Effects of Combined Hesperidin and BMP-2 Treatment on Craniofacial Bone Regeneration

Bone loss, especially in the jaw and alveolar region, is a silent threat to oral health. When left unchecked, the resulting reduction in bone density and volume can cause teeth to shift, loosen, and ultimately be lost, leading to partial or complete edentulism. Other complications include delayed post-extraction healing, worsening periodontal disease, and reduced success rates of dental implants. In response to the challenges posed by insufficient bone volume and quality, researchers have investigated biologic approaches to stimulate bone formation. Recombinant human bone morphogenetic protein-2 (BMP-2) emerged as a promising therapeutic agent. Despite BMP-2's efficacy in inducing bone growth, there have been numerous associated risks with its use (e.g., an increased risk of inflammation, ectopic bone growth, bone cyst formation, and even certain cancers). As BMP-2 is one of the few FDA-approved drugs to induce bone growth, this study aims to determine how BMP-2 can be used without its adverse clinical side effects.  Current literature points to hesperidin, a flavanone glycoside found primarily in citrus fruits, as a potential adjuvant. By itself, hesperidin is a compound that has demonstrated anti-inflammatory, antioxidant, anti-carcinogenic, and osteogenic properties. The question this study seeks to answer if HE can mitigate BMP-2, and understand how this potential mitigation occurs by comparing the type and maturity of bone formed after BMP-2 treatment with or without HE in a rat model. This study used a 5 mm mandibular critical-size defect in Sprague-Dawley rats. The rats were allocated into three groups: collagen scaffold (control), scaffold + 1 µg of BMP-2 with (BMP-2 + HE), and without dietary HE (BMP-2 only) at 100 mg/kg. Rats in the BMP-2 + HE group were given HE once daily via oral gavage for four weeks pre-mandible surgery and continued until euthanasia 6 weeks post-surgery. The rats were then sacrificed. Mandibles and femurs were harvested. The mandibles were stained by hematoxylin & eosin (H&E) and underwent histomorphometry to assess post-operative healing and bone quality. The femurs were stained with picrosirius red (PSR). The images were used to quantify red, yellow, and green color signals using MATLAB to evaluate DH’s effect on collagen maturity.    At six weeks, BMP-2 and BMP-2 + HE groups showed complete defect closure, while the control group did not. The BMP group demonstrated cyst and ectopic bone formation, unlike in the BMP-2 + HE group. PSR image quantifications showed significantly higher red color percentage in the BMP-2 + HE group compared to the BMP-2 only group (p < 0.05), indicating HE resulted in more mature collagen.    The results indicate the modulatory effect of hesperidin, its effect on mitigating BMP-2 side effects, and its potential in improving BMP-2-induced bone regeneration outcomes.