Study of the local immune response to Fasciola hepatica in the liver and hepatic lymph nodes of goats immunised with a peptide of the Sm14 antigen

The nature of the local immune response was assessed studying the distribution of CD2⁺, CD4⁺, CD8⁺, γδ⁺ T lymphocytes, IgM⁺ B cells, IL-4⁺ and IFN-γ⁺ cells in the liver and hepatic lymph nodes (HLN) of goats immunised with a synthetic peptide of the Sm14 antigen from Schistosoma mansoni and challenged with Fasciola hepatica. A morphometric study of HLN was also carried out in order to evaluate the hyperplasia of lymphoid follicles. Despite the decrease in fluke burdens found in the immunised group (45.9%) respect to the infected control group, this difference was not statistically significant due to the high individual variability. In liver, a significant increase of CD2⁺, CD4⁺, CD8⁺, γδ⁺ T lymphocytes was found in the infected groups respect to the uninfected control and in the infected control respect to the immunised group. HLN showed a significant enlargement due to the hyperplasia of lymphoid follicles and infiltration of CD2⁺, CD4⁺, CD8⁺, γδ⁺ T lymphocytes in both infected groups respect to the uninfected control, with no significant differences between the infected control and immunised group. IFN-γ⁺ lymphoid cells was absent or very occasional in HLN where the number of IL-4⁺ cells was higher than that of IFN-γ, suggesting a polarized Th2 response in immunised and in infected control group.     

Canine leishmaniasis transmission: higher infectivity amongst naturally infected dogs to sand flies is associated with lower proportions of T helper cells

The dog is the main reservoir of Leishmania infantum, which is a parasite spread among canine hosts by the bite of sand flies. Phlebotomus perniciosus is the sand fly acting as a major vector in the Mediterranean basin. As a consequence, the dog will suffer from leishmaniasis. In this work the infective capacity of infected dogs, established by direct xenodiagnosis, has been investigated in relation to their immunological status by determining the lymphocyte percentages present in peripheral blood mononuclear cells. We found a significant association between the percentages of T helper cells (CD4/TcRαβ⁺and CD4/CD45RA⁺) and the infection rates detected in the vector, while significant association was not detected in the case of the T cytotoxic cells (CD8/TcRαβ⁺and CD8/CD45RA⁺). The relationship discovered was that the lower the CD4⁺T cell count, the higher the rate of the infection in the vector.     

Effect of polyoxyethylene and polyoxypropylene nonionic block copolymers on performance and recruitment of immune cell subsets in weaned pigs

Background

Because European-wide directives are restricting the non-clinical use of antibiotics as in-feed growth promotors in swine production, there is an intensive search for alternative strategies for control and prevention of losses among young pigs. With the growing knowledge of the porcine immune system and its endogenous modulation, it has been clearly established that exogenous immunomodulation using adjuvants and immune response modifiers (IRMs) represents an important prophylactic/therapeutic approach in the prevention/treatment of both stress- and microbial-induced disorders that accompaning weaning. However, it is essential to select a fully evaluated agent which may act either as a nonspecific IRM or synergistically as an adjuvant with vaccines. The synthetic macromolecules with a long history as adjuvant and IRM are nonionic block copolymers which consist of polyoxyethylene (POE) and polyoxypropylene (POP) molecules.

Methods

The aim of this work was to evaluate the effectiveness of POE-POP given as a single peroral dose on productivity parameters such as body weight gain, feed intake and feed conversion ratio, and systemic and intestinal immune parameters by assessing the proportions of CD45⁺ lymphoid cells, CD4⁺ and CD8⁺ T cells, and CD21⁺ B cells in the peripheral blood as well as the number of CD45RA⁺ naive lymphoid cells residing in the ileal mucosa in weaned pigs during a follow-up study 5 weeks after the treatment.

Results

Pigs treated with POE-POP had better feed intake (+ 14.57%), higher average body mass at the end of the experiment (20.91 kg vs. 17.61 kg), and higher body weight gain in relation to Day 0 (191.63% vs. 144.58%) as well as in relation to nontreated pigs (+ 18.74%), with a lower feed conversion ratio (− 30.26%) in comparison to the control pigs. A much lower diarrhea severity score (5 vs. 54) was recorded in pigs treated with POE-POP (− 90.74%) than in the control pigs. A higher average diarrhea severity (ADS) was recorded in the control pigs (1.54 vs. 0.14), whereas the treatmant group had much a lower ADS ratio (− 90.91%) after 35 days of the experiment. The pigs that were treated with POE-POP had an increased proportion of CD45⁺, CD4⁺ and CD8⁺ cells at Day 21 (at p < 0.05, p < 0.05 or p < 0.01, respectively), Day 28 (at p < 0.01, respectively) and Day 35 (at p < 0.01, p < 0.05 or p < 0.01, respectively) as well as of CD21⁺ cells at Day 28 (p < 0.05) and Day 35 of the experiment (p < 0.01). Also, these pigs had more numerous CD45RA⁺ cells in interfollicular (p < 0.05) and follicular areas (p < 0.01) of the ileal Peyer’s patches than did control pigs.

Conclusion

This property of POE-POP to induce recruitment of circulating and intestinal immune cell subsets in weaned pigs may allow the use of IRM-active block copolymers as adjuvants for vaccines, particularly those orally delivered and targeted to the gut-associated lymphoid tissues that are well known to promote rather tolerogenic than protective immune responses.     

Isolation and characterization of equine nasal mucosal CD172a+ cells

The nasal mucosa surface is continuously confronted with a broad variety of environmental antigens, ranging from harmless agents to potentially harmful pathogens. This area is under rigorous control of professional antigen presenting cells (APCs), such as dendritic cells (DCs) and macrophages. Mucosal APCs play a crucial role in inducing primary immune responses and the establishment of an immunological memory. In the present study, a detailed characterization of CD172a⁺ cells, containing the APCs residing in the equine nasal mucosa was performed for the first time. CD172a⁺ cells were isolated from collagenase-treated equine nasal mucosa fragments by MACS. Expression of surface markers was determined by flow cytometry and functional analysis was done by measuring the uptake of FITC conjugated ovalbumin (FITC-OVA). Cell surface phenotype of the isolated cells was as follows: 90% CD172a⁺, 30% CD1c⁺, 46% CD83⁺, 42% CD206⁺ and 28% MHC II⁺. This clearly differs from the phenotype of blood-derived monocytes: 96% CD172a⁺, 4% CD1c⁺, 11% CD83⁺, 9% CD206⁺, 72% MHC II⁺ and blood monocyte derived DCs: 99% CD172a⁺, 13% CD1c⁺, 30% CD83⁺, 51% CD206⁺ and 93% MHC II⁺. The CD172a⁺ nasal mucosal cells were functionally able to endocytose FITC-OVA but to a lesser degree than monocyte-derived DCs. Together, these results demonstrate that the isolated CD172a⁺ nasal mucosal cells resemble immature DCs in the nasal area.     

Changes in Lymphocyte Subsets in the Bronchus‐associated Lymphoid Tissue of Goats Naturally Infected with Different Mycoplasma Species

The distribution of cells containing lysozyme, S‐100 protein, CD3, CD4, CD8, major histocompatibility complex class II antigen and immunoglobulin G (IgG) was analysed in the bronchus‐associated lymphoid tissue (BALT) of goats naturally infected with three Mycoplasma species. This study included the immunohistochemical characterization of the pneumonic lesions of 18 goats (3–5 months old) infected with one of the following Mycoplasma species: M. mycoides ssp. mycoides, Large Colony type (goats no. 1–6), M. mycoides ssp. capri (goats no. 7–12) and M. capricolum ssp. capricolum (goats no. 13–18). Microscopically, infected animals showed a moderate bronchointerstitial pneumonia, characterized by lymphoid hyperplasia of the BALT and infiltration of mononuclear cells in the alveolar walls and airways. The main cellular type in the BALT was represented by CD3⁺ T lymphocytes, and the ratio of CD4⁺:CD8⁺ cells was >2. The BALT showed large germinal centres mainly composed of IgG⁺ B lymphocytes, with numerous S‐100⁺ follicular dendritic cells. The presence of follicular dendritic cells confirmed the high degree of organization of this lymphoid tissue. The immunohistochemical results showed that activated T lymphocytes, particularly in the CD4 subset, and IgG⁺ B cells, play a major role in the immune response of the caprine lung infected with these species of mycoplasmas.     

Immunohistochemical characterization of Kisselev nodules (ectopic lymphoid follicles) in wild boar (Sus scrofa L.)

This paper describes the histopathological features and cellular distribution of T lymphocytes (CD3), B cells (CD79), follicular dendritic cells (FDC) and macrophages (alpha-1-antitrypsin, lysozyme) in lymphoid aggregates (Kisselev nodules) found in the lung, kidney and liver of wild boar (Sus scrofa L.). The distribution of immunoreactive cells, tested for antibodies, was similar to that found in the cortex of lymph nodes: lymphoid follicles with germinal centers mainly consisting of CD79⁺ B cells with sparse interfollicular tissue (CD3⁺ T lymphocytes). This finding and the association of these structures with helminthic infections suggests that local humoral immunity is central to the organism’s response to parasitic challenge. The presence of follicular dendritic cells confirms the high degree of organization of these lymphoid-like structures. The role of other pathogenic factors and the induction of chronic inflammatory reaction in these ectopic lymphoid sites is also discussed.     

Postnatal Development of Lymphocyte Subpopulations in the Intestinal Lymph Nodes in Goats

The incidence and location of CD2⁺, CD4⁺, CD8⁺ and γ/δ T lymphocytes and IgM⁺ B lymphocytes were studied in the intestinal lymph nodes in 1-week, 1-month, 3-month and 7-month-old goats, using monoclonal antibodies and immuno-histochemical methods. The cortical area of the intestinal lymph nodes in 1-week-old animals contains only primary follicles occupied by IgM⁺ B lymphocytes and some CD2⁺CD4⁺ T lymphocytes. In goats older than 1 month, secondary follicles, that increased in number and size with age, were observed; the light zone of the germinal centre was occupied by IgM⁺ lymphocytes and some CD2⁺ and CD4⁺ T lymphocytes. In the other compartments of the lymph nodes, B lymphocytes were scarce, their number increasing with age in the medulla and diminishing in the paracortex. The numerous CD2⁺ T lymphocytes in the interfollicular area increased in number in the paracortical area of the 7-month-old goats, simultaneously with an increase in the MHC II⁺ dendritic cells and the CD4/CD8 ratio, which was greater than 1. The γ/δ T lymphocytes represented a minor subpopulation scattered through the lymph nodes.     

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.     

Development and use of a polarized equine upper respiratory tract mucosal explant system to study the early phase of pathogenesis of a European strain of equine arteritis virus

The upper respiratory tract mucosa represents the first line of defense, which has to be overcome by pathogens before invading the host. Considering the economic and ethical aspects involved in using experimental animals for pathogenesis studies, respiratory mucosal explants, in which the tissue’s three-dimensional architecture is preserved, may be ideal alternatives. Different respiratory mucosal explant cultures have been developed. However, none of them could be inoculated with pathogens solely at the epithelium side. In the present study, equine nasal and nasopharyngeal explants were embedded in agarose (3%), leaving the epithelium side exposed to allow apical inoculation. Morphometric analysis did not show degenerative changes during 72 h of cultivation. The number of apoptotic cells in the mucosa slightly increased over time. After validation, the system was used for apical infection with a European strain (08P178) of equine arteritis virus (EAV) (10^7.6TCID₅₀/mL per explant). Impermeability of agarose to virus particles was demonstrated by the absence of labeled microspheres (40nm) and a lack of EAV-antigens in RK13 cells seeded underneath the agarose layer in which inoculated explants were embedded. At 72 hpi, 27% of the EAV-positive cells were CD172a⁺ and 19% were CD3⁺ in nasal explants and 45% of the EAV-positive cells were CD172a⁺ and 15% were CD3⁺ in nasopharyngeal explants. Only a small percentage of EAV-positive cells were IgM⁺. This study validates the usefulness of a polarized mucosal explant system and shows that CD172a⁺ myeloid cells and CD3⁺ T lymphocytes represent important EAV-target cells in the respiratory mucosa.     

Engraftment of canine peripheral blood lymphocytes into nonobese diabetic-severe combined immune deficient IL-2R common gamma chain null mice

To study the canine immune system we generated a mouse model engrafted with canine lymphocytes using NOD SCID IL2R common gamma chain ?/? (NSG) mice as recipients (Ca-PBL-SCID). Engraftment of canine peripheral blood lymphocytes (PBLs) was determined post-injection with 10⁷ peripheral blood mononuclear cells (PBMCs) into irradiated NSG mice using flow cytometry and fluorescently labeled antibodies specific to canine helper T cells (CD45⁺ CD4⁺), cytotoxic lymphocytes (CD45⁺ CD8⁺), regulatory T cells (CD45⁺ CD4⁺ Foxp3⁺), and B cells (CD45⁺ Ig⁺ CD21_lo). Canine CD45⁺ lymphocytes were detectable as early as day 1 in the peritoneal cavity, and beginning at 9 days in the blood, bone marrow, and spleen. CD4⁺ T cells, of which Foxp-3⁺ CD25_hi cells constituted a minor percentage, were the predominant lymphocyte population at 9 days post engraftment contrasting with increasing proportions of CD8⁺ CTL's and Ig⁺ B cells beginning at 16 days. Canine immunoglobulin was initially detected in the serum of Ca-PBL-SCID mice at 9 days post-engraftment and peaked in concentration at day 36. From day 28 to 52 post-engraftment 30% of the Ca-PBL-SCID mice became markedly anemic and thrombocytopenic, yet gross and histopathologic examination of bone marrow, kidneys, spleen, liver, and intestine revealed no obvious lesions. Blood smear evaluation revealed agglutination of mature red blood cells, reticulocytes and a regenerative anemia. These findings demonstrate that NSG mice are capable of engraftment of canine PBLs yet develop graft versus host disease similar to Hu-PBL-SCID mice.     

Replication of neurovirulent equine herpesvirus type 1 (EHV-1) in CD172a+ monocytic cells

Equine herpesvirus type 1 (EHV-1) is responsible for respiratory disorders, abortion and myeloencephalopathy (EHM) in horses. Two pathotypes of EHV-1 strains are circulating in the field: neurovirulent (N) and non-neurovirulent (NN). For both strains, CD172a⁺ monocytic cells are one of the main carrier cells of EHV-1 during primary infection, allowing the virus to invade the horse’s body. Recently, we showed that EHV-1 NN strains showed a restricted and delayed replication in CD172a⁺ cells. Here we characterize the in vitro replication kinetics of two EHV-1 N strains in CD172a⁺ cells and investigate if the replication of these strains is similarly silenced as shown for EHV-1 NN strains. We found that EHV-1 N replication was restricted to 7–8% in CD172a⁺ cells compared to 100% in control RK-13 cells. EHV-1 N replication was not delayed in CD172a⁺ cells but virus production was significant lower (10^3.0 TCID₅₀/10⁵ inoculated cells) than in RK-13 cells (10^8.5 TCID₅₀/10⁵ inoculated cells). Approximately 0.04% of CD172a⁺ cells produced and transmitted infectious EHV-1 to neighbour cells compared to 65% of RK-13 cells. Unlike what we observed for the NN strain, pretreatment of CD172a⁺ cells with histone deacetylases inhibitors (HDACi) did not influence the replication of EHV-1 N strains in these cells. Overall, these results show that the EHV-1 replication of N strains in CD172a⁺ cells differs from that observed for NN strains, which may contribute to their different pathogeneses in vivo.     

Porcine reproductive and respiratory syndrome virus induces pronounced immune modulatory responses at mucosal tissues in the parental vaccine strain VR2332 infected pigs

Porcine reproductive and respiratory syndrome (PRRS) is a chronic viral disease of pigs caused by PRRS virus (PRRSV). The PRRSV VR2332 is the prototype North American parental strain commonly used in the preparation of vaccines. Goal of this study was to understand missing information on VR2332 induced immune modulation at the lungs and lymphoid tissues, the sites of PRRSV replication. Pigs were infected intranasally and samples collected at post-infection day (PID) 15, 30, and 60. Microscopically, lungs had moderate interstitial pneumonia, and the virus was detected in all the tested tissues. Peak antibody response and the cytokine IFN-γ secretion were detected at PID 30, with increased TGF-β until PID 60. Population of CD8⁺, CD4⁺, and CD4⁺CD8⁺T cells, Natural killer (NK) cells, and γδ T cells in the lungs and lymphoid tissues were significantly modulated favoring PRRSV persistence. The NK cell-mediated cytotoxicity was significantly reduced in infected pigs. In addition, increased population of immunosuppressive T-regulatory cells (Tregs) and associated cytokines were also observed in VR2332 strain infected pigs.     

Comparative studies on the secondary lymphoid tissue areas in the chicken bursa of Fabricius and calf ileal Peyer's patch

The chicken bursa of Fabricius and calf ileal Peyer's patch are thought to be the primary lymphoid organs of B cell development. In the bursa, the existence of secondary lymphoid tissue, called the diffusely infiltrated area, has been recognized. Recently, we have found the presence of a region of secondary lymphoid tissue in the ileal Peyer's patch at the period of the most rapid growth of this organ. In this study, we compared the development of these secondary lymphoid tissue regions in the bursa and ileal Peyer's patch histologically. Before hatching, lymphatic follicle formation occurred in the bursa, but not in the diffusely infiltrated area, where only a small number of lymphoid cells were found. However, during fetal calf development, lymphatic follicle formation occurred not only in the primary lymphoid organ but also in the secondary lymphoid tissue regions. Therefore, the prenatal development of the secondary lymphoid tissue regions of the bursa and ileal Peyer's patch were distinct. After hatching, formation of the germinal center, which contained many CD4⁺ cells, was observed in the diffusely infiltrated area of the bursa. After birth, many CD4⁺ cells and IgG mRNA expression were observed in the lymphatic follicle of the secondary lymphoid tissue regions in the ileal Peyer's patch, but rarely in the ileal Peyer's patch lymphatic follicles. The change of character of these secondary lymphoid tissue regions at the postnatal stage might be dependent on external antigens.     

Assessing the immune state of dogs suffering from pituitary gland dependent hyperadrenocorticism by determining changes in peripheral lymphocyte subsets

In order to evaluate the immune state of dogs suffering from pituitary-dependent hyperadrenocorticism (PDH), peripheral lymphocyte subsets were examined. Twenty seven PDH dogs and eight healthy control dogs were used in the current study. Eight healthy dogs served as the control group. Twenty seven PDH dogs were categorized into 4 groups based on their post serum cortisol concentrations by ACTH stimulation test: 2−5, excellent control (n = 8); 5−20, fair control (n = 7); >20, poor control (n = 4); and untreated (n = 8). Cell counts were executed with white blood cells (WBC), lymphocytes, CD3⁺ (T lymphocytes), CD4⁺ (Helper T lymphocytes), CD8⁺ (Cytotoxic T lymphocytes), CD21⁺ (B lymphocytes) cells in addition to calculating CD4⁺/CD8⁺ ratio. Results indicated a significant difference in lymphocyte numbers and lymphocyte subset populations (CD3⁺, CD4⁺, CD8⁺, and CD21⁺ cells) between PDH and control dogs. Moreover, comparison of the PDH groups (excellent control; fair control; poor control; untreated) demonstrated that all groups had a significant decrease in lymphocytes numbers (CD3⁺, CD4⁺ and CD21⁺ cell counts) as compared to control group. Meanwhile, no significant differences were observed in WBC counts and CD4⁺/CD8⁺ ratio between groups. Furthermore, lymphocyte subset distribution in excellent control PDH dogs without concurrent disease (n = 4) better resembled that of control dogs as compared to PDH dogs with concurrent disease (n = 4). PDH dogs may be suffering from an immuno-depressed state as evidenced by significant differences in lymphocyte subset populations. Furthermore, treatment of both PDH and concurrent disease might improve lymphocyte subset distribution.     

Characterization of tuberculous granulomas in different stages of progression and associated tertiary lymphoid tissue in goats experimentally infected with Mycobacterium avium subsp. hominissuis

Oral infection of goats with Mycobacterium avium subsp. hominissuis (MAH) resulted in a large variety of granulomas in organized gut-associated lymphatic tissues and intestinal lymph nodes. To characterize the cellular composition of granulomas, CD4⁺, CD8⁺, γδ, B lymphocytes and plasma, CD25⁺, CD68⁺, MHC-II⁺, Ki67⁺ and endothelial cells were labeled in consecutive frozen sections by immunohistochemistry and acid fast bacilli (AFB) by Kinyoun stain. Granulomas with extensive necrosis, little mineralization and variable numbers of AFB surrounded by many CD4⁺ T cells, but only few epitheloid macrophages were observed in severely sick goats at 2–3 mpi. They were interpreted as exuberant immune reaction. Organized granulomas with very few AFB were seen in clinically healthy goats at 13 mpi. The necrotic cores were surrounded by a zone of granulomatous infiltrate with many epitheloid macrophages and few lymphocytes. This zone was initially wide and highly vascularized and became progressively smaller. It was enclosed by an increasing layer of connective tissue. All organized granulomas were surrounded by compartimentalized tertiary lymphoid tissue. The granulomas in experimental infection of goats with MAH reflect the heterogeneity of lesions seen in mycobacterial infections of humans and ruminants and are therefore valuable for comparative research.     

<Emphasis Type="Italic">Streptococcus uberis</Emphasis>-specific T cells are present in mammary gland secretions of cows and can be activated to kill <Emphasis Type="Italic">S. uberis</Emphasis>

The presence, phenotype and function of Streptococcus uberis-specific T cells in the mammary gland secretion (MGS) and blood of cows exposed to S. uberis were assessed. MGS T cells in the udder were purified and incubated with autologous blood monocytes as antigen-presenting cells (APC). Most cows, irrespective of prior S. uberis infection status and lactation status, were shown to have S. uberis-specific T cells both in MGS and in the blood. When cells from a subgroup of cows were studied, it was found that the S. uberis-specific T cells produced high levels of interferon-gamma (IFN-γ), but low levels of interleukin-10 (IL-10). A high percentage of responding T cells were of the CD8 ⁺ memory (CD45RO) subset. T cells from the MGS specific for S. uberis were propagated from animals during the drying off period and expanded in vitro using interleukin-2 (IL-2) and S. uberis antigens. This led to the accumulation of T cells of the CD8 ⁺ subset bearing memory cell markers (CD45A ⁻ , CD45RO ⁺ ), which released high levels of IFN-γ. Four of the five T cell lines derived from the MGS of three animals had substantial direct killing activity towards S. uberis in vitro. It is concluded that there is an emergence of S. uberis-specific bactericidal T cells in the MGS of cows after infection or environmental exposure to S. uberis. Vaccines aimed at activating and expanding this T cell population in the mammary glands of cattle may offer an avenue for the prevention of mastitis caused by S. uberis.     

Cytometric Analysis of Surface Molecules of Leucocytes and Phagocytic Activity of Granulocytes and Monocytes/Macrophages in Cows with Pyometra

Pyometra is a serious problem in dairy cow herds, causing large economic losses due to infertility. The development of pyometra depends mainly on the immunological status of the cow. The aim of the study was a comparative evaluation of selected indicators involving non‐specific and specific immunity in cows with pyometra and in cows without inflammation of the uterus. The study was performed in 20 cows, which were divided into two groups: pyometra group and healthy group, each comprising 10 cows, based on the results of cytological and ultrasonographic tests. A flow cytometric analysis was performed for the surface molecules CD4, CD8, CD14, CD21, CD25 and CD4⁺CD25⁺ on leucocytes, and the phagocytic activity was determined from granulocytes and monocytes/macrophages in the peripheral blood and uterine washings, respectively. It was demonstrated that the percentage of phagocytic granulocytes and monocytes/macrophages in both the peripheral blood and uterine washings was significantly lower in cows with pyometra compared with the healthy group (p < 0.001). Significantly (p ≤ 0.001) lower percentage of CD4⁺, CD14⁺, CD25⁺ and CD4⁺CD25⁺ phenotype leucocytes was also observed in the peripheral blood of cows from the pyometra group, along with a significantly higher (p < 0.001) percentage of CD8⁺ and CD21⁺ lymphocytes as compared to the healthy group. The results of work indicate that disfunction of cell immunity coexisting with pyometra may be caused by a bacterial infection and the presence of blocking agents (IL‐10), released by the increasing number of CD8⁺ lymphocytes what leads to the advanced inflammation of uterus.     

Cutaneous infiltrates and peripheral blood immune responses in dogs with immunomodulatory‐responsive lymphocytic–plasmacytic pododermatitis

This study characterizes T‐ and B‐lymphocyte responses in the peripheral blood and lesional skin of dogs with immunomodulatory‐responsive lymphocytic–plasmacytic pododermatitis (ImR‐LPP), a term previously proposed to denote a subpopulation of dogs with idiopathic pododermatitis. T‐cell (CD3⁺, CD4⁺ and CD8⁺) and B‐cell (CD21⁺) counts were significantly increased in both the epidermis and dermis of lesional ImR‐LPP skin compared with that in pedal skin from healthy controls. CD3⁺, CD4⁺, CD8⁺ and CD21⁺ cells were commonly observed in perivascular sites in the superficial dermis, periadnexally, beneath the dermal–epidermal (DE) junction and in the epidermis of lesional ImR‐LPP skin. The CD8⁺/CD3⁺ T‐cell ratio in peripheral blood was significantly increased in the ImR‐LPP group (0.42 versus 0.35 in controls). Serum IgA, IgG and IgM concentrations were all significantly elevated in affected dogs. Lymphocyte stimulation indices in ImR‐LPP dogs were comparable with control levels except for a lower response to ionomycin (6.0 versus 11.1). Dogs with ImR‐LPP had a higher incidence and mean (semi‐quantitative) score for IgA, IgG and IgM deposits in the epidermis, and a significantly increased incidence of dermal IgA⁺, IgG⁺ and IgM⁺ mononuclear inflammatory cells. The results indicate that upregulated T‐ and B‐lymphocyte responses may contribute to the pathogenesis of the skin lesions observed in dogs with ImR‐LPP.     

Canine CD8 T cells showing NK cytotoxic activity express mRNAs for NK cell-associated surface molecules

Natural killer (NK) cells have been considered to be a group of lymphocytes lacking clonally distributed receptors for antigens typical of T cells and B cells. In some mammalian species, including humans, a subpopulation of CD8⁺ peripheral blood lymphocytes (PBLs) exhibits NK activity. This NK subpopulation has not been well characterized in mammals and its characterization is particularly poor in the dog. In this study, we demonstrated that a subset of canine CD8⁺ cells derived from PBLs and lymphokine (IL-2)-activated killers (LAKs) of PBLs that was CD3⁺, CD4^?, CD21^?, CD5^lo, α/βTCR⁺, and γ/δTCR^? contained substantially higher levels of mRNAs for NK cell-related receptors (NKp30, NKp44, NKG2D, 2B4, and CD16 for PBL, and NKG2D and CD56 for LAK) than the corresponding CD8^? cells. This subset of CD8⁺ lymphocytes derived from LAKs also displayed significantly higher NK cytotoxic activity than the corresponding CD8^? cells. In contrast, CD8⁺ cells derived from nonstimulated PBLs showed very low levels of NK cytotoxic activity. Our results indicate that, in IL-2-stimulated PBLs, canine CD8⁺ cells are an important subset associated with NK cytotoxic activity.     

B‐cell lymphoma with Mott cell differentiation in a cat

A 12‐year‐old, male castrated Domestic Shorthair cat was presented to Animal Medical Center of Gifu Univeristy with anorexia and vomiting. Physical examination revealed an enlarged left tonsil and right mandibular lymph node (approximately 2–3× the normal size), and a submucosal mass on the right side of the epiglottis (1.5 × 2.0 cm). On computed tomography images, an enlarged left tonsil, and enlarged right mandibular, right pharyngeal, and left and right cervical lymph nodes were observed. Cytologic examination of smears of tonsil and lymph nodes revealed numerous medium‐ to large‐sized neoplastic lymphoid cells, approximately half of which contained one or several light‐blue homogenous globoid cytoplasmic inclusions (5–10 μm), which stained magenta with periodic acid–Schiff (PAS) stain. Histopathologic examination of the left tonsil revealed diffuse proliferation of medium‐ to large‐sized neoplastic lymphoid cells effacing the original lymphoid architecture. Half of the cells contained one or several eosinophilic globoid cytoplasmic inclusions, which stained magenta with PAS and showed positive immunohistochemical reactions for immunoglobulin M (IgM) and λ light chain. Neoplastic lymphoid cells were also CD20⁺, Pax5⁺, and MUM1⁺, and CD3^?. Thus, the neoplastic lymphoid cells expressed a B‐cell immunophenotype, and the globoid cytoplasmic inclusions represented an aberrant IgM λ light chain accumulation, similar to Russell bodies. B‐cell lymphoma with Mott cell differentiation was diagnosed based on cytologic, histopathologic, and immunohistochemical features. This is the first report of B‐cell lymphoma with Mott cell differentiation in a cat.