Table_1_Persistent Overactive Cytotoxic Immune Response in a Spanish Cohort of Individuals With Long-COVID: Identification of Diagnostic Biomarkers.docx

Long-COVID is a new emerging syndrome worldwide that is characterized by the persistence of unresolved signs and symptoms of COVID-19 more than 4 weeks after the infection and even after more than 12 weeks. The underlying mechanisms for Long-COVID are still undefined, but a sustained inflammatory response caused by the persistence of SARS-CoV-2 in organ and tissue sanctuaries or resemblance with an autoimmune disease are within the most considered hypotheses. In this study, we analyzed the usefulness of several demographic, clinical, and immunological parameters as diagnostic biomarkers of Long-COVID in one cohort of Spanish individuals who presented signs and symptoms of this syndrome after 49 weeks post-infection, in comparison with individuals who recovered completely in the first 12 weeks after the infection. We determined that individuals with Long-COVID showed significantly increased levels of functional memory cells with high antiviral cytotoxic activity such as CD8+ TEMRA cells, CD8±TCRγδ+ cells, and NK cells with CD56+CD57+NKG2C+ phenotype. The persistence of these long-lasting cytotoxic populations was supported by enhanced levels of CD4+ Tregs and the expression of the exhaustion marker PD-1 on the surface of CD3+ T lymphocytes. With the use of these immune parameters and significant clinical features such as lethargy, pleuritic chest pain, and dermatological injuries, as well as demographic factors such as female gender and O+ blood type, a Random Forest algorithm predicted the assignment of the participants in the Long-COVID group with 100% accuracy. The definition of the most accurate diagnostic biomarkers could be helpful to detect the development of Long-COVID and to improve the clinical management of these patients.

长新冠（Long-COVID）是一种在全球范围内新近出现的综合征，其特征为感染新型冠状病毒（SARS-CoV-2）4周以上甚至12周后，仍持续存在未缓解的新冠相关体征与症状。目前长新冠的潜在发病机制尚未明确，当前主流假说主要包括两类：一是SARS-CoV-2在器官及组织庇护所持续存在引发持续性炎症反应；二是该综合征与自身免疫性疾病存在相似性。本研究针对西班牙人群队列展开分析，该队列中部分受试者在感染后49周出现长新冠相关体征与症状，同时设置了感染后12周内完全康复的对照组，旨在探究多项人口统计学、临床及免疫学参数作为长新冠诊断生物标志物的应用价值。研究结果显示，长新冠患者体内具备高抗病毒细胞毒活性的功能性记忆细胞水平显著升高，此类细胞包括CD8+ TEMRA细胞、CD8±TCRγδ+细胞，以及表型为CD56+CD57+NKG2C+的自然杀伤细胞（NK）。这类持久存在的细胞毒性细胞群的持续性，可通过CD4+调节性T细胞（Tregs）水平升高以及CD3+ T淋巴细胞表面耗竭标志物程序性死亡受体1（PD-1）的表达增强得到佐证。结合上述免疫学参数、嗜睡、胸膜炎性胸痛、皮肤损伤等典型临床特征，以及女性性别、O型血等人口统计学因素，随机森林（Random Forest）算法可实现100%准确的受试者分组预测，将受试者归类至长新冠组。精准鉴定核心诊断生物标志物，将有助于早期发现长新冠的发生，并优化此类患者的临床管理方案。