Morphology,cytochemical staining and ultrastructural characteristics of reindeer (Rangifer tarandus) leukocytes

Peripheral blood smears from four adult reindeer (Rangifer tarandus) were examined after staining with Romanowsky's stain and cytochemical stains, including alpha-napthyl butyrate esterase (alpha-NBE), Sudan black B (SBB), chloroacetate esterase (CAE) and alkaline phosphatase (ALP). Romanowsky-stained eosinophils, neutrophils, lymphocytes and monocytes resembled those of cattle, sheep and goats. Basophils had two different staining patterns with Romanowsky's stain. Basophils that we termed "grey basophils" were similar in appearance to grey eosinophils in Greyhound dogs, with medium blue-grey to lavender-grey cytoplasm containing varying numbers of clear vacuoles or granules and variable numbers of small, intensely basophilic, perinuclear granules. The second basophil staining pattern was more typical of ruminant basophils, with uniform, pale to dark basophilic cytoplasmic granules. Basophils stained positive for alpha-NBE, SBB, CAE, and ALP. Eosinophils stained positive for SBB, and were negative for alpha-NBE, CAE, and ALP. Neutrophils were negative for SBB, CAE, and ALP. Monocytes stained positive for alpha-NBE, were rarely positive for CAE and SBB, and were negative for ALP. Transmission electron microscopy revealed matrix within all granulocytes granules, including those of basophils.     

Detection of retinoid receptors in non-neoplastic canine lymph nodes and in lymphoma

This study evaluated the difference in retinoid receptor expression between non-neoplastic lymph nodes and nodal lymphoma in dogs. Retinoid receptor expression was evaluated by immunohistochemistry in 32 canine lymph nodes. The lymph nodes had been previously diagnosed as non-neoplastic (6 normal and 7 hyperplastic lymph nodes) and B- and T-cell lymphoma (19 cases). Immunohistochemistry for retinoic acid receptors and retinoid-X receptors (and their subtypes α, β, and γ) was performed in all cases. In addition, immunohistochemistry for CD3 and CD79a was performed in all lymphoma cases. Non-neoplastic lymphocytes were negative for all retinoid receptors. Retinoic acid receptor-γ was detected in 100% of B-cell lymphoma and 78% of T-cell lymphoma, while retinoid X receptor-γ was positive in 78% of T-cell lymphoma cases. When normal lymph node architecture was still present, a contrast between retinoid-negative benign cells and retinoid-positive malignant cells was clear. Retinoid receptors were expressed in neoplastic, but not in benign lymphocytes, suggesting their value for both diagnosis and treatment of canine lymphoma.     

Lymphocyte 5'-nucleotidase activities in normal dogs and in dogs with malignant lymphoma

Plasma membrane 5'-nucleotidase (5'NT) activity was assayed in lymph node lymphocytes from seven normal control dogs and in malignant lymphocytes from 25 dogs with lymphoma. The lymphoid tumors were classified according to the NCI Working Formulation into five histologic subtypes. When compared with normal controls significantly lower 5'NT activities were found in the lymphoblastic, small lymphocytic and diffuse large (noncleaved) subtypes while no significant differences were observed in the immunoblastic or small noncleaved groups. In addition, dogs with hypercalcemia or paraproteinemia had decreased 5'NT levels. However, no significant differences were found between prognostic groups or histologic subtypes of the NCI classification. In conclusion, canine lymphoma subtypes had generally decreased 5'NT activities which appeared to reflect the B or T cell lineage, degree of maturation and enzyme activity of the cell of origin.     

Morphologic characterization of specific granules in Greyhound eosinophils

BACKGROUND: "Vacuolated" eosinophils (ie, eosinophils with empty, nonstaining granules) have been described previously in normal Greyhounds. However, to our knowledge, detailed studies of granules in vacuolated and normal eosinophils in this breed have not been performed. OBJECTIVE: The objective of this prospective study was to characterize some of the morphologic, ultrastructural, and cytochemical staining features of specific (primary) granules in both normal and vacuolated eosinophils in Greyhound blood. METHODS: Morphologic features of eosinophils in Wright's- and Diff-Quik-stained peripheral blood smears from 49 Greyhounds were compared with 200 blood smears from non-Greyhound dogs. Transmission electron microscopy was done on blood from 3 Greyhounds with vacuolated eosinophils and 3 with normal eosinophil granules. Blood smears from 4 of these dogs also were stained cytochemically with alkaline phosphatase (AP), chloracetate esterase (CAE), and alpha naphthyl butyrate esterase (ANBE). The morphologic features and tinctorial properties of vacuolated and normal eosinophils were compared. RESULTS: Twenty-six Greyhounds (53%) had vacuolated eosinophils and 23 (47%) had normal granulated eosinophils in smears stained with Wright's stain. Only 1% of eosinophils were vacuolated in non-Greyhound dogs. Twenty of the 23 (85%) Greyhounds with normal granulated eosinophils on Wright's-stained smears had vacuolated eosinophils in smears stained with Diff-Quik. Ultrastructurally, no morphologic differences were observed between granules of vacuolated and normal eosinophils. Both vacuolated and normal eosinophils in Greyhounds were positive for AP and negative for CAE and ANBE, as expected for normal dogs. CONCLUSION: Vacuolated eosinophils in Greyhounds likely reflect, at least in part, differential staining properties of the specific granules with different hematologic stains. Ultrastuctural and cytochemical features of eosinophil granules were similar in normal and vacuolated eosinophils from Greyhounds.     

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.     

Use of fine needle aspirates and flow cytometry for the diagnosis, classification, and immunophenotyping of canine lymphomas.

Fifty canine lymphomas were classified cytomorphologically using the updated Kiel classification scheme. Aspirates of lymph nodes from dogs with lymphoma were stained using 5 canine-specific antibodies and 3 human-specific antibodies that cross-react with canine lymphocytes. The antibody-stained aspirates were analyzed by flow cytometry. A total of 32 (64%) of the 50 lymphomas were characterized as B-cell origin and 18 (36%) were of T-cell origin. B-cell lymphomas were identified in 12 females and 20 males with a mean age of 8.35 years. T-cell lymphomas were identified in 8 females and 10 males with a mean age of 7.9 years. A minority of the lymphomas were low-grade B-cell and T-cell lymphomas (6/50, 12% and 4/50, 8%, respectively). The most common morphologic types were high-grade centroblastic and unclassifiable plasmacytoid for B- and T-cell lymphomas (18/50, 36% and 7/50, 14%, respectively).     

Exclusion of cytoplasmic fragments in flow cytometric analysis of lymph node samples from dogs with lymphoma using membrane-permeable violet laser-excitable DNA-binding fluorescent dye (DyeCycle Violet)

Background: Cytoplasmic fragments derived from fragile neoplastic lymphocytes are common in samples of lymph nodes collected from dogs with lymphoma. These cytoplasmic fragments interfere with accurate gating of target cells and quantification protocols used for flow cytometry because of their variable size and expression of lymphoid cell surface antigens on their membranes. Objective: The aim of this study was to develop a method to efficiently exclude cytoplasmic fragments from flow cytometric analysis of canine lymph nodes in which lymphoma was present. Methods: Single‐cell suspensions of neoplastic cells were prepared from biopsy samples and fine‐needle aspirates of lymph nodes from 23 dogs with lymphoma. Suspensions were stained using a violet laser‐excitable (405 nm) membrane‐permeable DNA‐binding fluorescent dye (DyeCycle Violet [DCV]), incubated with antibodies against CD3, CD5, CD21, CD22, and CD45, and then stained with 7‐amino‐actinomycin D (7‐AAD), an argon‐excitable (488 nm) membrane‐impermeable DNA‐binding fluorescent dye. Multiparameter flow cytometry was used for analysis based on selective uptake and laser‐activated fluorescence of these dyes. Results: Cytoplasmic fragments, which were DCV‐negative and CD45‐positive, and dead cells, which were positive for 7‐AAD, were efficiently separated from neoplastic cells. Conclusion: Staining with DCV is a useful method to improve flow cytometric gating methods and quantitative analyses of lymph node samples from dogs with lymphoma.     

Cytochemical characterization of peripheral blood cell populations of two Cyprinidae,Carassius auratus and Ctenopharyngodon idellus

Fish are the most diverse species of all vertebrate groups, and their blood cells have shown variable characteristics in terms of morphology. Cytochemical staining for enzyme activity in blood leukocytes will help assess the immune function of fish. We characterize blood cells from crucian carp (Carassius auratus) and grass carp (Ctenopharyngodon idellus) by using a Diff‐Quick stain as well as different cytochemical methods. Blood specimens obtained from crucian carp and grass carp were evaluated after cytochemical staining for acid phosphatase (ACP), alkaline phosphatase (ALP), naphthol AS chloroacetate esterase (AS‐DNCE), naphthyl acetate esterase (NAE), α‐naphthyl butyrate esterase (NBE), peroxidase (MPO) and periodic acid–Schiff's reaction (PAS) using commercial kits. Blood cell types were evaluated based on their morphological characteristics and the presence or absence of specific chromogen. The expression pattern of enzymes was similar between the two Cyprinidae and was also broadly consistent with other fish species. However, there were some interesting differences detected between crucian carp and grass carp, including naphthol AS chloroacetate esterase activity in monocytes, peroxidase activity and location in thrombocytes. The ACP, ALP and MPO expressions of different leukocytes of the two Cyprinidae were evaluated by Image Pro Plus and were analysed for statistical significant differences. This investigation provides basic haematology and enzyme activity analyses for crucian carp and grass carp and serves as an approach to evaluating the immune response of fish.     

Lineage differentiation of canine lymphoma/leukemias and aberrant expression of CD molecules

Multiparameter flow cytometry analysis and specific cluster differentiation (CD) molecules were used to determine the expression profiles of B- and T-cell antigens on lymph node preparations from 59 dogs with generalized or multisystemic lymphoma. Lymph node samples from 11 healthy dogs were labeled to validate the specificity of antibodies and to formulate guidelines for interpretation of the results obtained from lymphoma samples. In normal lymph nodes, T-lymphocytes expressing CD3, CD4, or CD8 beta represented 59+/-11%, 43+/-8%, or 16+/-5% of the total cells, whereas B-lymphocytes expressing either CD21 or surface IgM (IgM) represented 37+/-9% or 14+/-5%, respectively. Small lymphocytes could be distinguished from large lymphocytes by forward light scatter. Of the patient samples 29 different breeds were represented with Golden and Labrador retriever being the most common. The lymphoma samples segregated into three groups based on CD antigen expression. Thirty cases predominantly expressed one or more combinations of CD79a, IgM, and CD21 representing a B-cell lineage. Three B-cell cases also expressed the stem cell antigen, CD34. Sixteen cases expressed one or more combinations of CD3, CD4, and CD8 consistent with a T-cell lineage and CD3+CD4+CD8--phenotype was the most common. Thirteen cases showed a mixed expression profile for T- and B-cell antigens and in three cases CD14 was highly expressed. Clinical response was poorest for T-cell lymphomas. Leukemic states occurred in all three phenotypes; but mixed cell cases had the greatest proportion. Dual immunofluorescence staining confirmed co-expression of T-cell (CD3) and B-cell antigens (CD79a or CD21) on neoplastic lymphocytes of six mixed cell cases. In one mixed cell case, dual immunostaining identified lymphocyte populations that stained mutually exclusive for CD79a and CD3. Six mixed cell lymphomas tested by PCR showed clonality for rearranged antigen receptor. Four cases that were CD79a+CD3+ had TCRgamma chain gene rearrangements, whereas two cases that were CD3+CD8+CD21+ had Ig heavy chain rearrangement. One case expressing multiple CD molecules (CD3+CD8+CD21+CD14+) was PCR negative for both Ig and TCRgamma gene rearrangement and could not be classified into a B- or T-cell lineage. We show for the first time co-expression of B- and T-cell markers on lymphoma cells that had specific T- or B-cell gene rearrangements. These findings suggest that aberrant CD molecule expression is not an uncommon finding in canine lymphomas and is a useful diagnostic marker for malignancy.     

Precursor B-1 B cell lymphoma in a newborn calf.

A newborn Holstein female calf had neoplastic lesions in the skin and within the thoracic and abdominal cavities but not in the bone marrow, spleen, thymus, or most lymph nodes. Because the tumor cells were positive for CD79a (B cell marker), CD5 (B-1 cell marker) and terminal deoxynucleotidyl transferase (marker for immature lymphoid precursors), a diagnosis of precursor B-1 B cell lymphoma was made. The diagnosis was strongly supported by the fact that B-1 cells can develop in the fetus, unlike B-2 cells, which are produced after birth. The lymphoma was distinct from the typical calf form of lymphoma of B-2 cell origin, which does not express CD5 and is characterized by generalized lymphadenopathy and involvement of the bone marrow, blood and spleen.     

Identification of Equine Herpesvirus 5 in Horses with Lymphoma

Equine multinodular pulmonary fibrosis, equine herpesvirus 5 (EHV-5), and multicentric lymphoma were discovered in one patient. Review of gamma herpesvirus activity in humans revealed a propensity for lymphoproliferative disorders associated with infection. The objective was to determine the frequency of EHV-5 in lymphoma tissues and compare with the frequency found in the lymph nodes of clinically normal horses. Case control investigation of lymphoma-positive tissues and analysis via polymerase chain reaction (PCR) for EHV-5 was performed on 12 horses. Prospective collection and PCR analysis of lymph nodes (mesenteric or submandibular) for EHV-5 was performed on 21 control horses. Thirteen samples of lymphoma-positive tissues and fluid were submitted for PCR analysis for EHV-5. Of these, 67% was positive. In the control horse population, 14% was positive for EHV-5 (P = .004). Neoplastic samples positive for EHV-5 were classified as T-cell rich B-cell lymphoma (three), T-cell lymphoma (one), one was nondifferentiated, and two were not stained. Gamma herpesviruses in humans have been associated with lymphoproliferative diseases such as Kaposi sarcoma and Burkitt lymphoma. This study reveals an increased frequency of EHV-5 (gamma herpesvirus) in horses diagnosed with lymphoma compared with healthy control horses. Although the exact role this virus plays in the initiation or perpetuation of lymphoproliferative neoplasia is unknown, EHV-5 may be an etiologic agent associated with the development of some types of equine lymphoma.     

Morphologic and cytochemical characteristics of blood cells from the desert tortoise (Gopherus agassizii).

Morphologic and cytochemical staining characteristics of erythrocytes, leukocytes, and thrombocytes of the desert tortoise (Gopherus agassizii) were evaluated, using blood smears prepared from 23 healthy tortoises of Kern County, Calif. Special emphasis was placed on differentiating features of the various leukocytes and thrombocytes. A variety of cytochemical stains, including benzidine peroxidase, Sudan black B, chloroacetate esterase, alpha-naphthyl butyrate esterase, acid phosphatase, leukocyte alkaline phosphatase, periodic acid-Schiff, and toluidine blue were used. Heterophils had a characteristic, large, focal area of positive staining with chloroacetate esterase, alpha-naphthyl butyrate esterase, and acid phosphatase. Eosinophils stained diffusely positive with benzidine peroxidase, allowing differentiation of this leukocyte from heterophils. Thrombocytes stained focally positive with periodic acid-Schiff, allowing differentiation of these cells from lymphocytes, which stained uniformly negative. An intracytoplasmic body, commonly observed within erythrocytes, was considered ultrastructurally to represent a degenerate organelle.     

A clinical case of acute myelomonocytic leukemia in a Holstein cow

A 2-year, 3-month-old Holstein cow presented with anorexia and enlarged superficial lymph nodes. Fine needle aspiration cytology of the superficial lymph nodes revealed large blast cells. Hematological examination revealed anemia, neutropenia, and blast cells in peripheral blood. Blast cells were the predominant cell type in bone marrow aspirates. Of the non-erythroid cells, 26%, 58%, and 18% were positive for myeloperoxidase, α-naphthyl acetate esterase, and naphthol AS-D chloroacetate esterase, respectively. Pathological examination revealed the proliferation of neoplastic cells, which were positive for monocytic markers, in the affected lymph nodes. The cow was diagnosed with acute myelomonocytic leukemia based on these findings. This report highlights the importance of performing bone marrow aspiration cytology and cytochemical staining when diagnosing bovine myeloid leukemia.     

Prognostic significance of morphological subtypes in canine malignant lymphomas during chemotherapy

The aim of this study was to determine the response of different morphological subtypes of canine lymphoma to a standardized therapeutic protocol. Diagnosis of lymphoma was based on cytohistological analysis and immunophenotyping with antibodies against CD3 and CD79a of an enlarged lymph node or an extranodal mass. Fifty-seven cases were classified according to the updated Kiel classification adapted to the canine species, into 24 B-cell lymphomas (20 centroblastic polymorphic and four Burkitt-type subtypes), and 33 T-cell lymphomas (10 pleomorphic mixed, 10 lymphoblastic, eight unclassifiable high grade plasmacytoid, and five small clear-cell subtypes). All dogs were clinically staged at diagnosis. The protocol used l-asparaginase, vincristine, cyclophosphamide, doxorubicin, and prednisone. First remission duration and overall survival time were evaluated. Although the T-cell phenotype was associated, on the whole, with a poor prognosis, as previously reported in veterinary and human medicine, the study showed significant prognostic differences between the B- and the T-cell subtypes of canine lymphoma and suggests that clinico-morphological characterization of the disease is justified in dogs, as in humans.     

Argyrophilic nucleolar organizing regions and Ki67 equally reflect proliferation in fine needle aspirates of normal, hyperplastic, inflamed, and neoplastic canine lymph nodes (n = 101)

BACKGROUND: The count of argyrophilic nucleolar organizing regions (AgNOR) has been considered a useful variable that reflects cellular proliferation in canine lymph nodes, but it has not been compared with other markers of proliferation. Hypothesis: Ki67 and AgNORs are equally useful as markers of tissue proliferation in fine needle aspirates of canine lymph nodes. ANIMALS: A total of 101 dogs. MATERIAL AND METHODS: Prospective, observational study of a convenience sample of dogs. Two smears were prepared for a May-Gruenwald-Giemsa stain and a Ki67/AgNOR double stain. In addition, CD3/CD79a immunostaining was performed when cytologic examination revealed a lymphoma. The dogs were grouped as normal (n = 26), reactive hyperplasia (n = 25), lymphadenitis (n = 31), and lymphoma (n = 19), based on the physical examination and the cytologic findings. The AgNOR count/cell, AgNOR area/cell and the percentage of cells staining positive for Ki67 were evaluated in 100-167 cells (median, 113 cells) by using automatic image analysis. RESULTS: Mean (SD) AgNOR counts/cell were 1.36 +/- 0.19 in normal dogs, 1.55 +/- 0.26 in lymphadenitis, 1.65 +/- 0.32 in reactive hyperplasia, and 3.67 +/- 1.08 in lymphoma. The percentage of Ki67 positive cells was 2.67 +/- 0.99% in normal lymph nodes, 5.04 +/- 3.34% in lymphadenitis, 5.36 +/- 2.14% in reactive hyperplasia, and 30.2 +/- 10.8% in lymphoma. All variables were significantly higher in dogs with lymphoma compared with the other groups (P < .0001). The sensitivity and the specificity of the AgNOR count for diagnosing lymphoma were 95 and 96% at a cutoff value of >2.04 AgNORs/cell. The cutoff value for the Ki67 positive cells was >10.40% (sensitivity, 95%; specificity, 98%). CONCLUSION AND CLINICAL IMPORTANCE: The results indicated that both AgNOR and Ki67 counts were good diagnostic tools for assessment of proliferation in aspirates of canine lymph nodes.     

Hematologic, cytochemical, ultrastructural, and molecular findings of Hepatozoon-infected flat-headed cats (Prionailurus planiceps)

BACKGROUND: The flat-headed cat (Prionailurus planiceps) is a small wild cat of Southeast Asia and is considered extremely endangered. Little is known about the hematologic values, blood cell morphology, or hemoparasites of this species in relation to other Felidae. OBJECTIVES: The objective of this study was to report basic hematologic values and describe the light microscopic, cytochemical, and ultrastructural characteristics of blood cells in 2 wild-caught flat-headed cats. In addition, molecular analysis was done of a Hepatozoon organism found in the neutrophils of both cats. METHODS: Blood samples were collected into EDTA from the cephalic vein. A CBC, manual differential count, manual reticulocyte count, cytochemical stains (Sudan black B [SBB], alpha-naphthyl acetate esterase [ANAE], and beta-glucuronidase), and scanning and transmission electron microscopy were done using standard methods. RESULTS: HCT was slightly lower and reticulocyte counts and red cell distribution width were higher than the expected values for other species of cats. Hepatozoon organisms were found in the cytoplasm of neutrophils in both cats, but the number of infected neutrophils was very low (1%-2%). Neutrophils stained strongly positive for SBB, but were negative for ANAE and beta-glucuronidase. Hepatozoon-infected neutrophils were negative for SBB, but focally positive for ANAE and beta-glucuronidase. By transmission electron microscopy, gamonts of Hepatozoon sp were observed in neutrophils, and rarely free in plasma. Infected neutrophils had fewer specific granules and more mitochondria compared with noninfected neutrophils. PCR products of partial 18S rRNA revealed that the isolate of Hepatozoon in the flat-headed cats was closely related to that of the frog Hepatozoon sp. CONCLUSIONS: These results add to our understanding of hematologic values and blood cell morphology in Hepatozoon-infected flat-headed cats as well as the molecular analysis of the Hepatozoon organism, and may be useful for the health management and evaluation of hemoparasitic disease in this species.     

Flow cytometric immunophenotype of canine lymph node aspirates

Increasing availability of reagents able to distinguish subtypes of lymphocytes and other leukocytes has enabled greater understanding of lymphocyte biology and pathology in the dog. Lymphocytes in circulation most commonly are subjected to immunophenotypic assessment by flow cytometry, but needle aspirates of lymph nodes can be similarly suitable for immunophenotypic examination. In this investigation, the feasibility of immunophenotyping samples obtained by needle aspiration of lymph nodes from 32 dogs with no physical abnormalities and 6 dogs with lymphoma was determined. In addition, samples from 6 dogs were stored overnight at 4 degrees C and reanalyzed 24 hours later. For each sample, stained smear preparations were examined microscopically for lymphocyte morphology, neoplasia, and the presence of inflammatory cells. Expression of antigens on a corresponding sample of aspirated cells was determined by flow cytometric detection of antibody binding on a minimum of 10,000 events. The distribution of data was determined with Anderson-Darling tests, and reference intervals incorporating the central 95% of values were established. Adequate samples were obtained from 30 of 32 clinically normal dogs. Immunophenotypic results after 24 hours of storage were consistent with those obtained immediately after sampling. Reference intervals for lymphocyte subsets from normal dog lymph nodes were similar to the proportions of CD3+, CD4+, CD8+, and CD21+ lymphocytes found in blood. Aspirates of enlarged lymph nodes from dogs with lymphoma were readily classified by this technique. Aspiration of lymph nodes from dogs for comprehensive analysis by flow cytometry is feasible and applicable to immunophenotyping of lymphoma.     

Ultrasonography of the medial iliac lymph nodes in the dog

Sixty-one medial iliac lymph nodes of 38 different dogs (eight with adenocarcinoma of the apocrine glands of the anal sac, 13 with multicentric lymphoma, six with multicentric lymphoma but in clinical remission, and 11 control dogs) were evaluated to assess the ability of ultrasound to identify and interrogate these lymph nodes across the different groups and to differentiate these groups using different sonographic parameters. Ultrasound proved to be useful to assess canine medial iliac lymph nodes. An increase in size or number of detected lymph nodes or finding rounder or heterogeneous lymph nodes could differentiate lymph nodes of dogs of the control group from lymph nodes of dogs with lymphoma or an adenocarcinoma of the apocrine glands of the anal sac. Subcategories of malignancy could not be differentiated. More studies need to be performed, both with patients with reactive lymph nodes and also focusing on other canine superficial lymph nodes, before generalizing the results of this study to other areas or diseases.     

Development of the B- and T-cell compartments in porcine lymphoid organs from birth to adult life: an immunohistological approach.

Using immunohistological techniques, we studied the development over time of B- and T-cell compartments in the lymphoid organs of specific-pathogen-free pigs. Tissue samples were collected at various time-points, starting 2 days before the pigs were born until the pigs were 10 months old. The samples were collected from the spleen, thymus, peripheral lymph node, mesenteric lymph node, duodenum, jejunum, ileum, jejunal Peyer's patch and ileal Peyer's patch. Monoclonal antibodies specific to B- and T-cells were used to identify where the following cells were localized: IgM-B cells (cells positive to surface immunoglobulin), IgM-, IgG- and IgA-containing cells (cells positive to cytoplasmic immunoglobulin), and CD2-, CD4- and CD8-positive cells. The development of the B- and T-cell subpopulations in each organ was analysed. Two days before birth, most organs contained quantities of IgM-B cells. The spleen, lymph nodes, Peyer's patches and, notably, the thymus, contained some immunoglobulin-containing cells (Ig-CC); this finding indicates that pigs have cells that secrete immunoglobulins before birth. Just after birth, the incidence of Ig-CC increased in most organs; first IgM-CC increased, then either IgG- or IgA-CC increased, depending on the organ. T-cell development was observed clearly in spleen and in the lamina propria of the small intestine, in contrast to other organs, in which the T-cell compartments containing various T-cell subpopulations were well developed before birth. Comparison of the incidence of CD4+ and CD8+ cells showed that the CD4:CD8 ratio of these cells in the spleen, lymph nodes, Peyer's patches and small intestine is low, especially in adult pigs, compared with the CD4:CD8 ratio in other species. Weaning had little influence on the incidence of B- and T-cells in lymphoid organs. This study is the first immunohistological survey to describe the development of the major B- and T-cell subpopulations in various lymphoid organs of pigs, and it should be useful for future immunopathological and comparative immunological studies in pigs.