摘要
目的 探索铁死亡诱导剂咪唑酮埃拉斯汀(IKE)对胶原蛋白诱发关节炎(CIA)小鼠肺纤维化的作用及潜在机制.方法 选取8周龄～10周龄DBA/1小鼠,按照完全Freund佐剂(CFA)与鸡Ⅱ型胶原蛋白混合乳化方法建立CIA小鼠模型,设置对照组、CIA组、CIA联合IKE组.每2 d进行关节评分与爪垫厚度测量,39 d后收集小鼠各脏器组织.HE染色、番红-固绿染色、甲苯胺蓝染色评价关节处组织病理改变;Masson染色评价肺部组织病理改变.免疫组织化学染色法检测肺组织α平滑肌肌动蛋白(α-SMA)、成纤维细胞活化蛋白α(FAPα)、转化生长因子β(TGF-β)及I型胶原蛋白(Col1)、白细胞介素1(IL-1)、IL-6、IL-17及肿瘤坏死因子α(TNF-α)的表达水平;Olink mouse exploratory panel检测小鼠血清细胞因子IL-17α、IL-17F、TGF-β1、整合素亚基 β6(ITG-β6)、TNF 受体超家族成员 11B(TNFRSF11b)、TNF 受体超家族成员 12A(TNFRSF12a)、IL-6、IL-1α、IL-1β、IL-10、TNF-α、CC趋化因子配体5(CCL5)、CCL2、CXC趋化因子配体9(CXCL9)、CXCL1、烟酰胺腺嘌呤二核苷酸激酶(NADK)、促红细胞生成素(EPO)、集落刺激因子2(CSF2)、TGF-α、CCL20、CCL3水平.结果 与CIA组相比,CIA联合IKE组治疗后关节炎症及关节损伤明显缓解;肺组织α-SMA、FAPα、TGF-β和Col1表达水平均呈下降趋势,表明CIA小鼠肺部胶原蛋白聚集减少,组织病变明显缓解;IL-6、CCL5、CXCL9、IL-6水平显著下降表明CIA小鼠肺部炎症显著缓解.结论 IKE除缓解CIA小鼠关节炎症及关节损伤外,还可通过抑制IL-6、CCL5及CXCL9表达缓解CIA伴发的肺纤维化.
Abstract
Objective To investigate the impact of imidazole ketone erastin(IKE),a ferroptosis inducer,on pulmonary fibrosis progression in mice with collagen-induced arthritis(CIA),and to understand its potential mechanism.Methods Chick type Ⅱ collagen emulsified in complete Freunds adjuvant(CFA)was injected into DBA/1 mice,aged 8 to 10 weeks,to induce CIA.Fourteen days later,type Ⅱ collagen emulsified in incomplete Freund's adjuvant(IFA)was administered to the mice.The mice were randomly divided into a control group,a CIA group and a CIA combined IKE group.The development of arthritis was monitored by evaluating the arthritis scores every two days until day 39 and then the mice were sacrificed for organ collection.The histopathological changes of joints were evaluated by HE staining,Safranin O-fast green staining and toluidine blue staining.The histopathological changes of organs including heart,liver,spleen,lung,and kidney were evaluated by HE staining,and Masson's trichrome staining was used to assess pulmonary fibrosis.The expression levels of smooth muscle actin α(α-SMA),fibroblast activating protein α(FAPα),transforming growth factor β(TGF-β),type I collagen(Col1),interleukin 1(IL-1),IL-6,IL-17 and tumor necrosis factor α(TNF-α)were detected by immunohistochemical staining.The expression levels of serum cytokines including IL-17α,IL-17F,TGF-β1,ITG-β6,TNF receptor superfamily menber 11B(TNFRSF11B),TNFRSF12A,IL-6,IL-1α,IL-1β,IL-10,TNF-α,CCL5,CCL2,CXCL9,CXCL1,NADK,EPO,CSF2,TGF-α,CCL20 and CCL3 in serum were detected by Olink mouse exploratory panel.Results Histological staining in the CIA mice administered with IKE model demonstrated that IKE treatment reduced bone absorption and the degree of synovial inflammation when active inflammation was present.CIA mice administered with IKE showed lower expression levels of α-SMA,FAPα,TGF-β,Col1,IL-1,IL-6,IL-17 and TNF-α,according to the immunohistochemical staining of the lung.In addition,the expression levels of CCL5,CXCL9 and IL-6 were also decreased in serum of CIA mice treated with IKE.Conclusion IKE not only ameliorates joint inflammation and bone damage,but also alleviates the inflammation and the progression of pulmonary fibrosis in CIA mice.
基金项目
陕西省创新能力支撑计划(2023-CX-PT-18)
NETL
NSTL
万方数据