Modulation of interleukin-1 receptors followed by endotoxin lipopolysaccharide treatment in the mouse AtT-20 pituitary tumor cell line.
OBJECTIVE
We have previously reported the characterization and regulation of interleukin-1 (IL-1) receptors utilizing [125I]IL-1 binding assay in male C57BL/6 mice and the mouse AtT-20 pituitary tumor cells. In the present study, we examine IL-1 receptors using an immunoblotting method to further characterize the mechanisms regulating the interactions of IL-1 receptors with endotoxin, lipopolysaccharide (LPS).


METHODS
We established Western blotting for IL-1 receptors using AtT-20 mouse pituitary tumor cells.


RESULTS
Several bands were seen; however, only the 105-kD band was neutralized with a 5-fold excess of IL-1 receptor- blocking peptides, suggesting that this band is specific for IL-1 receptors. Next, we investigated the effect of LPS and IL-1beta on IL-1 receptors. Treatment of AtT-20 cells with 0.01 microg/ml of LPS did not affect IL-1 receptors. In contrast, 1 microg/ml of LPS significantly increased IL-1 receptors in AtT-20 cells compared with the control group. In addition, [125I]IL-1beta binding was markedly increased followed by 1 microg/ml of LPS. In contrast, 1 nM recombinant human IL-1beta significantly decreased IL-1 receptors in AtT-20 cells compared with the control group although treatment of AtT-20 cells with 0.01 nM IL-1beta did not affect IL-1 receptors. LPS (0.1 and 1 microg/ml) did not affect IL-1beta concentrations in the medium of AtT-20 cell culture. IL-1beta concentrations in the homogenates from AtT-20 cells were significantly decreased after 1 microg/ml of LPS treatment but not after 0.01 microg/ml LPS.


CONCLUSIONS
These data demonstrate that LPS and IL-1beta differentially modulate IL-1 receptors in AtT-20 cells and LPS-induced modulation of IL-1 receptors may provide a novel mechanism for the actions of LPS to alter pituitary function during endotoxemia. Additional in vivo studies are necessary to determine the physiological relevance of this in vitro phenomenon.
Kumagai
Chizuru
C
Second Department of Internal Medicine, Kochi Medical School, Nankoku, Kochi, Japan. kumagaic@kochi-ms.ac.jp
Takao
Toshihiro
T
Matsumoto
Reiko
R
Asaba
Koichi
K
Hashimoto
Kozo
K
D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects / Q000276:immunology; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects / Q000276:immunology; D019446:Endotoxemia / Q000276:immunology / Q000503:physiopathology; D007155:Immunologic Factors / Q000276:immunology / Q000494:pharmacology*; D007375:Interleukin-1 / Q000276:immunology / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000276:immunology / Q000494:pharmacology*; D008297:Male; D051379:Mice; D010911:Pituitary Neoplasms / Q000188:drug therapy / Q000276:immunology* / Q000378:metabolism; D017472:Receptors, Interleukin-1 / Q000187:drug effects / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000494:pharmacology
D007155:Immunologic Factors; D007375:Interleukin-1; D008070:Lipopolysaccharides; D017472:Receptors, Interleukin-1; D011993:Recombinant Fusion Proteins