Tissue-engineered pro-angiogenic fibroblast scaffold improves myocardial perfusion and function and limits ventricular remodeling after infarction

Abstract

Objective: Microvascular malperfusion after myocardial infarction leads to infarct expansion, adverse remodeling, and functional impairment. Native reparative mechanisms exist but are inadequate to vascularize ischemic myocardium. We hypothesized that a 3-dimensional human fibroblast culture (3DFC) functions as a sustained source of angiogenic cytokines, thereby augmenting native angiogenesis and limiting adverse effects of myocardial ischemia.

Methods: Lewis rats underwent ligation of the left anterior descending coronary artery to induce heart failure; experimental animals received a 3DFC scaffold to the ischemic region. Border-zone tissue was analyzed for the presence of human fibroblast surface protein, vascular endothelial growth factor, and hepatocyte growth factor. Cardiac function was assessed with echocardiography and pressure-volume conductance. Hearts underwent immunohistochemical analysis of angiogenesis by co-localization of platelet endothelial cell adhesion molecule and alpha smooth muscle actin and by digital analysis of ventricular geometry. Microvascular angiography was performed with fluorescein-labeled lectin to assess perfusion.

Results: Immunoblotting confirmed the presence of human fibroblast surface protein in rats receiving 3DFC, indicating survival of transplanted cells. Increased expression of vascular endothelial growth factor and hepatocyte growth factor in experimental rats confirmed elution by the 3DFC. Microvasculature expressing platelet endothelial cell adhesion molecule/alpha smooth muscle actin was increased in infarct and border-zone regions of rats receiving 3DFC. Microvascular perfusion was also improved in infarct and border-zone regions in these rats. Rats receiving 3DFC had increased wall thickness, smaller infarct area, and smaller infarct fraction. Echocardiography and pressure-volume measurements showed that cardiac function was preserved in these rats.

Conclusions: Application of a bioengineered 3DFC augments native angiogenesis through delivery of angiogenic cytokines to ischemic myocardium. This yields improved microvascular perfusion, limits infarct progression and adverse remodeling, and improves ventricular function.

FIGURE 1

Scanning electron microscopy images of the 3-dimensional fibroblast culture (3DFC). The 3DFC consists of a bioabsorbable scaffold, extracellular matrix proteins, and viable human dermal fibroblasts. A, 99×; B, 200×; C, 1000×.

FIGURE 2

A, In vitro VEGF-A concentration steadily increased over 24 hours of 3DFC culture. B, In vitro HGF concentration followed a similar pattern. VEGF, Vascular endothelial growth factor; HGF, hepatocyte growth factor.

FIGURE 3

A, Immunoblotting directed against human fibroblast surface protein 4 weeks after 3DFC scaffold implantation provides evidence of human fibroblast survival in experimental animals. B, Experimental animals exhibited statistically significant upregulation in vascular endothelial growth factor A expression 4 weeks after 3DFC implantation. C, 3DFC-treated animals also exhibited increase hepatocyte growth factor expression at 4 weeks. 3DFC, 3-dimensional human dermal fibroblast culture.

FIGURE 4

Representative images of myocardial sections depicting platelet endothelial cell adhesion molecule (PECAM, orange) and alpha smooth muscle actin (α-SMA, green) in control (A) and 3DFC-treated hearts (B).

FIGURE 5

After perfusion with lectin, 3-dimensional image stacks were obtained with confocal microscopy of control, remote myocardium (A), control, peri-infarct border-zone myocardium (B), control, infarct myocardium (C), 3DFC-treated, remote myocardium (D), 3DFC-treated, peri-infarct border-zone myocardium (E), and 3DFC-treated, infarct myocardium (F).

FIGURE 6

Representative pressure–volume loops obtained during steady state measurements from control (A) and 3DFC-treated animals (B). Representative pressure–volume loops obtained during inferior vena caval occlusion from control (C) and 3DFC-treated animals (D).