Niall Campbell

Cell Delivery Routes for Stem Cell Therapy to the Heart: Current and Future Approaches

An important factor to determine the success of stem cell therapy to the heart is the choice of cell delivery route. This will affect the fate of donor cells and subsequently influence the outcome of treatment; however, there is currently no optimum cell delivery route appropriate for every disease condition or every donor cell type. This review summarises currently available approaches for administering cells to the heart, with a particular focus on cell retention/survival and the therapeutic benefits seen in preclinical and clinical studies. Two major approaches are intracoronary and intramyocardial injection, which have been widely used for the delivery of various types of cells. Although there are advantages to both approaches, donor cell retention and survival are poor using these methods, potentially limiting therapeutic effects. Various attempts to improve current approaches, along with the development of emerging new approaches, are also described and discussed in this review.

Mild chronic kidney disease is an independent predictor of long-term mortality after emergency angiography and primary percutaneous intervention in patients with ST-elevation myocardial infarction

Objective Moderate renal impairment (RI) with a glomerular filtration rate (GFR) <60 ml/min/1.73 m2 is known to predict survival. The authors investigated whether mild RI with an estimated GFR of 60–89 ml/min/1.73 m2 independently predicts survival in a contemporary population with ST segment elevation myocardial infarction (STEMI). Design This is a single-centre, observational, retrospective cohort study. Patients 601 patients with STEMI who underwent emergency catheter laboratory admission met the inclusion criteria for this study. Methods Estimated glomerular filtration rate (eGFR) was obtained by the Modified Diet in Renal Disease equation, and preprocedure renal function was subdivided into chronic kidney disease stages. Univariate and multivariate Cox regression analyses were performed to assess which of 17 patient or procedural variables were independent risk factors for death. Results Longitudinal data were collated for 576 patients (96.3%). Median follow-up time was 2.6 years. 30-day and long-term death rates were 5.7% and 12.5%, respectively. Following multivariable analysis, mild RI with an eGFR of 60–89 ml/min/1.73 m2 was a strong independent predictor of death, compared with an eGFR ≥90 ml/min/1.73 m2 (HR 2.79, 95% CI 1.98 to 3.92, p<0.001), and increasing chronic kidney disease stage was a strong predictor of death after both 30 days and long-term follow-up. An eGFR of 60–89 ml/min/1.73 m2 had a greater independent effect on short- and long-term mortality than the presence of diabetes mellitus (HR 2.0, 95% CI 1.2 to 3.33). Conclusion Mild RI (eGFR=60–89 ml/min/1.73 m2) on admission is strongly predictive of short- and long-term mortality in patients with STEMI admitted to the catheter laboratory. A redefined threshold of clinically significant impairment is now required (GFR<90 ml/min/1.73 m2).

Epicardial Placement of Mesenchymal Stromal Cell-sheets for the Treatment of Ischemic Cardiomyopathy; In Vivo Proof-of-concept Study

Transplantation of bone marrow mesenchymal stromal cells (MSCs) is an emerging treatment for heart failure. We have reported that epicardial placement of MSC-sheets generated using temperature-responsive dishes markedly increases donor MSC survival and augments therapeutic effects in an acute myocardial infarction (MI) model, compared to intramyocardial (IM) injection. This study aims to expand this knowledge for the treatment of ischemic cardiomyopathy, which is likely to be more difficult to treat due to mature fibrosis and chronically stressed myocardium. Four weeks after MI, rats underwent either epicardial MSC-sheet placement, IM MSC injection, or sham treatment. At day 28 after treatment, the cell-sheet group showed augmented cardiac function improvement, which was associated with over 11-fold increased donor cell survival at both days 3 and 28 compared to IM injection. Moreover, the cell-sheet group showed improved myocardial repair, in conjunction with amplified upregulation of a group of reparative factors. Furthermore, by comparing with our own previous data, this study highlighted similar dynamics and behavior of epicardially placed MSCs in acute and chronic stages after MI, while the acute-phase myocardium may be more responsive to the stimuli from donor MSCs. These proof-of-concept data encourage further development of the MSC-sheet therapy for ischemic cardiomyopathy toward clinical application.

A simple and novel method for RNA-seq library preparation of single cell cDNA analysis by hyperactive Tn5 transposase.

Background: Deep sequencing of single cell‐derived cDNAs offers novel insights into oncogenesis and embryogenesis. However, traditional library preparation for RNA‐seq analysis requires multiple steps with consequent sample loss and stochastic variation at each step significantly affecting output. Thus, a simpler and better protocol is desirable. The recently developed hyperactive Tn5‐mediated library preparation, which brings high quality libraries, is likely one of the solutions. Results and Conclusions: Here, we tested the applicability of hyperactive Tn5‐mediated library preparation to deep sequencing of single cell cDNA, optimized the protocol, and compared it with the conventional method based on sonication. This new technique does not require any expensive or special equipment, which secures wider availability. A library was constructed from only 100 ng of cDNA, which enables the saving of precious specimens. Only a few steps of robust enzymatic reaction resulted in saved time, enabling more specimens to be prepared at once, and with a more reproducible size distribution among the different specimens. The obtained RNA‐seq results were comparable to the conventional method. Thus, this Tn5‐mediated preparation is applicable for anyone who aims to carry out deep sequencing for single cell cDNAs. Developmental Dynamics 241:1584–1590, 2012.

A highly effective technique for transseptal endocardial left ventricular lead placement for delivery of cardiac resynchronization therapy

BACKGROUND Implantation of a left ventricular (LV) lead fails in 5% to 10% of patients in whom cardiac resynchronization therapy (CRT) is attempted. Alternatives for delivery of CRT are surgical epicardial and endocardial transvenous leads. Endocardial transseptal LV lead delivery is challenging because of the absence of dedicated equipment designed for this procedure. OBJECTIVE The purpose of this study was to describe a new technique for delivery of a transseptal LV lead. METHODS This dual approach from the right femoral vein and left subclavian vein involves use of an Endrys transseptal needle and Mullins sheath to deliver a gooseneck snare from the left subclavian vein into the right atrium that can then be used to deliver a deflectable sheath into the left atrium. An active fixation lead is advanced into the LV through the sheath and screwed into the lateral wall. RESULTS The procedure was performed successfully in 12 patients in whom transvenous LV lead implantation had previously failed. The Endrys transseptal needle, ideally suited for this technique, facilitated passage of the gooseneck snare into the left atrium with no difficulty. Median procedure time was 148 minutes (interquartile range [IQR] 113-176 minutes), and median fluoroscopy time was 16 minutes (IQR 10-19 minutes). There was no need for repeat procedures after median follow-up of 97 days (IQR 36-313 days). CONCLUSION This approach using an Endrys needle and a gooseneck snare provides a reliable and effective alternative technique for delivery of an endocardial LV lead that is delivered easily through a deflectable sheath inserted transseptally into the LV.

Predictors of new onset atrial fibrillation in patients with heart failure

INTRODUCTION Stroke associated with atrial fibrillation (AF) is more frequent in heart failure. It is unknown what variables predict future AF in these patients and how AF might evolve over time. We investigated this in patients with implantable cardiac defibrillators (ICD) where AF detection is optimal. METHODS Single centre, retrospective, observational cohort study. All ischaemic cardiomyopathy patients with dual chamber, primary prevention ICD implants between Aug 2003 and Dec 2009 were screened and included if at implant, they had no known AF history. Nine variables were analysed. AF was defined as any atrial tachyarrhythmia ≥180 bpm and ≥30 s. Multivariable, binary logistic regression models were built by adding variables significant in the univariate models. Variables were retained in the final multivariate models if p<0.05. RESULTS n=197 met the inclusion criteria (85.8% male, median age: 66.8 years). After median follow-up for 2.8 years, 44.2% developed AF. After univariate analysis, the baseline variables associated with AF after implant were age, NYHA class and renal impairment (RI, defined eGFR<60 ml/min/1.73 m2) (p<0.05). After multivariable analysis, the only variable which was associated with AF was RI (HR: 2.04 (CI: 1.10-3.79)). Two baseline variables were independently associated with all-cause mortality: RI (HR: 2.42 (1.14-5.12)) and non-white ethnicity. CONCLUSION RI at time of implant was independently associated with both future AF and all-cause mortality during long-term follow-up. RI was a stronger predictor of AF than age. Those patients with heart failure and RI should be regularly screened for asymptomatic AF, regardless of age, to ensure that stroke prophylaxis may be initiated.

Telomere shortening and telomerase activity in ischaemic cardiomyopathy patients – Potential markers of ventricular arrhythmia

BACKGROUND Implantable cardioverter defibrillators (ICDs) reduce mortality in patients with ischaemic cardiomyopathy at high risk of ventricular arrhythmias (VA). However, the current indication for ICD prescription needs improvement. Telomere and telomerase in leucocytes have been shown to associate with biological ageing and pathogenesis of cardiovascular diseases. We hypothesised that leucocyte telomere length, load-of-short telomeres and/or telomerase activity are associated with VA occurrence in ischaemic cardiomyopathy patients. METHODS AND RESULTS 90 ischaemic cardiomyopathy patients with primary prevention ICDs were recruited. 35 had received appropriate therapy from the ICD for potentially-fatal VA while the remaining 55 patients had not. No significant differences in baseline demographic data relevant to telomere biology were seen between the two groups. There was no significant difference in the age and sex adjusted mean telomere length analysed by qPCR between the groups (p=0.88). In contrast, the load-of-short telomeres assessed by Universal-STELA method and telomerase activity by TRAP assay were both higher in patients who had appropriate ICD therapy and were significantly associated with incidence of ICD therapy (p=0.02, p=0.02). ROC analyses demonstrated that the sensitivity and specificity of these telomere dynamics in predicting potentially-fatal VA was higher than the current gold-standard - left ventricular ejection fraction (AUC 0.82 versus 0.47). CONCLUSION The load-of-short telomeres and telomerase activity had a significant association with ICD therapy (for VA) in ischaemic cardiomyopathy patients. These biomarkers should be tested in prospective studies to assess their clinical utility in predicting VA after myocardial infarction and guiding primary prevention ICD prescription.

Cell Size Critically Determines Initial Retention of Bone Marrow Mononuclear Cells in the Heart after Intracoronary Injection: Evidence from a Rat Model.

Intracoronary injection of bone marrow mononuclear cells (BMMNC) is an emerging treatment for heart failure. Initial donor cell retention in the heart is the key to the success of this approach, but this process remains insufficiently characterized. Although it is assumed that cell size of injected cells may influence their initial retention, no scientific evidence has been reported. We developed a unique model utilizing an ex-vivo rat heart perfusion system, enabling quantitative assessment of retention of donor cells after intracoronary injection. The initial (5 minutes after intracoronary injection) retention rate of BMMNC was as low as approximately 20% irrespective of donor cell doses injected (1×106, 8×106, 4×107). Quantitative cell-size assessment revealed a positive relationship between the size of BMMNC and retention ratio; larger subpopulations of BMMNC were more preferentially retained compared to smaller ones. Furthermore, a larger cell type—bone marrow-derived mesenchymal stromal cells (median size = 11.5μm versus 7.0μm for BMMNC)—had a markedly increased retention rate (77.5±1.8%). A positive relationship between the cell size and retention ratio was also seen in mesenchymal stromal cells. Flow-cytometric studies showed expression of cell-surface proteins, including integrins and selectin-ligands, was unchanged between pre-injection BMMNC and those exited from the heart, suggesting that biochemical interaction between donor cells and host coronary endothelium is not critical for BMMNC retention. Histological analyses showed that retained BMMNC and mesenchymal stromal cells were entrapped in the coronary vasculature and did not extravasate by 60 minutes after transplantation. Whilst BMMNC did not change coronary flow after intracoronary injection, mesenchymal stromal cells reduced it, suggesting coronary embolism, which was supported by the histological finding of intravascular cell-clump formation. These data indicate that cell-size dependent, passive (mechanical), intravascular entrapment is responsible for the initial donor cell retention after intracoronary injection of BMMNC in the heart having normal vasculatures (at least).

The lung impedance monitoring in treatment of chronic heart failure (the LIMIT-CHF study).

AIMS To assess the usefulness of intrathoracic impedance monitoring (IIM) alerts in guiding empirical treatment of chronic heart failure (CHF) patients to prevent heart failure (HF) hospitalizations and unplanned HF care. METHODS AND RESULTS Chronic heart failure patients with OptiVol or CorVue capable implantable cardioverter-defibrillators were randomized to either the active group (IIM alarm turned on and diuretic dose increased by 50% for 1 week in the event of alarm sounding) or the control group (IIM alarm turned off). The primary endpoint was the number of HF hospitalizations per patient at 1 year. The NYHA class, 6MWT, B-type natriuretic peptide (BNP), and MLWHF questionnaire score were collected at baseline and follow-up. Eighty patients were included and 71 reached 1-year follow-up. There were 1.7 ± 1.5 alerts in the active group and 1.1 ± 1.0 in the control group, P = 0.07. In the active group, 61% of alerts led to a diuretic dose increase. There was a total of 11 HF hospitalizations in the active group vs. 6 in the control group without significant differences in the number of episodes per patient (0.3 ± 0.9 vs. 0.2 ± 0.4, P = 0.95). There were no unplanned HF visits in the active group vs. 0.1 ± 0.3 per patient in the control group, P = 0.08. The total MLWHF scores were significantly increased at the final follow-up in the control group, whereas a trend towards reduction was observed in the active group. CONCLUSION In this study, an empirical HF treatment guided by IIM alerts did not reduce emergency treatment of HF. However, it seems to have a positive impact on quality of life. CLINICAL TRIAL REGISTRATIONURL: http://www.clinicaltrials.gov. Unique identifier: NCT01320007.

Association of genetic variation in telomere-related SNP and telomerase with ventricular arrhythmias in ischemic cardiomyopathy.

Telomeres are known to provide genomic stability and telomere length has been associated with cardiovascular diseases. Moreover, a higher telomerase activity has been shown to be associated with ventricular arrhythmias (VA) in ischemic cardiomyopathy. Increasing evidence suggests that genetic variation in key telomere genes has an impact on telomerase activity. Each copy of the minor allele of SNP rs12696304, at a locus including TERC (telomerase), has been associated with ∼75 base pairs reduction in mean telomere length likely mediated by an effect on TERC expression. We investigated the impact of genetic variation of this SNP on telomerase and its association with VA in ischemic cardiomyopathy patients.