Innovative In Vivo Approaches to Bone Regeneration: Combining Chitosan-Xanthan Gum Membranes, Electrical Stimulation, and Laser Therapy.

Critical bone defects present significant clinical challenges due to their debilitating nature and associated costs. Tissue engineering offers a promising solution, employing biomaterials like porous chitosan (CH) and xanthan gum (XG) membranes as scaffolds for tissue regeneration. Electrical stimulation (ES) and laser (L) therapy, known for osteogenic and anti-inflammatory effects, respectively, may synergistically enhance bone healing when combined with these membranes. This study evaluated CH/XG membranes and their association with ES and L for the treatment of critical bone defects (25 mm2) created in 80 male Wistar rats. The animals were divided into four groups: untreated, CH/XG, CH/XG + ES, and CH/XG + L, assessed at days 30 and 90. Histomorphometric and real-time PCR analyses revealed greater tissue and blood vessel formation in the CH/XG, CH/XG + ES, and CH/XG + L groups compared to the untreated group. At day 30, reduced Runx-2 and Osx expression in the treated groups suggested an initial high inflammatory response. The CH/XG group showed pronounced inflammation (elevated Il-1β and pro-inflammatory markers) alongside an anti-inflammatory response. Conversely, compared with the CH/XG-only group, the CH/XG + ES and CH/XG + L groups generally showed lower expression of the evaluated pro-inflammatory markers after 30 days, together with treatment- and time-dependent changes in anti-inflammatory markers. Although inflammation limited the initial effectiveness of CH/XG membranes, significant positive impacts were observed after 90 days. Combining membranes to cover bone defects and adjunctive therapies, as demonstrated in this study, emerges as a beneficial approach for enhancing bone repair outcomes. Adding osteoconductive or osteoinductive fillers may bring further healing improvements.