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Ganglioside GQ1b enhances anti–double-stranded DNA antibody and IgG production of PBMCs from patients with systemic lupus erythematosus  Naoko Kanda,

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Presentation on theme: "Ganglioside GQ1b enhances anti–double-stranded DNA antibody and IgG production of PBMCs from patients with systemic lupus erythematosus  Naoko Kanda,"— Presentation transcript:

1 Ganglioside GQ1b enhances anti–double-stranded DNA antibody and IgG production of PBMCs from patients with systemic lupus erythematosus  Naoko Kanda, MD, PhD, Shinichi Watanabe, MD, PhD  Journal of Allergy and Clinical Immunology  Volume 105, Issue 3, Pages (March 2000) DOI: /mai Copyright © 2000 Mosby, Inc. Terms and Conditions

2 Fig. 1 Dose dependency for the effect of GQ1b on the antibody production by PBMCs from patients with SLE who were seropositive for anti-dsDNA antibody. PBMCs from an active or inactive patient were cultured with GQ1b for 5 days and were assayed for IgG anti-dsDNA antibody and total IgG production. Values are the means ± SD of triplicate cultures. The dashed line indicates the cutoff point for the presence or absence of anti-dsDNA. Data represent 6 separate experiments with PBMCs from 3 active patients and 3 inactive patients. Analysis of data was performed with 1-way ANOVA with Dunnet’s multiple comparison test (*P < .01 versus control levels). Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2000 Mosby, Inc. Terms and Conditions

3 Fig. 2 Stimulatory effect of GQ1b on antibody production by PBMCs from 12 active and 8 inactive patients with SLE who were seropositive for anti-dsDNA and 6 active and 4 inactive patients with SLE who were seronegative for this antibody. PBMCs were cultured in the presence (+) or absence (–) of GQ1b (10 μmol/L) for 5 days and were assayed for anti-dsDNA (A) and total IgG (B) production. Each point represents the mean amount of triplicate cultures from individual donors. The solid lines show the group means. The dashed lines indicate the cutoff point for the presence or absence of anti-dsDNA. Analysis of data was performed with the paired t test. Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2000 Mosby, Inc. Terms and Conditions

4 Fig. 3 Inhibition of the GQ1b effect by various antibodies. PBMCs from active patients with SLE who were seropositive for anti-dsDNA were cultured with (+) or without (–) GQ1b (10 μmol/L) in the presence or absence of various antibodies (each 10 μg/mL) for 5 days and were assayed for anti-dsDNA (A) and total IgG (B) production. The concentration of antibodies was determined as recommended by the manufacturers. Values are means ± SD (n = 6; 3 with CNS lupus and 3 without). *P < .001 versus controls with medium alone. †P < .001 versus cultures with GQ1b alone. Analysis of data was performed with 1-way ANOVA with Scheffe’s multiple comparison test. Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2000 Mosby, Inc. Terms and Conditions

5 Fig. 4 Dose dependency of the GQ1b effect on IL-6 (A) and IL-10 (B) production. T cells, monocytes, and CD5+ or CD5– B cells from an active patient with SLE who was seropositive for anti-dsDNA were cultured with indicated doses of GQ1b for 2 days and were assayed for cytokine production. Values are the means ± SD of triplicate cultures. Analysis of data was performed with 1-way ANOVA with Dunnet’s multiple comparison test (*P < .05 versus control levels). Data represent 6 separate experiments with peripheral blood cells from 6 active patients with SLE who were seropositive for anti-dsDNA (3 with CNS lupus and 3 without). Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2000 Mosby, Inc. Terms and Conditions

6 Fig. 5 IL-6 and IL-10 amounts in the supernatant from GQ1b- or medium-treated T cells (A) and the effects of the supernatants on B-cell antibody production (B) . T cells (1 × 106 cells/mL) from 6 active patients with SLE who were seropositive for anti-dsDNA were cultured with or without GQ1b (10 μmol/L) for 2 days. The culture supernatants were harvested, and 200-μL aliquots were assayed for IL-6 and IL-10. One hundred fifty-microliter aliquots of the supernatants were added to CD5+ or CD5– B cells (2 × 105 cells in 50 μL of fresh medium) from the identical donors, and the B cell fractions were cultured in triplicate. The identical B-cell fractions were simultaneously cultured with medium alone or medium containing GQ1b (7.5 μmol/L). The B-cell antibody production was evaluated after 5 days. Values are means ± SD (n = 6; 3 with CNS lupus and 3 without). Analysis of data in A was performed with the paired t test, and analysis of data in B was performed with the 1-way ANOVA with Scheffe’s multiple comparison test (*P < .05 versus control cultures with medium alone; †P < .05 versus cultures with the supernatant from medium-treated T cells). Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2000 Mosby, Inc. Terms and Conditions

7 Fig. 6 The effects of signal transduction inhibitors on GQ1b-induced IL-6 and IL-10 production. T cells (2 × 105/200 μL/well) from 6 active patients with SLE who were seropositive for anti-dsDNA were preincubated for 1 hour with medium alone or with medium containing signal transduction inhibitors before the addition of GQ1b (10 μmol/L). After 2 days, the culture supernatants were assayed for IL-6 (A) and IL-10 (B) . The inhibitors were staurosporine (STS ; final 1.5 nmol/L), calphostin C (Cal C ; 0.1 μmol/L), H-89 (0.1 μmol/L), herbimycin A (HerA , 1 μmol/L), and calbidazolium chloride (R24571 , 1 μmol/L). The concentration of each inhibitor was determined to inhibit the aimed signal specifically as recommended by the manufacturer. Values are means ± SD (n = 6; 3 with CNS lupus and 3 without). Analysis of data was performed with the 1-way ANOVA with Scheffe’s multiple comparison test (*P < .001 versus control cultures with medium alone; †P < .001 versus cultures with GQ1b alone). Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2000 Mosby, Inc. Terms and Conditions

8 Fig. 7 A schematic diagram for putative immunologic pathways activated by GQ1b. Ab, Antibody. Journal of Allergy and Clinical Immunology  , DOI: ( /mai ) Copyright © 2000 Mosby, Inc. Terms and Conditions

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