Objectives Type I IFN (IFN-I) activation is a prominent feature of primary SS (pSS), SLE and SSc. Ultrasensitive single-molecule array (Simoa) technology has facilitated the measurement of subfemtomolar concentrations of IFNs. Here we aimed to measure IFN-α2 in serum from pSS, SLE and SSc using a Simoa immunoassay and correlate these levels to blood IFN-stimulated gene (ISG) expression and disease activity. Methods Serum IFN-α2 was measured in patients with pSS (n = 85 and n = 110), SLE (n = 24) and SSc (n = 23) and healthy controls (HCs; n = 68) using an IFN-α Simoa assay on an HD-X analyser. IFN-I pathway activation was additionally determined from serum by an IFN-I reporter assay and paired samples of whole blood ISG expression of IFI44, IFI44L, IFIT1, IFIT3 and MxA by RT-PCR or myxovirus resistance protein 1 (MxA) protein ELISA. Results Serum IFN-α2 levels were elevated in pSS (median 61.3 fg/ml) compared with HCs (median ≤5 fg/ml, P < 0.001) and SSc (median 11.6 fg/ml, P = 0.043), lower compared with SLE (median 313.5 fg/ml, P = 0.068) and positively correlated with blood ISG expression (r = 0.66–0.94, P < 0.001). Comparable to MxA ELISA [area under the curve (AUC) 0.93], IFN-α2 measurement using Simoa identified pSS with high ISG expression (AUC 0.90) with 80–93% specificity and 71–84% sensitivity. Blinded validation in an independent pSS cohort yielded a comparable accuracy. Multiple regression indicated independent associations of autoantibodies, IgG, HCQ treatment, cutaneous disease and a history of extraglandular manifestations with serum IFN-α2 concentrations in pSS. Conclusion Simoa serum IFN-α2 reflects blood ISG expression in pSS, SLE and SSc. In light of IFN-targeting treatments, Simoa could potentially be applied for patient stratification or retrospective analysis of historical cohorts.

中文翻译：

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通过单分子阵列测量的血清干扰素-α2与干燥综合征的全身性疾病表现相关

目标 I 型干扰素 (IFN-I) 激活是原发性 SS (pSS)、SLE 和 SSc 的一个突出特征。超灵敏单分子阵列 (Simoa) 技术有助于测量亚飞摩尔浓度的 IFN。在这里，我们旨在使用 Simoa 免疫测定法测量 pSS、SLE 和 SSc 血清中的 IFN-α2，并将这些水平与血液 IFN 刺激基因 (ISG) 表达和疾病活动相关联。方法 使用 IFN-α Simoa 测量 pSS（n = 85 和 n = 110）、SLE（n = 24）和 SSc（n = 23）和健康对照（HCs；n = 68）患者的血清 IFN-α2在 HD-X 分析仪上进行分析。另外，通过 IFN-I 报告基因测定从血清中测定 IFN-I 通路激活，并通过 RT-PCR 或粘病毒抗性蛋白 1 (MxA) 蛋白 ELISA 对 IFI44、IFI44L、IFIT1、IFIT3 和 MxA 的全血 ISG 表达的配对样品进行测定。结果 pSS（中位数 61.3 fg/ml）与 HC（中位数 ≤5 fg/ml，P < 0.001）和 SSc（中位数 11.6 fg/ml，P = 0.043）相比，血清 IFN-α2 水平升高，低于SLE（中位数 313.5 fg/ml，P = 0.068）并与血液 ISG 表达呈正相关（r = 0.66-0.94，P < 0.001）。与 MxA ELISA [曲线下面积 (AUC) 0.93] 相比，使用 Simoa 进行的 IFN-α2 测量确定了具有高 ISG 表达 (AUC 0.90) 的 pSS，特异性为 80-93%，敏感性为 71-84%。独立 pSS 队列中的盲法验证产生了相当的准确性。多元回归表明自身抗体、IgG、HCQ 治疗、皮肤病和腺外表现史与 pSS 中血清 IFN-α2 浓度的独立关联。结论 Simoa 血清 IFN-α2 反映了 pSS、SLE 和 SSc 中血液 ISG 的表达。