Tissue-engineered, hydrogel-based endothelial progenitor cell therapy robustly revascularizes ischemic myocardium and preserves ventricular function

Abstract

Objectives: Cell-based angiogenic therapy for ischemic heart failure has had limited clinical impact, likely related to low cell retention (<1%) and dispersion. We developed a novel, tissue-engineered, hydrogel-based cell-delivery strategy to overcome these limitations and provide prolonged regional retention of myocardial endothelial progenitor cells at high cell dosage.

Methods: Endothelial progenitor cells were isolated from Wistar rats and encapsulated in fibrin gels. In vitro viability was quantified using a fluorescent live-dead stain of transgenic enhanced green fluorescent protein(+) endothelial progenitor cells. Endothelial progenitor cell-laden constructs were implanted onto ischemic rat myocardium in a model of acute myocardial infarction (left anterior descending ligation) for 4 weeks. Intramyocardial cell injection (2 × 10(6) endothelial progenitor cells), empty fibrin, and isolated left anterior descending ligation groups served as controls. Hemodynamics were quantified using echocardiography, Doppler flow analysis, and intraventricular pressure-volume analysis. Vasculogenesis and ventricular geometry were quantified. Endothelial progenitor cell migration was analyzed by using endothelial progenitor cells from transgenic enhanced green fluorescent protein(+) rodents.

Results: Endothelial progenitor cells demonstrated an overall 88.7% viability for all matrix and cell conditions investigated after 48 hours. Histologic assessment of 1-week implants demonstrated significant migration of transgenic enhanced green fluorescent protein(+) endothelial progenitor cells from the fibrin matrix to the infarcted myocardium compared with intramyocardial cell injection (28 ± 12.3 cells/high power field vs 2.4 ± 2.1 cells/high power field, P = .0001). We also observed a marked increase in vasculogenesis at the implant site. Significant improvements in ventricular hemodynamics and geometry were present after endothelial progenitor cell-hydrogel therapy compared with control.

Conclusions: We present a tissue-engineered, hydrogel-based endothelial progenitor cell-mediated therapy to enhance cell delivery, cell retention, vasculogenesis, and preservation of myocardial structure and function.

Figure 1

Endothelial progenitor cells demonstrate characteristic spreading, in vitro, within the EPC Fibrin Hydrogel construct 2 days following Construct (10mg/ml Fibrin, 17×10⁶ EPCs/ml) creation (eGFP⁺ EPCs, confocal microscopy, 20× magnification).

Figure 2

Immunofluorescent imaging of eGFP⁺ transgenic EPCs migration from the EPC-Fibrin Construct to the myocardium. Sections were triple stained for nuclei (DAPI, blue), eGFP (green), α-smooth muscle actin (red). 20× magnification, scale bar= 75μm.

Figure 3

Quantification of myocardial neovasculogenesis within the ischemic peri-infarct myocardium.

Figure 4

Representative immunofluorescent images demonstrating vascular density in peri-infarct ischemic myocardium following therapy. (20× magnification; green = vWF, blue = DAPI, red= αSMA)

Figure 5

Representative myocardial cross sectional images following Masson’s Trichrome staining to delineate myocardial scar. (scar=blue, myocardium=red)