Abstract

Inhalation of crystalline silica (cSiO₂) in the workplace is etiologically linked to lupus and other autoimmune diseases. Exposing lupus-prone NZBWF1 mice to respirable cSiO₂ unleashes a vicious cycle of inflammation and cell death in the lung that triggers interferon-regulated gene expression, ectopic lymphoid structure (ELS) development, elevation of local and systemic autoantibodies (AAbs), and glomerulonephritis. However, cSiO₂-induced inflammation and onset of autoimmunity can be prevented by inclusion of the ω-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) into the diet of these mice. Since cSiO₂ both causes cell death and interferes with efferocytosis, secondary necrosis of residual cell corpses might provide a rich and varied autoantigen (AAg) source in the lung. While it is known that the particle induces anti-nuclear and anti-dsDNA AAbs in NZBWF1 mice, the full extent of the cSiO₂-induced AAb response relative to specificity and isotype is not yet understood. The purpose of this study was to test the hypotheses that cSiO₂ exposure induces a wide spectrum of AAbs in the pulmonary and systemic compartments, and that dietary DHA intervention prevents these changes. Archived tissue fluid samples were obtained from a prior study in which NZBWF1 mice were fed purified isocaloric diets containing no DHA (control) or DHA corresponding calorically to human doses of 2 and 5 g/day. Mice were intranasally instilled with 1 mg cSiO₂ or saline vehicle weekly for 4 weeks, then groups euthanized 1, 5, 9, or 13 weeks post-instillation (PI) of the last cSiO₂ dose. Bronchoalveolar lavage fluid (BALF) and plasma from each time point were subjected to AAb profiling using a microarray containing 122 AAgs. cSiO₂ triggered robust IgG and IgM AAb responses against lupus-associated AAgs, including DNA, histones, ribonucleoprotein, Smith antigen, Ro/SSA, La/SSB, and complement as early as 1 week PI in BALF and 5 weeks PI in plasma, peaking at 9 and 13 weeks PI, respectively. Importantly, cSiO₂ also induced AAbs to AAgs associated with rheumatoid arthritis (collagen II, fibrinogen IV, fibrinogen S, fibronectin, and vimentin), Sjögren’s syndrome (α-fodrin), systemic sclerosis (topoisomerase I), vasculitis (MPO and PR3), myositis (Mi-2, TIF1-γ, MDA5), autoimmune hepatitis (LC-1), and celiac disease (TTG). cSiO₂ elicited comparable but more modest IgA AAb responses in BALF and plasma. cSiO₂-induced AAb production was strongly associated with time dependent inflammatory/autoimmune gene expression, ELS development, and glomerulonephritis. AAb responses were dose-dependently suppressed by DHA supplementation and negatively correlated with the ω-3 index, an erythrocyte biomarker of ω-3 content in tissue phospholipids. Taken together, these findings suggest that cSiO₂ exposure elicits a diverse multi-isotype repertoire of AAbs, many of which have been reported in individuals with lupus and other autoimmune diseases. Furthermore, induction of this broad AAb spectrum could be impeded by increasing ω-3 tissue content via dietary DHA supplementation.

摘要

在工作场所吸入结晶二氧化硅 (cSiO ₂ ) 在病因学上与狼疮和其他自身免疫性疾病有关。将易患狼疮的 NZBWF1 小鼠暴露于可吸入的 cSiO ₂会引发肺部炎症和细胞死亡的恶性循环，从而引发干扰素调节的基因表达、异位淋巴结构 (ELS) 发育、局部和全身自身抗体 (AAbs) 升高以及肾小球肾炎. 然而，通过在这些小鼠的饮食中加入 ω-3 多不饱和脂肪酸二十二碳六烯酸 (DHA)，可以防止cSiO _{2诱导的炎症和自身免疫的发作。}由于 cSiO ₂两者都会导致细胞死亡并干扰细胞增多症，残留细胞尸体的继发性坏死可能会在肺中提供丰富多样的自身抗原 (AAg) 来源。虽然已知颗粒在 NZBWF1 小鼠中诱导抗核和抗 dsDNA AAbs，但 cSiO ₂诱导的 AAb 反应相对于特异性和同种型的全部程度尚不清楚。_{本研究的目的是检验 cSiO 2}的假设暴露会在肺和全身隔室中诱导广泛的 AAb，而饮食 DHA 干预可以防止这些变化。存档的组织液样本来自先前的研究，其中 NZBWF1 小鼠被喂食不含 DHA（对照）或 DHA 的纯化等热量饮食，其热量对应于 2 和 5 克/天的人类剂量。每周向小鼠鼻内滴注 1 mg cSiO ₂或盐水载体，持续 4 周，然后各组在最后一剂 cSiO ₂滴注后 (PI) 1、5、9 或 13 周实施安乐死。使用含有 122 个 AAg 的微阵列对每个时间点的支气管肺泡灌洗液 (BALF) 和血浆进行 AAb 分析。二氧化硅_{_}早在 BALF 中 PI 1 周和血浆 PI 5 周时，就触发了针对狼疮相关 AAg 的强烈 IgG 和 IgM AAb 反应，包括 DNA、组蛋白、核糖核蛋白、Smith 抗原、Ro/SSA、La/SSB 和补体，峰值在分别在 PI 9 周和 13 周。重要的是，cSiO ₂还诱导与类风湿性关节炎（胶原蛋白 II、纤维蛋白原 IV、纤维蛋白原 S、纤连蛋白和波形蛋白）、干燥综合征（α-fodrin）、系统性硬化症（拓扑异构酶 I）、血管炎（MPO 和 PR3）相关的 AAgs 的 Aabs 、肌炎（Mi-2、TIF1-γ、MDA5）、自身免疫性肝炎（LC-1）和乳糜泻（TTG）。cSiO ₂在 BALF 和血浆中引起相当但更温和的 IgA AAb 反应。二氧化硅_{_}诱导的 AAb 产生与时间依赖性炎症/自身免疫基因表达、ELS 发展和肾小球肾炎密切相关。AAb 反应受到 DHA 补充的剂量依赖性抑制，并且与 ω-3 指数负相关，ω-3 指数是组织磷脂中 ω-3 含量的红细胞生物标志物。总之，这些发现表明 cSiO ₂暴露会引发多种多同种型 AAb，其中许多已在狼疮和其他自身免疫性疾病患者中得到报道。此外，通过膳食 DHA 补充剂增加 ω-3 组织含量可能会阻碍这种广泛的 AAb 谱的诱导。