# Bone-Regenerating Capacity of Chitosan Membrane and Chitosan Foam Scaffolds in Critical Size Defects: In Vitro and In Vivo Study

## TL;DR

This study compares how well chitosan foam and chitosan membrane scaffolds help regenerate bone in rats with large bone defects.

## Contribution

The study introduces a direct comparison of bone regeneration between chitosan foam and membrane scaffolds using in vitro and in vivo methods.

## Key findings

- Chitosan foam had a higher pore size (65.42 µm) than chitosan membrane (6.44 µm).
- Chitosan membrane showed better cell proliferation at 72 hours, while chitosan foam showed better bone regeneration at 35 days.
- Both scaffolds outperformed the control group in bone regeneration and inflammation reduction.

## Abstract

Objectives: The objective of this study is to compare the bone-regenerating capacity between chitosan foam and chitosan membrane scaffolds. Methods: A medium-weight chitosan acidic mixture was used to prepare two scaffolds of freeze-dried chitosan foam (CF). One of the two CF scaffolds was physically crosslinked by NaHCO3 to obtain chitosan membrane (CM). A morphological assessment of the specimens’ porosity was carried out by scanning electron microscopy (SEM). An MTT assay of the CM and CF specimens using rats’ bone marrow mesenchymal stem cells (MSCs) was carried out. Then, 38 albino rats were subjected to surgical implantation in a critical-size defect of the femur bone. The rats were divided into three groups according to the type of implanted scaffold (Control (no scaffold) n = 10, CM (chitosan membrane) n = 14, CF (chitosan foam) n = 14). Each group was equally subdivided into two subgroups according to the time of euthanasia (21 d, 35 d). The femur bones were dissected for a histological analysis (hematoxylin and eosin, and Masson trichrome). The results of the histological analysis were graded according to a scoring system. A statistical analysis of the pore size and histological grading was carried out. Results: CF had a higher mean pore size (65.42 µm) compared to CM (6.44 µm); CM showed a significantly higher proliferation of MSCs at 72 h. Both the CM and CF groups showed a significantly higher bone regeneration and lower inflammation than the control group. The CF group showed a significantly higher bone regeneration score than the CM group, especially at 35 d with more dense compact lamellar bone structure. Conclusions: The higher mean pore size of CF allowed for a higher bone regenerating capacity than the crosslinked CM.