Stacked silk-cell monolayers as a biomimetic three dimensional construct for cardiac tissue reconstruction

Heart failure, due to necrosis of heart tissue, interminably poses a significant burden on world-wide health care systems. In this context, a facile tissue engineering approach using mulberry (Bombyx mori) and non-mulberry (Antheraea assama) silk fibroin (SF) has been delved into. Amalgamating the efficacious attributes of cell sheet tissue engineering and robusticity of silk biomaterial, we developed a 3-D construct using silk films to promote cardiac tissue regeneration. Herein, the fabricated patterned silk films were physico-chemically characterized and analysed for their compatibility with cardiomyocytes. The presence of nanogrooves on silk films provided contact guidance to the growing cardiomyocytes allowing them to form unidirectionally aligned cell monolayers. Non-mulberry silk films exhibiting significantly greater mechanical strength and low immunogenicity in vitro and in vivo supported better growth, proliferation and maturation of both primary rat cardiomyocytes (PCMs) and H9c2 cells. The directional cue and presence of cell binding motifs such as RGD in A. assama films favoured the growth and maturation of cardiomyocytes to their functional phenotype. Cardiomyocyte maturation was attested by significant (p≤ 0.05) upregulation of myosin heavy chain-α (∼1.25 fold), connexin 43 (∼2 fold), and troponin I (∼1.25 fold) genes in PCMs grown on non-mulberry silk films. The patterned silk-cardiomyocyte monolayers were then stacked onto each other while maintaining their alignment to form a 3-D construct which exhibited structural integrity and uniform cellular distribution. Taking together, this work attests the suitability of non-mulberry A. assama silk fibroin as a potential biomaterial and prospects of exploring silk-cardiomyocyte monolayers for cardiac tissue engineering applications.