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Regulation of CD154-induced interleukin-12 production in synovial fluid macrophages.

Möttönen M, Isomäki P, Luukkainen R, Lassila O - Arthritis Res. (2002)

Bottom Line: CD40 ligation induced the secretion of IL-12p40 but not IL-12p70.The expression of p40 mRNA was increased significantly and remained upregulated after CD40 ligation, whereas the increase of p35 transcript expression was observed only transiently and at a lower level.These results provide novel information about the regulation of IL-12p70 production and the function of the cytokine network in RA.

View Article: PubMed Central - HTML - PubMed

Affiliation: Turku Graduate School of Biomedical Sciences, Turku University, Finland. milja.mottonen@utu.fi

ABSTRACT
Interleukin (IL)-12, being a major cytokine that induces T helper (Th) 1 differentiation and inflammatory response, has been postulated to be an important mediator of synovial inflammation in rheumatoid arthritis (RA). However, the regulation of IL-12 production in RA has not been elucidated. Our knowledge is mainly based on studies of the production of IL-12p40 and not the functional IL-12p70 heterodimer. We have studied the CD154-induced IL-12p40 and IL-12p70 production by synovial fluid (SF) macrophages from patients with RA. CD40 ligation induced the secretion of IL-12p40 but not IL-12p70. The observed increase in IL-10 and tumor necrosis factor (TNF)-alpha production indicated that SF macrophages responded to CD40 ligation. The expression of p40 mRNA was increased significantly and remained upregulated after CD40 ligation, whereas the increase of p35 transcript expression was observed only transiently and at a lower level. We further observed that dendritic cells (DCs) derived in vitro from SF macrophages produced IL-12p70. Most importantly, IL-4 and IL-13 primed SF macrophages to produce IL-12p70, whereas IFN-gamma was not observed to activate IL-12p70 production in these cells, in contrast with normal peripheral blood monocytes. These results provide novel information about the regulation of IL-12p70 production and the function of the cytokine network in RA.

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Production of IL-12p40 and p70 by synovial fluid (SF) macrophages precultured with IL-4 and IL-13. SF macrophages were cultured in the presence of IL-4 (n = 9) or IL-13 (n = 6) (100 U/ml) or in medium only (n = 9) for 72 hours before addition of CD154 transfectants (at a ratio of 1:5 transfectants:SF macrophages) or control J558L cells (1:5) for a further 24 hours. Supernatants were collected at the end of the culture and the production of IL-12p70 (a) and p40 (b) was determined using ELISA. Individual values from each experiment are shown; each patient is represented by the same symbol in the two panels. Cells stimulated with control J558L did not produce IL-12p40 nor p70 (data not shown).
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Figure 3: Production of IL-12p40 and p70 by synovial fluid (SF) macrophages precultured with IL-4 and IL-13. SF macrophages were cultured in the presence of IL-4 (n = 9) or IL-13 (n = 6) (100 U/ml) or in medium only (n = 9) for 72 hours before addition of CD154 transfectants (at a ratio of 1:5 transfectants:SF macrophages) or control J558L cells (1:5) for a further 24 hours. Supernatants were collected at the end of the culture and the production of IL-12p70 (a) and p40 (b) was determined using ELISA. Individual values from each experiment are shown; each patient is represented by the same symbol in the two panels. Cells stimulated with control J558L did not produce IL-12p40 nor p70 (data not shown).

Mentions: IL-4 and IL-13 have been shown to enhance IL-12p70 production by DCs and monocytes/macrophages [1,6,7]. We have earlier shown that IL-4 increases the ability of SF macrophages to function as antigen-presenting cells [28]. Therefore, we studied whether IL-4 and IL-13 would induce IL-12p70 production. SF macrophages were precultured in the presence of IL-4 or IL-13 for 72 hours before stimulation with CD154 transfectants for 24 hours. IL-4 significantly primed the production of IL-12p40 and p70 (Fig. 3). Priming with IL-13 induced the production of IL-12p40 significantly. In addition, IL-13 primed the production of IL-12p70 in four of six samples studied. In two samples, the induction was clear, from <8 pg/ml to 255 and 384 pg/ml, and in two samples from <8 pg/ml to 23 and 28 pg/ml.


Regulation of CD154-induced interleukin-12 production in synovial fluid macrophages.

Möttönen M, Isomäki P, Luukkainen R, Lassila O - Arthritis Res. (2002)

Production of IL-12p40 and p70 by synovial fluid (SF) macrophages precultured with IL-4 and IL-13. SF macrophages were cultured in the presence of IL-4 (n = 9) or IL-13 (n = 6) (100 U/ml) or in medium only (n = 9) for 72 hours before addition of CD154 transfectants (at a ratio of 1:5 transfectants:SF macrophages) or control J558L cells (1:5) for a further 24 hours. Supernatants were collected at the end of the culture and the production of IL-12p70 (a) and p40 (b) was determined using ELISA. Individual values from each experiment are shown; each patient is represented by the same symbol in the two panels. Cells stimulated with control J558L did not produce IL-12p40 nor p70 (data not shown).
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Related In: Results  -  Collection

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Figure 3: Production of IL-12p40 and p70 by synovial fluid (SF) macrophages precultured with IL-4 and IL-13. SF macrophages were cultured in the presence of IL-4 (n = 9) or IL-13 (n = 6) (100 U/ml) or in medium only (n = 9) for 72 hours before addition of CD154 transfectants (at a ratio of 1:5 transfectants:SF macrophages) or control J558L cells (1:5) for a further 24 hours. Supernatants were collected at the end of the culture and the production of IL-12p70 (a) and p40 (b) was determined using ELISA. Individual values from each experiment are shown; each patient is represented by the same symbol in the two panels. Cells stimulated with control J558L did not produce IL-12p40 nor p70 (data not shown).
Mentions: IL-4 and IL-13 have been shown to enhance IL-12p70 production by DCs and monocytes/macrophages [1,6,7]. We have earlier shown that IL-4 increases the ability of SF macrophages to function as antigen-presenting cells [28]. Therefore, we studied whether IL-4 and IL-13 would induce IL-12p70 production. SF macrophages were precultured in the presence of IL-4 or IL-13 for 72 hours before stimulation with CD154 transfectants for 24 hours. IL-4 significantly primed the production of IL-12p40 and p70 (Fig. 3). Priming with IL-13 induced the production of IL-12p40 significantly. In addition, IL-13 primed the production of IL-12p70 in four of six samples studied. In two samples, the induction was clear, from <8 pg/ml to 255 and 384 pg/ml, and in two samples from <8 pg/ml to 23 and 28 pg/ml.

Bottom Line: CD40 ligation induced the secretion of IL-12p40 but not IL-12p70.The expression of p40 mRNA was increased significantly and remained upregulated after CD40 ligation, whereas the increase of p35 transcript expression was observed only transiently and at a lower level.These results provide novel information about the regulation of IL-12p70 production and the function of the cytokine network in RA.

View Article: PubMed Central - HTML - PubMed

Affiliation: Turku Graduate School of Biomedical Sciences, Turku University, Finland. milja.mottonen@utu.fi

ABSTRACT
Interleukin (IL)-12, being a major cytokine that induces T helper (Th) 1 differentiation and inflammatory response, has been postulated to be an important mediator of synovial inflammation in rheumatoid arthritis (RA). However, the regulation of IL-12 production in RA has not been elucidated. Our knowledge is mainly based on studies of the production of IL-12p40 and not the functional IL-12p70 heterodimer. We have studied the CD154-induced IL-12p40 and IL-12p70 production by synovial fluid (SF) macrophages from patients with RA. CD40 ligation induced the secretion of IL-12p40 but not IL-12p70. The observed increase in IL-10 and tumor necrosis factor (TNF)-alpha production indicated that SF macrophages responded to CD40 ligation. The expression of p40 mRNA was increased significantly and remained upregulated after CD40 ligation, whereas the increase of p35 transcript expression was observed only transiently and at a lower level. We further observed that dendritic cells (DCs) derived in vitro from SF macrophages produced IL-12p70. Most importantly, IL-4 and IL-13 primed SF macrophages to produce IL-12p70, whereas IFN-gamma was not observed to activate IL-12p70 production in these cells, in contrast with normal peripheral blood monocytes. These results provide novel information about the regulation of IL-12p70 production and the function of the cytokine network in RA.

Show MeSH
Related in: MedlinePlus