Pisani Francesco, Porciani Caterina, Croia Cristina, Pucino Valentina, Virdis Agostino, Puxeddu Ilaria, Migliorini Paola, Pratesi Federico
Clinical Immunology and Allergy Unit, Department of Clinical and Experimental Medicine, University of Pisa, 56126 Pisa, Italy.
Geriatrics Unit, Department of Clinical and Experimental Medicine, University of Pisa, 56126 Pisa, Italy.
Int J Mol Sci. 2025 Feb 28;26(5):2221. doi: 10.3390/ijms26052221.
Neutrophil extracellular traps (NETs) are web-like structures composed of chromatin and proteins from neutrophil granules. Several studies highlight the heterogeneity of NETs, underscoring the challenges associated with their detection. In patients with COVID-19, high levels of NET fragments, called NET remnants, are detected in the circulation but also in alveoli and bronchioles. NET remnants are usually measured as complexes of DNA and myeloperoxidase (DNA-MPO). Taking advantage of proteomic data on NET composition, we developed new solid-phase assays to detect NET remnants, measuring complexes of DNA with alpha enolase (DNA-eno) or calprotectin (DNA-cal). The two assays were compared with the DNA-MPO test for the detection of in vitro-generated NET and serum NET remnants; all of them showed similar sensitivity in the detection of in vitro-generated NET. In an analysis of 40 patients with severe COVID-19 and 25 healthy subjects, the results of the three assays were highly correlated, and all detected significantly higher levels of NET remnants in patient sera. Moreover, the level of NET remnants correlated with impaired gas exchange and increased with the progressive decline of pulmonary function. The proposed assays thus represent a novel tool with which to evaluate NETosis; using antibodies to different NET constituents may allow their fingerprinting in different disorders.
中性粒细胞胞外诱捕网（NETs）是由中性粒细胞颗粒中的染色质和蛋白质组成的网状结构。多项研究强调了NETs的异质性，凸显了其检测所面临的挑战。在新冠肺炎患者中，循环系统以及肺泡和细支气管中均可检测到高水平的NET片段，即所谓的NET残余物。NET残余物通常作为DNA与髓过氧化物酶的复合物（DNA-MPO）进行测量。利用关于NET组成的蛋白质组学数据，我们开发了新的固相检测方法来检测NET残余物，即测量DNA与α烯醇化酶（DNA-eno）或钙卫蛋白（DNA-cal）的复合物。将这两种检测方法与DNA-MPO检测法进行比较，以检测体外生成的NET和血清NET残余物；它们在检测体外生成的NET时均表现出相似的灵敏度。在对40例重症新冠肺炎患者和25名健康受试者的分析中，三种检测方法的结果高度相关，且均检测到患者血清中NET残余物水平显著升高。此外，NET残余物水平与气体交换受损相关，并随着肺功能的逐渐下降而升高。因此，所提出的检测方法代表了一种评估NETosis的新工具；使用针对不同NET成分的抗体可能有助于在不同疾病中对其进行特征分析。
