Circulating Th17, Th22, and Th1 Cells Are Increased in Psoriasis

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Circulating Th17, Th22, and Th1 Cells Are Increased in Psoriasis Shinji Kagami, Heather L. Rizzo, Jennifer J. Lee, Yoshinobu Koguchi, Andrew Blauvelt Journal of Investigative Dermatology Volume 130, Issue 5, Pages 1373-1383 (May 2010) DOI: 10.1038/jid.2009.399 Copyright © 2010 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Circulating CCR6+, IL-23R+, and CD161+ cells are increased in psoriasis. (a) The percentage of CD4+ cells expressing CCR6, IL-23R, and CD161 among unstimulated cells from 17 healthy individuals and 21 untreated psoriatics (horizontal bars, mean; each circle, single donor). (b) Representative dot plot analyses of CCR6, IL-23R, and CD161 expression in circulating unstimulated CD4+ cells from the same individual. (c) The percentages of each subset divided by the total number of circulating CCR6, IL-23R, and CD161-positive cells in unstimulated CD4+ cells of healthy individuals (white bars) and psoriatics (black bars). Data expressed as mean±SD; *P<0.05, **P<0.01, and ***P<0.001. Journal of Investigative Dermatology 2010 130, 1373-1383DOI: (10.1038/jid.2009.399) Copyright © 2010 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Circulating Th17, Th1, and Th17/Th1 cells are increased in psoriasis. (a) In single-color flow analyses, circulating IL-17A+, IL-22+, IFN-g+, and tumor necrosis factor (TNF)-alpha+ cells are increased in psoriasis patients. (b) Representative two-color dot plot analyses of IL-17A and IFN-γ expression in stimulated CD4+ cells from a healthy volunteer and a psoriasis patient. (c) The percentages of circulating Th17 (IL-17A+IFN-γ-), Th1 (IL-17A-IFN-γ+), and Th17/Th1 (IL-17A+IFN-γ+) cells among stimulated CD4+ cells from healthy individuals and psoriatics. For a and c, each circle, single donor; horizontal bars, mean; and *P<0.05, **P<0.01, and ***P<0.001. Journal of Investigative Dermatology 2010 130, 1373-1383DOI: (10.1038/jid.2009.399) Copyright © 2010 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Most Th17 cells do not simultaneously express both IL-17A and IL-22. (a) Representative two-color dot plot analyses of IL-17A, IL-22, and IFN-γ expression in stimulated CD4+ cells from a single patient with psoriasis. (b) The percentages of circulating CD4+ cells that coexpress IL-17A, IL-22, and IFN-γ in healthy individuals (white bars) and psoriatics (black bars). Data expressed as mean±SD; *P<0.05, **P<0.01, and ***P<0.001. Journal of Investigative Dermatology 2010 130, 1373-1383DOI: (10.1038/jid.2009.399) Copyright © 2010 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 On the basis of IL-17A production, CCR6 is a more reliable cell surface marker for Th17 cells when compared with IL-23R or CD161. IL-17A+ cells, IL-22+ cells, and IFN-γ+ cells from psoriatics were gated and the percentage of cells coexpressing CCR6, IL-23R, CD161, or tumor necrosis factor (TNF)-α was determined (black bars). Similarly, IL-17A- cells, IL-22- cells, and IFN-γ- cells from psoriatics were gated and the percentage of cells coexpressing CCR6, IL-23R, CD161, or TNF-α was determined (white bars). Data expressed as mean±SD; NS, not significant; *P<0.05, **P<0.01, and ***P<0.001. Journal of Investigative Dermatology 2010 130, 1373-1383DOI: (10.1038/jid.2009.399) Copyright © 2010 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 5 CCR6+IL-17A+, CCR6+IL-22+, and CCR6+TNF-α+ cells are increased in psoriatics compared with healthy individuals. Representative two-color dot plot analyses of CCR6 and IL-17A/IL-22/tumor necrosis factor (TNF)-α expression in stimulated CD4+ cells from a single healthy volunteer (a) and a psoriasis patient (b). (c) The percentages of circulating IL-17A+ cells that coexpress CCR6, IL-23R, and CD161 among stimulated CD4+ cells from healthy individuals and psoriatics. (d) The percentages of circulating IL-22+ cells that coexpress CCR6, IL-23R, and CD161 among stimulated CD4+ cells from healthy individuals and psoriatics. (e) The percentages of circulating TNF-α+ cells that coexpress CCR6, IL-23R, and CD161 among stimulated CD4+ cells from healthy individuals and psoriatics. Each circle, single donor; horizontal bars, mean; *P<0.05, **P<0.01, and ***P<0.001. Journal of Investigative Dermatology 2010 130, 1373-1383DOI: (10.1038/jid.2009.399) Copyright © 2010 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 6 NF-κB and STAT3 inhibition in vitro blocks cytokine production by both Th17 and Th1 cells in a dose-dependent manner. (a) CD4+ cells were cultured overnight with or without parthenolide or stattic at the indicated concentrations, and then activated with phorbol myristate acetate (PMA)/ionomycin for 6hours. Representative two-color dot plot analyses of IFN-γ and tumor necrosis factor (TNF)-α expression from a single healthy volunteer are shown. (b) The percentage of circulating CD4+ cells producing IL-17A, IL-22, IFN-γ, and TNF-α in healthy individuals (solid lines) and psoriatics (dotted lines) with or without 2.5μM of parthenolide or 1μM of stattic. Each line, single donor. Journal of Investigative Dermatology 2010 130, 1373-1383DOI: (10.1038/jid.2009.399) Copyright © 2010 The Society for Investigative Dermatology, Inc Terms and Conditions