Immunoglobulin M-based local production in skin-associated lymphoid tissue of flounder (Paralichthys olivaceus) initiated by immersion with inactivated Edwardsiella tarda

IF 4.1 2区农林科学 Q1 FISHERIES

Fish & shellfish immunology Pub Date : 2024-10-24 DOI:10.1016/j.fsi.2024.109982

Yuan Guo , Xiuzhen Sheng , Xiaoqian Tang , Jing Xing , Heng Chi , Wenbin Zhan

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Abstract

Fish skin, the mucosal site most exposed to external antigens, requires protection by an efficient local mucosal immune system. The mucosal reserve of IgM is recognized as an immune strategy that blocks pathogen invasion to maintain homeostasis, whereas the mechanism of skin-associated local IgM production induced by mucosal antigens is not well know. In this study, we found that the skin of flounder (Paralichthys olivaceus) was equipped with the immune cellular and molecular basis for processing mucosal antigens and triggering local specific responses, i.e., CD4⁺ Zap-70⁺ T cells, CD4⁻ Zap-70⁺ T/NK cells, IgM⁺ MHCII⁺ B cells, PNA⁺ MHCII⁺ antigen-presenting cells, UEA-1⁺ WGA⁺ and UEA-1⁺ WGA⁻ antigen-sampling cells, as well as secreted IgM and pIgR, as demonstrated by indirect immunofluorescence assay using different antibodies and lectins. After immersion immunization with inactivated Edwardsiella tarda, qPCR assay displayed up-regulation of immune-related genes in flounder skin. Flow cytometry analysis and EdU labeling demonstrated that the mucosal inactivated vaccine induced local proliferation and increased amounts of cutaneous IgM⁺ B cells. Skin explant culture proved the local production of specific IgM in the skin, which could bind to the surface of E. tarda. ELISA, laser scanning confocal microscopy, and Western blot revealed that, in addition to the elevated IgM levels, pIgR protein level was significantly up-regulated in skin tissue and surface mucus containing the pIgR (secretory component, SC)-tetrameric IgM complex, indicating that mucosal vaccine stimulated up-regulation of IgM and pIgR, which were secreted as a complex into skin mucus to exert the protective effects as secretory IgM. These findings deepen the understanding of IgM-based local responses in the mucosal immunity of teleosts, which will be critical for subsequent investigation into the protective mechanism of mucosal vaccines for fish health.

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用灭活的塔氏爱德华氏菌浸泡比目鱼（Paralichthys olivaceus）皮肤相关淋巴组织，可在局部产生以免疫球蛋白 M 为基础的抗体。

鱼皮是暴露于外部抗原最多的粘膜部位，需要高效的局部粘膜免疫系统的保护。粘膜储备的 IgM 被认为是一种阻止病原体入侵以维持体内平衡的免疫策略，而粘膜抗原诱导皮肤相关的局部 IgM 产生的机制尚不清楚。在这项研究中，我们发现比目鱼（Paralichthys olivaceus）的皮肤具备处理粘膜抗原并引发局部特异性反应的免疫细胞和分子基础，即通过使用不同抗体和凝集素进行间接免疫荧光检测，可发现CD4+ Zap-70+ T细胞、CD4- Zap-70+ T/NK细胞、IgM+ MHCII+B细胞、PNA+ MHCII+抗原递呈细胞、UEA-1+ WGA+和UEA-1+ WGA-抗原取样细胞，以及分泌的IgM和pIgR。用灭活的塔氏爱德华氏菌浸泡免疫后，qPCR 检测显示比目鱼皮肤中免疫相关基因上调。流式细胞术分析和 EdU 标记表明，粘膜灭活疫苗诱导了局部增殖和皮肤 IgM+ B 细胞数量的增加。皮肤外植体培养证明皮肤局部产生了特异性 IgM，可与 E. tarda 表面结合。ELISA、激光扫描共聚焦显微镜和Western印迹显示，除了IgM水平升高外，皮肤组织和含有pIgR（分泌成分，SC）-四聚体IgM复合物的表面粘液中pIgR蛋白水平也显著上调，这表明粘膜疫苗刺激了IgM和pIgR的上调，它们以复合物的形式分泌到皮肤粘液中，作为分泌型IgM发挥保护作用。这些发现加深了人们对远洋鱼类粘膜免疫中基于 IgM 的局部反应的理解，这对后续研究粘膜疫苗对鱼类健康的保护机制至关重要。

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来源期刊

Fish & shellfish immunology 农林科学-海洋与淡水生物学

CiteScore

7.50

自引率

19.10%

发文量

750

审稿时长

68 days

期刊介绍： Fish and Shellfish Immunology rapidly publishes high-quality, peer-refereed contributions in the expanding fields of fish and shellfish immunology. It presents studies on the basic mechanisms of both the specific and non-specific defense systems, the cells, tissues, and humoral factors involved, their dependence on environmental and intrinsic factors, response to pathogens, response to vaccination, and applied studies on the development of specific vaccines for use in the aquaculture industry.