Stefan Koudstaal

A Fast pH‐Switchable and Self‐Healing Supramolecular Hydrogel Carrier for Guided, Local Catheter Injection in the Infarcted Myocardium

Minimally invasive intervention strategies after myocardial infarction use state-of-the-art catheter systems that are able to combine mapping of the infarcted area with precise, local injection of drugs. To this end, catheter delivery of drugs that are not immediately pumped out of the heart is still challenging, and requires a carrier matrix that in the solution state can be injected through a long catheter, and instantaneously gelates at the site of injection. To address this unmet need, a pH-switchable supramolecular hydrogel is developed. The supramolecular hydrogel is switched into a liquid at pH > 8.5, with a viscosity low enough to enable passage through a 1-m long catheter while rapidly forming a hydrogel in contact with tissue. The hydrogel has self-healing properties taking care of adjustment to the injection site. Growth factors are delivered from the hydrogel thereby clearly showing a reduction of infarct scar in a pig myocardial infarction model.

Concise Review: Heart Regeneration and the Role of Cardiac Stem Cells

Acute myocardial infarction leads to irreversible loss of cardiac myocytes, thereby diminishing the pump function of the heart. As a result, the strenuous workload imposed on the remaining cardiac myocytes often gives rise to subsequent cell loss until the vicious circle ends in chronic heart failure (CHF). Thus, we are in need of a therapy that could ameliorate or even reverse the disease progression of CHF. Endogenous regeneration of the mammalian heart has been shown in the neonatal heart, and the discovery that it may still persist in adulthood sparked hope for novel cardioregenerative therapies. As the basis for cardiomyocyte renewal, multipotent cardiac stem/progenitor cells (CSCs) that reside in the heart have been shown to differentiate into cardiac myocytes, smooth muscle cells, and vascular endothelial cells. These CSCs do have the potential to actively regenerate the heart but clearly fail to do so after abundant and segmental loss of cells, such as what occurs with myocardial infarction. Therefore, it is vital to continue research for the most optimal therapy based on the use or in situ stimulation of these CSCs. In this review, we discuss the current status of the cardioregenerative field. In particular, we summarize the current knowledge of CSCs as the regenerative substrate in the adult heart and their use in preclinical and clinical studies to repair the injured myocardium.

Transendocardial cell injection is not superior to intracoronary infusion in a porcine model of ischaemic cardiomyopathy: a study on delivery efficiency

Stem cell therapy is a new strategy for chronic ischaemic heart disease in patients. However, no consensus exists on the most optimal delivery strategy. This randomized study was designed to assess cell delivery efficiency of three clinically relevant strategies: intracoronary (IC) and transendocardial (TE) using electromechanical mapping guidance (NOGA) compared to surgical delivery in a chronic pig model of ischaemic cardiomyopathy. Twenty‐four animals underwent delivery of 107 autologous Indium‐oxine‐labelled bone marrow‐derived mesenchymal stem cells (MSC) 4 weeks after infarction and were randomized to one of three groups (n = 8 each group): IC, TE or surgical delivery (reference group). Primary endpoint was defined as percentage (%) of injected dose per organ and assessed by in vivo gamma‐emission counting. In addition, troponin and coronary flow were assessed before and after MSC injection. Blinded endpoint analysis showed no significant difference in efficiency after surgical (16 ± 4%), IC (11 ± 1%) and TE (11 ± 3%) (P = 0.52) injections. IC showed less variability in efficiency compared with TE and surgical injection. Overall, TE injection showed less distribution of MSC to visceral organs compared with other modalities. Troponin rise and IC flow did not differ between the percutaneous groups. This randomized study showed no significant difference in cell delivery efficiency to the myocardium in a clinically relevant ischaemic large animal model between IC and TE delivery. In addition, no differences in safety profile were observed. These results are important in view of the choice of percutaneous cell delivery modality in future clinical stem cell trials.

Cell Therapy, a Novel Remedy for Dilated Cardiomyopathy? A Systematic Review

BACKGROUND Dilated cardiomyopathy (DCM) is the most common form of nonischemic cardiomyopathy worldwide and can lead to sudden cardiac death and heart failure. Despite ongoing advances made in the treatment of DCM, improvement of outcome remains problematic. Stem cell therapy has been extensively studied in preclinical and clinical models of ischemic heart disease, showing potential benefit. DCM is associated with a major health burden, and few studies have been performed on cell therapy for DCM. In this systematic review we aimed to provide an overview of preclinical and clinical studies performed on cell therapy for DCM. METHODS AND RESULTS A systematic search, critical appraisal, and summarized outcomes are presented. In total, 29 preclinical and 15 clinical studies were included. Methodologic quality of reported studies in general was low based on the Centre for Evidence Based Medicine, Oxford University, criteria. A large heterogeneity in inclusion criteria, procedural characteristics, and outcome measures was noted. The majority of studies showed a significant increase in left ventricular ejection fraction after cell therapy during follow-up. CONCLUSIONS Stem cell therapy has shown moderate but significant effects in clinical trials for ischemic heart disease, but it remains to be determined if we can extrapolate these results to DCM patients. There is a need for methodologically sound studies to elucidate underlying mechanisms and translate those into improved therapy for clinical practice. To validate safety and efficacy of cell therapy for DCM, adequate randomized (placebo) controlled trials using different strategies are mandatory.

Necrostatin‐1 alleviates reperfusion injury following acute myocardial infarction in pigs

In rodents, it has previously been shown that necrostatin‐1 (Nec‐1) inhibits RIP1, a central regulator of programmed necrosis, thereby decreasing cell death and reducing infarct size (IS) after ischaemia/reperfusion (I/R) injury. To address unanswered questions on feasibility and efficacy of Nec‐1 in a large animal model, we assessed the effects of Nec‐1 in a porcine I/R model, relevant to human disease.

Myocardial Infarction and Functional Outcome Assessment in Pigs

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One important prerequisite is a good understanding of all safety and efficacy aspects obtained in a large animal model that validly reflect the human scenario of myocardial infarction (MI). Pigs are widely used in this regard since their cardiac size, hemodynamics, and coronary anatomy are close to that of humans. Here, we present an effective protocol for using the porcine MI model using a closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. This approach is based on 90 min of myocardial ischemia, inducing large left ventricle infarction of the anterior, septal and inferoseptal walls. Furthermore, we present protocols for various measures of outcome that provide a wide range of information on the heart, such as cardiac systolic and diastolic function, hemodynamics, coronary flow velocity, microvascular resistance, and infarct size. This protocol can be easily tailored to meet study specific requirements for the validation of novel cardioregenerative biologics at different stages (i.e. directly after the acute ischemic insult, in the subacute setting or even in the chronic MI once scar formation has been completed). This model therefore provides a useful translational tool to study MI, subsequent adverse remodeling, and the potential of novel cardioregenerative agents.

Assessment of coronary microvascular resistance in the chronic infarcted pig heart

Pre‐clinical studies aimed at treating ischemic heart disease (i.e. stem cell‐ and growth factor therapy) often consider restoration of the impaired microvascular circulation as an important treatment goal. However, serial in vivo measurement hereof is often lacking. The purpose of this study was to evaluate the applicability of intracoronary pressure and flow velocity as a measure of microvascular resistance in a large animal model of chronic myocardial infarction (MI). Myocardial infarction was induced in Dalland Landrace pigs (n = 13; 68.9 ± 4.1 kg) by a 75‐min. balloon occlusion of the left circumflex artery (LCX). Intracoronary pressure and flow velocity parameters were measured simultaneously at rest and during adenosine‐induced hyperemia, using the Combowire (Volcano) before and 4 weeks after MI. Various pressure‐ and/or flow‐derived indices were evaluated. Hyperemic microvascular resistance (HMR) was significantly increased by 28% in the infarct‐related artery, based on a significantly decreased peak average peak flow velocity (pAPV) by 20% at 4 weeks post‐MI (P = 0.03). Capillary density in the infarct zone was decreased compared to the remote area (658 ± 207/mm2 versus 1650 ± 304/mm2, P = 0.017). In addition, arterioles in the infarct zone showed excessive thickening of the alpha smooth muscle actin (αSMA) positive cell layer compared to the remote area (33.55 ± 4.25 μm versus 14.64 ± 1.39 μm, P = 0.002). Intracoronary measurement of HMR successfully detected increased microvascular resistance that might be caused by the loss of capillaries and arteriolar remodelling in the chronic infarcted pig heart. Thus, HMR may serve as a novel outcome measure in pre‐clinical studies for serial assessment of microvascular circulation.

Advanced measurement techniques of regional myocardial function to assess the effects of cardiac regenerative therapy in different models of ischaemic cardiomyopathy

Cardiac regenerative therapy is still not used in daily clinical practice. A reason for this might be the modest effect on relevant global clinical endpoints [i.e. ejection fraction (EF)] in preclinical studies. To introduce proper improvement strategies, it is important to extend the focus from clinical endpoints to more detailed local measures of cardiac function. In this review, we discuss the measurement principles of all invasive and non-invasive techniques that are used to assess the local effects of cardiac regenerative therapy in order to improve feedback to researchers unravelling the dominant pathways that lead to effective cardiac regeneration. Generally adopted mechanisms of cardiac regenerative therapy are: (i) vasculogenesis, (ii) cardiomyogenesis, and (iii) matrix-assisted myocardium stabilization. Since direct in vivo measures of these mechanisms do not exist, we discuss the measurement techniques of local microvascular resistance, myocardial perfusion, viability, fibrosis, and deformation imaging. The ability of these techniques to reflect the mechanism of cardiac regenerative therapy, and the results of applications in stem cell studies are discussed, and critically commented upon. Special attention is given to applications of deformation imaging, since this has recently been suggested and used as a potential new technique to assess local changes of cardiac biomechanics, which requires special knowledge about cardiac physiology. We conclude that besides the clinically relevant EF measurements, detailed measures of local cardiac function provide information about the local changes induced by cardiac regenerative therapy. In particular, combination of deformation imaging, by ultrasound or magnetic resonance imaging, with simultaneously measured local geometry and pressure measurements is a promising approach to assess the effects of cardiac regenerative therapy on local cardiac biomechanics. This approach provides information about local tissue contractility, stiffness, and thereby remodelling. We recommend that researchers use this comprehensive approach in future studies.

Xenotransplantation of Human Cardiomyocyte Progenitor Cells Does Not Improve Cardiac Function in a Porcine Model of Chronic Ischemic Heart Failure. Results from a Randomized, Blinded, Placebo Controlled Trial.

Background Recently cardiomyocyte progenitor cells (CMPCs) were successfully isolated from fetal and adult human hearts. Direct intramyocardial injection of human CMPCs (hCMPCs) in experimental mouse models of acute myocardial infarction significantly improved cardiac function compared to controls. Aim Here, our aim was to investigate whether xenotransplantation via intracoronary infusion of fetal hCMPCs in a pig model of chronic myocardial infarction is safe and efficacious, in view of translation purposes. Methods & Results We performed a randomized, blinded, placebo controlled trial. Four weeks after ischemia/reperfusion injury by 90 minutes of percutaneous left anterior descending artery occlusion, pigs (n = 16, 68.5 ± 5.4 kg) received intracoronary infusion of 10 million fetal hCMPCs or placebo. All animals were immunosuppressed by cyclosporin (CsA). Four weeks after infusion, endpoint analysis by MRI displayed no difference in left ventricular ejection fraction, left ventricular end diastolic and left ventricular end systolic volumes between both groups. Serial pressure volume (PV-)loop and echocardiography showed no differences in functional parameters between groups at any timepoint. Infarct size at follow-up, measured by late gadolinium enhancement MRI showed no difference between groups. Intracoronary pressure and flow measurements showed no signs of coronary obstruction 30 minutes after cell infusion. No premature death occurred in cell treated animals. Conclusion Xenotransplantation via intracoronary infusion of hCMPCs is feasible and safe, but not associated with improved left ventricular performance and infarct size compared to placebo in a porcine model of chronic myocardial infarction.

Circulating Extracellular Vesicles Contain miRNAs and are Released as Early Biomarkers for Cardiac Injury

Plasma-circulating microRNAs have been implicated as novel early biomarkers for myocardial infarction (MI) due to their high specificity for cardiac injury. For swift clinical translation of this potential biomarker, it is important to understand their temporal and spatial characteristics upon MI. Therefore, we studied the temporal release, potential source, and transportation of circulating miRNAs in different models of ischemia reperfusion (I/R) injury. We demonstrated that extracellular vesicles are released from the ischemic myocardium upon I/R injury. Moreover, we provided evidence that cardiac and muscle-specific miRNAs are transported by extracellular vesicles and are rapidly detectable in plasma. Since these vesicles are enriched for the released miRNAs and their detection precedes traditional damage markers, they hold great potential as specific early biomarkers for MI.