Cow-to-cow variation in fibroblast response to a toll-like receptor 2/6 agonist and its relation to mastitis caused by intramammary challenge with Staphylococcus aureus

Abstract

Staphylococcus aureus is a common cause of chronic mammary gland infections in dairy cattle. However, the inflammatory response and duration of infection following pathogen exposure is variable between individual animals. To investigate interanimal differences in immune response, dermal fibroblast cultures were established from skin biopsies collected from 50 early lactation Holstein cows. The fibroblasts ability to produce IL-8 in response to a 24-h treatment with a synthetic toll-like receptor 2/6 agonist (Pam2CSK4) was used to assign a response phenotype to the animals. Five high-responding and 5 low-responding animals were then selected for an intramammary challenge with S. aureus to evaluate differences in the inflammatory response, chronicity of infection, and development of antibodies to the pathogen. All animals exhibited clinical symptoms of mastitis at 24h postchallenge. Animals previously classified as high responders experienced a greater inflammatory response characterized by elevated levels of milk somatic cell count, IL-8, and BSA following the challenge compared with low responders. In addition, antibodies toward the challenge strain of S. aureus reached higher levels in whey from the challenged gland of high responders compared with low responders. Despite the antibody response, all 5 high responders were chronically infected for the 6-wk duration of the study, whereas 2 of the low responders cleared the infection, although 1 of these did become reinfected. The observed differences between animals classified as low and high responders based on their fibroblast responsiveness suggests that this cell type can be used to further examine the causes of interanimal variation in response to mammary infection.

Variation in fibroblast expression of toll-like receptor 4 and lipopolysaccharide-induced cytokine production between animals predicts control of bacterial growth but not severity of Escherichia coli mastitis

“…Finally, no difference was measured between HR and LR animals in their E. coli-specific circulating IgG response and, importantly, antibody response did not correlate with ability to clear infection. This result agrees with previous literature stating that antibody response is not a major contributor to E. coli or S. aureus clearance within the mammary gland (Benjamin et al, 2015;Herry et al, 2017). The role of the adaptive response in determining between-cow differences in mastitis severity and duration should be the subject of future research, specifically the role of Th-17 cells, which have been shown to be the predominant T-cell responder in E. coli mastitis (Porcherie et al, 2016) and are associated with a primarily neutrophilic response.…”

“…Adjusted rank, ranging from −1.50 to 2.33 for IL-8 ( Figure 1A) and from −1.65 to 2.54 for IL-6 ( Figure 1B), was used as response variable. As with previous experiments (Kandasamy et al, 2011;Benjamin et al, 2015), a large range in response to LPS was measured in conditioned medium, with a significant correlation between LPS-induced IL-6 and IL-8 production (data not shown; R 2 = 0.28, P < 0.0001).…”

“…Nevertheless, a comparison of the area under the curve of each of the proteins showed a significant correlation (data not shown) in their production over the duration of the infection, such that animals with a prolonged cytokine response also exhibited more tissue damage, as measured by leakage of BSA into the milk. The concentration of IL-8 was significantly higher in HR animals at 24 h post-infection and likely contributed to the early elevation of milk SCC in HR animals at 12 h. Unfortunately, the lack of any real difference in milk IL-6 and IL-8 indicates that the fibroblast model may not accurately reflect the mammary gland environment, as suggested in our previous studies (Kandasamy et al, 2011;Benjamin et al, 2015). The strength and duration of pro-inflammatory cytokine and chemokine production in the mammary gland is, however, directly related to concentrations of serum albumin in the milk, a proxy measurement for tissue damage.…”

“…The HR animals challenged with E. coli also had a slower return to pre-infection milk production levels, and HR animals challenged with S. aureus had significantly elevated milk IL-8. In both experimental settings, no difference was measured in bacterial clearance, suggesting that HR animals do not benefit from the enhanced response and that the innate response contributes to mammary gland tissue damage, as measured by milk BSA levels, and lower milk production (Kandasamy et al, 2011;Benjamin et al, 2015).…”

Neonatal lipopolysaccharide exposure does not diminish the innate immune response to a subsequent lipopolysaccharide challenge in Holstein bull calves

“…Previous work from our laboratory and others has shown that dairy cattle manifest a wide range of immune responses following experimentally induced E. coli (Kornalijnslijper et al, 2003;Bannerman et al, 2004;Kandasamy et al, 2011) or Staphylococcus aureus (Schukken et al, 1999;Bannerman et al, 2004;Benjamin et al, 2015) mastitis. Some animals exhibit a much more robust inflammatory response during mastitis, characterized by high levels of IL-8, IL-6, and TNF-α, which can lead to severe clinical symptoms and damage to the mammary tissue.…”