肝泡型棘球蚴病患者中性粒细胞胞外诱捕网表达及吞噬功能初探
Expression of neutrophil extracellular traps and phagocytic functions among patients with hepatic alveolar echinococcosis
仁增卓嘎 1樊海宁 2杨康洁 3王志鑫 2张耀刚 4芦永良 5王海久2
作者信息
- 1. 青海大学附属医院肝胆胰外科(青海西宁 810001);青海省包虫病研究重点实验室(青海西宁 810001);西藏自治区拉萨市人民医院麻醉科(西藏拉萨 850000)
- 2. 青海大学附属医院肝胆胰外科(青海西宁 810001);青海省包虫病研究重点实验室(青海西宁 810001)
- 3. 山东大学第二医院检验医学中心(山东济南 250033)
- 4. 青海省包虫病研究重点实验室(青海西宁 810001)
- 5. 青海大学附属医院检验科(青海西宁 810001)
- 折叠
摘要
目的 探索肝泡型棘球蚴病(hepatic alveolar echinococcosis,HAE)患者外周血中中性粒细胞胞外诱捕网(neu-trophil extracellular traps,NETs)表达水平及吞噬功能变化,以及其与临床炎性指标和肝生化指标间的相关性.方法 选取2022年8月—2023年6月青海大学附属医院肝胆外科收治的50例HAE患者作为HAE患者组,选择同期在该院体检中心体检的年龄、性别匹配的50例健康志愿者作为健康对照组,采用酶联免疫吸附法(enzyme-linked immunosorbent as-say,ELISA)检测两组研究对象外周血中NETs标志物髓过氧化物酶(myeloperoxidase,MPO)和中性粒细胞弹性酶(neutro-phil elastase,NE)水平.同时,采用密度梯度离心法分离两组研究对象外周血中性粒细胞;经佛波酯(phorbol 12-myristate 13-acetate,PMA)体外刺激后,采用流式细胞术分别定量检测中性粒细胞胞外释放的MPO和瓜氨酸化组蛋白(citrullinat-ed histone H3,CitH3)水平.采用流式细胞术比较两组研究对象中性粒细胞吞噬能力差异,采用Spearman相关分析评价HAE患者NETs标志物含量与临床炎性指标及肝生化指标水平间的相关性.结果 HAE患者外周血中NETs标志物MPO、NE、CitH3水平分别为(417.15±76.08)、(23.16±6.75)、(33.93±18.93)ng/mL,均显著高于健康对照组[(255.70±80.84)、(11.92±3.17)、(19.52±13.89)ng/mL](t = 10.28、10.65、4.34,P均<0.05);但HAE组中性粒细胞吞噬率[(70.85±7.32)%]较健康对照组[(94.04±3.90)%]下降(t= 20.18,P<0.05);经PMA体外刺激后,HAE患者组中性粒细胞形成NETs能力较健康对照组显著提高.Spearman相关分析显示,HAE组外周血中性粒细胞吞噬水平和血小板与淋巴细胞比例(platelet-to-lymphocyte ratio,PLR)、中性粒细胞与淋巴细胞比例(neutrophil-to-lymphocyte ratio,NLR)、白细胞介素(in-terleukin,IL)-6、C反应蛋白(C reactive protein,CRP)水平呈负相关(rs =-0.515～-0.392,P均<0.05),NETs标志物MPO、NE水平与NLR、PLR、CRP、IL-6水平均呈正相关(rs = 0.333～0.445,P均<0.05),与天冬氨酸氨基转移酶、丙氨酸氨基转移酶、直接胆红素和总胆红素水平亦呈正相关(rs = 0.290～0.628,P均<0.001).结论 HAE患者体内NETs形成过多,影响中性粒细胞吞噬能力,导致机体炎症水平升高.NETs标志物有望作为肝脏疾病早期诊断、监测和严重程度评估的新型生物学标志物.
Abstract
Objective To investigate the expression of neutrophil extracellular traps(NETs)and phagocytic function in the peripheral blood of patients with hepatic alveolar echinococcosis(HAE),and to examine their correlations with clinical inflamma-tory indicators and liver functions.Methods A total of 50 patients with HAE admitted to Department of Hepatobiliary and Pan-creatic Surgery,The Affiliated Hospital of Qinghai University from August 2022 to June 2023 were enrolled,while 50 age-and gender-matched healthy individuals from the Centre for Healthy Examinations of the hospital during the same period served as controls.The levels of NETs markers neutrophil myeloperoxidase(MPO)and neutrophil elastase(NE)were measured using en-zyme-linked immunosorbent assay(ELISA).Peripheral blood neutrophils were isolated using density gradient centrifugation,stimulated in vitro using phorbol 12-myristate 13 acetate(PMA),and the levels of MPO and citrullination histone H3(CitH3)re-leased by neutrophils were quantified using flow cytometry.The phagocytic functions of neutrophils were examined using flow cy-tometry.In addition,the correlations of MPO and NE levels with clinical inflammatory indicators and liver biochemical indicators were examined using Spearman correlation analysis among HAE patients.Results The peripheral blood plasma MPO[(417.15±76.08)ng/mL vs.(255.70±80.84)ng/mL;t = 10.28,P<0.05],NE[(23.16±6.75)ng/mL vs.(11.92±3.17)ng/mL;t = 10.65,P<0.05]and CitH3 levels[(33.93±18.93)ng/mL vs.(19.52±13.89)ng/mL;t = 4.34,P<0.05]were all significantly higher among HAE patients than among healthy controls,and a lower phagocytosis rate of neutrophils was detected among HAE patients than among healthy controls[(70.85±7.32)%vs.(94.04±3.90)%;t = 20.18,P<0.05],and the ability to produce NETs by neu-trophils was higher among HAE patients than among healthy controls following in vitro PMA stimulation.Pearson correlation analysis showed that the phagocytosis rate of neutrophils correlated negatively with platelet-to-lymphocyte ratio(PLR),neutrophil-to-lymphocyte ratio(NLR),interleukin-6(IL-6)level and C-reactive protein(CRP)level(rs =-0.515 to-0.392,all P values<0.05),and the MPO and NE levels positively correlated with inflammatory markers NLR,PLR,CRP and IL-6(rs = 0.333 to 0.445,all P values<0.05)and clinical liver biochemical indicators aspartic transaminase,alanine aminotransferase,direct bilirubin and total bilirubin among HAE patients(rs = 0.290 to 0.628,all P values<0.001).Conclusions Excessive formation of NETs is found among HAE patients,which affects the phagocytic ability of neutrophils and results in elevated levels of inflammatory in-dicators.NETs markers may be promising novel biomarkers for early diagnosis,monitoring,and severity assessment of liver dis-ease.
关键词
肝泡型棘球蚴病/吞噬能力/中性粒细胞胞外诱捕网/炎症水平Key words
Hepatic alveolar echinococcosis/Phagocytosis/Neutrophil extracellular trap/Inflammatory level引用本文复制引用
基金项目
青海省科技厅临床医学研究中心项目(2017-SF-L2)
青海大学青年科研基金(2021)(2021-QYY-9)
出版年
2024
NETL
NSTL
万方数据