Data Sheet 1_Identification of cellular factors associated with inflammation and neurodegeneration in multiple sclerosis.docx

BackgroundSerum biomarkers as neurofilament light chain (sNfL) and glial fibrillary acidic protein (sGFAP) enabled early identification of multiple sclerosis (MS) patients at risk of relapse-associated worsening (RAW) or progression independent of relapses (PIRA). However, the immunological mechanisms underlying these clinical phenotypes remain unclear.
MethodsWe conducted a cross-sectional study including 117 MS patients and 84 healthy controls (HC). Patients were stratified as NLGL (low sNfL and sGFAP), NH (high sNfL at different levels of sGFAP), and NLGH (low sNfL and high sGFAP). Percentages of blood and cerebrospinal fluid (CSF) mononuclear cells, and intracellular production of cytokines by T and B cells after “in vitro” stimulation were analyzed by flow cytometry.
ResultsWe identified a common inflammatory profile present in the blood of all MS groups comprising significant increases of effector CD4+ and CD8+ T cells, of memory and antigen-presenting B cells, of CD4+ and CD8+ T cells producing interferon-gamma, interleukin-17 and tumor necrosis factor-alpha (TNF-α) and of B cells producing TNF-α. Additionally, the highly inflammatory NH group showed lower frequencies of different regulatory subsets (transitional B cells, PDL1+ monocytes and Treg cells) compared to HC and increased percentages of CD4+ and CD8+ T cells producing granulocyte-macrophage colony-stimulating factor and of effector CD56dim NK cells. They also showed lower percentages of Treg in blood and CSF compared to the low inflammatory NLGL group, which also displayed higher frequencies of regulatory CD56dim, NKG2A+ cells.
ConclusionAll MS patients share increased inflammatory B and T cells, but differ in regulatory or NK subsets, which identify highly inflammatory or benign disease courses.

背景 血清生物标志物如血清神经丝轻链（serum neurofilament light chain, sNfL）与血清胶质纤维酸性蛋白（serum glial fibrillary acidic protein, sGFAP）可早期识别存在复发相关恶化（relapse-associated worsening, RAW）风险或非复发依赖性进展（progression independent of relapses, PIRA）的多发性硬化（multiple sclerosis, MS）患者。然而，这些临床表型背后的免疫学机制仍未阐明。
方法 本研究纳入117例MS患者与84例健康对照（healthy controls, HC）开展横断面研究。按照血清sNfL与sGFAP水平将患者分为三组：NLGL组（sNfL与sGFAP均低表达）、NH组（不同sGFAP水平下sNfL高表达）以及NLGH组（sNfL低表达、sGFAP高表达）。采用流式细胞术分析血液与脑脊液（cerebrospinal fluid, CSF）单个核细胞的占比，以及T、B细胞经体外刺激后胞内细胞因子的产生情况。
结果 本研究发现所有MS患者的血液中均存在共同的炎症表型：效应性CD4+、CD8+T细胞，记忆B细胞与抗原呈递B细胞，分泌干扰素-γ、白细胞介素-17与肿瘤坏死因子-α（TNF-α）的CD4+、CD8+T细胞，以及分泌TNF-α的B细胞比例均显著升高。此外，高炎症表型的NH组与健康对照相比，多种调节性细胞亚群（过渡性B细胞、PDL1+单核细胞与调节性T细胞）的频率更低，而分泌粒细胞-巨噬细胞集落刺激因子的CD4+、CD8+T细胞以及效应性CD56dim NK细胞的比例更高。与低炎症的NLGL组相比，NH组血液与脑脊液中的调节性T细胞比例更低；而NLGL组的调节性CD56dim、NKG2A+细胞频率则更高。
结论 所有MS患者均存在炎症性B、T细胞增多的共性，但在调节性细胞或NK细胞亚群上存在差异，这些差异可用于区分高炎症性或良性疾病病程。