Jay B. Patel

Sustained Release of Engineered Stromal Cell–Derived Factor 1-α From Injectable Hydrogels Effectively Recruits Endothelial Progenitor Cells and Preserves Ventricular Function After Myocardial Infarction

Background— Exogenously delivered chemokines have enabled neovasculogenic myocardial repair in models of ischemic cardiomyopathy; however, these molecules have short half-lives in vivo. In this study, we hypothesized that the sustained delivery of a synthetic analog of stromal cell–derived factor 1-&agr; (engineered stromal cell–derived factor analog [ESA]) induces continuous homing of endothelial progenitor cells and improves left ventricular function in a rat model of myocardial infarction. Methods and Results— Our previously designed ESA peptide was synthesized by the addition of a fluorophore tag for tracking. Hyaluronic acid was chemically modified with hydroxyethyl methacrylate to form hydrolytically degradable hydrogels through free-radical–initiated crosslinking. ESA was encapsulated in hyaluronic acid hydrogels during gel formation, and then ESA release, along with gel degradation, was monitored for more than 4 weeks in vitro. Chemotactic properties of the eluted ESA were assessed at multiple time points using rat endothelial progenitor cells in a transwell migration assay. Finally, adult male Wistar rats (n=33) underwent permanent ligation of the left anterior descending (LAD) coronary artery, and 100 µL of saline, hydrogel alone, or hydrogel+25 µg ESA was injected into the borderzone. ESA fluorescence was monitored in animals for more than 4 weeks, after which vasculogenic, geometric, and functional parameters were assessed to determine the therapeutic benefit of each treatment group. ESA release was sustained for 4 weeks in vitro, remained active, and enhanced endothelial progenitor cell chemotaxis. In addition, ESA was detected in the rat heart >3 weeks when delivered within the hydrogels and significantly improved vascularity, ventricular geometry, ejection fraction, cardiac output, and contractility compared with controls. Conclusions— We have developed a hydrogel delivery system that sustains the release of a bioactive endothelial progenitor cell chemokine during a 4-week period that preserves ventricular function in a rat model of myocardial infarction.

Spatially Oriented, Temporally Sequential Smooth Muscle Cell-Endothelial Progenitor Cell Bi-Level Cell Sheet Neovascularizes Ischemic Myocardium

Background— Endothelial progenitor cells (EPCs) possess robust therapeutic angiogenic potential, yet may be limited in the capacity to develop into fully mature vasculature. This problem might be exacerbated by the absence of a neovascular foundation, namely pericytes, with simple EPC injection. We hypothesized that coculturing EPCs with smooth muscle cells (SMCs), components of the surrounding vascular wall, in a cell sheet will mimic the native spatial orientation and interaction between EPCs and SMCs to create a supratherapeutic angiogenic construct in a model of ischemic cardiomyopathy. Methods and Results— Primary EPCs and SMCs were isolated from Wistar rats. Confluent SMCs topped with confluent EPCs were spontaneously detached from the Upcell dish to create an SMC-EPC bi-level cell sheet. A rodent ischemic cardiomyopathy model was created by ligating the left anterior descending coronary artery. Rats were then immediately divided into 3 groups: cell-sheet transplantation (n=14), cell injection (n=12), and no treatment (n=13). Cocultured EPCs and SMCs stimulated an abundant release of multiple cytokines in vitro. Increased capillary density and improved blood perfusion in the borderzone elucidated the significant in vivo angiogenic potential of this technology. Most interestingly, however, cell fate–tracking experiments demonstrated that the cell-sheet EPCs and SMCs directly migrated into the myocardium and differentiated into elements of newly formed functional vasculature. The robust angiogenic effect of this cell sheet translated to enhanced ventricular function as demonstrated by echocardiography. Conclusions— Spatially arranged EPC-SMC bi-level cell-sheet technology facilitated the natural interaction between EPCs and SMCs, thereby creating structurally mature, functional microvasculature in a rodent ischemic cardiomyopathy model, leading to improved myocardial function.

Preclinical Evaluation of the Engineered Stem Cell Chemokine Stromal Cell–Derived Factor 1α Analog in a Translational Ovine Myocardial Infarction ModelNovelty and Significance

Rationale: After myocardial infarction, there is an inadequate blood supply to the myocardium, and the surrounding borderzone becomes hypocontractile. Objective: To develop a clinically translatable therapy, we hypothesized that in a preclinical ovine model of myocardial infarction, the modified endothelial progenitor stem cell chemokine, engineered stromal cell–derived factor 1&agr; analog (ESA), would induce endothelial progenitor stem cell chemotaxis, limit adverse ventricular remodeling, and preserve borderzone contractility. Methods and Results: Thirty-six adult male Dorset sheep underwent permanent ligation of the left anterior descending coronary artery, inducing an anteroapical infarction, and were randomized to borderzone injection of saline (n=18) or ESA (n=18). Ventricular function, geometry, and regional strain were assessed using cardiac MRI and pressure–volume catheter transduction. Bone marrow was harvested for in vitro analysis, and myocardial biopsies were taken for mRNA, protein, and immunohistochemical analysis. ESA induced greater chemotaxis of endothelial progenitor stem cells compared with saline (P<0.01) and was equivalent to recombinant stromal cell–derived factor 1&agr; (P=0.27). Analysis of mRNA expression and protein levels in ESA-treated animals revealed reduced matrix metalloproteinase 2 in the borderzone (P<0.05), with elevated levels of tissue inhibitor of matrix metalloproteinase 1 and elastin in the infarct (P<0.05), whereas immunohistochemical analysis of borderzone myocardium showed increased capillary and arteriolar density in the ESA group (P<0.01). Animals in the ESA treatment group also had significant reductions in infarct size (P<0.01), increased maximal principle strain in the borderzone (P<0.01), and a steeper slope of the end-systolic pressure–volume relationship (P=0.01). Conclusions: The novel, biomolecularly designed peptide ESA induces chemotaxis of endothelial progenitor stem cells, stimulates neovasculogenesis, limits infarct expansion, and preserves contractility in an ovine model of myocardial infarction.

Preclinical Evaluation of the Engineered Stem Cell Chemokine Stromal Cell–Derived Factor 1α Analog in a Translational Ovine Myocardial Infarction Model

Rationale: After myocardial infarction, there is an inadequate blood supply to the myocardium, and the surrounding borderzone becomes hypocontractile. Objective: To develop a clinically translatable therapy, we hypothesized that in a preclinical ovine model of myocardial infarction, the modified endothelial progenitor stem cell chemokine, engineered stromal cell–derived factor 1&agr; analog (ESA), would induce endothelial progenitor stem cell chemotaxis, limit adverse ventricular remodeling, and preserve borderzone contractility. Methods and Results: Thirty-six adult male Dorset sheep underwent permanent ligation of the left anterior descending coronary artery, inducing an anteroapical infarction, and were randomized to borderzone injection of saline (n=18) or ESA (n=18). Ventricular function, geometry, and regional strain were assessed using cardiac MRI and pressure–volume catheter transduction. Bone marrow was harvested for in vitro analysis, and myocardial biopsies were taken for mRNA, protein, and immunohistochemical analysis. ESA induced greater chemotaxis of endothelial progenitor stem cells compared with saline (P<0.01) and was equivalent to recombinant stromal cell–derived factor 1&agr; (P=0.27). Analysis of mRNA expression and protein levels in ESA-treated animals revealed reduced matrix metalloproteinase 2 in the borderzone (P<0.05), with elevated levels of tissue inhibitor of matrix metalloproteinase 1 and elastin in the infarct (P<0.05), whereas immunohistochemical analysis of borderzone myocardium showed increased capillary and arteriolar density in the ESA group (P<0.01). Animals in the ESA treatment group also had significant reductions in infarct size (P<0.01), increased maximal principle strain in the borderzone (P<0.01), and a steeper slope of the end-systolic pressure–volume relationship (P=0.01). Conclusions: The novel, biomolecularly designed peptide ESA induces chemotaxis of endothelial progenitor stem cells, stimulates neovasculogenesis, limits infarct expansion, and preserves contractility in an ovine model of myocardial infarction.

Layered smooth muscle cell–endothelial progenitor cell sheets derived from the bone marrow augment postinfarction ventricular function

Objective: The angiogenic potential of endothelial progenitor cells (EPCs) may be limited by the absence of their natural biologic foundation, namely smooth muscle pericytes. We hypothesized that joint delivery of EPCs and smooth muscle cells (SMCs) in a novel, totally bone marrow–derived cell sheet will mimic the native architecture of a mature blood vessel and act as an angiogenic construct to limit post infarction ventricular remodeling. Methods: Primary EPCs and mesenchymal stem cells were isolated from bone marrow of Wistar rats. Mesenchymal stem cells were transdifferentiated into SMCs by culture on fibronectin‐coated culture dishes. Confluent SMCs topped with confluent EPCs were detached from an Upcell dish to create a SMC‐EPC bi‐level cell sheet. A rodent model of ischemic cardiomyopathy was then created by ligating the left anterior descending artery. Rats were randomized into 3 groups: cell sheet transplantation (n = 9), no treatment (n = 12), or sham surgery control (n = 7). Results: Four weeks postinfarction, mature vessel density tended to increase in cell sheet‐treated animals compared with controls. Cell sheet therapy significantly attenuated the extent of cardiac fibrosis compared with that of the untreated group (untreated vs cell sheet, 198 degrees [interquartile range (IQR), 151‐246 degrees] vs 103 degrees [IQR, 92‐113 degrees], P = .04). Furthermore, EPC‐SMC cell sheet transplantation attenuated myocardial dysfunction, as evidenced by an increase in left ventricular ejection fraction (untreated vs cell sheet vs sham, 33.5% [IQR, 27.8%‐35.7%] vs 45.9% [IQR, 43.6%‐48.4%] vs 59.3% [IQR, 58.8%‐63.5%], P = .001) and decreases in left ventricular dimensions. Conclusions: The bone marrow‐derived, spatially arranged SMC‐EPC bi‐level cell sheet is a novel, multilineage cellular therapy obtained from a translationally practical source. Interactions between SMCs and EPCs augment mature neovascularization, limit adverse remodeling, and improve ventricular function after myocardial infarction.

An innovative biologic system for photon-powered myocardium in the ischemic heart

Solar-powered heart? Harnessing light to create myocardial renewable energy. Coronary artery disease is one of the most common causes of death and disability, afflicting more than 15 million Americans. Although pharmacological advances and revascularization techniques have decreased mortality, many survivors will eventually succumb to heart failure secondary to the residual microvascular perfusion deficit that remains after revascularization. We present a novel system that rescues the myocardium from acute ischemia, using photosynthesis through intramyocardial delivery of the cyanobacterium Synechococcus elongatus. By using light rather than blood flow as a source of energy, photosynthetic therapy increases tissue oxygenation, maintains myocardial metabolism, and yields durable improvements in cardiac function during and after induction of ischemia. By circumventing blood flow entirely to provide tissue with oxygen and nutrients, this system has the potential to create a paradigm shift in the way ischemic heart disease is treated.

SDF 1-alpha Attenuates Myocardial Injury Without Altering the Direct Contribution of Circulating Cells

Stromal cell-derived factor 1-alpha (SDF) is a potent bone marrow chemokine capable of recruiting circulating progenitor populations to injured tissue. SDF has known angiogenic capabilities, but bone marrow-derived cellular contributions to tissue regeneration remain controversial. Bone marrow from DsRed-transgenic donors was transplanted into recipients to lineage-trace circulating cells after myocardial infarction (MI). SDF was delivered post-MI, and hearts were evaluated for recruitment and plasticity of bone marrow-derived populations. SDF treatment improved ventricular function, border zone vessel density, and CD31+ cell frequency post-MI. Bone marrow-derived endothelial cells were observed; these cells arose through both cell fusion and transdifferentiation. Circulating cells also adopted cardiomyocyte fates, but such events were exceedingly rare and almost exclusively resulted from cell fusion. SDF did not significantly alter the proportion of circulating cells that adopted non-hematopoietic fates. Mechanistic insight into the governance of circulating cells is essential to realizing the full potential of cytokine therapies.

Bioengineered SDF-1a Analogue Delivered as an Angiogenic Therapy Significantly Normalizes Elastic and Viscoelastic Material Properties of Infarcted Cardiac Muscle

Heart disease is a leading cause of death worldwide, and coronary heart disease causes 1 of every 6 deaths in the United States [1]. Following a myocardial infarction, scar tissue gradually replaces myocardium that is lost through a process of collagen deposition and an increase in tensile strength of the tissue [2]. This leads to infarct expansion, adverse ventricular remodeling and dysfunction, and ultimately heart failure. Dilation of the left ventricle (LV) leads to increased LV wall stress and is ultimately responsible for adverse ventricular remodeling. LV dilation causes stretching and thereby increased wall stress, prohibiting cardiomyocytes from effectively contracting, which leads to further dilation, and ultimately a decrease in cardiac pump efficiency [3]. Previously, it has been shown that using a tissue filler to modify the material properties of an infarct limits the process of ventricular remodeling [4]. Angiogenesis is another mechanism by which adverse ventricular remodeling can be limited. Previously, our group developed engineered stromal cell-derived factor-1α (ESA), a computationally designed analog of an established endothelial progenitor cell chemokine, SDF-1α, and demonstrated that ESA injection enhances LV function by promoting angiogenesis and retains the native properties of the extracellular matrix (ECM) [5] [6]. In this study, we propose that injection of ESA to infarcted cardiac muscle improves the tensile strength and viscoelastic properties of ventricular muscle.Copyright

Preclinical Evaluation of the Engineered Stem Cell Chemokine Stromal Cell-Derived Factor 1-alpha Analogue in a Translational Ovine Myocardial Infarction Model

Rationale: After myocardial infarction, there is an inadequate blood supply to the myocardium, and the surrounding borderzone becomes hypocontractile. Objective: To develop a clinically translatable therapy, we hypothesized that in a preclinical ovine model of myocardial infarction, the modified endothelial progenitor stem cell chemokine, engineered stromal cell–derived factor 1&agr; analog (ESA), would induce endothelial progenitor stem cell chemotaxis, limit adverse ventricular remodeling, and preserve borderzone contractility. Methods and Results: Thirty-six adult male Dorset sheep underwent permanent ligation of the left anterior descending coronary artery, inducing an anteroapical infarction, and were randomized to borderzone injection of saline (n=18) or ESA (n=18). Ventricular function, geometry, and regional strain were assessed using cardiac MRI and pressure–volume catheter transduction. Bone marrow was harvested for in vitro analysis, and myocardial biopsies were taken for mRNA, protein, and immunohistochemical analysis. ESA induced greater chemotaxis of endothelial progenitor stem cells compared with saline (P<0.01) and was equivalent to recombinant stromal cell–derived factor 1&agr; (P=0.27). Analysis of mRNA expression and protein levels in ESA-treated animals revealed reduced matrix metalloproteinase 2 in the borderzone (P<0.05), with elevated levels of tissue inhibitor of matrix metalloproteinase 1 and elastin in the infarct (P<0.05), whereas immunohistochemical analysis of borderzone myocardium showed increased capillary and arteriolar density in the ESA group (P<0.01). Animals in the ESA treatment group also had significant reductions in infarct size (P<0.01), increased maximal principle strain in the borderzone (P<0.01), and a steeper slope of the end-systolic pressure–volume relationship (P=0.01). Conclusions: The novel, biomolecularly designed peptide ESA induces chemotaxis of endothelial progenitor stem cells, stimulates neovasculogenesis, limits infarct expansion, and preserves contractility in an ovine model of myocardial infarction.