In Vitro and In Ovo CAM Model Evaluation of Periosteum-Derived Micrografts

Despite advances in periodontal regenerative therapies, consistent tissue regeneration remains challenging, with cells playing an essential role in successful repair. Therefore, this study tested different dental bone substitutes embedded in the chorioallantoic membrane (CAM) combined with periosteum-derived micrografts obtained using a chair-side device (Rigenera HBW system). Cell populations within the micrografts were identified and characterised via immunofluorescence and flow cytometry (CD31, CD105, CD34, CD90, CD73, and CD45). A CAM model was employed to examine the angiogenic potential of micrografts combined with bone substitutes, which were analysed through quantitative blood vessel/vascularisation assessments using the Ikosa software (2025), along with histological and immunohistochemical evaluations such as smooth muscle actin (SMA), H&E, and Masson's trichrome staining. Statistical analysis was performed using GraphPad Prism 10. The addition of periosteum-derived micrografts resulted in angiogenic enhancement compared to the controls. Notable enhancement of total vessel area, total length, and branching points were obtained when Fisiograft^® (p = 0.0007, p = 0.0002, and p < 0.0001, respectively), New Shore^® (p = 0.0006, p = 0.0149, and p = 0.0083, respectively), and Bio-Oss^® (p = 0.0038 and p = 0.0010, respectively) were combined with micrografts, compared to the positive controls. The histological and immunohistochemical analyses confirmed increased vascularisation (positive staining for SMA) in the micrograft groups. Periosteum-derived micrografts represent a promising adjunct to conventional bone-grafting materials, promoting vascularisation and potentially enhancing tissue regeneration and healing outcomes.