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Ischemic heart disease, which is one of the top killers worldwide, encompasses a series of heart problems stemming from a compromised coronary blood supply to the myocardium. The severity of the disease ranges from an unstable manifestation of ischemic symptoms, such as unstable angina, to myocardial death, that is, the immediate life-threatening condition of myocardial infarction. Even though patients may survive myocardial infarction, the resulting ischemia-reperfusion injury triggers a cascade of inflammatory reactions and oxidative stress that poses a significant threat to myocardial function following successful revascularization. Moreover, despite evidence suggesting the presence of cardiac stem cells, the fact that cardiomyocytes are terminally differentiated and cannot significantly regenerate after injury accounts for the subsequent progression to ischemic cardiomyopathy and ischemic heart failure, despite the current advancements in cardiac medicine. In the last two decades, researchers have realized the possibility of utilizing stem cell plasticity for therapeutic purposes. Indeed, stem cells of different origin, such as bone-marrow- and adipose-derived mesenchymal stem cells, circulation-derived progenitor cells, and induced pluripotent stem cells, have all been shown to play therapeutic roles in ischemic heart disease. In addition, the discovery of stem-cell-associated paracrine effects has triggered intense investigations into the actions of exosomes. Notwithstanding the seemingly promising outcomes from both experimental and clinical studies regarding the therapeutic use of stem cells against ischemic heart disease, positive results from fraud or false data interpretation need to be taken into consideration. The current review is aimed at overviewing the therapeutic application of stem cells in different categories of ischemic heart disease, including relevant experimental and clinical outcomes, as well as the proposed mechanisms underpinning such observations. [ABSTRACT FROM AUTHOR]

Ischemic heart disease, which is one of the top killers worldwide, encompasses a series of heart problems stemming from a compromised coronary blood supply to the myocardium. The severity of the disease ranges from an unstable manifestation of ischemic symptoms, such as unstable angina, to myocardial death, that is, the immediate life-threatening condition of myocardial infarction. Even though patients may survive myocardial infarction, the resulting ischemia-reperfusion injury triggers a cascade of inflammatory reactions and oxidative stress that poses a significant threat to myocardial function following successful revascularization. Moreover, despite evidence suggesting the presence of cardiac stem cells, the fact that cardiomyocytes are terminally differentiated and cannot significantly regenerate after injury accounts for the subsequent progression to ischemic cardiomyopathy and ischemic heart failure, despite the current advancements in cardiac medicine. In the last two decades, researchers have realized the possibility of utilizing stem cell plasticity for therapeutic purposes. Indeed, stem cells of different origin, such as bone-marrow- and adipose-derived mesenchymal stem cells, circulation-derived progenitor cells, and induced pluripotent stem cells, have all been shown to play therapeutic roles in ischemic heart disease. In addition, the discovery of stem-cell-associated paracrine effects has triggered intense investigations into the actions of exosomes. Notwithstanding the seemingly promising outcomes from both experimental and clinical studies regarding the therapeutic use of stem cells against ischemic heart disease, positive results from fraud or false data interpretation need to be taken into consideration. The current review is aimed at overviewing the therapeutic application of stem cells in different categories of ischemic heart disease, including relevant experimental and clinical outcomes, as well as the proposed mechanisms underpinning such observations.

Objective. Improvement of the treatment results in patients with refractory angina, complicated by the heart insufficiency, using new methods of correction of the heart pumping function elaborated. Materials and methods. Clinical investigation was performed, basing on retro- and prospective analysis of the treatment results in 156 patients: efficacy of myocardial remodeling was studied, using mesenchymal stem cells in patients, suffering refractory angina. For determination of the cellular therapy efficacy a navigation system NOGA XP was applied. Results. There were presented the results of investigation of efficacy of intravenous injections of autologous mesenchymal stem cells to patients, suffering refractory angina, complicated by lowering of the heart pumping function, and immediate results of cardiomyoplasty using stem cells as well. For the first time a navigation system NOGA XP for determination of myocardial hibernation zones was applied in patients, suffering refractory angina. Also, using navigation system NOGA XP, the results of cellular cardiomyoplasty were obtained, while comparing electro-mechanical schemes before and after cellular therapy. Conclusion. There was established the improvement of indices of the heart pumping function and the patients’ quality of life.

Aims Cardiomyocytes are understood to possess a limited regenerative capacity. Any myocardial insult leads to an irreversible injury. Mesenchymal stem cell differentiation into cardiomyocyte-like cells stands as one of the leading experimental therapies. However, a candidate cell source has yet to be defined. Here, we examined the in vitro and in vivo cardiac differentiation potential of human placenta derived stem cells (hPDSCs); a unique, abundant, and non-immunogenic cell source. Main methods H9c2 cell culture medium was applied to hPDSCs at different ratios for a period of 4 weeks. In parallel, hPDSCs, human bone marrow stem cells, or cell free culture medium was injected in peri-infarcted regions induced in rat hearts. Key findings In vitro , hPDSCs pre-conditioned with H9c2 cell culture medium proportionally over-expressed alpha sarcoplasmic actinin and displaced connexin 43 from the cytoplasm to the cell membrane. Additionally, pre-conditioning promoted hPDSCs survival and triggered vascular endothelial growth factor (VEGF) dependent angiogenesis by activating the pAkt and p38MAPK/pSTAT3 pathways. In vivo , echocardiography analysis showed a significant improvement in cardiac parameters in the rats injected with hPDSCs, similar to the human bone marrow stem cells injected group. Moreover, hPDSCs detected within rat cardiac tissues expressed troponin I and myosin heavy chain. In accordance with the pre-conditioning findings, VEGF positive neovessels were observed in hearts injected with hPDSCs. Significance hPDSCs have the potential to differentiate into cardiac-like cells and induce angiogenesis via paracrine effects. With the advantages of easy availability and young age, these cells could be more suitable for clinical translation. [ABSTRACT FROM AUTHOR]

Although the induction of neovascularization by cell-based approaches has demonstrated substantial potential in treating myocardial infarction (MI), the process of cell-mediated angiogenesis and its correlation with therapeutic mechanisms of cardiac repair remain elusive. In this work, three-dimensional (3D) aggregates of human umbilical vein endothelial cells (HUVECs) and cord-blood mesenchymal stem cells (cbMSCs) are constructed using a methylcellulose hydrogel system. By maximizing cell–cell and cell–ECM communications and establishing a hypoxic microenvironment in their inner cores, these cell aggregates are capable of forming widespread tubular networks together with the angiogenic marker α v β 3 integrin; they secret multiple pro-angiogenic, pro-survival, and mobilizing factors when grown on Matrigel. The aggregates of HUVECs/cbMSCs are exogenously engrafted into the peri-infarct zones of rats with MI via direct local injection. Multimodality noninvasive imaging techniques, including positron emission tomography, single photon emission computed tomography, and echocardiography, are employed to monitor serially the beneficial effects of cell therapy on angiogenesis, blood perfusion, and global/regional ventricular function, respectively. The myocardial perfusion is correlated with ventricular contractility, demonstrating that the recovery of blood perfusion helps to restore regional cardiac function, leading to the improvement in global ventricular performance. These experimental data reveal the efficacy of the exogenous transplantation of 3D cell aggregates after MI and elucidate the mechanism of cell-mediated therapeutic angiogenesis for cardiac repair. [ABSTRACT FROM AUTHOR]

Abstract: In this work, the effectiveness of using Gadonanotubes (GNTs) with an external magnetic field to improve retention of transplanted adult mesenchymal stem cells (MSCs) during cellular cardiomyoplasty was evaluated. As a high-performance T1-weighted magnetic resonance imaging (MRI) cell tracking label, the GNTs are gadolinium-loaded carbon nanotube capsules that render MSCs magnetic when internalized. MSCs were internally labeled with either superparamagnetic GNTs or colloidal diamagnetic lutetium (Lu). In vitro cell rolling assays and ex vivo cardiac perfusion experiments qualitatively demonstrated increased magnetic-assisted retention of GNT-labeled MSCs. Subsequent in vivo epicardial cell injections were performed around a 1.3 T NdFeB ring magnet sutured onto the left ventricle of female juvenile pigs (n = 21). Cell dosage, magnet exposure time, and endpoints were varied to evaluate the safety and efficacy of the proposed therapy. Quantification of retained cells in collected tissues by elemental analysis (Gd or Lu) showed that the external magnet helped retain nearly three times more GNT-labeled MSCs than Lu-labeled cells. The sutured magnet was tolerated for up to 168 h; however, an inflammatory response to the magnet was noted after 48 h. These proof-of-concept studies support the feasibility and value of using GNTs as a magnetic nanoparticle facilitator to improve cell retention during cellular cardiomyoplasty. [Copyright &y& Elsevier]

Abstract: Transplantation of human cardiomyoblast-like cells (hCLCs) from human adipose tissue-derived multi-lineage progenitor cells improved left ventricular function and survival of rats with myocardial infarction. Here we examined the effect of intracoronary artery transplantation of human CLCs in a swine model of chronic heart failure. Twenty-four pigs underwent balloon-occlusion of the first diagonal branch followed by reperfusion, with a second balloon-occlusion of the left ascending coronary artery 1week later followed by reperfusion. Four weeks after the second occlusion/reperfusion, 17 of the 18 surviving animals with severe chronic MI (ejection fraction <35% by echocardiography) were immunosuppressed then randomly assigned to receive either intracoronary artery transplantation of hCLCs hADMPCs or placebo lactic Ringer’s solution with heparin. Intracoronary artery transplantation was followed by the distribution of DiI-stained hCLCs into the scarred myocardial milieu. Echocardiography at post-transplant days 4 and 8weeks showed rescue and maintenance of cardiac function in the hCLCs transplanted group, but not in the control animals, indicating myocardial functional recovery by hCLCs intracoronary transplantation. At 8week post-transplantation, 7 of 8 hCLCs transplanted animals were still alive compared with only 1 of the 5 control (p =0.0147). Histological studies at week 12 post-transplantation demonstrated engraftment of the pre DiI-stained hCLCs into the scarred myocardium and their expression of human specific alpha-cardiac actin. Human alpha cardiac actin-positive cells also expressed cardiac nuclear factors; nkx2.5 and GATA-4. Our results suggest that intracoronary artery transplantation of hCLCs is a potentially effective therapeutic strategy for future cardiac tissue regeneration. [Copyright &y& Elsevier]

Noninvasive or minimally invasive imaging techniques are essential for developing strategies and assessing outcomes of cell-based therapies for myocardial regeneration, also referred to as cellular cardiomyoplasty. Imaging-based monitoring of cell survival is useful for selection of optimal cell type and evaluating strategies to enhance engraftment. Imaging-derived surrogate end points including global and regional contractile function, myocardial blood flow, or perfusion and bioenergetics have been used in clinical trials or in relevant large animal models to evaluate the therapeutic effect and mechanisms of action of cellular cardiomyoplasty. New techniques are emerging to assess electrical integration of donor cells with host cardiomyocytes. This review will summarize and highlight important and informative findings revealed by imaging in clinical and preclinical cellular cardiomyoplasty studies over the past 3 years. [ABSTRACT FROM AUTHOR]

Our study aimed to elucidate whether bone marrow stem cell (BMC) treatment might result in a cellular response in cardiomyocytes in vitro. Subconfluent neonatal rat cardiomyocyte cultures were cocultured for three days with Vybrant CM‐DiI labeled BMC from human sternal bone marrow and underwent immunohistological staining for the proto-oncogene c-Myc and the cell cycle proteins CDK2, CDK4 and ATF-3. β-adrenoceptor density was analyzed using [125I]-iodocyanopindolol (ICYP) histoautoradiography. Quantitative analysis of immunohistochemical images revealed significantly increased expression and upregulation of c-Myc, and its downstream targets ATF-3, CDK2 and CDK4 in neighboring cardiomyocytes to BMC, depending on their distance to the BMC compared to cardiomyocytes far from the BMC. Histoautoradiography revealed a significantly higher β-adrenoceptor density in cardiomyocytes in the immediate vicinity to the BMC. With increasing distance to the BMC, β-adrenoceptor density in cardiomyocytes declined. Thus, a small number of BMC can affect a larger number of cardiomyocytes by activating an intracellular signaling cascade and enhancing β-adrenoceptor density. [ABSTRACT FROM AUTHOR]

This study was designed to compare the efficacy of intraventricular (IV), intramyocardial (IM) or combined IV and IM injections of human adipose tissue-derived stem cells (hADSCs) administered either immediately, 5, or 10 days after induction of acute myocardial infarction (AMI) in a nude rat model. Acute myocardial infarction was induced in 99, adult (250-350 g BW), male nude rats strain Crl:NIH-Fox1. Rats received either no cells (group I, n = 15) or 2 million, hADSCs as follows: group II ( n = 19) IV injection immediately after AMI; group III ( n = 18) IV injection 5 days after AMI; group IV ( n = 15) IV injection 5 days and IM injection 10 days after AMI; group V ( n = 17) IV injection immediately after AMI and IM injection 10 days after AMI and group VI ( n = 15) IM injection 10 days after AMI. Tissue sections from hearts were studied using H&E and immunohistochemistry. In the control group, there was a tendency toward granulation tissue formation, active phagocytosis, and variable angiogenesis when evaluated at 10 days, early fibrosis when evaluated at 30 days, and established fibrosis when evaluated at 60 days. However, hADSC-treated groups showed a tendency toward cardiomyocyte regeneration and prominent angiogenesis when evaluated at 10 days and smaller infarction size when evaluated at 30 and 60 days. The present study showed a significantly decreased amount of scar tissue following myocardial infarction and enhanced regenerative capacity of myocardial cells following a single, intraventricular injection of 2 million hADSCs immediately after AMI. [ABSTRACT FROM AUTHOR]

The mechanism(s) underlying cardiac reparative effects of bone marrow-derived mesenchymal stem cells (MSC) remain highly controversial. Here we tested the hypothesis that MSCs regenerate chronically infarcted myocardium through mechanisms comprising long-term engraftment and trilineage differentiation. Twelve weeks after myocardial infarction, female swine received catheterbased transendocardial injections of either placebo (n = 4) or male allogeneic MSCs (200 million; n = 6). Animals underwent serial cardiac magnetic resonance imaging, and in vivo cell fate was determined by co-localization of Y-chromosome (Ypos) cells with markers of cardiac, vascular muscle, and endothelial lineages. MSCs engrafted in infarct and border zones and differentiated into cardiomyocytes as ascertained by co-localization with GATA-4, Nkx2.5, and α-sarcomeric actin. In addition, Ypos MSCs exhibited vascular smooth muscle and endothelial cell differentiation, contributing to large and small vessel formation. Infarct size was reduced from 19.3 ± 1.7% to 13.9 ± 2.0% (P < 0.001), and ejection fraction (EF) increased from 35.0 ± 1.7% to 41.3 ± 2.7% (P < 0.05) in MSC but not placebo pigs over 12 weeks. This was accompanied by increases in regional contractility and myocardial blood flow (MBF), particularly in the infarct border zone. Importantly, MSC engraftment correlated with functional recovery in contractility (R = 0.85, P < 0.05) and MBF (R = 0.76, P < 0.01). Together these findings demonstrate long-term MSC survival, engraftment. and trilineage differentiation following transplantation into chronically scarred myocardium. MSCs are an adult stem cell with the capacity for cardiomyogenesis and vasculogenesis which contribute, at least in part, to their ability to repair chronically scarred myocardium. [ABSTRACT FROM AUTHOR]

Abstract: Cellular cardiomyoplasty is an investigational technique that may have the potential to reduce myocardial infarct size and improve cardiac function in patients with ischemic heart disease. However, experience with this approach is still limited to a few small trials, some of which were not blinded or had other design problems. The mechanism of benefit, long-term safety, and clinical efficacy of cellular cardiomyoplasty require further study. [Copyright &y& Elsevier]

Aim: A limitation of cell therapy for heart disease is the fact that stem cells injected directly into the myocardium are capable of entering the vasculature and migrating to remote organs. We determined whether retention of mesenchymal stem cells (MSCs) in the infarcted myocardium could be improved by implanting the cells in a collagen matrix. Methods: A myocardial infarction was induced by ligation of the left anterior descending coronary artery in Fischer rats. A total of 7 days after myocardial infarction, saline (n = 12), saline plus 2 million bone marrow-derived rat MSCs labeled with isotopic colloidal nanoparticles containing europium (n = 13), collagen (n = 13) or collagen plus 2 million labeled MSCs (n = 13) were directly injected into the infarcted myocardium. Tissues from the infarcted myocardium, noninfarcted myocardium, lung, liver, spleen and kidney were sampled 4 weeks later. Distribution of grafted MSCs was quantitatively analyzed by measuring the nanoparticle radioactivity in these tissues. Cardiac function was assessed by left ventriculography. Results: There were zero nanoparticles detected in the tissues that received saline or collagen alone into the heart. Nanoparticles were detected in the heart and remote organs in the saline plus MSC group. Labeled cells (expressed as cell number/g tissue weight) were present in three out of 13 lungs (mean of 12,724 ± 7060 cells/g), four out of 13 livers (12,301 ± 5924 cells/g), 11 out of 13 spleens (57,228 ± 11,483 cells/g), zero out of 13 kidneys, 13 out of 13 infarcted myocardium (8,006,835 ± 1,846,462 cells/g) and nine out of 13 noninfarcted myocardium (167,331 ± 47,007 cells/g). However, compared with the saline plus MSC group, nanoparticles were detected to a lesser extent in remote organs in collagen plus MSC group. Nanoparticles were detected in two out of 13 lungs (4631 ± 3176 cells/g; p = NS), zero out of 13 livers (0 cells/g; p <0.05 vs saline plus MSC), four out of 13 spleens (24,060 ± 17,373 cells/g; p <0.05), zero out of 13 kidneys (p = NS) and five out of 13 noninfarcted myocardium (51,522 ± 21,548 cells/g; p <0.05). In the collagen plus MSC group, nanoparticles were detected in 12 out of 13 infarcted myocardium (4,830,050 ± 592,215 cells/g), which did not significantly differ from that in the saline plus MSC group (p = NS). Both saline plus MSCs and collagen alone improved left ventricular ejection fraction compared with saline treatment. However, collagen plus MSCs failed to improve cardiac function. Conclusions: Collagen matrix as a delivery vehicle significantly reduced the relocation of transplanted MSCs to remote organs and noninfarcted myocardium. [ABSTRACT FROM AUTHOR]

During the past several years, there have been increasing experimental and early clinical observations indicating that allogeneic, and even xenogeneic, mesenchymal stem cells (MSCs) may be useful for cellular cardiomyoplasty. Although the immune tolerance of MSCs is well established in various in vitro studies, controversies on the in vivo immune tolerance of MSCs persist. Confirmation of the feasibility of allogeneic cellular cardiomyoplasty will have vast clinical implications, since it will provide convenient “off-the-shelf” donor cells, as a vehicle for gene therapy, and avoid the need to use dysfunctional autologous MSCs in senile and/or disabled patients. [Copyright &y& Elsevier]

Background: Cellular cardiomyoplasty is emerging as a potentially novel therapeutic option for heart failure and typically involves direct intramyocardial injection of donor cells into a beating heart. Yet, limited rates of cell engraftment remain an obstacle to be overcome before cell therapy is fully recognized. Mechanical and biological mechanisms may account for observed donor cell loss. This study examines acute mechanical loss during intramyocardial injections in beating and arrested hearts. Materials and methods: A porcine cardiopulmonary bypass model was used. Animals underwent either beating (n = 5) or arrested (n = 5) intramyocardial injections into the left ventricle. Fluorescent microspheres were used in lieu of cells because they are biologically inert. Thirty minutes after delivery, animals were euthanized. Microspheres in cardiac and peripheral tissues were quantified using flow cytometry. Results: Approximately 10% of microspheres were retained within the site of injection in both groups. There was no statistical difference between microsphere retention rates in either the beating or the arrested heart group. Microspheres were found in peripheral organs, pericardial fluid, and the delivery device. Conclusions: The majority of microspheres injected intramyocardially are lost in both beating and arrested hearts. Cardiac standstill does not enhance microsphere retention. Possible mechanisms include leakage from the injection site and washout via the cardiac venous/lymphatic system. Delivery strategy will need to be modified if more cells are to be retained within the target organ. [Copyright &y& Elsevier]

Abstract: Background: Efficacy of cellular cardiomyoplasty seems to occur in a dose-related manner. However, the safety of multiple transendomyocardial transplantation procedures to administer high cell dosages has not been previously reported. The aims of this study were to assess the short- and intermediate-term results of a repeated cell administration strategy and evaluate the safety of an “off-the-shelf” allogeneic mesenchymal stem cell (MSC) source. Methods: Porcine bone marrow-derived MSCs were culture-expanded through three cycles in vitro before transplantation. Yorkshire swine weighing 30–40 kg were allocated to receive the total dose during 1 injection procedure or divided over 2 procedures separated by 14 days, as follows: (i) 400×106 allogeneic MSC (n =5), (ii) 800×106 allogeneic MSC divided in 2 doses (n =5), (iii) cryopreservant vehicle containing 10% DMSO, 5% porcine serum and 85% PlasmaLyte A, 14 days apart (n =2), or (iv) sterile saline 14 days apart (n =2). During each procedure, twenty 0.5 ml aliquots of the assigned injectant were administered using the Stiletto™ Endocardial Direct Injection Catheter System, targeting at the left ventricular anterior, septal and lateral walls under fluoroscopy. Vital signs and electrocardiograms were recorded during the procedure and at 24 h. The animals were examined daily and cardiac enzymes were measured immediately post-procedure, and on days 1, 15 and 90. Necropsy and histopathology were performed at day 90. Results: Mean transendocardial injection procedure time was 40±10 min. All ventricular target areas were accessed by the Stiletto™ system. Ventricular ectopic beats and/or non-sustained ventricular tachycardia associated with catheter contact or endomyocardial injections were observed in all cases. However, no sustained ventricular arrhythmia, anaphylaxis, or significant cardiac enzyme release was seen. One mortality resulted from air embolism during the procedure. All other swine survived from the time of recovery until planned sacrifice at day 90 and had normal physical examination findings. The 3-month histopathology showed no evidence of rejection, calcification, teratoma or myocardial infarction. Conclusion: Repeated endomyocardial transplantation of high dose, bone marrow-derived allogeneic cells appeared safe in a large animal, human surrogate model. Such cellular cardiomyoplasty strategy warrants further investigation. [Copyright &y& Elsevier]

Objective: Transplantation of autologous skeletal myoblasts (SKMB) into infarcted heart (or cellular cardiomyoplasty, CCM) augments myocardial performance in animal models of myocardial infarction. However, the effect of CCM in the setting of ventricular aneurysm has not been evaluated. This study analyzes the effects of transplanted SKMB on regional wall motion in a rabbit model of postinfarct ventricular aneurysm. We hypothesize that CCM, performed early after myocardial infarction, prevents the progression of dyskinetic wall motion. Methods: Twenty-six rabbits underwent apical left ventricular cryoinfarction and soleus muscle biopsy for in vitro isolation of skeletal myoblasts. At 2 weeks postinfarct, the presence of ventricular aneurysm was detected in 23/26 animals by sonomicrometry and micromanometry. Seventeen of 23 animals were randomized to receive either 108 autologous myoblasts (CCM) or vehicle. Regional stroke work, percent systolic shortening, and synchronicity of regional wall motion were determined prior to, and four weeks following, injection (CCM; n = 9; vehicle, n = 8). Wall motion was evaluated under baseline and stress (dobutamine, 10 (g/kg/min) conditions. Six animals did not undergo randomization, but their hearts were used to measure the size of infarction. Results: Four weeks following treatment of animals with ventricular aneurysm, systolic contractile activity was present in most animals treated with myoblasts but in none treated with vehicle (5/7 versus 0/6, respectively, P < 0.05). Dobutamine tended to accentuate the differences seen at baseline between the groups. Conclusions: This study demonstrates a high incidence of systolic contractile activity in a previously aneurysmal region of myocardium following CCM and may represent a novel therapy for the prevention and treatment of postinfarct aneurysm. [Copyright &y& Elsevier]

Abstract: We tested the hypothesis that granulocyte colony-stimulating factor (G-CSF) administration would enhance the efficacy of cellular cardiomyoplasty with embryonic stem (ES) cell-derived cardiomyocytes in infarcted myocardium. Three weeks after myocardial infarction by cryoinjury, Sprague–Dawley rats were randomized to receive either an injection of medium, ES cell-derived cardiomyocyte transplantation, G-CSF administration, or a combination of G-CSF administration and ES cell-derived cardiomyocyte transplantation. Eight weeks after treatment, the cardiac tissue formation, neovascularization, and apoptotic activity in the infarct regions were evaluated by histology and immunohistochemistry. The left ventricular (LV) dimensions and function of the treated heart were evaluated by echocardiography. Transplanted ES cell-derived cardiomyocytes survived and participated in the myocardial regeneration in the infarcted heart. A combination of G-CSF treatment and ES cell-derived cardiomyocyte transplantation significantly promoted angiogenesis and reduced the infarct area and cell apoptosis in the infarcted myocardium compared with ES cell-derived cardiomyocyte transplantation alone. The combination therapy also attenuated LV dilation, as compared with ES cell-derived cardiomyocyte transplantation alone. G-CSF treatment can enhance the efficacy of cellular cardiomyoplasty by ES cell-derived cardiomyocyte transplantation to treat myocardial infarction. [Copyright &y& Elsevier]

Cell transplantation holds promise as a potential treatment for cardiac dysfunction. Our group has isolated populations of murine skeletal muscle-derived stem cells (MDSCs) that exhibit stem cell-like properties. Here, we investigated the fate of MDSCs after transplantation into the hearts of dystrophin-deficient mdx mice, which model Duchenne muscular dystrophy (DMD). Transplanted MDSCs generated large grafts consisting primarily of numerous dystrophin-positive myocytes and, to a lesser degree, dystrophin-negative nonmyocytes that expressed an endothelial phenotype. Most of the dystrophin-positive myocytes expressed a skeletal muscle phenotype and did not express a cardiac phenotype. However, some donor myocytes, located at the graft–host myocardium border, were observed to express cardiac-specific markers. More than half of these donor cells that exhibited a cardiac phenotype still maintained a skeletal muscle phenotype, demonstrating a hybrid state. Sex-mismatched donors and hosts revealed that many donor-derived cells that acquired a cardiac phenotype did so through fusion with host cardiomyocytes. Connexin43 gap junctions were not expressed by donor-derived myocytes in the graft. Scar tissue formation in the border region may inhibit the fusion and gap junction connections between donor and host cells. This study demonstrates that MDSC transplantation warrants further investigation as a potential therapy for cardiac dysfunction in DMD.Gene Therapy (2005) 12, 1264–1274. doi:10.1038/sj.gt.3302521; published online 21 April 2005 [ABSTRACT FROM AUTHOR]

Abstract: Objective: Recent progress in the field of cellular cardiomyoplasty has opened new prospects for the treatment of ischemic heart disease and currently moves from bench to bedside. The aim of the present study was to develop a novel cell delivery technique, reducing target tissue damage and improving cell dispersion and engraftment. Methods: In 30 male Fischer F344 rats an infarction of the left ventricle was generated by ligation of the left anterior descendent artery. Seven days after infarction, either 15 microdepots of 10μl myoblast cell suspension (microdepot group) or culture medium (control group) were injected into the infarcted region using an automatic pressure injection device, or three depots of 50μl myoblast cell suspension (macrodepot group) were injected using the standard surgical technique. Echocardiography was performed in all rats before and 6 weeks after cell injection. In all groups the perioperative mortality was below 20%. Six weeks after cell transplantation, a significant improvement of ejection fraction was seen in both myoblast treated groups compared to controls (macrodepot, microdepot, control; 53.7±11.9, 70.7±2.0, 39.1±6.4; P=0.026, P&#60;0.001). The microdepot group showed a more decent improvement than the macrodepot group (70.7±2.0 vs. 53.7±11.9, P=0.013). In both treated groups, grafted myoblasts differentiated into multinucleated myotubes within host myocardium, however, the engraftment pattern was different and angiogenesis was enhanced in the microdepot group. Conclusions: Intramyocardial multisite pressure injection allows the safe and reliable transplantation of several myoblast microdepots into an infarcted myocardium and improves the efficacy of myoblast transplantation compared to the standard technique. [Copyright &y& Elsevier]

Background: Contraction of transplanted myoblasts and their effects on function and remodeling after myocardial infarction remain controversial. Aim: We used magnetic resonance imaging (MRI) to study wall thickening and left ventricular (LV) function and geometry after myoblast transplantation. Methods and results: Three weeks after cryo-infarction rabbits were randomized to receive an injection of ∼2×108 myoblasts (n=8) or medium (n=9) into the scar. Cine MRI and contrast enhanced (ce) MRI images were acquired before injection (baseline) and 4 weeks later (endpoint). Regional wall thickening was measured at the site of transmural hyperenhancement. In the control group, regional wall thickening decreased to -15.3±8.6% at baseline, which further decreased to -18.3±5.7% at endpoint. Further, end-diastolic volume increased from 3.96±0.27 to 5.00±0.46 ml and end-systolic volume from 2.23±0.19 to 2.96±0.30 ml (both P<0.05 vs. baseline), which was accompanied by increased LV wall volumes (P<0.05 vs. baseline). In contrast, myoblast transplantation increased regional wall thickening from -11.9±15.9% at baseline to 26.9±17.0% (P<0.05 vs. control), which resulted in significantly improved two-dimensional ejection fractions at the infarct level and prevented the increase in end-diastolic and end-systolic volumes and wall volume. Conclusion: Intracardiac myoblast transplantation after myocardial infarction improves regional wall thickening and prevents progressive left ventricular remodeling. [Copyright &y& Elsevier]

Some recent studies have demonstrated that epicardial injection of autologous myoblasts, obtained from satellite cells of skeletal muscle, in association to coronary artery bypass graft surgery (CABG) in patients with decreased left ventricular function secondary to ischaemic disease could be of some utility to get a better recovery of ventricular function due to the ability of these cells to grow and generate new muscle fibers over the previous fibrotic scar. The aims are the setting up of a process for the collection of the cellular cardiomyoplasty in samples of multiorganic donations and to carry out this technique in the same surgical moment as the revascularisation is performed in two patients. For this purpose we obtained muscle through biopsy of 15 human multiorgan donors and of two patients. Separation of fatty tissue, minced, and further digestion with collagenase type I (1.5 mgr/ml/2 gr by weight) and trypsin 1 ×. Filtration of the cellular suspension, centrifugation and sowing of this suspension in culture medium, with 20% of human serum. Culture for three weeks until obtainment of between 200–300 million cells. Inmunohistochemistry and flow cytometry for the identification of the myoblasts was carried out. The results were obtained through flow cytometry, using CD56 as an indicator of the presence of myoblasts, between 70 and 80% of these types of cells were obtained after three weeks of culture. By inmunohistochemistry analyses, different markers were analyzed: desmin and myogenin. The results indicated the presence of a great number of positive cells with these markers, possibly myoblasts. Skeletal myoblast implant was not associated with adverse effects. The culture of autologous myoblasts is a rapid and simple technique where after three weeks of culture a great number of cells for implantation are obtained. In patients with old myocardial infarction, treatment with skeletal myoblast in conjunction with coronary artery bypass is safe and feasible. and it is easy to obtain myoblasts from muscle tissue for transplant into patients. [ABSTRACT FROM AUTHOR]

An adult heart injured by an ischemic episode has a limited capacity to regenerate. We administered three types of adult guinea pig cells [cardiomyocytes (CMs), cardiac fibroblasts (CFs), and skeletal myoblasts (Mbs)] to compare their suitability for repair of acute myocardial infarction. We used confocal fluorescent microscopy and a variety of specific immunomarkers and echocardiography to provide anatomic evidence for the viability of such cells and their possible functional beneficial effects. All cells were transfected with adenovims-containing β-galactosidase gene so that migration from the injection sites could be traced. Both freshly isolated CMs as well as CFs were found concentrated in the infarcted zone; these cells survived for at least 2 wk posttransplantation. Transplanted CMs were regularly striated and grew long projections that could form gap junctions with native CMs, which was evidenced by connexin43 labeling. In addition, CM transplantation resulted in increased angiogenesis in the infarcted areas. In contrast, transplanted CFs did not appear to make any gap junctional contacts with native CMs nor did they enhance local angiogenesis. Mbs cultured for 7 days and transfected Mbs were identified 7 days posttransplantation in the infarcted area. During that time and thereafter, Mbs proliferated and differentiated into myotubes that formed new, regularly striated myofibers that occupied most (50-70%) of the infarcted area by 2-3 wk. These newly formed myofibers maintained their Mb skeletal muscle origin as evidenced by their capacity to express myogenin and fast skeletal myosin. This skeletal phenotype appeared to downregulate with time, and Mbs partially transdifferentiated into a cardiac phenotype as indicated by labeling for cardiac-specific troponin T and cardiac myosin heavy chain. By the third week posttransplantation, new myofibers formed apparent contacts with the native CMs via putative gap junctions that expressed connexin43. Myocardial performance of animals that were successfully transplanted with Mbs was improved. [ABSTRACT FROM AUTHOR]

Objectives: To prove whether intramyocardial transplantation of combined skeletal myoblasts (SM) and mononuclear bone marrow stem cells is superior to the isolated transplantation of these cell types after myocardial infarction in rats. Methods: In 67 male Fischer rats myocardial infarction was induced by direct ligature of the LAD. Seven days postinfarction baseline echocardiography and intramyocardial cell transplantation were performed. Via lateral thoracotomy 200 μl containing either 107 SMs or 107 bone marrow-derived mononuclear cells (BM-MNC) or a combination of 5×106 of both cell types (MB) were injected in 10–15 sites in and around the infarct zone. In controls (C) 200 μl of cell-free medium were injected in the same manner. Before injection both cell types were stained using a fluorescent cell linker kit (PKH, Sigma). In addition, SMs were transfected with green fluorescent protein. Nine weeks postinfarction follow-up echocardiography was performed and animals were sacrificed for further analysis. Results: At baseline echocardiography there was no difference in left ventricular ejection fraction (LVEF; C, SM, BM-MNC, MB: 60.1±3.2, 53.3±10.2, 53.1±8.7, 49±9.0%) and left ventricular end diastolic diameter (LVEDD; C, SM, BM-MNC, MB: 6.5±0.8, 5.17±0.8, 5.77±1.4, 6.25±0.8 mm) between the different therapeutic groups. Eight weeks after cell transplantation LVEDD was significantly increased in all animals except those that received a combination of myoblasts and bone marrow stem cells (MB; C, SM, BM-MNC, MB: 7.7±0.6 mm, P=0.001; 7.7±1.5 mm, P<0.001; 7.7±1.1 mm, P=0.005; 6.6±1.7 mm, P=0.397). At the same time LVEF decreased significantly in the control group (C), stayed unchanged in animals that received bone marrow stem cells (BM-MNC) and increased in animals that received myoblasts (SM) and a combination of both cell types (MB; C, SM, BM-MNC, MB: 45.3±7.0%, P=0.05; 63.9±15.4%, P=0.044; 54.3±6.3%, P=0.607; 63.0±11.5%, P=0.039). Conclusions: The present data show that the concept of combining SMs with bone marrow-derived stem cells may be of clinical relevance by merging the beneficial effects of each cell line and potentially reducing the required cell quantity. Further studies are required to identify the exact mechanisms underlying this synergy and to allow full exploitation of its therapeutic potential. [Copyright &y& Elsevier]

Autologous skeletal myoblast transplantation may be used to ameliorate the healing process following myocardium infarct and, hopefully, cardiomyopathies. Despite successful animal experimentation, several issues need to be addressed in clinical settings, i.e., the impact of the delivery route, the extent of short- and long-term survival, and differentiation of the injected skeletal myoblasts. The authors offer some new hypotheses resulting from basic research, i.e., where and when to inject the myogenic cells, whatever their source, how to decrease new myofiber atrophy and improve their regeneration. Although these new hypotheses still need to be tested in humans, they may be decisive for future experimental studies and will lead to making endovascular cell implantation a more effective way to treat ischemic heart disease and failure. [Copyright &y& Elsevier]

Background: Cellular cardiomyoplasty is evolving as a new strategy to treat cardiac diseases. A prerequisite is a reliable source of pure cardiomyocytes, which could also help in the exploitation of recent advances in genomics and drug screening. Our goal was to establish a robust lab-scale process for the generation of embryonic stem (ES)-cell-derived cardiomyocytes in suspension. Methods: A J1 ES cell clone carrying a construct consisting of the α-cardiac myosin heavy chain (αMHC) promoter driving the neomycin resistance gene was used for antibiotic-driven cardiomyocyte enrichment. Rotating suspension culture was established to initiate embryoid body (EB) formation. To track growth and differentiation kinetics, cell count and flow cytometry for SSEA-1, E-cadherin (stem-cell marker) and sarcomeric myosin (cardiomyocytes marker) was performed. Oct4 expression was measured via real time (RT)-PCR. Results: Cultures comprising 2.5 - 8 × 10[sup6] differentiating ES cells/mL were obtained after 9 days in rotating suspension. Upon G418 addition, vigorous contracting spheres, termed cardiac bodies (CB), developed. These cultures consisted of about 2.1 × 10[sup5] enriched cardiomyocytes/ mL after 6 - 10 days of selection. Suspensions comprising 90 - 95% viable single cells were generated using an improved dissociation method. Seeding of cardiomyocytes with 7 &times 10[sup4] cell/cm[sup2] resulted in homogeneous monolayer of synchronously contracting cells. Myocyte specific immunohistochemistry indicated purity of > 99%. Discussion: We have established a reliable lab-scale protocol to generate cultures highly enriched cardiomyocytes in suspension. This will facilitate development of larger-scale processes for stem-cell based cardiomyocyte supply. An improved method is provided to derive vital suspensions cardiomyocytes, which could be utilized for transplantation as well as for drug screening purposes. [ABSTRACT FROM AUTHOR]

Background: Cellular cardiomyoplasty is the method of transplanting myogenic cells into injured myocardium to restore the lost heart muscle cells and to improve ventricular function. Method: Three patients, all with a history of coronary heart disease, underwent coronary artery bypass grafting and implantation of autologous satellite cells. A muscle biopsy of 2–4 g from the right vastus lateralis muscle was obtained for satellite cell (myogenic stem cell from skeletal muscle) isolation and proliferation before implanted into the donor''s heart. The cells were suspended in serum-free medium and injected into 30–40 sites at and around the ischemic areas just before reversing the hypothermic cardioplegia to eliminate arrhythmia and to improve retention. After recovery, each patient was maintained at the intensive care unit for 3–4 days with ECG monitoring before transferring to the patient floor. Results: All patients survived the procedure with an uneventful recovery and were discharged from the hospital. At 3–4 months follow-up examination, increased left ventricular ejection fraction of 11% (35–46%), 5.4% (40–45.4%) and 1% (40–41%) and decreased left ventricular diastolic diameter of 4, 2 and 9 mm were observed for the patients, respectively. Arrhythmia was not detected during the follow-up evaluation by ECG. Improved perfusion (99mTC-MIBI) and increased metabolic activity (18F-deoxyglucose) were found at the sites of satellite cell implantation. Significant increase of wall thickness and movement at the areas of cell injection was also observed using 2D-echo. Conclusion: Cellular cardiomyoplasty using autologous satellite cells is a safe procedure with encouraging beneficial outcomes in patients. [Copyright &y& Elsevier]

Background: A 73-year-old man with a history of myocardial infarction and hypertension for 5 years suffered heart failure (NYHA III–IV). Methods: 2D echo indicated hypokinesia at septal, left ventricular anterior wall and apical regions. Coronary angiograms demonstrated 60% stenosis in distal left main and 99% stenosis in proximal and distal left anterior descending coronary arteries (LAD). Both proximal artery and middle left circumflex coronary artery (LC) had 90% stenosis, and diffuse stenosis of right coronary artery (RC) was found. Myocardial perfusion imaging using 99mTc-MIBI indicated defective perfusion of left ventricular apex, anterior wall and septal region and severe reduced perfusion of posterior inferior wall. Myocardial metabolic activities (18F-deoxyglucose) also showed comparable reductions. After exposing the heart, LAD, LC, and RC were all completely occluded and bypass procedure could not be completed. Autologous satellite cells were implanted without any complication and the patient had an uneventful recovery. Results: During the first 2 months, he remained in heart failure, and by the third month, he gradually improved and reached NYHA II. At fifth month after the procedure, significant increased ejection fraction (37.1–48.6%) and wall movement with modest reduction of left ventricular systolic diameter (48–45 mm) were observed. Imaging with 18F-deoxyglucose showed dramatic improvement in myocardial metabolic activity with similar improvement in myocardial perfusion (99mTc-MIBI). Conclusion: This is the first successful case of cellular cardiomyoplasty without any conjunctional procedure for patient with severe coronary heart disease and heart failure. [Copyright &y& Elsevier]

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