José Anastasio Montero

Human Dental Pulp Stem Cells Improve Left Ventricular Function, Induce Angiogenesis, and Reduce Infarct Size in Rats with Acute Myocardial Infarction

Human dental pulp contains precursor cells termed dental pulp stem cells (DPSC) that show self‐renewal and multilineage differentiation and also secrete multiple proangiogenic and antiapoptotic factors. To examine whether these cells could have therapeutic potential in the repair of myocardial infarction (MI), DPSC were infected with a retrovirus encoding the green fluorescent protein (GFP) and expanded ex vivo. Seven days after induction of myocardial infarction by coronary artery ligation, 1.5 × 106 GFP‐DPSC were injected intramyocardially in nude rats. At 4 weeks, cell‐treated animals showed an improvement in cardiac function, observed by percentage changes in anterior wall thickening left ventricular fractional area change, in parallel with a reduction in infarct size. No histologic evidence was seen of GFP+ endothelial cells, smooth muscle cells, or cardiac muscle cells within the infarct. However, angiogenesis was increased relative to control‐treated animals. Taken together, these data suggest that DPSC could provide a novel alternative cell population for cardiac repair, at least in the setting of acute MI.

Cardiac Differentiation Is Driven by NKX2.5 and GATA4 Nuclear Translocation in Tissue-Specific Mesenchymal Stem Cells

Myocardial infarction is a major public health problem that causes significant mortality despite recent advances in its prevention and treatment. Therefore, approaches based on adult stem cells represent a promising alternative to conventional therapies for this life-threatening condition. Mesenchymal stem cells (MSCs) are self-renewing pluripotent cells that have been isolated from multiple tissues and differentiate to various cell types. Here we have analyzed the capacity of MSCs from human bone marrow (BMSC), adipose tissue (ATSC), and dental pulp (DPSC) to differentiate to cells with a cardiac phenotype. Differentiation of MSCs was induced by long-term co-culture with neonatal rat cardiomyocytes (CMs). Shortly after the establishment of MSC-CM co-cultures, expression of connexin 43 and the cardiac-specific markers troponin I, beta-myosin heavy chain, atrial natriuretic peptide, and alpha-sarcomeric actinin was detected in BMSCs, ATSCs, and DPSCs. Expression of differentiation markers increased over time in the co-cultures, reaching the highest levels at 4 weeks. Translocation of the transcription factors NKX2.5 and GATA4 to the nucleus was observed in all three cultures of MSCs during the differentiation process; moreover, nuclear localization of NKX2.5 and GATA4 correlated with expression of alpha-sarcomeric actinin. These changes were accompanied by an increase in myofibril organization in the resulting CM-like cells as analyzed by electron microscopy. Thus, our results provide novel information regarding the differentiation of tissue-specific MSCs to cardiomyocytes and support the potential use of MSCs in cell-based cardiac therapies.

Human progenitor cells derived from cardiac adipose tissue ameliorate myocardial infarction in rodents.

Myocardial infarction caused by vascular occlusion results in the formation of nonfunctional fibrous tissue. Cumulative evidence indicates that cell therapy modestly improves cardiac function; thus, novel cell sources with the potential to repair injured tissue are actively sought. Here, we identify and characterize a cell population of cardiac adipose tissue-derived progenitor cells (ATDPCs) from biopsies of human adult cardiac adipose tissue. Cardiac ATDPCs express a mesenchymal stem cell-like marker profile (strongly positive for CD105, CD44, CD166, CD29 and CD90) and have immunosuppressive capacity. Moreover, cardiac ATDPCs have an inherent cardiac-like phenotype and were able to express de novo myocardial and endothelial markers in vitro but not to differentiate into adipocytes. In addition, when cardiac ATDPCs were transplanted into injured myocardium in mouse and rat models of myocardial infarction, the engrafted cells expressed cardiac (troponin I, sarcomeric α-actinin) and endothelial (CD31) markers, vascularization increased, and infarct size was reduced in mice and rats. Moreover, significant differences between control and cell-treated groups were found in fractional shortening and ejection fraction, and the anterior wall remained significantly thicker 30days after cardiac delivery of ATDPCs. Finally, cardiac ATDPCs secreted proangiogenic factors under in vitro hypoxic conditions, suggesting a paracrine effect to promote local vascularization. Our results indicate that the population of progenitor cells isolated from human cardiac adipose tissue (cardiac ATDPCs) may be valid candidates for future use in cell therapy to regenerate injured myocardium.

IL1β Induces Mesenchymal Stem Cells Migration and Leucocyte Chemotaxis Through NF-κB

Mesenchymal stem cells are often transplanted into inflammatory environments where they are able to survive and modulate host immune responses through a poorly understood mechanism. In this paper we analyzed the responses of MSC to IL-1β: a representative inflammatory mediator. Microarray analysis of MSC treated with IL-1β revealed that this cytokine activateds a set of genes related to biological processes such as cell survival, cell migration, cell adhesion, chemokine production, induction of angiogenesis and modulation of the immune response. Further more detailed analysis by real-time PCR and functional assays revealed that IL-1β mainly increaseds the production of chemokines such as CCL5, CCL20, CXCL1, CXCL3, CXCL5, CXCL6, CXCL10, CXCL11 and CX3CL1, interleukins IL-6, IL-8, IL23A, IL32, Toll-like receptors TLR2, TLR4, CLDN1, metalloproteins MMP1 and MMP3, growth factors CSF2 and TNF-α, together with adhesion molecules ICAM1 and ICAM4. Functional analysis of MSC proliferation, migration and adhesion to extracellular matrix components revealed that IL-1β did not affect proliferation but also served to induce the secretion of trophic factors and adhesion to ECM components such as collagen and laminin. IL-1β treatment enhanced the ability of MSC to recruit monocytes and granulocytes in vitro. Blockade of NF-κβ transcription factor activation with IκB kinase beta (IKKβ) shRNA impaired MSC migration, adhesion and leucocyte recruitment, induced by IL-1β demonstrating that NF-κB pathway is an important downstream regulator of these responses. These findings are relevant to understanding the biological responses of MSC to inflammatory environments.

Midterm Clinical and Echocardiographic Results With Patch Glue Repair of Left Ventricular Free Wall Rupture

Background—Left ventricular free wall rupture (LVFWR) is a dramatic complication after myocardial infarction. We present our mid-term clinical and echocardiographic results of LVFWR with an epicardial patch without cardiopulmonary bypass. Methods—From February 1993 to May 2001, 17 patients underwent surgery for LVFWR. The mean age±SD of 12 males and 5 females was 68±10 years. All patients presented for emergency surgery with cardiac tamponade confirmed on echocardiography. After opening the chest and identification of the site of rupture, a Goretex patch was fashioned and applied with enbucrilate surgical glue. Results—Effective control of bleeding was achieved in all cases. There were no on-table deaths. The operative (30 day) mortality was 23.5% (4/17). One death occurred because of patch failure, two because of cardiogenic shock, and one from pneumonia. On follow-up at a median of 2.2 years (interquartile range, 1.1 to 4.3 years), two further deaths occurred, one from myocardial infarction and another of undetermined etiology. Echocardiography did not reveal any evidence of restriction to left ventricular free wall motion. Conclusions—Patch glue repair is expedient, simple and effective; with no adverse effects on mid-term ventricular dynamics. In view of superior published results to infarctectomy and repair with extra corporeal circulation, it should be considered to be the initial procedure of choice for the surgical repair of LVFWR.

Hypoxia-inducible factor 1 alpha contributes to cardiac healing in mesenchymal stem cells-mediated cardiac repair

Mesenchymal stem cells (MSC) are effective in treating myocardial infarction (MI) and previous reports demonstrated that hypoxia improves MSC self-renewal and therapeutics. Considering that hypoxia-inducible factor-1 alpha (HIF-1α) is a master regulator of the adaptative response to hypoxia, we hypothesized that HIF-1α overexpression in MSC could mimic some of the mechanisms triggered by hypoxia and increase their therapeutic potential without hypoxia stimulation. Transduction of MSC with HIF-1α lentivirus vectors (MSC-HIF) resulted in increased cell adhesion and migration, and activation of target genes coding for paracrine factors. When MSC-HIF were intramyocardially injected in infarcted nude rats, significant improvement was found (after treatment of infarcted rats with MSC-HIF) in terms of cardiac function, angiogenesis, cardiomyocyte proliferation, and reduction of fibrotic tissue with no induction of cardiac hypertrophy. This finding provides evidences for a crucial role of HIF-1α on MSC biology and suggests the stabilization of HIF-1α as a novel strategy for cellular therapies.

Cardiac Cell Therapy: Boosting Mesenchymal Stem Cells Effects

Acute myocardial infarction is a major problem of world public health and available treatments have limited efficacy. Cardiac cell therapy is a new therapeutic strategy focused on regeneration and repair of the injured cardiac muscle. Among different cell types used, mesenchymal stem cells (MSC) have been widely tested in preclinical studies and several clinical trials have evaluated their clinical efficacy in myocardial infarction. However, the beneficial effects of MSC in humans are limited due to poor engraftment and survival of these cells, therefore ways to overcome these obstacles should improve efficacy. Different strategies have been used, such as genetically modifying MSC, or preconditioning the cells with factors that potentiate their survival and therapeutic mechanisms. In this review we compile the most relevant approaches used to improve MSC therapeutic capacity and to understand the molecular mechanisms involved in MSC mediated cardiac repair.

Hypoxia Inducible Factor‐1α Potentiates Jagged 1‐Mediated Angiogenesis by Mesenchymal Stem Cell‐Derived Exosomes

Insufficient vessel growth associated with ischemia remains an unresolved issue in vascular medicine. Mesenchymal stem cells (MSCs) have been shown to promote angiogenesis via a mechanism that is potentiated by hypoxia. Overexpression of hypoxia inducible factor (HIF)‐1α in MSCs improves their therapeutic potential by inducing angiogenesis in transplanted tissues. Here, we studied the contribution of exosomes released by HIF‐1α‐overexpressing donor MSCs (HIF‐MSC) to angiogenesis by endothelial cells. Exosome secretion was enhanced in HIF‐MSC. Omics analysis of miRNAs and proteins incorporated into exosomes pointed to the Notch pathway as a candidate mediator of exosome communication. Interestingly, we found that Jagged1 was the sole Notch ligand packaged into MSC exosomes and was more abundant in HIF‐MSC than in MSC controls. The addition of Jagged1‐containing exosomes from MSC and HIF‐MSC cultures to endothelial cells triggered transcriptional changes in Notch target genes and induced angiogenesis in an in vitro model of capillary‐like tube formation, and both processes were stimulated by HIF‐1α. Finally, subcutaneous injection of Jagged 1‐containing exosomes from MSC and HIF‐MSC cultures in the Matrigel plug assay induced angiogenesis in vivo, which was more robust when they were derived from HIF‐MSC cultures. All Jagged1‐mediated effects could be blocked by prior incubation of exosomes with an anti‐Jagged 1 antibody. All together, the results indicate that exosomes derived from MSCs stably overexpressing HIF‐1α have an increased angiogenic capacity in part via an increase in the packaging of Jagged1, which could have potential applications for the treatment of ischemia‐related disease. Stem Cells 2017;35:1747–1759

Surgical ablation of permanent atrial fibrillation by means of maze radiofrequency: mid-term results.

Abstract  Background: The maze procedure can be performed surgically with radiofrequency, generating transmural ablation lines. We report our experience with a biatrial pattern of lesions based on the use of epicardial and endocardial radiofrequency ablation in an effort to minimize maze procedure. Method: In 85 patients undergoing cardiac surgery for established permanent atrial fibrillation (>3 months), a biauricular pattern of epicardic–endocardic maze lesions was performed. The main surgical procedures were diverse: 42 mitral valve surgeries, 7 mitrotricuspid valves, 18 mitroaortics, 4 mitroaortic and tricuspids, 2 aortic valves, 3 CABGs, 5 CABG and valve procedures, and 4 atrial septal defects. The mean age of the patients was 61 ± 12 (range 39–78). The mean duration of atrial fibrillation was 5.8 years (range 0.3 to 24). Results: Sixty‐two (72.9%) patients presented postoperative supraventricular arrhythmia. Hospital mortality was seen in five patients (5.8%). Two patients died after a 12‐month mean follow‐up (range 2 to 32). A total of 14.1% of patients remained with their previous atrial fibrillation and 85.9% recovered and maintained sinus rhythm, with two patients having a permanent pacemaker. A total of 56% patients have been followed‐up for a period of more than 6 months, and among them prevalence of sinus rhythm is 87.5%. Echocardiography detected biauricular contraction in 65% of them. After analyzing the data, factors involved in postoperative recurrence of atrial fibrillation after radiofrequency surgery were oldness of the atrial fibrillation (p < 0.01) and pre and postoperative left auricle volume (p < 0.04). Conclusion: Intraoperative radiofrequency has permitted us to perform the maze procedure in a simple way, with a low surgical morbid‐mortality. We have obtained an 85.9% electrographic effectiveness and a 65% recovery of atrial contraction. Postoperative incidence of arrhythmia is the main postoperative problem.

Cardiac Transcription Factors Driven Lineage-Specification of Adult Stem Cells

Differentiation of human bone marrow mesenchymal stem cells (hBMSC) into the cardiac lineage has been assayed using different approaches such as coculture with cardiac or embryonic cells, treatment with factors, or by seeding cells in organotypic cultures. In most cases, differentiation was evaluated in terms of expression of cardiac-specific markers at protein or molecular level, electrophysiological properties, and formation of sarcomers in differentiated cells. As observed in embryonic stem cells and cardiac progenitors, differentiation of MSC towards the cardiac lineage was preceded by translocation of NKX2.5 and GATA4 transcription factors to the nucleus. Here, we induce differentiation of hBMSC towards the cardiac lineage using coculture with neonatal rat cardiomyocytes. Although important ultrastructural changes occurred during the course of differentiation, sarcomerogenesis was not fully achieved even after long periods of time. Nevertheless, we show that the main cardiac markers, NKX2.5 and GATA4, translocate to the nucleus in a process characteristic of cardiac specification.