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Figure 8 C5a-induced IL-10 regulates IL-23 production by engaging Jnk and AP-1. (a) IL-10 production by medium- or HDM- treated BMDCs (BALB/c) in the presence of medium alone (Ctrl) or recombinant C3a (rC3a) or C5a (rC5a). *P < 0.05 and **P < 0.001 (one-way ANOVA). (b) IL-23 production by medium- or HDM- treated BMDCs (BALB/c) with (rC5a) or without (Ctrl) recombinant C5a in the presence of IgG1, anti-IL-10 (α-IL-10) or recombinant IL-10 (rIL-10). (c,d) Production of IL-23 (c) and IL-10 (d) by BMDCs (BALB/c) pretreated for 1 h with dimethyl sulfoxide (DMSO), SP600125, AS602868 or rapamycin, then treated with medium or HDM (100 μg/ml) in the presence (rC5a) or absence (Ctrl) of recombinant C5a. (e) Immunoblot analysis of c-Jun phosphorylated at Ser63 (p-c-Jun (Ser63)) and total c-Jun in medium- or C5a-treated RAW264.7 cells. (f) AP-1 activity in RAW264.7 cells treated with medium or various combinations of HDM, recombinant C5a and SP600125 (SP). (g) AP-1 activity in RAW264.7 cells stimulated with medium or various combinations of HDM, recombinant C5a, IgG1, anti-IL-10 and recombinant IL-10. (h) Il23a promoter activity in RAW264.7 cells cotransfected with pcDNA3.1 vector alone <t>(p-cDNA)</t> or vector containing c-Jun cDNA (p-c-Jun) or c-Fos cDNA (p-c-Fos), then treated with medium or HDM. (i) Jun and Fos mRNA in medium- or HDM-treated A/J and C3H/HeJ BMDCs, presented relative to S14 expression. Results are representative of two to three independent experiments (mean and s.e.m. of four to eight individual samples).
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Figure 8 C5a-induced IL-10 regulates IL-23 production by engaging Jnk and AP-1. (a) IL-10 production by medium- or HDM- treated BMDCs (BALB/c) in the presence of medium alone (Ctrl) or recombinant C3a (rC3a) or C5a (rC5a). *P < 0.05 and **P < 0.001 (one-way ANOVA). (b) IL-23 production by medium- or HDM- treated BMDCs (BALB/c) with (rC5a) or without (Ctrl) recombinant C5a in the presence of IgG1, anti-IL-10 (α-IL-10) or recombinant IL-10 (rIL-10). (c,d) Production of IL-23 (c) and IL-10 (d) by BMDCs (BALB/c) pretreated for 1 h with dimethyl sulfoxide (DMSO), SP600125, AS602868 or rapamycin, then treated with medium or HDM (100 μg/ml) in the presence (rC5a) or absence (Ctrl) of recombinant C5a. (e) Immunoblot analysis of c-Jun phosphorylated at Ser63 (p-c-Jun (Ser63)) and total c-Jun in medium- or C5a-treated RAW264.7 cells. (f) AP-1 activity in RAW264.7 cells treated with medium or various combinations of HDM, recombinant C5a and SP600125 (SP). (g) AP-1 activity in RAW264.7 cells stimulated with medium or various combinations of HDM, recombinant C5a, IgG1, anti-IL-10 and recombinant IL-10. (h) Il23a promoter activity in RAW264.7 cells cotransfected with pcDNA3.1 vector alone (p-cDNA) or vector containing c-Jun cDNA (p-c-Jun) or c-Fos cDNA (p-c-Fos), then treated with medium or HDM. (i) Jun and Fos mRNA in medium- or HDM-treated A/J and C3H/HeJ BMDCs, presented relative to S14 expression. Results are representative of two to three independent experiments (mean and s.e.m. of four to eight individual samples).

Journal: Nature immunology

Article Title: Complement-mediated regulation of the IL-17A axis is a central genetic determinant of the severity of experimental allergic asthma.

doi: 10.1038/ni.1926

Figure Lengend Snippet: Figure 8 C5a-induced IL-10 regulates IL-23 production by engaging Jnk and AP-1. (a) IL-10 production by medium- or HDM- treated BMDCs (BALB/c) in the presence of medium alone (Ctrl) or recombinant C3a (rC3a) or C5a (rC5a). *P < 0.05 and **P < 0.001 (one-way ANOVA). (b) IL-23 production by medium- or HDM- treated BMDCs (BALB/c) with (rC5a) or without (Ctrl) recombinant C5a in the presence of IgG1, anti-IL-10 (α-IL-10) or recombinant IL-10 (rIL-10). (c,d) Production of IL-23 (c) and IL-10 (d) by BMDCs (BALB/c) pretreated for 1 h with dimethyl sulfoxide (DMSO), SP600125, AS602868 or rapamycin, then treated with medium or HDM (100 μg/ml) in the presence (rC5a) or absence (Ctrl) of recombinant C5a. (e) Immunoblot analysis of c-Jun phosphorylated at Ser63 (p-c-Jun (Ser63)) and total c-Jun in medium- or C5a-treated RAW264.7 cells. (f) AP-1 activity in RAW264.7 cells treated with medium or various combinations of HDM, recombinant C5a and SP600125 (SP). (g) AP-1 activity in RAW264.7 cells stimulated with medium or various combinations of HDM, recombinant C5a, IgG1, anti-IL-10 and recombinant IL-10. (h) Il23a promoter activity in RAW264.7 cells cotransfected with pcDNA3.1 vector alone (p-cDNA) or vector containing c-Jun cDNA (p-c-Jun) or c-Fos cDNA (p-c-Fos), then treated with medium or HDM. (i) Jun and Fos mRNA in medium- or HDM-treated A/J and C3H/HeJ BMDCs, presented relative to S14 expression. Results are representative of two to three independent experiments (mean and s.e.m. of four to eight individual samples).

Article Snippet: Some cells were transfected with a 1,096–base pair fragment of the A/J Il23a promoter region inserted into the pGL3 vector (Promega) in combination with 50 ng empty pcDNA3.1 (Invitrogen), human c-Jun cDNA (SC118762; Origene) or mouse c-Fos cDNA (MC203181; Origene).

Techniques: Recombinant, Western Blot, Activity Assay, Plasmid Preparation, Expressing