Identification of very early inflammatory markers in a porcine myocardial infarction model.

BACKGROUND: Acute myocardial infarction (AMI) is one of the most deleterious conditions leading to cardiovascular diseases and mortality. The importance of an early and accurate diagnosis assures immediate medical treatments, which are fundamental to reduce mortality and improve prognoses. AMI is associated to an inflammatory response which includes the increase of circulating inflammatory cytokines, chemokines and immune cell activation. This study aimed to identify which are the very early immune-related biomarkers that may be used as predictors of myocardial infarction severity. In order to mimic the pathophysiological events involved in human myocardial infarction, a temporary occlusion (90 min) of the mid-left anterior descending coronary artery was performed in a swine animal model. RESULTS: Lymphocyte subsets analysis in peripheral blood revealed significant alterations in CD4+/CD8+ ratio and naïve and effector/memory T cell percentages at 1 h post-myocardial infarction. Changes in TH1/TH2-related cytokine, monocyte and neutrophil markers gene expression were observed in peripheral blood lymphocytes, as well. Additionally, significant correlations between cardiac parameters (cardiac enzymes, left ventricular ejection fraction and % infarct) and blood-derived parameters (cytokine expression and lymphocyte subset distribution) were found. CONCLUSIONS: Peripheral blood lymphocyte alterations are easily and swiftly detectable, so they may be good biomarkers for a very early prognosis and to predict myocardial infarction severity.

Low CD4/CD8 ratio in classical swine fever postnatal persistent infection generated at 3 weeks after birth.

Classical swine fever virus (CSFV) is one of the most important pathogens affecting swine. After infection with a moderate virulence strain at 8 hours after birth, CSFV is able to induce viral persistence. These animals may appear clinically healthy or showed unspecific clinical signs despite the permanent viremia and high viral shedding, in absence of immune response to the virus. Given the role played by this infection in disease control, we aimed to evaluate the capacity of CSFV to induce postnatal persistent infection at 3 weeks after birth. Nine pigs were CSFV infected and sampled weekly during 6 weeks and viral, clinical, pathological and immunological tests were carried out. Also, the CD4/CD8 ratio was calculated with the purpose to relate this marker with the CSFV persistent infection. The IFN-α response was detected mainly 1 week after infection, being similar in all the infected animals. However, 44.4% of animals were CSFV persistently infected, 33.3% died and 22.2% developed specific antibody response. Interestingly, in persistently infected pigs, the T-CD8 population was increased, the T-CD4 subset was decreased and lower CD4/CD8 ratios were detected. This is the first report of CSFV capacity to confer postnatal persistent infection in pigs infected at 3 weeks after birth, an age in which the weaning could be carried out in some swine production systems. This type of infected animals shed high amounts of virus and are difficult to evaluate from the clinical and anatomopathological point of view. Therefore, the detection of this type of infection and its elimination in endemic areas will be relevant for global CSF eradication. Finally, the low CD4/CD8 ratios found in persistently infected animals may be implicated in maintaining high CSFV replication during persistence and further studies will be performed to decipher the role of these cells in CSFV immunopathogenesis.

Prior Puma Lentivirus Infection Modifies Early Immune Responses and Attenuates Feline Immunodeficiency Virus Infection in Cats.

We previously showed that cats that were infected with non-pathogenic Puma lentivirus (PLV) and then infected with pathogenic feline immunodeficiency virus (FIV) (co-infection with the host adapted/pathogenic virus) had delayed FIV proviral and RNA viral loads in blood, with viral set-points that were lower than cats infected solely with FIV. This difference was associated with global CD4⁺ T cell preservation, greater interferon gamma (IFN-γ) mRNA expression, and no cytotoxic T lymphocyte responses in co-infected cats relative to cats with a single FIV infection. In this study, we reinforced previous observations that prior exposure to an apathogenic lentivirus infection can diminish the effects of acute infection with a second, more virulent, viral exposure. In addition, we investigated whether the viral load differences that were observed between PLV/FIV and FIV infected cats were associated with different immunocyte phenotypes and cytokines. We found that the immune landscape at the time of FIV infection influences the infection outcome. The novel findings in this study advance our knowledge about early immune correlates and documents an immune state that is associated with PLV/FIV co-infection that has positive outcomes for lentiviral diseases.

Specific immunotypes of canine T cell lymphoma are associated with different outcomes.

Canine lymphoma is a heterogeneous disease with many different subtypes. Lymphoma of T cell type in particular is variable in outcome, and includes subtypes with non-progressive, slowly- and rapidly-progressive disease course. Association of immunotype with disease course is incompletely defined. Here, results of flow cytometric immunotyping of 127 canine T cell lymphomas were analyzed in relation to survival and progression free interval. Samples originated from 101 multicentric, 8 mediastinal, 6 cutaneous, 5 hepatosplenic, 5 gastrointestinal and 2 other anatomic subtypes of T cell lymphoma. Compared to multicentric T cell lymphoma, gastrointestinal lymphoma had shorter survival and progression free interval, and hepatosplenic lymphoma had shorter survival. Among dogs with multicentric T cell lymphoma, immunotypes of CD4+/CD8-/MHCII+, CD4-/CD8+/MHCII+ and CD4-/CD8+/MHCII- were associated with longer survival times than the immunotype of CD4+/CD8-/MHCII-, and immunotypes of CD4+/CD8-/MHCII+, CD4-/CD8+/MHCII-, and CD4-/CD8-/MHCII+ were associated with longer progression free intervals. Dogs with multicentric T cell lymphoma and concurrent leukemia had shorter survival but similar progression free interval compared to those without leukemia. Body weight, sex, hypercalcemia, cell size, expression of CD3 and use of combination or single agent chemotherapy did not significantly affect outcome of multicentric TCL.

Effect of whole yeast cell product supplementation (CitriStim®) on immune responses and cecal microflora species in pullet and layer chickens during an experimental coccidial challenge.

Three separate experiments were conducted to study the effects of whole yeast cell product supplementation in pullets and layer hens. Body weight gain, fecal and intestinal coccidial oocyst counts, cecal microflora species, cytokine mRNA amounts, and CD4+ and CD8+ T-cell populations in the cecal tonsils were analyzed following an experimental coccidial infection. In Experiment I, day-old Leghorn layer chicks were fed 3 experimental diets with 0, 0.1, or 0.2% whole yeast cell product (CitriStim®, ADM, Decatur, IL). At 21 d of age, birds were challenged with 1 × 105 live coccidial oocysts. Supplementation with whole yeast cell product decreased the fecal coccidial oocyst count at 7 (P = 0.05) and 8 (P < 0.01) d post-challenge. In Experiment II, 27-week old Leghorn layer hens were fed 3 experimental diets with 0, 0.05 or 0.1% whole yeast cell product and challenged with 1 × 105 live coccidial oocysts on d 25 of whole yeast cell product feeding. Supplementation with whole yeast cell product decreased the coccidial oocyst count in the intestinal content (P < 0.01) at 5, 13, and 38 d post-coccidial challenge. Supplementation with whole yeast cell product increased relative proportion of Lactobacillus (P < 0.01) in the cecal tonsils 13 d post-coccidial challenge. Supplementation with whole yeast cell product decreased CD8+ T cell percentages (P < 0.05) in the cecal tonsils at 5 d post-coccidial challenge. In Experiment III, 32-week-old Leghorn layer hens were fed 3 experimental diets with 0, 0.1, or 0.2% whole yeast cell product and challenged with 1 × 105 live coccidial oocysts on d 66 of whole yeast cell product feeding. At 5 d post-coccidial challenge, whole yeast cell product supplementation down-regulated (P = 0.01) IL-10 mRNA amount. It could be concluded that supplementing whole yeast cell product can help minimize coccidial infection in both growing pullets and layer chickens.

Longitudinal assessment of T-lymphocyte subpopulations during generalized demodicosis in dogs and their relationship with remission.

BACKGROUND: The pathogenesis of canine generalized demodicosis is poorly understood but is thought to involve dysfunction of the immune system. Previous studies showed diminished CD4+ T lymphocyte counts in affected dogs, but none has evaluated this subpopulation through resolution of the disease. HYPOTHESIS/OBJECTIVES: In this longitudinal study, we tested whether quantification of CD4+ cells, CD8+ cells and the ratio of CD4+ to CD8+ cells are good indicators of immunological status and could be used as biomarkers of treatment efficacy and prognosis. ANIMALS: Sixteen dogs of several breeds with diagnoses of generalized demodicosis, plus 30 age/breedmatched healthy dogs. METHODS: Total lymphocytes, CD4+, CD8+ and CD4+:CD8+ ratio were quantified at four time points: at diagnosis, 30 days after diagnosis (during treatment), at first negative parasitological examination and at clinical cure. RESULTS: Absolute numbers of CD4+ cells were significantly lower in affected dogs at the time of diagnosis. Absolute numbers of CD4+ and CD8+ cells were significantly augmented in affected animals compared with control dogs after treatment was established, and this persisted until the first negative parasitological examination, at which time the CD4+ counts equalled those of the control group. CONCLUSIONS AND CLINICAL IMPORTANCE: Our findings suggest that longitudinal quantification of CD4+ and CD8+ T lymphocytes is a useful indicator of the efficacy of demodicosis treatment.

Influence of endogamy and mitochondrial DNA on immunological parameters in cattle.

BACKGROUND: Endogamy increases the risk of manifestation of deleterious recessive genes. Mitochondrial DNA allows the separation of American Zebu (Bos indicus and Bos taurus) and evaluate the effect of mitochondrial DNA on productive traits of cattle. However, the effect of endogamy and mitochondrial DNA (mtDNA) on the immune system remains unclear. The aim of this study was to evaluate the association between endogamy, mtDNA and immune parameters. RESULTS: A total of 86 cattle (43 cows and 43 calves) were used in this study. Age, endogamy, milk yield, and origin of mtDNA were measured and their influence on immunological parameters was evaluated. Older cows had increased CD4+ T cells, decreased CD21+ and γδhigh T cells as well as increased CD4+/CD8+ and T/B ratio. Multiple regression analysis indicated that endogamy in calves was associated with increased CD8+ T and CD21+ B lymphocytes, and decreased γδhigh T cells in peripheral blood. Cows with medium and lower endogamy had a lower percentage of B lymphocytes and γδlow T cells and cows with lower endogamy had higher levels of γδ T cells and γδhigh T cells, as well as the CD4+/CD48+ cell ratio. Calves with higher endogamy had higher levels of CD8+ T lymphocytes, whereas calves with lower endogamy had lower levels of γδlow T cells. CONCLUSIONS: These results demonstrated for the first time that endogamy influences the immune system of cattle.

Diagnostic utility of CD4%:CD8 low% T-lymphocyte ratio to differentiate feline immunodeficiency virus (FIV)-infected from FIV-vaccinated cats.

Antibody testing based on individual risk assessments is recommended to determine feline immunodeficiency virus (FIV) status, but ELISA and Western blot tests cannot distinguish between anti-FIV antibodies produced in response to natural infection and those produced in response to FIV vaccination. The aim of this cross-sectional study was to test the hypothesis that FIV-infected cats could be differentiated from FIV-vaccinated uninfected cats using lymphocyte subset results, specifically the CD4%:CD8(low)% T-lymphocyte ratio. Comparisons of the CD4%:CD8(low)% T-lymphocyte ratio were made among the following four groups: Group 1 - FIV-infected cats (n=61; FIV-antibody positive by ELISA and FIV PCR positive); Group 2 - FIV-uninfected cats (n=96; FIV-antibody negative by ELISA); Group 3 - FIV-vaccinated uninfected cats (n=31; FIV-antibody negative by ELISA before being vaccinated against FIV, after which they tested FIV ELISA positive); and Group 4 - FIV-uninfected but under chronic/active antigenic stimulation (n=16; FIV-antibody negative by ELISA; all had active clinical signs of either upper respiratory tract disease or gingival disease for ≥ 21 days). The median CD4%:CD8(low)% T-lymphocyte ratio was lower in Group 1 (1.39) than in each of the other three groups (Group 2 - 9.77, Group 3 - 9.72, Group 4 - 5.64; P<0.05). The CD4%:CD8(low)% T-lymphocyte ratio was also the most effective discriminator between FIV-infected cats and the other three groups, and areas under ROC curves ranged from 0.91 (compared with Group 4) to 0.96 (compared with Group 3). CD4%:CD8(low)% shows promise as an effective test to differentiate between FIV-infected cats and FIV-vaccinated uninfected cats.

Identification of immune traits correlated with dairy cow health, reproduction and productivity.

Detailed biological analyses (e.g. epidemiological, genetic) of animal health and fitness in the field are limited by the lack of large-scale recording of individual animals. An alternative approach is to identify immune traits that are associated with these important functions and can be subsequently used in more detailed studies. We have used an experimental dairy herd with uniquely dense phenotypic data to identify a range of potentially useful immune traits correlated with enhanced (or depressed) health and fitness. Blood samples from 248 dairy cows were collected at two-monthly intervals over a 10-month period and analysed for a number of immune traits, including levels of serum proteins associated with the innate immune response and circulating leukocyte populations. Immune measures were matched to individual cow records related to productivity, fertility and disease. Correlations between traits were calculated using bivariate analyses based on animal repeatability and random regression models with a Bonferroni correction to account for multiple testing. A number of significant correlations were found between immune traits and other recorded traits including: CD4(+):CD8(+) T lymphocyte ratio and subclinical mastitis; % CD8(+) lymphocytes and fertility; % CD335(+) natural killer cells and lameness episodes; and serum haptoglobin levels and clinical mastitis. Importantly these traits were not associated with reduced productivity and, in the case of cellular immune traits, were highly repeatable. Moreover these immune traits displayed significant between-animal variation suggesting that they may be altered by genetic selection. This study represents the largest simultaneous analysis of multiple immune traits in dairy cattle to-date and demonstrates that a number of immune traits are associated with health events. These traits represent useful selection markers for future programmes aimed at improving animal health and fitness.

Flow cytometric analysis of lymphocyte subset kinetics in Bali cattle experimentally infected with Jembrana disease virus.

Jembrana disease virus (JDV) is an unusual bovine lentivirus that causes an acute and sometimes fatal disease after a short incubation period in Bali cattle (Bos javanicus). The pathological changes occur primarily in lymphoid tissues, which feature proliferating lymphoblastoid-like cells predominantly throughout parafollicular (T-cell) areas, and atrophy of follicles (B-cell) areas. Five Bali cattle were experimentally infected with JDV and all developed typical clinical signs of Jembrana disease characterised by a transient febrile response, enlargement of superficial lymph nodes and a significant leukopenia. Flow cytometric analysis of PBMC during the acute (febrile) disease phase showed that the reduced number of lymphocytes was due to a significant decrease in both the proportion and absolute numbers of CD4(+) T cells, but not CD8(+) T-cells or CD21(+) B-cells. At the end of the febrile phase, total numbers of both CD8(+) T-cells and CD21(+) B-cells increased significantly, while CD4(+) T-cell numbers remained below normal values, resulting in a significantly reduced CD4(+):CD8(+) ratio. We speculate that the persistent depletion of CD4(+) T cells following JDV infection, through lack of CD4(+) T cell help to B cells, may explain the lack of production of JDV-specific antibodies for several weeks after recovery despite an increase in CD21(+) B cell numbers. Further, our previous data showing that IgG(+) plasma cells are targets for JDV infection, correlated with our current data demonstrating an increase in CD8(+) T cell numbers, supports the suggestion that anti-viral cytotoxic T cell or other cell-mediated immune responses may be critical in the recovery process, although this remains to be formally demonstrated for JDV.

Oxygenated drinking water enhances immune activity in pigs and increases immune responses of pigs during Salmonella typhimurium infection.

It has been considered that drinking oxygenated water improves oxygen availability, which may increase vitality and improve immune functions. The present study evaluated the effects of oxygenated drinking water on immune function in pigs. Continuous drinking of oxygenated water markedly increased peripheral blood mononuclear cell proliferation, interleukin-1ß expression level and the CD4(+):CD8(+) cell ratio in pigs. During Salmonella Typhimurium infection, total leukocytes and relative cytokines expression levels were significantly increased in pigs consuming oxygenated water compared with pigs consuming tap water. These findings suggest that oxygenated drinking water enhances immune activity in pigs and increases immune responses of pigs during S. Typhimurium Infection.

Immunopathological effects of experimental T-2 mycotoxocosis in broiler chicken co-infected with infectious bronchitis virus (IBV).

A total of 128 1-week-old chicks were classified into four groups; T-2 toxin fed (T-2), IBV infected (IBV), T-2 toxin fed and co-infected with IBV (T-2+IBV), and untreated (control) for a period of 6 weeks. Within their respective groups, the birds belonged to T-2 and T-2+IBV were exposed to 2 ppm of T-2 toxin contaminated feed for 6 weeks, and 0.2 ml of 10 EID(50) (10(5.69)/0.2 ml) inoculums of IBV isolate (India/LKW/56/IVRI/08) was used to challenge the chicks belonged to IBV alone and T-2+IBV groups after 3 weeks of the experiment. To study immunopathological effects, parameters such as lymphocyte stimulation indices (SI), haemagglutination inhibition, enzyme linked immunosorbant assay (ELISA), peripheral lymphocytes CD(4)(+) and CD(8)(+) analysis, and histopathological examination of lymphoid organs were done. Accordingly, SI values were significantly (P<0.05) lower in all the treatment groups as compared to control, however, the SI values of IBV infected group were significantly higher than the values in toxin fed groups. The mean HI titres to ND vaccine was significantly (P<0.05) lower in the toxin groups at all the intervals, and the antibody titres in IBV infected group were significantly (P<0.05) higher than that of T-2 toxin fed and co-infected with IBV group but were significantly (P<0.05) lower than the control at 21 (3) and 28 (10) days of toxin feeding (DTF) (days post infection (DPI)). Similarly, the mean IBV ELISA antibody titres in the toxin fed groups were significantly (P<0.01) reduced as compared with the IBV ELISA antibody titres of IBV infected but not toxin fed group, at all intervals. Peripheral CD(4)(+):CD(8)(+) ratios in T-2+IBV group and number of CD(4)(+) and CD(8)(+) peripheral lymphocytes in all treatment groups were significantly reduced as compared to the values in control birds. However, CD(4)(+):CD(8)(+) ratios of IBV infected group at 42 (21) DTF (DPI) were found significantly (P<0.05) higher than the values in control birds. Histopathologically, lymphoid organs (bursa of Fabricius, spleen, thymus, caecal tonsils and Harderian glands) showed moderate to severe necrosis (lymphocytolysis) and extensive lymphocyte depletion in all the toxin groups (T-2 and T-2+IBV groups) where the severity and extent of the lesions were more in T-2+IBV group. The findings of the present experiment revealed immunosuppressive effects of T-2 toxin and aggravated the pathology and pathogenesis of IBV infection.

Peripheral lymphocyte subsets as a prognostic indicator of mortality and morbidity in healthy dogs.

To evaluate the relationship among immune status and increased morbidity and mortality, peripheral blood lymphocytes (CD3(+), CD4(+), CD8(+) and CD21(+) cells) from 32 healthy dogs over 8 years of age were analyzed. Twenty-five of the 32 dogs were followed-up for 3 years after the analysis; and 14 dogs were found to be diseased, and nine dogs died. There was no notable difference between the ages of the dogs that died compared with the ones that survived. The relative percentage of CD4(+) and the CD4(+):CD8(+) ratio decreased notably in dogs falling ill compared with healthy dogs. The relative percentage of CD3(+) lymphocytes showed a notable decrease in dogs that died within 3 years in comparison with dogs that survived. In a discriminant analysis of morbidity and mortality, most patients were correctly classified as diseased or not and surviving or dead, respectively. These results indicate that the immunophenotypes of peripheral blood lymphocytes in older dogs offer promise as parameters for evaluating mortality and morbidity.

Renal disease in cats infected with feline immunodeficiency virus.

BACKGROUND: Feline immunodeficiency virus (FIV) and human immunodeficiency virus (HIV) infection cause similar clinical syndromes of immune dysregulation, opportunistic infections, inflammatory diseases, and neoplasia. Renal disease is the 4th most common cause of death associated with HIV infection. OBJECTIVE: To investigate the association between FIV infection and renal disease in cats. ANIMALS: Client-owned cats (153 FIV-infected, 306 FIV-noninfected) and specific-pathogen-free (SPF) research colony cats (95 FIV-infected, 98 FIV-noninfected). METHODS: A mixed retrospective/prospective cross-sectional study. Blood urea nitrogen (BUN), serum creatinine, urine specific gravity (USG), and urine protein:creatinine ratio (UPC) data were compared between FIV-infected and FIV-noninfected cats. In FIV-infected cats, total CD4+ and CD8+ T lymphocytes were measured using flow cytometry, and CD4+:CD8+ T lymphocyte ratio was calculated. Renal azotemia was defined as a serum creatinine ≥ 1.9 mg/dL with USG ≤ 1.035. Proteinuria was defined as a UPC > 0.4 with an inactive urine sediment. RESULTS: Among the client-owned cats, no association was detected between FIV infection and renal azotemia (P = .24); however, a greater proportion of FIV-infected cats were proteinuric (25.0%, 16 of 64 cats) compared to FIV-noninfected cats (10.3%, 20 of 195 cats) (P < .01). Neither neuter status nor health status were risk factors for proteinuria in FIV-infected cats, but UPC was positively correlated with the CD4+:CD8+ T lymphocyte ratio (Spearman's rho = 0.37, P = .01). Among the SPF research colony cats, no association was detected between FIV infection and renal azotemia (P = .21) or proteinuria (P = .25). CONCLUSIONS AND CLINICAL IMPORTANCE: Proteinuria but not azotemia was associated with natural FIV infection.

Improvements of immune status, intestinal integrity and gain performance in the early-weaned calves parenterally supplemented with L-alanyl-L-glutamine dipeptide.

Glutamine (Gln) is an important substrate for the innate immune cells including lymphocytes and macrophages. In this report, the effects of alanyl-glutamine dipeptide (Ala-Gln) on the naïve immune system, intestinal integrity and gain performance of early-weaned calves were investigated. Early-weaned Chinese Holstein calves were intravenously administered different dosages of Ala-Gln. The effects of Ala-Gln administration were ascertained by evaluating the blood for naïve T lymphocyte subpopulations, the concentrations of serum IgG, serum IgA and intestinal mucosal secretory IgA (s-IgA), the intestinal integrity, as well as the gain performance. Results demonstrated that intravenous administration of Ala-Gln dipeptide (1.01 g/kg×d(-1)) for 7 days had a positive effect on gain performance, intestinal integrity and the immune system. Calves administered doses of Ala-Gln displayed an improvement in gain performance and health status concurrent with increases in blood CD2(+) and CD4(+) lymphocytes, the ratio of CD4(+)/CD8(+), serum IgA and IgG, intestinal mucosal s-IgA while decreasing the occurrence of diarrhea. Moreover, we found that animals given the effective dose (1.01 g/kg×d(-1)) of Ala-Gln resulted in improved immune status and intestinal integrity relative to those given a lower (0.49 g/kg×d(-1)) or higher dose (1.99 g/kg×d(-1)) of Ala-Gln. These findings suggest that maintaining a certain concentration of plasma and/or tissue glutamine in the early stages of weaning is an effective alternative approach for improvement of growth performance in early-wean calves.

Use of recombinant interferon omega in feline retrovirosis: from theory to practice.

Type-I interferons (IFNs) are cytokines that have non-specific antiviral activity, participating mostly in innate defense mechanisms. Their administration has been proposed to treat several viral and immunomediated diseases as an immunomodulatory therapy. Due to its availability, recombinant human interferon-alpha (rHuIFN-α) has been studied in relation to feline retrovirosis, both in vitro and in vivo. However, IFNs are species-specific and antibodies have been shown to develop in response to the high rHuIFN-α doses necessary for an effective therapy. A recombinant feline IFN has been developed, which has been characterized as interferon-omega (rFeIFN-ω), designed to overcome these problems. Nonetheless, very few studies have been undertaken to evaluate its efficacy in cats naturally infected with FIV or FeLV. In an initial study, we here demonstrated that rFeIFN-ω can dramatically improve the clinical condition of infected cats, and induce improvement of hematologic parameters. Minor changes or no change was observed for hypergammaglobulinemia, CD4/CD8 ratio, proviral load, viremia and RT activity, suggesting that the overall effect of IFN was on innate immunity. More studies are needed in order to better understand its in vivo mechanisms.

Alterations of lymphocyte subpopulations in healthy dogs with aging and in dogs with cancer.

Changes in an individual's immune status are considered major contributing factors towards the morbidity of cancer and mortality of aging. To evaluate age-related changes in the immune status of dogs, the immunophenotypes (CD3, CD4, CD8 and CD21) of peripheral blood lymphocytes were measured in 160 healthy dogs aged from 1 to 17 years, and in 365 dogs with various tumors and at various stages. In healthy dogs, the absolute numbers of white blood cells, lymphocytes, and CD3(+), CD4(+) and CD21(+) lymphocytes decreased significantly with age. The relative percentages of lymphocytes and CD4(+) cells decreased significantly, while CD8(+) cells increased significantly with age. The CD4:CD8 ratio showed a significant age-related decrease. In contrast, dogs with tumors possessed significantly lower absolute numbers and relative percentages of all lymphocyte phenotypes, while the CD4:CD8 ratio was significantly higher than in age-matched controls. The relative percentages of CD3(+) and CD8(+) lymphocytes were significantly lower in dogs with distant metastases compared with dogs without metastases, and the CD4:CD8 ratio increased with advanced stage. These observations illustrate the significant changes in immune status with age and the presence of marked immunological defects in a large-scale study of dogs with advanced tumors.

Humoral and cellular immune responses in dogs with inapparent natural Leishmania infantum infection.

Molecular analysis, serology and immunophenotyping for T lymphocytes and their subsets, B lymphocytes and monocytes were performed on dogs naturally infected with Leishmania infantum. Animals were categorised as asymptomatic dogs I (AD-I), with negative serology and positive molecular results, and asymptomatic dogs II (AD-II), with positive serology and positive molecular results, and these were compared to symptomatic dogs (SD) and control dogs (CD). AD-I exhibited immunophenotypic features similar to those of CD, including isotype profiles and concentrations of monocytes. Similar biomarkers were found in AD-II and SD, such as, higher levels of immunoglobulins IgG, IgG2, IgM and IgA and higher concentrations of eosinophils. High frequencies of T lymphocytes and CD4(+) T cells were observed in both AD-I and AD-II compared to SD, whereas CD8(+) T cells were higher only in AD-II compared with SD. Analysis of B lymphocytes revealed an increased frequency of this cell type only in AD-II animals compared with SD. Asymptomatic dogs appear to have a dichotomous infection spectrum that can influence the humoral and cellular immunological status during canine visceral leishmaniasis.

Effects of n-6:n-3 polyunsaturated fatty acid ratio on heterophil: lymphocyte ratio and T lymphocyte subsets in the peripheral blood of the Yangzhou gosling.

The objective of this study was to determine the effects of dietary n-6:n-3 polyunsaturated fatty acid (PUFA) ratios on heterophil:lymphocyte (H:L) ratios and T cell subsets in the peripheral blood of the growing Yangzhou goose. A total of 160 healthy Yangzhou geese (0.407 ± 0.023 kg), aged 21 d, were randomly divided into 4 groups consisting of diets with different n-6:n-3 PUFA ratios: 12:1, 9:1, 6:1, and 3:1. White blood cell counts and H:L ratios were measured at 42, 56, and 70 d of age; T lymphocyte subsets were also analyzed using monoclonal antibodies and flow cytometry simultaneously. The results showed that differences were found in white blood cells between groups (P < 0.05) but not within groups (P > 0.05), and white blood cells of 70-d-old geese were significantly higher than those of other ages within the same group. The H:L ratios of all groups decreased as age increased, and differences were found in 6:1 and 3:1 groups between ages (P < 0.05). As for the comparison of H:L ratio between treatments, 6:1 and 3:1 were lower than the other 2 groups at 42 or 70 d of age (P < 0.05), and 3:1 was lower than the other 3 groups at 56 d of age (P < 0.05). Moreover, H:L ratios had a tendency to decrease with the decrease of n-6:n-3 PUFA ratios. Furthermore, cluster of differentiation (CD) 3(+) and CD8(+) increased gradually, whereas CD4(+) and CD4(+):CD8(+) ratios first increased and then decreased as age increased. Differences were detected in T cell subsets among ages (P < 0.05). In addition, CD4(+) and CD4(+):CD8(+) ratios were affected remarkably by n-6:n-3 PUFA ratios but CD3(+) and CD8(+) were not, and 6:1 had much higher CD4(+) percentage and CD4(+):CD8(+) ratio compared with the other groups (P < 0.05). Taken together, diets containing low n-6:n-3 PUFA ratios could decrease H:L ratios in the peripheral blood. Additionally, CD4(+) percentage and CD4(+):CD8(+) ratio were much higher in the 6:1 group, which could be relevant for improving positive immune responses for Yangzhou goslings from 42 to 70 d of age.

Relationship between cortisol response to stress and behavior, immune profile, and production performance of dairy ewes.

The existence of a relationship between cortisol levels, after an acute stress, and behavioral activities, immunological profile, and production performance in sheep was studied. An initial flock of 30 Comisana ewes was involved in the experiment, and each of the 30 ewes was individually subjected to an isolation test in a novel environment. Subsequently, from the initial flock, 2 groups of 8 Comisana ewes were each retrospectively selected, and the animals were divided, according to their cortisol concentration 10 min after the isolation test, into high cortisol (HC) ewes, having a peak of cortisol concentration >90 ng/mL (average: 119.3 ng/mL +/- 11.8), and low cortisol (LC) ewes having a peak of cortisol concentration <80 ng/mL (average: 52.4+/-11.8). During the isolation test, the behavior of each animal was video-recorded and behavioral activities were registered. Blood samples were collected before the isolation test, immediately after the test (10 min), and at 60, 120, 300 min, 24 h, and 48 h after the test to evaluate percentages of T-helper (CD4(+)) and T-cytotoxic (CD8(+)) cells, CD4(+)/CD8(+) ratio, and IL-1beta and IL-6 levels. The ewes were milked for 3 d after the isolation test to determine cortisol levels and IL-1beta and IL-6 concentrations in whey. Milk yield was recorded at each milking, and milk samples were analyzed for pH, nutritional parameters, renneting properties, and somatic cell count. During the isolation test, HC ewes exhibited a shorter duration of movement and fewer bleats than LC ewes. The average plasma IL-1beta concentration was higher in HC than in LC ewes. The average whey IL-1beta and IL-6 concentrations were higher in whey from HC ewes than in LC ewes. A positive correlation emerged between plasma and whey IL-1beta concentrations. The average CD4(+)/CD8(+) ratio in blood was lower in HC than in LC ewes. Time from isolation affected the CD4(+)/CD8(+) ratio: at 120 min, the CD4(+)/CD8(+) ratio increased compared with that at 10 min after isolation and then decreased until 300 min after isolation. On average, ewes with low cortisol concentrations showed higher milk production and lower SCC than ewes with high cortisol concentrations. Results suggest that plasma cortisol concentration is connected to the behavioral response and immune competence of dairy ewes and cytokine concentrations. Both whey IL-1beta and IL-6 can be considered reliable indicators of the magnitude of hypothalamic-pituitary-adrenal axis activation. The stress-induced changes in CD4(+)/CD8(+) ratio are critical for controlling disease incidence and planning appropriate vaccination programs. High reactivity of the hypothalamic-pituitary-adrenal axis is also associated with a reduction in milk production and an increased predisposition to develop intramammary inflammatory processes.