Human Dental Follicle Cell-Derived Conditioned Media Enhance Periodontal Regeneration

This study aimed to investigate whether conditioned media (CM) from human Dental follicle cells (hDFCs) could regenerate impaired periodontal tissue and the underlying mechanisms. hDFC-CM was obtained via ultracentrifugation. Human periodontal ligament stem cells (hPDLSCs) from 18-25 years old donors were treated with hDFC-CM under normal and inflammatory microenvironments (lipopolysaccharide-induced), with proliferation and migration assessed by CCK-8 and scratch assays, osteogenesis through ALP and Alizarin Red staining, and Wnt/β-catenin pathway analyzed via Western blot. 6-week-old male Sprague-Dawley rats (n=16) with induced periodontitis received hDFC-CM treatment for 4 weeks. Micro-CT analysis quantified bone regeneration, while H&E staining and CD68+/CD206+ immunohistochemistry evaluated tissue repair and macrophage polarization. All procedures were approved by the Institutional Ethics Committee (certificate number, 20221118-028 and 20211209004). Data were analyzed using ANOVA with p<0.05 considered significant. hDFC-CM enhanced hPDLSCs' proliferation, migration, and osteogenic differentiation (evidenced by ALP/Alizarin Red staining and upregulated osteogenic markers) by regulating the Wnt/β-catenin pathway in an inflammatory environment. It concurrently inhibited inflammatory responses in hPDLSCs and induced macrophage polarization from M1 to M2 phenotype. In vivo experiments confirmed hDFC-CM's efficacy in promoting periodontal tissue regeneration. hDFC-CM enhance periodontal regeneration by regulating the osteogenic differentiation and inflammation of periodontal ligament stem cells and macrophage polarization, which provided new biochemical cues for the treatment of periodontitis.