当前位置: X-MOL 学术Allergy › 论文详情
Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)
Tissue eosinophils express the IL-33 receptor ST2 and type 2 cytokines in patients with eosinophilic esophagitis
Allergy ( IF 12.6 ) Pub Date : 2021-10-06 , DOI: 10.1111/all.15127
Amiko M Uchida 1, 2, 3 , Patrick J Lenehan 2, 3 , Praveen Vimalathas 1 , Kaia C Miller 1 , Mabel Valencia-Yang 1 , Li Qiang 3, 4 , Lauren A Canha 1 , Lestat R Ali 2, 3 , Michael Dougan 1, 2, 3, 4 , John J Garber 1, 2 , Stephanie K Dougan 3, 4
Affiliation  

Eosinophilic esophagitis (EoE) is a type 2 inflammatory disease of the esophagus largely driven by food antigens and is characterized by esophageal eosinophilia, fibrosis, and clinically by dysphagia and food impactions 1. The alarmin interleukin (IL)-33 is elevated in the esophageal epithelium and endothelium of patients with EoE 2, 3. Exogenous IL-33 causes esophageal eosinophilia and basal hyperplasia in mice 3, implicating IL-33 in EoE pathogenesis. The IL-33 receptor, suppression of tumorigenicity 2 (IL1RL1/ST2), induces type 2 cytokines IL-4, IL-5, and IL-13. As blockade of IL-4/IL-13 improves both histologic and clinical EoE 4, and trials of anti-IL-5R are ongoing, we aimed to identify cells that can respond to IL-33 and the cellular sources of pathogenic Type 2 cytokines in EoE. T-helper 2 (Th2) CD4 lymphocytes are known producers of IL-4, IL-5, and IL-13, as are group 2 innate lymphoid cells (ILC2), but whether tissue eosinophils, mast cells, or basophils contribute type 2 cytokines in active EoE is under explored 5, 6.

We examined biopsies and blood samples from patients undergoing esophagogastroduodenoscopy at our institution (Table S1). We found eosinophils, mast cells, basophils, and Th2 cells; however, ILC2s were not reliably detected in esophageal tissue and were withheld from further analyses (Figure 1A-B). ST2 was robustly detected on tissue eosinophils from patients with active EoE (Figure 1C, 1E), whereas blood eosinophils from healthy control, remission, and active EoE were uniformly low for ST2 (Figure 1D, 1F-G; Figure S1). Similarly, serum levels of soluble ST2 (sST2), which is shed as a decoy receptor, were not significantly different among healthy controls, patients with EoE in remission or active disease (Figure 1H, 1I).

Details are in the caption following the image
FIGURE 1
Open in figure viewerPowerPoint
Tissue eosinophils express ST2. (A) Flow cytometry of esophageal biopsies. (B) Cell type as percent of CD45+ leukocytes in patients with active (n=8–9) and remission (n=4–7) EoE. Flow cytometry showing ST2 expression across esophageal immune cells (C) and blood eosinophils (D). Quantification of ST2 in tissue (E) and blood eosinophils (F). (G) ST2 on blood versus tissue eosinophils. (H) Concentration of sST2 in healthy non-EoE, remission, and active EoE patients shown overall (H) and by sex (I). N=34 EoE (16 active, 16 remission), n=34 controls [Color figure can be viewed at wileyonlinelibrary.com]

We next measured IL-4, IL-5, and IL-13 by flow cytometry and immunofluorescence from esophageal tissue in patients with active EoE (Figure 2A-H). Eosinophils (which unlike T cells were not stimulated with PMA/ionomycin) were frequent expressors of these cytokines (Figure 2A-E). IL-4 was detected in most unstimulated eosinophils (60%) compared to mast cells (8.0%) and basophils 3.8% (Figure 2C). We found IL-13 in 74% of eosinophils compared to 17% of stimulated CRTH2+ Th2 cells, 3.5% of mast cells, and 3.4% of basophils (Figure 2D-F). Although the proportion of unstimulated mast cells positive for type 2 cytokines was relatively low, mast cells may be pathogenic in EoE and warrant further investigation. Tissue eosinophil expression of type 2 cytokine protein was further verified by immunofluorescent staining for IL-4, IL-13, and eosinophil peroxidase (EPX). A majority of the IL-4+ cells in the esophageal epithelium were eosinophils (80.7%); this was similar for IL-13+ cells (82.7%) (Figure 2G-H; Figure S2). To assess whether eosinophils were necessary for type 2 cytokine induction, we administered IL-33 to eosinophil-sufficient (BALB/c WT) and deficient (BALB/c ΔdblGATA-1) mice using a previously established model of EoE 3 (Figure 2I). Immunofluorescence revealed eosinophil infiltration in IL-33-treated WT esophagi but not PBS-treated WT or ΔdblGATA-1 mice, as expected (Figure 2J). Although IL-13 was below the limit of detection in this model, multiplex cytokine analysis of esophagi showed IL-33-induced IL-4 was critically dependent on the presence of eosinophils (Figure 2K).

Details are in the caption following the image
FIGURE 2
Open in figure viewerPowerPoint
Human esophageal eosinophils express type 2 cytokines IL-4, IL-5, and IL-13 by flow cytometry and immunofluorescence. (A) Flow cytometry of intracellular cytokine IL-4, IL-5, and IL-13 on esophageal immune cells, quantification shown in (B) with IL-5 and IL-13 co-expression, (C) IL-4, (D) IL-13, and (E) IL-13 MFI. T-cell cytokines shown in (F). (G-H) Immunofluorescence of human esophageal tissue identifies eosinophils by eosinophil peroxidase (EPX), IL-4- and IL-13-expressing cells with quantification shown to the right. (I) IL-33 EoE mouse model; n=5/group. (J) Immunofluorescence of mouse esophagi. (K) Esophageal cytokine concentrations for IL-4, IL-5, and IL-13 from various treatment groups [Color figure can be viewed at wileyonlinelibrary.com]

Here, we report frequent expression of the IL-33 receptor ST2 on esophageal-infiltrating eosinophils compared to blood eosinophils, other granulocytes, and Th2 cells. Noting eosinophils lack antigen-specific T-cell receptors, their ability to respond to IL-33 may explain continued inflammation observed in patients after removal of dietary antigen. Future studies are warranted to identify factors influencing ST2 and IL-33 expression and determine whether modulation of this pathway may be a novel therapeutic approach to this increasingly diagnosed yet poorly understood allergic disease.



中文翻译:

嗜酸性食管炎患者的组织嗜酸性粒细胞表达 IL-33 受体 ST2 和 2 型细胞因子

嗜酸性食管炎 (EoE) 是一种主要由食物抗原驱动的 2 型食管炎性疾病,其特点是食管嗜酸性粒细胞增多、纤维化,临床上表现为吞咽困难和食物嵌塞1。EoE 2, 3患者的食管上皮和内皮中的警报素白细胞介素 (IL)-33 升高。外源性 IL-33 导致小鼠食管嗜酸性粒细胞增多和基底细胞增生3,表明 IL-33 参与 EoE 发病机制。IL-33 受体抑制致瘤性 2 (IL1RL1/ST2),诱导 2 型细胞因子 IL-4、IL-5 和 IL-13。由于 IL-4/IL-13 的阻断改善了组织学和临床 EoE 4, 并且抗 IL-5R 的试验正在进行中,我们旨在确定可以对 IL-33 和 EoE 中致病性 2 型细胞因子的细胞来源有反应的细胞。T 辅助 2 (Th2) CD4 淋巴细胞是已知的 IL-4、IL-5 和 IL-13 的产生者,第 2 组先天性淋巴细胞 (ILC2) 也是如此,但组织嗜酸性粒细胞、肥大细胞或嗜碱性粒细胞是否贡献 2 型正在探索活性 EoE 中的细胞因子5, 6

我们检查了在我们机构接受食管胃十二指肠镜检查的患者的活检和血液样本(表 S1)。我们发现了嗜酸性粒细胞、肥大细胞、嗜碱性粒细胞和Th2细胞;然而,在食管组织中未能可靠地检测到 ILC2,因此未进行进一步分析(图 1A-B)。ST2 在来自活动性 EoE 患者的组织嗜酸性粒细胞中稳健地检测到(图 1C,1E),而来自健康对照、缓解和活动性 EoE 的血液嗜酸性粒细胞在 ST2 中均较低(图 1D,1F-G;图 S1)。同样,作为诱饵受体脱落的可溶性 ST2 (sST2) 的血清水平在健康对照组、缓解期或活动性疾病的 EoE 患者中没有显着差异(图 1H、1I)。

详细信息在图片后面的标题中
图1
在图形查看器中打开微软幻灯片软件
组织嗜酸性粒细胞表达 ST2。(A) 食管活检的流式细胞术。(B)活动性 (n=8-9) 和缓解 (n=4-7) EoE 患者的细胞类型占 CD45 +白细胞的百分比。流式细胞术显示 ST2 在食管免疫细胞 (C) 和血液嗜酸性粒细胞 (D) 中的表达。组织 (E) 和血液嗜酸性粒细胞 (F) 中 ST2 的定量。(G) 血液与组织嗜酸性粒细胞的 ST2。(H) 总体 (H) 和按性别 (I) 显示的健康非 EoE、缓解和活动性 EoE 患者中 sST2 的浓度。N=34 EoE(16 个活动,16 个缓解),n=34 个对照 [可在 wileyonlinelibrary.com 上查看彩色图]

接下来,我们通过流式细胞术和来自活动性 EoE 患者食管组织的免疫荧光测量了 IL-4、IL-5 和 IL-13(图 2A-H)。嗜酸性粒细胞(与 T 细胞不同,没有用 PMA/离子霉素刺激)是这些细胞因子的常见表达者(图 2A-E)。与肥大细胞 (8.0%) 和嗜碱性粒细胞 3.8% (图 2C) 相比, 在大多数未刺激的嗜酸性粒细胞 (60%) 中检测到 IL-4。我们在 74% 的嗜酸性粒细胞中发现了 IL-13,而在受刺激的 CRTH2+ Th2 细胞中为 17%,在肥大细胞中为 3.5%,在嗜碱性粒细胞中为 3.4%(图 2D-F)。尽管未刺激的 2 型细胞因子阳性肥大细胞的比例相对较低,但肥大细胞可能在 EoE 中具有致病性,值得进一步研究。2 型细胞因子蛋白的组织嗜酸性粒细胞表达通过对 IL-4、IL-13、和嗜酸性粒细胞过氧化物酶 (EPX)。大多数 IL-4+食管上皮细胞为嗜酸性粒细胞(80.7%);这与 IL-13 +细胞 (82.7%) 相似(图 2G-H;图 S2)。为了评估 2 型细胞因子诱导是否需要嗜酸性粒细胞,我们使用先前建立的 EoE 3模型向嗜酸性粒细胞充足 (BALB/c WT) 和缺乏 (BALB/c ΔdblGATA-1) 小鼠施用 IL-33 (图 2I) . 正如预期的那样,免疫荧光显示 IL-33 处理的 WT 食管中的嗜酸性粒细胞浸润,而不是 PBS 处理的 WT 或 ΔdblGATA-1 小鼠(图 2J)。尽管 IL-13 在该模型中低于检测限,但食管的多重细胞因子分析显示 IL-33 诱导的 IL-4 严重依赖于嗜酸性粒细胞的存在(图 2K)。

详细信息在图片后面的标题中
图 2
在图形查看器中打开微软幻灯片软件
人食管嗜酸性粒细胞通过流式细胞术和免疫荧光表达 2 型细胞因子 IL-4、IL-5 和 IL-13。(A) 食管免疫细胞上细胞内细胞因子 IL-4、IL-5 和 IL-13 的流式细胞术,在 (B) 中显示的定量,其中 IL-5 和 IL-13 共表达,(C) IL-4, (D) IL-13 和 (E) IL-13 MFI。(F) 中显示的 T 细胞细胞因子。(GH) 人类食管组织的免疫荧光通过嗜酸性粒细胞过氧化物酶 (EPX)、IL-4 和 IL-13 表达细胞识别嗜酸性粒细胞,其量化结果如右图所示。(I) IL-33 EoE 小鼠模型;n=5/组。(J) 小鼠食管的免疫荧光。(K) 来自不同治疗组的 IL-4、IL-5 和 IL-13 的食管细胞因子浓度 [可在 wileyonlinelibrary.com 上查看彩色图]

在这里,我们报告了与血液嗜酸性粒细胞、其他粒细胞和 Th2 细胞相比,IL-33 受体 ST2 在食管浸润性嗜酸性粒细胞上的频繁表达。注意到嗜酸性粒细胞缺乏抗原特异性 T 细胞受体,它们对 IL-33 的反应能力可以解释在去除饮食抗原后在患者中观察到的持续炎症。未来的研究有必要确定影响 ST2 和 IL-33 表达的因素,并确定该途径的调节是否可能是一种新的治疗方法,用于治疗这种越来越多地被诊断出但知之甚少的过敏性疾病。

更新日期:2021-10-06
down
wechat
bug