Proliferation Of Cardiac Stem Cells Research Articles

Restoring Ravaged Heart: Molecular Mechanisms and Clinical Application of miRNA in Heart Regeneration.

Human heart development is a complex and tightly regulated process, conserving proliferation, and multipotency of embryonic cardiovascular progenitors. At terminal stage, progenitor cell type gets suppressed for terminal differentiation and maturation. In the human heart, most cardiomyocytes are terminally differentiated and so have limited proliferation capacity. MicroRNAs (miRNAs) are non-coding single-stranded RNA that regulate gene expression and mRNA silencing at the post-transcriptional level. These miRNAs play a crucial role in numerous biological events, including cardiac development, and cardiomyocyte proliferation. Several cardiac cells specific miRNAs have been discovered. Inhibition or overexpression of these miRNAs could induce cardiac regeneration, cardiac stem cell proliferation and cardiomyocyte proliferation. Clinical application of miRNAs extends to heart failure, wherein the cell cycle arrest of terminally differentiated cardiac cells inhibits the heart regeneration. The regenerative capacity of the myocardium can be enhanced by cardiomyocyte specific miRNAs controlling the cell cycle. In this review, we focus on cardiac-specific miRNAs involved in cardiac regeneration and cardiomyocyte proliferation, and their potential as a new clinical therapy for heart regeneration.

Abstract 3694: Differential expressions of miR-17 and complement factor D in carfilzomib-related cardiotoxicity in multiple myeloma patients: Results of microRNA array and proteomics integrated analysis

Abstract Background: Carfilzomib (CFZ), a proteasome inhibitor used to treat multiple myeloma (MM), has been associated with risk for cardiotoxicity including heart failure (HF). There is a lack of data on the mechanism underlying CFZ-induced cardiotoxicity. Our study aims to better understand molecular mechanisms of CFZ-related cardiotoxicity by determining the differential expression of circulating miRNAs and proteins in MM patients. Methods: Plasma samples were collected from 13 MM patients after treated with CFZ (including 7 with cardiotoxicity and 6 with no cardiotoxicity) at the University of Florida Health Cancer Center. TaqMan OpenArray Human MicroRNA Panel containing 754 human miRNA was used for miRNA analysis, and iTRAQ analysis on LC/MS/MS platforms was used for global proteomic profiling. Results: The overall mean age was 61 and 38.5% were men. A total of 374 proteins were identified and 57 were differentially expressed between cardiotoxicity and non-cardiotoxicity groups (false discovery rate <0.05). Differential expression analysis of miRNAs profiling yielded 5 miRNAs that were dysregulated (p-value <0.05 and fold change (FC) < 0.5): miR-17, miR-30c, miR-30b, miR-106a, and miR-92a. Among the top proteins that differentiate the two groups were complement factor D (CFD) (P= 5.5*10-5, and FC= 6), which is targeted by miR-17. CFD circulates in an active form and cleaves complement factor B (FB) to Ba and Bb, which promote the alternative pathway complement (AP) activation, the complement activation has shown to occur in chronic HF, and is associated with adverse clinical events. Also, CFD has a function in the activation of notch signaling, which is required for the proliferation of cardiac stem cells and the survival of cardiomyocytes. Conclusion: These findings suggest that CFZ may induce cardiac dysfunction via increasing CFD activity that is regulated by miR-17, leading to activation of the complement system. Dysregulation in components of the AP of complement activation could impact the diastolic dysfunction associated with HF progression. Citation Format: Marwa Tantawy, Lakshmi Manasa Chekka, Taimour Langaee, Timothy J Garrett, Michael Fradley, Robert Frank Cornell, Rachid C. Baz, Sonal Singh, Lihui Yuan, Chintan P. Shah, Nida Waheed, Carl J. Pepine, Jan S. Moreb, Yan Gong. Differential expressions of miR-17 and complement factor D in carfilzomib-related cardiotoxicity in multiple myeloma patients: Results of microRNA array and proteomics integrated analysis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3694.

MiR-155 regulates placental development and fetal growth, probably by inhibiting the expression of angiotensin II type 1 receptor (AT1R).

The adult myocardium, including human, harbours a population of resident multi-potent cardiac stem cells (CSCs), which when stimulated under the right conditions can give rise to new cardiomyocytes and vasculature. Elucidation of the cellular and molecular mechanisms that govern CSC biology and their role in myocardial regeneration will allow the design and development of optimal therapeutic interventions. It is now evident that different growth factors and cytokines govern CSC survival, proliferation, migration and differentiation, as well as playing a role in activating cardiac repair mechanisms such as improving angiogenesis, cardiomyocyte survival and limiting fibrosis. This review article will summarize the evidence for a role of VEGF, NRG-1, IGF-1, HGF, EGF, FGF and TGF-β1 in modulating the repair and regeneration of cardiac tissue. It will also discuss the use of exosomes and exercise training as interventions to stimulate the endogenous repair and regenerative mechanisms in the damaged heart.

Combination of Mesenchymal Stromal Cells and Cardiac Stem Cells in a Multilayer Cell Construct Promotes Activation of Notch Signaling and Initiation of Endothelial Differentiation

We showed the possibility of generating combined tissue-engineered cell consisting of layers of rat cardiac stem cells and mesenchymal stromal cells from the adipose tissue. Cell-cell interaction within the cell sheet promoted proliferation of cardiac stem cells, expression of endothelial differentiation marker ETS1, and Notch signaling activation. The obtained results provide new insights into possible mechanisms of stimulation of endogenous regeneration processes after myocardial damage and demonstrate potential of cell-based cardiomyoplasty with the use of these combined cell sheets.

Insulin-like growth factor-1-mediated regulation of miR-193a expression promotes the migration and proliferation of c-kit-positive mouse cardiac stem cells

BackgroundC-kit-positive cardiac stem cells (CSCs) have been shown to be a promising candidate treatment for myocardial infarction and heart failure. Insulin-like growth factor (IGF)-1 is an anabolic growth hormone that regulates cellular proliferation, differentiation, senescence, and death in various tissues. Although IGF-1 promotes the migration and proliferation of c-kit-positive mouse CSCs, the underlying mechanism remains unclear.MethodsCells were isolated from adult mouse hearts, and c-kit-positive CSCs were separated using magnetic beads. The cells were cultured with or without IGF-1, and c-kit expression was measured by Western blotting. IGF-1 induced CSC proliferation and migration, as measured through Cell Counting Kit-8 (CCK-8) and Transwell assays, respectively. The miR-193a expression was measured by quantitative real-time PCR (qPCR) assays.ResultsIGF-1 enhanced c-kit expression in c-kit-positive CSCs. The activities of the phosphoinositol 3-kinase (PI3K)/AKT signaling pathway and DNA methyltransferases (DNMTs) were enhanced, and their respective inhibitors LY294002 and 5-azacytidine (5-AZA) blunted c-kit expression. Based on the results of quantitative real-time PCR (qPCR) assays, the expression of miR-193a, which is embedded in a CpG island, was down-regulated in the IGF-1-stimulated group and negatively correlated with c-kit expression, whereas c-kit-positive CSCs infected with lentivirus carrying micro-RNA193a displayed reduced c-kit expression, migration and proliferation.ConclusionsIGF-1 upregulated c-kit expression in c-kit-positive CSCs resulting in enhanced CSC proliferation and migration by activating the PI3K/AKT/DNMT signaling pathway to epigenetically silence miR-193a, which negatively modifies the c-kit expression level.

Modulation of cardiac stem cell characteristics by metoprolol in hypertensive heart disease.

Cardiac stem cells (CSCs) play a vital role in cardiac remodeling. Uncontrolled hypertension leads to cardiac hypertrophy, followed by cardiac failure. Pathological remodeling is associated with enhanced oxidative stress. Decreased cardiac stem cell efficiency is speculated in heart diseases. Maintaining a healthy stem cell population is essential for preventing progressive cardiac remodeling. Some anti-hypertensive drugs are cardioprotective. However, the effect of these drugs on CSCs has not been investigated. Metoprolol is a cardioprotective anti-hypertensive agent. To examine whether metoprolol can prevent the deterioration of CSC efficiency, spontaneously hypertensive rats (SHRs) were treated with this drug, and the effects on stem cell function were evaluated. Six-month-old male SHRs were treated with metoprolol (50 mg × kg-1per day) for 2 months. The effectiveness of the treatment at reducing blood pressure and reducing hypertrophy was ensured, and the animals were killed. Cardiac stem cells were isolated from the atrial tissue, and the effect of metoprolol on stem cell migration, proliferation, differentiation, and survival was evaluated by comparing the treated SHRs with untreated SHRs and normotensive Wistar rats. Compared to the Wistar rats, the SHR rats presented with a decrease in stem cell migration and proliferation and an increase in intracellular oxidative stress and senescence. Treating SHRs with metoprolol increased CSC migration and proliferation potential and stemness retention. Cellular senescence and oxidative stress were reduced. The attributes of stem cells from the metoprolol-treated SHRs were comparable to those of the Wistar rats. The restoration of stem cell efficiency is expected to prevent hypertension-induced progressive cardiac remodeling.

Bmi1 and BRG1 drive myocardial repair by regulating cardiac stem cell function in acute rheumatic heart disease.

Rheumatic heart disease (RHD) occurs due to the accumulation of complications associated with rheumatic fever, and it results in high morbidity and mortality. The majority of cases of RHD are diagnosed in the chronic stages, when treatment options are limited. A small reservoir of cardiac stem cells is responsible for maintaining cardiac homeostasis and repairing tissue damage. Understanding the role of cardiac stem cells and the various proteins responsible for their functions in different pathological stages of RHD is an important area of investigation. Polycomb complex protein BMI-1 (Bmi1) and transcription activator BRG1 (BRG1) are associated with the maintenance of stemness in various types of stem cells. The present study investigated the role served by Bmi1 and BRG1 in cardiac stem cells during various pathological stages of RHD through immunohistochemistry and western blotting. A rat model of RHD was established via immunization with the Group A Streptococcus M5 protein. The rat was demonstrated to develop acute RHD 2 months after the final immunization, characterized by cardiac inflammation and tissue damage. Chronic RHD was identified 4 months after the final immunization, revealed by cardiac tissue compression and shrinkage. Expression of the cardiac stem cell marker mast/stem cell growth factor receptor kit was identified to be elevated during acute RHD, but downregulated in the chronic stages of RHD. A similar pattern of expression was revealed for Bmi1 and BRG1, indicating that they serve a role in regulating cardiac stem cell proliferation during acute RHD. These results suggest that cardiac stem cells serve a supportive role in the acute, but not chronic, stages of RHD via expression of Bmi1 and BRG1.

MiR-21 increases c-kit+ cardiac stem cell proliferation in vitro through PTEN/PI3K/Akt signaling.

The low survival rate of cardiac stem cells (CSCs) in the ischemic myocardium is one of the obstacles in ischemic cardiomyopathy cell therapy. The MicroRNA (miR)-21 and one of its target protein, the tensin homolog deleted on chromosome ten (PTEN), contributes to the proliferation of many kinds of tissues and cell types. It is reported that miR-21 promotes proliferation through PTEN/PI3K/Akt pathway, but its effects on c-kit+ CSC remain unclear. The authors hypothesized that miR-21 promotes the proliferation in c-kit+ CSC, and evaluated the involvement of PTEN/PI3K/Akt pathway in vitro. miR-21 up-regulation with miR-21 efficiently mimics accelerated cell viability and proliferation in c-kit+ CSC, which was evidenced by the CCK-8, EdU and cell cycle analyses. In addition, the over-expression of miR-21 in c-kit+ CSCs notably down-regulated the protein expression of PTEN although the mRNA level of PTEN showed little change. Gain-of-function of miR-21 also increased the phosphor-Akt (p-Akt) level. Phen, the selective inhibitor of PTEN, reproduced the pro-proliferation effects of miR-21, while PI3K inhibitor, LY294002, totally attenuated the pro-survival effect of miR-21. These results indicate that miR-21 is efficient in promoting proliferation in c-kit+ CSCs, which is contributed by the PTEN/PI3K/Akt pathway. miR-21 holds the potential to facilitate CSC therapy in ischemic myocardium.

Engineered extracellular microenvironment with a tunable mechanical property for controlling cell behavior and cardiomyogenic fate of cardiac stem cells.

The extracellular matrix (ECM) provides a physical framework of myocardial niches in which endogenous cardiac stem cells (CSCs) reside, renew, differentiate, and replace cardiac cells. Interactions between ECM and CSCs might be critical for the maintenance of myocardial homeostasis in the adult heart. Yet most studies done so far have used irrelevant cell types and have been performed at the individual cell level, none able to reflect the in vivo situation. By the use of a chemically defined hydrogel to create a tunable 3D microenvironment, we succeeded in controlling CSC behavior at the myocardial and individual cell level and directing the cardiomyogenic fate. Our work may provide insight into the design of biomaterials for in situ myocardial regeneration as well as for tissue engineering.

Comparison of cardiac stem cell sheets detached by Versene solution and from thermoresponsive dishes reveals similar properties of constructs.

Cell sheets (CS) from c-kit+ cardiac stem cell (CSC) hold a potential for application in regenerative medicine. However, manufacture of CS may require thermoresponsive dishes, which increases cost and puts one in dependence on specific materials. Alternative approaches were established recently and we conducted a short study to compare approaches for detachment of CS from c-kit+ CSC. Our in-house developed method using chelation by Versene solution was compared to UpCell™ thermoresponsive plates in terms of CSC proliferation, viability, gap junction formation and engraftment in a model of myocardial infarction. Use of Versene solution instead of thermoresponsive dishes resulted in comparable CS thickness (approximately 100mcm), cell proliferation rate and no signs of apoptosis detected in both types of constructs. However, we observed a minor reduction of gap junction count in Versene-treated CS. At day 30 after delivery to infarcted myocardium both types of CS retained at the site of transplantation and contained comparable amounts of proliferating cells indicating engraftment. Thus, we may conclude that detachment of CS from c-kit+ CSC using Versene solution followed by mechanical treatment is an alternative to thermoresponsive plates allowing use of routinely available materials to generate constructs for cardiac repair.

Corrigendum.

CorrigendaCorrigendumPublished Online:02 Dec 2016https://doi.org/10.1152/ajpheart.H-zh4-2124-corr.2016Original articleMoreSectionsPDF (36 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInWeChat Volume 311, December 2016Volume 80, December 2016Bellio MA, Rodrigues CO, Landin A, Hatzistergos KE, Kuznetsov J, Florea V, Valasaki K, Khan A, Hare JM, Schulman IH. Physiological and hypoxic oxygen concentration differentially regulates human c-Kit+ cardiac stem cell proliferation and migration. Am J Physiol Hert Circ Physiol 311: pages TBD, 2016. First published September 30, 2016; doi: https://doi.org/10.1152/ajpheart.00449.2016.— In the Articles-in-Press version of the manuscript only, posted online ahead-of-print on September 30, 2016, there has been a change of authorship with the addition of a previously undeclared coauthor, Jeffim Kuznetsov.As such, the new author line of the manuscript reads as follows: Michael A. Bellio,1 Claudia O. Rodrigues,1 Ana Marie Landin,1 Konstantinos E. Hatzistergos,1 Jeffim Kuznetsov,1 Victoria Florea,1 Krystalenia Valasaki,1 Aisha Khan,1 Joshua M. Hare,1 and Ivonne Hernandez Schulman1,2.The revised author line in the Abstract is as stated above in the Abstract citation for this corrigendum.Finally, the revised Author Contribution line includes J. Kuznetsov as contributing to the performing of experiments, as follows: M.A.B., J.M.H., and I.H.S. conception and design of research; M.A.B., K.E.H., V.F., K.V., J.K., and A.K. performed experiments; M.A.B., A.M.L., and K.E.H. analyzed data; M.A.B., C.O.R., A.M.L., K.E.H., V.F., J.M.H., and I.H.S. interpreted results of experiments; M.A.B. prepared figures; M.A.B. drafted manuscript; M.A.B., C.O.R., A.M.L., J.M.H., and I.H.S. edited and revised manuscript; M.A.B., C.O.R., A.M.L., K.E.H., V.F., K.V., A.K., J.M.H., and I.H.S. approved final version of manuscript.This article has no references to display. Download PDF Previous Back to Top FiguresReferencesRelatedInformation Collections Related ArticlesPhysiological and hypoxic oxygen concentration differentially regulates human c-Kit+ cardiac stem cell proliferation and migration 02 Dec 2016American Journal of Physiology-Heart and Circulatory Physiology More from this issue > Volume 311Issue 6December 2016Pages H1572-H1572 Copyright & PermissionsCopyright © 2016 the American Physiological Societyhttps://doi.org/10.1152/ajpheart.H-zh4-2124-corr.2016PubMed27913555History Published online 2 December 2016 Published in print 1 December 2016 Metrics

Physiological and hypoxic oxygen concentration differentially regulates human c-Kit+ cardiac stem cell proliferation and migration.

Cardiac stem cells (CSCs) are being evaluated for their efficacy in the treatment of heart failure. However, numerous factors impair the exogenously delivered cells' regenerative capabilities. Hypoxia is one stress that contributes to inadequate tissue repair. Here, we tested the hypothesis that hypoxia impairs cell proliferation, survival, and migration of human CSCs relative to physiological and room air oxygen concentrations. Human endomyocardial biopsy-derived CSCs were isolated, selected for c-Kit expression, and expanded in vitro at room air (21% O2). To assess the effect on proliferation, survival, and migration, CSCs were transferred to physiological (5%) or hypoxic (0.5%) O2 concentrations. Physiological O2 levels increased proliferation (P < 0.05) but did not affect survival of CSCs. Although similar growth rates were observed in room air and hypoxia, a significant reduction of β-galactosidase activity (-4,203 fluorescent units, P < 0.05), p16 protein expression (0.58-fold, P < 0.001), and mitochondrial content (0.18-fold, P < 0.001) in hypoxia suggests that transition from high (21%) to low (0.5%) O2 reduces senescence and promotes quiescence. Furthermore, physiological O2 levels increased migration (P < 0.05) compared with room air and hypoxia, and treatment with mesenchymal stem cell-conditioned media rescued CSC migration under hypoxia to levels comparable to physiological O2 migration (2-fold, P < 0.05 relative to CSC media control). Our finding that physiological O2 concentration is optimal for in vitro parameters of CSC biology suggests that standard room air may diminish cell regenerative potential. This study provides novel insights into the modulatory effects of O2 concentration on CSC biology and has important implications for refining stem cell therapies.

Enhancement of Anti-Hypoxic Activity and Differentiation of Cardiac Stem Cells by Supernatant Fluids from Cultured Macrophages that Phagocytized Dead Mesenchymal Stem Cells.

Background: Most mesenchymal stem cells (MSCs) die shortly after transplantation into a myocardial infarcted area. Dead MSCs (dMSCs) are phagocytized by macrophages (pMΦ) in vivo and in vitro; however, the effects of pMΦ on cardiac stem cells (CSCs) remain unknown. Methods: MSCs, CSCs, and macrophages were obtained from bone marrow, hearts, and peritoneal cavity of mice, respectively. dMSCs were harvested after hypoxia for 24 h, and incubated with macrophages (2:1) for another 2 days with or without lipopolysaccharide (LPS, 50 ng/mL) and sorted by flow cytometry to obtain pMΦ. Viability and apoptosis of CSCs were respectively evaluated with the cell counting kit-8 (CCk-8) assay and Annexin V-PE/7-AAD staining at 0, 6, 12, and 24 h of culture with supernatant fluids from macrophages (MΦ), LPS-stimulated macrophages (LPS-pMΦ), pMΦ, and MSCs. GATA-4 and c-TnI expression was measured by flow cytometry on the seventh day. Expression of inflammation and growth factors was assessed by real-time polymerase chain reaction (RT-PCR) in MΦ, LPS-pMΦ, and pMΦ cells. Results: pMΦ expressed higher levels of interleukin-10 (IL-10) and transforming growth factor-β (TGF-β)and lower levels of tumor necrosis factor-α(TNF-α)and IL-6 than LPS-pMΦ, higher levels of growth factors and of GATA-4 and c-TnI at the 7th day, which were similar to those in MSCs. CSCs cultured with supernatant fluids of pMΦ exhibited higher proliferative, anti-hypoxic, and differentiation activities. Conclusion: The supernatant fluids of macrophages that had phagocytized dead MSCs encouraged changes in phenotype and growth factor expression, enhanced proliferation, differentiation, and anti-hypoxic activity of CSCs, which is relevant to understanding the persistent therapeutic effect of MSCs after their massive demise upon transplantation in myocardial infarction. Furthermore, some miRNAs or proteins which were extracted from the supernatant fluids may give us a new insight into the treatment of myocardial infarction in the future.

Macrophage migration inhibitory factor promotes cardiac stem cell proliferation and endothelial differentiation through the activation of the PI3K/Akt/mTOR and AMPK pathways.

Macrophage migration inhibitory factor (MIF) has pleiotropic immune functions in a number of inflammatory diseases. Recent evidence from expression and functional studies has indicated that MIF is involved in various aspects of cardiovascular disease. In this study, we aimed to determine whether MIF supports in vitro c-kit+CD45− cardiac stem cell (CSC) survival, proliferation and differentiation into endothelial cells, as well as the possible mechanisms involved. We observed MIF receptor (CD74) expression in mouse CSCs (mCSCs) using PCR and immunofluorescence staining, and MIF secretion by mCSCs using PCR and ELISA in vitro. Increasing amounts of exogenous MIF did not affect CD74 expression, but promoted mCSC survival, proliferation and endothelial differentiation. By contrast, treatment with an MIF inhibitor (ISO-1) or siRNA targeting CD74 (CD74-siRNA) suppressed the biological changes induced by MIF in the mCSCs. Increasing amounts of MIF increased the phosphorylation of Akt and mammalian target of rapamycin (mTOR), which are known to support cell survival, proliferation and differentiation. These effects of MIF on the mCSCs were abolished by LY294002 [a phosphoinositide 3-kinase (PI3K) inhibitor] and MK-2206 (an Akt inhibitor). Moreover, adenosine monophosphate-activated protein kinase (AMPK) phosphorylation increased following treatment with MIF. The AMPK inhibitor, compound C, partly blocked the pro-proliferative effects of MIF on the mCSCs. In conclusion, our results suggest that MIF promotes mCSC survival, proliferation and endothelial differentiation through the activation of the PI3K/Akt/mTOR and AMPK signaling pathways. Thus, MIF may prove to be a potential therapeutic factor in the treatment of heart failure and myocardial infarction by activating CSCs.

MiR218 Modulates Wnt Signaling in Mouse Cardiac Stem Cells by Promoting Proliferation and Inhibiting Differentiation through a Positive Feedback Loop.

MiRNA expression was determined in both proliferating and differentiated cardiac stem cells (CSCs) through a comprehensive miRNA microarray analysis. We selected miR218 for functional follow-up studies to examine its significance in CSCs. First, we observed that the expression of miR218 was altered in CSCs during differentiation into cardiomyocytes, and transfection of an miR218 mimic or miR218 inhibitor affected the myocardial differentiation of CSCs. Furthermore, we observed that a negative regulator of Wnt signaling, sFRP2, was a direct target of miR218, and the protein levels of sFRP2 were increased in cells transfected with the synthetic miR218 inhibitor. In contrast, transfection with the miR218 mimic decreased the expression of sFRP2 and potentiated Wnt signaling. The subsequent down-regulation of sFRP2 by shRNA potentiated Wnt signaling, contributing to a gene expression program that is important for CSC proliferation and cardiac differentiation. Specifically, canonical Wnt signaling induced miR218 transcription. Thus, miR218 and Wnt signaling were coupled through a feed-forward positive feedback loop, forming a biological regulatory circuit. Together, these results provide the first evidence that miR218 plays an important role in CSC proliferation and differentiation through the canonical Wnt signaling pathway.

Inside Cover, Volume 133, Issue 14

Journal of Applied Polymer ScienceVolume 133, Issue 14 Cover ImageFree Access Inside Cover, Volume 133, Issue 14 First published: 08 January 2016 https://doi.org/10.1002/app.43406AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat Graphical Abstract Sasan Jalili-Firoozinezhad, Sareh Rajabi-Zeleti, Anna Marsano, Nasser Aghdami and Hossein Baharvand discuss the preparation of decellularized pericardium membranes by a mild devitalization process on human pericardium tissue, where most of the native extracellular cues are preserved, while more than 99% of cellular components are removed. Decellularized pericardium, as a bioactive material, not only supports survival, metabolism and proliferation of cardiac stem cells, but also promotes their differentiation into mature cardiomyocytes. This native biomaterial could find use in various cardiovascular applications, such as valve replacement, cardiac patches and vascular grafts. (DOI: 10.1002/app.43255) Volume133, Issue14April 10, 2016 RelatedInformation

Necroptosis Induced by Ad-HGF Activates Endogenous C-Kit+ Cardiac Stem Cells and Promotes Cardiomyocyte Proliferation and Angiogenesis in the Infarcted Aged Heart

Background/Aims: The discovery of c-kit⁺ cardiac stem cells (CSCs) provided us with new therapeutic targets to repair the damaged heart. However, the precise mechanisms regulating CSC proliferation and differentiation in the aged heart remained elusive. Necroptosis, a type of regulated cell death, has recently been shown to occur following myocardial infarction (MI); however, its effect on c-kit⁺ CSCs remains unknown. We investigated the effects of hepatocyte growth factor (HGF) and necroptosis on the proliferation and differentiation of endogenous c-kit⁺ CSCs in aged rat hearts following MI. Methods: The c-kit⁺ CSCs and HGF/p-Met expression levels in neonatal, adult and aged rats were compared using immunofluorescence and Western blotting. Immediately after MI, adenovirus carrying the HGF gene (Ad-HGF) was injected into the left ventricular wall surrounding the infarct areas of the aged rat heart. The proliferation and differentiation of the endogenous c-kit⁺ CSCs were studied using immunofluorescence. The signalling pathways were analysed via Western blotting and ELISA. Results: HGF/p-Met expression levels and c-kit⁺ CSC abundance gradually decreased with age. Ad-HGF promoted c-kit⁺ CSC differentiation into precursor cells of cardiomyocyte, endothelial and smooth muscle cell lineages and enhanced cardiomyocyte proliferation and angiogenesis in aged rats; these effects were reversed by the inhibition of necroptosis. Ad-HGF administration induced necroptosis by increasing the expression of receptor interacting protein kinase (RIP) 1 and receptor interacting protein kinase (RIP) 3 proteins in the infarcted heart. Moreover, Ad-HGF-induced necroptosis increased high-mobility group box 1 protein (HMGB1) levels and enhanced the abundance of c-kit⁺ cells in the bone marrow, which may partly account for the beneficial effect of necroptosis on the c-kit⁺ CSCs. Conclusion: Ad-HGF-induced necroptosis facilitated aged heart repair after MI by promoting c-kit⁺ CSC proliferation and differentiation. These findings may lead to the development of new methods for the treatment of ischaemic heart disease in aged populations.

Mesenchymal Stem Cells in Cardiology.

Cardiovascular disease (CVD) accounts for more deaths globally than any other single disease. There are on average 1.5 million episodes of myocardial infarction (heart attack) each year in the United States alone with roughly one-third resulting in death. There is therefore a major need for developing new and effective strategies to promote cardiac repair. Intramyocardial transplantation of mesenchymal stem cells (MSCs) has emerged as a leading contender in the pursuit of clinical intervention and therapy. MSCs are potent mediators of cardiac repair and are therefore an attractive tool in the development of preclinical and clinical trials. MSCs are capable of secreting a large array of soluble factors, which have had demonstrated effects on pathogenic cardiac remolding, fibrosis, immune activation, and cardiac stem cell proliferation within the damaged heart. MSCs are also capable of differentiation into cardiomyocytes, endothelial cells, and vascular smooth muscle cells, although the relative contribution of trilineage differentiation and paracrine effectors on cardiac repair remains the subject of active investigation.

REPETITIVE TARGETED DELIVERY OF GDF11 BY ULTRASOUND-MEDIATED CATIONIC MICROBUBBLE DESTRUCTION REJUVENATES AND PROTECTS THE AGED MOUSE HEART

Preclinical studies demonstrated dramatic improvements in ventricular function with the implantation of stem cells into the damaged myocardium. However, stem cells in aged recipients have reduced regenerative potential, and aged tissues have diminished renewal capacity even with potent stem cells. Growth differentiation factor 11 (GDF11), a member of the TGF-β superfamily, has been shown to reverse age-related cardiac hypertrophy and skeletal muscular dysfunction, remodel the cerebral vasculature of aged mice and enhance olfactory neurogenesis. However, systemic delivery of recombinant GDF11 protein may not be the ideal method to rejuvenate specific organs. To evaluate a non-invasive method to protect aged hearts from ischemic injury, we tested whether a GDF11 expression plasmid construct could be delivered via ultrasound-targeted microbubble destruction (UTMD) using our effective cationic microbubbles (CMBs). Young (aged 3 months) and old (aged 20-22 months) C57BL/6 mice were used to evaluate the expression of GDF11 mRNA and protein at baseline and after ischemia/reperfusion (I/R) injury. GDF11 mRNA and protein expression at baseline in old mice was significantly lower than in young mice. GDF11 transcript and protein expression were significantly upregulated in the ischemic hearts of young and old mice 3 days after I/R compared to baseline, but the increase in the aged ischemic hearts was less than in young mice after I/R injury. To enhance the expression of GDF11 in aged mouse hearts, UTMD was used to deliver a mouse GDF11 expression plasmid to the ischemic hearts of aged mice 3 and 6 days after I/R. A control group received the empty vector. UTMD-mediated delivery of GDF11 effectively increased the expression of GDF11 specifically in cardiac tissue, but not in other organs of the old mice. UTMD-mediated delivery of GDF11 improved cardiac systolic and diastolic function and reduced infarct size in the aged mice 21 days after I/R. Additional investigations to determine the underlying mechanism of this functional benefit revealed that UTMD-mediated delivery of GDF11 stimulated cKit+ and Sca-1+ cardiac stem cell proliferation and decreased the expression of senescence markers (p16, p53) and the number of p16+ CSCs in the hearts of old mice post-I/R. Repetitive targeted delivery of GDF11 plasmid via UTMD using CMBs can rejuvenate aged mouse hearts and protect them from I/R injury.

The roles of nanocarriers on pigment epithelium-derived factor in the differentiation of human cardiac stem cells.

Over the past decade, adult stem cells have attracted great attention because of their ability to potentially regenerate desired tissues or entire organs. With the emergence of nanomaterial-based gene therapy, adult stem cells have been considered as a proper tool for the biomedical field. In this study, we utilized organically modified silica (ORMOSIL) nanoparticles to deliver small interfering RNA (siRNA) against pigment epithelium-derived factor (PEDF) and induce the differentiation of human cardiac stem cells (CSCs). We found that the down-regulation of PEDF can inhibit the proliferation of human CSCs and induce cell differentiation. To further study the mechanism, we have tested the Notch signalling pathway genes, Hes1 and Hes5, and found that their expressions were inhibited by the PEDF down-regulation. Furthermore, with the restoration of PEDF, both the proliferation of human CSCs and expressions of Hes1 and Hes5 were recovered. Our results suggest for the first time the use of ORMOSIL as nanocarriers for the delivery of PEDF siRNA in human CSCs, and demonstrated the cooperation between PEDF and the Notch signalling pathway in maintaining the self-renewal and pluripotency of stem cells. PEDF as the essential controller in differentiation may be a promising target for the regulation of cardiac homeostasis and damage repair, which opens new treatment strategies using nanomaterials for heart disease therapy.