中性粒细胞诱捕网在小鼠胸主动脉瘤和夹层形成中的作用及机制研究  

The role and mechanism of neutrophil extracellular traps in the formation of thoracic aortic aneurysm and dissection in mice

作　　者：张彬 朱翰朝 孙赫 江丽青 宋凡 蒋鹏程 金振晓 段维勋 俞世强 Zhang Bin;Zhu Hanzhao;Sun He;Jiang Liqing;Song Fan;Jiang Pengcheng;Jin Zhenxiao;Duan Weixun;Yu Shiqiang(Department of Cardiovascular Surgery,the First Affiliated Hospital of the Air Force Medical University,Shaanxi Xi’an 710032,China)

机构地区：[1]空军军医大学第一附属医院心血管外科,西安710032 [2]空军军医大学药学系中药与天然药物学教研室 [3]中国人民解放军95195部队,襄阳441000

出　　处：《中国体外循环杂志》2025年第2期153-161,共9页Chinese Journal of Extracorporeal Circulation

基　　金：国家自然科学基金专项项目(82241204);国家自然科学基金面上项目(82270420);国家自然科学基金青年科学基金项目(82300388);西京创新研究院-联合基金“临床医学+药学”科研项目(LHJJ2023-YX15)。

摘　　要：目的探讨中性粒细胞诱捕网(NETs)在小鼠胸主动脉瘤和夹层(TAAD)形成中的作用及机制。方法饮水加入β-氨基丙腈(BAPN)[1g/(kg·day)]构建小鼠TAAD模型,采用腹腔注射脱氧核糖核酸酶I(DNaseⅠ)[20mg/(kg·day)]清除NETs。80只3周龄雄性C57BL/6小鼠随机分为对照组(Con+V)、对照+DNaseⅠ组(Con+DNaseⅠ)、BAPN+V组和BAPN+DNaseⅠ组。4周后记录各组小鼠TAAD发生率和破裂率,主动脉弓超声测量胸主动脉最大内径,苏木精-伊红染色观察主动脉壁结构改变,纤维组织染色观察主动脉壁弹力纤维断裂,试剂盒检测血清瓜氨酸组蛋白H3(citH3)、循环核酸(cfDNA)和核小体水平,免疫荧光染色检测citH3、髓过氧化物酶和α-平滑肌肌动蛋白荧光表达,末端标记法观察细胞凋亡,实时定量聚合酶链式反应检测白介素-1β、白介素-6和肿瘤坏死因子αmRNA表达,蛋白印迹法检测citH3、肽酰基精氨酸脱亚氨酶4、基质金属蛋白酶2、基质金属蛋白酶9、沉默信息调节因子1(SIRT1)、乙酰化叉头转录因子1和乙酰化肿瘤抗原p53表达。结果Con+V组及Con+DNaseⅠ组TAAD发生率与破裂率均为0,BAPN+V组TAAD发生率和破裂率为90.0%和55.0%,BAPN+DNaseⅠ组TAAD发生率和破裂率为50.0%和30.0%。与Con+V组比较,BAPN+V组小鼠主动脉内径增大,血管壁扩张伴弹力纤维断裂明显;血清citH3、cfDNA和核小体水平上升,主动脉NETs生成增多;主动脉炎症和平滑肌细胞凋亡增加,SIRT1信号通路表达下调(P<0.01)。与BAPN+V组比较,BAPN+DNaseⅠ组小鼠主动脉内径减小,血管壁损伤及弹力纤维断裂减轻;血清citH3、cfDNA和核小体水平下降,主动脉NETs生成减少;主动脉炎症和平滑肌细胞凋亡减弱,SIRT1信号通路表达上调(P<0.01)。结论小鼠TAAD的发生与NETs抑制SIRT1信号通路进而上调胸主动脉炎症和促进平滑肌细胞凋亡有关,而NETs抑制剂DNaseⅠ能减少TAAD的形成,具有显著的血管保护效应。Objective To investigate the role and mechanism of neutrophil extracellular traps(NETs)in the formation of thoracic aortic aneurysm and dissection(TAAD)in mice.Methods A mouse TAAD model was induced byβ-aminopropionitrile(BAPN)administration(orally,diluted in drinking water,1 g/kg/day),and DNaseⅠ(20 mg/kg/day)was intraperitoneal injected to remove the generated NETs in vivo.Eighty 3-week-old male C57BL/6 mice were randomly divided into Con+V group,Con+DNaseⅠgroup,BAPN+V group and BAPN+DNaseⅠgroup.After 4 weeks,the incidence and rupture rate of TAAD were recorded.The maximum internal diameter of thoracic aorta was measured by the aortic ultrasonography.H&E and EVG staining were used to describe the aortic histological changes and elastin degradation.Serum levels of citH3,cfDNA and nucleosome were detected by special kits.citH3,MPO andα-SMA fluorescence intensity were detected by immunofluorescence staining.Apoptosis was observed by TUNEL staining.The mRNA expressions of IL-1β,IL-6 and TNF-αwere detected by qRT-PCR.The protein expressions of citH3,PAD4,MMP2,MMP9,SIRT1,Ac-foxo1 and Ac-p53 were detected by Western Blot.Results The incidence and rupture rate of TAAD in Con+V group and Con+DNaseⅠgroup were 0,the incidence and rupture rate of TAAD in BAPN+V group were 90.0%and 55.0%,and the incidence and rupture rate of TAAD in BAPN+DNaseⅠgroup were 50.0%and 30.0%.Compared with Con+V group,the maximum internal diameter of thoracic aorta in BAPN+V group was increased,the vascular wall became dilated,and the elastic fibers broken obviously.Besides,the serum levels of citH3,cfDNA and nucleosome were increased,and the formation of NETs in aorta in BAPN+V group was increased.Additionally,the inflammation of aorta tissue and apoptosis of smooth muscle cell were significantly enhanced,while the expression of SIRT1 signaling pathway was significantly down-regulated in BAPN+V group(P<0.01).Compared with BAPN+V group,the maximum internal diameter of thoracic aorta in BAPN+DNaseⅠgroup was decreased,the damage of va

关 键 词：中性粒细胞诱捕网 胸主动脉瘤 夹层动脉瘤 炎症 凋亡 沉默信息调节因子1 小鼠 

分 类 号：R543.5[医药卫生—心血管疾病]