This study describes, to our knowledge, a previously unreported mouse model of vitiligo that targets the epidermis but spares the hair. A recent study in humans with vitiligo reported a direct correlation among the severity of vitiligo, involvement of the hair, and autoreactive CD8+
T cell response to stimulation in vitro
(van den Boorn et al., 2009
). Two other models of melanocyte-targeted immunity resulted in prominent loss of the hair (Becker et al., 1996
; Gilhar et al., 2001
), however we do not observe this in our model. Studies in a mouse model of melanoma immunotherapy report hair depigmentation with a treatment protocol that includes PMEL CD8+
T cell adoptive transfer into RAG1−/−
hosts (Antony et al., 2005
), a result that we have reproduced (not shown). These data demonstrate that melanocytes within the hair follicle can serve as targets for PMEL CD8+
T cells under certain immunologic conditions, arguing against altered PMEL antigen presentation by follicular melanocytes as a cause for sparing of the hair in vitiligo. Because the hair follicle is an immune privileged organ (Gilhar et al., 2007
) and is often protected from depigmentation in patients, the involvement of the hair in existing mouse models of vitiligo suggests a more robust T cell response than the model presented here. Thus, these models are complimentary, and may provide an opportunity to further dissect the mechanisms of immune tolerance in the hair follicle.
Flow cytometry of GFP+
T cells in the skin revealed that autoreactive T cells accumulated gradually after transfer and became maximal at 5 weeks. The loss of melanocytes paralleled this accumulation, as levels decreased beginning soon after transfer and continued for 5 weeks. These data reveal an early loss of melanocytes prior to visible depigmentation, suggesting subclinical disease induced by very few infiltrating T cells. Fluorescence microscopy in our model revealed microscopic clusters of T cells in areas of grossly normal pigmentation, a pattern that is reminiscent of patchy depigmentation but on a microscopic scale. A previous report by Wankowicz-Kalinska, et al.
described the microscopic infiltration of T cells in clinically normally pigmented skin of patients with vitiligo, which was accompanied by focal melanocyte loss by immunohistochemistry. They labeled this phenomenon “microdepigmentation”, and hypothesized that it may represent the earliest stage in the development of macroscopic depigmented lesions (Wankowicz-Kalinska et al., 2003
). Our data describing the early accumulation and microscopic clustering of autoreactive T cells in the skin with a quantifiable decrease in melanocytes, all prior to macroscopic depigmentation, support this concept.
Similar to human disease (van den Boorn et al., 2009
), the mouse model of vitiligo described here revealed that IFN-γ RNA transcript was elevated in affected skin and that autoreactive CD8+
T cells produced IFN-γ. Furthermore, treatment with anti-IFN-γ antibody abrogated depigmentation in this model, which was associated with a prominent deficiency in T cell accumulation in the skin. Additional studies including BrdU incorporation and staining for Ki-67 reveal limited evidence for proliferation within the skin (not shown), suggesting that migration is a major component of the T cell accumulation during disease, and that IFN-γ may induce T cell recruitment to the skin during vitiligo. A study conducted by Gregg, et al
. reported that depigmentation of the hair and epidermis in their TCR transgenic mouse model targeting tyrosinase was impaired in IFN-γ−/−
mice (Gregg et al., 2010
), consistent with our results. IFN-γ-dependent mechanisms related to T cell homing to peripheral tissues include the local induction of chemokines and the expression of adhesion molecules on endothelial cells (Bromley et al., 2008
). Their studies identified a role for the chemokine receptors CCR5 and CXCR3 in depigmentation. However their contribution, revealed using individual receptor-deficient mice, was mild compared to IFN-γ-deficient mice, suggesting that additional IFN-γ-dependent mechanisms contributed to disease. Future studies will further define the role of IFN-γ in T cell recruitment to the skin, and determine whether IFN-γ neutralization can reverse existing skin depigmentation, which depends not only on inhibiting the autoimmune response, but on the ability of melanocytes to repigment the epidermis.
The clinical significance of the treatment response to IFN-γ neutralization reported here lies in its potential use as a new therapy for vitiligo. However, this must be balanced with the understanding that IFN-γ is required for clearance of a number of pathogens (Zhang et al., 2008
), and thus future studies to implicate critical events downstream of IFN-γ signaling may provide more targeted, and therefore safer, systemic treatments for vitiligo and related autoimmune diseases. In addition to testing systemic treatments for vitiligo, future studies will take advantage of the prominent epidermal depigmentation observed in this model of vitiligo to test new topical treatments.