E3泛素连接酶TRIM65通过调控NLRP3炎症小体的活化改善DSS诱导的结肠炎

E3 ubiquitin ligase alleviates DSS-induced colitis via regulating NLRP3 inflammasome activation

目的 TRIM65对葡聚糖硫酸钠(DSS)诱导的结肠炎的作用及机制探讨。方法 Trim65^(-/-)和Trim65-/-小鼠连续5d给予含3%(w/v)DSS的饮用水,然后给予无菌水2d。每天监测小鼠结肠炎的临床症状(体重、粪便稠度和直肠出血评分),第7d处死小鼠。测量结肠长度,收集结肠远端组织,HE染色和免疫组化判定组织损伤和炎症程度;匀浆结肠组织,评估MPO活性,免疫印迹法(western blotting)和酶联免疫吸附试验(ELISA)检测成熟caspase-1和IL-1β的变化。Trim65^(-/-)小鼠腹腔注射NLRP3炎症小体抑制剂MCC950,同时给予上述处理,以判断TRIM65对小鼠结肠炎的影响是否通过NLRP3炎症小体来实现。结果 在葡聚糖硫酸钠(DSS)诱导的小鼠的结肠炎模型中,Trim65基因的缺失可显著增强DSS诱导的小鼠的体重减轻和结肠缩短,促进疾病活动指数和组织病理学评分增加,促进结肠MPO活性和F4/80+免疫细胞浸润增加、caspase-1激活和成熟IL-1β分泌增加。NLRP3炎症小体抑制剂MCC950在Trim65基因缺陷小鼠中缓解DSS诱导小鼠的结肠炎症状和炎症水平。综上,TRIM65可通过抑制NLRP3炎症小体激活减轻DSS诱导的结肠炎。结论TRIM65通过抑制NLRP3炎症小体的激活,在DSS诱导的结肠炎小鼠中发挥抗炎作用。

Objective To investigate the role of TRIM65 on DSS induced colitis and the underlying molecular mechanisms. Methods Trim65^(-/-)and Trim65^(-/-)mice were administered with 3%(w/v) DSS in their drinking water for 5 consecutive days and thenwere switched to sterile water for 2 days.DSS treatedmice were monitored daily for the clinical symptoms(bodyweight, stool consistency and rectal bleeding score).Micewere sacrificed on day 7 to measure colon length.Colon homogenates were collected to measure MPO activity and detect cleaved caspase-1 and mature IL-1β by Enzyme linked immunosorbent assay(ELISA) and Western blot.Trim65^(-/-)mice were intraperitoneally injected with NLRP3 inflammasome inhibitor MCC950,and were given the above treatment to determine the effect of MCC950 on colitis in Trim65^(-/-)mice. Results The results showed that deletion of Trim65 significantly enhanced weight loss and colon shortening in DSS mice, increased disease activity index and histopathological score, induced the activity of MPO,and promoted the F4/80+ immune cell infiltration, the activation of caspase-1 and the secretion of mature IL-1(in the colon of DSS mice.The NLRP3 inflammasome inhibitor MCC950 alleviated DSS induced colitis symptoms and inflammation levels in trim65 deficient mice. Conclusion TRIM65 plays an anti-inflammatory role in DSS induced colitis mice by inhibiting the activation of NLRP3 inflammasome.