Abstract: In this study, we developed a novel bilayered scaffold consisting of a bottom layer composed of the DecellularizedBovine Pericardium (DP) coated with Polyaniline Nanoparticles (PANINPs) and a top layer made of an electrospunPoly(lactic-co-glycolic acid)/Gelatin (PLGA/Gel) membrane incorporated with Vascular Endothelial Growth Factor (VEGF) and hawthorn extract. Functionally, the DP supplies native Extracellular Matrix (ECM) componentsand mechanical support, while PANINPs provide conductivity. The electrospun PLGA/Gel layer mimics fibrousECM. It incorporates bioactives, with VEGF promoting pro-angiogenic stimulation and hawthorn extract enhancing anticoagulant activity, as well as increasing surface hydrophilicity. The tissue adhesive ensures the interfacialintegrity between the two layers. Decellularization efficiency was confirmed histologically using 4′,6-diamidino-2-phenylindole (DAPI) and Hematoxylin–Eosin (H&E) staining. The DP exhibited a DNA content of 115.9 ± 47.8ng/mg DNA, compared to 982.88 ± 395.42 ng/mg in Native Pericardium (NP). The PANINPs had an average particle size of 104.94 ± 13.7 nm. The conductivity of PANINPs-coated decellularized pericardium was measured to be9.093 ± 8.6 × 10- 4 S/cm using the four-point probe method. PLGA/Gel membranes containing hawthorn extract (1%,5%, 10%, and 15% w/v) and VEGF (0.1 μg/mL, 0.5 μg/mL, and 1 μg/mL) were fabricated by electrospinning, resulting in fiber diameters between 850 and 1200 nm and pore sizes between 14 and 20 μm. The anticoagulant efficiencyof the membranes containing hawthorn extract reached 430 s in the Activated Partial Thromboplastin Time Assay(aPTT). Mechanical testing revealed a tensile strength of 22.70 ± 6.33 MPa, an elongation of 53.58 ± 10.63%, andYoung’s modulus of 0.67 ± 0.10 MPa. The scaffold also exhibited over 91% cell viability and excellent cardiomyocyte adhesion. The hemolysis ratio was determined to be 0.421 ± 0.191%, which confirms its blood compatibility.Our results indicate that the proposed bilayered scaffold can be a promising candidate for cardiac patch applications.

Key words: Cardiac patch, Bilayered scaffold, Decellularization, Electrospinning, Hawthorn extract, VEGF')">Cardiac patch, Bilayered scaffold, Decellularization, Electrospinning, Hawthorn extract, VEGF