Nordic Bioscience A/S, Herlev, Denmark.
Lund University, Rheumatology and Molecular Skeletal Biology, Department of Clinical Sciences, Lund, Sweden.
J Crohns Colitis. 2022 Sep 8;16(9):1447-1460. doi: 10.1093/ecco-jcc/jjac047.
Endoscopy and the use of faecal calprotectin [faecal CP] are among the least-favoured methods for assessing disease activity by inflammatory bowel disease [IBD] patients; the handling/processing of faecal samples is also impractical. Therefore, we sought to develop a novel neo-epitope serum calprotectin enzyme-linked immunosorbent assay [ELISA], CPa9-HNE, with the aim of quantifying neutrophil activity and neutrophil extracellular trap [NET]-osis and proposing a non-invasive method for monitoring disease activity in IBD patients.
In vitro cleavage was performed by mixing calprotectin [S100A9/S100A8] with human neutrophil elastase [HNE], and a novel HNE-derived calprotectin neo-epitope [CPa9-HNE] was identified by mass spectrometry for ELISA development. The CPa9-HNE ELISA was quantified in supernatants from ex vivo activated neutrophils and serum samples from patients with ulcerative colitis [UC, n = 43], Crohn's disease [CD, n = 93], and healthy subjects [HS, n = 23]. For comparison, faecal CP and MRP8/14 biomarkers were also measured.
CPa9-HNE was specific for activated neutrophils ex vivo. Serum CPa9-HNE levels were 4-fold higher in CD [p <0.0001] and UC [p <0.0001] patients than in HS. CPa9-HNE correlated well with the Simple Endoscopic Score [SES]-CD score [r = 0.61, p <0.0001], MES [r = 0.46, p = 0.0141], and the full Mayo score [r = 0.52, p = 0.0013]. CPa9-HNE was able to differentiate between CD and UC patients in endoscopic remission and moderate/severe disease activity (CD: area under the curve [AUC] = 0.82 [p = 0.0003], UC: AUC = 0.87 [p = 0.0004]). The performance of CPa9-HNE was equipotent or slightly better than that of faecal CP.
Serum CPa9-HNE levels were highly associated with CD and UC patients. CPa9-HNE correlated with the SES-CD score and the full Mayo score, indicating a strong association with disease activity.
内镜检查和粪便钙卫蛋白（粪便 CP）检测是炎症性肠病（IBD）患者评估疾病活动度的最不优选方法之一；粪便样本的处理也不切实际。因此，我们试图开发一种新型的新型neo-epitope 血清钙卫蛋白酶联免疫吸附试验（ELISA），CPa9-HNE，旨在定量中性粒细胞活性和中性粒细胞细胞外陷阱（NET）形成，并提出一种非侵入性方法来监测 IBD 患者的疾病活动度。
通过混合钙卫蛋白（S100A9/S100A8）与人中性粒细胞弹性蛋白酶（HNE）进行体外切割，并通过质谱鉴定一种新型的 HNE 衍生的钙卫蛋白新表位（CPa9-HNE）用于 ELISA 开发。在体外激活的中性粒细胞上清液和溃疡性结肠炎（UC，n=43）、克罗恩病（CD，n=93）和健康受试者（HS，n=23）的血清样本中定量 CPa9-HNE ELISA。为了比较，还测量了粪便 CP 和 MRP8/14 生物标志物。
CPa9-HNE 是体外激活中性粒细胞的特异性标志物。CD [p<0.0001] 和 UC [p<0.0001] 患者的血清 CPa9-HNE 水平比 HS 高 4 倍。CPa9-HNE 与简单内镜评分（SES-CD）评分 [r=0.61，p<0.0001]、MES [r=0.46，p=0.0141] 和完整 Mayo 评分 [r=0.52，p=0.0013] 相关性良好。CPa9-HNE 能够区分内镜缓解和中度/重度疾病活动度的 CD 和 UC 患者（CD：曲线下面积 [AUC]=0.82 [p=0.0003]，UC：AUC=0.87 [p=0.0004]）。CPa9-HNE 的性能与粪便 CP 相当或略优。
血清 CPa9-HNE 水平与 CD 和 UC 患者高度相关。CPa9-HNE 与 SES-CD 评分和完整 Mayo 评分相关，表明与疾病活动度有很强的关联。
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