NLR家族Pyrin域蛋白3炎症小体对糖尿病大鼠牙槽骨缺损愈合的作用及其机制研究

The role of NLRP3 inflammasome in the healing of alveolar bone defects in diabetic rats and its underlying mechanisms

摘要:

目的 探究含NLR家族Pyrin域蛋白3（NLRP3）炎症小体对糖尿病大鼠牙槽骨缺损愈合的作用及其机制。方法 将30只大鼠随机分为对照组、模型组、炎症小体抑制组，每组10只。除对照组外，采用高糖饲养+腹腔注射链脲佐菌素复制糖尿病模型，模型复制成功后用球钻毁损大鼠部分牙槽骨骨质，复制牙槽骨缺损模型。模型复制成功后，炎症小体抑制组大鼠立即尾静脉注射20 mg/kg NLRP3抑制剂，1次/d，连续给药28 d。给药结束后，检测各组大鼠空腹血糖。采用酶联免疫吸附试验测定各组大鼠血清护骨因子（OPG）和碱性磷酸酶（ALP）含量。分别采用苏木精-伊红（HE）和抗酒石酸酸性磷酸酶（TARP）染色观察牙槽骨缺损愈合情况及破骨细胞数，进行骨再生Lane-Sandhu评分。采用实时荧光定量聚合酶链反应（qRT-PCR）和Western blotting检测NLRP3、Caspase-1和白细胞介素1β（IL-1β）mRNA和蛋白的表达。结果 模型组OPG、ALP低于对照组和炎症小体抑制组（P <0.05）。HE染色结果显示，模型组大鼠牙槽骨缺损严重，可见大量炎症浸润细胞，炎症小体抑制组牙槽骨缺损程度低，炎症浸润细胞较少。模型组Lane-Sandhu评分低于对照组和炎症小体抑制组（P <0.05）。TARP染色结果显示，模型组破骨细胞数高于对照组和炎症小体抑制组（P <0.05）。模型组大鼠上颌骨组织NLRP3、Caspase-1、IL-1β mRNA和蛋白相对表达量高于对照组和炎症小体抑制组（P <0.05）。结论 抑制NLRP3炎症小体可改善骨代谢，促进糖尿病大鼠牙槽骨缺损愈合，其作用可能与抑制NLRP3/IL-1β通路有关。

Abstract:

Objective To explore the role of NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome in the healing of alveolar bone defects in diabetic rats and its underlying mechanisms.Methods Thirty rats were randomly divided into the control group, the model group and the inflammasome inhibition group, with 10 rats in each group. All rats except those in the control group were fed with high-glucose diets and intraperitoneally injected with streptozotocin to establish the diabetic models, and the alveolar bones of rats were destructed to establish the alveolar bone defects models. After the model establishment, the rats in the inflammasome inhibition group were administrated 20 mg/kg of NLRP3 inhibitor via tail vein injection once a day for twenty-eight days, followed by the measurement of the fasting blood glucose of rats in each group. The contents of serum osteoprotegerin (OPG) and alkaline phosphatase (ALP) were measured by enzyme-linked immunosorbent assay. The hematoxylin and eosin (HE) staining and tartrate-resistant acid phosphatase staining (TARP) were applied to observe the healing of alveolar bone defects and the number of osteoclasts, as well as grading the bone formation according to the Lane and Sandhu histopathological scoring system. The mRNA and protein expressions of NLRP3, caspase-1 and interleukin (IL)-1β were detected by quantitative real-time polymerase chain reaction and Western blotting.Results Compared with the control group and the inflammasome inhibition group, the contents of OPG and ALP were lower in the model group (P < 0.05). The HE staining showed that the alveolar bone defects in the model group were severe, with the infiltration of a large number of inflammatory cells. In contrast, the degree of alveolar bone defects in the inflammasome inhibition group was low, with the infiltration of only a few of inflammatory cells. Compared with the control group and the inflammasome group, the Lane and Sandhu score in the model group decreased (P < 0.05). The TARP staining exhibited that the number of osteoclasts in the model group was higher than that in the control group and the inflammasome inhibition group (P < 0.05). The mRNA and protein expressions of NLRP3, caspase-1 and IL-1β were higher in the model group relative to those in the control group and the inflammasome inhibition group (P < 0.05).Conclusions Inhibition of NLRP3 inflammasome can improve bone metabolism and promote the healing of alveolar bone defects in diabetic rats, which may be related to the suppression of NLRP3/IL-1β pathway.