Non-mulberry silk gland fibroin protein 3-D scaffold for enhanced differentiation of human mesenchymal stem cells into osteocytes

This study investigates the potential of three-dimensional (3-D) scaffolds of wild non-mulberry tropical tasar silk gland fibroin protein as substratum for osteogenic differentiation of human mesenchymal stem cells (hMSCs). The novelty of the study lies in the fabrication of scaffolds from non-bioengineered silk fibroin directly extracted from the glands of non-mulberry tropical tasar silkworms using sodium dodecyl sulfate dissolution protocol and its osteogenic application using single- and double-seeding methods. The scaffolds were mechanically robust and showed homogenous pore distribution within the scaffold. hMSCs were seeded on the scaffolds and were cultured for up to 28 days under static conditions in osteogenic media. Osteogenic differentiation of hMSCs seeded on fibroin scaffolds resulted in extensive mineralization with the formation of large calcium nodules, higher alkaline phosphatase activity and intense von Kossa staining. Real-time studies revealed higher transcript levels for osteopontin (OS) and bone sialoprotein (IBSP) under double-seeded conditions as compared to single-seeded scaffolds. Histological analysis showed the development of osteoblastic cells and large calcified nodules. The development and spreading of nuclei and actin filaments on fibroin matrices were revealed through confocal studies. The results suggest the suitability of non-mulberry silk-fibroin protein 3-D scaffolds as natural biomaterial for potential in vitro bone-tissue engineering applications.