3D printed neobavaisoflavone-coated scaffolds promote bone regeneration by regulating osteoblast/osteoclast activities

Abstract

BACKGROUND: Neobavaisoflavone could promote bone formation and may be a potential small molecule drug for bone regeneration. The use of 3D printed bone tissue engineering scaffolds as drug delivery carriers for neobavaisoflavone is expected to enhance the potential application of bone regeneration. OBJECTIVE: To explore the effects of polylactic acid/polydopamine/neobavaisoflavone bone scaffold on osteoclast and osteoblast activity. METHODS: (1) Fused deposition modeling technology was used to manufacture a 3D printed polylactic acid scaffold. These polylactic acid scaffolds were immersed in a dopamine solution containing or without neobavaisoflavone to produce polylactic acid/polydopamine/neobavaisoflavone scaffolds and polylactic acid/polydopamine scaffolds, respectively. The surface morphology, surface hardness, and compressive strength of the three groups of scaffolds were characterized, and the drug release properties of the polylactic acid/polydopamine/neobavaisoflavone scaffolds were investigated. (2) Mouse embryonic osteoblast MC3T3-E1 cells were co-cultured with the three groups of scaffolds. CCK-8 assay and live/dead staining were used to evaluate the cytocompatibility of the scaffolds. Transwell assay was used to evaluate the effect of scaffolds on osteoblast migration. Alkaline phosphatase quantitative assay and alizarin red staining were used to evaluate the effect of scaffolds on osteoblast differentiation. RAW264.7 cells were co-cultured with the three groups of scaffolds. After osteoclast induction, tartrate-resistant acid phosphatase staining was used to evaluate the effect of scaffolds on osteoclast differentiation. RESULTS AND CONCLUSION: (1) Scanning electron microscopy showed that all three groups of scaffolds had three-dimensional structure and regular interconnected porous structure with an average pore size of 400 µm. The surface hardness and compressive strength of polylactic acid/polydopamine scaffolds and polylactic acid/polydopamine/neobavaisoflavone scaffolds were higher than those of polylactic acid scaffolds (P < 0.05). Polylactic acid/polydopamine/neobavaisoflavone scaffolds had good drug release behavior and could continuously release drugs for more than 14 days in vitro. (2) CCK-8 assay and live/dead staining showed that all three groups of scaffolds had good cytocompatibility, and polylactic acid/polydopamine scaffolds and polylactic acid/polydopamine/neobavaisoflavone scaffolds could promote MC3T3-E1 cell proliferation. Transwell assay showed that compared with polylactic acid scaffolds, polylactic acid/polydopamine scaffolds and polylactic acid/polydopamine/neobavaisoflavone scaffolds could promote MC3T3-E1 cell migration. Alkaline phosphatase quantitative assay and alizarin red staining showed that compared with the other two groups, polylactic acid/polydopamine scaffolds and polylactic acid/polydopamine/neobavaisoflavone scaffolds could promote osteogenic differentiation of MC3T3-E1 cells. Tartrate-resistant acid phosphatase staining showed that polylactic acid/polydopamine/neobavaisoflavone scaffolds could inhibit osteoclast differentiation of RAW264.7 cells. (3) These results indicate that polylactic acid/polydopamine/neobavaisoflavone scaffolds have good biosafety and can promote bone regeneration by regulating osteoblast and osteoclast activities.

1. Introduction

Bone transplantation, as a traditional and effective method for repairing bone defects, has achieved remarkable success in clinical practice. However, with the widespread application of bone transplantation, problems such as donor shortage, surgical complications, and graft rejection have gradually become prominent, prompting research and innovation in bone graft materials [1-3]. In recent years, with the aging population and the increasing incidence of bone-related diseases, the demand for bone biomaterials has become more urgent. The rapid development of bone tissue engineering and bone repair has made the search for new, sustainable bone biomaterials a research focus [4-6].

In recent years, the rapid advancement of 3D printing technology has introduced a revolution in the field of bone transplantation. Traditional bone graft materials often face issues such as donor scarcity, morphological mismatch, and post-implantation biocompatibility problems [7]. With continuous innovation in 3D printing technology, new design and fabrication methods for bone graft materials provide new ideas to solve traditional problems. The advantage of 3D printing lies in its highly personalized manufacturing capability; by precisely controlling printing parameters, bone scaffolds can better adapt to the patient's anatomical shape, improving the adaptability between bone graft materials and surrounding tissues [8]. Furthermore, 3D printing offers opportunities to design bone graft materials with complex structures, such as mimicking the porous structure of natural bone, which positively influences cell adhesion and angiogenesis [9-10].

Although 3D printed bone scaffolds show great potential in bone defect repair and bone reconstruction, bone healing is a complex and comprehensive process that requires consideration not only of bone formation but also of osteoclastogenesis. Osteoclast-related bone resorption may be a key factor leading to unsatisfactory bone repair outcomes. Recently, an increasing number of studies have confirmed that osteoclasts play an indispensable role in bone formation, maintenance, and remodeling [11-13]. During bone mineralization, osteogenesis without osteoclast activity may inhibit bone remodeling, thereby impairing the natural mechanism of bone repair and increasing the risk of unnecessary bone overgrowth and osteosclerosis [14].