PDF(Pubmed)
Abstract：
BACKGROUND: Neutrophil extracellular traps (NETs) have repeatedly been related to COVID-19 severity and mortality. However, there is no consensus on their quantification, and there are scarce data on their evolution during the disease. We studied circulating NET markers in patients with COVID-19 throughout their hospitalization.
METHODS: We prospectively included 93 patients (201 blood samples), evaluating the disease severity in 3 evolutionary phases (viral, early, and late inflammation). Of these, 72 had 180 samples in various phases. We also evaluated 55 controls with similar age, sex and comorbidities. We measured 4 NET markers in serum: cfDNA, CitH3, and MPO-DNA and NE-DNA complexes; as well as neutrophil-related cytokines IL-8 and G-CSF.
RESULTS: The COVID-19 group had higher CitH3 (28.29 vs 20.29 pg/mL, p = 0.022), and cfDNA, MPO-DNA, and NE-DNA (7.87 vs 2.56 ng/mL; 0.80 vs 0.52 and 1.04 vs 0.72, respectively, p < 0.001 for all) than the controls throughout hospitalisation. cfDNA was the only NET marker clearly related to severity, and it remained higher in non-survivors during the 3 phases. Only cfDNA was an independent risk factor for mortality and need for intensive care. Neutrophil count, IL-8, and G-CSF were significantly related to severity. MPO-DNA and NE-DNA showed significant correlations (r: 0.483, p < 0.001), including all 3 phases and across all severity grades, and they only remained significantly higher on days 10-16 of evolution in those who died. Correlations among the other NET markers were lower than expected.
CONCLUSIONS: The circulating biomarkers of NETs were present in patients with COVID-19 throughout hospitalization. cfDNA was associated with severity and mortality, but the three other markers showed little or no association with these outcomes. Neutrophil activity and neutrophil count were also associated with severity. MPO-DNA and NE-DNA better reflected NET formation. cfDNA appeared to be more associated with overall tissue damage; previous widespread use of this marker could have overestimated the relationship between NETs and severity. Currently, there are limitations to accurate NET markers measurement that make it difficult to assess its true role in COVID-19 pathogenesis.
METHODS: We prospectively included 93 patients (201 blood samples), evaluating the disease severity in 3 evolutionary phases (viral, early, and late inflammation). Of these, 72 had 180 samples in various phases. We also evaluated 55 controls with similar age, sex and comorbidities. We measured 4 NET markers in serum: cfDNA, CitH3, and MPO-DNA and NE-DNA complexes; as well as neutrophil-related cytokines IL-8 and G-CSF.
RESULTS: The COVID-19 group had higher CitH3 (28.29 vs 20.29 pg/mL, p = 0.022), and cfDNA, MPO-DNA, and NE-DNA (7.87 vs 2.56 ng/mL; 0.80 vs 0.52 and 1.04 vs 0.72, respectively, p < 0.001 for all) than the controls throughout hospitalisation. cfDNA was the only NET marker clearly related to severity, and it remained higher in non-survivors during the 3 phases. Only cfDNA was an independent risk factor for mortality and need for intensive care. Neutrophil count, IL-8, and G-CSF were significantly related to severity. MPO-DNA and NE-DNA showed significant correlations (r: 0.483, p < 0.001), including all 3 phases and across all severity grades, and they only remained significantly higher on days 10-16 of evolution in those who died. Correlations among the other NET markers were lower than expected.
CONCLUSIONS: The circulating biomarkers of NETs were present in patients with COVID-19 throughout hospitalization. cfDNA was associated with severity and mortality, but the three other markers showed little or no association with these outcomes. Neutrophil activity and neutrophil count were also associated with severity. MPO-DNA and NE-DNA better reflected NET formation. cfDNA appeared to be more associated with overall tissue damage; previous widespread use of this marker could have overestimated the relationship between NETs and severity. Currently, there are limitations to accurate NET markers measurement that make it difficult to assess its true role in COVID-19 pathogenesis.
摘要：
背景：中性粒细胞胞外陷阱（NETs）与COVID-19的严重程度和死亡率反复相关。然而,对它们的量化没有共识，关于它们在疾病期间的演变的数据很少。我们研究了COVID-19患者在整个住院期间的循环NET标记。
方法：我们前瞻性纳入93例患者（201份血液样本），评估3个进化阶段的疾病严重程度(病毒，早期，和晚期炎症）。其中,72具有180个不同相的样品。我们还评估了55名年龄相似的对照，性和合并症。我们测量了血清中的4个NET标记：cfDNA，CitH3和MPO-DNA和NE-DNA复合物;以及中性粒细胞相关细胞因子IL-8和G-CSF。
结果：COVID-19组的CitH3较高（28.29比20.29pg/mL，p=0.022),和cfDNA，MPO-DNA,和NE-DNA（分别为7.87对2.56ng/mL；0.80对0.52和1.04对0.72，在整个住院期间，p<0.001）均高于对照组。cfDNA是唯一与严重程度明显相关的NET标记，在三个阶段中，非幸存者的比例仍然更高。只有cfDNA是死亡和需要重症监护的独立危险因素。中性粒细胞计数,IL-8和G-CSF与严重程度显著相关。MPO-DNA和NE-DNA呈显著相关(r:0.483,p<0.001),包括所有3个阶段和所有严重性等级，在进化的第10-16天，它们仅在死亡的人中保持明显更高的水平。其他NET标记之间的相关性低于预期。
结论：COVID-19患者在整个住院期间都存在NETs的循环生物标志物。cfDNA与严重程度和死亡率相关，但其他三个标记与这些结果几乎没有或没有关联.中性粒细胞活性和中性粒细胞计数也与严重程度相关。MPO-DNA和NE-DNA更好地反映了NET的形成。cfDNA似乎与整体组织损伤更相关；以前广泛使用该标记可能高估了NET和严重程度之间的关系。目前,准确的NET标志物测量存在局限性，这使得很难评估其在COVID-19发病机制中的真正作用。
方法：我们前瞻性纳入93例患者（201份血液样本），评估3个进化阶段的疾病严重程度(病毒，早期，和晚期炎症）。其中,72具有180个不同相的样品。我们还评估了55名年龄相似的对照，性和合并症。我们测量了血清中的4个NET标记：cfDNA，CitH3和MPO-DNA和NE-DNA复合物;以及中性粒细胞相关细胞因子IL-8和G-CSF。
结果：COVID-19组的CitH3较高（28.29比20.29pg/mL，p=0.022),和cfDNA，MPO-DNA,和NE-DNA（分别为7.87对2.56ng/mL；0.80对0.52和1.04对0.72，在整个住院期间，p<0.001）均高于对照组。cfDNA是唯一与严重程度明显相关的NET标记，在三个阶段中，非幸存者的比例仍然更高。只有cfDNA是死亡和需要重症监护的独立危险因素。中性粒细胞计数,IL-8和G-CSF与严重程度显著相关。MPO-DNA和NE-DNA呈显著相关(r:0.483,p<0.001),包括所有3个阶段和所有严重性等级，在进化的第10-16天，它们仅在死亡的人中保持明显更高的水平。其他NET标记之间的相关性低于预期。
结论：COVID-19患者在整个住院期间都存在NETs的循环生物标志物。cfDNA与严重程度和死亡率相关，但其他三个标记与这些结果几乎没有或没有关联.中性粒细胞活性和中性粒细胞计数也与严重程度相关。MPO-DNA和NE-DNA更好地反映了NET的形成。cfDNA似乎与整体组织损伤更相关；以前广泛使用该标记可能高估了NET和严重程度之间的关系。目前,准确的NET标志物测量存在局限性，这使得很难评估其在COVID-19发病机制中的真正作用。
