Expression analysis of candidate genes for chronic subclinical mastitis in Norwegian Red cattle.

Chronic subclinical mastitis (SCM) is characterized by a long-term inflammation in the udder with high somatic cell count (SCC) in milk. Previously, several novel alternative SCM traits for Norwegian Red (NR) cattle have been defined to improve breeding strategies against chronic SCM. Quantitative trait loci and candidate genes affecting chronic SCM in NR have been identified. The aim of this study was to analyze the expression profiles of 14 selected candidate genes (RAD17, ACOT2, ACOT4, FOS, CXCL1, CXCL8, CCNB1, CDK7, TGFB3, SEL1L, STAT4, C6, GLI2, and SLC18A2). Twenty healthy NR cows with official genomic estimated breeding values (GEBV) for lactation average somatic cell scores (LSCS) were selected. Ten cows had high GEBV for LSCS (cows with low probability to have high SCC in milk during lactation) and 10 cows had low GEBV for LSCS (cows with high probability of having high SCC in milk). We isolated RNA from unstimulated peripheral blood mononuclear cells from these. Two out of the 14 analyzed genes showed significantly different results between groups. The group with high GEBV for LSCS displayed significantly higher expression of the CXCL1 gene than the low GEBV group. Grouping by lactation stage revealed significant differential expression of the FOS gene, with higher expression in early lactation (2-3 mo after calving) compared with late lactation (7-8 mo after calving). In addition, flow cytometry was performed on the peripheral blood mononuclear cells samples to analyze if number and type of isolated cells influenced the gene expression in the groups. The results in the current study provide identified genes that can be considered as possible candidate genes for chronic SCM in NR cows.

Surgical stress influences cytokine content in autologous conditioned serum.

REASONS FOR PERFORMING STUDY: No recommendations have been made regarding the relative timing of blood collection for autologous conditioned serum (ACS) preparation and surgical procedures. OBJECTIVES: 1) To identify effects of surgical stress on cytokine levels in ACS, 2) identify haematological markers for prediction of cytokine production in ACS and 3) investigate the necessity for specialised ACS containers when preparing a cytokine-rich serum. STUDY DESIGN: Experimental in vitro study. METHODS: Blood was drawn from 15 stallions admitted for elective castration preoperatively and 22-24 h post operatively and incubated in ACS containers and plastic vacutainer tubes containing Z Serum Clot Activator. Concentrations of interleukin (IL)-1 receptor agonist (IL-1Ra), IL-10, IL-1ß, tumour necrosis factor (TNF)-α, insulin-like growth factor (IGF)-1 and transforming growth factor (TGF)-ß were determined in all serum samples and compared between preparation methods and sampling time by ANOVA. Changes in cytokine levels induced by incubation, defined as delta cytokine, were calculated by subtracting the baseline levels from the levels in incubated samples. Based on post operative serum amyloid A (SAA), horses were grouped into 'mild', moderate' and 'marked' surgical stress; delta cytokine levels in post operative samples were compared between these groups by ANOVA. RESULTS: Delta IGF-1 was significantly lower in post operative samples compared with preoperative. Horses in the 'marked' surgical stress group had significantly lower delta IL-1Ra and delta TGF-ß than the 'moderate' group and significantly lower delta IGF-1 than the 'mild' group. No association between cytokine levels and haematology variables were identified. Cytokine levels were comparable between serum prepared in blood tubes and in specialised ACS containers. CONCLUSIONS: Surgical stress influences the cytokine content in ACS. Useful predictors of cytokine production in ACS were not identified. Specialised ACS containers may not be necessary for preparation of a cytokine-rich serum.

Modulation of leukocyte populations and immune responses in sheep experimentally infected with Anaplasma (formerly Ehrlichia) phagocytophilum.

Anaplasma phagocytophilum infection in sheep is characterized by an immune suppression as indicated by impaired antibody response, reduced lymphocyte response and reduced oxidative burst. The effect of A. phagocytophilum infection on leucocyte populations, especially lymphocytes, was therefore investigated in six sheep experimentally infected with A. phagocytophilum, and compared with leucocyte populations from control animals.To investigate the ability of the infection to interfere with the cellular and humoral responses to specific antigens, the animals were vaccinated with commercial vaccines at the time of experimental infection, and monitored for 56 days. There were reduced percentages of gammadelta T-cells and CD4+ T-cells in peripheral blood of infected animals throughout the study period, and these cell populations showed a down-regulation of CD25 expression; while there was a relative increase in CD8+ T-cells. The reduction in CD25+ gammadelta T-cells involved a subpopulation of WC1+ gammadelta T-cells. During the first 2 weeks of the study there were reduced percentages of B-cells and leukocytes expressing MHC II and CD11b, though this decrease changed to a relative increase later in the study. The relative reductions in leucocyte populations corresponded with the observed leucopenia during the first 3 weeks post-infection, which involved lymphocyte, neutrophil and eosinophil subsets [Vet. Immunol. Immunopathol. 86 (2002) 183]. There was a reduced expression of CD11b and CD14 on granulocytes during the first 2 weeks of the study, which corresponded with the previously reported leucopenia involving neutrophils and eosinophils. Antibody responses to vaccines, lymphocyte in vitro proliferative responses to antigens and mitogens, and in vitro IFN-gamma responses to antigens were reduced up to 4 weeks after infection.

Immunosuppression in sheep experimentally infected with Ehrlichia phagocytophila.

The effect of tick-borne fever (TBF) on antibody formation and lymphocyte proliferation in sheep was studied following experimental infection with Ehrlichia phagocytophila. All infected sheep developed fever within three to four days. The sheep recovered clinically within eight days. Both infected and non-infected control sheep were immunised twice with different antigens, that is, on days 9 and 35 following the experimental infection. The levels of antibodies produced against tetanus toxoid and influenza virus in the infected sheep were significantly lower than in the control animals. The findings indicated that a TBF-infection may impair both primary and secondary antibody responses for up to six weeks. Immunisation with Actinomyces pyogenes resulted in significantly higher antibody titres in the TBF-infected group than in the control group, as measured by an enzyme-linked immunosorbent assay (ELISA). It is believed that TBF-induced neutropenia may lead to increased exposure to A pyogenes-antigens and thereby enhance antibody production. Antibodies to E phagocytophila were measured by the indirect fluorescent antibody test and by an ELISA. The inoculated sheep responded with the formation of antibodies to E phagocytophila at one week (P < 0.025), and showed a peak response at four weeks (P < 0.0005) after inoculation. The antibody titre decreased between four and six weeks, but was still high at six weeks (P < 0.0005). The lymphocyte responses to phytohaemagglutinin (PHA) concanavalin A (Con A) and pokeweed mitogen (PWM) were lower than in the control group and this difference was significant at most time points after infection.(ABSTRACT TRUNCATED AT 250 WORDS)