Ultrastructure of the palatine tonsils of the donkey (Equus asinus): New insights by light, scanning, and transmission electron microscopy.

The study aimed to explore the ultrastructure of the donkeys' palatine tonsils. Palatine tonsils of five male donkeys (5 years old) were investigated macroscopically and microscopically. The tonsils appeared as a dome shape with slight elevation and a circular opening on the surface of the oropharynx. The central tonsillar crypt appeared on the medial side of the palate-pharyngeal folds and the floor of the oropharynx. The external surface of the palatine tonsil had different sizes of mucosal folds, some grooves directed to drainage at the tonsillar opening, and the tonsil crypt opening was a crescentic or irregular oval shape. The outer surface was covered by stratified squamous epithelium and modified to be reticular epithelium invaded by lymphocytes in the crypt called lympho-epithelium. The tonsil crypt had aggregated lymphoid nodules, and the cryptal epithelium has surrounded by diffused lymphocytes and hassles corpuscles-like structures. The lymphocytes infiltrated into different layers of the cryptal epithelium and transformed into reticular or lympho-epithelium. The organized lymphoid nodules were primary and secondary, and the secondary ones had a light germinal center. The interfollicular area had many high endothelial venules and blood capillaries. The endothelial venules were lined by simple cuboidal epithelium and had lymphocytes. The blood capillaries had red blood cells and neutrophils. The tonsil was surrounded incompletely by a connective tissue capsule with mucous glands under that capsule. In conclusion, the epithelial lymphocyte infiltration, crypt epithelium, lymphoid nodules, and intra-follicular area of the donkey's palatine tonsils indicate the humoral and cell-mediated immunological process.

Morphological changes in the tongue as a consequence of manganese inhalation in a murine experimental model: light and scanning electron microscopic analysis.

Air pollution by suspended particles has become a worldwide health problem. The main sources of these particles are fossils and additives combustion. Mn enters the body through inhalation, but part of the particles accesses contact with tongue's posterior surface where lingual tonsils and lingual papillae are placed. We decided to explore in a mouse model, the impact that the deposit of inhaled Mn has on the tongue's surface. Atrophy of the lingual tonsil, filiform papillae, as well as the swelling of taste buds in fungiform papillae, were the predominant changes. Ferropenic anemia is associated with the changes described and could be related to the interference of Mn in iron metabolism and riboflavin absorption. More research should be done to explore the participation of suspended particles trapped in the oral cavity in toxicology of Mn or other inhaled pollutants.

Histology, immunohistochemistry and ultrastructure of the bovine palatine tonsil with special emphasis on reticular epithelium.

The paired palatine tonsils, located at the junction of the nasopharynx and oropharynx, are ideally positioned to sample antigens entering through the nasal cavity or oral cavity. Entering antigens will first contact tonsilar epithelium. To better understand the cellular composition of this important epithelial layer, palatine tonsils were collected from six, 7-month-old calves and examined by light microscopy, immunohistochemistry and electron microscopy. Morphometric analysis showed that epithelium overlying lymphoid follicles (reticular epithelium) contained more B-cells, CD4+, CD8+, CD11c+, CD172a+ and gamma/delta TCR+ cells than non-reticular epithelium, with B-cells, CD4+ cells and CD11c+ cells being most numerous. Scanning and transmission electron microscopy of reticular epithelium identified an interrupted basement membrane and vascular elements within the epithelium, as well as cells with characteristics morphologically consistent with cells identified as M-cells in other species. Bovine palatine tonsilar reticular epithelium contains key immune cells, as well as potential M-cell-like cells; elements essential for antigen uptake, antigen processing and initiation of immune responses.

Yak (Bos grunniens) Tonsils: Morphological Description and Expression of IgA and IgG.

This study aimed to describe the morphology, expression of IgA and IgG in adult yak tonsils. The 12 clinically healthy yak tonsils [3- to 6-year old, n = 12] were examined for morphology using light, and transmission electron microscopes. Expression of IgA and IgG was measured by qRT-PCR, ELISA, and immunohistochemistry. The results showed that the palatine tonsil, the tonsil of the soft palate, and the lingual tonsil were oropharyngeal tonsils. The stratified squamous epithelia covering them had a thick underlying layer of connective tissue and their crypts were heavily infiltrated by lymphocytes. The pharyngeal tonsil and the tubal tonsil were nasopharyngeal tonsils. The epithelia of them was predominantly pseudostratified columnar ciliary epithelium, which were loosely arranged with a number of desmosomes or intermediate junctions variably connecting them. The expression levels of IgA and IgG mRNA and protein from high to low was in the pharyngeal tonsil, palatine tonsil, tonsil of the soft palate, lingual tonsil, and tubal tonsil, respectively. Interestingly, the expression of IgG was very significantly higher than that of IgA in yak tonsils (P < 0.01). Both the IgA and IgG ASCs were distributed in the subepithelial areas of the non-reticular crypt epithelium, especially areas of pseudostratified columnar ciliary epithelium, the reticular crypt epithelium, lymphoid follicles, interfollicular areas, and with some of the positive cells aggregating around the glands. The results indicated that the tonsils were not only typical secondary lymphoid organs but also lymphoepithelial structures. IgG could be a significant component of mucosal immune responses in yak tonsils. Anat Rec, 302:999-1009, 2019. © 2018 Wiley Periodicals, Inc.

Hyperplastic lymphoid tissue in HIV/AIDS: an electron microscopic study.

The purpose of this study was to characterize the ultrastructure of lymphoid tissue from HIV/AIDS patients and to evaluate it as a reservoir and source of HIV. HIV has been demonstrated in lymph nodes and tonsils and adenoids, by immunohistochemistry (IHC), in situ hybridization (ISH), and transmission electron microscopy (TEM), to be associated with germinal center (GC) follicular dendritic cells (FDC). The presence of HIV in the larger gastrointestinal tract-associated lymphoid tissue (GALT) has been much less studied. Whether FDC themselves are productively infected by HIV in any of the lymphoid sites is controversial. Lymph nodes, tonsils, and gastrointestinal biopsies were fixed in neutral buffered glutaraldehyde and prepared for TEM. Mature HIV particles were abundant in GC of hyperplastic lymph nodes, tonsils, and the GALT. They were enmeshed within an electron-dense matrix associated with an all-encompassing branching FDC network of processes. HIV particles were seen budding from both FDC and lymphocytes. The greatest numbers of particles were seen in hyperplastic lymphoid tissue from untreated individuals and in lymph nodes co-infected with opportunistic organisms, such as Mycobacterium avium complex. In addition to HIV, unidentifiable "particles" of varying sizes, possibly including other viruses, were regularly seen in association with FDC. Ultrastructural study graphically demonstrated the abundance of HIV particles associated with the complex FDC network of hyperplastic lymph nodes, tonsils, and GALT. HIV was shown to productively infect FDC, as well as lymphocytes.

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.

Detection of genetic alterations by immunoFISH analysis of whole cells extracted from routine biopsy material.

The detection of genetic abnormalities (eg, translocations, amplifications) in paraffin-embedded samples by the fluorescence in situ hybridization (FISH) technique is usually performed on tissue sections. FISH analysis of nuclei extracted from paraffin-embedded samples is also possible, but the technique is not widely used, principally because of the extra labor involved and the loss of information on tissue architecture. In this article, we report that nuclei extracted from paraffin-embedded tissue often retain at least part of the surrounding cytoplasm. Consequently, immunocytochemical labeling for a range of cellular markers (eg, of lineage or proliferation) can be performed in combination with FISH labeling, allowing specific cell populations to be analyzed for genetic abnormalities. These cell preparations are largely free of the problems associated with tissue sections (eg, truncation artifact, signals in different focal planes) so that interpretation is easy and numerical chromosomal abnormalities are readily assessed. Cells isolated from paraffin sections can be stored in suspension so that arrays can be created as and when needed from a range of neoplasms for investigation by the immunoFISH technique (for example, for studying a new genetic abnormality). This procedure represents a novel methodology, which in some settings offers clear advantages over analysis of tissue sections.

Expression of glucocorticoid receptors in non-neoplastic lymphoid follicles and B cell type malignant lymphomas.

OBJECTIVE: To evaluate the expression of human glucocorticoid receptors (hGRs), such as hGR (4H2), hGR-alpha, and hGR-beta, in non-neoplastic lymphoid follicles and B cell type malignant lymphomas. METHODS: The expression of hGRs in non-neoplastic lymphoid follicles and malignant lymphomas, including diffuse large cell lymphoma, mantle cell lymphoma, and follicular lymphoma, was examined immunohistochemically. HGR (4H2) expression was confirmed by double immunostaining of tissues and in isolated cells from tonsillar germinal centres, and by immunoelectronmicroscopy. RESULTS: In secondary lymphoid follicles of any non-neoplastic diseases--such as chronic tonsillitis, reactive lymphadenitis, and Kimura's disease--the germinal centre cells often expressed hGR (4H2) and hGR-alpha. Double immunocytochemical staining of isolated germinal centre cells showed that the majority of hGR (4H2) positive cells were CD20 positive B cells, and that follicular dendritic cells also expressed hGR. Immunoelectronmicroscopy revealed the presence of nuclear hGR (4H2) in the binucleated follicular dendritic cells and germinal centre cells. The frequency of hGR (4H2) expression in diffuse large B cell lymphoma was higher, that in mantle cell lymphoma was lower, and that in follicular lymphoma was intermediate among the types of malignant lymphoma. The hGR (4H2) expression was less frequent in cases of grade I follicular lymphoma. CONCLUSIONS: There are differences in hGR expression between the germinal centre and the mantle zone in non-neoplastic lymphoid follicles, and differences of hGR (4H2) expression among the types of malignant lymphoma and grades of follicular lymphoma, which probably contribute to the different steroid sensitivities.

Neurotrophins and neurotransmitters in human palatine tonsils: an immunohistochemical and RT-PCR analysis.

Lymphoid organs are supplied by many nerve endings associated with different kinds of cells and macrophages. The role of these neuromediators on the release of locally active molecules is still unknown. Here we focused our attention on the expression of some neurotrophins (NTs), their high- and low-affinity receptors and several neurotransmitters in human palatine tonsils. Light and electron microscopy immunohistochemistry showed that human tonsillar samples were positive for all analyzed neurotrophins (NGF, BDNF and NT-3) and their high-affinity receptors (TrkA, TrkB and TrkC, respectively). All of these molecules were strongly expressed in macrophages whereas, in some patients, a weaker specific staining of lymphocytes and blood vessels was also found. The low-affinity receptor for NGF (p75) was always absent in the analysed samples. RT-PCR confirmed the occurrence of specific transcripts for NTs and their high-affinity receptors as well as the absence of mRNA for p75 protein. Also, specific immunoreactivity for neurotransmitters SP, VIP, CGRP, ChAT and nNOS was mainly expressed by macrophagic cells. These results suggest the presence of an extensive network of innervation in the human palatine tonsils which may play a role in the regulation of some immune functions as well as in the modulation of a possible functional scenario of interactions among different immune cellular subtypes.

Morphology of mucosa-associated lymphoid tissue in odontocetes.

This study describes the mucosa-associated lymphoid tissue (MALT) in odontocetes from the Brazilian coast and freshwater systems. Seven species were evaluated and tissue samples were analyzed by light, scanning and transmission electron microscopy, and immunohistochemistry. Laryngeal tonsil was a palpable oval mass located in the larynx, composed of a lymphoepithelial complex. Dense collections of lymphocytes were found in the skin of male fetus and calf. Clusters of lymphoid tissue were found in the uterine cervix of a reproductively active juvenile female and along the pulmonary artery of an adult female. Lymphoid tissues associated with the gastrointestinal tract were characterized by diffusely arranged or organized lymphocytes. The anal tonsil was composed of an aggregate of lymphoid tissue occurring exclusively in the anal canal, being composed of squamous epithelium branches. MALT was present in different tissues and organic systems of cetaceans, providing constant protection against mucosal pathogens present in their environment.

Ex vivo modeling of oral HIV transmission in human palatine tonsil.

The majority of newly acquired HIV infections are believed to occur following transmission of virus infectivity across mucosal surfaces, although many mechanistic details still remain unresolved. We have used human ex vivo organ cultures and primary cell populations to analyze the cellular and molecular basis for mucosal HIV transmission. By using human palatine tonsil from routine tonsillectomies and semen from HIV-positive donors, we have created an experimental equivalent to oral HIV transmission. HIV infection was readily transferred into tonsillar lymphocytes, but this transmission into lymphocytes was dramatically reduced when the exposed lymphocyte populations were protected by intact mucosal surfaces. In this study, we consider the impact that leukocyte activation and morphological aberrations in surface structure may have on susceptibility to primary HIV infection and introduce novel time-lapse confocal microscopy procedures that begin to reveal the dynamic complexity associated with cell-mediated HIV transmission.

Ultrastructure of the human palatine tonsil and its functional significance.

The human palatine tonsils represent a mucosa-associated lymphoid tissue with a significant function in mucosal protection against alimentary and airborne pathogens. The ultrastructure of different morphological compartments in the human palatine tonsil was studied in eighteen tonsils obtained from the patients who had undergone elective tonsillectomy due to chronic tonsillitis. The tonsillar specimens were analyzed by scanning and transmission electron microscopy. The results showed the presence of tight junctions between superficial epithelial cells of the oropharyngeal tonsillar surface. The crypt epithelium is a sponge-like structure infiltrated by non-epithelial cells, mostly lymphocytes, and is characterized by the presence of small pores - microcrypts occupied by large microvillus cells and/or lymphocytes. Antigen-presenting Langerhans cells with typical intracytoplasmic Birbeck granules were also found in the crypt epithelium. The lymphoid follicles are composed of lymphocytes and two types of non-lymphoid follicular cells: small fibroblast-like cells and large cells, morphologically consistent with antigen-bearing follicular dendritic cells or macrophages. The interfollicular areas consisted of a dense network of reticular cells and reticular fibers; many lymphocytes were interspersed between the reticular fibers. In addition to arterioles and high endothelial venules in the interfollicular lymphoid tissue, some fenestrated capillaries were seen intraepithelially and subepithelially. The complex ultrastructure of the human palatine tonsil provides a microenvironment necessary for antigen uptake, antigen processing and immune response.

C1q binding to mitochondria: a possible artefact?

Mitochondrial preparations, obtained from human tonsils and from rat spleen, liver, heart, and kidney tissues, bound [125I]C1q with affinities of 10(7)-10(8)M-1. The binding of C1q was not affected by treatment of the mitochondrial preparations with pronase, trypsin, or phospholipase D, but it was lowered 5-6-fold following treatment of the mitochondria with DNase and RNase. Binding of C1q to mitochondrial preparations was also greatly diminished by limited chemical modification of C1q with cyclohexane-1,2-dione. It is suggested that the reported binding of C1q to mitochondria may have arisen from the protein binding to DNA and/or RNA contaminating the mitochondrial preparations.

Ultrastructural demonstration of immunoglobulin in glutaraldehyde fixed resin embedded human tonsil by an indirect immunoperoxidase method.

A post-embedding method has been established for demonstrating intracellular immunoglobulin using an immunocytochemical technique on material which has been fixed to glutaraldehyde and resin embedded. This allows localisation at both a light and ultrastructural level with good preservation of tissue structure.

The intracellular localisation of immunoglobulin in human lymphoid cells and haematopoietic cell lines by immunoperoxidase electron microscopy.

A technique is described for the ultrastructural localisation of intracellular immunoglobulin (Ig) in human lymphoid cell suspensions by the immunoperoxidase method. The technique involves restricted saponin digestion of glutaraldehyde prefixed cells to enhance conjugate penetration. With this method of staining Ig was located in the rough endoplasmic reticulum, perinuclear space and Golgi apparatus in human lymphoid cells from a variety of sources, which is consistent with previously published observations using other ultrastructural techniques. In contrast, diffuse intracytoplasmic staining was predominant in cells prefixed with glutaraldehyde but not treated with saponin. These differences in patterns are discussed in terms of membrane permeability. Although saponin treatment was necessary for consistent localisation of intracellular Ig it resulted in unavoidable loss of Ig from the surface of the cells.

Isolation of follicular dendritic cells from human tonsils and adenoids. I. Procedure and morphological characterization.

Follicular dendritic cells have been isolated from human tonsils and adenoids and characterized at the ultrastructural level. Follicles were dissected and digested with different hydrolytic enzymes. The cells were separated by sedimentation at unit gravity. By this procedure we obtained follicular dendritic cells enveloping lymphocytes with their cytoplasmic extensions in a way analogous to that described for isolated thymic nurse cells. The ultrastructural features of isolated follicular dendritic cells are similar to those observed in situ. Prolonged enzymatic action caused loss of the enveloped lymphocytes.

The localisation of intracellular immunoglobulin and alpha-1-antitrypsin by immunoelectron staining of post-osmicated, resin-embedded tissue.

An indirect immunoperoxidase method is described to demonstrate intracellular immunoglobulins and alpha-1-antitrypsin in semithin and ultrathin sections from human tissue. The tissue was primarily fixed in glutaraldehyde, post-osmicated and resin-embedded.

Multifocal verruciform xanthoma of the upper aerodigestive tract in a child with a systemic lipid storage disease.

We report the clinical, light-microscopic, and ultrastructural features of a case of multifocal verruciform xanthoma in the upper aerodigestive tract of a child with a systemic lipid disorder. Lipid storage cells were found in liver, bone marrow, and as a component of verruciform xanthomas. To our knowledge this represents the first case of verruciform xanthoma reported in (a) a child, (b) as a multifocal lesion in the upper aerodigestive tract, (c) associated with a systemic lipid disorder, and (d) with ultrastructural evidence of lipid accumulation within endothelial cells. Although this patient presented with lesions involving the tongue and larnyx, subsequently lesions were found in the bone marrow and liver. Two months later more lesions were discovered on the epiglottis, posteior tongue, right glottis, and in grossly normal peritonsillar mucosa. Six months later a new oral lesion developed. Based upon these observations, we speculate that the pathogenesis of verruciform xanthoma involves accumulation of excess lipid in subepithelial sites which is scavenged by macrophages. Lipid-laden macrophages release epithelial growth factors that lead to epithelial hyperplasia. Depending on the degree of epithelial hyperplasia, the gross appearance of verruciform xanthomas may be flat, sessile, papillary, or verrucous.

Immunoelectron microscopic labeling of immunoglobulin in plasma cells after osmium fixation and epoxy embedding.

Previous studies have found that immunoglobulin cannot be immunolabeled in tissues prepared for electron microscopy by usual methods. To test this conclusion, we used a protein A-gold postembedding immunolabeling method on tissues that were fixed in glutaraldehyde, post-fixed in osmium tetroxide, and embedded in epoxy resin; sections were pretreated with sodium metaperiodate. A variety of common fixation protocols were also used and the most suitable conditions for immunolabeling were determined. This technique permitted the ultrastructural localization of immunoglobulin light chains in optimally preserved and contrasted plasma cells from human tonsil, lymph nodes, plasmacytomas, and a renal biopsy. We were able to demonstrate multiple antigens in the same tissue and label antigens in tissues that had been stored for many years in epoxy resin. The technique allows quantitation of the gold label over plasma cell organelles and therefore gives information about the immunoglobulin secretory pathway in these cells. We found that the protein A-gold procedure compares favorably in technical ease with the immunoperoxidase, avidin-biotin peroxidase, and immunoglobulin-colloidal gold immunolabeling methods, and has added advantages in allowing precise localization and quantitation of the labeled antigen.

Bauhinia purpurea lectin (BPA) binding spectra in hyperplastic human tonsil and in peripheral blood: immunohistochemical, immunoelectron microscopic, and flow cytometric analyses.

We studied the binding of Bauhinia purpurea lectin (BPA) to human tonsils and peripheral blood mononuclear cells by immunohistochemical, immunoelectron microscopic, and flow cytometric techniques. The effect of several fixatives (acetone, ethanol, buffered and non-buffered formalin, B5, Bodian 11, Bouin's, Carnoy's, Zenker's, and glutaraldehyde) was also examined. BPA was reactive with germinal center lymphocytes, macrophages/histiocytes, follicular dendritic cells, squamous epithelial cells, and a subset of endothelial cells. Mantle zone and paracortical lymphocytes were non-reactive with BPA. The profile of the specific binding characteristic of BPA lectin was found to be influenced by the fixatives. Most significantly, formalin fixation greatly reduced overall binding intensity, particularly making germinal center lymphocytes totally non-reactive. The reaction intensity was most prominent in frozen sections or those fixed in Carnoy's or ethanol solution. The combination of heavy metal salt-containing fixatives with acetic acid usually did not enhance BPA binding. Glutaraldehyde solution used for immunoelectron microscopic study also preserved BPA receptors fairly well, and BPA was confined to the membrane in lymphocytes and to both the membrane and cytoplasm in macrophages/histiocytes and follicular dendritic cells. Neuraminidase treatment of tissues resulted in binding of BPA to lymphocytes that were non-reactive before treatment. Double-staining studies on cell suspensions from tonsils with FITC-BPA and PE-conjugated anti-CD3 or SIg reagents revealed that 28.4% of CD3+ cells and 61.3% or SIg+ cells were BPA reactive. In PBL, 65.6% and 81.4% of CD3+ and SIg+ cells, respectively, were BPA reactive. Neuraminidase treatment also increased the percentage of BPA-reactive lymphocytes. In conclusion, BPA is a marker for macrophages/histiocytes and germinal center lymphocytes, provided that the tissues are unfixed or fixed with suitable fixatives such as ethanol or Carnoy's solution.