Cardiac stem cells transplantation enhances the expression of connexin 43 via the ANG II/AT1R/TGF-beta1 signaling pathway in a rat model of myocardial infarction☆
Introduction
Myocardial infarction (MI) with resultant congestive heart failure and malignant arrhythmia continues to be a major cause of death and disability worldwide (Mozaffarian et al., 2015). Various approaches have been taken to improve the survival of patients with MI over the past decade, however, the morbidity and mortality remain significant (Hausenloy et al., 2013). In recent years, cell therapy has emerged as a potentially novel strategy for the treatment of MI. Series of preclinical and clinical studies have displayed the benefits of cell interventions in MI (Sanganalmath and Bolli, 2013, Pavo et al., 2014). Substantial evidence has already revealed that stem cell transplantation can reduce infarct size, preserve cardiac function and improve clinical outcomes in patients with MI (Schutt et al., 2015, Heldman et al., 2014). Several types of stem cells, such as embryonic stem cells (ESCs), bone marrow stem cells (BMSCs) and induced pluripotent stem cells (iPSCs) have been used in the therapy studies of MI. In spite of this, the adverse effects and ethical issues brought about by them have limited their further clinical applications (Hou et al., 2013).
Cardiac stem cells (CSCs) represent a logical candidate for cardiac regeneration therapy after MI. they participate in the turnover of myocytes in the absence of pathological state and they are intrinsically programmed to form cardiac tissues rapidly upon activation (Hou et al., 2013, Bergmann et al., 2009, Davis et al., 2010). Our previous work has demonstrated that CSCs transplantation can improve the electrophysiological stability in MI (Zheng et al., 2011). One further study shows that CSCs are superior to MSCs in modulating the electrophysiological abnormality and improving the VFT in rats with MI (Zheng et al., 2013). They can express connexin 43 (Cx43) in the infarct zone and border zone after cells implantation. However, it is still unknown how CSCs may regulate the expression of Cx43 in the procedure.
Cx43 is, by far, the most predominant type of gap junction proteins expressed in the conductive and working myocardial cells (Akar et al., 2004). The downregulation or heterogeneous redistribution of Cx43 causes the structural and electrical changes that impair electrical stability (Kim and Fishman, 2013). The expression and remodeling of Cx43 post MI is dominated by numerous factors. Specific molecular mechanisms involved are intricate and not completely understood.
The renin-angiotensin system (RAS) exerts a pivotal role in the pathogenesis of myocardial infarction (MI) (Morales-Suárez-Varela et al., 2011). Angiotensin II (ANG II) is a key component of the RAS and has been implicated in the development of various cardiovascular ailments including cardiac hypertrophy, remodeling and heart failure (Lang and Struthers, 2013). Most of the classical and biological functions of angiotensin II are mediated by the angiotensin II type 1 receptor (AT1R), whereas the angiotensin II type 1 receptor (AT2R) plays a counter-regulatory role. Blockade of AT1R has been reported to reduce the incidence of ventricular arrhythmias (Jiao et al., 2012). Accumulating evidence has indicated that the activation of ANG II is closely linked with the loss of Cx43 in vivo (Hussain et al., 2010, Yasuno et al., 2013).
ANG II performs its functions via activating several signaling pathways. TGF-β1 is a crucial downstream molecule of ANG II. It mediates a variety of biological processes such as cell proliferation, immune suppression, and inflammation (Shi and Massagué, 2003). It is also vital in the pathogenesis of infarct healing, cardiac remodeling, and interstitial fibrosis (Vivar et al., 2013). TGF-β1 expression has been shown to be markedly increased in experimental models of myocardial infarction and in patients with cardiomyopathy (Dobaczewski et al., 2011). Recent studies manifest that TGF-β1 modulates the formation of Cx43 (Neuhaus et al., 2009). Nevertheless, relevant signaling pathways involved are still undetermined.
In this study, we hypothesized that CSCs transplantation promoted the expression of Cx43 via the inhibition of ANGII and its downstream signaling molecules including AT1R and TGFβ1 signaling pathways after MI.
Section snippets
Ethics statement
Twenty adult male Sprague–Dawley (SD) rats weighing 250 g–350 g were obtained from the Animal Experimental Center of the Sun Yat-sen University and housed in a standard animal facility with 12-hour on/off light conditions. All animals were acclimatized for at least one week prior to surgery and allowed free access to standard food and water. All animal handling and procedures were performed in accordance with protocols approved by the Animal Ethics Committee of Sun Yat-sen University
CSCs transplantation enhanced the expression of Cx43
CSCs marked with PHK26 were found to be surviving in the infarct zone and border zone and expressed Cx43 by immunofluorescence staining (Fig.1A, c and e), which was accorded with the results of our previous work. In the CSCs group, the expression of Cx43 was detected in different zones of the left ventricle (Fig.1A; a, c and e) whereas the PBS group rarely expressed Cx43 (Fig.1A, b, d and f). Western blot analysis and qRT-PCR showed that the expression of Cx43 was distinctly increased not only
Discussion
The present study revealed that CSCs transplantation could enhance the expression of Cx43 after MI. They could probably function through the blockade of ANGII/AT1R/TGF-β1 signaling pathway during the process.
Malignant ventricular arrhythmia is the main cause of sudden cardiac death following MI (Ersbøll et al., 2013). Over the past half-century, remarkable progresses in pharmacological and interventional therapeutic approaches have improved the survival and prognosis of patients with MI (
Conclusions
CSCs transplantation enhanced the expression of Cx43. They might function via ANGII/AT1/TGF-β1 signaling pathway to regulate the expression of Cx43.
Competing interests
The authors declare that there are no competing interests.
Authors' contributions
Dr. Jingying Hou and Dr. Ping Yan carried out the experiments and drafted the manuscript; Yue Xing, Tianzhu Guo, Shaoxin Zheng and Changqing Zhou participated in the preparation of the animal model, tissue staining, and molecular assay; Hui Huang and Jingfeng Wang provided the technical assistance; Huibao Long, Tingting Zhong and Quanhua Wu participated and statistical analysis; Tong Wang conceived the study and participated in the study design. All authors read and approved the final
Acknowledgments
This study was supported by Grant [2013]163 from Key Laboratory of Malignant Tumor Molecular Mechanism and Translational Medicine of Guangzhou Bureau of Science and Information Technology; Grant KLB09001 from the Key Laboratory of Malignant Tumor Gene Regulation and Target Therapy of Guangdong Higher Education Institutes; National Natural Science Foundation of China (Nos. 81070125, 81270213), Science and Technology Foundation in Guangdong Province (No: 2010B031600032, 2014A020211002) and the
References (62)
- et al.
Cardiac stem cells in patients with ischaemic cardiomyopathy (SCIPIO): initial results of a randomised phase 1 trial
Lancet
(2011) - et al.
Isolation and expansion of functionally-competent cardiac progenitor cells directly from heart
J. Mol. Cell. Cardiol.
(2010) - et al.
Transforming growth factor (TGF)-β signaling in cardiac remodeling
J. Mol. Cell. Cardiol.
(2011) - et al.
Early echocardiographic deformation analysis for the prediction of sudden cardiac death and life-threatening arrhythmias after myocardial infarction
JACC Cardiovasc. Imaging
(2013) - et al.
Mesenchymal stem cells attenuate angiotensin II-induced aortic aneurysm growth in apolipoprotein E-deficient mice
J. Vasc. Surg.
(2011) - et al.
Meta-analysis of stem cell therapy in chronic ischemic cardiomyopathy
Am. J. Cardiol.
(2013) - et al.
Designer gap junctions that prevent cardiac arrhythmias
Trends Cardiovasc. Med.
(2013) - et al.
The clinical application of mesenchymal and cardiac stem cells as a therapy for cardiovascular disease
Pharmacol. Ther.
(2015) - et al.
In vitro and in vivo characteristics of connexin 43-modified human skeletal myoblasts as candidates for prospective stem cell therapy for the failing heart
Int. J. Cardiol.
(2014) - et al.
Angiotensin II upregulates Kv1.5 expression through ROS-dependent transforming growth factor-beta1 and extracellular signal-regulated kinase 1/2 signalings in neonatal rat atrial myocytes
Biochem Biophys Res Commun
(2014)
Pirfenidone mitigates left ventricular fibrosis and dysfunction after myocardial infarction and reduces arrhythmias
Heart Rhythm
AT2 receptor mediates the cardioprotective effects of AT1 receptor antagonist in post-myocardial infarction remodeling
Life Sci.
Cell therapy for human ischemic heart diseases: critical review and summary of the clinical experiences
J. Mol. Cell. Cardiol.
Mechanisms of TGF-beta signaling from cell membrane to the nucleus
Cell
Dudley SC. Inhibition of c-Src tyrosine kinase prevents angiotensin II-mediated connexin-43 remodeling and sudden cardiac death
J. Am. Coll. Cardiol.
Engraftment of human embryonic stem cell derived cardiomyocytes improves conduction in an arrhythmogenic in vitro model
J. Mol. Cell. Cardiol.
A novel high throughput approach to screen for cardiac arrhythmic events following stem cell treatment
Med. Hypotheses
Effect of human umbilical cord blood cells on Ang-II-induced hypertrophy in mice
Biochem. Biophys. Res. Commun.
TGF-β1 prevents simulated ischemia/reperfusion-induced cardiac fibroblast apoptosis by activation of both canonical and non-canonical signaling pathways
Biochim. Biophys. Acta
Down-regulation of connexin43 gap junction by serum deprivation in human endothelial cells was improved by (−)-epigallocatechin gallate via ERK MAP kinase pathway
Biochem. Biophys. Res. Commun.
Fibrosis-related biomarkers and incident cardiovascular disease in older adults: the cardiovascular health study
Am. J. Epidemiol.
Mechanisms underlying conduction slowing and arrhythmogenesis in nonischemic dilated cardiomyopathy
Circ. Res.
Cardiac c-kit + AT2 + cell population is increased in response to ischemic injury and supports cardiomyocyte performance
Stem Cells
Evidence for cardiomyocyte renewal in humans
Science
Growth factor-induced mobilization of cardiac progenitor cells reduces the risk of arrhythmias, in a rat model of chronic myocardial infarction
PLoS ONE
Intracoronary delivery of autologous cardiac stem cells improves cardiac function in a porcine model of chronic ischemic cardiomyopathy
Circulation
Electrophysiological challenges of cell based myocardial repair
Circulation
Cx43 mediates TGF-beta signaling through competitive SMADs binding to microtubules
Mol. Biol. Cell
Insulin-like growth factor-1 receptor identifies a pool of human cardiac stem cells with superior therapeutic potential for myocardial regeneration
Circ Res
Gualou Xiebai decoction prevents myocardial fibrosis by blocking TGF-beta/Smad signalling
J. Pharm. Pharmacol.
Meta-analysis of cell therapy trials for patients with heart failure
Circ. Res.
Cited by (11)
Long noncoding RNAs: Novel molecules in cardiovascular biology, disease and regeneration
2016, Experimental and Molecular PathologyCitation Excerpt :Previous studies have revealed that implantation of these cells results in cardiomyocytes differentiation and neovascularization. ( Carvalho et al., 2015; Hou et al., 2015). However, optimizing their differentiation into desired cell types for cardiac repair may be a tough task due to their pluripotency and unique origins.
Experimental induction of reparative morphogenesis and adaptive reserves in the ischemic myocardium using multipotent mesenchymal bone marrow-derived stem cells
2016, PathophysiologyCitation Excerpt :We used an aliquot of 1–5 × 106 cells, which was applied to the MCS transplantation, as this is the amount recommended as ideal for experimental and clinical research [14,29,30]. Clinical investigations have shown that intravenous injection of 1–5 × 106 MCS (isolated from bone marrow aspirate and after 5–6 passages in culture) effectively replaces damaged non-hematopoietic tissue function (myocardium, skeletal muscles, nervous tissue) without complications or side effects [31–33]. After administration of narcotics, the animal was fixed on its back.
Activin receptor-like kinase 4 haplodeficiency mitigates arrhythmogenic atrial remodeling and vulnerability to atrial fibrillation in cardiac pathological hypertrophy
2018, Journal of the American Heart AssociationMetoprolol protects cardiomyocytes in rabbit model of heart failure by regulating cx43
2018, Experimental and Therapeutic MedicineThe research progress on treatment of myocardial fibrosis by regulating TGF-pl/Smads pathway
2018, Chinese Pharmacological Bulletin
- ☆
This work was done by the investigators of the Sun Yat-sen Memorial Hospital of Sun Yat-sen University. The authors took responsibility for all aspects of the reliability and had no differences in data presentation and interpretation.
- 1
Dr. Jingying Hou and Dr. Ping Yan played equally important roles in the development of the experimental protocol, in the interpretation of the results, and in the texture of the present article.