ORCID Profile
0000-0002-7266-6720
Current Organisations
University of Sydney
,
Westmead Hospital
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Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 25-03-2008
DOI: 10.1161/CIRCULATIONAHA.107.732073
Abstract: Background— Stem cell labeling with iron oxide (ferumoxide) particles allows labeled cells to be detected by magnetic resonance imaging (MRI) and is commonly used to track stem cell engraftment. However, the validity of MRI for distinguishing surviving ferumoxide-labeled cells from other sources of MRI signal, for ex le, macrophages containing ferumoxides released from nonsurviving cells, has not been thoroughly investigated. We sought to determine the relationship between the persistence of iron-dependent MRI signals and cell survival 3 weeks after injection of syngeneic or xenogeneic ferumoxides-labeled stem cells (cardiac-derived stem cells) in rats. Methods and Results— We studied nonimmunoprivileged human and rat cardiac-derived stem cells and human mesenchymal stem cells doubly labeled with ferumoxides and β-galactosidase and injected intramyocardially into immunocompetent Wistar-Kyoto rats. Animals were imaged at 2 days and 3 weeks after stem cell injection in a clinical 3-T MRI scanner. At 2 days, injection sites of xenogeneic and syngeneic cells (cardiac-derived stem cells and mesenchymal stem cells) were identified by MRI as large intramyocardial signal voids that persisted at 3 weeks (50% to 90% of initial signal). Histology (at 3 weeks) revealed the presence of iron-containing macrophages at the injection site, identified by CD68 staining, but very few or no β-galactosidase–positive stem cells in the animals transplanted with syngeneic or xenogeneic cells, respectively. Conclusions— The persistence of significant iron-dependent MRI signal derived from ferumoxide-containing macrophages despite few or no viable stem cells 3 weeks after transplantation indicates that MRI of ferumoxide-labeled cells does not reliably report long-term stem cell engraftment in the heart.
Publisher: Wiley
Date: 06-2020
DOI: 10.1111/IMJ.14416
Abstract: Cardioembolism (CE) contributes to a large proportion of ischaemic stroke. To evaluate the demographic and clinical profile of CE stroke in Western Sydney. A retrospective analysis of ischaemic stroke patients presenting to Westmead Hospital (January-October 2016) was performed. Strokes were classified by TOAST (Trial of Org 10172 in Acute Stroke Treatment) criteria into different categories. Clinical and demographic data were collected on all stroke patients, and differences between CE and other stroke causes were identified. Two hundred and twenty-eight consecutive patients (70.9 years 53% male) were identified. By TOAST criteria, 21 (9%) had large-artery atherosclerosis, 94 (41%) CE, 10 (5%) small-vessel disease, 2 (1%) other aetiology and 101 (44%) undetermined aetiology. A significant proportion of CE stroke patients had cardiovascular risk factors including hypertension (66%), hypercholesterolaemia (50%), diabetes (26%) and ischaemic heart disease (28%). The majority (81%) of patients with CE had atrial flutter/flutter. CE stroke, compared with other types of stroke, was more common in females (56 vs 41%, P = 0.022) and patients with CE stroke were more likely to have previous cerebral ischaemia (34 vs 21%, P = 0.026), suggesting increased recurrence in this group. Of the patients with atrial flutter/flutter (n = 56), the majority (87%) had a high CHA Cardioembolic stroke remains a significant burden in Western Sydney, and it is likely that a significant proportion may be preventable, as evidenced by the substantial presence of modifiable cardiovascular risk factors, and inadequate anticoagulation of patients with atrial arrhythmias.
Publisher: Wiley
Date: 31-05-2019
DOI: 10.1111/ENE.13981
Abstract: Ischaemic stroke frequently has a cardioembolic (CE) source. Clinical and echocardiographic parameters associated with CE stroke were evaluated. In all, 93 consecutive ischaemic stroke patients who underwent a transthoracic echocardiogram were retrospectively analysed strokes were classified by TOAST (Trial of Org 10172 in Acute Stroke Treatment) criteria. Echocardiographic parameters related to CE stroke, including left atrial volumes and function, were compared to 73 healthy controls. Of 93 patients (mean age 66.1 years, 56% male), nine (10%) had large artery atherosclerosis, 38 (41%) CE stroke, two (2%) small vessel disease, two (2%) other and 42 (45%) undetermined aetiology. Left atrial (LA) maximum volumes (LAVI Left atrial enlargement with reduced LA function was associated with CE stroke and LAFI was the best independent predictor. LA parameters were also altered in the undetermined aetiology group, suggesting an underlying LA myopathy in this subset.
Publisher: Elsevier BV
Date: 05-2009
Publisher: Elsevier BV
Date: 12-2021
Publisher: Springer Science and Business Media LLC
Date: 29-08-2019
DOI: 10.1038/S41434-019-0097-0
Abstract: Differences between mouse and human hearts pose a significant limitation to the value of small animal models when predicting vector behavior following recombinant adeno-associated viral (rAAV) vector-mediated cardiac gene therapy. Hence, sheep have been adopted as a preclinical animal, as they better model the anatomy and cardiac physiological processes of humans. There is, however, no comprehensive data on the shedding profile of rAAV in sheep following intracoronary delivery, so as to understand biosafety risks in future preclinical and clinical applications. In this study, sheep received intracoronary delivery of rAAV serotypes 2/6 (2 × 10
Publisher: Wiley
Date: 12-04-2021
DOI: 10.1111/JCE.15026
Publisher: MDPI AG
Date: 26-07-2022
DOI: 10.3390/V14081623
Abstract: Gene therapy is making significant impact on a modest, yet growing, number of human diseases. Justifiably, the preferred viral vector for clinical use is that based on recombinant adeno-associated virus (rAAV). There is a need to scale up rAAV vector production with the transition from pre-clinical models to human application. Standard production methods based on the adherent cell type (HEK293) are limited in scalability and other methods, such as those based on the baculovirus and non-adherent insect cell (Sf9) system, have been pursued as an alternative to increase rAAV production. In this study, we compare these two production methods for cardiotropic rAAVs. Transduction efficiency for both production methods was assessed in primary cardiomyocytes, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), and in mice following systemic delivery. We found that the rAAV produced by the traditional HEK293 method out-performed vector produced using the baculovirus/Sf9 system in vitro and in vivo. This finding provides a potential caveat for vector function when using the baculovirus/Sf9 production system and underscores the need for thorough assessment of vector performance when using erse rAAV production methods.
Publisher: Elsevier BV
Date: 10-2023
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 19-11-2013
DOI: 10.1161/CIRCULATIONAHA.113.002238
Abstract: Collagen has been attributed as the principal structural substrate of ventricular tachycardia (VT) after myocardial infarction (MI), even though adiposity of myocardium after MI is well recognized histologically. We investigated the effects of intramyocardial adiposity compared with collagen on electrophysiological properties, connexin43 expression, and VT induction after MI. Simultaneous left ventricular plunge-needle, noncontact mapping was performed in sheep without MI (MI− n=5), with MI and inducible VT (MI+VT+ n=7), and with MI and no inducible VT (MI+VT− n=8). Histological intramyocardial quantity of adipose and collagen and degree of discontinuity were coregistered with electrophysiological parameters (MI+ 290 specimens). Additional assessment of connexin43 expression was performed. Left ventricular scar contained a body mass–independent abundance of adipocytes (adipose:collagen=0.8). Increased adipose density and discontinuity contributed to a greater inverse correlation ( r ) with conduction velocity ( r for adipose=0.39, r for discontinuity=0.45, r for collagen=0.26) and electrogram litude ( r for adipose=0.73, r for contiguity=0.77, r for collagen=0.68) compared with collagen. Collagen density was similar between the MI+ groups ( P .29). However, the MI+VT+ group demonstrated a significant (all P ≤0.01) increase in adipose (8%) and discontinuity (qualitative) and decrease in conduction velocity (13%) and electrogram litude (21%) at MI borders compared with the MI+VT− group. In scar, myocytes adjacent to fibrofatty interfaces demonstrated increased connexin43 lateralization. A gradient increase in adipose was observed at sites that supported preferential presystolic VT activation and exhibited attenuation of excitation wavelength ( P .001). Intramyocardial adiposity, in association with myocardial discontinuity within left ventricular scar borders, is a significant factor associated with altered electrophysiological properties, aberrant connexin43 expression, and increased propensity for VT after MI.
Publisher: Elsevier BV
Date: 04-2015
DOI: 10.1016/J.IJCARD.2015.01.068
Abstract: Patients with left ventricular diastolic impairment (LVDI) have higher rates of arrhythmia recurrence following atrial fibrillation (AF) ablation. Past studies have implicated the posterior left atrium (LA) in atrial arrhythmia maintenance in conditions that cause LVDI. We prospectively compared posterior LA isolation (PLAI) with wide antral isolation (WAI) in patients with LVDI having AF ablation. We conducted a sub-study of a previously published large randomized control study that compared PLAI with WAI. Two hundred and twenty consecutive consenting patients referred for catheter ablation of AF (paroxysmal 135, persistent 48, long standing persistent 37) were recruited (female 43, mean age 59 ± 10 years). Transthoracic echocardiography identified 50 (23%) patients with LVDI and preserved left ventricular systolic function (ejection fraction ≥ 50%). Cox regression analysis was utilized to identify independent predictors of atrial arrhythmia after ablation. Patients were followed for median 4.6 (inter quartile range 4.0-5.5) years. Patients with LVDI having PLAI had better arrhythmia free survival than patients randomized to conventional ablation (Log rank P=0.028). The only independent predictor of recurrence utilizing Cox regression analysis was ablation strategy (2.3 [1.15-4.74], P=0.026). Posterior isolation of the LA results in superior atrial arrhythmia free survival in patients with LVDI. Further investigation is required to determine potential mechanisms. www.anzctr.org.au ACTRN12606000467538.
Publisher: Elsevier BV
Date: 11-2018
Publisher: Springer Science and Business Media LLC
Date: 22-07-2019
DOI: 10.1038/S41598-019-47022-W
Abstract: We have previously reported a subpopulation of mesenchymal stromal cells (MSCs) within the platelet-derived growth factor receptor-alpha (PDGFRα)/CD90 co-expressing cardiac interstitial and adventitial cell fraction. Here we further characterise PDGFRα/CD90-expressing cardiac MSCs (PDGFRα + cMSCs) and use human telomerase reverse transcriptase (hTERT) over-expression to increase cMSCs ability to repair the heart after induced myocardial infarction. hTERT over-expression in PDGFRα + cardiac MSCs (hTERT + PDGFRα + cMSCs) modulates cell differentiation, proliferation, survival and angiogenesis related genes. In vivo , transplantation of hTERT + PDGFRα + cMSCs in athymic rats significantly increased left ventricular function, reduced scar size, increased angiogenesis and proliferation of both cardiomyocyte and non-myocyte cell fractions four weeks after myocardial infarction. In contrast, transplantation of mutant hTERT + PDGFRα + cMSCs (which generate catalytically-inactive telomerase) failed to replicate this cardiac functional improvement, indicating a telomerase-dependent mechanism. There was no hTERT + PDGFRα + cMSCs engraftment 14 days after transplantation indicating functional improvement occurred by paracrine mechanisms. Mass spectrometry on hTERT + PDGFRα + cMSCs conditioned media showed increased proteins associated with matrix modulation, angiogenesis, cell proliferation/survival/adhesion and innate immunity function. Our study shows that hTERT can activate pro-regenerative signalling within PDGFRα + cMSCs and enhance cardiac repair after myocardial infarction. An increased understanding of hTERT’s role in mesenchymal stromal cells from various organs will favourably impact clinical regenerative and anti-cancer therapies.
Publisher: Elsevier BV
Date: 04-2021
Publisher: Public Library of Science (PLoS)
Date: 10-02-2020
Publisher: Elsevier BV
Date: 10-2018
Publisher: Elsevier BV
Date: 08-2016
DOI: 10.1016/J.HLC.2016.04.011
Abstract: Atrial Fibrillation (AF) is one of the most common types of cardiac arrhythmias experienced in clinical practice, increasing the risk of stroke, dementia, myocardial infarction and death. Currently available options for the treatment of AF use either pharmacological agents or catheter-based ablation therapies to restore sinus rhythm or control the ventricular response rate. These current treatment options are suboptimal at best, motivating research into discovering more effective and innovative ways to treat AF. Gene therapy is being explored for its potential to treat various human conditions including cardiac arrhythmias. Gene transfer vectors with increasing transduction efficiency and biosafety have been developed and trialled for cardiovascular disease treatment. With an improved understanding of the molecular mechanisms of AF, several gene therapy targets have been identified and evaluated in an attempt to rate or rhythm control the heart during AF. This review will discuss the gene therapy vectors in use today and methods for delivery of these vectors to the atrium. Further, it will evaluate several gene therapy strategies and approaches for sinus rhythm restoration and ventricular rate control that have the potential to emerge as a therapy for AF.
Publisher: Elsevier BV
Date: 06-2007
DOI: 10.1016/J.HLC.2007.02.106
Abstract: Cardiovascular diseases remain a major cause of morbidity and mortality. Despite significant gains in medical and surgical treatment of these conditions, the burden imposed continues to increase, particularly as the population ages. Hence, there exists an impetus to explore novel therapeutic approaches. Gene therapy, broadly defined as the use of nucleic acid sequences as medicine, is an ex le of such a novel approach. The potential of gene therapy has been explored in several models of both inherited monogenic and acquired polygenic cardiovascular diseases, such as heart failure and arrhythmia. Exciting developments in gene transfer technology and important insights into the molecular basis of these common diseases have placed them within reach of gene-based therapy.
Publisher: Elsevier BV
Date: 07-2023
Publisher: Elsevier BV
Date: 08-2019
Publisher: Elsevier BV
Date: 2020
DOI: 10.1016/J.JACEP.2019.09.013
Abstract: This study sought to summarize the procedural characteristics and outcomes of patients with structural heart disease (SHD) who have focal ventricular tachycardia (VT). Scar-mediated re-entry is the predominant mechanism of VT in SHD. Some SHD patients may have a focal VT mechanism that remains poorly described. An extended induction protocol incorporating programmed electrical stimulation, right ventricular burst pacing and isoprenaline was used to elucidate both re-entrant and focal VT mechanisms. Eighteen of 112 patients (16%) with SHD undergoing VT ablation over 2 years had a focal VT mechanism elucidated (mean age 66±13 years ejection fraction 46±14% nonischemic cardiomyopathy 10). Repetitive failure of termination with antitachycardia pacing (ATP) (69% of patients) or defibrillator shocks (56%) was a common feature of focal VTs. A median of 3 VTs per patient were inducible (28 focal VTs, 34 re-entrant VTs 53% of patients had both focal and re-entrant VT mechanism). Focal VTs more commonly originated from the right ventricle (RV) than the left ventricle (LV) (67% vs. 33%, respectively). In the RV, the RV outflow tract was the most common site (33% of all focal VTs), followed by the RV moderator band (22%), apical septal RV (6%), and lateral tricuspid annulus (6%). The lateral LV (non-Purkinje) was the most common LV focal VT site (16%), followed by the papillary muscles (17%). After median follow-up of 289 days, 78% of patients remained arrhythmia-free no patients had recurrence of focal VT at repeat procedure. In patients with recurrence, defibrillator therapies were significantly reduced from a median of 53 ATP episodes pre-ablation to 10 ATP episodes post-ablation. During follow-up, 2 patients (11%) underwent repeat VT ablation none had recurrence of focal VT. Focal VTs are common in patients with SHD and often coexist with re-entrant forms of VT. High failure rate of defibrillator therapies was a common feature of focal VT mechanisms. Uncovering and abolishing focal VT may further improve outcomes of catheter ablation in SHD.
Publisher: American Physiological Society
Date: 11-2007
DOI: 10.1152/AJPHEART.00477.2007
Abstract: Recombinant lentiviral vectors (LVs) are capable of transducing neonatal rat ventricular myocytes (NRVMs) and providing stable, long-term transgene expression. The goal of the present study was to comprehensively test whether transduction of NRVMs by LVs results in cytotoxicity and to examine the electrophysiological consequences of gene modification of NRVM monolayers by two vectors: one encoding a putatively inert enhanced green fluorescent protein (eGFP) and the other a major ion channel protein, inward rectifier K + channel (Kir) 2.1. Freshly isolated NRVMs were transduced and cultured in monolayers. Immunohistochemistry, Trypan blue exclusion, annexin V binding followed by flow cytometry (FCM), and terminal transferase dUTP nick-end labeling assays were performed to assess for cytotoxicity. Optical mapping studies of action potential propagation in NRVM monolayers were performed to characterize the electrophysiological alterations following transduction. The cytotoxicity assays revealed that transduction had no adverse effects on NRVM cultures. However, eGFP-transduced monolayers exhibited a decrease in conduction velocity (CV) and action potential duration (APD) compared with monolayers transduced with LVs encoding LacZ or devoid of a transgene. In addition, small interfering RNA-mediated knockdown of eGFP expression corrected this phenotype. In contrast, Kir2.1 gene-modified monolayers showed an increase in CV and a predictable decrease in APD. This study demonstrates that LVs transduce NRVMs without cytotoxic effects. However, eGFP has a significant effect on APD and CV in this experimental system and calls into question the widely held belief that GFP is physiologically inert. In addition, LV-mediated overexpression of Kir2.1 opens up the prospect of studying the functional role of inward rectifier K + current in cardiac arrhythmias.
Publisher: Elsevier BV
Date: 05-2011
DOI: 10.1038/MT.2011.32
Publisher: Elsevier BV
Date: 04-2017
DOI: 10.1016/J.HLC.2016.11.002
Abstract: Numerous pharmacological and device therapies have improved adverse cardiac remodelling and mortality in heart failure. However, none are able to regenerate damaged cardiac tissue. Stem cell based therapies using multipotent (adult) stem cells and pluripotent stem cells are new approaches that could potentially achieve the elusive goal of true cardiac regeneration. Over the past two decades, various stem cell based approaches have been shown to improve left ventricular function in pre-clinical animal models. Promising results rapidly led to clinical trials, initially using bone marrow-derived mononuclear cells, then mesenchymal stromal cell populations and, more recently, progenitor cells from the adult heart itself. These have been shown to be safe and have advanced our understanding of potential suitable recipients, cell delivery routes, and possible mechanisms of action. However, efficacy in these trials has been inconsistent. Human pluripotent stem cells (hPSCs) are another potential source of stem cells for cardiac regeneration. They could theoretically provide an unlimited source of cardiomyocytes or cardiac progenitors. Pre-clinical studies in both small and large animal models have shown robust engraftment and improvements in cardiac function. The first clinical trial using hPSC-derived cardiac derivatives has now commenced and others are imminent. In this brief review article, we summarise recent developments in stem cell therapies aimed at cardiac regeneration, including discussion of types of cell and non-cell-based strategies being explored.
Publisher: Proceedings of the National Academy of Sciences
Date: 18-03-2008
Abstract: Congenital long- or short-QT syndrome may lead to life-threatening ventricular tachycardia and sudden cardiac death. Apart from the rare disease-causing mutations, common genetic variants in CAPON, a neuronal nitric oxide synthase (NOS1) regulator, have recently been associated with QT interval variations in a human whole-genome association study. CAPON had been unsuspected of playing a role in cardiac repolarization indeed, its physiological role in the heart (if any) is unknown. To define the biological effects of CAPON in the heart, we investigated endogenous CAPON protein expression and protein–protein interactions in the heart and performed electrophysiological studies in isolated ventricular myocytes with and without CAPON overexpression. We find that CAPON protein is expressed in the heart and interacts with NOS1 to accelerate cardiac repolarization by inhibition of L-type calcium channel. Our findings provide a rationale for the association of CAPON gene variants with extremes of the QT interval in human populations.
Publisher: Wiley
Date: 2007
DOI: 10.1002/JGM.1039
Abstract: Development of effective and durable gene therapy for treatment of the respiratory manifestations of cystic fibrosis remains a formidable challenge. Obstacles include difficulty in achieving efficient gene transfer to mature airway epithelium and the need to stably transduce self-renewing epithelial progenitor cells in order to avoid loss of transgene expression through epithelial turnover. Targeting the developing airway epithelium during fetal life offers the prospect of circumventing these challenges. In the current study we investigated vesicular stomatitis virus glycoprotein (VSVg)-pseudotyped HIV-1-derived lentivirus vector-mediated gene transfer to the airway epithelium of mid-gestation fetal lambs, both in vitro and in vivo. In the in vitro studies epithelial sheet explants and lung organ culture were used to examine transduction of the proximal and more distal airway epithelium, respectively. For the in vivo studies, vector was delivered directly into the proximal airway. We found that even during the early pseudoglandular and canalicular phases of lung development, occurring through mid-gestation, the proximal bronchial airway epithelium was relatively mature and highly resistant to lentivirus-mediated transduction. In contrast, the more distal bronchiolar airway epithelium was relatively permissive for transduction although the absolute levels achieved remained low. This result is promising as the bronchiolar airway epithelium is a major site of pathology in the cystic fibrosis airway, and much higher levels of transduction are likely to be achieved by developing strategies that increase the amount of vector reaching the more distal airway after intratracheal delivery.
Publisher: Oxford University Press (OUP)
Date: 02-05-2015
Abstract: Early atrial arrhythmia following atrial fibrillation (AF) ablation is associated with higher recurrence rates. Few studies explore the impact of early AF (EAF) and atrial tachycardia (EAT) on long-term outcomes. Furthermore, EAF/EAT have not been characterized after wide pulmonary vein isolation. We aimed to characterize EAF and EAT and its impact on late AF (LAF) and AT (LAT) after single ring isolation (SRI). We recruited 119 (females 21, age 58 ± 10 years) consecutive patients with AF (paroxysmal 76, persistent 43) undergoing SRI. Early atrial fibrillation/ early atrial tachycardia was defined as AF/AT within 3 months post-procedure (blanking period). Patients were followed for median 2.8[2.2-4] years. Early atrial fibrillation occurred in 28% (n = 33) and EAT in 25% (n = 30). At follow-up, 25% (n = 30) had LAF and 28% (n = 33) had LAT. Patients with EAF and EAT had higher rates of LAF (48 vs. 16%, P<0.0001) and LAT (60 vs. 16%, P < 0.0001), respectively. Independent predictors of LAF were EAF (3.53(1.72-7.29) P = 0.001) and of LAT were EAT (5.62(2.88-10.95) P < 0.0001) and procedure time (1.38/ h(1.07-1.78) P = 0.04). Importantly, EAF did not predict LAT and EAT did not predict LAF. Early atrial fibrillation late in the blanking period was associated with higher rates of LAF (73% for month 3 vs. 25% for Months 1-2, P = 0.004). However, EAT timing did not predict LAT. Early atrial fibrillation and EAT are predictive of LAF and LAT, respectively. Early atrial fibrillation late in the blanking period has greater predictive significance for LAF. This timing is not relevant for LAT. Early arrhythmia type and timing have important prognostic significance following SRI. www.anzctr.org.au ACTRN12606000467538.
Publisher: Elsevier BV
Date: 12-2021
DOI: 10.1016/J.JCYT.2021.07.016
Abstract: Mesenchymal stromal cells (MSCs) have been shown to improve cardiac function after injury and are the subject of ongoing clinical trials. In this study, the authors tested the cardiac regenerative potential of an induced pluripotent stem cell-derived MSC (iPSC-MSC) population (Cymerus MSCs) in a rat model of myocardial ischemia-reperfusion (I/R). Furthermore, the authors compared this efficacy with bone marrow-derived MSCs (BM-MSCs), which are the predominant cell type in clinical trials. Four days after myocardial I/R injury, rats were randomly assigned to (i) a Cymerus MSC group (n = 15), (ii) a BM-MSC group (n = 15) or (iii) a vehicle control group (n = 14). For cell-treated animals, a total of 5 × 10 One month after cell transplantation, Cymerus MSCs improved LV function (assessed by echocardiography) compared with vehicle and BM-MSCs. Interestingly, Cymerus MSCs enhanced angiogenesis without sustained engraftment or significant impact on infarct scar size. Suggesting safety, Cymerus MSCs had no effect on inducible tachycardia or the ventricular scar heterogeneity that provides a substrate for cardiac re-entrant circuits. The authors here demonstrate that intra-myocardial administration of iPSC-MSCs (Cymerus MSCs) provide better therapeutic effects compared with conventional BM-MSCs in a rodent model of myocardial I/R. Because of its manufacturing scalability, iPSC-MSC therapy offers an exciting opportunity for an "off-the-shelf" stem cell therapy for cardiac repair.
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 23-06-2021
Publisher: Wiley
Date: 06-08-2020
DOI: 10.1111/PACE.14025
Publisher: Elsevier BV
Date: 2018
DOI: 10.1016/J.HLC.2017.02.010
Abstract: Patch cl ing studies using non-cardiomyocytes revealed that the human connexin40 mutations P88S, G38D, and A96S are associated with reduced gap junction conductances compared to wild type connexin40 (wtCx40). Their effects within myocytes however are unclear. We aimed to characterise P88S, G38D, and A96S after expression in rat hearts and primary cardiomyocyte cultures. Adult Sprague-Dawley rat atria were transduced with a lentivector containing a transgene encoding wtCx40, P88S, G38D, A96S, or eGFP (n=6 per transgene). Electrophysiology studies (EPS) were performed just prior to and 7 days after surgery. Left atria were assessed for connexin expression, mRNA levels, inflammation and fibrosis. Primary cardiomyocyte cultures were also transduced with the abovementioned vectors (n=6 per transgene) and monolayer conduction velocities (CV) and protein expression were assessed at 96hours. At day 7 EPS, P wave and induced atrial fibrillation (AF) durations were significantly longer in the mutant groups when compared to wtCx40 controls (p<0.05). There were no significant differences in inflammation, fibrosis, or heart to body weight ratios. Monolayer CV's were reduced in the A96S group compared to the wtCx40 group. While similar to wtCx40 controls, P88S velocities were reduced compared to eGFP controls. G38D monolayers possessed spontaneous fibrillatory activity and could not be paced. Immunofluorescence revealed that P88S and G38D reduced native connexin43 myocyte coupling while A96S appeared to co-localise with connexin43 in gap junctions. Connexin43 mRNA levels were similar between groups. The A96S, G38D, and P88S Cx40 mutations slow conduction and increased the propensity for inducible AF.
Publisher: Elsevier BV
Date: 10-2020
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 02-2005
DOI: 10.1161/01.CIR.0000153812.64956.EF
Abstract: Background— Cardiac conduction occurs in an electrical syncytium of excitable cells connected by gap junctions. Disruption of these electrophysiological properties causes conduction slowing or block. Depending on the location of affected cells within the heart, this has the potential to result in clinical syndromes such as atrioventricular block. With a view to developing gene therapy strategies for repairing cardiac conduction defects, we sought to establish whether the phenotype of fibroblasts can be modified by gene transfer to produce cells capable of electrical excitation and coupling. Methods and Results— High-titer lentiviral vectors encoding MyoD, a myogenic transcription factor, and connexin43, a gap junction protein, were produced by established methods. Human dermal fibroblasts (HDFs) were efficiently ( %) transduced at a multiplicity of infection of 50. HDFs transduced with the MyoD-encoding vector underwent myogenic conversion, as evidenced by myotube formation and detection of muscle-specific proteins. Importantly, calcium transients indicative of membrane excitability were observed in MyoD-induced myotubes after loading with a calcium-sensitive dye and electrical stimulation. Transients from adjacent myotubes displayed different excitation thresholds, indicating an absence of coupling between cells, consistent with skeletal muscle biology. In contrast, simultaneous transduction of HDFs with MyoD and connexin43-encoding vectors resulted in the appearance of transients in adjacent myotubes with identical thresholds, indicative of electrical coupling. Notably, dye transfer studies confirmed gap junctional intercellular communication. Conclusions— Fibroblasts can be genetically modified to produce excitable cells capable of electrical coupling. These observations strengthen the prospect of developing gene-based strategies for repairing cardiac conduction defects.
Publisher: Wiley
Date: 2004
DOI: 10.1002/CM.20010
Abstract: We describe a simple culture method for obtaining highly differentiated clonal C2C12 myotubes using a feeder layer of confluent fibroblasts, and document the expression of contractile protein expression and aspects of myofibre morphology using this system. Traditional culture methods using collagen- or laminin-coated tissue-culture plastic typically results in a cyclic pattern of detachment and reformation of myotubes, rarely producing myotubes of a mature adult phenotype. C2C12 co-culture on a fibroblast substratum facilitates the sustained culture of contractile myotubes, resulting in a mature sarcomeric register with evidence for peripherally migrating nuclei. Immunoblot analysis demonstrates that desmin, tropomyosin, sarcomeric actin, alpha-actinin-2 and slow myosin are detected throughout myogenic differentiation, whereas adult fast myosin heavy chain isoforms, members of the dystrophin-associated complex, and alpha-actinin-3 are not expressed at significant levels until >6 days of differentiation, coincident with the onset of contractile activity. Electrical stimulation of mature myotubes reveals typical and reproducible calcium transients, demonstrating functional maturation with respect to calcium handling proteins. Immunocytochemical staining demonstrates a well-defined sarcomeric register throughout the majority of myotubes (70-80%) and a striated staining pattern is observed for desmin, indicating alignment of the intermediate filament network with the sarcomeric register. We report that culture volume affects the fusion index and rate of sarcomeric development in developing myotubes and propose that a fibroblast feeder layer provides an elastic substratum to support contractile activity and likely secretes growth factors and extracellular matrix proteins that assist myotube development.
Publisher: Elsevier BV
Date: 10-2021
DOI: 10.1016/J.IJCARD.2021.07.021
Abstract: Novel therapies that can limit or reverse damage caused by myocardial infarction (MI) could ease the increasing burden of heart failure. In this regard Platelet Derived Growth Factor (PDGF) has been previously shown to contribute to cardiac repair after MI. Here, we use a rodent model of MI and recombinant adeno-associated virus 9 (rAAV9)-mediated gene transfer to overexpress Pdgf-a in the injured heart and assess its therapeutic potential. Sprague Dawley rats underwent temporary occlusion of the left anterior descending coronary artery, followed immediately by systemic delivery of 1 × 10^11 vector genomes of either rAAV9 Pdgf-a or rAAV9 Empty vector (control). At day 28 post-MI echocardiography showed significantly improved left ventricular (LV) function (fractional shortening) after rAAV9 Pdgf-a (0.394 ± 0.019%) treatment vs control (0.304 ± 0.018%). Immunohistochemical analysis demonstrated significantly increased capillary and arteriolar density in the infarct border zone of rAAV9 Pdgf-a treated hearts together with a significant reduction in infarct scar size (rAAV9 Pdgf-a 6.09 ± 0.94% vs Empty 12.45 ± 0.92%). Western blot and qPCR analyses confirmed overexpression of PDGF-A and showed upregulation of smooth muscle alpha actin (Acta2), collagen type III alpha 1 (Col3a1) and lysyl oxidase (Lox) genes in rAAV9 Pdgf-a treated infarcts. Overexpression of Pdgf-a in the post-MI heart can modulate scar composition and improve LV function. Our study highlights the potential of rAAV gene transfer of Pdgf-a as a cardio-reparative therapy.
Publisher: MDPI AG
Date: 16-09-2023
DOI: 10.3390/BIOM13091397
Publisher: Springer US
Date: 2022
DOI: 10.1007/978-1-0716-2707-5_26
Abstract: Gene therapy based on recombinant adeno-associated viral (rAAV) vectors has recently made significant progress as a clinical therapeutic. Unlike most traditional medications, gene therapy vectors can be biologically active when shed into the surrounding environment. Here we describe methods for collection and storage of multiple biological specimen s les and a PCR-based method for detection of shed adeno-associated viral (AAV) particles. We also describe a method for use of an infectious replication assay utilizing a cell line stably expressing AAV Rep and Cap genes and superinfection with adenovirus 5 to detect functionality in shed AAV particles.
Publisher: Elsevier BV
Date: 05-2008
DOI: 10.1038/MT.2008.30
Publisher: Elsevier BV
Date: 04-2007
DOI: 10.1016/J.NMD.2006.12.010
Abstract: The objective of this study is to expand the applications of MyoD-forced myogenesis for research and diagnosis of human muscle disorders using a lentiviral vector (LVhMyoD) for efficient trans-differentiation of patient primary cells. LVhMyoD transduced cells readily formed striated, multinucleate myotubes expressing a wide range of genes associated with muscular dystrophy (dystrophin, dysferlin, sarcoglycans, caveolin-3) and congenital myopathy (nebulin, actin, desmin, tropomyosin, troponin). We demonstrate that MyoD gene-modified fibroblasts reproduce protein deficiencies associated with different forms of muscular dystrophy, and confirm that LVhMyoD gene-modified chorionic villus can be used successfully to determine the dystrophin status of the developing fetus, augmenting prenatal diagnosis of dystrophinopathy patients. Using muscle-specific cDNA derived from LVhMyoD gene-modified patient cells, we identified a female carrier bearing a large dystrophin deletion and a previously unidentified non-coding splice-site mutation within dystrophin in a Becker muscular dystrophy patient. This study highlights the significant potential of lentiviral MyoD-forced myogenesis for study of a wide range of human muscle disorders a field constrained by the limited availability of human tissue. LVhMyoD gene-modified patient cells provide a renewable source of mutant protein and muscle-specific mRNA, facilitating accelerated mutation screening of large genes, molecular analyses of splicing abnormalities and study of disease-causing mutations.
Publisher: Elsevier BV
Date: 06-2023
Publisher: S. Karger AG
Date: 2017
DOI: 10.1159/000477436
Abstract: 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.
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 20-07-2007
DOI: 10.1161/CIRCRESAHA.107.155721
Abstract: Calcium cycling figures prominently in excitation-contraction coupling and in various signaling cascades involved in the development of left ventricular hypertrophy. We hypothesized that genetic suppression of the L-type calcium channel accessory β-subunit would modulate calcium current and suppress cardiac hypertrophy. A short hairpin RNA template sequence capable of mediating the knockdown of the L-type calcium channel accessory β-subunit gene was incorporated into a lentiviral vector (PPT.CG.H1.β 2 ). Transduction of ventricular myocytes in vivo with the active short hairpin RNA partially inhibited the L-type calcium current. In neonatal rat cardiomyocytes, L-type calcium channel accessory β-subunit gene knockdown reduced calcium transient litude. Similarly, [ 3 H]leucine incorporation was attenuated in PPT.CG.H1.β 2 -transduced neonatal rat cardiomyocytes compared with nonsilencing controls in a phenylephrine-induced hypertrophy model. In vivo gene transfer attenuated the hypertrophic response in an aortic-banded rat model of left ventricular hypertrophy, with reduced left ventricular wall thickness and heart weight/body weight ratios in PPT.CG.H1.β 2 -injected rats at four weeks post transduction. Fractional shortening was preserved in rats treated with PPT.CG.H1.β 2 . These findings indicate that knockdown of L-type calcium channel accessory β-subunit is capable of attenuating the hypertrophic response both in vitro and in vivo without compromising systolic performance. Suppression of the calcium channel β subunit may represent a novel and useful therapeutic strategy for left ventricular hypertrophy.
Publisher: Wiley
Date: 11-1996
DOI: 10.1111/J.1440-1673.1996.TB00448.X
Abstract: To evaluate the role of radiological imaging of meningiomas in confirming the diagnosis and as a neuroanatomical aid to surgical planning, 115 patients with surgically excised meningiomas between 1990 and 1993 were studied. Computed tomography (CT), magnetic resonance imaging (MRI) (on a 0.5 T unit) and angiography were reviewed, and compared with histopathology (when available). Seventy-eight CT, 89 MRI and 85 angiographic studies were reviewed, and correlated with histopathology in 67 cases. In 48 cases, the surgical specimens could not be pathologically classified. The most common lesion sites were the cerebral convexities, falx and sphenoidal ridges. True demarcation of cleavage planes was seen on 73% of MRI and 10% of CT studies. Computed tomography showed hyperostosis in 27% and MRI in 7% of studies. Tumours enhanced strongly with contrast in 98% of CT scans. On MRI there were variable signal intensities on different sequences, and no correlation between signal intensities and histological subtype was found. Oedema was present in 59% of CT and 66% of MRI studies, and was most pronounced in lesions > 3 cm in diameter. Tumour calcification was seen in 62% of CT and 8% of MRI studies. Vascular abnormalities were seen on 65% of MRI, 21% of CT and 84% of angiogram studies. Angiographic tumour vascularity did not correlate with histologic subtype. All three imaging modalities have management roles: CT for bony changes and calcification, MRI for multiplanar and vessel anatomy imaging, and angiography for vessel delineation and embolization if required.
Publisher: Oxford University Press (OUP)
Date: 04-2012
Abstract: 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.
Publisher: Elsevier BV
Date: 08-2010
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 06-01-2023
DOI: 10.1161/CIRCRESAHA.122.321123
Abstract: Myocardial infarction (MI) is among the leading causes of death worldwide. Following MI, necrotic cardiomyocytes are replaced by a stiff collagen-rich scar. Compared to collagen, the extracellular matrix protein elastin has high elasticity and may have more favorable properties within the cardiac scar. We sought to improve post-MI healing by introducing tropoelastin, the soluble subunit of elastin, to alter scar mechanics early after MI. We developed an ultrasound-guided direct intramyocardial injection method to administer tropoelastin directly into the left ventricular anterior wall of rats subjected to induced MI. Experimental groups included shams and infarcted rats injected with either PBS vehicle control or tropoelastin. Compared to vehicle treated controls, echocardiography assessments showed tropoelastin significantly improved left ventricular ejection fraction (64.7±4.4% versus 46.0±3.1% control) and reduced left ventricular dyssynchrony (11.4±3.5 ms versus 31.1±5.8 ms control) 28 days post-MI. Additionally, tropoelastin reduced post-MI scar size (8.9±1.5% versus 20.9±2.7% control) and increased scar elastin (22±5.8% versus 6.2±1.5% control) as determined by histological assessments. RNA sequencing (RNAseq) analyses of rat infarcts showed that tropoelastin injection increased genes associated with elastic fiber formation 7 days post-MI and reduced genes associated with immune response 11 days post-MI. To show translational relevance, we performed immunohistochemical analyses on human ischemic heart disease cardiac s les and showed an increase in tropoelastin within fibrotic areas. Using RNA-seq we also demonstrated the tropoelastin gene ELN is upregulated in human ischemic heart disease and during human cardiac fibroblast-myofibroblast differentiation. Furthermore, we showed by immunocytochemistry that human cardiac fibroblast synthesize increased elastin in direct response to tropoelastin treatment. We demonstrate for the first time that purified human tropoelastin can significantly repair the infarcted heart in a rodent model of MI and that human cardiac fibroblast synthesize elastin. Since human cardiac fibroblasts are primarily responsible for post-MI scar synthesis, our findings suggest exciting future clinical translation options designed to therapeutically manipulate this synthesis.
Publisher: Wiley
Date: 20-01-2020
DOI: 10.1111/JCE.14351
Abstract: Minimal data exist on the Advisor HD Grid (HDG) catheter and the Precision electroanatomic mapping (EAM) system for ventricular arrhythmia (VA) procedures. Using the HDG catheter, the EAM uses the high-density (HD) wave mapping and best duplicate software to compare the maximum peak-to-peak bipolar voltages within a small zone independent of wavefront direction and catheter orientation. This study aimed to summarize the procedural experience for VAs using the HDG catheter. Clinical and procedural characteristics of VA ablation procedures were retrospectively reviewed that used the HDG catheter and the Precision EAM over a 12-month period. A total of 22 patients, 18 with sustained ventricular tachycardia and 4 with premature ventricular contractions were included. Clinically indicated left and/or right ventricular (LV, RV, respectively), and aortic maps were created. LV substrate maps (n = 13) used a median 1700 points (interquartile range [IQR] HD wave mapping using the novel HDG catheter integrated with the Precision EAM is safe and feasible in VA procedures in the LV, RV, and aorta. Mapping times are consistent with other multielectrode mapping catheters.
Publisher: Springer Science and Business Media LLC
Date: 09-2011
DOI: 10.1038/NATURE10407
Publisher: Springer Science and Business Media LLC
Date: 11-06-2009
DOI: 10.1038/GT.2009.64
Abstract: In mammalian cells, small regulatory RNA molecules are able to modulate gene expression in a cell-autonomous manner. In contrast, this mechanism of gene regulation can occur systemically in plants and nematodes. The existence of similar cell-to-cell transmission in mammalian cells has been explored, but generalizibilty and mechanistic insights have remained elusive. Here, we show that small regulatory RNA molecules are capable of a non-cell-autonomous effect between primary cardiac myocytes through a gap-junction-dependent mechanism. Co-culture experiments showed that both Dicer-processed small-interfering RNAs (siRNAs) and Drosha-processed microRNAs (miRNAs) were capable of target gene knockdown and physiological effects in a non-cell-autonomous manner. Target gene siRNA molecules were detected in recipient cells, indicating transfer of the primary effector molecule. All of these effects were abrogated by dominant-negative molecular suppression of gap junction function. Our results show that both siRNAs and miRNAs are capable of a non-cell-autonomous effect between mammalian cells through gap junctions. The recognition of this biological process raises the novel therapeutic prospect of a bystander effect after gene transfer to tissues bearing gap junctions and for cell engineering with a view to creating regulatory RNA donor cells that exert their influence throughout a syncytium.
Publisher: Springer Science and Business Media LLC
Date: 13-07-2006
Abstract: In an earlier study exploring the potential of gene transfer to repair myocardial conduction defects, we observed that myotubes, generated by forced expression of MyoD, exhibit reduced excitability when also modified to express connexin43 (Cx43). We hypothesized that this effect was caused by gap junction-mediated coupling between myotubes and the underlying fibroblast feeder layer. This intriguing possibility has important implications for ongoing efforts to develop strategies for repairing myocardial conduction defects by gene transfer, and also provides novel insights into the electrophysiological function of naturally occurring heterologous cell coupling within the heart. Although a conductive function for fibroblasts through heterologous coupling has previously been reported, the current study provides novel evidence that fibroblasts can modulate cardiomyocyte excitability in a Cx43-dependent manner. In a co-culture study system, neonatal rat cardiomyocytes were grown on monolayers of mouse fibroblasts with genetically altered Cx43 expression and the effect on intrinsic beat frequency examined. Cardiomyocytes grown on wild-type (WT) fibroblasts expressing native levels of Cx43 beat significantly slower than cells grown on fibroblasts devoid of this molecule (germline knockout) or with dominant-negative functional suppression. Expression of Cx43 in fibroblasts from Cx43 knockout mice restored cardiomyocyte beat frequency, to rates comparable with those observed in co-culture with WT fibroblasts.
Publisher: Europa Digital & Publishing
Date: 06-2014
DOI: 10.4244/EIJV10I2A46
Publisher: Wiley
Date: 2011
Publisher: Elsevier BV
Date: 11-2008
Publisher: Elsevier BV
Date: 08-2018
DOI: 10.1016/J.HLC.2018.01.006
Abstract: Vitamin D has been implicated in the prevention of heart failure. However the underlying mechanism remains unclear. We hypothesised that these effects may be partially mediated by cardiac stem rogenitor cells (CPCs). Therefore, we examined the effects of 1,25-dihydroxyvitamin D3 (1,25D) on cell cycle activity and differentiation of a previously described CPC population called cardiac colony-forming unit fibroblasts (cCFU-Fs). cCFU-Fs were isolated from adult male C57Bl/6 mouse hearts using fluorescence-activated cell sorting. The effect of 1,25D on cell proliferation and differentiation were was assessed by colony-forming and fibroblast differentiation assays. Cell cycle was analysed by flow cytometry. Mice with induced myocardial infarction (MI) were treated with 1,25D or vehicle controls and cardiac function assessed by echocardiography. 1,25D dose-dependently increased expression of vitamin D receptor (Vdr) and reduced large colony formation. Addition of 1,25D to cCFU-Fs slowed cell proliferation, promoted cell cycle arrest and decreased expression of pro-fibrotic factors during TGF-β-induced fibroblast differentiation of cCFU-Fs. After MI, 1,25D-treated mice had less left ventricular wall thinning and significant improvement in left ventricular systolic function compared to vehicle-treated controls. Although no significant changes in myocardial fibrotic area and cardiomyocyte size were noted, treatment with 1,25D significantly inhibited cardiac interstitial cell proliferation after MI. Vitamin D signalling promotes cardioprotection after myocardial infarction. This may be through modulation of cCFU-F cell cycle. The role of 1,25D and VDR in regulating cardiac stem rogenitor cell function therefore warrants further investigation.
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 08-06-2007
DOI: 10.1161/CIRCRESAHA.106.144956
Abstract: Modification of electrical conduction would be a useful principle to recruit in preventing or treating certain arrhythmias, notably ventricular tachycardia (VT). Here we pursue a novel gene transfer approach to modulate electrical conduction by reducing gap junctional intercellular communication (GJIC) and hence potentially modify the arrhythmia substrate. The ultimate goal is to develop a nondestructive approach to uncouple zones of slow conduction by focal gene transfer. Lentiviral vectors encoding connexin43 (Cx43) internal loop mutants were produced and studied in vitro. Transduction of neonatal rat ventricular myocytes (NRVMs) revealed the expected subcellular localization of the mutant gene product. Fluorescent dye transfer studies showed a significant reduction of GJIC in NRVMs that had been genetically modified. Additionally, adjacent mutant gene-modified NRVMs displayed delayed calcium transients, indicative of electrical uncoupling. Multi-site optical mapping of action potential (AP) propagation in gene-modified NRVM monolayers revealed a 3-fold slowing of conduction velocity (CV) relative to nontransduced NRVMs. In conclusion, lentiviral vector–mediated gene transfer of Cx43 mutants reduced GJIC in NRVMs. Electrical charge transfer was also reduced as evidenced by delayed calcium transients in adjacent NRVMs and reduced CV in NRVM monolayers. These data validate a molecular tool that opens the prospect for gene transfer targeting gap junctions as an approach to modulate cardiac conduction.
Publisher: Proceedings of the National Academy of Sciences
Date: 11-08-2009
Abstract: Gap junction intercellular communication (GJIC) is ubiquitous in the majority of vertebrate cells and is required for the proper development of most tissues. The loss of gap junction-mediated cell-to-cell communication leads to compromised development in many tissues and organs. Because cells constantly interact through gap junctions to coordinate tissue functions and homeostasis, we hypothesized that increasing cell-to-cell communication, via genetically engineering cells to overexpress gap junction proteins, could enhance cell differentiation in the interior regions of 3D tissue equivalents, thereby increasing the ability to regenerate larger and more uniform volumes of tissue. To test this hypothesis, we used bone as a model tissue because of the difficulty in achieving spatially uniform bone regeneration in 3D. In bone marrow stromal cells (BMSC), GJIC and osteogenic differentiation were compromised in 3D cultures relative to 2D monolayers and in the core of 3D cultures relative to the surface. Overexpression of connexin 43 (Cx43) via transduction of BMSCs with a lentivirus overcame this problem, enhancing both the magnitude and spatial distribution of GJIC and osteogenic differentiation markers throughout 3D constructs. Transplantation of cells overexpressing Cx43 resulted in an increased volume fraction and spatial uniformity of bone in vivo, relative to nontransduced BMSCs. Increased GJIC also enhanced the effect of a potent osteoinductive agent (BMP-7), suggesting a synergism between the soluble factor and GJIC. These findings present a platform to improve cell-to-cell communication in 3D and to achieve uniformly distributed tissue regeneration in 3D.
Publisher: Elsevier BV
Date: 07-2023
Publisher: Wiley
Date: 16-05-2014
DOI: 10.1111/PACE.12423
Abstract: Organized atrial tachycardias (OATs) after pulmonary vein isolation (PVI) procedure are common. Arrhythmia mechanisms include mitral annular, ring gap, or roof-dependent gap-related flutters. In this series, we describe a mechanism of arrhythmia utilizing the ridge between left pulmonary vein (PV) and left atrial appendage (LAA) in the Ligament of Marshall (LOM) region. Five tachycardias involving the LOM region were identified from a group of 240 patients who underwent a single ring PVI procedure for symptomatic atrial fibrillation. The common characteristics of these tachycardias were the endocardial breakout over a broad area adjacent to the LOM region, presence of presystolic or mid-diastolic potentials, and abolition by ablation of the presystolic or mid-diastolic potentials remote from the endocardial breakout site. In all five cases, tachycardias were present after isolation of the veins and posterior left atria. All demonstrated characteristic areas of very slow conduction in the LOM region highlighted by presence of either low voltage, long duration fractionated potentials, or mid-diastolic potentials with a fixed temporal relationship to the subsequent endocardial activation. The pattern of activation and termination of tachycardia during ablation was consistent with an arrhythmia utilizing an electrically insulated tract within LOM and the PV-LAA ridge region. We identified a pattern of arrhythmias involving a concealed presystolic component and a broad endocardial breakout site related to the LOM region. Successful ablation site involved careful identification of small diastolic potentials in the LAA/ridge region or adjacent to the coronary sinus.
Publisher: Bentham Science Publishers Ltd.
Date: 10-2003
Abstract: Cardiovascular disease remains a major cause of morbidity and mortality in modern societies. While contemporary treatment modalities are making steady inroads to reduce this disease burden there remains a pressing need to vigorously explore novel therapeutic strategies. Rapid advances in our understanding of molecular pathology and the evolution of increasingly efficient gene transfer technology offer the imminent prospect of gene-based approaches to, at least, a subset of cardiovascular pathophysiologies. Initially envisaged as a treatment strategy for inherited monogenic disorders, it is now apparent that gene therapy has broader potential that encompasses acquired polygenic diseases, including many that affect the cardiovascular system. Extensive in vitro and animal studies are providing an increasingly sound scientific basis for cautious human evaluation. This review focuses on gene therapy of diseases primarily afflicting the heart, and provides an overview of gene and vector delivery systems with particular emphasis on systems suited to in idual cardiac conditions. The pathophysiology underlying these conditions and molecular targets for therapeutic intervention are also reviewed.
Publisher: MyJove Corporation
Date: 10-02-2023
DOI: 10.3791/64177
Publisher: Oxford University Press (OUP)
Date: 05-2018
Publisher: Frontiers Media SA
Date: 03-07-2023
DOI: 10.3389/FCVM.2023.1183485
Abstract: Differentiating phenotypes of cardiac “hypertrophy” characterised by increased wall thickness on echocardiography is essential for management and prognostication. Transthoracic echocardiography is the most commonly used screening test for this purpose. We sought to identify echocardiographic markers that distinguish infiltrative and storage disorders that present with increased left ventricular (LV) wall thickness, namely, cardiac amyloidosis (CA) and Anderson–Fabry disease (AFD), from hypertensive heart disease (HHT). Patients were retrospectively recruited from Westmead Hospital, Sydney, and Princess Alexandra Hospital, Brisbane. LV structural, systolic, and diastolic function parameters, as well as global (LVGLS) and segmental longitudinal strains, were assessed. Previously reported echocardiographic parameters including relative apical sparing ratio (RAS), LV ejection fraction-to-strain ratio (EFSR), mass-to-strain ratio (MSR) and amyloidosis index (AMYLI) score (relative wall thickness × E / e ′) were evaluated. A total of 209 patients {120 CA [58 transthyretin amyloidosis (ATTR) and 62 light-chain (AL) amyloidosis], 31 AFD and 58 HHT patients mean age 64.1 ± 13.7 years, 75% male} comprised the study cohort. Echocardiographic measurements differed across the three groups, The LV mass index was higher in both CA {median 126.6 [interquartile range (IQR) 106.4–157.9 g/m 2 ]} and AFD [median 134 (IQR 108.8–152.2 g/m 2 )] vs. HHT [median 92.7 (IQR 79.6–102.3 g/m 2 ), p & 0.05]. LVGLS was lowest in CA [median 12.29 (IQR 10.33–15.56%)] followed by AFD [median 16.92 (IQR 14.14–18.78%)] then HHT [median 18.56 (IQR 17.51–19.97%), p & 0.05]. Diastolic function measurements including average e ′ and E / e ′ were most impaired in CA and least impaired in AFD. Indexed left atrial volume was highest in CA. EFSR and MSR differentiated secondary (CA + AFD) from HHT [receiver operating curve–area under the curve (ROC-AUC) of 0.80 and 0.91, respectively]. RAS and AMYLI score differentiated CA from AFD (ROC-AUC of 0.79 and 0.80, respectively). A linear discriminant analysis with stepwise variable selection using linear combinations of LV mass index, average e ′, LVGLS and basal strain correctly classified 79% of all cases. Simple echocardiographic parameters differentiate between different “hypertrophic” cardiac phenotypes. These have potential utility as a screening tool to guide further confirmatory testing.
Publisher: Elsevier BV
Date: 12-2021
Publisher: American Association for the Advancement of Science (AAAS)
Date: 2020
DOI: 10.1126/SCITRANSLMED.AAY2140
Abstract: Recombinant human platelet-derived growth factor-AB improves cardiac function and survival after myocardial infarction in a porcine model.
Publisher: MDPI AG
Date: 17-08-2022
DOI: 10.3390/IJMS23169230
Abstract: Sinoatrial node dysfunction can manifest as bradycardia, leading to symptoms of syncope and sudden cardiac death. Electronic pacemakers are the current standard of care but are limited due to a lack of biological chronotropic control, cost of revision surgeries, and risk of lead- and device-related complications. We therefore aimed to develop a biological alternative to electronic devices by using a clinically relevant gene therapy vector to demonstrate conversion of cardiomyocytes into sinoatrial node-like cells in an in vitro context. Neonatal rat ventricular myocytes were transduced with recombinant adeno-associated virus vector 6 encoding either hTBX18 or green fluorescent protein and maintained for 3 weeks. At the endpoint, qPCR, Western blot analysis and immunocytochemistry were used to assess for reprogramming into pacemaker cells. Cell morphology and Arclight action potentials were imaged via confocal microscopy. Compared to GFP, hTBX18-transduced cells showed that hTBX18, HCN4 and Cx45 were upregulated. Cx43 was significantly downregulated, while sarcomeric α-actinin remained unchanged. Cardiomyocytes transduced with hTBX18 acquired the tapering morphology of native pacemaker cells, as compared to the block-like, striated appearance of ventricular cardiomyocytes. Analysis of the action potentials showed phase 4 depolarization and a significant decrease in the APD50 of the hTBX18-transduced cells. We have demonstrated that rAAV-hTBX18 gene transfer to ventricular myocytes results in morphological, molecular, physiological, and functional changes, recapitulating the pacemaker phenotype in an in vitro setting. The generation of these induced pacemaker-like cells using a clinically relevant vector opens new prospects for biological pacemaker development.
Publisher: Elsevier BV
Date: 07-2023
Publisher: Ovid Technologies (Wolters Kluwer Health)
Date: 10-2017
DOI: 10.1161/CIRCEP.116.004998
Abstract: Recent studies have demonstrated that intramyocardial adipose tissue (IMAT) may contribute to ventricular electrophysiological remodeling in patients with chronic myocardial infarction. Using an ovine model of myocardial infarction, we aimed to determine the influence of IMAT on scar tissue identification during endocardial contact mapping and optimal voltage-based mapping criteria for defining IMAT dense regions. In 7 sheep, left ventricular endocardial and transmural mapping was performed 84 weeks (15–111 weeks) post-myocardial infarction. Spearman rank correlation coefficient was used to assess the relationship between endocardial contact electrogram litude and histological composition of myocardium. Receiver operator characteristic curves were used to derive optimal electrogram thresholds for IMAT delineation during endocardial mapping and to describe the use of endocardial mapping for delineation of IMAT dense regions within scar. Endocardial electrogram litude correlated significantly with IMAT (unipolar r =−0.48±0.12, P .001 bipolar r =−0.45±0.22, P =0.04) but not collagen (unipolar r =−0.36±0.24, P =0.13 bipolar r =−0.43±0.31, P =0.16). IMAT dense regions of myocardium reliably identified using endocardial mapping with thresholds of .7 and .6 mV, respectively, for unipolar, bipolar, and combined modalities (single modality area under the curve=0.80, P .001 combined modality area under the curve=0.84, P .001). Unipolar mapping using optimal thresholding remained significantly reliable (area under the curve=0.76, P .001) during mapping of IMAT, confined to putative scar border zones (bipolar litude, 0.5–1.5 mV). These novel findings enhance our understanding of the confounding influence of IMAT on endocardial scar mapping. Combined bipolar and unipolar voltage mapping using optimal thresholds may be useful for delineating IMAT dense regions of myocardium, in postinfarct cardiomyopathy.
Publisher: MDPI AG
Date: 19-10-2018
DOI: 10.3390/JCDD5040050
Abstract: Bradycardia arising from pacemaker dysfunction can be debilitating and life threatening. Electronic pacemakers serve as effective treatment options for pacemaker dysfunction. They however present their own limitations and complications. This has motivated research into discovering more effective and innovative ways to treat pacemaker dysfunction. Gene therapy is being explored for its potential to treat various cardiac conditions including cardiac arrhythmias. Gene transfer vectors with increasing transduction efficiency and biosafety have been developed and trialed for cardiovascular disease treatment. With an improved understanding of the molecular mechanisms driving pacemaker development, several gene therapy targets have been identified to generate the phenotypic changes required to correct pacemaker dysfunction. This review will discuss the gene therapy vectors in use today along with methods for their delivery. Furthermore, it will evaluate several gene therapy strategies attempting to restore biological pacing, having the potential to emerge as viable therapies for pacemaker dysfunction.
Publisher: Elsevier BV
Date: 09-2023
Location: United States of America
No related grants have been discovered for Eddy Kizana.