Effects of Low‐Dose Doxycycline and Bisphosphonate Clodronate on Alveolar Bone Loss and Gingival Levels of Matrix Metalloproteinase‐9 and Interleukin‐1β in Rats With Diabetes: A Histomorphometric and Immunohistochemical Study

Abstract

Bisphosphonates (BPs) and low-dose doxycycline (LDD) have been shown to inhibit bone resorption and to improve the levels of proinflammatory mediators and destructive enzymes in gingival tissues, respectively. The purpose of this study is to evaluate the effect of mono and combined BP clodronate and LDD therapies in reducing gingival levels of matrix metalloproteinase-9 (MMP-9), interleukin-1β (IL-1β), and alveolar bone loss in rats with diabetes. Fifty adult Wistar rats were divided into five study groups as follows: 1) group 1 = diabetes control; 2) group 2 = diabetes + periodontitis; 3) group 3 = diabetes + periodontitis + LDD; 4) group 4 = diabetes + periodontitis + clodronate; and 5) group 5 = diabetes + periodontitis + LDD + clodronate. LDD and clodronate were given as a single agent or as combination therapy during the 7 days of the post-experimental periodontitis period. On day 7, the rats were sacrificed, the mobility of the tooth was recorded, and block biopsies were removed. The gingival tissues were analyzed histologically and immunohistochemically for expression of MMP-9 and IL-1β. Alveolar bone loss was evaluated morphometrically under a light microscope. Data analysis was performed statistically by Kruskal-Wallis and post hoc Tukey and Spearman correlation tests. Alveolar bone loss was significantly greater in groups 2 through 5 than group 1 (P <0.05) but was not significantly different among groups 2 through 5 (P >0.05). Animals with periodontitis (group 2) expressed significantly higher levels of MMP-9 and IL-1β compared with those without periodontitis (group 1) (P <0.05). MMP-9 expression was significantly lower in group 3 than groups 1, 2, and 5 (P <0.05). IL-1β expression was significantly lower in the groups 1, 3, 4, and 5 than 2 (P <0.01) but was not significantly different among groups 1, 3, 4, and 5. Positive correlations were found between alveolar bone loss and density of inflammation (ρ = 0.319, P = 0.021) and between MMP-9 and IL-1β (ρ = 0.418, P = 0.002), respectively. Our findings suggest that ligature-induced periodontitis in animals with diabetes results in significantly higher levels of MMP-9 and IL-1β expression in gingiva. The use of mono and combined clodronate and LDD administrations may significantly reduce levels of MMP-9 and IL-1β expression. However, drug administration did not affect alveolar bone levels during the study period.