Henricus J. Duckers

Adipose-derived cells

Heart failure is by far the most common cause of hospitalization in Western countries, with onerous economic consequences. Cell therapy holds great promise for use in tissue regeneration and is increasingly used in an effort to improve outcomes in cardiac disease. Recently it has been shown that adipose tissue, in addition to committed adipogenic, endothelial progenitor cells and pluripotent vascular progenitor cells, also contains multipotent cell types (adipose-derived stem cells, ADSCs) that, in cell culture conditions, have shown to have an impressive developmental plasticity including the ability to undergo multilineage differentiation and self-renewal. ADSCs express multiple CD marker antigens similar to those observed on MSCs and are also capable of secreting a large number of angiogenesis-related cytokines, including vascular endothelial growth factor, granulocyte/macrophage colony stimulating factor, stromal-derived factor-1α, and hepatocyte growth factor. Adipose tissue can be harvested in large quantities with minimal morbidity in several regions of the body and, on average, 100 ml of human adipose tissue yields about 1 × 106 stem cells. Studies conducted in porcine AMI models have shown a significant LV functional improvement, with no report of any potentially fatal arrhythmias. The APOLLO trial, a prospective, double blind, randomized, placebo-controlled trial currently in the recruiting phase, is a “first-in-man” study that explores the safety and feasibility of ADSC transplantation in patients with acute MI.

Intracoronary Infusion of Allogeneic Mesenchymal Precursor Cells Directly After Experimental Acute Myocardial Infarction Reduces Infarct Size, Abrogates Adverse Remodeling, and Improves Cardiac Function

Rationale: Mesenchymal precursor cells (MPCs) are a specific Stro-3+ subpopulation of mesenchymal stem cells isolated from bone marrow. MPCs exert extensive cardioprotective effects, and are considered to be immune privileged. Objective: This study assessed the safety, feasibility, and efficacy of intracoronary delivery of allogeneic MPCs directly after acute myocardial infarction in sheep. Methods and Results: Initially, intracoronary delivery conditions were optimized in 20 sheep. These conditions were applied in a randomized study of 68 sheep with an anterior acute myocardial infarction. Coronary flow was monitored during MPC infusion, and cardiac function was assessed using invasive hemodynamics and echocardiography at baseline and during 8 weeks follow-up. Coronary flow remained within thrombolysis in myocardial infarction III definitions in all sheep during MPC infusion. Global left ventricular ejection fraction as measured by pressure–volume loop analysis deteriorated in controls to 40.7±2.6% after 8 weeks. In contrast, MPC treatment improved cardiac function to 52.8±0.7%. Echocardiography revealed significant improvement of both global and regional cardiac functions. Infarct size decreased by 40% in treated sheep, whereas infarct and border zone thickness were enhanced. Left ventricular adverse remodeling was abrogated by MPC therapy, resulting in a marked reduction of left ventricular volumes. Blood vessel density increased by >50% in the infarct and border areas. Compensatory cardiomyocyte hypertrophy was reduced in border and remote segments, accompanied by reduced collagen deposition and apoptosis. No microinfarctions in remote myocardial segments or histological abnormalities in unrelated organs were found. Conclusions: Intracoronary infusion of allogeneic MPCs is safe, feasible, and markedly effective in a large animal model of acute myocardial infarction.

Capture of circulatory endothelial progenitor cells and accelerated re-endothelialization of a bio-engineered stent in human ex vivo shunt and rabbit denudation model

Aims The Genous™ Bio-engineered R™ stent (GS) aims to promote vascular healing by capture of circulatory endothelial progenitor cells (EPCs) to the surface of the stent struts, resulting in accelerated re-endothelialization. Here, we assessed the function of the GS in comparison to bare-metal stent (BMS), when exposed to the human and animal circulation. Methods and results First, 15 patients undergoing coronary angiography received an extracorporeal femoral arteriovenous (AV) shunt containing BMS and GS. Macroscopical mural thrombi were observed in BMS, whereas GS remained visibly clean. Confocal and scanning electron microscopic (SEM) analysis of GS showed an increase in strut coverage. Quantitative polymerase chain reaction (qPCR) analysis of captured cells on the GS demonstrated increased expression of endothelial markers KDR/VEGFR2 and E-selectin, and a decrease in pro-thrombogenic markers tissue factor pathway inhibitor and plasminogen activator inhibitor-1 compared with BMS. Secondly, a similar primate AV shunt model was used to validate these findings and occlusion of BMS was observed, while GS remained patent, as demonstrated by live imaging of indium-labelled platelets. Thirdly, in an in vitro cell-capture assay, GS struts showed increased coverage by EPCs, whereas monocyte coverage remained similar to BMS. Finally, the assessment of re-endothelialization was studied in a rabbit denudation model. Twenty animals received BMS and GS in the aorta and iliac arteries for 7 days. Scanning electron microscopic analysis showed a trend towards increased strut coverage, confirmed by qPCR analysis revealing increased levels of endothelial markers (Tie2, CD34, PCD31, and P-selectin) in GS. Conclusion In this proof-of-concept study, we have demonstrated that the bio-engineered EPC-capture stent, Genous™ R™ stent, is effective in EPC capture, resulting in accelerated re-endothelialization and reduced thrombogenicity.

Final results of the HEALING IIB trial to evaluate a bio-engineered CD34 antibody coated stent (Genous™Stent) designed to promote vascular healing by capture of circulating endothelial progenitor cells in CAD patients

OBJECTIVEnTo assess the safety and efficacy of the Genous™ endothelial progenitor cell (EPC) capturing stent in conjunction with HmG-CoA-reductase inhibitors (statins) to stimulate EPC recruitment, in the treatment of patients with de novo coronary artery lesions.nnnMETHODS AND RESULTSnThe HEALING IIB study was a multi-center, prospective trial, including 100 patients. The primary efficacy endpoint was late luminal loss by QCA at 6-month follow-up (FU). Although statin therapy increased relative EPC levels by 5.6-fold, the angiographic outcome at 6 month FU was not improved in patients with an overall in-stent late luminal loss of 0.76±0.50 mm. The composite major adverse cardiac events (MACE) rate was 9.4%, whereas 6.3% clinically justified target lesion revascularizations (TLRs) were observed. 2 Patients died within the first 30 days after stent implantation due to angiographically verified in-stent thrombosis. At 12 month FU, MACE and TLR increased to 15.6% and 11.5% respectively and stabilized until 24 month FU. 18 Month angiographic FU showed a significant decrease in late luminal loss (0.67±0.54, 11.8% reduction or 10% by matched serial analysis, P=0.001).nnnCONCLUSIONnThe HEALING IIB study suggests that statin therapy in combination with the EPC capture stent does not contribute to a reduction of in-stent restenosis formation for the treatment of de novo coronary artery disease. Although concomitant statin therapy was able to stimulate EPC recruitment, it did not improve the angiographic outcome of the bio-engineered EPC capture stent. Remarkably, angiographic late loss was significantly reduced between 6 and 18 months.

Biological mechanisms of microvessel formation in advanced atherosclerosis: The big Five

Advanced atherosclerotic lesions prone to rupture are characterized by a distinct histomorphology and pathobiology that became in recent years, increasingly related to the process of intraplaque neovascularization. Molecular mechanisms that regulate angiogenesis and that are active in the plaque region may destabilize advanced lesions by promoting microvessel growth and thus providing an entry route for inflammatory cells secondary to the luminal endothelium. In addition, angiogenic factors can also define intraplaque microvessel integrity and endothelial barrier function, determining the prevalence of intraplaque hemorrhaging. Here, we aim to compose a hypothetical model for angiogenic regulation of vulnerable plaque development, based on the evidence of clinical correlation and experimental functional studies that are provided for five of the most well-described angiogenic pathways in the current literature.

Feasibility of Intracoronary GLP-1 Eluting CellBead™ Infusion in Acute Myocardial Infarction:

Cell therapy is a field of growing interest in the prevention of post acute myocardial infarction (AMI) heart failure. Stem cell retention upon local delivery to the heart, however, is still unsatisfactory. CellBeads were recently developed as a potential solution to this problem. CellBeads are 170-μm alginate microspheres that contain mesenchymal stem cells (MSCs) genetically modified to express glucagon-like peptide-1 (GLP-1) supplementary to inherent paracrine factors. GLP-1 is an incretin hormone that has both antiapoptotic and cardioprotective effects. Transplanting CellBeads in the post-AMI heart might induce cardiomyocyte salvage and ultimately abrogate adverse cardiac remodeling. We aimed to investigate the feasibility of intracoronary infusion of CellBeads in a large animal model of AMI. Four pigs were used in a pilot study to assess the maximal safe dose of CellBeads. In the remaining 21 animals, an AMI was induced by balloon occlusion of the left circumflex coronary artery for 90 min. During reperfusion, 60,000 CellBeads (n = 11), control beads (n = 4), or lactated Ringers (n = 6) were infused. Animals were sacrificed after 2 or 7 days, and the hearts were excised for histological analyses. Intracoronary infusion did not permanently affect coronary flow in any of the groups. Histological analysis revealed CellBeads containing viable MSCs up to 7 days. Viability and activity of the MSCs was confirmed by qPCR analysis that showed expression of recombinant GLP-1 and human genes after 2 and 7 days. CellBeads reduced inflammatory infiltration by 29% (p = 0.001). In addition, they decreased the extent of apoptosis by 25% (p = 0.001) after 2 days. We show that intracoronary infusion of 5 million encapsulated MSCs is safe and feasible. Also, several parameters indicate that the cells have paracrine effects, suggesting a potential therapeutic benefit of this new approach.

Regulation of Vulnerable Plaque Development by the Heme Oxygenase/Carbon Monoxide System

Plaque rupture and luminal thrombosis is the most common cause of coronary occlusion that leads to acute coronary syndromes. High-risk plaques, or vulnerable plaques, are defined as lesions that are prone to rupture, also known as thin cap fibroatheroma (TCFA), or lesions prone to erosion or with calcified cores. This review will focus mainly on the vulnerable plaque, which is thought to be the precursor of the thrombogenic or ruptured plaque. Heme oxygenase 1 (HO-1) protein expression is specifically increased in lesions with a vulnerable plaque phenotype resembling TCFAs and correlates with a rise in expression levels of intimal proinflammatory markers. Data from several human and animal studies imply an important function for HO-1 in the genetic regulation of early, as well as late atherogenesis, and plaque destabilization toward a vulnerable phenotype. Although a direct association between HO-1, vulnerable plaque development, and clinical outcome is for now missing, the correlations that have been reported for HO-1 and coronary artery disease point to a possible link.

Do we have a future with transcatheter adventitial delivery of stem cells

Critically evaluating the methodology of the adventitial delivery of stem cells, some specific options should be underlined. Adventitia as the most superficial layer, consisting of connective tissue has to be distinguished of perivascular tissues. By strict definition, an adventitia is the outermost connective tissue covering any organ, or vessel. The adventitial delivery of stem cells with a 1mm micro-needle means a delivery to superficial so called pericardial myocardium, perivascular fat tissues, including a risk of perforation and injury of soft tissues. In fact, the mapping of the artery with visualization of the three-layer vessel structure and perivascular tissues as well as pericardial space with the state-of-the-art imaging approaches including IVUS (intravascular ultrasound) or OCT (optical coherence tomography) allows to find an optimal site for injection, prevents any technical complications and improves efficacy. NOGA magnetic navigation system still remains the optimal tool for the stem cell delivery to myocardium with appropriate visualization of necrosis and peri-infarct tissues. Potentially, more advanced imaging provides a chance to deliver infusate to the adventitial layer, which is a gate to the vessel wall for inflammation as well as a source of stem and progenitor cells, and myofibroblasts.

Promoting Vascular Regeneration as an Alternative to Conventional Angioplasty-Based Intervention

Technologies in interventional Cardiology have evolved from balloon to mechanical ablation, atherectomy, stenting, and brachytherapy to current drug eluting interventional strategies. New challenges are to develop techniques that not only prevent restenosis, but also promote vascular and endothelial healing after (balloon) injury. Endothelial healing approaches range from preventing endothelial injury to restoring endothelial function and reendothelialization by pharmacotherapy and cell therapy. These novel healing strategies warrant further exploration as they may represent an alternative to drug-eluting stent approaches.