo experiments demonstrated that hESC-VCs effectively inhibit myocyte apoptosis. In the mouse model, a fibrin patch-based cell delivery resulted in a significantly better cell engraftment rate that was accompanied by a better ejection fraction. In the swine model of ischemia-reperfusion, the patch-enhanced delivery of hESC-VCs resulted in alleviation of abnormalities including border zone myocardial perfusion, contractile dysfunction, and LV wall stress. These results were also accompanied by a pronounced recruitment of endogenous c-kit(+) cells to the injury site. These improvements were directly associated with a remarkable improvement in myocardial energetics, as measured by a novel in vivo (31)P magnetic resonance spectroscopy-2-dimensional chemical shift imaging technology. CONCLUSIONS: The findings of this study demonstrate that a severely abnormal heterogeneity of myocardial bioenergetics in hearts with postinfarction LV remodeling can be alleviated by the hESC-VCs therapy. These findings suggest an important therapeutic target of peri-scar border zone and a promising therapeutic potential for using hESC-VCs together with the fibrin patch-based delivery system.

Ex vivo diffusion tensor MRI of human hearts: relative effects of specimen decomposition.

Recently, diffusion tensor MRI has been utilized as a tool to reconstruct, with high resolution, the three-dimensional myofiber orientation of hearts ex vivo. However, because of limited availability of human hearts, it is common to recover specimens postmortem, after some degree of decomposition has occurred. In this study, we describe the ability to reconstruct local fiber orientation within a unique set of human hearts using diffusion tensor MRI; relative effects of specimen decomposition were also assessed. The mean diffusivity, fractional anisotropy, and principal eigenvalues (E1, E2, and E3) were measured in hearts with varying postmortem intervals from 0 to 27 days (n = 17), and local fiber orientations were reconstructed. Mean diffusivities and principal eigenvalues nonlinearly increased as a function of postmortem interval, while fractional anisotropies nonlinearly decreased. There were good correlations between postmortem intervals and changes in diffusion properties after linear transformation of the data (R(2) > 0.7). Thus, we determined that there were regional discontinuities in the reconstruction of local fiber orientation in hearts where postmortem intervals were greater than 3 days. Therefore, any delay in tissue fixation after death (if not properly preserved using organ transplantation techniques) will compromise measurement of fiber orientation and delineation of diseased pathology, e.g., based, in part, on fractional anisotropy.

Circadian dependence of infarct size and left ventricular function after ST elevation myocardial infarction.

RATIONALE: In rodents, infarct size after ischemia/reperfusion exhibits a circadian dependence on the time of coronary occlusion. It is not known if a similar circadian dependence of infarct size occurs in humans. OBJECTIVE: To determine if humans exhibit a circadian dependence of infarct size in the setting of ST elevation myocardial infarction (STEMI). METHODS AND RESULTS: A retrospective analysis of 1031 patients with STEMI referred for primary percutaneous coronary intervention with known ischemic times between 1 and 6 hours identified 165 patients with occluded arteries on presentation without evidence of preinfarction angina or collateral blood flow. Both ischemic duration and angiographic area at risk were not dependent on time of infarct onset. We observed that the extent of infarct size measured by creatine kinase release was significantly associated with time of day onset of infarction (P<0.0001). The greatest myocardial injury occurred at 1:00 am onset of ischemia and 5:00 am onset of reperfusion, with the peak creatine kinase measured at the peak of the curve being 82% higher than that recorded at the trough. Similarly, left ventricular ejection fraction measured within 2 days of infarction was also dependent on time of onset of STEMI with the absolute left ventricular ejection fraction at peak >7% higher than at trough (43% vs 51%; P<0.03). These findings were supported by a subgroup of patients (n = 45) who underwent cardiac MRI measurements of infarct size and area-at-risk measurements. CONCLUSIONS: The results of this study demonstrate for the first time in humans that myocardial infarct size and left ventricular function after STEMI have a circadian dependence on the time of day onset of ischemia.

A fibrin patch-based enhanced delivery of human embryonic stem cell-derived vascular cell transplantation in a porcine model of postinfarction left ventricular remodeling.

It is unknown how to use human embryonic stem cell (hESC) to effectively treat hearts with postinfarction left ventricular (LV) remodeling. Using a porcine model of postinfarction LV remodeling, this study examined the functional improvement of enhanced delivery of combined transplantation of hESC-derived endothelial cells (ECs) and hESC-derived smooth muscle cells (SMCs) with a fibrin three-dimensional (3D) porous scaffold biomatrix. To facilitate tracking the transplanted cells, the hESCs were genetically modified to stably express green fluorescent protein and luciferase (GFP/Luc). Myocardial infarction (MI) was created by ligating the first diagonal coronary artery for 60 minutes followed by reperfusion. Two million each of GFP/Luc hESC-derived ECs and SMCs were seeded in the 3D porous biomatrix patch and applied to the region of ischemia/reperfusion for cell group (MI+P+C, n = 6), whereas biomatrix without cell (MI+P, n = 5), or saline only (MI, n = 5) were applied to control group hearts with same coronary artery ligation. Functional outcome (1 and 4 weeks follow-up) of stem cell transplantation was assessed by cardiac magnetic resonance imaging. The transplantation of hESC-derived vascular cells resulted in significant LV functional improvement. Significant engraftment of hESC-derived cells was confirmed by both in vivo and ex vivo bioluminescent imaging. The mechanism underlying the functional beneficial effects of cardiac progenitor transplantation is attributed to the increased neovascularization. These findings demonstrate a promising therapeutic potential of using these hESC-derived vascular cell types and the mode of patch delivery.

Long-term preservation of myocardial energetic in chronic hibernating myocardium.

We previously reported that the myocardial energetic state, as defined by the ratio of phosphocreatine to ATP (PCr/ATP), was preserved at baseline (BL) in a swine model of chronic myocardial ischemia with mild reduction of myocardial blood flow (MBF) 10 wk after the placement of an external constrictor on the left anterior descending coronary artery. It remains to be seen whether this stable energetic state is maintained at a longer-term follow-up. Hibernating myocardium (HB) was created in minipigs (n = 7) by the placement of an external constrictor (1.25 mm internal diameter) on the left anterior descending coronary artery. Function was assessed with MRI at regular intervals until 6 mo. At 6 mo, myocardial energetic in the HB was assessed by (31)P-magnetic resonance spectrometry and myocardial oxygenation was examined from the deoxymyoglobin signal using (1)H-magnetic resonance spectrometry during BL, coronary vasodilation with adenosine, and high cardiac workload with dopamine and dobutamine (DpDb). MBF was measured with radiolabeled microspheres. At BL, systolic thickening fraction was significantly lower in the HB compared with remote region (34.4 ± 9.4 vs. 50.1 ± 10.7, P = 0.006). This was associated with a decreased MBF in the HB compared with the remote region (0.73 ± 0.08 vs. 0.97 ± 0.07 ml · min(-1) · g, P = 0.03). The HB PCr/ATP at BL was normal. DpDb resulted in a significant increase in rate pressure product, which caused a twofold increase in MBF in the HB and a threefold increase in the remote region. The systolic thickening fraction increased with DpDb, which was significantly higher in the remote region than HB (P < 0.05). The high cardiac workload was associated with a significant reduction in the HB PCr/ATP (P < 0.02), but this response was similar to normal myocardium. Thus HB has stable BL myocardial energetic despite the reduction MBF and regional left ventricular function. More importantly, HB has a reduced contractile reserve but has a similar energetic response to high cardiac workload like normal myocardium.

Long-term functional improvement and gene expression changes after bone marrow-derived multipotent progenitor cell transplantation in myocardial infarction.

The study examined the long-term outcome of cardiac stem cell transplantation in hearts with postinfarction left ventricular (LV) remodeling. Myocardial infarction (MI) was created by ligating the first and second diagonal branches of the left anterior descending coronary artery in miniature swine. Intramyocardial injections of 50 million LacZ-labeled bone marrow-derived multipotent progenitor cells (MPC) were performed in the periscar region (Cell, n = 7) immediately after MI, whereas, in control animals (Cont, n = 7), saline was injected. Functional outcome was assessed monthly for 4 mo with MRI and (31)P-magnetic resonance spectroscopy. Engraftment was studied on histology, and gene chip (Affymetrix) array analysis was used to study differential expression of genes in the two groups. MPC treatment resulted in improvement of ejection fraction as early as 10 days after MI (Cell, 43.4 +/- 5.1% vs. Cont, 32.2 +/- 5.5%; P < 0.05). This improvement was seen each month and persisted to 4 mo (Cell, 51.2 +/- 4.8% vs. Cont, 35.7 +/- 5.0%; P < 0.05). PCr-to-ATP ratio (PCr/ATP) improved with MPC transplantation, which was most pronounced at high cardiac work states (subendocardial PCr/ATP was 1.70 +/- 0.10 vs. 1.34 +/- 0.14, P < 0.05). There was no significant difference in scar size (scar/LV area * 100) at 10 days postinfarction. However, at 4 mo, there was a significant decrease in scar size in the Cell group (Cell, 4.6 +/- 1.0% vs. Cont, 8.6 +/- 2.4%; P < 0.05). No significant engraftment of MPC was observed. MPC transplantation was associated with a downregulation of mitochondrial oxidative enzymes and increased levels of myocyte enhancer factor 2a and zinc finger protein 91. In conclusion, MPC transplantation leads to long-term functional and bioenergetic improvement in a porcine model of postinfarction LV remodeling, despite no significant engraftment of stem cells in the heart. MPC transplantation reduces regional wall stresses and infarct size and mitigates the adverse effects of LV remodeling, as seen by a reduction in LV hypertrophy and LV dilatation, and is associated with differential expression of genes relating to metabolism and apoptosis.