Abstract
External root resorption following avulsion injury is a complex process wherein differentiation of Macrophages (Mφ) to multinucleated osteoclasts is temporally regulated by resident periodontal fibroblasts (PDLF). The current study aims to assess the effect of engineered bioactive chitosan nanoparticles (CSNP), sustained released Dexamethasone conjugated CSNP (CS-DEX) and CSNP functionalised with photosensitizer Rose Bengal (CSRB) for application in root resorption using an in-vitro PDLF-Mφ direct coculture model, and in-vivo delayed reimplantation model. PDLF-Mφ direct coculture system was exposed to LPS, MCSF, RANKL with or without CSNP/CS-DEX for 7 days. Clastic differentiation was assessed by tartrate resistant acid phosphatase (TRAP) staining on day 7. On day 2 and 7, immunofluorescence analysis was done to assess the expression of Mφ polarization markers (CD80, CD206), multinucleation markers (NFATc1, STAT6) in Mφ and matricellular protein periostin in PDLF and cytokine profiling in cell culture supernatants. Delayed replantation model with extraoral air dry / LPS exposure for 1h followed by root surface treatment with CS-DEX/CSRB was used in Wistar rats. After 21 days, rats were euthanized for histological and immunofluorescence analysis. Statistical analysis One-way ANOVA with Tukey's multiple comparisons was used to analyze the data (p<0.05). CS-DEX significantly reduced TRAP+ multinucleated cells and CSNP treatment showed no TRAP+ cells. Immunofluorescence analysis showed that CSNP/CS-DEX reduced CD80, NFATc1 and STAT6 expression and increased periostin as expressed by fluorescence intensity. CSNP/CS-DEX significantly reduced TNFα, MMP9 and increased IL10, TGFβ1. OPG was upregulated only by CSNP. Root surface treatment in delayed replantation model showed that CS-DEX and CSRB substantially reduced the degree of resorption and ankylosis. Further, CD80, CD206, MMP2 expression in groups with root surface treatment with CS-DEX and CSRB was lower than airdry/ LPS group and similar to healthy control and NFATc1, STAT6, MMP9 expression were lower than healthy control. The engineered nanosized immunomodulatory bioactive materials chitosan nanoparticles functionalised with photosensitizer and dexamethasone effectively reduced the clastic differentiation of Mφ in in-vitro coculture and minimized the resorption and ankylosis in a delayed reimplantation model. These biomaterials have the potential to serve as root modification agents, promoting favourable healing outcomes in cases of dental avulsion.
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