Detection of interleukin-1 and interleukin-6 on cryopreserved bovine mammary epithelial cells in vitro.
This investigation was performed to determine whether primary cultures of mammary cells from lactating cows would sustain production of interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF), and express mRNA for cytokines interleukin (IL)-1alpha, IL-1beta, IL-2, IL-4, IL-6, IL-10, interferon (INF)-tau, TNF-alpha, and granulocyte-macrophage colony-stimulating factor (GM-CSF) by the reverse transcription-polymerase chain reaction (RT-PCR) in vitro. Cryopreserved mammary epithelial cells collected from cows at 1 week post calving were plated in collagen-coated 24-well culture plates (250,000 cells/well). IL-1 and IL-6 productions were measured using a A375 cell growth inhibition assay and a 7TD1 hybridoma proliferation assay, respectively. Production of IL-1 was demonstrated in mammary epithelial cells cultured with unsupplemented medium, but was not produced by cells cultured in medium supplemented with fetal bovine serum. IL-6 production in the conditioned medium was continued at steady level until day 14, whereas IL-6-like bioactivity was not detected in medium alone. TNF-like activity was not detectable in any experiments. This study also demonstrated the expression of mRNA for multiple cytokines including IL-1alpha, IL-1beta, IL-6, IL-10, TNF-alpha, and GM-CSF by RT-PCR in mammary cell cultures. The results indicate that bovine mammary epithelial cells of lactating cows produce IL-1 and IL-6 and have gene expression for multiple cytokines. This in vitro model will be useful to investigate the function and regulation of IL-1 and IL-6 in the lactating mammary gland.
Okada
H
H
Department of Veterinary Pathology, Rakuno Gakuen University, Ebetsu, Hokkaido, Japan.
Ito
T
T
Ohtsuka
H
H
Kirisawa
R
R
Iwai
H
H
Yamashita
K
K
Yoshino
T
T
Rosol
T J
TJ
D000818:Animals; D002417:Cattle; D002455:Cell Division; D002478:Cells, Cultured; D015925:Cryopreservation; D004847:Epithelial Cells; D004848:Epithelium / Q000276:immunology; D005260:Female; D007375:Interleukin-1 / Q000032:analysis / Q000096:biosynthesis*; D015850:Interleukin-6 / Q000032:analysis / Q000096:biosynthesis*; D007700:Kinetics; D007774:Lactation; D008321:Mammary Glands, Animal / Q000166:cytology* / Q000276:immunology*; D013997:Time Factors
D007375:Interleukin-1; D015850:Interleukin-6