Protective Effects of the Phytocystatin CaneCPI-5 on Periodontal Ligament Cell Osteogenic Activity and Alveolar Bone Loss under Inflammatory Challenge

Abstract Periodontal disease is characterized by a chronic inflammatory environment that compromises the regenerative capacity of periodontal tissues, particularly by impairing the osteogenic potential of periodontal ligament cells. Phytocystatins have emerged as bioactive molecules with potential regenerative and immunomodulatory properties; however, their effects on periodontal tissues under inflammatory conditions remain unexplored. In this study, we investigated the effects of CaneCPI-5 on the osteogenic behavior of human periodontal ligament cells (hPDLCs) under basal and inflammatory conditions in vitro, as well as its impact on alveolar bone preservation and periodontal tissue responses in a murine model of experimental periodontitis. In vitro, CaneCPI-5 promoted the expression of osteogenic markers and supported mineralized matrix formation under basal conditions. Notably, CaneCPI-5 preserved osteogenic gene expression and mineral deposition in hPDLCs exposed to Filifactor alocis, indicating a sustained osteogenic response despite inflammatory stimulation. In vivo, treatment with CaneCPI-5 significantly attenuated alveolar bone loss, increased bone mineral density and bone volume fraction, and was associated with histological features indicative of a more preserved periodontal tissue architecture compared with untreated disease controls. Stereometric analyses further revealed reduced inflammatory cell infiltration and vascular density, alongside increased fibroblast presence and extracellular matrix components in treated animals. Collectively, these findings demonstrate that CaneCPI-5 supports osteogenic activity and contributes to periodontal tissue preservation in inflammatory settings. The convergence of in vitro and in vivo data suggests that CaneCPI-5 may be a promising bioactive molecule for the development of adjunctive regenerative strategies to mitigate inflammation-associated periodontal tissue destruction.