Systemic Autoimmunity Branch, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD, United States.
Department of Immunology and Rheumatology, Instituto Nacional de Ciencias Medicas y de la Nutricion, Salvador Zubiran, Mexico City, Mexico.
Front Immunol. 2021 Sep 14;12:715997. doi: 10.3389/fimmu.2021.715997. eCollection 2021.
Antibodies against carbamylated proteins (anti-CarP) are associated with poor prognosis and the development of bone erosions in rheumatoid arthritis (RA). RA neutrophils externalize modified autoantigens through the formation of neutrophil extracellular traps (NETs). Increased levels of the cathelicidin LL37 have been documented in the synovium of RA patients, but the cellular source remains unclear. We sought to determine if post-translational modifications of LL37, specifically carbamylation, occur during NET formation, enhance this protein's autoantigenicity, and contribute to drive bone erosion in the synovial joint.
ELISA and Western blot analyses were used to identify carbamylated LL37 (carLL37) in biological samples. Anti-carLL37 antibodies were measured in the serum of HLA-DRB1*04:01 transgenic mice and in human RA synovial fluid.
Elevated levels of carLL37 were found in plasma and synovial fluid from RA patients, compared to healthy controls. RA NETs release carLL37 and fibroblast-like synoviocytes (FLS) internalized NET-bound carLL37 and loaded it into their MHCII compartment. HLA-DRB1*04:01 transgenic mice immunized with FLS containing NETs developed autoantibodies against carLL37. Anti-carLL37 antibodies were present in RA sera and synovial fluid and they correlated with radiologic bone erosion scores of the hands and feet in RA patients. CarLL37-IgG immune complexes enhanced the ability of monocytes to differentiate into osteoclasts and potentiated osteoclast-mediated extracellular matrix resorption.
NETs are a source of carLL37 leading to induction of anti-carbamylated autoantibody responses. Furthermore, carLL37-IgG immune complexes may be implicated in the bone damage characteristic of RA. These results support that dysregulated NET formation has pathogenic roles in RA.
抗碳化蛋白抗体（anti-CarP）与类风湿关节炎（RA）的预后不良和骨侵蚀的发展有关。RA 中性粒细胞通过形成中性粒细胞细胞外陷阱（NETs）将修饰的自身抗原外化。已经在 RA 患者的滑膜中记录到 cathelicidin LL37 的水平增加，但细胞来源尚不清楚。我们试图确定 LL37 的翻译后修饰，特别是碳化，是否发生在 NET 形成过程中，增强这种蛋白质的自身抗原性，并有助于驱动滑膜关节的骨侵蚀。
ELISA 和 Western blot 分析用于鉴定生物样品中的碳化 LL37（carLL37）。在 HLA-DRB1*04:01 转基因小鼠的血清和人类 RA 滑膜液中测量抗 carLL37 抗体。
与健康对照相比，RA 患者的血浆和滑膜液中发现 carLL37 水平升高。RA NETs 释放 carLL37，成纤维样滑膜细胞（FLS）内化 NET 结合的 carLL37，并将其加载到其 MHCII 隔室中。用含有 NET 的 FLS 免疫的 HLA-DRB1*04:01 转基因小鼠产生针对 carLL37 的自身抗体。抗 carLL37 抗体存在于 RA 血清和滑膜液中，与 RA 患者手和脚的放射学骨侵蚀评分相关。CarLL37-IgG 免疫复合物增强单核细胞分化为破骨细胞的能力，并增强破骨细胞介导的细胞外基质吸收。
NETs 是导致抗碳化自身抗体反应的 carLL37 的来源。此外，carLL37-IgG 免疫复合物可能与 RA 特征性的骨损伤有关。这些结果支持失调的 NET 形成在 RA 中具有致病性作用。
