Serum anti-PAD4 autoantibodies are present in cystic fibrosis children and increase with age and lung disease severity

Cystic fibrosis (CF) lung disease begins early in childhood and is characterized by neutrophilic inflammation of the airways. Neutrophil extracellular traps (NETs) represent one mechanism by which neutrophils contribute to lung damage. The enzyme peptidylarginine deiminase 4 (PAD4) is required for NET formation. Our overall concept is that NET formation delivers PAD4 outside the neutrophil resulting in autoantibody generation, and this autoimmunity may be a novel mechanism contributing to CF lung disease progression. The aim of this study was to investigate clinical predictors of serum anti-PAD4 autoantibody (PAD4 Ab) levels in CF subjects with a wide range of ages from early childhood through middle age. We measured PAD4 Ab levels in sera from 104 CF subjects. PAD4 Abs were detectable among CF children as young as one year of age and elevated compared to paediatric healthy controls. PAD4 Ab levels increased significantly with age (r = 0.584, p r = −0.481, n = 99, p Pseudomonas aeruginosa airways infection (p CFTR genotype, sweat chloride, pancreatic enzyme use, nutritional status, recent pulmonary exacerbations, Staphylococcus aureus, or CF-related diabetes. PAD4 Ab levels were also correlated with serum anti-double-stranded DNA IgA autoantibodies, which have similarly been shown to be elevated in CF subjects and associated with lung damage. In multivariable analysis, age and lung function remained correlated with PAD4 Ab levels. In summary, we describe novel findings of anti-PAD4 autoantibodies in CF that are present early in childhood, increase over time with age, and correlate with lung disease severity. Autoimmunity to antigens extruded by NETs appears to be an early event in CF lung disease, and airway autoimmunity related to NET formation is a potential mechanism of lung disease progression in CF.HighlightsSerum anti-PAD4 autoantibodies are detected in paediatric CF serum and are elevated compared to healthy paediatric controlsAnti-PAD4 autoantibodies increase with ageAnti-PAD4 autoantibodies correlate with lower lung function, Pseudomonas aeruginosa airway infection and anti-dsDNA IgA autoantibodies, but not with other key clinical CF co-variatesAge and lung function remain correlated with anti-PAD4 autoantibodies in multivariable analysis Serum anti-PAD4 autoantibodies are detected in paediatric CF serum and are elevated compared to healthy paediatric controls Anti-PAD4 autoantibodies increase with age Anti-PAD4 autoantibodies correlate with lower lung function, Pseudomonas aeruginosa airway infection and anti-dsDNA IgA autoantibodies, but not with other key clinical CF co-variates Age and lung function remain correlated with anti-PAD4 autoantibodies in multivariable analysis

囊性纤维化（Cystic Fibrosis, CF）肺部疾病起病于儿童早期，以气道中性粒细胞性炎症为核心病理特征。中性粒细胞胞外陷阱（Neutrophil Extracellular Traps, NETs）是中性粒细胞介导肺损伤的关键机制之一。肽酰精氨酸脱亚胺酶4（Peptidylarginine Deiminase 4, PAD4）是NET形成所必需的酶。本研究的核心假说为：NET形成会将PAD4释放至中性粒细胞外，进而诱导自身抗体生成，而这种自身免疫可能是驱动CF肺部疾病进展的全新机制。
本研究旨在探究覆盖儿童早期至中年的宽年龄段CF受试者血清抗PAD4自身抗体（PAD4 Ab）水平的临床预测因子。我们对104名CF受试者的血清样本进行了PAD4 Ab水平检测。结果显示，年仅1岁的CF儿童即可检出PAD4 Ab，且其水平较儿科健康对照显著升高。PAD4 Ab水平随年龄增长显著升高（r=0.584, p<0.001），同时与肺功能指标呈显著负相关（r=-0.481, n=99, p<0.001）。此外，PAD4 Ab水平与铜绿假单胞菌（Pseudomonas aeruginosa）气道感染（p<0.001）显著相关，但与囊性纤维化跨膜传导调节因子（CFTR）基因型、汗液氯离子浓度、胰酶使用情况、营养状况、近期肺部急性加重史、金黄色葡萄球菌（Staphylococcus aureus）感染及囊性纤维化相关糖尿病无显著关联。
PAD4 Ab水平还与血清抗双链DNA IgA自身抗体呈显著相关，此类自身抗体在CF受试者中同样升高，且既往研究已证实其与肺损伤密切相关。多变量分析显示，年龄与肺功能仍与PAD4 Ab水平保持显著相关性。
综上，本研究揭示了CF患者体内抗PAD4自身抗体的全新发现：此类抗体在儿童早期即可检出，随年龄增长逐渐升高，且与肺部疾病严重程度显著相关。由NETs释放的抗原介导的自身免疫似乎是CF肺部疾病的早期事件，而与NET形成相关的气道自身免疫可能是CF肺部疾病进展的潜在机制。

研究亮点：
1. 在儿科CF患者的血清中可检测到抗PAD4自身抗体，且其水平较健康儿科对照显著升高
2. 抗PAD4自身抗体水平随年龄增长而升高
3. 抗PAD4自身抗体与肺功能降低、铜绿假单胞菌气道感染及抗dsDNA IgA自身抗体水平呈显著相关，但与其他关键临床CF协变量无关联
4. 多变量分析显示，年龄与肺功能仍与抗PAD4自身抗体水平保持显著相关性