Light and Electron Microscopic Studies on Alpha Naphtyl Acetate Esterase Activity of the Periphral Blood T Lymphocytes in Van Cats

The purpose of this study was to determine the percentage of peripheral blood T lymphocytes and the localization of ANAE enzyme at the electron microscopic level in Van cats by using an alpha-naphtyl acetate esterase procedure. Peripheral blood samples taken from 20 Van cats were used. The percentage of ANAE positive lymphocytes was 83.0%. Neutrophilic gra-nulocytes gave a negative reaction, whereas monocytes, eosinophilic granulocytes showed a diffuse granular positivity. In the electron microscopic examination, ANAE positive reactions were seen in lysosomal granules found in lymphocytes.     

感染网状内皮组织增殖病病毒SPF雏鸡局部免疫组织的免疫学变化

研究１日龄ＳＰＦ雏鸡感染网状内皮组织增殖病病毒后局部免疫组织均浆涂片的免疫学变化。结果表明：雏鸡感染ＲＥＶ后其局部免疫组织中,淋巴细胞数、ＡＮＡＥ、Ｔ细胞数以及颗粒型和弥散型ＡＮＡＥ＾＋Ｔ淋巴细胞数明显低于未感染雏鸡。说明病消化道和呼吸道相关的局部免疫组织的细胞免疫呈现抑制。     

Morphology of blood cells in carp (Cyprinus carpio L.)

The morphology of blood cells in the carp was investigated by light and electron microscopy. Erythrocytes, thrombocytes, lymphocytes, granulocytes and monocytes were identified as the peripheral blood cells. Thrombocytes were round to long oval, each containing vesicular and microtubular structures and an oval nucleus with abundant heterochromatins. Lymphocytes were divided into three types in size, small, medium and large. Some of the small and medium lymphocytes were alpha-naphthyl-acetate esterase (ANAE) positive, while large lymphocytes were pyroninophilic. Granulocytes were distinguished into three types (type I, type II and type III) according to the morphology of the nucleus and granules. Type I granulocytes possessed lobulated nuclei and a large number of cytoplasmic granules, each of which was oval and contained electron-dense materials and a crystalloid. Type II granulocytes had small eccentric nuclei and were subdivided into IIa and IIb granulocytes by electron microscopic analysis. Granules of type IIa granulocytes were furnished with an electron-dense rim. Granules of type IIb granulocytes were larger than those of type IIa, containing randomly distributed electron-dense and electron-lucent materials. Type III granulocytes possessed round nuclei and a few large granules. The granules were filled with regularly arranged fibriform materials and some needle-like structures. Monocytes were morphologically similar to those of mammals.     

鸡马立克病CVI988疫苗不同接种部位对免疫效果的影响

探讨鸡马立克病CVI988疫苗不同的接种部位对免疫效果的影响,将96只1日龄海兰褐公雏鸡随机分成对照组(C)、腹腔注射接种组(PI)和颈部皮下接种组(HI),C组腹腔注射0.2mL疫苗稀释液,PI组和HI组注射0.2mL CVI988疫苗。在第3、6、10、15、20、26、35、45天对3组各4只鸡进行心脏采血,制作血涂片,分别进行ANAE和甲基绿-派洛宁染色,观察血液中阳性B淋巴细胞和T淋巴细胞百分率的变化。结果显示,在免疫早期(3日龄～6日龄),免疫组的阳性T淋巴细胞显著低于对照组,而阳性B淋巴细胞在第3天就显著高于对照组;在免疫中后期,免疫组的阳性T淋巴细胞和B淋巴细胞基本都显著高于对照组;并且PI组的阳性T淋巴细胞高于或显著高于HI组,阳性B淋巴细胞在第15天显著高于HI组。由此可见,CVI988疫苗接种第3天产生体液免疫效应,10d后细胞免疫效应高于对照组,即早期免疫效果主要靠体液免疫,腹部免疫接种效果优于颈部免疫,为马立克病更有效的免疫接种提供了理论依据。     

感染网状内皮组织增殖病病毒SPF雏鸡中枢和外周免疫器官的免疫学变化

实验研究了１日龄ＳＰＦ雏鸡感染网状内皮组织增殖病病毒（Ｒｅｔｉｃｕｌｏｅｎｄｏｔｈｅｌｉｏｓｉｓｖｉｒｕｓ,ＲＥＶ）后免疫器官匀浆涂片的免疫学变化。结果发现,雏鸡感染ＲＥＶ后其免疫器官法氏囊、胸腺、脾脏中淋巴细胞数、酯酶阳性（ＡＮＡＥ＋）Ｔ细胞数以及颗粒型和弥散型ＡＮＡＥ＋Ｔ淋巴细胞数显著降低（Ｐ＜０．０５,Ｐ＜０．０１）。表明１日龄ＳＰＦ雏鸡感染ＲＥＶ后,其中枢免疫器官和外周免疫器官的细胞免疫呈现抑制。     

Alpha-naphthyl acetate esterase activity is not a specific marker for ovine T lymphocytes

Alpha-naphthyl acetate esterase (ANAE) activity was demonstrated in ovine lymphocytes harvested from blood on Ficoll-metrizoate gradients. The enzyme's specificity for T lymphocytes, identified by immunofluorescent staining of T cell-specific antigens, was assessed. Correlation analysis of the results obtained using unfractionated lymphocytes from 12 sheep showed no correlation between ANAE activity and the expression of T cell antigens (r = 0.22). When lymphocytes from 4 sheep were fractionated on nylon wool columns a mean of only 43.2% of the cells in the non-adherent population were ANAE-positive whereas 94.7% of these cells were identified as T lymphocytes. Blood lymphocytes from 5 animals were separated into 3 fractions using Percoll discontinuous density gradients. No significant relationship was seen between ANAE activity and T cells in Fractions 1 and 3 (r = 0.41 and 0.21). Fraction 2 cells, however, did show a significant positive relationship (r = 0.91) between these two features but the biological significance of this relationship is unknown. It was concluded that ANAE activity is not a specific marker for ovine T lymphocytes.     

复方中草药"毒菌杀"对感染IBDV雏鸡白细胞数量及淋巴细胞ANAE+百分率的影响

试验观察了复方中草药“毒菌杀”对感染法氏囊病毒(IBDV)雏鸡的外周血液白细胞及α-醋酸萘酯酶(α-acid naphthyl acetate esterase,ANAE)阳性淋巴细胞百分率的影响。结果发现,“毒菌杀”可以使感染IBD V的雏鸡维持外周血白细胞数量的相对稳定,提高ANAE阳性淋巴细胞的活性水平,具有明显的免疫保护作用。     

Morphology, cytochemical staining, and ultrastructural characteristics of the blood cells of the giant lizard of El Hierro (Gallotia simonyi)

The object of this study was to examine the erythrocytes, leukocytes and thrombocytes of the giant lizard of El Hierro (Gallotia simonyi) by light and electron (TEM) microscopy, and cytochemical staining. Smears were prepared from blood from the ventral coccygeal vein of 10 healthy adult lizards (five males and five females) from the Giant Lizard of El Hierro Reproduction and Research Centre, Canary Islands, Spain. The cytochemical stains used were: benzidine peroxidase (BP), chloroacetate esterase (CAE), alpha-naphthyl acetate esterase (ANAE), acid phosphatase (AP), periodic acid-Schiff (PAS), toluidine blue (TB) and May-Grünwald-Giemsa (MGG). Electron microscopy was also performed on all samples. Heterophils had granules that were heterogeneous in both size and electron density, and stained with BP, PAS and ANAE. Eosinophil granules were homogeneously electron-dense and stained for AP, CAE and ANAE. Basophils had both highly and moderately electron-dense granules, and stained with TB and ANAE. Azurophil granules were of low electron-density and stained for AP, CAE and ANAE. Azurophil cytoplasm was vacuolated on TEM. The cytoplasm of lymphocytes contained many ribosomes and was positive for AP. Monocytes had a large nucleus and a vacuolated cytoplasm but did not stain by any of the cytochemical methods used. Thrombocytes had a relatively large nucleus but little cytoplasm; they did not stain cytochemically. The blood cells of the giant lizards of El Hierro differ from those of other members of the Order Squamata both morphologically and cytochemically. The variation in cytochemical responses in the blood of reptiles makes it necessary to study species individually if meaningful clinical decisions are to be made.     

阻断淋巴细胞糖皮质激素受体对鸡免疫功能的影响

为进一步证明糖皮质激素受体（ＧＲ）在介导淋巴细胞免疫功能中的作用,观察ＧＲ阻断鸡对马立克氏病病毒（ＭＤＶ）的感受性,用ＲＵ４８６阻断鸡淋巴细胞ＧＲ,当ＧＲ被阻断６０％以上时（ＲＵ４８６剂量为５０ｍｇ／ｋｇ）,鸡脾细胞诱生ＩＬ－２和ＩＦＮ活性及Ｔ、Ｂ细胞丝裂原刺激增殖活性均明显降低;外周血淋巴细胞、单核细胞、ＡＮＡＥ＋细胞减少;脾脏、法氏囊、胸腺重量与体重之比的相对重量减轻,淋巴细胞系统免疫功能呈低下状态。此时雏鸡对ＭＤＶ的易感性增高。当ＧＲ被阻断不足３５％时（ＲＵ４８６剂量为２５ｍｇ／ｋｇ）,对淋巴细胞系统免疫功能无影响。     

禽网状内皮组织增生病病毒感染对SPF雏鸡血液和局部淋巴组织CD4+/CD8+细胞及相关细胞因子表达的影响

旨在探讨禽网状内皮组织增生病病毒（reticuloendotheliosis virus,REV）对SPF雏鸡血液和局部淋巴组织中T淋巴细胞数量以及相关细胞因子表达的影响。将96只1日龄SPF雏鸡随机分为REV感染组和对照组,应用流式细胞术、酸性α-醋酸萘酯酶染色（acid α-naphthyl acetate esterase,ANAE）、荧光定量PCR等方法对上述指标进行检测。试验数据表明,与对照组相比,感染组雏鸡血液CD4⁺T淋巴细胞数量在第7~35天显著降低、CD8⁺T淋巴细胞数量在第7~28天显著升高,CD4⁺/CD8⁺比值在第7~28天显著降低（均P<0.05或P<0.01）;局部淋巴组织哈德尔腺（Hader’s gland,HG）、派伊尔结（Peyer’s patch,PP）和盲肠扁桃体（caecal tonsil,CT）中ANAE⁺T淋巴细胞数量均显著降低（均P<0.05或P<0.01）;GH、PP和CT中细胞因子IL-2、IFN-γ和TNF-α 转录量都有不同程度升高。本研究表明,REV感染引起雏鸡血液中CD4⁺ T淋巴细胞数量降低、CD4⁺/CD8⁺T淋巴细胞比例失衡、局部淋巴组织中T淋巴细胞数量相对减少及细胞因子TNF-α转录持续升高与REV造成感染雏鸡细胞免疫功能显著降低密切相关。     

Staining of pig lymphocytes subpopulations with acid alpha-naphthyl acetate esterase

The use of alpha-naphthyl acetate esterase (ANAE) as a T cell marker in some other species and the broad correlation of incidence of ANAE-positivity and E rosette-formation in the pig suggest that ANAE-staining may be a T-cell marker in the pig. However, by studying the staining of lymphocytes within a variety of rosettes in fixed preparations a similar incidence of pig blood lymphocytes were found to be ANAE+ among T cells (E rosettes formed in dextran), B cells (antiglobulin rosettes) and Fc-gamma receptor-bearing B and T cells (EA rosettes in saline and dextran): complement (C') receptor-bearing cells showed a higher incidence of staining than other lymphocytes. Analysis of staining morphology suggested that certain morphologies within the B and T lineages may be confined to subpopulations. Thus ANAE positivity is certainly not a marker identifying blood T lymphocytes but could be of some value indicating subpopulations of B and T lymphocytes.     

疑核对细胞免疫机能调节作用的研究

实验用3～4月龄1.5～2.5kg灰色家兔22只,麻醉后于疑核内插入同心圆电极,在刺激前和刺激后采血检测PHA诱导的淋巴细胞转化率、T细胞百分率,并测定全血葡萄糖浓度;刺激疑核后0.5h,采6例胸腺组织作超微结构观察;刺激疑核后3.5h,采胸腺作石蜡切片ANAE阳性率检测。电极位置均作组织学检查。结果:刺激疑核后T细胞百分率、PHA诱导的淋巴细胞转化率持续升高(P<0.05);胸腺各部位ANAE阳性淋巴细胞百分率升高,胸腺后微静脉内淋巴细胞增多,均与对照组及非刺激对照组相差非常显著(P<0.005);胸腺髓质ANAE阳性淋巴细胞率与外周血液T细胞百分率相关显著;胸腺超微结构示上皮性网状细胞机能活跃,分泌旺盛,皮质处于分裂状态的淋巴细胞增多,髓质淋巴细胞活动能力增强。由此认为,疑核与外周细胞免疫存在机能联系,刺激疑核可引起细胞免疫机能增强。本实验还证明,刺激疑核引起血糖浓度降低,因而除神经途径外,疑核还通过体液途径调节免疫机能。实验结果提示,疑核可能是大脑皮层、下丘脑调节免疫机能的中继核。实验确定了疑核定位坐标是AP_(20～21),L_(1～1.5),H_(20～21)。     

A study of peripheral blood in hedgehogs in Turkey

The aim of this study was to determine diameters of blood cells, differential counts of peripheral blood leukocytes, alpha-naphthyl acetate esterase (ANAE), acid phosphatase (ACP-ase) activity of some leukocyte types, and enzymatic positivity percentages of peripheral blood lymphocytes in two hedgehogs species, Hemiechinus auritus, the long-eared hedgehog, and Erinaceus concolor, the southern white-breasted hedgehog. Air-dried peripheral blood smears were stained with May-Grünwald-Giemsa stain. ANAE and ACP-ase were stained in glutaraldehyde-acetone-fixed smears. ANAE-positive lymphocytes displayed a dot-like positivity pattern characterized with 1-5 reddish brown cytoplasmic granules, whereas ACP-ase positive lymphocytes displayed a dot-like positivity pattern characterized with 1-3 pinkish cytoplasmic granules. Monocytes gave a diffuse and strong reaction while neutrophils displayed a weak positive reaction for ANAE and ACP-ase. No difference was observed in mean diameters of peripheral blood cells of these species. It was found that lymphocytes made up the majority (64.3% and 65.5%) of leukocytes, followed by neutrophils (23.9% and 23.3%), eosinophils (9.0% and 7.6%), monocytes (1.8% and 2.3%), and basophils (1.0% and 1.3%) in H. auritus and E. concolor, respectively. Mean ANAE positivity oflymphocytes was 36.6% and 51.3% and ACP-ase positivity was 32.1% and 37.5% for H. auritus and E. concolor, respectively. The ANAE positivity of lymphocytes in E. concolor was significantly (P < 0.05) higher than that of H. auritus.     

雏鸡一次免疫鸡球虫早熟株三价苗后外周血T细胞变化的观察

通过α－醋酸萘酯酶（ＡＮＡＥ）组织化学染色法，对雏鸡接种鸡球虫早熟株三价苗引起细胞免疫情况进行了研究。试验结果表明，免疫组雏鸡Ｔ细胞百分率上升３９．２％，对照组上升８．７％，免疫组明显高于对照组（Ｐ〈０．０１）。     

芪芍增免散对鸡法氏囊病疫苗免疫增强作用的研究

将520只7日龄海兰灰蛋鸡雏随机分成4组,分别设为3个剂量组（0．5％组、1.0％组和1．5％组）和-个空白对照组,给药组分别于14日龄常规剂量滴鼻点眼免疫法必要（鸡传染性法氏囊病中等毒力活疫苗）,28日龄2倍剂量饮水法必妥。通过测定法氏囊病抗体水平、脾指数、法氏囊指数、外周血液酸性-醋酸萘酯酶（ANAE）阳性淋巴细胞...     

Use of alpha-naphthyl acetate esterase staining to identify T lymphocytes in cattle

Peripheral blood lymphocytes (PBL), lymph nodes, and/or spleens from clinically normal cattle were examined for cytochemical staining of alpha-naphthyl acetate esterase (ANAE). Two types of positive-staining patterns in ANAE staining resulted. By a combination of ANAE staining and latex-ingesting test, diffuse ANAE-positive cells were considered as mononuclear phagocytic cells. Using erythrocyte rosettes, erythrocyte antibody complement rosettes, nylon-wool column technique, surface immunoglobulin (SIg) staining, and the ANAE staining technique, granular ANAE-positive lymphocytes were shown to be T lymphocytes. The frequency of T and B lymphocytes in PBL, spleens, and lymph nodes of clinically normal cattle was measured, using ANAE staining and SIg staining. In PBL, 47.7% were ANAE-positive and 26.9% were SIg-positive; in spleens, 22.4% were ANAE-positive and 53.7% were SIg-positive; and in lymph nodes, 38.5% were ANAE-positive and 28.3% were SIg-positive. The frequencies of T and B lymphocytes in PBL, spleens, and/or lymph nodes from cattle with enzootic bovine leukosis (EBL) and cattle with persistent lymphocytosis and in tumor cells from cattle with EBL were measured. When compared with those of clinically normal cattle, PBL, spleens, and lymph nodes of cattle with EBL and the PBL of cattle with persistent lymphocytosis contained numerous SIg-positive cells and few ANAE-positive cells. Tumor cells from cattle with EBL contained 7.3% ANAE-positive and 78.0% SIg-positive cells.     

雏鸡脑软化症细胞免疫功能变化的研究

以1日龄海兰蛋鸡为试验动物，用低VE水平且添加鱼油日粮饲喂雏鸡，复制脑软化症动物模型，研究脑软化症雏鸡血液白细胞数（WBC）、T淋巴细胞总数、T淋巴细胞酸性非特异性α-醋酸萘脂酶（ANAE+）及血清白介素-1β和白介素-2含量的变化。结果表明，患雏的WBC数、ANAE+T淋巴细胞总数、血清白介素含量均明显低于健康对照组雏鸡。结果说明，VE缺乏对雏鸡细胞免疫功能有明显的抑制作用。     

感染IBDV雏鸡血液激素水平和ANAE阳性淋巴细胞动态变化

为探讨内分泌活动及细胞免疫反应在鸡抗传染性法氏囊病毒（ＩＢＤＶ）感染中的调节作用机制而进行了本研究。结果表明,攻毒后１～５ｄ内,未免疫攻毒鸡（Ａ组）、免疫攻毒鸡（Ｂ组）血浆皮质酮均明显上升,而未免疫未攻毒鸡（Ｃ组）则否。Ａ、Ｂ、Ｃ３组血浆Ｔ４水平攻毒前后无明显变化,Ｔ３除在攻毒后第３天Ａ组明显高于Ｂ、Ｃ组外,其他时间无明显差异性变化。血液ＡＮＡＥ阳性淋巴细胞（％）在攻毒后的１～５ｄ内,Ａ组和Ｂ组明显下降,以后回升,至２８ｄ回复到攻毒前水平,并接近于Ｃ组。Ａ组的ＡＮＡＥ阳性淋巴细胞（％）与皮质酮呈显著负相关,与Ｔ３和Ｔ４无明显相关性。     

Decreased expression of L‐selectin (CD62L) on lymphocytes in enzootic bovine leukaemia

Expression of L‐selectin was determined by single‐ and two‐colour immunofluorescence on granulocytes, peripheral blood mononuclear cells (PBMC) and blasts of bovine origin by means of a monoclonal antibody IVA94 which recognizes bovine L‐selectin (CD62L). Cells were separated from peripheral blood of healthy cattle and colleagues infected with bovine leukaemia virus (BLV). BLV‐infected animals comprised lymphocytotic and non‐lymphocytotic cows. L‐selectin was expressed on 90–98 % of granulocytes in all tested animals. The percentage of PBMC expressing L‐selectin was lower in cattle with persistent lymphocytosis than in non‐lymphocytotic or BLV‐free cattle, and inversely correlated with lymphocyte counts. The ratio of B lymphocytes stained for L‐selectin was significantly decreased from 60.2 ± 1.9 % in BLV‐free cattle to 43.8 ± 3.6 and 22.5 ± 5.7 % in non‐lymphocytotic and lymphocytotic cattle, respectively. B‐lymphocytes stained for L‐selectin exhibited about 50 % reduction in L‐selectin expression in BLV‐infected cattle compared with BLV‐free cattle, as judged by the mean fluorescence intensity (MFI). The percentage of L‐selectin‐positive PBMC not bearing surface immunoglobulin M (predominantly T lymphocytes) was comparable in BLV‐free and BLV‐infected cattle. However, L‐selectin expression on T lymphocytes was reduced (about 50 %) in BLV‐infected cattle, as judged by the MFI. We suppose that BLV infection results in a decreased L‐selectin expression on lymphocytes, and accordingly, it may contribute to deregulation of the host immune system.     

谷氨酰胺和牛磺酸对断奶仔猪外周血液免疫功能的影响

试验旨在研究甘氨酰谷氨酰胺和牛磺酸对早期断奶仔猪外周血液免疫功能的影响。选择28日龄断奶杜×长×大仔猪为研究对象,通过饲养试验,研究甘氨酰谷氨酰胺和牛磺酸对早期断奶仔猪外周血液活化T、B淋巴细胞的影响。研究结果表明,日粮中分别或同时添加甘氨酰谷氨酰胺和牛磺酸提高了血液中的T、B淋巴细胞阳性率(P＜0.05)。在早期断奶仔猪日粮中分别或同时添加0.25％甘氨酰谷氨酰胺和0.1％牛磺酸对仔猪T、B淋巴细胞免疫功能有一定作用。