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Toll-like receptor 7–induced naive human B-cell differentiation and immunoglobulin production  Mark C. Glaum, MD, PhD, Shilpi Narula, MD, Decheng Song,

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Presentation on theme: "Toll-like receptor 7–induced naive human B-cell differentiation and immunoglobulin production  Mark C. Glaum, MD, PhD, Shilpi Narula, MD, Decheng Song,"— Presentation transcript:

1 Toll-like receptor 7–induced naive human B-cell differentiation and immunoglobulin production 
Mark C. Glaum, MD, PhD, Shilpi Narula, MD, Decheng Song, MD, Yi Zheng, MD, Amy L. Anderson, BS, MS, C. Hank Pletcher, BA, MCI, Arnold I. Levinson, MD  Journal of Allergy and Clinical Immunology  Volume 123, Issue 1, Pages e4 (January 2009) DOI: /j.jaci Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 Resiquimod-induced IgM and IgG production by naive B cells. IgM and IgG production by CD19+CD27− naive B cells cultured with medium alone (M), resiquimod (R), IL-2 and IL-10 (C), and resiquimod + IL-2 and IL-10 (R + C) is shown. The results of experiments from 5 individual donors (EXP1-5) are shown on the scatter plot. There were insufficient cells in experiment 1 to establish cultures with resiquimod plus cytokines, accounting for 4 data points. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 Resiquimod-induced IgM and IgG production by memory B cells. IgM and IgG production by CD19+CD27+ memory B cells cultured with medium alone (M) or resiquimod (R) is shown. The results of experiments from 5 individual donors (EXP1-5) are shown on the scatter plot. There were insufficient memory B cells to establish cultures with added cytokines. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig 3 Effect of resiquimod on B-cell proliferation. A total of 5 × 104 highly purified naive B cells (A) or memory B cells (B) were cultured in triplicate in the presence of the additives indicated in the graphs for 72 hours, at which time 1 μC [3H]thymidine was added for 16 hours. Uptake of [3H]thymidine was expressed as counts per minute. Bars represent the means ± SDs of 3 independent experiments. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig 4 Resiquimod-induced expression of AID and Iγ1-Cμ circle transcripts in naive human B cells. Expression of AID, Iγ1-Cμ circle, and β-actin transcripts in naive B cells cultured for 7 days in the presence of indicated additives. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig 5 Cytokine production induced by resiquimod. The supernatants of highly purified naive B cells cultured with medium alone or resiquimod were recovered after 24 hours, and levels of IFN-α, IL-6, and IL-10 were analyzed by Searchlight multiplex ELISA. Bars represent the means ± SDs of 3 independent experiments. IFN-α was not detected in any of the culture supernatants and is therefore not shown. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

7 Bivariate dot plot showing sort purity of the CD19+CD27−SIgG− CD123− B cells (highly purified naive B cells). PBMCs were sorted into CD19+CD27−SIgG− CD123− B cells as described in Methods. The highly purified cells were reanalyzed by flow cytometry and found to contain <0.1% non-B cells, which include T cells, natural killer cells, and pDCs (A) and <0.8% SIgG+ cells (B). Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

8 Bivariate dot plot showing percentage of isotype-switched memory B cells in the PBMCs of a patient with X-linked hyper-IgM syndrome. The percentage of CD27−SIgM− cells (isotype-switched) were enumerated by flow cytometry using mAbs specific for CD19, CD27, and SIgM. The gated CD19 cells contained 0.5% CD27+SIgM− cells (quadrant C4). PE, Phycoerythrin; PC5, PE-Cy5. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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