鱼类鼻黏膜相关淋巴组织的研究进展

鱼类黏膜相关淋巴组织是抵御病原入侵的第一道防线。其中,鱼类鼻黏膜相关淋巴组织（Nasal-associated lymphoid tissue,NALT）近年来被国内外学者所关注,并被证明是嗅觉器官中抗原识别和启动黏膜免疫应答的重要场所,在细菌、病毒和寄生虫感染后可发挥快速的局部免疫响应。以NALT为靶点,对鱼类开展疫苗鼻内接种可起到良好的免疫保护效果。但目前,对NALT中复杂的免疫细胞与分子网络及其互作机制知之甚少。本文对鱼类嗅觉器官结构与功能、NALT的细胞与分子网络及免疫应答、鼻内接种的应答及免疫保护效果等方面的最新研究进展进行综述,旨在阐明鱼类NALT在黏膜局部发挥的免疫防御机制,以期为新型黏膜疫苗的设计与研发提供参考。     

硬骨鱼类鳃黏膜免疫相关分子的研究进展

鳃为鱼类重要的呼吸器官，是鱼类进行离子交换、酸碱调节和含氮废物排泄的重要结构基础，也是鱼类重要的外周黏膜免疫器官之一，在抵御病原微生物侵染过程中发挥重要的免疫屏障作用。当前，硬骨鱼类鳃黏膜免疫反应是研究热点之一。本文首先对硬骨鱼类鳃的结构和特点进行分析，之后综述了抗菌肽、干扰素、白细胞介素、Toll样受体、补体等先天性免疫相关分子以及T细胞受体和免疫球蛋白等适应性免疫相关分子在硬骨鱼类鳃黏膜中的表达规律、分子功能，最后探讨了化学因素(重金属、杀虫剂等)、生物因素(细菌、病毒、真菌、和寄生虫等)以及营养物质和疫苗等对硬骨鱼类鳃黏膜结构的影响，以期为深入研究鳃在鱼类黏膜免疫反应中的角色和应答机制提供指导，为硬骨鱼类病原性疾病的免疫防控策略的制定提供理论基础。     

鱼类的免疫与鱼病的防治(I)

１硬骨鱼类免疫系统的组成 免疫是机体识别“自身”与“非己”抗原，对“自身”抗原形成天然免疫耐受，对“非己”抗原产生排异作用的一种生理功能。生物体在识别和清除“非己”抗原过程中所产生的各种生物学作用总称为免疫功能。主宰和执行机体的免疫功能，使机体发生免疫应答的物质基础是生物的免疫系统。 鱼类的免疫功能还没有象人及高等哺乳动物那样复杂发达，但鱼类已经具备了一个相当完善的免疫系统，在抵抗不良环境的影响，保护鱼体健康的过程中发挥着重要的作用。 鱼类的免疫系统由免疫组织和器官，免疫细胞，免疫分子３部分组成。…     

鱼类microRNA在先天免疫中的研究进展

先天免疫是宿主识别病原及消除病原感染的第一道防线。模式识别受体是参与识别病原入侵的主要分子，主要包括Toll样受体、RIG-I样受体、NOD样受体和C型凝集素受体等。模式识别受体在识别病原相关分子模式后，激活机体的先天免疫信号通路，诱导炎症细胞因子和干扰素的产生，从而启动抵抗病原入侵的免疫应答。越来越多的证据表明，免疫应答的激活、维持和终止受到了严格的调节，使机体在保持一定免疫强度的同时避免产生过度的免疫反应。microRNA是一类长度为18～23 nt的微小非编码RNA，是鱼类先天免疫应答网络中的重要调控因子。近年来，microRNA在鱼类免疫学领域已开展了大量的研究，但缺乏对其进行及时地全面性的总结。本文综述了近年来miRNA在鱼类先天免疫反应中的研究进展，以期为鱼类的分子抗病育种及疾病防控研究提供一些思路。     

鱼类的非特异性免疫概述

对于鱼类免疫的研究很早就出现,鱼类是特异性免疫和非特异性免疫并存的脊椎动物。但与哺乳类相比,鱼类的特异免疫系统尚不够发达,特异性免疫机制还不完善。因此,在鱼类对外界刺激及病原生物侵袭的防御反应中,非特异性免疫起着重要作用。现已知参与鱼类非特异性免疫反应的因子主要包括巨噬细胞、粒细胞和细胞毒性细胞等一些具有吞噬作用的细胞,以及介导一系列免疫或应激反应的蛋白分子,如补体、细胞因子、趋化因子及溶菌酶和抗菌肽等。     

军曹鱼黏膜免疫组织发育的形态学研究

运用组织学及组织化学的染色方法,对出膜后第l-44天（days post-hatch,dph）的军曹鱼（Rachycentron canadum）黏膜免疫组织的发育过程进行研究。结果显示,白细胞在黏膜免疫组织出现的时间顺序由早到晚依次为鳃、皮肤、胃、肠,同大量抗原接触鱼体组织时白细胞出现的时间顺序相符。军曹鱼胃固有层中丰富的白细胞说明胃可能在黏膜免疫中发挥着很大的作用。黏液细胞最早出现在胃;其后是皮肤和肠道;鳃最晚。在胃肠、皮肤内的黏液细胞几天后就可分泌中性至酸性黏液物质,且分泌量随鱼体分发育而增多,但在鳃组织内,黏液的分泌早于黏液细胞的出现。在采样的时间段里,黏膜免疫组织未发现浆细胞,可能是由于B细胞还没有发育成熟并具备免疫功能。从白细胞在军曹鱼黏膜组织内发育及分布的状况,可见黏膜组织具有完成免疫应答的细胞基础。     

无脊椎动物适应性免疫的研究进展

无脊椎动物免疫学是一门新兴学科,近十余年来,由于对虾等水产养殖动物受到了前所未有的病害侵袭,无脊椎动物的免疫机制引起国内外学者的高度重视。一般认为,无脊椎动物由于缺乏Ig (immunoglobulin)、TCR (T cell recep-tor)和MHC (major histocompatibility complex)等分子而不具备适应性免疫。但随着研究的深入,人们发现无脊椎动物不仅具有较完备的非特异性免疫系统,而且在一定范围内具备脊椎动物所特有的适应性免疫现象,同时还发现了很多结构或功能上类似于脊椎动物适应性免疫分子的免疫因子。由此,许多研究者认为无脊椎动物体内也应…     

鱼类树突状细胞研究进展

树突状细胞(DCs)是目前已知的体内功能最强的抗原递呈细胞,是唯一能够激活初始T淋巴细胞反应的细胞,在先天性免疫、适应性免疫以及维持自身免疫耐受方面具有重要的作用,因此一直是免疫学研究的重要领域。本文简要综述了DCs的类型及其在动物体内的功能、各类DCs的细胞标记。总结了鱼类DCs的分离、纯化方法和形态学观察方法;现有研究表明,鱼类DCs具有吞噬细菌、刺激T细胞增殖、诱导CD4+T细胞的活化、表达DCs的标记基因、被Toll样受体的配体激活、迁移能力、引起混合淋巴细胞反应等生物学功能;不同鱼类DCs的分子标记并不完全一样;鱼类的头肾、肾、鳃、皮肤、胸腺、脾、肠等均有DCs的分布。目前,对鱼类DCs的研究虽然取得了一定进展,但仍有许多重要问题需要解决:①鱼类DCs目前缺乏明确的细胞标记,加强这方面的研究有助于提高鱼类DCs的分离、体内分布与功能的研究水平;②加强和完善鱼类DCs的分离、培养技术的研究,掌握各种鱼类DCs的分离培养方法;③加强鱼类DCs在抗原递呈中的功能研究,对深入分析鱼类免疫机理,合理设计和应用疫苗,具有重要的理论指导意义。     

疫苗佐剂的研究进展

正佐剂最早来源于拉丁语"adjuvare"一词,为"帮助",是指能够提高机体对抗原的适应性免疫应答的物质,能够在免疫反应中诱导全面持久的免疫应答。在疫苗制剂中,佐剂的功能主要包括:增强疫苗抗原的免疫原性;促进细胞免疫和体液免疫,优化免疫应答,促进免疫能力较弱人群中的免疫应答;增进抗原与黏膜之间的传递以及免疫接触;减少疫苗成分中抗原的     

海洋贝类Toll样受体及信号通路的研究进展

Toll样受体(Toll-like receptor,TLR)是模式识别受体家族(PRR)中重要的一员,它可以特异地识别病原体相关的分子模式(PAMP),激活信号级联、诱发固有免疫反应,在抵御病原微生物过程中发挥重要作用。TLRs最早在果蝇胚胎中发现,是一类进化高度保守的免疫受体家族,在果蝇及脊椎动物中研究得较为透彻。海洋贝类具有较高的经济价值及重要系统进化地位,近年来,大量贝类的Toll样受体及信号通路相关因子被发现。本文综述了Toll样受体的结构、信号通路和海洋贝类TLR及信号通路相关因子的研究进展,有助于丰富海洋贝类固有免疫理论研究。     

鱼类粘膜免疫研究进展

Fish immunology has achieved great progress in recent years. While before 1990s, most researches focused on the fish systematic immunity, and the mucosal immunity of fish had not been given enough attention. Indeed, it has been shown that fish mucosal immunity plays an important role in disease defense. Fish mucosal immunity research has made some exciting progress in this decade. This review will focus on such progress: Constitution of mucosal-associated tissues and distribution of different immune cells, including T/B lymphocytes, granules, monocytes, macrophages, goblet cells, etc, in these sites have been well described with the development of some monoclonal antibody to these cells and associated techniques. Non-specific immune response mechanism of mucosal tissues reported these years, such as secretion of non-specific anti-bacteria and anti-fungi substances in mucus, the respiratory burst, enzyme activity of immune cells and so on, is believed important for fish disease defense. The specific immunity of mucosal tissues also attracts much interest and makes great achievement in antigen presenting, MHC genes, antibody producing and antibody secreting cells, comparison of serum and mucus immunoglobulin, relationships of immune response between different mucosal immune tissues. Whether mucosal immune system is independent of systematic immune system is another interesting question and causes great concern. In recent years, some evidences from phyletic evolution and ontogenesis show that mucosal immunity is prior to systematic immunity in evolution. Dynamics of antibody producing of mucosal tissues and serum in immersion or oral vaccines immunized fish also shows immune response can be elicited in mucosal tissues independent of systematic immune system. Some researchers also begin to pay attention to factors involved in mucosal immune regulations, for instance, neuromodulators and cytokines. The level of these factors changes in fish immune response process but the mechanisms of regulation still remain unknown. Prospect of the promising future of fish mucosal immunity has also been discussed in this review.     

The role of Toll‐like Receptors in innate immunity and infectious diseases of teleost

The immune defence mechanism depends mainly on germ‐line encoded pattern‐recognition receptors (PRRs). These PRRs respond to many exogenous pathogens and/or endogenous serious signals, by recognizing some highly conserved structures such as pathogen‐associated molecular patterns (PAMPs) and danger/damage associated molecular patterns (DAMPs). Till date, the most studied PRRs are Toll‐like receptors (TLRs). Upon activation of TLRs, there is production of inflammatory cytokines and type I interferons (IFNs) via myeloid differentiation primary response gene 88 (MyD88)‐dependent or ‐independent signalling, respectively, modulating innate and adaptive immunity, as well as inflammatory responses. In fish species studied to date, there are more than 17 TLRs that are identified with some showing homology to mammals, and some are unique for teleost. In the present review, more light are to be shed on the classification, structure and specific ligands of TLRs, with focuses on their signal pathways and different biological activities. Studies of TLRs and their role in the innate immune will potentially have implications for the prevention and treatment of fish diseases.     

Development of a new antibody for detecting natural killer enhancing factor (NKEF)-like protein in infected salmonids

The main cellular responses of innate immunity are phagocytic activity and the respiratory burst, which produces a high amount of reactive oxygen species. Natural killer enhancing factor (NKEF) belongs to the peroxiredoxin family that has an antioxidant function and enhances cytotoxic cell activity. This molecule may play a key role in macrophage and cytotoxic cell communication during the innate immune response of fish against pathogens. In fish, the NKEF gene has been characterized in some species as showing an up-regulation in infected fish, suggesting a trigger effect upon NK-like cells. To detect and localize this molecule in salmonids at protein level, a monospecific polyclonal antibody was generated. A probable NKEF-like protein epitope region was identified and characterized using bioinformatic tools, and the sequence was chemically synthesized using Fmoc strategy, analysed by RP-HPLC and its molecular weight confirmed by mass spectrometry. The synthetic peptide was immunized and antibodies from ascitic fluid were obtained. The resulting antibody is a versatile tool for detecting NKEF by different immune techniques such as ELISA, Western blotting and immunohistochemistry. Analysis of NKEF-like protein is a useful method for characterizing immune properties of this molecule in fish during response to pathogens.     

Fish immune responses against endoparasitic nematodes - experimental models

Vertebrates mount a series of immune reactions when invaded by helminths but antihelmintic immune strategies allow, in many cases, the first invaders of the non-immune host to survive for prolonged periods, whereas subsequent larval invaders of the same parasite species face increased host resistance and thereby decreased colonization success. This concomitant immunity may represent a trade-off between adverse side effects (associated with killing of large helminths in the host tissue) and the need for future protection against invasion. Encapsulation and isolation of large live endoparasitic larvae may be associated with less pathology compared to coping with excess dead parasite tissue in host organs. Likewise, live adult nematodes may be accepted in tissues at a certain activity level for the same reasons. Various host cell receptors bind helminth molecules after which signal-transducing events lead to mobilization of specific reaction patterns depending on the combination of receptors and ligands involved. Both innate and adaptive responses (humoral and cellular) are prominent actors, but skewing of the Th1 lymphocyte response towards a Th2 type is a characteristic element of antihelminthic responses in mammalian hosts. Similar patterns may be expected also to occur in at least some fish species, such as salmonids, producing relevant cytokines, MHCII and CD4+ cells required for these lymphocyte subpopulations. Atlantic cod, Gadus morhua L., is without these immunological elements that indicate that alternative reaction pathways exist in at least some fish groups. Recent achievements within teleost immunology have made it possible to track these host responses in fish and the present work outlines the main immune reactions in fish against helminths and suggests three experimental fish models for exploration of these immune pathways in fish infected with nematodes.     

Characterization of serum and mucosal antibody responses and relative per cent survival in rainbow trout, Oncorhynchus mykiss (Walbaum), following immunization and challenge with Flavobacterium psychrophilum

Serum and mucosal antibody responses of juvenile rainbow trout, Oncorhynchus mykiss, were characterized by enzyme‐linked immunosorbent assay (ELISA) following immunization with various preparations of formalin‐killed Flavobacterium psychrophilum cells. The protective nature of these preparations was then determined by immunizing rainbow trout fry and challenging with the bacterium. Juvenile rainbow trout immunized intraperitoneally (i.p.) with formalin‐killed F. psychrophilum emulsified with Freund's complete adjuvant (FCA), and i.p. with formalin‐killed F. psychrophilum either with or without culture supernatant generated significant serum antibody responses by 6 and 9 weeks, respectively. Significant mucosal antibody responses were detected by 9 weeks only in fish immunized i.p. with killed F. psychrophilum/FCA. Following immunization and bacterial challenge of rainbow trout fry, protective immunity was conferred in F. psychrophilum/FCA and saline/FCA groups with relative per cent survival values of up to 83 and 51, respectively. Significant protection was not observed in treatment groups immunized by immersion or i.p. without adjuvant at the challenge doses tested. Results suggest that stimulation of non‐specific immune factors enhances the ability of fish to mount a protective immune response, but specific antibody appears necessary to provide near complete protection. In this study, an ELISA was developed to monitor anti‐F. psychrophilum antibody production in trout. The relationship of such responses to protective immunity suggests that future vaccination strategies against coldwater disease may require stimulation of both the innate and adaptive arms of the immune response.     

Lysozyme: an important defence molecule of fish innate immune system

The innate immune system of fish is considered to be the first line of defence against a broad spectrum of pathogens and is more important for fish as compared with mammals. Lysozyme level or activity is an important index of innate immunity of fish and is ubiquitous in its distribution among living organisms. It is well documented that fish lysozyme possess lytic activity against both Gram‐positive bacteria and Gram‐negative bacteria. It is also known to be opsonic in nature and activates the complement system and phagocytes. It is present in mucus, lymphoid tissue, plasma and other body fluids of freshwater and marine fish. It is also expressed in a wide variety of tissues. Lysozyme activity has been shown to vary depending on the sex, age and size, season, water temperature, pH, toxicants, infections and degree of stressors. Here, we review our current understanding of different types of lysozyme and their expression and its role in fish innate immune system.     

中国明对虾C 型凝集素基因(Fclectin)的重组表达及活性分析

研究拟通过分析对虾C型凝集素的活性特点,探讨其在对虾先天免疫应答过程中的潜在功能以及在养殖生产实践中的应用。实验利用原核表达系统对中国明对虾C-型凝集素基因的两个串联的糖识别结构域(carbohydrate recognition domain,CRD)进行了重组表达,并通过纯化复性获得了重组目的蛋白(rFclectin-CRD1和rFclectin-CRD2)。活性分析结果显示,重组目的蛋白对多种病原菌有凝集和抑制生长的作用,并且具有Ca2+依赖活性;其凝集活性可被半乳糖、肽聚糖、脂多糖等多种病原相关分子模式所抑制,研究结果证实,Fclectin是一种典型的C-型凝集素,它可能作为中国明对虾先天免疫中重要的模式识别受体,在一定程度上参与了机体应答病原微生物的防御过程。