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Published on: May 2, 2018
Interleukin 8 response by bovine mammary epithelial cells to lipopolysaccharide stimulation
N Boudjellab1, H S Chan-Tang, X Li
1Department of Animal Science, McGill University, Ste-Anne-de-Bellevue, Québec, Canada.
Insights
Bovine mammary epithelial cells (MAC-T) produce interleukin 8 (IL-8) when stimulated with lipopolysaccharide (LPS). This suggests a role for IL-8 in early mastitis stages and bacterial infection resolution.
Area of Science:
- Veterinary Immunology
- Cell Biology
- Bovine Mastitis Research
Background:
- Bovine mastitis is a significant economic concern in dairy farming.
- Interleukin 8 (IL-8) is a key chemokine involved in inflammatory responses.
- Understanding epithelial cell responses is crucial for managing mastitis.
Purpose of the Study:
- To investigate if a bovine mammary epithelial cell line (MAC-T) expresses IL-8 mRNA and protein.
- To determine if lipopolysaccharide (LPS) can stimulate IL-8 production in MAC-T cells.
Main Methods:
- MAC-T cells were stimulated with varying concentrations of LPS.
- Reverse transcriptase polymerase chain reaction (RT-PCR) was used to detect IL-8 mRNA.
- Sandwich ELISA quantified antigenic IL-8 protein in the cell culture medium.
Main Results:
- RT-PCR confirmed IL-8 mRNA expression in MAC-T cells within one hour of LPS stimulation.
- IL-8 protein production was detected and correlated with mRNA expression.
- IL-8 secretion increased in a dose- and time-dependent manner, peaking at 48 hours.
Conclusions:
- MAC-T cells secrete IL-8 in response to LPS stimulation.
- Mammary epithelial cells can be an early source of IL-8 during mastitis.
- IL-8 may play a critical role in resolving bacterial infections in the mammary gland.
Objective:
To determine whether an established bovine mammary epithelial cell line expresses interleukin 8 (IL-8) mRNA and synthesizes antigenic IL-8 in response to lipopolysaccharide (LPS) stimulation.
Sample Population:
A bovine mammary epithelial cell line (MAC-T).
Procedure:
mRNA was isolated from cells stimulated with graded concentrations of LPS. The first strand of IL-8 cDNA was synthesized, using a reverse transcriptase (RT) reaction with a specific oligonucleotide. Amplification of IL-8 cDNA was obtained by use of polymerase chain reaction (PCR). The MAC-T-derived antigenic IL-8 was quantified by use of a commercial anti-human IL-8 kit in a sandwich ELISA.
Results:
RT-PCR revealed expression of MAC-T-derived mRNA within the first hour after stimulation with LPS. Expression of IL-8 mRNA was correlated to production of IL-8 protein detected in medium by use of the sandwich ELISA. Amounts of antigenic IL-8 increased in a dose- and time-dependent manner, and were maximal (57 pg/ml) at 48 hours after stimulation with 20 microg of LPS/ml.
Conclusions:
MAC-T cells secrete IL-8 in response to stimulation with LPS in a dose- and time- dependent manner. The results were consistent with our hypothesis that mammary gland epithelial cells can be a source of IL-8 during the early stage of mastitis. Therefore, IL-8 may have a pivotal role in resolving bacterial infections.
