第一部分 免疫治疗方案及停药对重症肌无力复发风险的影响

背景与目的：糖皮质激素是重症肌无力的一线免疫治疗方案，但最佳的激素剂量方案及停用激素对复发风险的影响仍不明确。本研究旨在探讨激素剂量方案和激素停药对接受激素单药治疗的患者复发风险的影响。

研究方法：本研究是一项基于前瞻性登记数据库的队列研究，纳入在激素单药治疗后达到最小临床表现或更好状态的患者。主要结局为复发。采用组轨迹模型识别不同激素剂量方案，并结合Cox比例风险模型和倾向评分匹配分析评估不同激素剂量方案和激素停药的影响。

研究结果：本研究共纳入209例患者（中位随访时间为54.0个月），其中113例（54.1%）发生复发，68例（32.5%）停用了激素。组轨迹模型识别三种治疗方案：“大剂量起始快速减量”（方案1）、“中等剂量起始逐渐减量”（方案2）以及“小剂量起始缓慢减量”（方案3）。这些剂量方案与根据疾病严重程度调整剂量方案的临床实践一致。与方案1相比，方案2（HR = 0.26, 95% CI 0.14-0.48, p < 0.001）和方案3（HR = 0.14, 95% CI 0.07-0.31, p < 0.001）的复发风险更低。激素停药后的中位复发时间为7.0个月。倾向评分匹配分析显示，与小剂量维持激素相比，激素停药显著增加了1年内的复发风险（HR = 1.61, p = 0.031）。

结论：需要大剂量激素治疗患者的疾病严重程度与复发风险更高。与小剂量维持激素相比，激素停药显著增加了复发风险，因此在激素单药治疗的患者中停药应谨慎。

第二部分 基于质谱流式细胞技术构建重症肌无力患者外周血免疫细胞图谱及临床相关生物标志物的探索

背景与目的：重症肌无力（Myasthenia gravis，MG）是一种T细胞依赖、B细胞介导的自身免疫性疾病，特征为针对神经肌肉接头的自身抗体的产生。然而，目前关于MG患者外周血免疫细胞图谱的全面分析仍然缺乏，也缺少能反映疾病严重程度和预测治疗反应的生物标志物。本研究通过质谱流式细胞术，构建并比较了MG患者与健康对照、治疗前后以及不同治疗反应患者之间的外周血免疫细胞图谱，旨在探索MG致病过程中的关键细胞亚群及生物标志物。

研究方法：本研究纳入了7例未经免疫治疗的乙酰胆碱受体抗体阳性的全身型MG患者和7例年龄性别匹配的健康对照。通过质谱流式技术，检测MG患者基线及治疗后、健康对照的外周血单个核细胞，分析37个抗体标记的免疫细胞亚群。对所得高通量数据进行了聚类、降维、注释和差异分析。

研究结果：在MG患者中，CD4+T细胞比例显著升高，治疗后显著下降；调节性T细胞（Regulatory T cells，Treg）比例及其IL-10的表达显著升高，治疗后Treg细胞比例显著下降。与治疗后症状控制达标的患者相比，治疗后无改善的患者CD4+T细胞比例显著更高，辅助性T细胞17型（T helper 17 cells，Th17 cells）的比例显著升高，Treg细胞比例显著降低。此外，CD57+CXCR3+CD8+ 效应T细胞在患者中显著升高，并在治疗后显著下降。

结论：本研究通过质谱流式技术构建了MG患者外周血治疗前后的免疫细胞图谱变化。CD4+T细胞及其亚群在MG患者的外周血中的变化呈主导地位，Th17/Treg失衡可能是MG治疗反应不佳的潜在机制，Th17和Treg细胞具有作为预测治疗反应的生物标志物的潜力。CD57+CXCR3+CD8+效应T细胞可能参与MG的致病过程。

Part I: Impact of Immunotherapy Regimens and Withdrawal on Relapse Risk in Myasthenia Gravis

Background and objective: Corticosteroids (steroids) are the first-line immunotherapy for myasthenia gravis, but the optimal steroid dosing regimen and effects of discontinuation remain unclear. This study aimed to investigate the impact of steroid regimens and steroid withdrawal in patients on steroid monotherapy.

Methods: This cohort study, based on a prospective registry, included patients who achieved minimal manifestations or better status with steroid monotherapy. The primary outcome was relapse. Group-based trajectory modeling identified distinct regimens, and Cox proportional hazards models along with propensity score matching assessed the impact of regimens and steroid withdrawal.

Results: Among 209 patients (median follow-up: 54.0 months), 113 (54.1%) experienced relapse, and 68 (32.5%) discontinued steroids. Three regimens were identified: "High Start, Fast Taper" (Regimen 1), "Moderate Start, Gradual Taper" (Regimen 2), and " Low Start, Slow Taper" (Regimen 3). These regimens aligned with our practice of adjusting doses based on disease severity. Regimen 2 (HR = 0.26, 95% CI 0.14-0.48, p < 0.001) and Regimen 3 (HR = 0.14, 95% CI 0.07-0.31, p < 0.001) had lower relapse risks than Regimen 1. The median time to relapse after steroid withdrawal was 7.0 months. Propensity score matching revealed steroid withdrawal increased 1-year relapse risk versus low-dose maintenance (HR = 1.61, p = 0.031).

Conclusion: Patients with higher disease severity needing high-dose steroids have a higher relapse risk. Steroid withdrawal significantly increases relapse risk compared to low-dose maintenance, necessitating caution when discontinuing steroids in patients on monotherapy.

Part II: Utilizing Mass Cytometry to Profile Peripheral Blood Immune Cells and Investigate Clinical Biomarkers in Myasthenia Gravis Patients

Background and objective: Myasthenia gravis (MG) is a T-cell dependent, B-cell mediated autoimmune disease characterized by the production of autoantibodies targeting the neuromuscular junction. However, comprehensive analysis of the peripheral blood immune cell profiles in MG patients is still lacking, as well as biomarkers that reflect disease severity and predict treatment response. This study employed mass cytometry (CyTOF) to construct and compare the peripheral blood immune cell profiles of MG patients with healthy controls, before and after treatment, and across different treatment responses, aiming to identify key cell subsets and biomarkers involved in the pathogenesis of MG.

Methods: This study included 7 acetylcholine receptor antibody-positive generalized MG patients who had not undergone immunotherapy, and 7 age- and sex-matched healthy controls. Peripheral blood mononuclear cells from MG patients at baseline and post-treatment, as well as from healthy controls, were analyzed using CyTOF with 37 antibody markers to examine immune cell subsets. High-throughput data were subjected to clustering, dimensionality reduction, annotation, and differential analysis.

Results: In MG patients, the proportion of CD4+ T cells was significantly increased, and decreased significantly after treatment. The proportion of regulatory T cells (Treg) and the expression of IL-10 in Treg cells were significantly higher, with a notable decrease in Treg cell proportion after treatment. Compared to patients with symptom control after treatment, those without improvement had significantly higher CD4+ T cell proportions, with significantly increased proportions of T helper 17 (Th17) cells and significantly decreased Treg cells. Additionally, CD57+CXCR3+CD8+ effector T cells were significantly elevated in patients and decreased significantly after treatment.

Conclusion: This study constructed a peripheral blood immune cell profile in MG patients before and after treatment using CyTOF technology. Changes in CD4+ T cells and their subsets dominate in the peripheral blood of MG patients, with Th17/Treg imbalance potentially serving as a mechanism for poor treatment response. Th17 and Treg cells hold promise as biomarkers for predicting treatment response. CD57+CXCR3+CD8+ effector T cells may play a role in the pathogenesis of MG.