1. SOCS1 Prevents Potentially Skin-Reactive Cytotoxic T Lymphocytes from Gaining the Ability to Cause Inflammatory Lesions 
Galaxia Maria Rodriguez, Dante D'Urbano, Diwakar Bobbala, Xi Lin Chen, Mehdi Yeganeh, Sheela Ramanathan, Subburaj Ilangumaran 
Journal of Investigative Dermatology 
Volume 133, Issue 8, Pages (August 2013)
DOI: /jid
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2. Figure 1
Socs1−/-Pmel-1 cells display signs of in vivo antigen stimulation. (a) Appearance of 4-week-old Socs1−/-Pmel-1 and Socs1+/+ Pmel-1 control mice. (b–e) Pooled brachial, inguinal, cervical, and mesenteric lymph nodes from 3-week-old mice of the indicated genotype from the same litter were stained for CD4, CD8, TCR Vβ13 (b) and the indicated memory cell markers (c) or molecules that are modulated following antigen stimulation (d and e), and evaluated by flow cytometry. Numbers within quadrants of dot plots indicate the proportion of cells. Numbers within histograms denote mean channel values. Dotted lines were placed in histograms for visual comparison. Data shown are representative of similar results from more than three mice from different litters. SOCS1, suppressor of cytokine signaling 1.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Socs1−/- CD8+ T cells show impaired antigen (Ag)-induced proliferation. (a) Lymph node cells (left panel) from 2- to 3-week-old Socs1−/- and control Pmel-1 mice were stimulated with mgp10025–33 with or without IL-2. Purified CD8+ T cells (right panel) were stimulated with mgp10025–33 presented by irradiated C57BL/6 splenocytes. (b, c) Lymph node cells were stimulated with hgp10025–33 with or without IL-2, anti-CD3ε, and anti-CD28 mAb-coated Dynabeads or cytokines. (d) Lymph node cells from female Socs1−/- or control Rag1−/-H-Y TCR transgenic mice were stimulated with H-Y peptide or cytokines. Cell proliferation was evaluated by [3H]-thymidine incorporation after 2 days (for Ag) or 3 days (cytokines) stimulation. Representative data from at least three independent experiments are shown. c.p.m., counts per minute; SOCS1, suppressor of cytokine signaling 1.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Socs1−/-Pmel-1 cells display increased antigen (Ag)-specific cytolytic activity after Ag or cytokine stimulation. (a) Lymph node cells from control and Socs1−/- Pmel-1 mice were stimulated with 1μgml−1 of mouse (upper panel) or human (lower panel) gp100-derived peptide. After 2 days, stimulated cells were equalized for CD8+ T cell numbers, and incubated with 51Cr-loaded EL4 target cells that were pulsed with hgp10025–33 peptide to measure cytotoxic T lymphocyte (CTL) activity. Freshly isolated cells were used as controls. (b) Lymph node cells from Socs1−/- Pmel-1 mice were stimulated with IL-15 and IL-21, either alone or together for 36hours, and equivalent numbers of CD8 cells were tested for CTL activity against hgp100 peptide–loaded EL4 targets. Representative data from two independent experiments with similar results are shown. SOCS1, suppressor of cytokine signaling 1.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Socs1−/-Pmel-1 cells undergo massive lymphopenia-induced proliferation. (a) Lymph node cells from Socs1−/- and control Pmel-1 cells were labeled with 5-(6)carboxyfluorescein diacetate succinimidyl ester (CFSE) and 10 × 106 cells were adoptively transferred to Rag1−/- mice. After 5 days, pooled lymph node cells were stained for CD8, and proliferation of CD8+ T cells was evaluated by flow cytometry. (b) In parallel, markers of lymphopenia-induced proliferation were evaluated on CD8+ T cells. (c) Rag1−/- mice lacking IL-15 were used as recipients of CFSE-labeled Socs1−/- and control Pmel-1 cells, and proliferation of donor CD8+ T cells was evaluated after 5 days. Representative data from three independent experiments are shown. SOCS1, suppressor of cytokine signaling 1.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
Socs1−/-Pmel-1 cells cause severe skin lesions following lymphopenia-induced proliferation. (a) Lymph node cells containing 5 × 106 CD8+ T cells from Socs1−/- or control Pmel-1 mice were adoptively transferred to Rag1−/- or Rag1−/-Il15−/- recipients and monitored for pathological manifestations. (b) (A) Representative Rag1−/- mice that received Socs1−/- (knockout (KO)) or control (wild type (WT)) Pmel-1 cells at 8 weeks after cell transfer. (B–D) Inflammatory lesions of the muzzle in Rag1−/- mice harboring KO cells (B), showing lymphocytic and histiocytic infiltration in the dermis (C, between braces) with unaffected epidermis (C, arrowhead) and vibrissae (C, arrow), and dense lymphocytic and histiocytic infiltration around hair follicles (D). (E) Skin section of Rag1−/- mice harboring control Pmel-1 cells. SOCS1, suppressor of cytokine signaling 1.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions

7. Figure 6
Socs1−/-Pmel-1 cells show increased Blimp-1 expression after antigen stimulation. (a) Socs1−/- and control Pmel-1 cells recovered 6 weeks after transfer to Rag1−/- recipients were evaluated for the indicated cell surface markers. Freshly isolated control Pmel-1 cells were included for comparison. (b) Freshly isolated Socs1−/- and control Pmel-1 cells were stimulated with antigen or the indicated cytokines. After 48hours, cell lystaes were analyzed by western blot to evaluate Blimp-1 expression. Actin was used to ensure equivalent protein loading. Data shown are representative of two independent experiments with similar results. SOCS1, suppressor of cytokine signaling 1.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2013 The Society for Investigative Dermatology, Inc Terms and Conditions