Biomimetic mineralization of novel hydroxyethyl cellulose/soy protein isolate scaffolds promote bone regeneration in vitro and in vivo.

Although various strategies have been utilized to accelerate bone regeneration in bone tissue engineering (BTE), the treatment and repair of large bone defects remains a clinical challenge worldwide. Inspired by the natural extracellular matrix of bone tissue, organic-inorganic composite scaffolds with three-dimensional (3D) porous structures, sufficient mechanical properties, excellent cytocompatibility, osteoconductivity, and osteogenic potential have received considerable attention within the field of bone engineering. In this work, a novel epichlorohydrin (ECH)-crosslinked hydroxyethyl cellulose (HEC)/soy protein isolate (SPI) porous bi-component scaffold (EHSS) with hydroxyapatite (HAp) functionalization (EHSS/HAp) was constructed for bone defect repair via the combination of lyophilization and in situ biomimetic mineralization. Systematic characterization experiments were performed to assess the morphology, HAp-forming properties, mechanical properties and degradation rate of the scaffold. The results indicated that the prepared scaffolds exhibited an interconnected porous structure, a biomimetic HAp coating on their surfaces, improved mechanical properties in compression and a controllable degradation rate. In particular, semiquantitative analysis showed that the calcium/phosphorus (Ca/P) ratio of EHSS/HAp with 70% SPI content (1.65) was similar to that of natural bone tissue (1.67) according to energy dispersive X-ray spectroscopy analysis. In vitro cell culture experiments indicated that the EHSS/HAp with 70% SPI content showed improved cytocompatibility and was suitable for MC3T3-E1 cell attachment, proliferation and growth. Consistently, in vitro osteogenic differentiation studies showed that EHSS/HAp with 70% SPI content can significantly accelerate the expression of osteogenesis-related genes (Col-1, Runx2, OPN, and OCN) during osteogenic differentiation of MC3T3-E1 cells. Furthermore, when applied to the repair of critical-sized cranial defects in a rat model, EHSS/HAp with 70% SPI was capable of significantly promoting tissue regeneration and integration with native bone tissue. Microscopic computed tomography (micro-CT) results demonstrated that the bone defect site was nearly occupied with newly formed bone at 12 weeks after implantation of EHSS/HAp with 70% SPI content into the defect. Hematoxylin and eosin (H&E) staining and Masson's trichrome staining of histological sections further confirmed that EHSS/HAp with 70% SPI markedly promoted new bone formation and maturation. Collectively, our results demonstrate the potential of EHSS/HAp scaffolds with 70% SPI for successful bone defect repair and regeneration.