Effect of different mineralization processes on in vitro and in vivo bone regeneration and osteoblast-macrophage cross-talk in co-culture system using dual growth factor mediated non-mulberry silk fibroin grafted poly (Є-caprolactone) nanofibrous scaffold

Abstract

This study evaluates mineralized nanofibrous polymeric scaffolds at supporting in vitro osteogenic differentiation of human mesenchymal stem cells (hMSCs) and in vivo bone tissue regeneration. Co-electrospin, alternative soaking, and electrodeposition were used to introduce hydroxyapatite in non-mulberry silk fibroin grafted poly(Є-caprolactone) nanofibrous scaffolds. Bone morphogenic protein-2 and Transforming growth factor-β, at a potency ratio of 1:1, are covalently coupled onto the scaffolds. hMSCs proliferation and interactions are studied through MTT and Alamar blue assay and scanning electron and confocal microscopy. Alkaline phosphatase activity, mineralization assays, and real-time PCR studies substantiate hMSCs’ osteogenic differentiation. Co-cultures of human macrophages and osteoblasts exhibit insignificant pro-inflammatory cytokines production. In vivo trials are conducted in rabbit femur (distal metaphysis region). Bone regeneration ability of the scaffolds’ is assessed using chronological radiography, micro-CT analysis, host tissue immuno-compatibility, histology, scanning electron microscope imagery, and fluorochrome labelling. In vitro and in vivo characterizations for osteogenesis and osseointegration show best results for scaffolds mineralized by electrodeposition, followed by alternate soaking and co-electrospinning. Non-mulberry silk fibroin grafted poly(Є-caprolactone) nanofibrous scaffold, mineralized by electrodeposition, could provide promising platform for bone healing and regeneration.