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Title: The use of dermal fibroblasts as a predictive tool of the TLR4 response pathway and its development in Holstein heifers

Author
item GREEN, B - University Of Vermont
item KANDASAMY, S - University Of Vermont
item Elsasser, Theodore
item KERR, D - University Of Vermont

Submitted to: Journal of Dairy Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/8/2011
Publication Date: 11/1/2011
Citation: Green, B.B., Kandasamy, S., Elsasser, T.H., Kerr, D.E. 2011. The use of dermal fibroblasts as a predictive tool of the TLR4 response pathway and its development in Holstein heifers. Journal of Dairy Science. 94: 5502-5514.

Interpretive Summary: The innate immune response potential of heifers was analyzed using dermal fibroblast cultures established from 15 animals when they were 5, 11, and 16 months of age. Production of interleukin-8 by the fibroblasts in response to lipopolysaccharide (LPS) revealed substantial between-animal variation. The relative ranking of heifers remained consistent with age, although there was an increased responsiveness with fibroblasts from older animals. Four low- and four high-responsive heifers were also challenged with intravenous LPS in vivo. High-responsive heifers exhibited higher plasma levels of the cytokines IL-8 and TNF-', suggesting that the fibroblast model is predictive of the animal response.

Technical Abstract: The innate immune system comprises the host’s first line of defense against invading pathogens, and variation in the magnitude of this response between animals has been shown to affect susceptibility to mastitis. The toll-like receptor (TLR) family of proteins initiates the response to invading bacteria, specifically with TLR4 recognizing lipopolysaccharide (LPS) of Gram-negative microbes. The underlying genetic variation in the TLR4 pathway leading to differential response is not well understood; therefore, the objective of this work was to determine the efficacy in which the response to LPS by dermal fibroblasts could be used to predict the actual systemic response of that animal to an intravenous endotoxin challenge. To accomplish this, dermal fibroblasts were isolated from 15 Holstein heifers at 5, 11, and 16 months of age, exposed to either LPS or IL-1', and the production of IL-8 in media quantified by ELISA. Animals were ranked based upon the magnitude of the fibroblast IL-8 response and 8 heifers were selected (4 low-, 4 high-responsive) for challenge with an intravenous bolus dose (0.5 'g/kg BW) of LPS. Overall between-animal variation in fibroblast IL-8 production following LPS or IL-1' was high, indicating appreciable differences in the TLR4 pathway of the animals. Ranking of the fibroblast responses was consistent across the three sampling times for each animal, however the absolute response increased with age at which the fibroblasts were obtained indicating a possible developmental aspect. Following systemic LPS challenge, heifers selected as high responders (HR) had higher plasma concentrations of TNF-' and IL-8 than heifers selected as low responders (LR). LR heifers had a stronger febrile response than HR however. The use of dermal fibroblasts under laboratory settings appears to represent a practical model for predicting the innate immune response in vivo, and could act as an important tool in mapping genetic differences of the TLR4 pathway.