Aligned electrospun gelatin-carbon black nanofibers for cardiac tissue modelling

Replicating the complex myocardial microenvironment remains a fundamental challenge in cardiac tissue engineering. Although nanomaterials are increasingly incorporated into electrospun scaffolds to enhance biomaterials properties, it remains unclear whether improvements in cardiac function result primarily from increased conductivity or from broader modulations of the microenvironment. Here, we investigated how carbon black nanoparticles regulate the structural, physicochemical, mechanical, electrical, and biological properties of aligned electrospun gelatin scaffolds. Aligned gelatin scaffolds containing 0.1%–2% (w/v) carbon black were fabricated with fiber diameters of 0.15–0.20 μm. Directionality analysis confirmed preferential fiber orientation in all formulations. Carbon black addition preserved scaffold hydration at approximately 500%, improved hydrolytic stability, and produced hydrated elastic moduli similar to the microscale stiffness of native myocardium. The carbon black concentrations tested did not significantly enhance bulk scaffold conductivity, despite successful nanoparticle incorporation. Nevertheless, carbon black concentration was associated with differences in selected structural, molecular, and contractile readouts. All formulations supported H9c2 cardiomyoblast metabolic activity and alignment, while primary neonatal cardiomyocyte–fibroblast co-cultures developed organized sarcomeres, elongated myotubes, maintained cardiac-marker expression, and improved spontaneous contractions. This study, integrating multiscale materials characterization with functional biological evaluation, indicates that, under the tested conditions, the differences among formulations were associated with the combined scaffold microenvironment rather than with a demonstrated conductivity-dependent mechanism. These findings suggest that Gel-CB formulations, with their alignment and mechanical characteristics, preserve primary neonatal cardiac cells phenotype and functionality over early-stages cardiac models’ development.