Hierarchical clustering of expression profiles.

<p>Mammary gland tissue was subjected to microarray analysis at 24(n = 2) and 48 h (n = 3) post challenge with 100 cfu <i>E. coli</i>. Per individual cow one quarter served as un-challenged control (green), one quarter was challenged but not treated (red) and two challenged quarters were treated at 4, 12, 24 and 36 h post challenge with 300 mg cefapirin (dark gray) or 300 mg cefapirin plus 20 mg prednisolone (light gray). Gene expression profiles of individual samples were compared and the degree of similarity of the expression patterns is shown as a heatmap of the distance matrix (metric: Pearson correlation). The darker the color (smaller distance) the more similar are the expression pattern between any two samples.</p

Levels of chemokines in milk whey.

<p>A) Levels (pg/ml) of chemokines (CXCL1, CXCL2 and CXCL8) in milk whey were determined by ELISA at time of (0), 6, 12, 18, 24 (n = 6) and 48 h (n = 3) post challenge with 100 cfu <i>E. coli</i>. Per cow one quarter was left unchallenged (open circles) and one quarter was challenged but not treated (filled circles), one quarter was challenged and cefapirin treated (open triangles) and one quarter was challenged and treated with cefapirin plus prednisolone (filled squares) at 4, 12 and 24 h post challenge. Data are shown as LSM of chemokine levels plus standard errors. In the case of CXCL8 LSM of the log10 transformed levels are shown. Statistically significant differences (P<0.05) are indicated by <b>*</b> (significantly different to all other groups), <b>†</b> (significantly different to not infected, not treated) and <b>‡</b> (significantly different to infected, cefapirin treated). B) Example of H&E stained sections from four quarters of one animal at 24 h post challenge. Arrows indicate PMN in tissue.</p

Negative fold inductions of DE genes from challenged treated quarters compared to challenged not treated quarters associated to enriched pathways.

<p>^a log2 fold induction compared to challenged, not treated quarters.</p

Significantly enriched Gene Ontology terms in up-regulated DE gene list (positive FI) and down-regulated DE gene list (negative FI) in challenged not treated quarters compared to not challenged quarters.

<p>^a significance of gene term enrichment examined by modified Fisher’s exact test.</p

Validation of microarray results for selected genes by qPCR.

<p>Expression levels of DE genes in treated quarters belonging to significantly down-regulated pathways were confirmed by qPCR. Data are shown as median plus range of relative expression in challenged, not treated quarters (filled circle), challenged, cefapirin treated quarters (open triangle) and challenged, cefapirin plus prednisolone treated quarters (filled square). A) Samples obtained 24 h post challenge (n = 2). B) Samples obtained 48 h post challenge (n = 3). Statistically significant differences (P<0.05) are indicated by *.</p

Primer sequences for quantitative RT-PCR.

<p>^a bp, length of amplicons in base pairs.</p><p>^b NCBI National Center for Biotechnology Information.</p