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脓毒症小鼠急性肺损伤时ASAH1与细胞焦亡的关系

Association between ASAH1 and pyroptosis during acute lung injury in septic mice
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摘要 目的评价脓毒症小鼠急性肺损伤时N-酰基鞘氨醇酰胺水解酶1(ASAH1)与细胞焦亡的关系。方法SPF级健康雄性C57BL/6小鼠40只,6~8周龄,体质量18~23 g,采用随机数字表法将小鼠分为4组(n=10):假手术组(Sham组)、急性肺损伤组(ALI组)、HCFU溶剂+急性肺损伤组(HA组)和ASAH1抑制剂HCFU+急性肺损伤组(AA组)。Sham组仅开腹腔,ALI组、HA组和AA组行盲肠结扎穿孔术,HA组小鼠术前2 h腹腔注射HCFU溶剂0.2 ml,AA组小鼠术前2 h腹腔注射HCFU 10 mg/kg。24 h时深麻醉下处死小鼠,摘除眼球取血,收集支气管肺泡灌洗液(BALF)、肺组织和血清标本。计算肺湿质量/干质量(W/D)比值,采用BCA法检测BALF蛋白浓度,HE染色观察肺组织病理学结果并行肺损伤评分;ELISA法测定BALF IL-1β、IL-6和TNF-α浓度;Western blot法检测肺组织NOD样受体热蛋白结构域相关蛋白3(NLRP3)、Gasdermin D蛋白(GSDMD)、ASAH1和半胱氨酸蛋白酶-1(caspase-1)的表达。结果与Sham组相比,ALI组、HA组和AA组肺损伤评分、W/D比值、BALF蛋白、血清IL-1β、IL-6和TNF-α浓度升高,肺组织NLRP3、GSDMD和caspase-1表达上调,ALI组和HA组ASAH1表达上调(P<0.05);与ALI组和HA组相比,AA组肺损伤评分、W/D比值、BALF蛋白、血清IL-1β、IL-6和TNF-α浓度升高,肺组织NLRP3、GSDMD及caspase-1表达上调,ASAH1表达下调(P<0.05或0.01)。结论ASAH1参与了脓毒症小鼠急性肺损伤的内源性保护机制,可能与其抑制细胞焦亡有关。 Objective To evaluate the association between N-acylsphingosine amide hydrolase 1(ASAH1)and pyroptosis during acute lung injury(ALI)in septic mice.Methods Forty SPF-grade healthy male C57BL/6 mice,aged 6-8 weeks,weighing 18-23 g,were divided into 4 groups(n=10 each)by a random number table method:sham operation group(Sham group),ALI group,HCFU solvent+ALI group(HA group)and ASAH1 inhibitor HCFU+ALI group(AA group).The abdominal cavity was only opened in Sham group,and cecal ligation puncture was performed in ALI,HA and AA groups.HCFU solvent 0.2 ml was intraperitoneally injected at 2 h before operation in HA group,and HCFU 10 mg/kg was intraperitoneally injected at 2 h before operation in AA group.The mice were sacrificed at 24 h under deep anesthesia,the eyeballs were removed to collect the blood,the bronchoalveolar lavage fluid(BALF)was collected,and lung tissues and blood samples were collected for microscopic examination of the pathological changes(using HE staining)which were scored and for determination of concentrations of protein in BALF(by BCA method),concentrations of interleukin-1beta(IL-1β),IL-6 and tumor necrosis factor-alpha(TNF-α)in BALF(by enzyme-linked immunosorbent assay),and expression of NOD-like receptor thermoprotein structural domain-related protein 3(NLRP3)in lung tissues(by Western blot),gasdermin D protein(GSDMD),ASAH1 and cysteine protease-1(caspase-1)(by Western blot).The wet/dry lung weight(W/D)ratio was calculated.Results Compared with Sham group,the lung injury score,W/D ratio and concentrations of protein in BALF,IL-1β,IL-6 and TNF-αin serum were significantly increased,and the expression of NLRP3,GSDMD and caspase-1 in lung tissues was up-regulated in ALI,HA and AA groups,and the expression of ASAH1 was significantly up-regulated in ALI and HA groups(P<0.05).Compared with ALI and HA groups,the lung injury score,W/D ratio,and concentrations of protein in BALF,IL-1β,IL-6 and TNF-αin serum were significantly increased,the expression of NLRP3,GSDMD and caspase-1 in lung tissues was up-regulated,and the expression of ASAH1 was down-regulated in AA group(P<0.05 or 0.01).Conclusions ASAH1 is involved in the endogenous protective mechanism underlying ALI in septic mice,which may be related to the inhibition of cell pyroptosis.
作者 王蓓莹 李大鹏 张丹丹 秦伟伟 孙立新 韩伟 Wang Beiying;Li Dapeng;Zhang Dandan;Qin Weiwei;Sun Lixin;Han Wei(Qingdao Clinical Medical College,Nanjing Medical University,Qingdao 266071,China;Department of Trauma Surgery,Qingdao Municipal Hospital,Qingdao 266071,China;Department of Anesthesiology,Qingdao Municipal Hospital,Qingdao 266071,China;Department of Respiratory and Critical Care Medicine,Qingdao Municipal Hospital,Qingdao 266071,China)
出处 《中华麻醉学杂志》 CAS CSCD 北大核心 2024年第7期876-880,共5页 Chinese Journal of Anesthesiology
关键词 脓毒症 急性肺损伤 N-酰基鞘氨醇酰胺水解酶1 细胞焦亡 Sepsis Acute lung injury ASAH1 Pyroptosis
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