The effect of negative energy balance on bovine blood PMN gene expression

Our objectives were to compare gene expression profiles in neutrophils (PMN) during a Streptococcus uberis mastitis challenge between lactating cows subjected to feed restriction to induce negative energy balance (NEB; n = 5) and cows fed ad libitum to maintain positive energy balance (PEB; n = 5). The NEB cows were feed-restricted to 60% of calculated net energy for lactation requirements for 7 d, whereas PEB cows were fed the same diet for ad libitum intake. After 5 d of feed restriction, one rear mammary quarter of each cow was inoculated with 5,000 cfu of Streptococcus uberis (strain O140J). Blood PMN were isolated at 24 h post-inoculation from all cows for RNA extraction and microarray analysis. NEB resulted in 94 differentially expressed genes compared with PEB. Of these, 51 genes were down-regulated, including genes involved with antigen presentation (HLADRA and HLAA), respiratory burst (SOD1), and the pro-inflammatory response (TNFA and IRAK-1). The most affected genes up-regulated by NEB (n = 43) included IL1R2 and IL6, toll-like receptors (TLR2 and TLR4), and THY1. Network analysis by Ingenuity Pathway Analysis ® revealed that TNFA was associated with the expression of numerous differentially expressed genes involved with immune response in NEB cows compared with PEB cows. Energy balance alters PMN expression of several genes involved with immune response, which provides new information on transcriptomic mechanisms associated with post-partal NEB and immune response during early lactation. Briefly, 10 multiparous Holstein cows in PEB and past peak lactation were used for this study. The NEB cows were feed-restricted to 60% of calculated NEL requirements for 7 d, whereas PEB cows were fed the same diet for ad libitum intake. After 5 d of feed restriction, one rear mammary quarter of each cow was inoculated with 5,000 cfu of Streptococcus uberis (strain O140J). Blood PMN were isolated at 24 h post-inoculation from all cows. A 13,257-oligonucleotide (70-mers) array was used for transcript profiling. Cy3- and Cy5 labelled cDNA from PMN and a reference standard were used for hybridizations. All samples were hybridized to duplicate slides with reverse labeling (20 microarrays total).