1. Topical Mechlorethamine Restores Autoimmune-Arrested Follicular Activity in Mice with an Alopecia Areata-Like Disease by Targeting Infiltrated Lymphocytes 
Liren Tang, Liping Cao, Olga Bernardo, Harvey Lui, Jerry Shapiro 
Journal of Investigative Dermatology 
Volume 120, Issue 3, Pages (March 2003)
DOI: /j x
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
Mechlorethamine increased hair indices in C3H/HeJ mice. Each data point represents the average hair indices of 24 mice over 10wk of mechlorethamine treatment. There was no baseline difference in hair indices between the two sides before treatment. Significant increase in hair indices was observed on the treated sides after the fourth week of mechlorethamine treatment, whereas the control-vehicle-treated sides remained unchanged over the course of the experiment. Error bars represent SD.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
A female C3H/HeJ mouse was successfully treated unilaterally with mechlorethamine. There was no difference in hair density 1wk after the treatment (A, C, E). After 6wk, the mechlorethamine-treated right side (B, F) showed normal hair density, whereas the vehicle-treated left side (B, D) showed more hair loss than at baseline. A normal mouse showed no difference in hair growth between mechlorethamine-treated right side and control-vehicle-treated left side either before (G) or after 6wk of treatment (H). Hair regrowth was present on both sides after 6wk of treatment (H).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Immunohistochemistry of C3H/HeJ mouse skin with CD4+ (A, B, C, D) and CD8+ (E, F, G, H) antibodies on the control (A, B, E, F) and mechlorethamine-treated (C, D, G, H) sides. Both CD4+ and CD8+ subpopulations of lymphocytes were depleted on the treated sides from perifollicular and intrafollicular areas (compare A, B with C, D; E, F with G, H). Scale bars: 25μM.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Viability (A) and cell proliferation (B) of different cell types treated with mechlorethamine in vitro. Human peripheral blood mononuclear cells (PBMC), T cell line Jurkat cells, epidermal keratinocytes (EKC), dermal fibroblasts (DFB), follicular keratinocytes (FKC) and follicular dermal papilla fibroblasts (FDP), mouse dermal fibroblasts (MDFB), and mouse spleen lymphocytes (MSL) were treated with different concentrations of mechlorethamine as indicated. The cell viability was determined by MTT assay and cell proliferation was assayed by 3H-thymidine incorporation. The average of three independent tests was used and the percentage over the control culture was plotted against the treated groups with different concentrations of mechlorethamine.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
TNF-α/β and IL-12 were inhibited upon successful treatment by mechlorethamine in AA-affected C3H/HeJ mice. Total RNA was extracted from AA-affected C3H/HeJ mice after 10wk of mechlorethamine treatment. The RNA from the control (C) and treated (T) sides of each mouse was assayed for gene expression by RPA using the templates described in Materials and Methods. TNF-α/β and IL-12 were consistently and dramatically inhibited by mechlorethamine in all 12 mice tested. Data of six mice are shown. L32 was used as an internal control and showed no significant difference between two sides among animals.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

7. Figure 6
Expression of IFN-γ, TNF-α, and IL-12 was downregulated by mechlorethamine using real-time RT-PCR. Total RNA extracted from skin biopsies of treated and control sides from seven mice after 10wk of mechlorethamine therapy was used for PCR amplification. The cross-point cycle number of a specific cytokine from each specimen was normalized to that of β-actin from the same specimen. The average difference in the cycle number between mechlorethamine-treated and control sides from seven mice was used to calculate the fold change. The “+” indicates increased by mechlorethamine and “–” represents inhibitory expression by mechlorethamine treatment. Error bars represent SD.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions