Division of Pulmonary and Critical Care Medicine, Department of Medicine, Mayo Clinic, Rochester, MN, United States of America.
Department of Immunology, Mayo Clinic, Rochester, MN, United States of America.
PLoS One. 2024 Apr 19;19(4):e0301659. doi: 10.1371/journal.pone.0301659. eCollection 2024.
Clinical prediction of nontuberculous mycobacteria lung disease (NTM-LD) progression remains challenging. We aimed to evaluate antigen-specific immunoprofiling utilizing flow cytometry (FC) of activation-induced markers (AIM) and IFN-γ enzyme-linked immune absorbent spot assay (ELISpot) accurately identifies patients with NTM-LD, and differentiate those with progressive from nonprogressive NTM-LD. A Prospective, single-center, and laboratory technician-blinded pilot study was conducted to evaluate the FC and ELISpot based immunoprofiling in patients with NTM-LD (n = 18) and controls (n = 22). Among 18 NTM-LD patients, 10 NTM-LD patients were classified into nonprogressive, and 8 as progressive NTM-LD based on clinical and radiological features. Peripheral blood mononuclear cells were collected from patients with NTM-LD and control subjects with negative QuantiFERON results. After stimulation with purified protein derivative (PPD), mycobacteria-specific peptide pools (MTB300, RD1-peptides), and control antigens, we performed IFN-γ ELISpot and FC AIM assays to access their diagnostic accuracies by receiver operating curve (ROC) analysis across study groups. Patients with NTM-LD had significantly higher percentage of CD4+/CD8+ T-cells co-expressing CD25+CD134+ in response to PPD stimulation, differentiating between NTM-LD and controls. Among patients with NTM-LD, there was a significant difference in CD25+CD134+ co-expression in MTB300-stimulated CD8+ T-cells (p <0.05; AUC-ROC = 0.831; Sensitivity = 75% [95% CI: 34.9-96.8]; Specificity = 90% [95% CI: 55.5-99.7]) between progressors and nonprogressors. Significant differences in the ratios of antigen-specific IFN-γ ELISpot responses were also seen for RD1-nil/PPD-nil and RD1-nil/anti-CD3-nil between patients with nonprogressive vs. progressive NTM-LD. Our results suggest that multiparameter immunoprofiling can accurately identify patients with NTM-LD and may identify patients at risk of disease progression. A larger longitudinal study is needed to further evaluate this novel immunoprofiling approach.
临床预测非结核分枝杆菌肺病 (NTM-LD) 的进展仍然具有挑战性。我们旨在评估利用流式细胞术 (FC) 检测激活诱导标记物 (AIM) 和 IFN-γ 酶联免疫吸附斑点检测 (ELISpot) 的抗原特异性免疫谱是否能准确识别 NTM-LD 患者，并区分进展性和非进展性 NTM-LD 患者。进行了一项前瞻性、单中心、实验室技术员盲法的初步研究，以评估 NTM-LD 患者（n=18）和对照者（n=22）的 FC 和 ELISpot 基于免疫谱的检测。在 18 名 NTM-LD 患者中，根据临床和影像学特征，10 名 NTM-LD 患者被分为非进展性，8 名患者被分为进展性 NTM-LD。从 NTM-LD 患者和阴性 QuantiFERON 结果的对照者中采集外周血单核细胞。在刺激后用纯化蛋白衍生物 (PPD)、分枝杆菌特异性肽库 (MTB300、RD1-肽) 和对照抗原，我们进行 IFN-γ ELISpot 和 FC AIM 检测，通过接收者操作曲线 (ROC) 分析来评估它们在研究组中的诊断准确性。NTM-LD 患者对 PPD 刺激的 CD4+/CD8+ T 细胞共表达 CD25+CD134+的百分比显著更高，可将 NTM-LD 患者与对照者区分开。在 NTM-LD 患者中，MTB300 刺激的 CD8+ T 细胞中 CD25+CD134+的共表达存在显著差异（p<0.05；AUC-ROC=0.831；灵敏度=75% [95%CI:34.9-96.8]；特异性=90% [95%CI:55.5-99.7]），这一差异在进展者和非进展者之间存在。在非进展性与进展性 NTM-LD 患者之间，还观察到 RD1-nil/PPD-nil 和 RD1-nil/抗-CD3-nil 的抗原特异性 IFN-γ ELISpot 反应的比值存在显著差异。我们的结果表明，多参数免疫谱分析可准确识别 NTM-LD 患者，并可能识别出有疾病进展风险的患者。需要进一步进行更大的纵向研究来进一步评估这种新型免疫谱分析方法。
