活性维生素D缺乏导致颞下颌关节骨关节病的作用机制研究

Temporomandibular joint osteoarthritis (TMJ-OA) is a common disease, however, its pathogenesis is still unclear. Our previous study found that 1alpha, 25dihydroxylvitaminD [1,25(OH)2D] deficiency can lead to TMJ-OA, and the expressions of DNA damage and aging-related indicators were higher. To clarify the possible mechanism, we plan to use gene silencing method to harvest 1,25(OH)2D deficiency condylar chondrocytes in vitro, give exogenous oxidative stimuli, and then the 1,25(OH)2D and p53 inhibitor, detecting the level of oxidative stress, cell senescence and osteoarthritis-related indicators. To investigate the further antioxidate effect of 1,25(OH)2D on TMJ, 1α(OH)ase-/- mice will be maintained on a diet of rescue diet with N-acetyl-L-cysteine (NAC) solution, or subcutaneous injection of 1,25(OH)2D, and their wild-type littermates will be fed on a diet of regular mouse chow. The mice will be sacrificed at 6-month-old, and the results will be analyzed by imageology, histomorphology immunohistochemistry and biochemical detection. At last, to investigate the role of p53 in the TMJ-OA induced by 1,25(OH)2D deficiency, we will delete p53 gene in 1α(OH)ase-/- mice, their phenotypes will be compared in TMJ with those of p53 deficient, 1α(OH)ase-/- and wild-type mice. Results from this study will be helpful to elucidate the mechanism of 1,25(OH)2D deficiency-induced TMJ-OA, and will provide theoretical and experimental basis for the applications of 1,25(OH)2D in clinical study on treatment of TMJ-OA.

颞下颌关节骨关节病（TMJ-OA）为临床常见疾病，但其发病机制尚不明确。我们的前期研究发现活性维生素D缺乏可导致TMJ-OA，且髁状突软骨细胞内DNA损伤及衰老相关指标p53表达水平明显增高。为了进一步明确其发病机制，本研究拟采用基因沉默的方法于体外构建维生素D受体缺乏的TMJ-OA软骨细胞模型，予以活性维生素D、p53抑制剂进行干预，检测氧化应激及衰老相关指标。同时，给予活性维生素D缺乏的1α(OH)ase-/-小鼠抗氧化剂喂养，或构建1α(OH)ase-/-p53-/-双基因突变小鼠，将其与同窝野生型小鼠比较，观察颞下颌关节的表型差异。本研究结果将从细胞及动物水平明确活性维生素D缺乏导致氧化应激水平增高，激活p53-p21信号通路，加速细胞衰老，进而导致TMJ-OA的作用机制，并为其临床应用于治疗TMJ-OA提供理论和实验依据。

颞下颌关节骨关节病（TMJ-OA）为临床常见疾病，但其发病机制尚不明确。我们的前期研究发现活性维生素D缺乏可导致TMJ-OA，且髁状突软骨细胞内DNA损伤及衰老相关指标p53表达水平明显增高。为了进一步明确其发病机制，本研究拟使用慢病毒转染的方法在体外构建VDR缺乏的软骨细胞，再通过给予H2O2刺激的方法构建颞下颌关节骨关节病的软骨细胞模型，予以活性维生素D、p53抑制剂进行干预，各组细胞通过Western blot方法检测DNA损伤及相关炎症因子表达的变化情况，发现p53作为1,25(OH)2D的重要下游靶标，在1,25(OH)2D缺乏导致TMJ-OA中促进衰老，进而导致软骨细胞产生SASP。进一步运用1,25(OH)2D 缺乏的1α(OH)ase-/-小鼠模型，通过分笼后给予正常饮食，或高钙高磷饮食，或高钙高磷饮食并于饮水中补充抗氧化剂NAC（1g/L）至6月龄，将其与同窝WT小鼠的颞下颌关节表型进行比较，发现1,25(OH)2D 缺乏导致的氧化应激水平增高，进而导致DNA损伤增加，使得髁状突软骨细胞产生SASP。此外，我们构建了1α(OH)ase-/-p53+/-双基因突变小鼠，将其与同窝野生型小鼠比较，观察颞下颌关节的表型差异，发现活性维生素D缺乏导致氧化应激水平增高，激活p53-p21信号通路，加速细胞衰老，进而导致TMJ-OA。