ABSTRACT

Whereas central nervous system (CNS) homeostasis is highly dependent on tissue surveillance by immune cells, dysregulated entry of leukocytes during autoimmune neuroinflammation causes severe immunopathology and neurological deficits. To invade the CNS parenchyma, encephalitogenic T helper (T_H) cells must encounter their cognate antigen(s) presented by local major histocompatibility complex (MHC) class II-expressing antigen-presenting cells (APCs). The precise mechanisms by which CNS-associated APCs facilitate autoimmune T cell reactivation remain largely unknown. We previously showed that mice with conditional deletion of the gene encoding the essential autophagy protein ATG5 in dendritic cells (DCs) are resistant to EAE development. Here, we report that the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2, also known as CYBB/NOX2, in conventional DCs (cDCs) regulates endocytosed MOG (myelin oligodendrocyte protein) antigen processing and supports MOG-antigen presentation to CD4⁺ T cells through LC3-associated phagocytosis (LAP). Genetic ablation of Cybb in cDCs is sufficient to restrain encephalitogenic T_H cell recruitment into the CNS and to ameliorate clinical disease development upon the adoptive transfer of MOG-specific CD4⁺ T cells. These data indicate that CYBB-regulated MOG-antigen processing and LAP in cDCs licenses encephalitogenic T_H cells to initiate and sustain autoimmune neuroinflammation.

Abbreviations: Ag: antigen; APC: antigen-presenting cell; AT: adoptive transfer; ATG/Atg: autophagy-related; BAMs: border-associated macrophages; BMDC: bone marrow-derived DC; CD: cluster of differentiation; CNS: central nervous system; CSF2/GM-CSF: colony stimulating factor 2 (granulocyte-macrophage); CYBB/NOX2/gp91^phox: cytochrome b-245, beta polypeptide; DC: dendritic cell; EAE: experimental autoimmune encephalomyelitis; fl: floxed; FOXP3: forkhead box P3; GFP: green fluorescent protein; H2-Ab: histocompatibility 2, class II antigen A, beta 1; IFN: interferon; IL: interleukin; ITGAX/CD11c: integrin subunit alpha X; LAP: LC3-associated phagocytosis; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MFI: median fluorescence intensity; MG: microglia; MHCII: major histocompatibility complex class II; MOG: myelin oligodendrocyte glycoprotein; MS: multiple sclerosis; NADPH: nicotinamide adenine dinucleotide phosphate; ODC: oligodendroglial cell; OVA: ovalbumin; pDC: plasmacytoid DC; Ptd-L-Ser: phosphatidylserine; PTPRC: protein tyrosine phosphatase, receptor type, C; ROS: reactive oxygen species; SLE: systemic lupus erythematosus; T_H cells: T helper cells; TLR: toll-like receptor; ZBTB46: zinc finger and BTB domain containing 46

摘要

虽然中枢神经系统 (CNS) 稳态高度依赖于免疫细胞的组织监测，但自身免疫性神经炎症期间白细胞的失调进入会导致严重的免疫病理学和神经功能缺陷。侵入中枢神经系统实质，致脑炎T辅助（T _H) 细胞必须遇到由表达局部主要组织相容性复合物 (MHC) II 类抗原呈递细胞 (APC) 呈递的同源抗原。中枢神经系统相关的 APC 促进自身免疫 T 细胞再激活的确切机制仍然很大程度上未知。我们之前表明，在树突状细胞 (DC) 中条件性缺失编码必需自噬蛋白 ATG5 的基因的小鼠对 EAE 发展具有抗性。在这里，我们报告了烟酰胺腺嘌呤二核苷酸磷酸 (NADPH) 氧化酶 2，也称为 CYBB/NOX2，在常规 DC (cDC) 中调节内吞 MOG（髓鞘少突胶质细胞蛋白）抗原加工并支持 MOG 抗原呈递给 CD4 ⁺ T 细胞通过 LC3 相关吞噬作用 (LAP)。Cybb的基因消融在 cDCs 中足以抑制脑炎性 T _H细胞募集到 CNS 中，并在 MOG 特异性 CD4 ⁺ T 细胞过继转移后改善临床疾病发展。这些数据表明，cDC 中 CYBB 调节的 MOG 抗原加工和 LAP 允许致脑炎 T _H细胞启动和维持自身免疫性神经炎症。

缩写：Ag：抗原；APC：抗原呈递细胞；AT：收养转移；ATG/ Atg：自噬相关；BAMs：边界相关巨噬细胞；BMDC：骨髓来源的 DC；CD：分化簇；CNS：中枢神经系统；CSF2/GM-CSF：集落刺激因子 2（粒细胞-巨噬细胞）；CYBB/NOX2/gp91 ^phox：细胞色素b-245，β多肽；DC：树突状细胞；EAE：实验性自身免疫性脑脊髓炎；fl：floxed；FOXP3：叉头箱P3；GFP：绿色荧光蛋白；H2-Ab：组织相容性 2，II 类抗原 A，β 1；干扰素：干扰素；IL：白细胞介素；ITGAX/CD11c：整合素亚基α X；LAP：LC3相关的吞噬作用；MAP1LC3/LC3：微管相关蛋白1轻链3；MFI：中值荧光强度；MG：小胶质细胞；MHCII：主要组织相容性复合物 II 类；MOG：髓鞘少突胶质细胞糖蛋白；MS：多发性硬化症；NADPH：烟酰胺腺嘌呤二核苷酸磷酸盐；ODC：少突胶质细胞；OVA：卵清蛋白；pDC：浆细胞样 DC；Ptd-L-Ser：磷脂酰丝氨酸；PTPRC：蛋白酪氨酸磷酸酶，受体类型，C；ROS：活性氧；SLE：系统性红斑狼疮；吨_H细胞：T辅助细胞；TLR：toll样受体；ZBTB46：锌指和 BTB 结构域，包含 46