Bone marrow-derived cells can acquire cardiac stem cells properties in damaged heart

Experimental data suggest that cell-based therapies may be useful for cardiac regeneration following ischaemic heart disease. Bone marrow (BM) cells have been reported to contribute to tissue repair after myocardial infarction (MI) by a variety of humoural and cellular mechanisms. However, there is no direct evidence, so far, that BM cells can generate cardiac stem cells (CSCs). To investigate whether BM cells contribute to repopulate the Kit+ CSCs pool, we transplanted BM cells from transgenic mice, expressing green fluorescent protein under the control of Kit regulatory elements, into wild-type irradiated recipients. Following haematological reconstitution and MI, CSCs were cultured from cardiac explants to generate 'cardiospheres', a microtissue normally originating in vitro from CSCs. These were all green fluorescent (i.e. BM derived) and contained cells capable of initiating differentiation into cells expressing the cardiac marker Nkx2.5. These findings indicate that, at least in conditions of local acute cardiac damage, BM cells can home into the heart and give rise to cells that share properties of resident Kit+ CSCs

Functional impairment of human resident cardiac stem cells by the cardiotoxic antineoplastic agent trastuzumab

Trastuzumab (TZM), a monoclonal antibody against the ERBB2 protein, increases survival in ERBB2-positive breast cancer patients. Its clinical use, however, is limited by cardiotoxicity. We sought to evaluate whether TZM cardiotoxicity involves inhibition of human adult cardiac-derived stem cells, in addition to previously reported direct adverse effects on cardiomyocytes. To test this idea, we exposed human cardiosphere-derived cells (hCDCs), a natural mixture of cardiac stem cells and supporting cells that has been shown to exert potent regenerative effects, to TZM and tested the effects in vitro and in vivo. We found that ERBB2 mRNA and protein are expressed in hCDCs at levels comparable to those in human myocardium. Although clinically relevant concentrations of TZM had no effect on proliferation, apoptosis, or size of the c-kit-positive hCDC subpopulation, in vitro assays demonstrated diminished potential for cardiogenic differentiation and impaired ability to form microvascular networks in TZM-treated cells. The functional benefit of hCDCs injected into the border zone of acutely infarcted mouse hearts was abrogated by TZM: infarcted animals treated with TZM + hCDCs had a lower ejection fraction, thinner infarct scar, and reduced capillary density in the infarct border zone compared with animals that received hCDCs alone (n = 12 per group). Collectively, these results indicate that TZM inhibits the cardiomyogenic and angiogenic capacities of hCDCs in vitro and abrogates the morphological and functional benefits of hCDC transplantation in vivo. Thus, TZM impairs the function of human resident cardiac stem cells, potentially contributing to TZM cardiotoxicity

Early and long-term outcomes after manual and remote magnetic navigation guided catheter ablation for ventricular tachycardia

Aims Remote magnetic navigation (RMN) is a safe and effective means of performing ventricular tachycardia (VT) ablation. It may have advantages over manual catheter ablation due to ease of manoeuvrability and catheter stability. We sought to compare the safety and efficacy of RMN vs. manual VT ablation. Methods and results Retrospective study of procedural outcomes of 139 consecutive VT ablation procedures (69 RMN, 70 manual ablation) in 113 patients between 2009 and 2015 was performed. Remote magnetic navigation was associated with overall higher acute procedural success (80% vs. 60%, P = 0.01), with a trend to fewer major complications (3% vs. 9% P = 0.09). Seventy-nine patients were followed up for a median of 17.0 [interquartile range (IQR) 3.0–41.0] months for the RMN group and 15.5 (IQR 6.5–30.0) months for manual ablation group. In the ischaemic cardiomyopathy subgroup, RMN was associated with longer survival from the composite endpoint of VT recurrence leading to defibrillator shock, re-hospitalization or repeat catheter ablation and all-cause mortality; single-procedure adjusted hazard ratio (HR) 0.240 (95% CI 0.070–0.821) P = 0.023, multi-procedure HR 0.170 (95% CI 0.046–0.632) P = 0.002. In patients with implanted defibrillators, multi-procedure VT-free survival was superior with RMN, HR 0.199 (95% CI 0.060–0.657) P = 0.003. Conclusion Remote magnetic navigation may improve clinical outcomes after catheter ablation of VT in patients with ischaemic cardiomyopathy. Further prospective clinical studies are required to confirm these findings

Cellular heterogeneity of pluripotent stem cell-derived cardiomyocyte grafts is mechanistically linked to treatable arrhythmias

Preclinical data have confirmed that human pluripotent stem cell-derived cardiomyocytes (PSC-CMs) can remuscularize the injured or diseased heart, with several clinical trials now in planning or recruitment stages. However, because ventricular arrhythmias represent a complication following engraftment of intramyocardially injected PSC-CMs, it is necessary to provide treatment strategies to control or prevent engraftment arrhythmias (EAs). Here, we show in a porcine model of myocardial infarction and PSC-CM transplantation that EAs are mechanistically linked to cellular heterogeneity in the input PSC-CM and resultant graft. Specifically, we identify atrial and pacemaker-like cardiomyocytes as culprit arrhythmogenic subpopulations. Two unique surface marker signatures, signal regulatory protein α (SIRPA)+CD90−CD200+ and SIRPA+CD90−CD200−, identify arrhythmogenic and non-arrhythmogenic cardiomyocytes, respectively. Our data suggest that modifications to current PSC-CM-production and/or PSC-CM-selection protocols could potentially prevent EAs. We further show that pharmacologic and interventional anti-arrhythmic strategies can control and potentially abolish these arrhythmias

Gap junction intercellular communication: A review of a potential platform to modulate craniofacial tissue engineering

Defects in craniofacial tissues, resulting from trauma, congenital abnormalities, oncologic resection or progressive deforming diseases, may result in aesthetic deformity, pain and reduced function. Restoring the structure, function and aesthetics of craniofacial tissues represents a substantial clinical problem in need of new solutions. More biologically-interactive biomaterials could potentially improve the treatment of craniofacial defects, and an understanding of developmental processes may help identify strategies and materials that can be used in tissue engineering. One such strategy that can potentially advance tissue engineering is cell–cell communication. Gap junction intercellular communication is the most direct way of achieving such signaling. Gap junction communication through connexin-mediated junctions, in particular connexin 43 (Cx43), plays a major role bone development. Given the important role of Cx43 in controlling development and differentiation, especially in bone cells, controlling the expression of Cx43 may provide control over cell-to-cell communication and may help overcome some of the challenges in craniofacial tissue engineering. Following a review of gap junctions in bone cells, the ability to enhance cell–cell communication and osteogenic differentiation via control of gap junctions is discussed, as is the potential utility of this approach in craniofacial tissue engineering. © 2008 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2009Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/61547/1/31127_ftp.pd

Myoblast sensitivity and fibroblast insensitivity to osteogenic conversion by BMP-2 correlates with the expression of Bmpr-1a

<p>Abstract</p> <p>Background</p> <p>Osteoblasts are considered to primarily arise from osseous progenitors within the periosteum or bone marrow. We have speculated that cells from local soft tissues may also take on an osteogenic phenotype. Myoblasts are known to adopt a bone gene program upon treatment with the osteogenic bone morphogenetic proteins (BMP-2,-4,-6,-7,-9), but their osteogenic capacity relative to other progenitor types is unclear. We further hypothesized that the sensitivity of cells to BMP-2 would correlate with BMP receptor expression.</p> <p>Methods</p> <p>We directly compared the BMP-2 sensitivity of myoblastic murine cell lines and primary cells with osteoprogenitors from osseous tissues and fibroblasts. Fibroblasts forced to undergo myogenic conversion by transduction with a MyoD-expressing lentiviral vector (LV-MyoD) were also examined. Outcome measures included alkaline phosphatase expression, matrix mineralization, and expression of osteogenic genes <it>(alkaline phosphatase, osteocalcin </it>and <it>bone morphogenetic protein receptor-1A) </it>as measured by quantitative PCR.</p> <p>Results</p> <p>BMP-2 induced a rapid and robust osteogenic response in myoblasts and osteoprogenitors, but not in fibroblasts. Myoblasts and osteoprogenitors grown in osteogenic media rapidly upregulated <it>Bmpr-1a </it>expression. Chronic BMP-2 treatment resulted in peak <it>Bmpr-1a </it>expression at day 6 before declining, suggestive of a negative feedback mechanism. In contrast, fibroblasts expressed low levels of <it>Bmpr-1a </it>that was only weakly up-regulated by BMP-2 treatment. Bioinformatics analysis confirmed the presence of myogenic responsive elements in the proximal promoter region of human and murine <it>BMPR-1A/Bmpr-1a</it>. Forced myogenic gene expression in fibroblasts was associated with a significant increase in <it>Bmpr-1a </it>expression and a synergistic increase in the osteogenic response to BMP-2.</p> <p>Conclusion</p> <p>These data demonstrate the osteogenic sensitivity of muscle progenitors and provide a mechanistic insight into the variable response of different cell lineages to BMP-2.</p

Vascularized Cardiac Spheroids as Novel 3D in vitro Models to Study Cardiac Fibrosis.

Spheroid cultures are among the most explored cellular biomaterials used in cardiovascular research, due to their improved integration of biochemical and physiological features of the heart in a defined architectural three-dimensional microenvironment when compared to monolayer cultures. To further explore the potential use of spheroid cultures for research, we engineered a novel in vitro model of the heart with vascularized cardiac spheroids (VCSs), by coculturing cardiac myocytes, endothelial cells, and fibroblasts isolated from dissociated rat neonatal hearts (aged 1-3 days) in hanging drop cultures. To evaluate the validity of VCSs in recapitulating pathophysiological processes typical of the in vivo heart, such as cardiac fibrosis, we then treated VCSs with transforming growth factor beta 1 (TGFβ1), a known profibrotic agent. Our mRNA analysis demonstrated that TGFβ1-treated VCSs present elevated levels of expression of connective tissue growth factor, fibronectin, and TGFβ1 when compared to control cultures. We demonstrated a dramatic increase in collagen deposition following TGFβ1 treatment in VCSs in the PicroSirius Red-stained sections. Doxorubicin, a renowned cardiotoxic and profibrotic agent, triggered apoptosis and disrupted vascular networks in VCSs. Taken together, our findings demonstrate that VCSs are a valid model for the study of the mechanisms involved in cardiac fibrosis, with the potential to be used to investigate novel mechanisms and therapeutics for treating and preventing cardiac fibrosis in vitro

Promoter Optimization Circumvents Bcl-2 Transgene-Mediated Suppression of Lentiviral Vector Production

Lentiviral vectors are a robust gene delivery tool for inducing transgene expression in a variety of cells. They are well suited to facilitate the testing of therapeutic candidate genes in vitro, due to relative ease of packaging and ability to transduce dividing and non-dividing cells. Our goal was to identify a gene that could be delivered to the heart to protect against cancer-therapy-induced cardiotoxicity. We sought to generate a lentivirus construct with a ubiquitous CMV promoter driving expression of B-cell lymphocyte/leukemia 2 gene (Bcl-2), a potent anti-apoptotic gene. Contrary to our aim, overexpression of Bcl-2 induced cell death in the producer HEK293T cells, resulting in failure to produce usable vector titre. This was circumvented by exchanging the CMV promoter to the cardiac-specific NCX1 promoter, leading to the successful production of a lentiviral vector which could induce cardioprotective expression of Bcl-2. In conclusion, reduced expression of Bcl-2 driven by a weaker promoter improved vector yield, and led to the production of functional cardioprotective Bcl-2 in primary cardiomyocytes

Development of a sheep model of atrioventricular block for the application of novel therapies.

INTRODUCTION:Sheep have been adopted as a pre-clinical large animal for scientific research as they are good models of cardiac anatomy and physiology, and allow for investigation of pathophysiological processes which occur in the large mammalian heart. There is, however, no defined model of atrioventricular block in sheep to allow for pre-clinical assessment of new cardiac treatment options. We therefore aimed to develop an adult sheep model of atrioventricular block with the focus on future novel applications. METHODS AND RESULTS:We utilized six sheep to undergo two procedures each. The first procedure involved implantation of a single chamber pacemaker into the right ventricular apex, for baseline assessment over four weeks. The second procedure involved creating atrioventricular block by radiofrequency ablation of the His bundle, before holding for a further four weeks. Interrogation of pacemakers and electrocardiograms determined the persistence of atrioventricular block during the follow up period. Pacemakers were inserted, and atrioventricular block created in 6 animals using a conventional approach. One animal died following ablation of the His bundle, due to procedural complications. Four unablated sheep were assessed for baseline data over four weeks and showed 5.53 ± 1.28% pacing reliance. Five sheep were assessed over four weeks following His bundle ablation and showed continuous (98.89 ± 0.81%) ventricular pacing attributable to persistent atrioventricular block, with no major complications. CONCLUSION:We have successfully developed, characterized and validated a large animal model of atrioventricular block that is stable and technically feasible in adult sheep. This model will allow for the advancement of novel therapies, including the development of cell and gene-based therapies