| BackgroundPeriodontitis is one of the chronic infectious diseases with the highest incidence rate in human beings.Periodontitis induces the recruitment of a variety of immune cells by activating the host’s internal immunity,resulting in the loss of periodontal tissues such as periodontal ligament,cementum,and alveolar bone.Bringing great inconvenience to patients’ lives and seriously affecting oral health.There are many triggering factors for periodontitis,such as poor lifestyle habits such as smoking,the impact of systemic diseases,and genetic inheritance.At present,the impact of systemic diseases on periodontitis is the focus of research in recent years.The complications of systemic diseases such as diabetes,liver disease,osteoporosis,and neurological diseases will reduce the patient’s immunity and increase the risk of infection by pathogenic microorganisms such as bacteria and fungi,activate innate immunity,inhibit bone formation,and increase bone resorption.Periodontitis also increases the risk of systemic disease development,for example,periodontitis promotes the secretion of insulin,affects downstream pancreatic islet organs,reduces insulin secretion,increases the risk of hyperglycemia,and further damages the immune system.Periodontitis is the sixth complication of diabetes.Successful periodontal treatment has a significant effect on blood glucose control in diabetes patients,which suggests a close relationship between the two diseases.Diabetes-related serum metabolites,hyperglycemia levels,and advanced glycation end products(AGEs)are the main factors that aggravate periodontitis and oral microbiome disorders.There are many patients with diabetes periodontitis,and more than 80% of type 2 diabetes patients have periodontitis,which leads to alveolar bone absorption and tooth loss,which is highly harmful.Therefore,the treatment of diabetes periodontitis has become an important issue.The main solution to prevent tooth loss is to enhance bone regeneration ability and reduce alveolar bone absorption.There are three kinds of cells in bone tissue: osteoblasts,osteoclasts,and osteocytes.Osteoblasts have the ability of bone regeneration to promote bone tissue differentiation,osteoclasts have the ability to bone absorption,and osteocytes regulate osteoblast and osteoclast functions.Osteocytes secrete cytokines to regulate the differentiation of osteoblasts and osteoclast,regulate bone regeneration and bone absorption,and maintain bone homeostasis.Ferroptosis is a kind of programmed cell death related to lipid peroxide metabolism,which is involved in the process of diabetes,inflammation,and other diseases.The cysteine/glutamate transporter system(Xc-)containing glutathione peroxidase 4(GPX4)and member 11 of the solute carrier family 7(SLC7A11)is a classic pathway of ferroptosis,and downregulating the expression of GPX4 and SLC7A11 triggers ferroptosis.Cells that undergo ferroptosis may experience pathological phenomena such as mitochondrial damage,oxidative stress,and lipid peroxidation accumulation.According to literature reports,osteoblasts in osteoarthritis and diabetes undergo ferroptosis,but there is no report on osteocyte ferroptosis during diabetes periodontitis.Therefore,this study aimed to explore whether diabetic periodontitis induces osteocyte ferroptosis.Resveratrol(RSV)is a natural compound molecule with strong anti-inflammatory effects,which has been proven to treat various diseases.RSV downregulates interleukin-6(IL-6)and tumor necrosis factor-ɑ(Tumor necrosis factor-α,TNF-ɑ)inhibiting the occurrence and development of arthritis.Moreover,RSV inhibits ferroptosis in atrial fibrillation induced by excessive ethanol.Similarly,RSV reduces the damage of autoimmunity to islet B cells and effectively alleviates diabetes by increasing the transfer of glucose transporter 4 to the cell membrane.However,there is no report on the effect of RSV treatment on alveolar osteocyte functions during diabetes or periodontitis.ObjectiveThis topic will clarify the occurrence of osteocyte ferroptosis during diabetic periodontitis using animal models and in vitro studies.We will further evaluate the effect of local RSV treatment on diabetic periodontitis-induced osteocyte ferroptosis.Material and methodsThe first part: The effect of diabetic periodontitis on osteocyte ferroptosis(1)Establishment of the diabetic periodontitis models in vivo and Evaluate osteocyte ferroptosis.The diabetes was induced in mice by a high sugar/fat diet and the second molar ligation was adopted to induce periodontitis in diabetic mice.Micro-CT,IHC,and HE staining were performed to evaluate the periodontal status and osteocyte ferroptosis.(2)Establishment of the diabetic periodontitis models in vitro and Evaluate osteocyte ferroptosis.MLOY4(mice osteocytes)were cultured with different concentrations of AGEs and LPS for 24 h.The expressions of inflammatory factors and ferroptosis markers were evaluated by RT-q PCR,ELISA,and capillary-based immunoassay.The second part: The effect of RSV treatment on diabetic periodontitis-induced alveolar osteocyte ferroptosisLPS and AGEs-treated MLOY4 cells were treated with RSV and the cell viability,ferroptosis markers’ expression,and inflammatory factors’ expression were analyzed.Similarly,RSV was injected in the periodontal tissue of diabetic periodontitis mice,and periodontal status and osteocyte ferroptosis were evaluated as mentioned earlier.The third part: Molecular mechanism of the resveratrol-mediated rescue of diabetic periodontitis-induced osteocyte ferroptosismRNA seq,bioinformatic analysis,and further verification of TNF-α related NF-κB signaling were performed in osteocytes treated with diabetic periodontitis condition and/or RSV.ResultsThe first part: Diabetic periodontitis promoted alveolar osteocytes ferroptosis(1)Diabetic periodontitis mice promoted ferroptosis in alveolar osteocytes.The expression of GPX4 and SLC7A11 in the alveolar osteocytes of diabetes periodontitis mice was significantly downregulated compared to control mice,indicating that diabetic periodontitis activates ferroptosis in alveolar osteocytes in vivo.(2)Diabetic periodontitis osteocytes promoted ferroptosis.The expression of GPX4 and SLC7A11 in the diabetes periodontitis osteocytes was significantly downregulated compared to control osteocytes,indicating that diabetic periodontitis activates ferroptosis in alveolar osteocytes in vitro.The addition of Fer-1reversed this phenomenon,indicating that diabetic periodontitis activates ferroptosis in osteocytes in vitro.The second part: RSV alleviated diabetic periodontitis-induced alveolar osteocyte ferroptosisRSV inhibited diabetic periodontitis-induced alveolar osteocyte inflammation and ferroptosis in vivo.The distance between enamel cementum junction and alveolar bone crest CEJ-ABC in the DP+RSV group was significantly lower than that in the diabetic periodontitis group,and the expression of GPX4 and SLC7A11 was significantly increased,suggesting that RSV inhibits diabetic periodontitis induced inflammation and ferroptosis in vivo.The number of osteocytes death in the AGEs+LPS+RSV group decreased compared to the disease group.The expression of pro-inflammatory factors significantly decreased.and anti-inflammatory factors significantly increased.The expression of GPX4 and SLC7A11 significantly increased.It suggests that RSV inhibits diabetic periodontitisinduced inflammation and ferroptosis in vitro.The third part: RSV alleviates the ferroptosis of alveolar osteocytes induced by diabetes periodontitis possibly via downregulated NF-κBRSV inhibits lipid peroxidation accumulation via downregulated NF-κB,thereby inhibiting diabetic periodontitis-induced ferroptosis in alveolar osteocytes and alleviating diabetic periodontitis-induced osteocyte inflammation.The accumulation of lipid peroxidation in the AGEs+LPS+RSV and NF-κB expression group were significantly reduced compared with the AGEs+LPS group.Conclusion1.Diabetic periodontitis triggered alveolar osteocyte ferroptosis.2.RSV alleviated the osteocyte inflammatory pathogenicity induced by diabetes periodontitis.3.RSV inhibited diabetic periodontitis-induced osteocyte ferroptosis via activating SLC7A11/GPX4,and alleviated periodontitis pathogenicity.4.RSV downregulated NF-κB signaling in osteocytes cultured in diabetic periodontitis conditions. |
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