Histological characteristics and stereological volume assessment of the ovine tonsils

Tonsils form a first line of defence against foreign antigens and therefore play a key role in immunity. Since documented information about ovine tonsils is limited, a study was performed in which the morphological characteristics and the volume of lymphoid tissue present in each ovine tonsil were determined. The tonsils of five adult healthy sheep were examined histologically and the volumes were estimated using the Cavalieri method. The pharyngeal tonsil had a mean volume of 1296.1 ± 205.9 mm³ and was by far the largest ovine tonsil, followed by the paired palatine tonsil with a mean volume of 715.0 ± 110.5 mm³. The tonsil of the soft palate, the paired tubal and paraepiglottic tonsils and the lingual tonsil were much smaller with a mean volume of, respectively, 90.3 ± 24.9 mm³, 80.1 ± 24.3 mm³, 29.7 ± 11.8 mm³ and 10.1 ± 2.8 mm³. The folds and crypts of the pharyngeal and palatine tonsils were covered by a reticular and a non-reticular epithelium. Both tonsils were mainly composed of primary and secondary lymph follicles. The palatine tonsils contained 1–3 crypts with a few secondary infoldings. Lymphoid tissue in the tonsil of the soft palate was located at the nasopharyngeal (dorsal) side of the soft palate. The tubal tonsil was lined with a pseudostratified columnar ciliated epithelium and consisted of scattered lymphoid cells and lymph follicles. The paraepiglottic tonsil consisted of lymph follicles and aggregated lymphoid cells. Its overlying keratinized stratified squamous epithelium was folded and often heavily infiltrated by lymphocytes. The ovine lingual tonsil was not macroscopically visible and did not contain clearly distinguishable lymph follicles. It consisted of aggregations of lymphoid cells that were mainly located within the vallate lingual papillae.     

山羊咽扁桃体和咽鼓管扁桃体的组织结构观察

选取健康10月龄奶山羊10头,断头宰杀后取咽扁桃体和咽鼓管扁桃体,应用组织学光镜和电镜制片技术研究咽扁桃体和咽鼓管扁桃体的显微和亚显微组织结构.结果表明:山羊咽扁桃体和咽鼓管扁桃体的黏膜上皮主要由2～3层多边形上皮细胞组成,部分区域只有单层扁平细胞,相邻上皮细胞间空隙很大,上皮细胞表面有丰富的微绒毛.上皮细胞之间和黏膜上皮下方固有层内有大量淋巴细胞浸润.扁桃体的实质部分由数个次级淋巴小结和弥散淋巴组织构成,弥散淋巴组织中有大量分布的淋巴管和毛细血管后微静脉.此外,在紧贴黏膜上皮细胞下方的固有层和淋巴滤泡中可观察到少量的树突状细胞.结果提示山羊的咽扁桃体和咽鼓管扁桃体可作为鼻腔免疫的主要诱导位点和效应部位.     

Bovine lingual tonsil: histomorphological characteristics with special reference to the follicular dendritic cells

In the bovine, the lingual tonsil is localized at the lingual radix. However, as the expansion of the tonsillar tissue was presumed to be much larger than the macroscopically visible part, selected specimens from the bovine tongue were examined to clarify if lymphatic tissue of the lingual tonsil was present rostral of a defined macroscopical landmark, i.e. rostral of the most caudal of the papillae vallatae. The identification of the lymphoreticular system was validated by means of the detection of reticulum cells, especially follicular dendritic cells (FDC). They characterize the tonsillar lymph nodules and distinguish them structurally from unspecific accumulations of lymphocytes. In our study, two monoclonal antibodies, i.e. CNA.42 and D46, detected bovine FDC cells in situ. 'Organized' lymphoid tissue--i.e. solitary lymph nodules and aggregations of lymph nodules, both either with or without surrounding connective tissue, and with or without crypts, but with immunopositive reactions for FDC--could be observed up to 30 mm rostral of the most caudal of the papillae vallatae. As these formations were not identified macroscopically, they shall be referred to as the disseminate part of the lingual tonsil.     

猪呼吸道淋巴组织发育的亚显微结构变化

应用组织学和电镜技术研究猪呼吸道发育过程中淋巴组织的变化。结果表明：扁桃体和咽部是呼吸道进入机体的第一个淋巴组织集中的部位，弥散淋巴组织在出生时就存在，淋巴小结不明显；20日龄时扁桃体中淋巴组织增生，淋巴小结清晰可见；120日龄淋巴小结数量增加，紧靠鳞状上皮密集排列，淋巴小结发育很好，并出现生发中心。扁桃体复层鳞状上皮中含有大量的上皮内淋巴细胞。气管叉是呼吸道进入机体的第二个淋巴组织集中的部位，出生时气管叉外膜中淋巴组织直接与气管支气管淋巴结相连，淋巴组织明显可见。20日龄时气管叉外膜中淋巴组织已分开，形成气管叉外膜密集的淋巴组织和气管支气管淋巴结两个部分。120日龄时气管叉处淋巴组织特别发达，黏膜上皮中上皮内淋巴细胞数量也显著增加。肺内气管和细支气管固有膜中均有较多的淋巴细胞，其中浆细胞数量增加，上皮中仍存在少量的上皮内淋巴细胞。本试验结果提示猪呼吸道是黏膜免疫较理想的诱导位点和效应位点，新生仔猪通过鼻腔免疫可提高呼吸道局部黏膜免疫力。     

Anatomical and histological aspects of the bovine lingual tonsil

The localization of the bovine lingual tonsil is described as a prerequisite for the removal of specified risk material from the tongue meat in order to restrict the risk arising from bovine spongiform encephalopathy (BSE) to public health. The major part of this tonsil can be located macroscopically by the openings of its follicular crypts at the root of tongue. This part consists of organized aggregations of lymph nodules. Additional solitary primary lymph nodules and diffuse accumulations of lymphocytes are macroscopically invisible but are bilaterally present in the area extending 2 cm caudal to 3 cm rostral to the last vallate papillae. By sectioning the tongue 3 cm rostral to the last vallate papillae, undermining the lingual mucosa to the level of these papillae and making a transverse cut towards the lingual process of the basihyoid bone, the greater part of the lingual tonsil can efficiently be removed. Finally, immunohistochemical staining demonstrated the presence of T and B lymphocytes, suggesting that the bovine lingual tonsil can be considered as a site where an immune response can be induced.     

Differences between the ovine tonsils based on an immunohistochemical quantification of the lymphocyte subpopulations

In sheep, the pharyngeal first defence line against oral and inhaled antigens is organized in six tonsils. Since tonsils are regarded as secondary lymphoid tissue and part of the acquired immune system which is subjected to induction through contact with antigens, an evaluation of the different lymphocyte populations in tonsils is useful to determine a tendency of the specific tonsils to more inductive or more effective immunity. By means of immunohistochemistry, different lymphocyte populations were quantified and localized using a panel of eight antibodies, i.e. anti-CD45, anti-CD21, anti-CD2, anti-CD3, anti-CD4, anti-CD8, anti-WC1 and anti-Ki67. The CD21+ B lymphocytes were localized within the tonsillar lymphoid follicles. The CD2+/CD3+ T lymphocytes were numerous in the interfollicular regions and were aligned underneath and within the epithelium but were also observed at the CD21+ pole of the lymphoid follicles. Near the lingual and tubal tonsils, and the tonsil of the soft palate, the CD45+ cells around the seromucous glands and in the lamina propria were mainly CD3+ T cells. In all tonsils, the WC1+ gamma delta T cells formed a small lymphocyte population which harboured the lamina propria and the interfollicular region. The relative percentages of the different lymphocyte populations of the large palatine and pharyngeal tonsils, which are macroscopically the most developed, were comparable. In contrast, the lingual tonsil was significantly different from the other tonsils not only by its small size and lack of lymphoid follicles, but also by the lymphocyte populations. Based on the lymphocyte populations, the ovine tonsils can be divided in three groups with the tonsil of the soft palate, the tubal and paraepiglottic tonsil forming an intermediate between the palatine and pharyngeal tonsils as true tonsils on the one side, and the lingual tonsil as a scattered lymphocyte aggregation on the other side.     

The microanatomy of the palatine tonsils of the buffalo (Bos bubalus)

The palatine tonsils play a key role in initiating immune responses against antigens entering the body through the mouth. They are also replication sites of some pathogens. There is no data available about the structure of the palatine tonsils of the Egyptian water buffalo. Therefore, palatine tonsils of 14 clinically healthy buffalo bulls (2–3 years old) were examined macroscopically and microscopically using light, and transmission electron microscopes. The tonsils had an elongated kidney shape with a central invagination (tonsillar fossa) containing a single macroscopic opening leading to a small central cavity (tonsillar sinus). A number of macroscopic crypts originated from this sinus (internal crypts). Besides the tonsillar fossa, also small macroscopic crypts (external crypts) were present. The tonsils were enclosed by a thin connective tissue capsule and septa divided the tonsils into incomplete lobes. Within these encapsulated organs mucous glands were very obvious. Each crypt was highly branched and lined with stratified squamous non-keratinized epithelium. Several lymphoid cells infiltrated between the epithelial cells forming patches of lymphoepithelium. The crypt lumen contained lymphocytes, neutrophils and erythrocytes. Lymph nodules with clear germinal centers extended under the epithelial surface. Diffusely distributed lymphocytes were found in the narrow interfollicular region. High endothelial venules, interdigitating dendritic cells, macrophages and plasma cells were observed among the diffuse lymphocytes. Lymphatics filled with lymphocytes drained the tonsils.     

Shortening of the bovine tongue according to regulation (EC) 999/2001 is not complying with the current legal definition of specified risk material - a macroscopical and histological preliminary study

The full elimination of all specified risk material (SRM) in food of animal origin is crucial for consumer protection and is of high priority in inner EU trade. Among other tissues, the tonsils of cattle are considered as SRM. The aim of this study was to evaluate whether the 'cut at the back of the tongue just before the tongue bones' required by EC regulation is sufficient to remove tonsils and lymphatic tissue completely. Eight skulls from cattle were collected for the simulation of a vertical cut according to the EC regulation and the detection of the target at the back of the tongue. Further, specimens of the lingual mucosa were cut out from two tongues and examined microscopically. The most caudal of these specimens was from the macroscopically visible part of the lingual tonsil. The most rostral specimen contained the most caudal Papilla vallata. Simulation of the obligatory ventro-dorsal cut yielded hits at varying locations on the dorsal surface of the tongue, sometimes including tissue of the lingual tonsil. Histological examination of the lingual mucosa gave clear evidence that lymphatic tissue resembling the tissue of a tonsil in terms of its histological organization and infiltration of the mucosal epithelium could even be found in areas with no macroscopically visible lingual tonsils.     

Distribution of the lingual lymphoid tissue in domestic ruminants

The distribution and organisation of the intralingual lymphoid tissue was studied in sheep, goat and cattle. For each species, the tongues of two animals were harvested and divided in sample blocks extending over the total surface of the tongue. With 2.5 mm intervals, ten serial histological sections were made for conventional histological staining (haematoxylin-eosin, Van Gieson, Masson's trichrome) and immunohistochemical staining of lymphoid cells (anti-CD3, anti-CD21, anti-CD45). Lymphocytes were scattered in the subepithelial propria-submucosa and in the connective tissue cores of the lingual papillae. The connective tissue cores of fungiform papillae, including those located on the lingual apex, and vallate papillae showed relatively more lymphocytes than the propria-submucosa. Lymphoid cell aggregations were even more abundant beneath the grooves surrounding the vallate papillae in small ruminants. In cattle, a well-organised lingual tonsil was additionally found at the root of the tongue. CD3-positive lymphocytes were observed in all species examined. CD21-positive lymphocytes were numerous in the lymphoid nodules of the bovine lingual tonsil but very scarce in the ovine and caprine tongues. Therefore, the lymphoid cell aggregations in the tongues of small ruminants should not be referred to by the term 'lingual tonsil'.     

Shortening of the Bovine Tongue According to Regulation (EC) 999/2001 is not Complying with the Current Legal Definition of Specified Risk Material – a Macroscopical and Histological Preliminary Study

The full elimination of all specified risk material (SRM) in food of animal origin is crucial for consumer protection and is of high priority in inner EU trade. Among other tissues, the tonsils of cattle are considered as SRM. The aim of this study was to evaluate whether the ‘cut at the back of the tongue just before the tongue bones’ required by EC regulation is sufficient to remove tonsils and lymphatic tissue completely. Eight skulls from cattle were collected for the simulation of a vertical cut according to the EC regulation and the detection of the target at the back of the tongue. Further, specimens of the lingual mucosa were cut out from two tongues and examined microscopically. The most caudal of these specimens was from the macroscopically visible part of the lingual tonsil. The most rostral specimen contained the most caudal Papilla vallata. Simulation of the obligatory ventro‐dorsal cut yielded hits at varying locations on the dorsal surface of the tongue, sometimes including tissue of the lingual tonsil. Histological examination of the lingual mucosa gave clear evidence that lymphatic tissue resembling the tissue of a tonsil in terms of its histological organization and infiltration of the mucosal epithelium could even be found in areas with no macroscopically visible lingual tonsils.     

日粮中添加铁对绵羊免疫器官组织结构和铁含量的影响

为研究日粮中不同铁水平对绵羊免疫功能的影响，探究绵羊饲养过程中铁过量的风险，试验在绵羊基础日粮中分别添加铁500、1 000、1 500 mg/kg（硫酸亚铁形式），饲养75 d后宰杀，采取绵羊免疫器官（脾脏、胸腺、肝门淋巴结、十二指肠淋巴结及腭扁桃体），应用传统病理学方法观察以上器官的组织结构变化、含铁血黄素分布，并测定组织铁含量的变化情况。结果显示，绵羊饲喂过量铁后，脾脏白髓占比减小，淋巴细胞数量减少、排列疏松，伴有肿大、空泡化现象，红髓内有大量含铁血黄素沉积；胸腺皮质区变薄，髓质区变厚且融合，胸腺小体数量减少、结构模糊；肝门淋巴结、十二指肠淋巴结皮质与髓质区变薄、界限模糊，皮质淋巴窦与髓窦间隙增大，淋巴小结结构模糊；腭扁桃体实质内淋巴小结数量减少，毛细血管后微静脉有充血现象。饲喂过量铁后，各免疫器官组织铁含量均升高，其中脾脏内铁含量最多，腭扁桃体内最少，铁低、中、高剂量组脾脏铁含量极显著高于对照组（P<0.01）。结果表明，饲喂过量铁后，绵羊免疫器官组织结构均遭到不同程度破坏，且随着铁添加量越多，组织结构破坏程度越明显；各免疫器官中铁蓄积量与日粮中铁添加水平呈正相关，摄入过量铁严重影响绵羊免疫功能，因此在绵羊饲养过程中，要严格遵守美国NRC（1985）饲料标准，日粮中铁水平不要超过500 mg/kg。     

Differential expression of homing receptors and vascular addressins in tonsils and draining lymph nodes: Effect of Brucella infection in sheep

The differential expression of homing receptors (HR) and complementary vascular addressins was studied in T and B lymphocytes from ovine tonsils and draining lymph nodes (LN) in uninfected and Brucella melitensis-infected sheep. In uninfected sheep, CD4+CD25+ T cells expressed proportionally more L-selectin and beta1 integrin than beta7 integrin in pharyngeal and palatine tonsils and in parotid LN (PLN), retropharyngeal LN (RLN) and the peripheral prescapular LN (PSLN). In contrast, memory CD4+CD45RA- T cells expressed an equivalent proportion of the three HR in PLN and PSLN, whereas beta1 and beta7 integrins were proportionally more expressed than L-selectin in pharyngeal tonsil. beta7 integrin was proportionally more expressed than beta1 integrin or L-selectin in palatine tonsils, RLN and the mucosal mesenteric LN (MLN). beta1 integrin was proportionally more expressed in IgG+ and IgA+ cells than beta7 integrin and L-selectin in tonsils, PLN and RLN. The main endothelial addressin expressed on venules in both pharyngeal and palatine tonsils, the PLN and RLN, as well as in the PSLN, was the peripheral PNAd, while in the MLN it was MAdCAM-1. Conjunctival infection by Brucella resulted in an increase of CD4+CD25+ and CD4+CD45RA- T cell subsets, which was associated to modifications of HR expression. CD4+CD45RA- T cells expressed proportionally more beta1 and beta7 integrins than L-selectin in regional PLN and RLN, but also in PSLN. The infection induced an increase of IgG+ and IgA+ cell percentages expressing beta1 integrin in all LN, and also beta7 integrin in the RLN. PNAd continued to be expressed on venules of tonsils and draining LN after Brucella infection, and MAdCAM-1 was also weakly expressed on RLN venules. These results suggest that lymphocyte trafficking through tonsils and draining LN could involve L-selectin/PNAd interactions, as well as beta1 or beta7 integrin, possibly in interaction with VCAM-1 or MAdCAM-1. The homing of antigen-specific lymphocytes in these tissues could be modulated after conjunctival infection with Brucella, which induces the recruitment of lymphocytes that express both beta1 and/or beta7 integrin in regional and more distant LN.     

Stereological assessment of the epithelial surface area of the ovine palatine and pharyngeal tonsils

Although ovine tonsils play a major role in prion diseases, their morphology is poorly documented and morphometric data are sparse. Therefore, a stereological assessment of the surface area of the palatine and pharyngeal tonsils of five sheep was performed. The epithelial surface area of both tonsils is considerably increased by the presence of tonsillar crypts and folds. The mean epithelial surface area of the paired palatine tonsil was 7.14 cm(2) and that of the pharyngeal tonsil was 23.5 cm(2). In comparison with the paired palatine tonsil, the volume of the pharyngeal tonsil was almost two times larger, whilst its surface area was more than three times larger.     

Pathogenesis of experimental bovine spongiform encephalopathy: preclinical infectivity in tonsil and observations on the distribution of lingual tonsil in slaughtered cattle

The infectivity in tissues from cattle exposed orally to the agent of BSE was assayed by the intracerebral inoculation of cattle. In addition to the infectivity in the central nervous system and distal ileum at stages of pathogenesis previously indicated by mouse bioassay, traces of infectivity were found in the palatine tonsil of cattle killed 10 months after exposure. Because the infectivity may therefore be present throughout the tonsils in cattle infected with BSE, observations were made of the anatomical and histological distribution of lingual tonsil in the root of the tongue of cattle. Examinations of tongues derived from abattoirs in Britain and intended for human consumption showed that macroscopically identifiable tonsillar tissue was present in more than 75 per cent of them, and even in the tongues in which no visible tonsillar tissue remained, histological examination revealed lymphoid tissue in more than 90 per cent. Variations in the distribution of the lingual tonsil suggested that even after the most rigorous trimming of the root of the tongue, traces of tonsillar tissue may remain.     

Histology, immunohistochemistry and ultrastructure of the equine tubal tonsil

The tubal tonsil of the horse surrounds the pharyngeal opening of the eustachian tube and is lined by pseudostratified columnar ciliated epithelium interspersed with areas of follicle-associated epithelium (FAE) heavily infiltrated by lymphocytes but devoid of goblet and ciliated cells. Scanning and transmission electron microscopy revealed microvillous cells and cells with features characteristic of M cells such as reduced microvilli or depressed bare surface, more numerous mitochondria, small vesicles and lysosomes, as well as vimentin filaments and epitopes specific for GS 1-B4 as previously seen in the nasopharyngeal tonsil. M cells were also identified in areas of respiratory epithelium not associated with lymphoid follicles and appeared to be the nasal mucosal counterparts of recently described intestinal villous M cells in the mouse. The underlying lymphoid tissue of the FAE was generally organized as solitary lymphoid follicles without germinal centres in contrast to the diffuse and large amount of organized lymphoid follicles with germinal centres that characterize the nasopharyngeal tonsil. CD8+ T and B-lymphocytes were much fewer than in the nasopharyngeal tonsil. High endothelial venules were mainly oriented towards the parafollicular area and contained much fewer endothelial pores and vesiculo-vacuolar organelles. Finally, scattered small clusters of mucus acini and striated muscles were other features that differentiated the tubal and nasopharyngeal tonsils.     

Ultrasound-guided percutaneous lung biopsy in sheep

Lung biopsies were taken through the ninth intercostal space on the two sides of the thorax of 10 clinically healthy sheep, using a Tru-Cut needle on one side and the Bard Biopty-System on the other. Nine of the 10 sheep remained clinically healthy, but one coughed transiently and had a mild bloody discharge from the right nostril immediately after the biopsy. The sheep were slaughtered 10 days later and the lungs and pleura were examined macroscopically; there were either no lesions or only small scars visible at the sites of the biopsies. However, well-developed subpleural nodules due to parasites were observed in some of the lungs. There were no adhesions between the costal and pulmonary pleurae. Of the 20 biopsy specimens, 18 were ideal for histological examination, and none of them was histologically normal. Mild interstitial pneumonia was diagnosed in 15 specimens, chronic bronchiolitis in nine specimens, and hyperplasia of smooth muscle or connective tissue in 17.     

Perinatal development of lymphoid tissue and its associated epithelium in the ovine pharyngeal tonsil: A morphological study

The perinatal development of lymphoid tissue and its associated epithelium in the pharyngeal tonsil of sheep was investigated by light and electron microscopy. The lymphoid cells first appeared in the subepithelium in a scattered form at about 92 days gestation. These cells proliferated rapidly during the last trimester of gestation, and had formed many dense aggregates at the time of parturition. At birth, the epithelium overlying the aggregates was extensively infiltrated with lymphocytes and showed early transformation of follicle-associated epithelium. The lymphoid tissue and its associated epithelium did not, however., fully develop until after birth, when well-differentiated follicle-associated epithelium and lymphoid follicles with vigorous lymphopoiesis were present. In l–2 week old lambs, these structures were ultrastructurally similar to those seen in adult sheep although their size was smaller. The results of this study suggest that the lymphoid tissue of the ovine pharyngeal tonsil and its associated epithelium are morphologically ready to cope with antigens in the extra-uterine environment at birth, but that their full development and maturation are dependent on postnatal antigen stimulation.     

Mastitis associated with ovine progressive pneumonia virus infection in sheep

Eighteen ewes were experimentally infected with ovine progressive pneumonia virus and their mammary glands were examined for lesions and virus at 2.5 to 10 years postinoculation. Lesions were seen in 14 of 18 sheep; virus was isolated from 4 of 8 sheep. Lesion consisted of an interstitial accumulation of lymphocytes with periductal lymphoid nodules, and epithelial vacuolation and necrosis at the site of the lymphoid nodules.     

Perinatal development of lymphoid tissue and its associated epithelium in the ovine pharyngeal tonsil: a morphological study

The perinatal development of lymphoid tissue and its associated epithelium in the pharyngeal tonsil of sheep was investigated by light and electron microscopy. The lymphoid cells first appeared in the subepithelium in a scattered form at about 92 days gestation. These cells proliferated rapidly during the last trimester of gestation, and had formed many dense aggregates at the time of parturition. At birth, the epithelium overlying the aggregates was extensively infiltrated with lymphocytes and showed early transformation of follicle-associated epithelium. The lymphoid tissue and its associated epithelium did not, however, fully develop until after birth, when well-differentiated follicle-associated epithelium and lymphoid follicles with vigorous lymphopoiesis were present. In 1-2 week old lambs, these structures were ultrastructurally similar to those seen in adult sheep although their size was smaller. The results of this study suggest that the lymphoid tissue of the ovine pharyngeal tonsil and its associated epithelium are morphologically ready to cope with antigens in the extra-uterine environment at birth, but that their full development and maturation are dependent postnatal antigen stimulation.