RESUMO
The junctional epithelium (JE) serves a crucial protective role in the periodontium. High glucose-related aging results in accelerated barrier dysfunction of the gingival epithelium, which may be associated with diabetic periodontitis. Metformin, an oral hypoglycemic therapeutic, has been proposed as a anti-aging agent. This study aimed to clarify the effect of metformin on diabetic periodontitis and explore its mechanism in ameliorating senescence of JE during hyperglycemia. The db/db mice was used as a diabetic model mice and alterations in the periodontium were observed by hematoxylin-eosin staining and immunohistochemistry. An ameloblast-like cell line (ALC) was cultured with high glucose to induce senescence. Cellular senescence and oxidative stress were evaluated by SA-ß-gal staining and Intracellular reactive oxygen species (ROS) levels. Senescence biomarkers, P21 and P53, and autophagy markers, LC3-II/LC3-I, were measured by western blotting and quantitative real-time PCR. To construct a stable SIRT1 (Sirtuin 1) overexpression cell line, we transfected ALCs with lentiviral vectors overexpressing the mouse SIRT1 gene. Cellular senescence was increased in the JE of db/db mice and the periodontium was destroyed, which could be alleviated by metformin. Moreover, oxidative stress and cellular senescence in a high glucose environment were reduced by metformin in in-vitro assays. The autophagy inhibitor 3-MA and SIRT1 inhibitor EX-527 could dampen the effects of metformin. Overexpression of SIRT1 resulted in increased autophagy and decreased oxidative stress and cellular senescence. Meanwhile, AMPK (AMP-activated protein kinase) inhibition reversed the anti-senescence effects of metformin. Overall, these results suggest that metformin alleviates periodontal damage in db/db mice and cellular senescence in ALCs under high glucose conditions via the AMPK/SIRT1/autophagy pathway.
RESUMO
Excessive fluoride intake during enamel development can affect enamel mineralization, leading to dental fluorosis. However, its potential mechanisms remain largely unexplored. In the present study, we aimed to investigate the impact of fluoride on the expressions of RUNX2 and ALPL during mineralization and the effect of TGF-ß1 administration on fluoride treatment. A dental fluorosis model of newborn mice and an ameloblast cell line ALC were both used in the present study. The mice of the NaF group, including the mothers and newborns, were fed with water containing 150 ppm NaF after delivery to induce dental fluorosis. The mandibular incisors and molars showed significant abrasion in the NaF group. Immunostaining, qRT-PCR, and Western blotting analysis indicated that exposure to fluoride markedly down-regulated RUNX2 and ALPL in mouse ameloblasts and ALCs. Besides, fluoride treatment significantly decreased the mineralization level detected by ALP staining. Furthermore, exogenous TGF-ß1 up-regulated RUNX2 and ALPL and promoted mineralization, while the addition of SIS3 could block such TGF-ß1-induced up-regulation. In TGF-ß1 conditional knockout mice, the immunostaining of RUNX2 and ALPL was weaker compared with wild-type mice. Exposure to fluoride inhibited the expressions of TGF-ß1 and Smad3. Co-treatment of TGF-ß1 and fluoride up-regulated RUNX2 and ALPL compared with the fluoride alone treatment, promoting mineralization. Collectively, our data indicated that TGF-ß1/Smad3 signaling pathway was necessary for the regulatory effects of fluoride on RUNX2 and ALPL, and the fluoride-induced suppression of ameloblast mineralization was mitigated by activating TGF-ß1/Smad3 signaling pathway.
