Figure 5.

AAV-VEGFC increases local lymphatic drainage and reduces alveolar bone loss in ligature-induced periodontitis. Two-month-old WT mice were randomly divided into two groups (six mice per group). One group received local injection of AAV-GFP control virus in periodontal tissues of both the left and right maxillary first molar. Another group received AAV-VEGFC virus injection. Two weeks later, a 5–0 silk ligature was tied around the right maxillary first molar and the contralateral tooth was left unligated to serve as the baseline control. All the mice were sacrificed and analyzed 2 weeks after placement of the ligature. (A) Paraffin sections of maxillae were subjected to IF for PDPN (red) and αSMA (green). Representative images of periodontal tissue showing the distribution of PDPN⁺/αSMA⁻ lymphatic capillaries (red), PDPN⁺/αSMA⁺ collecting LVs (yellow), and PDPN⁻/αSMA⁺ blood vessels (green). Quantification of the PDPN⁺αSMA⁻ area or PDPN⁺αSMA⁺ area to tissue area (%) was determined. The number of PDPN⁺αSMA⁺ collecting LVs per mm² tissue areas (#/mm²) was calculated. (B) An example of in vivo washout of ICG in periodontal tissues reflecting lymph flow through all time points. ICG clearance (%) was quantified. Values shown are mean ± SEM. *p < 0.05 versus control + AAV-GFP, ^#p < 0.05 versus periodontitis + AAV-VEGFC, at the same time point. (C) Representative 3D scanned sections (upper panels) and reconstructed sections (lower panels) along the longitudinal direction of the maxillae. Bone mineral density (BMD, g/cm²) was analyzed. (D) Representative images of H&E-stained paraffin sections. Measurement of the bone levels by comparing the distance from the cemento-enamel junction (CEJ) to the alveolar bone crest (ABC) in μm was determined. (E) Representative images of TRAP-stained paraffin sections. The percentage of alveolar bone surface covered by osteoclasts (Oc.S/B.S) was determined. (F) Representative images of ALP-stained paraffin sections. The percentage of alveolar bone surface covered by osteoblasts (Ob.S/B.S) was determined. *p < 0.05 in the indicated groups. N = 6. One-way ANOVA followed by Dunnett’s post hoc multiple comparisons test was performed.