Positive Cardiac Cells Research Articles

Comparison of Repeated Doses of C-kit-Positive Cardiac Cells versus a Single Equivalent Combined Dose in a Murine Model of Chronic Ischemic Cardiomyopathy

Using a murine model of chronic ischemic cardiomyopathy caused by an old myocardial infarction (MI), we have previously found that three doses of 1 × 106 c-kit positive cardiac cells (CPCs) are more effective than a single dose of 1 × 106 cells. The goal of this study was to determine whether the beneficial effects of three doses of CPCs (1 × 106 cells each) can be fully replicated by a single combined dose of 3 × 106 CPCs. Mice underwent a 60-min coronary occlusion; after 90 days of reperfusion, they received three echo-guided intraventricular infusions at 5-week intervals: (1) vehicle × 3; (2) one combined dose of CPCs (3 × 106) and vehicle × 2; or (3) three doses of CPCs (1 × 106 each). In the combined-dose group, left ventricular ejection fraction (LVEF) improved after the 1st CPC infusion, but not after the 2nd and 3rd (vehicle) infusions. In contrast, in the multiple-dose group, LVEF increased after each CPC infusion; at the final echo, LVEF averaged 35.2 ± 0.6% (p < 0.001 vs. the vehicle group, 27.3 ± 0.2%). At the end of the study, the total cumulative change in EF from pretreatment values was numerically greater in the multiple-dose group (6.6 ± 0.6%) than in the combined-dose group (4.8 ± 0.8%), although the difference was not statistically significant (p = 0.08). Hemodynamic studies showed that several parameters of LV function in the multiple-dose group were numerically greater than in the combined-dose group (p = 0.08 for the difference in LVEF). Compared with vehicle, cardiomyocyte cross-sectional area was reduced only in the multiple-dose group (−32.7%, 182.6 ± 15.1 µm2 vs. 271.5 ± 27.2 µm2, p < 0.05, in the risk region and −28.5%, 148.5 ± 12.1 µm2 vs. 207.6 ± 20.5 µm2, p < 0.05, in the noninfarcted region). LV weight/body weight ratio and LV weight/tibia length ratios were significantly reduced in both cell treated groups vs. the vehicle group, indicating the attenuation of LV hypertrophy; however, the lung weight/body weight ratio was significantly reduced only in the multiple-dose group, suggesting decreased pulmonary congestion. Taken together, these results indicate that in mice with chronic ischemic cardiomyopathy, the beneficial effects of three doses of CPCs on LV function and hypertrophy cannot be fully replicated with a single dose, notwithstanding the fact that the total number of cells delivered with one or three doses is the same. Thus, it is the multiplicity of doses, and not the total number of cells, that accounts for the superiority of the repeated-dose paradigm. This study supports the idea that the efficacy of cell therapy in heart failure can be augmented by repeated administrations.

Abstract 368: Ex vivo Fabrication of Heart Tissue Model With Perfusable Blood Vessels Using Cell Sheet

Introduction: Creation of 3-D tissue model similar to a biological tissue using tissue engineering method are expected as an alternative approach to 2-D culture or animal test in a research for developing a new medicine. We have already developed functional 3-D heart tissue with perfusable blood vessels in vitro using cell sheet based tissue engineering approach. Present study assessed the possibility as 3-D tissue model by performing drug evaluation in the engineered heart tissues. Methods: Firefly luciferase positive cardiac cells co-cultured with endothelial cells were harvested as cell sheets from temperature-responsive culture dishes. Triple-layer cell sheets were overlaid on the vascular bed with an artery and a vein in vitro. Layered cardiac cell sheets were maintained in custom made bioreactor system with perfusion medium. After neovascularization within the first triple-layer tissue, the second triple-layer cardiac cells sheets were overlaid on the first graft after cultivation for 3 days and then the whole construct was further perfused for an additional 3 days. After 6-day perfusion culture, isoproterenol or doxorubicin were administrated to the engineered heart tissues from artery side. The sympathetic and toxic effects were measured by in vitro bioluminescence imaging, electrical potential, and blood pressure. Results: In case of isoproterenol administration, bioluminescence imaging (BLI) demonstrated that adenosine triphosphate (ATP) of the heart tissue improved. In addition, electrograms demonstrated increasing beating rate and amplitude. Moreover, sphygmomanometer showed decreasing arterial pressure. In case of doxorubicin administration, BLI showed that the ATP of the heart tissue worsened after 24 hours. It is now possible to analyze the pharmacological effects such as metabolism and the blood pressure as well as action potential by using our vascularized heart tissue model. Conclusions: We believe that our 3-D heart tissue model offers a new possibilities for evaluation of drug efficacy.

Flos Puerariae extract prevents myocardial apoptosis via attenuation oxidative stress in streptozotocin-induced diabetic mice.

BackgroundDiabetic cardiomyopathy (DCM) suggests a direct cellular insult to myocardium. Apoptosis is considered as one of the hallmarks of DCM. Oxidative stress plays a key role in the pathogenesis of DCM. In this study, we explored the prevention of myocardial apoptosis by crude extract from Flos Puerariae (FPE) in experimental diabetic mice.MethodsExperimental diabetic model was induced by intraperitoneally injection of streptozotocin (STZ, 50 mg/kg/day) for five consecutive days in C57BL/6J mice. FPE (100, 200 mg/kg) was orally administrated once a day for ten weeks. Cardiac structure changes, apoptosis, superoxide production, NADPH oxidase subunits expression (gp91phox, p47phox, and p67phox), and related regulatory factors were assessed in the heart of mice.ResultsDiabetic mice were characterized by high blood glucose (≥11.1 mmol/L) and reduced body weight. In the end of the experiment, aberrant myofilament structure, as well as TUNEL positive cardiac cells coupled with increased Bax/Bcl-2 ratio and Caspase-3 expression was found in diabetic mice. Moreover, ROS formation, the ratio of NADP+/NADPH and NADPH oxidase subunits expression of gp91phox and p47phox, lipid peroxidation level was significantly increased, while antioxidant enzyme SOD and GSH-Px activity were reduced in the myocardial tissue of diabetic mice. In contrast, treatment with FPE resulted in a normalized glucose and weight profile. FPE administration also preserved myocardial structure and reduced apoptotic cardiac cell death in diabetic mice. The elevated markers of oxidative stress were significantly reversed by FPE supplementation. Further, FPE treatment markedly inhibited the increased Bax/Bcl-2 ratio and Caspase-3 expression, as well as suppressed JNK and P38 MAPK activation in the heart of diabetic mice.ConclusionsOur data demonstrate for the first time that FPE may have therapeutic potential for STZ-induced diabetic cardiomyopathy through preventing myocardial apoptosis via attenuation oxidative stress. And this effect is probably mediated by JNK and P38 MAPK signaling pathway.

Injectable Small Intestine Submucosal Extracellular Matrix in an Acute Myocardial Infarction Model

The mechanisms involved in myocardial regeneration and cardiac remodeling were examined by injecting porcine-derived small intestine submucosal extracellular matrix (SIS-ECM), with and without circulating angiogenic cells (CACs), in a mouse model of acute myocardial infarction (MI). Nine- to 10-week-old female C57BL/6J mice had the left anterior descending (LAD) coronary artery ligated. Seven days after ligation, 38 randomly allocated animals received echocardiographically guided intramyocardial injections of phosphate buffered saline (PBS), CACs, SIS-ECM, or SIS-ECM + CACs. Repeated echocardiography and immunohistochemical analysis were performed at 28 days after ligation. Baseline postligation left ventricular ejection fraction (LVEF) was equivalent in all groups. Twenty-one days after treatment, ejection fraction improved in the SIS-ECM + CAC treatment group (by 38% ± 2.12%) and the SIS-ECM treatment group (by 36% ± 3.71%), compared with the CAC-alone and PBS treatment groups (p < 0.1). Masson's trichrome staining showed reduced infarct size in SIS-ECM + CACs (34.2% ± 3.1%) and SIS-ECM alone (34.5% ± 4.7%) compared with CACs alone (47.3% ± 6.0%) and PBS (61.9% ± 5.5%; p < 0.002). Arteriolar density in periinfarct regions was enhanced in both SIS-ECM-treated groups (by ≥ 78% ± 7%; p = 0.03). More GATA4- and β-catenin-positive cardiac cells were found in the myocardium of SIS-ECM-treated animals. Intramyocardial delivery of SIS-ECM 7 days after MI in a mouse model reduced infarct size and improved myocardial vessel density and function; when combined with CACs it helped restore myocardial cellularity, suggesting a potential therapeutic role for SIS-ECM in cardiac regeneration.

Multiple Antioxidants Improve Cardiac Complications and Inhibit Cardiac Cell Death in Streptozotocin-Induced Diabetic Rats

Diabetic cardiomyopathy, a disorder of the heart muscle in diabetic patients, is one of the major causes of heart failure. Since diabetic cardiomyopathy is now known to have a high prevalence in the asymptomatic diabetic patient, prevention at the earliest stage of development by existing molecules would be appropriate in order to prevent the progression of heart failure. In this study, we investigated the protective role of multiple antioxidants (MA), on cardiac dysfunction and cardiac cell apoptosis in streptozotocin (STZ)-induced diabetic rat. Diabetic cardiomyopathy in STZ-treated animals was characterized by declined systolic, diastolic myocardial performance, oxidative stress and apoptosis in cardiac cells. Diabetic rats on supplementation with MA showed decreased oxidative stress evaluated by the content of reduced levels of lipid per-oxidation and decreased activity of catalase with down-regulation of heme-oxygenase-1 mRNA. Supplementation with MA also resulted in a normalized lipid profile and decreased levels of pro-inflammatory transcription factor NF-kappaB as well as cytokines such as TNF-α, IFN-γ, TGF-β, and IL-10. MA was found to decrease the expression of ROS-generating enzymes like xanthine oxidase, monoamine oxidase-A along with 5-Lipoxygenase mRNA and/or protein expression. Further, left ventricular function, measured by a microtip pressure transducer, was re-established as evidenced by increase in ±dp/dtmax, heart rate, decreased blood pressure, systolic and diastolic pressure as well as decrease in the TUNEL positive cardiac cells with increased Bcl-2/Bax ratio. In addition, MA supplementation decreased cell death and activation of NF-kappaB in cardiac H9c2 cells. Based on our results, we conclude that MA supplementation significantly attenuated cardiac dysfunction in diabetic rats; hence MA supplementation may have important clinical implications in terms of prevention and management of diabetic cardiomyopathy.

Cardiotoxicity induced in mice by long term ketamine and ketamine plus alcohol treatment

Increased use of ketamine as a recreational drug in rave party has raised lots of social concerns. Although its toxicity on the central nervous system (CNS) and urinary system had been reported, its potential adverse effects on the heart is still not addressed. Using our long term ketamine addiction model in which, 1-month-old ICR mice were given ketamine intraperitoneally (i.p. 30mg/kg) for different study periods (6 weeks, 16 weeks and 28 weeks). The electrocardiogram (ECG) and the level of troponin as well as the presence of lactate dehydrogenase were used together to determine the cardiac function of mice. ST elevations were observed in 30% of mice from 6 weeks onwards. Concomitant increase in troponin in serum and presence of lactate dehydrogenase positive cardiac cells were taken together to indicate cardiac damages. The potential interactive effects of alcohol (ethanol) and ketamine were also addressed and 0.5ml of 10% ethanol was administrated additionally to the mice daily and orally for the last 4 weeks in the 28-week long-term ketamine treated animals. The cardiac functions of these mice deteriorated more severely as indicated by further rise in troponin levels. In addition, high number of lactate dehydrogenase positive cardiac cells and abnormal ECGs in both ketamine and ketamine plus alcohol treated models also gave coherent conclusions.

Micro and nano‐scale in vitro 3D culture system for cardiac stem cells

Despite the success to prevent or limit cardiovascular diseases, the restoration of the function of a damaged heart remains a formidable challenge. Cardiac stem cells (CSCs), with the capacity to differentiate into cardiomyocytes, hold great potential as a source of cells for regenerative medicine. A major challenge facing the clinical application of differentiated CSCs, however, is theability to generate sufficient numbers of cells with the desired phenotype. We previously established cell lines of CSCs using a c-kit antibody from adult rat hearts for use in regenerative medicine. C-kit -positive cardiac cells are well recognized as CSCs and have the potential to differentiate into cardiomyocytes. Here, before implant these cells in vivo, we first developed three-dimensional culture system (3D) using micro- and nano-scaled material. Sheets of poly(glycolic acid) (PGA) were fabricated by electrospinning. Composites of collagen-PGA were prepared that contained 0, 1.5, 3 or 6 mg of electrospun PGA nanofibers. The nanofibers were added as a sheet that formed a layer within the collagen sponge. The sponges were freeze-dried and then dehydrothermally crosslinked. A scanning electron microscopy (SEM)-based analysis of the surface of the sponges demonstrated a uniform collagenous structure regardless of the amount of PGA nanofibres included. The PGA nanofibers significantly enhanced the compressive strength of the collagen sponge. More CSCs attached to the collagen sponge incorporating 6 mg of PGA nanofibers than the sponge without PGA nanofibers. The attachment and proliferation of CSCs in the 3D culture was enhanced by incubation in a bioreactor perfusion system compared with 3D static and two-dimensional (2D; i.e. tissue culture plates) culture systems. The use of micro- and nano-scale materials in the fabrication of composites together with a 3D culture system is a very promising way to promote the culture of stem cells. (c) 2009 Wiley Periodicals, Inc. J Biomed Mater Res, 2010.