PTHrP Overexpression in Periosteum Promotes Alveolar Bone Formation

Alveolar bone regions with low innate thickness are prone to dehiscence, potentially causing harmful gingival recession. In situ alveolar bone thickening via local bioagent administration has recently emerged as a promising research area. Parathyroid hormone-related protein (PTHrP), encoded by the Pthlh gene, plays a pivotal role in skeletal development and bone remodeling. In this study, we developed a recombinant adeno-associated virus (rAAV) vector overexpressing Pthlh (rAAV9-Pthlh). Using a split-mouth design, we administered rAAV9-Pthlh via a single submucosal injection into the right maxillary buccal gingiva of rats. Micro-computed tomography scans, both in vivo and ex vivo, revealed a significant increase in alveolar bone thickness on the PTHLH sides 6 wk post-transfection. This enhanced alveolar bone formation was accompanied by marked periosteal thickening, with elevated expression of PTHrP, parathyroid hormone 1 receptor (PTH1R), OCN, and COL1A1. RNA sequencing of the buccal bone tissue showed upregulation of osteogenesis-related pathways along with increased mitophagy activity. Immunofluorescence confirmed enhanced colocalization of mitochondrial and lysosomal markers in the PTHLH group, primarily in the periosteum. Overexpressing Pthlh in primary rat periosteal stem cells via lentiviral transduction promoted cell proliferation and migration and elevated expression of COL1A1, as well as mitophagy markers BNIP3 and LC3-II. These effects were mitigated by Mdivi-1, an inhibitor of mitochondrial fission, suggesting potential involvement of mitophagy-dependent mitochondrial homeostasis maintenance. Finally, we established a rat model of repeated maxillary expansion and validated the effects of prophylactic rAAV9-Pthlh injection to protect against alveolar crest loss in this scenario. In conclusion, local rAAV9-Pthlh injection enhances in situ alveolar bone formation, holding potential for preventing dehiscence in aging or following orthodontic interventions. Additionally, overexpression of PTHrP in the alveolar periosteum has reiterated its well-documented role in expanding the pool of chondroskeletal progenitors, a process vital for the development of long bones and the mandibular condyle.