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Intracellular IL-1 receptor antagonist (icIL-1Ra1) does not antagonize growth inhibitory effects of pre-IL-1α in SaOS-2 cells
Arthritis Research & Therapy volume 7, Article number: P91 (2005)
Background
IL-1α is synthesized as a precursor (preIL-1α), which is processed into mature IL-1α and a N-terminal propeptide by calpain-like proteases. Besides its classical effects elicited upon IL-1 receptor binding, preIL-1α exerts intracellular functions, including the modulation of cell growth and apoptosis. Nuclear translocation of preIL-1α, mediated by the N-terminal propeptide, is required for these effects. IL-1 receptor antagonist (IL-1Ra) inhibits the classical effects of IL-1 by preventing the interaction of IL-1 with its receptor. Four different isoforms of IL-1Ra have been described, of which one is secreted and three others are intracellular (icIL-1Ra1, icIL-1Ra 2, icIL-1Ra 3). Due to their intracellular localization, icIL-1Ras cannot interact with cell surface receptors and have been suggested to carry out specific functions inside cells. The description of nuclear functions for preIL-1α suggested that icIL1Ra variants might antagonize intracellular effects of preIL-1α.
Objective and methods
The aim of this study was to investigate effects of preIL-1α and icIL-1Ra1 on cell growth using stably transfected SaOS-2 osteosarcoma cells. IL-1α and IL-1Ra expression was quantified by ELISA and their localization was examined by immunofluorescence. Cell counts, lactate dehydrogenase activity and [3H]-thymidine incorporation were used to monitor cell growth.
Results
IL-1α expression ranged from 5.3 to 9.4 ng/106 cells in culture supernatants and from 39.8 to 87.3 ng/106 cells in lysates of preIL-1α transfected SaOS-2 cells. Immunostaining showed nuclear localization of preIL-1α, which was not modified by co-transfection of icIL-1Ra1. Expression levels of IL-1Ra ranged from 53 to 219 ng/106 cells in supernatants and from 1678 to 4414 ng/106 cells in lysates of icIL-1Ra1 transfected cells. IL-1Ra staining was essentially cytoplasmic. Transfection of SaOS-2 cells with preIL-1α significantly decreased cell growth, as indicated by reduced cell counts and lactate dehydrogenase activity. PreIL-1α also decreased thymidine incorporation, indicating an inhibition of cell proliferation. In contrast, addition of exogenous mature IL-1α (500 pg/ml) had no inhibitory effect on SaOS-2 proliferation, suggesting that the effect of preIL-1α on cell growth is intracellular. Transfection of SaOS-2 cells with icIL-1Ra1 alone did not affect cell growth. Moreover, co-expression of icIL-1Ra1 did not reverse the growth inhibitory effect of preIL-1α. However the induction of IL-6 secretion, a classical, receptor-mediated effect of preIL-1α, was inhibited in icIL-1Ra1 co-transfected cells.
Conclusions
Expression of preIL-1α decreased the growth of SaOS-2 cells. Co-transfection of icIL-1Ra1 did not antagonize this effect. These observations suggest that intracellular effects of preIL-1α are not necessarily susceptible to inhibition by icIL-1Ra.
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Palmer, G., Trolliet, S., Talabot-Ayer, D. et al. Intracellular IL-1 receptor antagonist (icIL-1Ra1) does not antagonize growth inhibitory effects of pre-IL-1α in SaOS-2 cells. Arthritis Res Ther 7, P91 (2005). https://doi.org/10.1186/ar1612
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Keywords
- Osteosarcoma
- Thymidine Incorporation
- Growth Inhibitory Effect
- Intracellular Effect
- Lactate Dehydrogenase Activity
