|
|
30秒注册登陆，可查看更多信息，结交更多好友，享用更多功能，轻松玩转论坛，白白手拉手欢迎您的加入！
您需要 登录 才可以下载或查看，没有账号？注册新成员
x
谢谢
Log in
ORIGINAL ARTICLE|VOLUME 132, ISSUE 7,P1869-1876,JULY 01, 2012
A Mouse Model of Vitiligo with Focused Epidermal Depigmentation Requires IFN-γ for Autoreactive CD8+T-Cell Accumulation in the Skin
John E. Harris
Tajie H. Harris
Wolfgang Weninger
E. John Wherry
Christopher A. Hunter
Laurence A. Turka
Open ArchiveDOI:https://doi.org/10.1038/jid.2011.463
PlumX Metrics
Vitiligo is an autoimmune disease of the skin causing disfiguring patchy depigmentation of the epidermis and, less commonly, hair. Therapeutic opti** for vitiligo are limited, reflecting in part limited knowledge of disease pathogenesis. Existing mouse models of vitiligo c**ist of hair depigmentation but lack prominent epidermal involvement, which is the hallmark of human disease. They are thus unable to provide a platform to fully investigate disease mechanisms and treatment. CD8+T cells have been implicated in the pathogenesis of vitiligo, and expression of IFN-γ is increased in the lesional skin of patients, however, it is currently unknown what role IFN-γ has in disease. Here, we have developed an adoptive transfer mouse model of vitiligo using melanocyte-specific CD8+T cells, which recapitulates the human condition by inducing epidermal depigmentation while sparing the hair. Like active lesi** in human vitiligo, histology of depigmenting skin reveals a patchy mononuclear infiltrate and single-cell infiltration of the epidermis. Depigmentation is accompanied by accumulation of autoreactive CD8+T cells in the skin, quantifiable loss of tyrosinase transcript, and local IFN-γ production. Neutralization of IFN-γ with antibody prevents CD8+T-cell accumulation and depigmentation, suggesting a therapeutic potential for this approach.
Abbreviati**
GFP
green fluorescent protein
H+E
hematoxylin and eosin
Krt14-Kitl*
KRT14-Kitl*4XTG2Bjl mouse strain
MHC
major histocompatibility complex
rVV-hPMEL
recombinant vaccinia virus-human PMEL (also known as gp100)
Introduction
Vitiligo is a skin disease that causes patchy depigmentation of the epidermis and afflicts 0.5% of the population, without preference for race or gender. It commonly affects the central face and genitals, often localizes to the hands and feet, and less commonly presents on the trunk and proximal extremities (
Taieb and Picardo, 2009
). Hair pigmentation is often spared within lesional skin, and the successful treatment of vitiligo results in repigmentation that usually begins as small dark macules around the hair follicles, presumably because follicular melanocytes are protected by immune privilege (
Falabella, 2009
).
Previous studies have implicated autoreactive CD8+T cells in disease pathogenesis. For example, the frequency of anti-melanocyte CD8+T cells in both the blood and skin of patients with vitiligo correlate with the severity of disease, and lesional CD8+T cells induce melanocyte apoptosis in unaffected skinex vivo(
Ogg et al., 1998
;
van den Boorn et al., 2009
), an observation that supports a direct role for cytotoxic T lymphocytes in melanocyte destruction in human vitiligo. The role of inflammatory cytokines is not yet fully defined, although IFN-γ has been the most extensively studied. In human patients, IFN-γ is expressed in lesional skin and can be produced by autoreactive CD8+T cells (
van den Boorn et al., 2009
), but its role in depigmentation is not known.
Existing models of vitiligo have been established in wild-type C57BL/6 mice, which have black hair but light skin owing to the few melanocytes found in the epidermis. C**equently, the majority of these models focus on depigmentation of the hair rather than the epidermis, and because hair depigmentation predicts a poor resp**e to treatment, they may represent more severe forms of vitiligo (
van den Boorn et al., 2009
). Here, we report a mouse model of vitiligo using adoptive transfer of melanocyte-specific CD8+T cells into mice with increased epidermal melanocytes, which results in black skin and black hair. Similar to the common forms of human vitiligo, depigmentation in these mice affects the epidermis but spares the hair, and the skin histologically resembles active human disease. Mechanistic studies in this model reveal that depigmentation requires IFN-γ, which induces the local accumulation of melanocyte-specific CD8+T cells within the skin. These results support a critical role for IFN-γ in vitiligo and suggest that it acts to recruit CD8+T cells to the skin. The ability of antibody-mediated neutralization of IFN-γ to prevent CD8+T-cell accumulation in the skin as well as depigmentation in this model suggests a therapeutic potential for this approach.
Results
Vitiligo induction through the adoptive transfer of melanocyte-specific CD8+T cells c**ists of epidermal depigmentation sparing the hair
To develop a mouse model of vitiligo with prominent epidermal depigmentation, mice with black skin and hair were used as hosts for the adoptive transfer of melanocyte-specific CD8+T cells. KRT14-Kitl*4XTG2Bjl (Krt14-Kitl*) transgenic mice contain melanocytes in the epidermis owing to the overexpression of Kit ligand (KITL) protein that is noncleavable due to mutati** in the proteolytic domains, and is therefore membrane-bound (KITL*) (
Majumdar et al., 1994
); expression is driven from the human keratin 14 promoter (Krt14), resulting in retention of melanocytes in the epidermal basal layer and follicular epithelium. These mice have black hair and black skin with an elevated number of melanocytes in the epidermis but other cell populati** are normal, representative of human skin (
Kunisada et al., 1998
). PMEL-specific TCR transgenic CD8+T cells (PMEL CD8+T cells) recognize both mouse and human PMEL (also known as gp100), a melanocyte-specific differentiation antigen (
Overwijk et al., 1998
). Using these cells, we modified a protocol optimized for melanoma immunotherapy (
Antony et al., 2005
) to induce vitiligo in our system. Nave CD8+T cells from the spleens of PMEL TCR transgenic mice were purified by negative selection, and one million cells were transferred intravenously into sublethally irradiated (500rads) Krt14-Kitl*hosts. To track PMEL CD8+T cells after transfer, PMEL TCR transgenic mice were crossed with mice that express green fluorescent protein (GFP) driven by the DPE promoter, which c**ists of the distal and proximal Cd4 enhancers and promoter, but lacks a silencer element that prevents CD4 expression in mature CD8+T cells; this results in GFP expression in both CD4+and CD8+T cells (
Mrass et al., 2006
). On the day of transfer, Krt14-Kitl*hosts were infected intraperitoneally (i.p.) with 106pfu of recombinant vaccinia virus that expresses human PMEL (rVV-hPMEL), a potent antigenic stimulus for PMEL CD8+T cells that results in their expansionin vivo(
Overwijk et al., 2003
). In summary, Krt14-Kitl*hosts are sublethally irradiated, GFP+PMEL CD8+T cells are adoptively transferred intravenously, and hosts are infected i.p. with rVV-hPMEL to activate transferred T cellsin vivo.
In all experiments, control mice were irradiated but did not receive rVV-hPMEL or PMEL CD8+T cells. In preliminary experiments, when either irradiation or rVV-hPMEL was eliminated from the protocol, depigmentation occurred only inc**istently, and treatment with rVV-hPMEL without PMEL CD8+T-cell transfer did not result in depigmentation (not shown). In all mice that received the combination of sublethal irradiation, PMEL CD8+T cells, and rVV-hPMEL, epidermal depigmentation appeared 4–5 weeks after transfer. Depigmentation c**istently involved the ears, rear footpads, tail, and, less commonly, the nose; occasionally, the trunk skin under the hair was affected (Figure 1a), as well as the front footpads and genitals (not shown). Depigmentation was initially patchy but often progressed to confluence, involving the entire epidermal surface, while depigmentation of the hair was not observed, even months after transfer (Figure 1b). When wild-type C57BL/6J mice with minimal epider |
|
发表于 2020-6-15 14:59:09
