雌二醇对虹鳟免疫功能的影响*

腺的成熟,大麻哈鱼属鱼类的抗病能力降低,其中的许多种在排卵(精)后死亡.虹鳟在繁殖季节后虽然不至于死亡,但极容易受许多病原菌的感染而发生病患.我们以前的研究提示,疾病的易感性可能与性激素的水平相关,但那些研究主要集中于雄激素对虹鳟免疫功能的影响.雌二醇是重要的雌激素之一,本研究运用酶联吸附免疫法、酶联吸附斑点法以及微量活性氧测定法,主要探讨了雌二醇在体内或体外对抗体的水平、产生抗体的细胞的数目和巨噬细胞产生活性氧的水平的影响.实验结果表明:(1)口服雌二醇能够提高未成熟虹鳟血浆中的雌二醇水平,这种提高能显著地降低血浆中的免疫球蛋白M(IgM)的浓度;(2)在离体的条件下雌二醇也能按剂量依赖方式降低来自成熟红鳟血液、头肾和脾脏的淋巴细胞中产生抗体的细胞数量以及抑制头肾的巨噬细胞产生各种活性氧.这些结果提示随着性腺成熟,免疫功能的降低可能与血液中雌二醇水平的升高有关,免疫功能的降低可能是导致生殖季节雌性虹鳟易受许多病原菌感染的原因之一.     

雌二醇对鳟免疫功能的影响

伴随着性腺的成熟,大麻哈鱼属鱼类的抗病能力降低,其中的许多种在排卵（精）后死亡。虹鳟在繁殖季节后虽然不至于死亡,但极容易受许多病原菌的感染而发生病患,我们以前的研究提示,疾病的易感性可能与性激素的水平相关,但那些研究主要集中于雄激素对虹鳟免疫功能的影响,雌二醇是重要的雌激素之一,本研究运用酶联吸附免疫法、酶联吸附斑点法以及微量活性氧测定法,主要探讨了雌二醇在体内或体外对抗体的水平、产生抗体的细胞的数目和噬细胞产生活性氧的水平的影响。实验结果表明：（1）口服雌二醇能够提高未成熟虹鳟血浆中的雌二醇水平,这种提高能显著地降低血浆中的免疫球蛋白M（IgM）的浓度;（2）在离体的条件下雌二醇也能按剂量依赖方式降低自成熟红鳟血液、头肾和脾脏的淋巴细胞中产生抗体的细胞数量以及抑制头肾的巨噬细胞产生各种活性氧。这些结果提示随着性腺成熟,免疫功能的降低可有与血液中雌二醇水平的升高有关,免疫功能的降低可能是导致生殖季节雌性虹鳟易受许多病原菌感染的原因之一。     

17α，20β-二羟黄体酮降低虹鳟鱼血浆免疫球蛋白M的水平(英文)

本文首次报道１７α,２０β－二羟黄体酮对虹鳟免疫功能的影响。在对生殖季节虹鳟血浆１７α,２０β－二羟黄体酮、免疫球蛋白Ｍ和总蛋白水平的调查时发现,雌雄虹鳟血浆１７α,２０β－二羟黄体酮的水平与免疫球蛋白Ｍ的水平负相关,即在雌性相关系数ｒ＞－０．７９７,Ｐ＞０．０１;而在雄性ｒ＞－０．４７７,Ｐ＞０．０５。这提示１７α,２０β－二羟黄体酮可能抑制免疫球蛋白Ｍ的合成或分泌。产卵或排精后的血浆总蛋白量低     

低鱼粉饲料中添加苏氨酸对三倍体虹鳟抗氧化能力、消化生理及肠道炎症因子基因表达的影响

研究低鱼粉饲料中不同水平苏氨酸对三倍体虹鳟(Oncorhynchus mykiss)生长、抗氧化能力、消化生理及肠道炎症因子基因表达的影响, 以初始体质量(18.42±0.20) g的三倍体虹鳟为研究对象, 在室内200 L流水水族箱中进行摄食生长试验56d。在基础饲料中添加L-苏氨酸, 配制苏氨酸水平分别为0.45%(对照)、0.76%、1.09%、1.29%和1.64%的5种等氮等能饲料。将450尾三倍体虹鳟随机分为5组, 每组处理3个重复, 每个重复30尾鱼。结果表明, 当添加量为1.09%、1.29%和1.64%时, 血清超氧化物歧化酶(SOD)活力显著提高, 同时显著降低了丙二醛(MDA)的含量(P<0.05), 过氧化氢酶(CAT)在1.29%组含量最高, 相比对照组差异显著(P<0.05)。在饲料中添加苏氨酸(0.76%—1.64%)时, 肠道脂肪酶(LPS)和胰蛋白酶的消化酶活性显著高于对照组(P<0.05), 但随着饲料苏氨酸水平达到1.64%时, 消化酶活性有所降低。1.09%和1.65%组的肠道组织形态最佳, 绒毛发达, 排列整齐, 无融合和脱落现象。苏氨酸水平对肠道炎症因子IL-2、IL-8、IL-10、IgM、TNF-α和PepT1都有显著影响(P<0.05), 随着苏氨酸水平的增加, IL-2、IgM和PepT1呈现先升高后降低的变化趋势, 在1.29%和1.09%组的表达量达到最高; IL-8呈逐渐升高趋势; 而IL-10和TNF-α呈下降的变化趋势。研究结果表明, 在低鱼粉饲料(15%)中添加适量水平的苏氨酸, 对提高三倍体虹鳟幼鱼生长、消化酶活性、促进肠道形态结构发育和增强免疫具有积极的作用。     

粪便激素水平反映不同出生时期雄性布氏田鼠的繁殖策略

布氏田鼠是我国内蒙古草原的主要害鼠之一,具有明显的季节繁殖特征,不同季节出生个体可能具有不同的繁殖策略,但尚缺乏内分泌证据支持。本研究采用标志重捕法,连续监测大型自然围栏中不同年龄雄性布氏田鼠的繁殖状态和应激水平的季节变化,分析各年龄组雄鼠的繁殖发育策略。结果表明,越冬雄鼠可保持较高的睾丸下降率和睾酮水平至8月初;而部分5月生雄鼠的繁殖期睾酮水平较接近越冬鼠,但8月初已降至年内最低水平,而6月及以后出生雄鼠睾酮始终处于较低水平;这说明越冬鼠的繁殖状态可贯穿繁殖期始终,只有部分5月生雄鼠可能参与当年繁殖,而6月及以后出生雄鼠则不能在当年繁殖。越冬鼠皮质醇水平高于当年鼠,繁殖期高于非繁殖期,这可能是越冬鼠由于繁殖需要而保持较高应激状态,从而造成繁殖盛期后死亡率升高。这些结果说明,不同时期出生的雄性布氏田鼠具有不同的出生后性腺发育模式和繁殖策略,反映出生存与繁殖之间的权衡。     

黄芪多糖免疫草鱼母本对子代IgM、C3和LSZ表达特性研究

为探究黄芪多糖免疫草鱼母本所产子代早期发育阶段体液免疫因子IgM、C3和LSZ的表达特性及代间传递效率。采用ELISA、RT-qPCR等方法分析了饲喂黄芪多糖草鱼母本血液及其子代早期发育阶段3种免疫因子的蛋白质活性及mRNA水平。结果显示,黄芪多糖免疫草鱼母本血液中IgM、C3和LSZ蛋白活性均显著高于对照组。在子代早期阶段中,3种免疫因子蛋白活性呈先下降后上升趋势,实验组IgM蛋白活性在各阶段均显著高于对照组,在卵子、5d和28d仔鱼中分别提高了2.2倍、1.7倍和1.8倍。实验组C3活性在卵子、24h胚胎分别提高了1.9倍和1.6倍。实验组LSZ活性在卵子、5d和28d仔鱼中分别提高了2.4倍、2.0倍和1.9倍;在卵子和受精卵时期3种免疫因子mRNA水平显著高于对照组。在24h胚胎至5d仔鱼没有检测到IgM和LSZ mRNA,而在14d后3种免疫因子的mRNA均呈上调表达,但Ig M和C3 mRNA水平与对照组无显著差异。GCRV攻毒后,实验组2月龄草鱼脾脏和头肾中IgM mRNA水平显著高于对照组。结果表明,黄芪多糖能够提高草鱼母本免疫能力并向子代垂直传递,在应对GCRV感染时发挥一定免疫保护作用。     

圈养雄性麋鹿血清睾酮和皮质醇含量在发情期的变化

2000年7-8月在北京麋鹿苑采集不同繁殖状态雄鹿（发情前3头、发情3头和发情后4头）血液样品,用放射免疫分析法测定血清睾酮和皮质醇含量。结果显示发情雄鹿的睾酮浓度显著高于发情前、后,而3种状态雄鹿的皮质醇浓度无差异,并且与睾酮浓度无关。本苑麋鹿睾酮水平的变化与同类野生动物一致。     

皮质醇的周年变化与雌性虹鳟性腺成熟的关系

利用放射免疫分析法对饲养于恒定水温和自然光照下的雌性虹鳟血浆中皮质醇和性激素含量的周年变化进行了测定．结果表明：１）根据性腺结构指数和性激素分泌量判断,三龄时,雌性虹鳟达到性成熟;２）在血浆中不仅性激素而且皮质醇的变化水平与性腺结构指数的变化高度相关．排卵前性激素的水平都较高,伴随着排卵的进行性激素水平下降．而且在产卵季节虹鳟血浆中皮质醇水平也较高,三龄时皮质醇水平与性腺结构指数的相关系数为０．８６．这些结果提示,皮质醇在虹鳟的繁殖过程中可能发挥某种作用．     

注射LHRHa对泥鳅血清性类固醇激素变化研究

研究了注射促黄体激素类似物(LHRHa)后,泥鳅(Misgurnus anguillicaudatus)血清性类同醇激素的变化规律,并探讨在泥鳅繁殖季节时.孕酮(P)、睾酮(T)和雌二醇(E2)对性腺发育的作用及调节机制.实验共分两组,对照组和实验组;对照组只注射生理盐水;实验组注射LHRHa,雌鱼0.2 μg/g,雄鱼减半.注射前尾静脉采血,作为血液样本分析基础水平(Oh),注射药物后分别在7、24、48、72和96h尾静脉采血.测定雌鱼睾酮、雌二醇和孕酮,雄鱼孕酮和睾酬血清浓度.实验结果表明:注射LHRHa,雌鱼血清睾酮和雌二醇浓度显著高于对照组,雄鱼血清睾酮和孕酮显著高于对照组(P<0.05);24h浓度较高.雌鱼孕酬、睾酮和雌二醇分别为(0.710±0.082)ng/mL、(9.00±0.57)ng/mL和(696.4±26.2)pg/mL,雄鱼孕酮和睾酮分别为(0.527±0.121)ng/mL和(9.62±0.62)ng/mL.实验组雌鱼孕酮变化基本规律为,基础水平(0-7h)-逐渐升至最高(7-24h)-逐渐降至基础水平(24-48h)-维持基础水平(48-96h).实验组雌鱼睾酮和雌二醇与雄鱼孕酮和睾酮变化规律基本相似,其规律为,逐渐上升至最高(0-24h)-逐渐降至基础水平(24-72h)-维持基础水(72-96h).24h对照组雌鱼睾酮和雌二醇显著升高,浓度分别为:睾酮(2.20±0.18)ng/mL,雌二醇(269.1±36.6)pg/mL.对照组雄鱼血清孕酮和睾酮浓度实验期间均无显著变化.研究认为:LHRHa能够刺激泥鳅性类同醇激素分泌,特别是睾酮的分泌,显著提高雌鱼性腺指数(GSI).但刺激P的分泌调控能力有限,实验期间处于较低水平,诱导排卵效果差,泥鳅的性类固醇激素可能有特殊的调节机制.雌二醇和睾酮对性腺成熟有重要作用,孕酮对介导卵细胞最终成熟和排卵可能起重要作用,而雌二醇和睾酮无明显效果.     

脾切除后免疫功能与血流变的相关性研究

目的:观察大鼠脾切除后免疫功能与血流变的相关性。方法:在水合氯醛麻醉下给大鼠行脾切除术制备模型大鼠,并分组给与不同处理研究脾切除后免疫功能与血流变的相关性。结果:通过统计学分析证明脾切除能明显升高脾切除模型大鼠的全血黏度、全血还原黏度、聚集指数以及电泳指数,血小板,降低免疫功能,而肝素钠能在一定程度上降低这种改变,进而提高机体的免疫功能。结论:脾切除能升高大鼠的全血黏度、全血还原黏度、聚集指数以及电泳指数,血小板,从而增加大鼠患血栓性疾病的机率,并可以降低大鼠的免疫功能,而肝素钠能在一定程度上降低这种血液高黏度的改变,进而提高机体的免疫功能,证明血流变与免疫功能有正相关性。     

Influence of salinity and ratio of lipid to protein in diets on certain enzyme activities in rainbow trout (Salmo gairdneri Richardson)

The connection between metabolic and sea water adaptation of the rainbow trout was investigated. The rainbow trout were kept in fresh water and diluted sea water of 8 and 20 0/00 S at 16 degrees C and fed on three different diets for 51 days. Hyperosmotic salinity (20 0/00) tends to inhibit growth in rainbow trout by reducing the food conversion efficiency. A higher protein concentration in the diet can partly compensate for this effect. The liver IDH, G6PDH and 6PGDH activities of the rainbow trout are influenced only by food quality, whereas the liver G1DH, AspT and A1T activities, like the muscle A1T, are also affected by salinity. The salinity had no significant effect on the activities of the kidney enzymes we investigated (Na/K-ATPase, G1DH, A1T, AspT) or of the muscle AspT in these experiments.     

Triiodothyronine is needed for the acclimatization of brown trout (Salmo trutta) or rainbow trout (Oncorhynchus mykiss) to seawater]

Brown and rainbow trout, held in freshwater at 13 +/- 1 degrees, were injected, every 3 days, with iopanoic acid (IOP: 5 mg/100 g body wt), an inhibitor of deiodination of thyroxine (T4) to triiodothyronine (T3). One group of IOP-treated rainbow trout was immersed in T3 (20 micrograms/l water). In IOP trout, plasma T3 fell to very low levels by day 7, while changes in T4 levels were less marked. In IOP + T3 trout plasma T3 increased fivefold, plasma T4 being unchanged. No mortality occurred and plasma osmolarity (OP) was not altered by any treatment. After direct transfer to seawater (30/1000), IOP trout were unable to acclimate to salinity: all died within 2 or 3 days, while the survival at day 3 was 100% in control brown trout and 45 and 74% in control and IOP + T3 rainbow trout respectively. OP increased more in IOP and less in IOP + T3 than in controls. There was a significant inverse correlation between T3, but not T4, plasma level, at the time of transfer and the OP 1 day later. In conclusion, although T3 does not play a significant role in osmoregulation in freshwater, T3 and therefore the deiodination of T4 into T3, were required for the development of hypo-osmoregulatory capacity involved in acclimation of trout to seawater.     

The phylogeography of salmonid proliferative kidney disease in Europe and North America

Salmonid proliferative kidney disease (PKD) is caused by the myxozoan Tetracapsuloides bryosalmonae. Given the serious and apparently growing impact of PKD on farmed and wild salmonids, we undertook a phylogeographic study to gain insights into the history of genealogical lineages of T. bryosalmonae in Europe and North America, and to determine if the global expansion of rainbow trout farming has spread the disease. Phylogenetic analyses of internal transcribed spacer 1 sequences revealed a clade composed of all North American sequences plus a subset of Italian and French sequences. High genetic diversity in North America and the absence of genotypes diagnostic of the North American clade in the rest of Europe imply that southern Europe was colonized by immigration from North America; however, sequence divergence suggests that this colonization substantially pre-dated fisheries activities. Furthermore, the lack of southern European lineages in the rest of Europe, despite widespread rainbow trout farming, indicates that T. bryosalmonae is not transported through fisheries activities. This result strikingly contrasts with the commonness of fisheries-related introductions of other pathogens and parasites and indicates that fishes may be dead-end hosts. Our results also demonstrate that European strains of T. bryosalmonae infect and induce PKD in rainbow trout introduced to Europe.     

虹鳟补体活化调节因子CD46的分子特征及功能

为进一步研究硬骨鱼类中补体活化调节因子的分子特征和功能,研究克隆了虹鳟(Oncorhynchus mykiss)的CD46基因,对其分子特征进行了系统分析,结果显示:虹鳟CD46基因由10个外显子和9个内含子组成,cDNA序列全长2812 bp,编码317个氨基酸,蛋白序列由1个信号肽、4个SCR结构域、1个跨膜区和1个胞内区组成,预测分子量为33.9 kD。基因组共线性分析显示,虹鳟CD46基因位于16号染色体,其基因座在脊椎动物中具有保守的共线性。组织和白细胞亚群表达分析显示,虹鳟CD46基因在各种组织和白细胞亚群中均有表达。为了进一步阐明虹鳟CD46的免疫功能,研究原核表达纯化了标签蛋白GST和融合蛋白GST-CD46。溶血活性实验表明,与GST相比, GST-CD46能够显著抑制虹鳟血清对兔红细胞的溶血活性,且呈现剂量依赖效应,表明虹鳟CD46是补体活化的调节因子。此外,研究用HEK293T细胞过表达了GFP和GFP-CD46。细胞损伤实验显示,与GFP相比, GFP-CD46能够显著抑制虹鳟血清对HEK293T细胞的损伤,进一步表明虹鳟CD46是补体活化的调节因子,能够保护细...     

Rhabdovirus infection induces public and private T cell responses in teleost fish.

Many viruses induce a strong T cell response that contributes to the elimination of infected cells presenting viral peptides by MHC molecules. The structure and expression of genes encoding molecules homologous to mammalian alphabeta TCRs have been recently characterized in rainbow trout and in several teleost species, but the alphabeta T cell response against pathogens has not been directly demonstrated. To study the modifications of the T cell repertoire during an acute viral infection in rainbow trout, we adapted the immunoscope methodology, which consists of spectratyping the complementarity-determining region 3 length of the TCRbeta chain. We showed that the naive T cell repertoire is polyclonal and highly diverse in the naive rainbow trout. Using viral hemorrhagic septicemia virus (VHSV), which provokes an acute infection in rainbow trout, we identified skewed complementarity-determining region 3 size profiles for several VbetaJbeta combinations, corresponding to T cell clonal expansions during primary and secondary response to VHSV. Both public and private T cell expansions were shown by immunoscope analysis of spleen cells from several infected individuals of a rainbow trout clone sharing the same genetic background. The public response to VHSV consisted of expansion of Vbeta4Jbeta1 T cell, which appeared early during the primary response and was strongly boosted during the secondary response.     

The correlation between fish growth and several biochemical characteristics with reference to the steelhead <Emphasis Type="Italic">Parasalmo mykiss</Emphasis> Walb.

Correlation analysis demonstrated a statistically significant correlation of linear-weight characteristics of the steelhead rainbow trout (cultivated steelhead form) with an RNA/DNA ratio and the expression level of the gene encoding cytochrome c oxidase (CCO) in two-year-old individuals (1+) as well as the expression level of the gene encoding the myosin heavy chain (MyHC) and activities of the enzymes CCO and lactate dehydrogenase (LDH) in muscles and 1-glycerophosphate dehydrogenase (1-GPDH) and glucose-6-phosphate dehydrogenase (G6PDH) in the liver of two- and three-year-old individuals (1+ and 2+). With age, the correlation of 1-GPDH and G6PDH activities in the rainbow trout liver with the fish body weight increased, whereas their correlations with the body length reduced. The age- and sex-related distinctions in the MyHC gene expression and activities of the white muscle enzyme LDH and the liver enzymes 2-GPDH and G6PDH were detected in rainbow trout of both age cohorts.     

Susceptibility of rainbow trout Oncorhynchus mykiss,Atlantic salmon Salmo salar and coho salmon Oncorhynchus kisutch to experimental infection with sea lice Lepeophtheirus salmonis

Physiological, immunological and biochemical parameters of blood and mucus, as well as skin histology, were compared in 3 salmonid species (rainbow trout Oncorhynchus mykiss, Atlantic salmon Salmo salar and coho salmon O. kisutch) following experimental infection with sea lice Lepeophtheirus salmonis. The 3 salmonid species were cohabited in order to standardize initial infection conditions. Lice density was significantly reduced on coho salmon within 7 to 14 d, while lice persisted in higher numbers on rainbow trout and Atlantic salmon. Lice matured more slowly on coho salmon than on the other 2 species, and maturation was slightly slower on rainbow trout than on Atlantic salmon. Head kidney macrophages from infected Atlantic salmon had diminished respiratory burst and phagocytic capacity at 14 and 21 d post-infection (dpi), while infected rainbow trout macrophages had reduced respiratory burst and phagocytic capacities at 21 dpi, compared to controls. The slower development of lice, coupled with delayed suppression of immune parameters, suggests that rainbow trout are slightly more resistant to lice than Atlantic salmon. Infected rainbow trout and Atlantic salmon showed increases in mucus lysozyme activities at 1 dpi, which decreased over the rest of the study. Mucus lysozyme activities of infected rainbow trout, however, remained higher than controls over the entire period. Coho salmon lysozyme activities did not increase in infected fish until 21 dpi. Mucus alkaline phosphatase levels were also higher in infected Atlantic salmon compared to controls at 3 and 21 dpi. Low molecular weight (LMW) proteases increased in infected rainbow trout and Atlantic salmon between 14 and 21 dpi. Histological analysis of the outer epithelium revealed mucus cell hypertrophy in rainbow trout and Atlantic salmon following infection. Plasma cortisol, glucose, electrolyte and protein concentrations and hematocrit all remained within physiological limits for each species, with no differences occurring between infected and control fish. Our results demonstrate that significant differences in mucus biochemistry and numbers of L. salmonis occur between these species.     

Transmission of Tetracapsuloides bryosalmonae (Myxozoa: Malacosporea) to Fredericella sultana (Bryozoa: Phylactolaemata) by various fish species

Tetracapsuloides bryosalmonae is a myxozoan parasite of salmonids and freshwater bryozoans, which causes proliferative kidney disease (PKD) in the fish host. To test which fish species are able to transmit T. bryosalmonae to bryozoans, an infection experiment was conducted with 5 PKD-sensitive fish species from different genera. Rainbow trout Oncorhynchus mykiss, brown trout Salmo trutta, brook trout Salvelinus fontinalis, grayling Thymallus thymallus and northern pike Esox lucius were cohabitated with T. bryosalmonae-infected Fredericella sultana colonies and then subsequently cohabitated with statoblast-reared parasite free Bryozoa. Statoblasts from infected colonies were tested by PCR to detect cryptic stages of T. bryosalmonae, which may indicate vertical transmission of the parasite. In this study, brown trout and brook trout were able to infect Bryozoa, while there was no evidence that rainbow trout and grayling were able to do so. Few interstitial kidney stages of the parasite were detected by immunohistochemistry in brown trout and brook trout, while rainbow trout and grayling showed marked proliferation of renal interstitial tissue and macrophages with numerous parasite cells. Intraluminal stages in the kidney tubules were only detected in brown trout and rainbow trout. In contrast to previous observations, pike was not susceptible to PKD in these trials according to the results of T. bryosalmonae-specific PCR. No DNA of T. bryosalmonae was detected in any statoblast.