Engineered 3D bioimplants using elastomeric scaffold, self-assembling peptide hydrogel, and adipose tissue-derived progenitor cells for cardiac regeneration

Abstract

Contractile restoration of myocardial scars remains a challenge with important clinical implications. Here, a combination of porous elastomeric membrane, peptide hydrogel, and subcutaneous adipose tissue-derived progenitor cells (subATDPCs) was designed and evaluated as a bioimplant for cardiac regeneration in a mouse model of myocardial infarction. SubATDPCs were doubly transduced with lentiviral vectors to express bioluminescent-fluorescent reporters driven by constitutively active, cardiac tissue-specific promoters. Cells were seeded into an engineered bioimplant consisting of a scaffold (polycaprolactone methacryloyloxyethyl ester) filled with a peptide hydrogel (PuraMatrix™), and transplanted to cover injured myocardium. Bioluminescence and fluorescence quantifications showed de novo and progressive increases in promoter expression in bioactive implant-treated animals. The bioactive implant was well adapted to the heart, and fully functional vessels traversed the myocardium-bioactive implant interface. Treatment translated into a detectable positive effect on cardiac function, as revealed by echocardiography. Thus, this novel implant is a promising construct for supporting myocardial regeneration.

Keywords: Cardiac regeneration; RECATABI; elastomeric membrane; self-assembling peptide hydrogel; subcutaneous ATDPCs.

Figure 1

BLI and fluorescent evaluation of cTnI expression and survival of subATDPCs implanted over the mouse infarcted myocardium. A: Representative BLI of labeled subATDPCs within the bioactive implant displays the luciferase signal from the cell differentiation reporter (PLuc-eGFP) that is regulated by the hcTnI promoter. Images of the hcTnI-specific reporter are superimposed on black-and-white dorsal images of the recipient animal. Color bars illustrate relative light intensities from PLuc. B: Histograms of the PLuc/RLuc ratio calculated from photon fluxes recorded via BLI from bioactive implant-treated infarcted animals. C: Representative fluorescence images from subATDPCs within the bioactive implant pre-implantation and post-sacrifice over the excised hearts. Upper images show fluorescence from the cell differentiation reporter (PLuc-eGFP) regulated by hcTnIp (green). Bottom images are representative of constitutive fluorescence from the cell-number reporter (CMVp-RLuc-mRFP1; red). Color bars illustrate the relative fluorescence intensities from eGFP (green) and RFP (red). D: Immunofluorescence staining of mouse heart cross-sections shows the bioactive implant filled with human subATDPCs. Transplanted cells were detected via RFP immunostaining (red), and cTnI expression was detected with anti-eGFP (green) and anti-cTnI (white) antibodies. (BI, bioactive implant; My, myocardium). Scale bars, 20 μm.

Figure 3

Cardiac functional analysis. Representative images and measurements of the LVEF of control-MI (A) and bioactive implant-treated animals (MI-bioimplant) (B) in short axis M-mode. (C) Percentage of LVEF relative change in infarcted groups (control-MI, MI-scaffold, and MI-bioimplant) expressed as the differences between the post-MI and pre-sacrifice values.

Figure 2

Adaptability and vascularization of the bioimplant in the mouse model of MI. A: Representative image of a heart excised from a bioactive implant-treated animal. Dotted lines indicate localization of the implant. Scale bars, 1 mm. B: Masson’s trichrome staining in a heart cross-section reveals the presence of a myocardial scar (blue) and the good adaptability of the bioactive implant. Scale bars, 1 mm. C: Cells inside the bioactive implant three days post-implantation were detected via the constitutively active reporter CMVp-RLuc-mRFP1 (red). C’: Detail of the spindle-shape morphology of subATDPCs. Scale bars, 25 μm. D and E: Functional vessel detection with FITC-dextran (green) in mice treated with subATDPCs. Arrowheads indicate a vessel connection between the mouse myocardium and the bioactive implant in a cell-treated animal. Nuclei were counterstained with Hoechst 33342. Scale bars, 20 μm and 250 μm.