Cardiac specification of embryonic stem cells

Over the past decade, cell transplantation has been recognized as a mean of repairing infarcted myocardium. Both adult stem cells and differentiated cells have yielded encouraging results with regard to engraftment into postinfarction scars. However, these cells now feature serious restrictions. Asan alternative, embryonic stem (ES) cells are particularly attractive, because of their plasticity and the subsequent possibility to drive them towards a cardiomyogenic phenotype after exposure to appropriate growth factors. An additional theoretical advantage of ES cells is their expected immune privilege. In this article, we summarize the findings obtained in cell therapy using ES cells and discuss the molecular mechanisms of cardiac specification of the cells.     

Molecular imaging,biodistribution and efficacy of mesenchymal bone marrow cell therapy in a mouse model of Chagas disease

Chagasic cardiomyopathy, resulting from infection with the parasite Trypanosoma cruzi, was discovered more than a century ago and remains an incurable disease. Due to the unique properties of mesenchymal stem cells (MSC) we hypothesized that these cells could have therapeutic potential for chagasic cardiomyopathy. Recently, our group pioneered use of nanoparticle-labeled MSC to correlate migration with its effect in an acute Chagas disease model. We expanded our investigation into a chronic model and performed more comprehensive assays. Infected mice were treated with nanoparticle-labeled MSC and their migration was correlated with alterations in heart morphology, metalloproteinase activity, and expression of several proteins. The vast majority of labeled MSC migrated to liver, lungs and spleen whereas a small number of cells migrated to chagasic hearts. Magnetic resonance imaging demonstrated that MSC therapy reduced heart dilatation. Additionally metalloproteinase activity was higher in heart and other organs of infected mice. Protein expression analyses revealed that connexin 43, laminin γ1, IL-10 and INF-γ were affected by the disease and recovered after cell therapy. Interestingly, MSC therapy led to upregulation of SDF-1 and c-kit in the hearts. The beneficial effect of MSC therapy in Chagas disease is likely due to an indirect action of the cells of the heart, rather than the incorporation of large numbers of stem cells into working myocardium.     

Cardiac stem cell therapy to modulate inflammation upon myocardial infarction

Background

After myocardial infarction (MI) a local inflammatory reaction clears the damaged myocardium from dead cells and matrix debris at the onset of scar formation. The intensity and duration of this inflammatory reaction are intimately linked to post-infarct remodeling and cardiac dysfunction. Strikingly, treatment with standard anti-inflammatory drugs worsens clinical outcome, suggesting a dual role of inflammation in the cardiac response to injury. Cardiac stem cell therapy with different stem or progenitor cells, e.g. mesenchymal stem cells (MSC), was recently found to have beneficial effects, mostly related to paracrine actions. One of the suggested paracrine effects of cell therapy is modulation of the immune system.

Scope of review

MSC are reported to interact with several cells of the immune system and could therefore be an excellent means to reduce detrimental inflammatory reactions and promote the switch to the healing phase upon cardiac injury. This review focuses on the potential use of MSC therapy for post-MI inflammation. To understand the effects MSC might have on the post-MI heart the cellular and molecular changes in the myocardium after MI need to be understood.

Major conclusions

By studying the general pathways involved in immunomodulation, and examining the interactions with cell types important for post-MI inflammation, it becomes clear that MSC treatment might provide a new therapeutic opportunity to improve cardiac outcome after acute injury.

General significance

Using stem cells to target the post-MI inflammation is a novel therapy which could have considerable clinical implications. This article is part of a Special Issue entitled Biochemistry of Stem Cells.     

Pretreatment of rat bone marrow mesenchymal stem cells with a combination of hypergravity and 5-azacytidine enhances therapeutic efficacy for myocardial infarction

Background and Purpose: The in vivo cardiac differentiation and functional effects of unmodified adult bone marrow mesenchymal stem cells (BMSCs) after myocardial infarction (MI) is controversial. Our previous results suggested that hypergravity promoted the cardiomyogenic differentiation of BMSCs, and thus we postulated that ex vivo pretreatment of BMSCs using hypergravity and 5‐azacytidine (5‐Aza) would lead to cardiomyogenic differentiation and result in superior biological and functional effects on cardiac regeneration of infarcted myocardium. Methods: We used a rat MI model generated by ligation of the coronary artery. Homogeneous rat BMSCs were isolated, culture expanded, and differentiated into a cardiac lineage by adding hypergravity (2G) for 3 days and 5‐Aza (50 lmol/L, 24 h). Rats underwent BMSCs (labeled with DAPI) injection after the infarction and were randomized into five groups. Group A rats received the control medium, Group B rats received unmodified BMSCs, Group C rats received BMSCs treated with hypergravity, Group D rats received BMSCs treated with 5‐Aza, and Group E rats received BMSCs treated with 5‐Aza and hypergravity (n = 6). Results: After hypergravity and 5‐Aza treatment, BMSCs showed positive for the early muscle and cardiac markers GATA‐4, MEF‐2, and Nkx2‐5 with RT‐PCR. We also found that hypergravity could enhance the activities of MEF‐2 via promoting the nuclear export of HDAC5. The frozen section showed that the implanted BMSCs labeled with DAPI survived and angiogenesis was identified at the implantation site. In Groups B, C, D, and E rats, pre‐treated BMSCs colocalized with α‐actinin, and Group E rats showed a significantly larger increase in left ventricular function. Conclusions: The biological ex vivo cardiomyogenic differentiation of adult BMSCs with hypergravity and 5‐Aza prior to their transplantation is feasible and appears to improve their in vivo cardiac differentiation as well as the functional recovery in a rat model of the infarcted myocardium. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011     

Differentiation of human adipose‐derived stem cells into beating cardiomyocytes

Human adipose‐derived stem cells (ASCs) may differentiate into cardiomyocytes and this provides a source of donor cells for tissue engineering. In this study, we evaluated cardiomyogenic differentiation protocols using a DNA demethylating agent 5‐azacytidine (5‐aza), a modified cardiomyogenic medium (MCM), a histone deacetylase inhibitor trichostatin A (TSA) and co‐culture with neonatal rat cardiomyocytes. 5‐aza treatment reduced both cardiac actin and TropT mRNA expression. Incubation in MCM only slightly increased gene expression (1.5‐ to 1.9‐fold) and the number of cells co‐expressing nkx2.5/sarcomeric α‐actin (27.2%versus 0.2% in control). TSA treatment increased cardiac actin mRNA expression 11‐fold after 1 week, which could be sustained for 2 weeks by culturing cells in cardiomyocyte culture medium. TSA‐treated cells also stained positively for cardiac myosin heavy chain, α‐actin, TropI and connexin43; however, none of these treatments produced beating cells. ASCs in non‐contact co‐culture showed no cardiac differentiation; however, ASCs co‐cultured in direct contact co‐culture exhibited a time‐dependent increase in cardiac actin mRNA expression (up to 33‐fold) between days 3 and 14. Immunocytochemistry revealed co‐expression of GATA4 and Nkx2.5, α‐actin, TropI and cardiac myosin heavy chain in CM‐DiI labelled ASCs. Most importantly, many of these cells showed spontaneous contractions accompanied by calcium transients in culture. Human ASC (hASC) showed synchronous Ca²⁺ transient and contraction synchronous with surrounding rat cardiomyocytes (106 beats/min.). Gap junctions also formed between them as observed by dye transfer. In conclusion, cell‐to‐cell interaction was identified as a key inducer for cardiomyogenic differentiation of hASCs. This method was optimized by co‐culture with contracting cardiomyocytes and provides a potential cardiac differentiation system to progress applications for cardiac cell therapy or tissue engineering.     

Prolyl hydroxylase inhibitor dimethyloxalylglycine enhances mesenchymal stem cell survival

Mesenchymal stem cell (MSC) transplantation is a promising approach in the therapy of ischemic heart or CNS diseases; however, the poor viability of MSCs after transplantation critically limits the efficacy of this new strategy. Prolyl hydroxylase inhibition followed by HIF‐1α up‐regulation participates in the regulation of apoptosis and cell survival, which have been shown in cancer cells and neurons. The role of prolyl hydroxylase inhibition by dimethyloxalylglycine (DMOG) in regulation of cell survival has not been investigated in MSCs. In the present investigation with MSCs, apoptosis and cell death induced by serum deprivation were assessed by caspase‐3 activation and trypan blue staining, respectively. The mitochondrial apoptotic pathway and PI3K/Akt cell survival pathway were evaluated. DMOG significantly attenuated apoptosis and cell death of MSCs, stabilized HIF‐1α and induced downstream glucose transport 1 (Glut‐1) synthesis. DMOG treatment reduced mitochondrial cytochrome c release, nuclear translocation of apoptosis inducing factor (AIF), and promoted Akt phosphorylation. A specific PI3K inhibitor, wortmannin, blocked Akt phosphorylation and abrogated the beneficial effect of DMOG. These data suggest that the DMOG protection of MSCs may provide a novel approach to promote cell survival during cell stress. J. Cell. Biochem. 106: 903–911, 2009. © 2009 Wiley‐Liss, Inc.     

Interleukin‐6 downregulation with mesenchymal stem cell differentiation results in loss of immunoprivilege

Allogeneic mesenchymal stem cell (MSC) transplantation improves cardiac function, but cellular differentiation results in loss of immunoprivilege and rejection. To explore the mechanism involved in this immune rejection, we investigated the influence of interleukin‐6 (IL‐6), a factor secreted by MSCs, on immune privilege after myogenic, endothelial and smooth muscle cell differentiation induced by 5‐azacytidine, VEGF, and transforming growth factor‐β (TGF‐β), respectively. Both RT‐PCR and ELISA showed that myogenic differentiation of MSCs was associated with significant downregulation of IL‐6 expression (P < 0.01), which was also observed following endothelial (P < 0.01) and smooth muscle cell differentiation (P < 0.05), indicating that IL‐6 downregulation was dependent on differentiation but not cell phenotype. Flow cytometry demonstrated that IL‐6 downregulation as a result of myogenic differentiation was associated with increased leucocyte‐mediated cell death in an allogeneic leucocyte co‐culture study (P < 0.01). The allogeneic reactivity associated with IL‐6 downregulation was also observed following MSC differentiation to endothelial and smooth muscle cells (P < 0.01), demonstrating that leucocyte‐mediated cytotoxicity was also dependent on differentiation but not cell phenotype. Restoration of IL‐6 partially rescued the differentiated cells from leucocyte‐mediated cell death. These findings suggest that rejection of allogeneic MSCs after implantation may be because of a reduction in cellular IL‐6 levels, and restoration of IL‐6 may be a new target to retain MSC immunoprivilege.     

Comparative osteogenic transcription profiling of various fetal and adult mesenchymal stem cell sources

Human mesenchymal stem cells (MSC) from adult and fetal tissues are promising candidates for cell therapy but there is a need to identify the optimal source for bone regeneration. We have previously characterized MSC populations in first trimester fetal blood, liver, and bone marrow and we now evaluate their osteogenic differentiation potential in comparison to adult bone marrow MSC. Using quantitative real-time RT-PCR, we demonstrated that 16 osteogenic-specific genes (OC, ON, BSP, OP, Col1, PCE, Met2A, OPG, PHOS1, SORT, ALP, BMP2, CBFA1, OSX, NOG, IGFII) were expressed in both fetal and adult MSC under basal conditions and were up-regulated under osteogenic conditions both in vivo and during an in vitro 21-day time-course. However, under basal conditions, fetal MSC had higher levels of osteogenic gene expression than adult MSC. Upon osteogenic differentiation, fetal MSC produced more calcium in vitro and reached higher levels of osteogenic gene up-regulation in vivo and in vitro. Second, we observed a hierarchy within fetal samples, with fetal bone marrow MSC having greater osteogenic potential than fetal blood MSC, which in turn had greater osteogenic potential than fetal liver MSC. Finally, we found that the level of gene expression under basal conditions was positively correlated with both calcium secretion and gene expression after 21 days in osteogenic conditions. Our findings suggest that stem cell therapy for bone dysplasias such as osteogenesis imperfecta may benefit from preferentially using first trimester fetal blood or bone marrow MSC over fetal liver or adult bone marrow MSC.     

Local activation of cardiac stem cells for post‐myocardial infarction cardiac repair

The prognosis of patients with myocardial infarction (MI) and resultant chronic heart failure remains extremely poor despite continuous advancements in optimal medical therapy and interventional procedures. Animal experiments and clinical trials using adult stem cell therapy following MI have shown a global improvement of myocardial function. The emergence of stem cell transplantation approaches has recently represented promising alternatives to stimulate myocardial regeneration. Regarding their tissue‐specific properties, cardiac stem cells (CSCs) residing within the heart have advantages over other stem cell types to be the best cell source for cell transplantation. However, time‐consuming and costly procedures to expanse cells prior to cell transplantation and the reliability of cell culture and expansion may both be major obstacles in the clinical application of CSC‐based transplantation therapy after MI. The recognition that the adult heart possesses endogenous CSCs that can regenerate cardiomyocytes and vascular cells has raised the unique therapeutic strategy to reconstitute dead myocardium via activating these cells post‐MI. Several strategies, such as growth factors, mircoRNAs and drugs, may be implemented to potentiate endogenous CSCs to repair infarcted heart without cell transplantation. Most molecular and cellular mechanism involved in the process of CSC‐based endogenous regeneration after MI is far from understanding. This article reviews current knowledge opening up the possibilities of cardiac repair through CSCs activation in situ in the setting of MI.     

iPSC‐derived mesenchymal stem cells exert SCF‐dependent recovery of cigarette smoke‐induced apoptosis/proliferation imbalance in airway cells

Mesenchymal stem cells (MSCs) have emerged as a potential cell‐based therapy for pulmonary emphysema in animal models. Our previous study demonstrated that human induced pluripotent stem cell–derived MSCs (iPSC‐MSCs) were superior over bone marrow–derived MSCs (BM‐MSCs) in attenuating cigarette smoke (CS)‐induced airspace enlargement possibly through mitochondrial transfer. This study further investigated the effects of iPSC‐MSCs on inflammation, apoptosis, and proliferation in a CS‐exposed rat model and examined the effects of the secreted paracrine factor from MSCs as another possible mechanism in an in vitro model of bronchial epithelial cells. Rats were exposed to 4% CS for 1 hr daily for 56 days. At days 29 and 43, human iPSC‐MSCs or BM‐MSCs were administered intravenously. We observed significant attenuation of CS‐induced elevation of circulating 8‐isoprostane and cytokine‐induced neutrophil chemoattractant‐1 after iPSC‐MSC treatment. In line, a superior capacity of iPSC‐MSCs was also observed in ameliorating CS‐induced infiltration of macrophages and neutrophils and apoptosis/proliferation imbalance in lung sections over BM‐MSCs. In support, the conditioned medium (CdM) from iPSC‐MSCs ameliorated CS medium‐induced apoptosis/proliferation imbalance of bronchial epithelial cells in vitro. Conditioned medium from iPSC‐MSCs contained higher level of stem cell factor (SCF) than that from BM‐MSCs. Deprivation of SCF from iPSC‐MSC‐derived CdM led to a reduction in anti‐apoptotic and pro‐proliferative capacity. Taken together, our data suggest that iPSC‐MSCs may possess anti‐apoptotic/pro‐proliferative capacity in the in vivo and in vitro models of CS‐induced airway cell injury partly through paracrine secretion of SCF.     

Enhanced infarct myocardium repair mediated by thermosensitive copolymer hydrogel-based stem cell transplantation

Mesenchymal stem cell (MSC) transplantation by intramyocardial injection has been proposed as a promising therapy strategy for cardiac repair after myocardium infarction. However, low retention and survival of grafted MSCs hinder its further application. In this study, copolymer with N-isopropylacrylamide/acrylic acid/2-hydroxylethyl methacrylate-poly(ɛ-caprolactone) ratio of 88:9.6:2.4 was bioconjugated with type I collagen to construct a novel injectable thermosensitive hydrogel. The injectable and biocompatible hydrogel-mediated MSC transplantation could enhance the grafted cell survival in the myocardium, which contributed to the increased neovascularization, decreased interstitial fibrosis, and ultimately improved heart function to a significantly greater degree than regular MSC transplantation. We suggest that this novel hydrogel has the potential for future stem cell transplantation.     

PKC-delta induces cardiomyogenic gene expression in human adipose-derived stem cells

Stromal cells from fat tissues exhibit properties of mesenchymal stem cells from other sources with the ability to differentiate towards multiple cell types. However, effective differentiation of these mesenchymal cells, called adipose-derived stem cells (ADSCs), towards cardiomyogenic lineage has been limited to a small number of isolated clones in an extended culture. Previously, we reported that treatment with phorbol ester induces the expression of several cardiomyogenic genes in the absence of serum. This study was performed to identify the roles of PKC isoforms in cardiomyogenic gene expression of ADSCs. Treatment with 10 nM phorbol myristate acetate (PMA) for 24 h caused sustained increases in mRNA levels for various cardiomyogenic genes, such as Mef2C, cardiac actin and troponin, for at least 6 days following the drug removal. The use of various inhibitors specific for PKC isoforms demonstrated that the novel PKC-theta/delta isoforms mediate the PMA effects. RT-PCR revealed that ADSCs express significant mRNA for PKC-delta, but not theta isoform. Overexpression of cDNA for PKC-delta resulted in marked increases in cardiac mRNA expression. These results indicate that activation of PKC-delta induces the expression of multiple cardiomyogenic genes in ADSCs.     

Preconditioning influences mesenchymal stem cell properties in vitro and in vivo

Various diseases and toxic factors easily impair cellular and organic functions in mammals. Organ transplantation is used to rescue organ function, but is limited by scarce resources. Mesenchymal stem cell (MSC)‐based therapy carries promising potential in regenerative medicine because of the self‐renewal and multilineage potency of MSCs; however, MSCs may lose biological functions after isolation and cultivation for a long time in vitro. Moreover, after they are injected in vivo and migrate into the damaged tissues or organs, they encounter a harsh environment coupled with death signals due to the inadequate tensegrity structure between the cells and matrix. Preconditioning, genetic modification and optimization of MSC culture conditions are key strategies to improve MSC functions in vitro and in vivo, and all of these procedures will contribute to improving MSC transplantation efficacy in tissue engineering and regenerative medicine. Preconditioning with various physical, chemical and biological factors is possible to preserve the stemness of MSCs for further application in studies and clinical tests. In this review, we mainly focus on preconditioning and the corresponding mechanisms for improving MSC activities in vitro and in vivo; we provide a glimpse into the promotion of MSC‐based cell therapy development for regenerative medicine. As a promising consequence, MSC transplantation can be applied for the treatment of some terminal diseases and can prolong the survival time of patients in the near future.     

WJSC 6th Anniversary Special Issues（2）:Mesenchymal stem cells Neurotrauma and mesenchymal stem cells treatment:From experimental studies to clinical trials

Mesenchymal stem cell(MSC)therapy has attracted the attention of scientists and clinicians around the world.Basic and pre-clinical experimental studies have highlighted the positive effects of MSC treatment after spinal cord and peripheral nerve injury.These effects are believed to be due to their ability to differentiate into other cell lineages,modulate inflammatory and immunomodulatory responses,reduce cell apoptosis,secrete several neurotrophic factors and respond to tissue injury,among others.There are many pre-clinical studies on MSC treatment for spinal cord injury(SCI)and peripheral nerve injuries.However,the same is not true for clinical trials,particularly those concerned with nerve trauma,indicating the necessity of more well-constructed studies showing the benefits that cell therapy can provide for individuals suffering the consequences of nerve lesions.As for clinical trials for SCI treatment the results obtained so far are not as beneficial as those described in experimental studies.For these reasons basic and pre-clinical studies dealing with MSC therapy should emphasize the standardization of protocols that could be translated to the clinical set with consistent and positive outcomes.This review is based on pre-clinical studies and clinical trials available in the literature from 2010 until now.At the time of writing this article there were 43 and 36 pre-clinical and 19 and 1 clinical trials on injured spinal cord and peripheral nerves,respectively.     

Transendocardial cell injection is not superior to intracoronary infusion in a porcine model of ischaemic cardiomyopathy: a study on delivery efficiency

Stem cell therapy is a new strategy for chronic ischaemic heart disease in patients. However, no consensus exists on the most optimal delivery strategy. This randomized study was designed to assess cell delivery efficiency of three clinically relevant strategies: intracoronary (IC) and transendocardial (TE) using electromechanical mapping guidance (NOGA) compared to surgical delivery in a chronic pig model of ischaemic cardiomyopathy. Twenty‐four animals underwent delivery of 10⁷ autologous Indium‐oxine‐labelled bone marrow‐derived mesenchymal stem cells (MSC) 4 weeks after infarction and were randomized to one of three groups (n = 8 each group): IC, TE or surgical delivery (reference group). Primary endpoint was defined as percentage (%) of injected dose per organ and assessed by in vivo gamma‐emission counting. In addition, troponin and coronary flow were assessed before and after MSC injection. Blinded endpoint analysis showed no significant difference in efficiency after surgical (16 ± 4%), IC (11 ± 1%) and TE (11 ± 3%) (P = 0.52) injections. IC showed less variability in efficiency compared with TE and surgical injection. Overall, TE injection showed less distribution of MSC to visceral organs compared with other modalities. Troponin rise and IC flow did not differ between the percutaneous groups. This randomized study showed no significant difference in cell delivery efficiency to the myocardium in a clinically relevant ischaemic large animal model between IC and TE delivery. In addition, no differences in safety profile were observed. These results are important in view of the choice of percutaneous cell delivery modality in future clinical stem cell trials.     

Survival and function of mesenchymal stem cells (MSCs) depend on glucose to overcome exposure to long‐term,severe and continuous hypoxia

Use of mesenchymal stem cells (MSCs) has emerged as a potential new treatment for various diseases but has generated marginally successful results. A consistent finding of most studies is massive death of transplanted cells. The present study examined the respective roles of glucose and continuous severe hypoxia on MSC viability and function with respect to bone tissue engineering. We hereby demonstrate for the first time that MSCs survive exposure to long‐term (12 days), severe (pO₂ < 1.5 mmHg) hypoxia, provided glucose is available. To this end, an in vitro model that mimics the hypoxic environment and cell‐driven metabolic changes encountered by grafted sheep cells was established. In this model, the hallmarks of hypoxia (low pO₂, hypoxia inducible factor‐1α expression and anaerobic metabolism) were present. When conditions switched from hypoxic (low pO₂) to ischemic (low pO₂ and glucose depletion), MSCs exhibited shrinking, decreased cell viability and ATP content due to complete exhaustion of glucose at day 6; these results provided evidence that ischemia led to the observed massive cell death. Moreover, MSCs exposed to severe, continuous hypoxia, but without any glucose shortage, remained viable and maintained both their in vitro proliferative ability after simulation with blood reperfusion at day 12 and their in vivo osteogenic ability. These findings challenge the traditional view according to which severe hypoxia per se is responsible for the massive MSC death observed upon transplantation of these cells and provide evidence that MSCs are able to withstand exposure to severe, continuous hypoxia provided that a glucose supply is available.     

Human adult skeletal muscle stem cells differentiate into cardiomyocyte phenotype in vitro

Cell transplantation to repair or regenerate injured myocardium is a new frontier in the treatment of cardiovascular disease. Most studies on stem cell transplantation therapy in both experimental heart infarct and in phase-I human clinical trials have focused on the use of undifferentiated stem cells. Based on our previous observations demonstrating the presence of multipotent progenitor cells in human adult skeletal muscle, in this study we investigated the capacity of these progenitors to differentiate into cardiomyocytes. Here we show an efficient protocol for the cardiomyogenic differentiation of human adult skeletal muscle stem cells in vitro. We found that treatment with Retinoic Acid directed cardiomyogenic differentiation of skeletal muscle stem cells in vitro. After Retinoic Acid treatment, cells expressed cardiomyocyte markers and acquired spontaneous contraction. Functional assays exhibited cardiac-like response to increased extracellular calcium. When cocultured with mouse cardiomyocytes, Retinoic Acid-treated skeletal muscle stem cells expressed connexin43 and when transplanted into ischemic heart were detectable even 5 weeks after injection. Based on these results, we can conclude that human adult skeletal muscle stem cells, if opportunely treated, can transdifferentiate into cells of cardiac lineage and once injected into infarcted heart can integrate, survive in cardiac tissue and improve the cardiac function.     

Mesenchymal stem cell‐loaded cardiac patch promotes epicardial activation and repair of the infarcted myocardium

Cardiac patch is considered a promising strategy for enhancing stem cell therapy of myocardial infarction (MI). However, the underlying mechanisms for cardiac patch repairing infarcted myocardium remain unclear. In this study, we investigated the mechanisms of PCL/gelatin patch loaded with MSCs on activating endogenous cardiac repair. PCL/gelatin patch was fabricated by electrospun. The patch enhanced the survival of the seeded MSCs and their HIF‐1α, Tβ4, VEGF and SDF‐1 expression and decreased CXCL14 expression in hypoxic and serum‐deprived conditions. In murine MI models, the survival and distribution of the engrafted MSCs and the activation of the epicardium were examined, respectively. At 4 weeks after transplantation of the cell patch, the cardiac functions were significantly improved. The engrafted MSCs migrated across the epicardium and into the myocardium. Tendency of HIF‐1α, Tβ4, VEGF, SDF‐1 and CXCL14 expression in the infarcted myocardium was similar with expression in vitro. The epicardium was activated and epicardial‐derived cells (EPDCs) migrated into deep tissue. The EPDCs differentiated into endothelial cells and smooth muscle cells, and some of EPDCs showed to have differentiated into cardiomyocytes. Density of blood and lymphatic capillaries increased significantly. More c‐kit⁺ cells were recruited into the infarcted myocardium after transplantation of the cell patch. The results suggest that epicardial transplantation of the cell patch promotes repair of the infarcted myocardium and improves cardiac functions by enhancing the survival of the transplanted cells, accelerating locality paracrine, and then activating the epicardium and recruiting endogenous c‐kit⁺ cells. Epicardial transplantation of the cell patch may be applied as a novel effective MI therapy.     

Graphene enhances the cardiomyogenic differentiation of human embryonic stem cells

Graphene has drawn attention as a substrate for stem cell culture and has been reported to stimulate the differentiation of multipotent adult stem cells. Here, we report that graphene enhances the cardiomyogenic differentiation of human embryonic stem cells (hESCs) at least in part, due to nanoroughness of graphene. Large-area graphene on glass coverslips was prepared via the chemical vapor deposition method. The coating of the graphene with vitronectin (VN) was required to ensure high viability of the hESCs cultured on the graphene. hESCs were cultured on either VN-coated glass (glass group) or VN-coated graphene (graphene group) for 21 days. The cells were also cultured on glass coated with Matrigel (Matrigel group), which is a substrate used in conventional, directed cardiomyogenic differentiation systems. The culture of hESCs on graphene promoted the expression of genes involved in the stepwise differentiation into mesodermal and endodermal lineage cells and subsequently cardiomyogenic differentiation compared with the culture on glass or Matrigel. In addition, the culture on graphene enhanced the gene expression of cardiac-specific extracellular matrices. Culture on graphene may provide a new platform for the development of stem cell therapies for ischemic heart diseases by enhancing the cardiomyogenic differentiation of hESCs.     

骨髓间充质干细胞移植对心衰大鼠心肌结构和功能的影响

研究骨髓间充质干细胞(MSC)移植对心力衰竭(简称心衰)大鼠心肌结构和功能的影响以及在病损心肌体内分化为心肌细胞的情况。将96只Wistar大鼠,用阿霉素成功诱导了54只心衰模型,随机分成3组,移植组为左室前壁注射MSC,对照组注射培养基,心衰组不给予任何干预措施。由彩色超声心动图(TTE)监测左室心功能参数。8周检测完成后取出心脏标本,做冰冻切片脏染色观察病损心肌结构的变化及免疫荧光检查植入MSC心肌肌球蛋白重链(MHC)及心肌特有的连接蛋白(Cx43)表达情况。结果表明植入的MSC存活并表达了MHC及Cx43,其周围宿主心肌细胞肿胀明显减轻。在移植MSC2周后,心功能开始改善,至8周时,心功改善能更明显。由此得出结论：MSC在病损心肌体内不仅能存活、分化为心肌细胞,使病损心肌组织病变减轻。而且可显著改善心衰大鼠的心功能。