Quercetin-loaded hydrogel materials for treatment of infected bone defects

Abstract

BACKGROUND: Traditional topical administration of antibiotics for infected bone defects is limited by the selective proliferation of drug-resistant strains, burst drug release, and a lack of osteoinductive activity. Simple bone repair materials are ineffective in controlling the infection process. Therefore, the development of intelligent drug delivery systems with multiple biological functions has become a research hotspot in this field. OBJECTIVE: To construct a polyethylenimine/oxidized dextran dynamic cross-linked hydrogel-loaded quercetin composite system to achieve a temporally synergistic antibacterial and osteogenic effect and to investigate the efficacy of this composite system in treating infected bone defects. METHODS: (1) Oxidized dextran was prepared by sodium periodate oxidation, and quercetin nanocrystals were prepared by antisolvent precipitation. Quercetin nanocrystals were then added to a polyethylenimine solution and a Schiff base reaction was used to prepare a quercetin-loaded polyethylenimine/oxidized dextran hydrogel. The in vitro drug release from this hydrogel was characterized. (2) In vitro experiments: Rabbit bone marrow mesenchymal stem cells were seeded onto the surfaces of polyethyleneimine/oxidized dextran hydrogels and quercetin-loaded polyethyleneimine/oxidized dextran hydrogels, respectively. Cells cultured alone served as controls, and the cytocompatibility of the materials was assessed using CCK-8 assay and live/dead cell staining. After osteogenic induction, alkaline phosphatase staining, alizarin red staining, and osteogenic gene detection were used to evaluate the osteogenic ability of the materials. Staphylococcus aureus (or Escherichia coli, methicillin-resistant Staphylococcus aureus) were co-cultured with polyethyleneimine/oxidized dextran hydrogels and quercetin-loaded polyethyleneimine/oxidized dextran hydrogels, respectively, with bacteria cultured alone as controls. The antibacterial properties of the materials were evaluated by measuring the absorbance of bacterial suspensions and colony counting on agar plates. (3) Animal experiments: An infected femoral defect model was established in SD rats by drilling a hole below the left femoral greater trochanter, extracting bone marrow, and injecting 5% sodium morrhuate plus Staphylococcus aureus suspension into the medullary cavity. Four weeks after modeling, rats were randomly divided into 3 groups for intervention: control group (n=9) received only thorough debridement, hydrogel group (n=9) and quercetin-loaded hydrogel group (n=9) received injection of polyethyleneimine/oxidized dextran hydrogel or quercetin-loaded polyethyleneimine/oxidized dextran hydrogel after debridement, respectively. At 8 weeks post-surgery, samples were harvested for Micro-CT scanning and histological observation. RESULTS AND CONCLUSION: (1) The polyethyleneimine/oxidized dextran hydrogel showed rapid drug release in the initial period (within 3 days), followed by sustained release for up to 42 days. (2) CCK-8 assay and live/dead cell staining showed that compared with polyethyleneimine/oxidized dextran hydrogel, quercetin-loaded polyethyleneimine/oxidized dextran hydrogel promoted cell proliferation and had good cytocompatibility. Bacterial suspension absorbance and colony counting showed that polyethyleneimine/oxidized dextran hydrogel had no antibacterial activity, while quercetin-loaded polyethyleneimine/oxidized dextran hydrogel significantly inhibited the growth and reproduction of Staphylococcus aureus, Escherichia coli, and methicillin-resistant Staphylococcus aureus. Alkaline phosphatase staining, alizarin red staining, and osteogenic gene detection showed that polyethyleneimine/oxidized dextran hydrogel had no osteogenic ability, while quercetin-loaded polyethyleneimine/oxidized dextran hydrogel had good osteogenic ability. (3) Micro-CT scanning showed that the quercetin-loaded hydrogel group had significantly more new bone tissue than the control and hydrogel groups; hematoxylin-eosin and Masson staining showed that the quality of bone defect repair in the quercetin-loaded hydrogel group was better than that in the control and hydrogel groups; Giemsa staining showed a large number of bacteria in the control and hydrogel groups, while almost no bacteria were observed in the quercetin-loaded hydrogel group. (4) These results indicate that quercetin-loaded polyethyleneimine/oxidized dextran hydrogel has good antibacterial and osteogenic abilities and can promote the repair of infected bone defects.

1. Introduction

Infected bone defects require simultaneous management of bacterial infection, modulation of the inflammatory microenvironment, and bone tissue regeneration. Traditional local antibiotic delivery suffers from limitations such as selective proliferation of drug-resistant strains, burst drug release, and lack of osteoinductive activity, while simple bone repair materials are inadequate for controlling infection. Therefore, the development of intelligent drug delivery systems with multiple biological functions has become a research hotspot in this field.

Quercetin, a natural flavonoid, exhibits broad-spectrum antibacterial properties, particularly against methicillin-resistant Staphylococcus aureus. It can inhibit osteoclast differentiation via the nuclear factor κB signaling pathway and promote osteoblast mineralization by activating the Wnt/β-catenin pathway. However, its clinical application is limited by poor water solubility and short half-life. Various strategies, such as nanocrystallization, liposome encapsulation, or combination with biomaterials, have been explored to improve quercetin delivery. For instance, JAFARBEGLOU et al. loaded quercetin into poly(lactic-co-glycolic acid) nanoparticles embedded in silk fibroin/chitosan hydrogels, achieving 72-hour sustained release, but with prolonged gelation time and insufficient mechanical properties. Recently, dynamic cross-linked hydrogels have attracted attention due to their self-healing ability and tunable degradation. For example, YU et al. reported a Schiff base-based gelatin/oxidized alginate hydrogel that adapts to the mechanical microenvironment of bone defects via dynamic bond reorganization, but its antibacterial function relies on exogenous antibiotics and fails to integrate the multiple advantages of natural active ingredients. Therefore, developing an effective delivery system to overcome these limitations of quercetin is crucial.