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We compared the morphology and differentiation characteristics of the human cells from bone marrow, adipose tissue, hair papilla and skin dermis. All cell types showed fibroblastic morphology. Immunofluorescent analysis showed that adipose tissue derived stem cells (ADAS) and hair papilla cells (HPC) expressed CD105, CD49d and STRO-1, bone marrow mesenchymal stem cells (BMSC) were absent for CD49, dermal fibroblasts (DFb) expressed CD49 and STRO-1 at low level. Populations of ADAS, BMSC and HPC had similar capacities to lipid and bone differentiation. Following exposure to appropriate induction stimuli, these cells changed phenotype and expressed specific cell markers. However, the rate and extent of HPC differentiation were lower in comparison with populations of ADAS and BMSC. We propose that all investigated cell populations contain primitive progenitor cells with mesenchymal stem cell properties.

Background: Insect bites produce diverse skin reactions. Although quite common, the histopathologic features of arthropod assaults have not ever been studied systemically., Materials and Methods: Twenty biopsies from cases, clinically diagnosed as arthropod bite reactions between January 2003 and June 2007 were reviewed retrospectively. The aim of the study was to verify as to whether reliable histopathologic criteria could be established based on the frequency of findings observed., Results: Epidermal spongiosis (present in 16 of 20 cases), in particular spongiosis of the infundibular epithelium and acrosyringia as well as eosinophilic spongiosis, emerge as relevant diagnostic clues. A moderately dense, superficial and deep infiltrate consisting mainly of lymphocytes and eosinophils was prevalent in the dermis, with eosinophils tending to interstitial and periadnexal distribution. Of note, 19 of 20 (95%) cases revealed periadnexal involvement, whereas 16 of 20 (80%) had the infiltrate extending particularly along the sweat ducts and the coiled glands. In three biopsies, concomitant involvement of sweat glands, hair follicles and sebaceous glands was noted., Conclusion: A practical histopathologic algorithm of arthropod bite recognition is proposed. The involvement of the sweat glands in the pattern of arthropod bite reaction is suggested as a new reliable diagnostic clue.

Follicular mycosis fungoides (FMF) is a recognized variant of mycosis fungoides. In this review, the authors characterize the distinct histopathological and immunohistochemical patterns of FMF that have been reported in the literature. This article is an extensive review of the literature cited in Medline and own data of the authors. The major patterns of FMF histopathology, immunohistochemistry, and molecular genetics are summarized in this review. Histologically, the quintessential finding in FMF is small to medium atypical CD3+ CD4+ CD8- T lymphocytes around and within the epithelium of the hair follicles. This finding is requisite to the diagnosis. However, this finding may be obscured by a host of other patterns often identified in FMF. This includes basaloid folliculo-lymphoid hyperplasia, a granulomatous reaction, eosinophilic folliculitis, and follicular cystic changes with subtle atypical lymphocytes in the cyst wall. Follicular mucinosis (MF) and syringo-tropism are also variably present. Immunohistochemistry of all reported cases uniformly show a CD4+ T cell infiltrate. This review emphasizes and discusses the broad spectrum of histologic changes which may be seen in FMF, clues to the diagnosis, and some potential mimickers.

Background: Recent gene profiling data suggest that, besides the anagen hair bulb, the epithelial stem cell region in the outer root sheath of hair follicles (HFs), termed the bulge, may also represent an area of relative immune privilege (IP)., Objectives: To investigate whether the human HF bulge is a site of relative IP within anagen VI HFs., Methods: Anagen VI HFs from normal human scalp skin were analysed using immunohistological staining techniques, quantitative histomorphometry and statistical analysis. For functional evidence we performed full-thickness human scalp skin organ cultures to investigate whether interferon (IFN)-gamma, a key inducer of IP collapse in hair bulbs, has a similar effect on the putative bulge IP., Results: Major histocompatibility complex (MHC) class Ia, beta(2)-microglobulin and MHC class II immunoreactivity are downregulated in the human bulge. The immunosuppressants alpha-melanocyte stimulating hormone, transforming growth factor-beta2, macrophage migration inhibitory factor and indoleamine-2,3-dioxygenase (IDO) are upregulated in the CD200+, stem cell-rich bulge region. These CD200+ cells also co-express HLA-E. Furthermore, IFN-gamma induces significant ectopic MHC class Ia expression in bulge cells of organ-cultured human scalp skin., Conclusions: These data suggest that the bulge of human anagen HFs represents a hitherto unrecognized site of relative IP in human skin. Simultaneously, we present the first evidence of IDO and HLA-E protein expression in normal human HFs. Bulge IP presumably protects the HF epithelial stem cell reservoir from autoaggressive immune attack whereas a loss of bulge IP may play a central role in the pathogenesis of cicatricial alopecias.

Desmosomal cadherins are essential cell adhesion molecules expressed in the epidermis. We identified a mutation of a cadherin superfamily member, namely, desmoglein 4 (Dsg4), in early onset of death (EOD)( hage ) mice with hypotrichosis. The mutation was induced by the insertion of an early transposon II-beta into intron 8 of Dsg4. Mast cell hyperplasia was observed in the skin of EOD( hage ) mice. The abnormally expanded population of lpr T cells, i.e., CD4(-)CD8(-)B220(+)Thy1.2(+) alphabetaT cells, in the splenocytes of EOD mice was reduced in EOD( hage ) mice. Therefore, it was suspected that the long-living mutant EOD( hage ) mice were selected from lupus-prone EOD mice because of their immunological immaturity. These findings clearly indicate that Dsg4 is an important molecule for the formation of hair follicles and hypothesize that unorganized hyperplastic hair follicles in anagen due to the Dsg4 mutation provide niches for mast cell precursors in the skin.

While the pathogenesis of most scarring alopecias is poorly understood, one recent study indicates destruction of follicular stem cells as a possible mechanism in lichen planopilaris, the prototypic scarring alopecia. The aim of this cross-sectional study was to ascertain the target of inflammation and to more precisely characterize the inflammatory infiltrate in various stages of primary scarring alopecias. Immunohistochemical studies were performed using a panel of antibodies that included anti-cytokeratin 15, an antibody that specifically targets follicular bulge stem cells and CD4, CD8, CD1a and human leukocyte antigen-DR to characterize the inflammatory infiltrate. Our data showing absence of follicular bulge stem cells in cases with moderate to heavy inflammation suggest involvement of the bulge region in 'early' active stages of primary scarring alopecia. The paucity of CD8+ T cells in the inflammatory infiltrate in the majority of these cases argues against a cell-mediated cytotoxic destruction of follicular bulge stem cells. Preservation of CK15+ cells in 'late' fibrotic stages of primary scarring alopecia further supports this and implies that the irreversible loss of hair follicles, the sine qua non of primary scarring alopecia, is not necessarily a consequence of T cell-mediated destruction of follicular bulge stem cells.

Background: CD10 and CD34 have been detected in both epithelial and mesenchymal components of anagen human hair follicles., Objectives: To analyse the expression of CD10 and CD34 in human hair follicle development as well as in different phases of the hair cycle., Methods: Fetal and adult hair follicles at different stages of the hair cycle were examined by immunohistochemistry for CD10 and CD34., Results: In fetal follicles, CD10 is expressed by the cells of the placodes, and CD34 by the mesenchymal cells of the dermal condensate. As the follicle matures, CD10 can be seen in the matrix cells, inner root sheath and dermal sheath. In adult follicles, the expression of CD10 in the follicular epithelium is present in anagen follicles, but tends to disappear in catagen, and is not detected in telogen. The CD10 positivity of the dermal sheath is more intense in catagen than in anagen follicles. CD34 immunostaining of the external root sheath was seen in adult anagen follicles but not in fetal follicles. This staining of the anagen outer sheath tends to disappear in catagen and is not detected in telogen., Conclusions: CD10 and CD34 are not proteins constantly present in a specific cell type of the hair follicle, but are proteins that can be expressed by both epithelial and mesenchymal cells depending on the stage of development and hair cycle. The distribution of the immunoreactivity to CD10 in the placode and CD34 in the dermal condensate suggests a role of these proteins in initial stages of hair formation.

The hair follicle is a repository of different types of somatic stem cells. However, even though the hair follicle is both a prominent target organ and a potent, non-classical site of production and/or metabolism of numerous polypetide- and steroid hormones, neuropeptides, neurotransmitters and neurotrophins, the (neuro-)endocrine controls of hair follicle epithelial stem cell (HFeSC) biology remain to be systematically explored. Focussing on HFeSCs, we attempt here to offer a "roadmap through terra incognita" by listing key open questions, by exploring endocrinologically relevant HFeSC gene profiling and mouse genomics data, and by sketching several clinically relevant pathways via which systemic and/or locally generated (neuro-)endocrine signals might impact on HFeSC. Exemplarily, we discuss, e.g. the potential roles of glucocorticoid and vitamin D receptors, the hairless gene product, thymic hormones, bone morphogenic proteins (BMPs) and their antagonists, and Skg-3 in HFeSC biology. Furthermore, we elaborate on the potential role of nerve growth factor (NGF) and substance P-dependent neurogenic inflammation in HFeSC damage, and explore how neuroendocrine signals may influence the balance between maintenance and destruction of hair follicle immune privilege, which protects these stem cells and their progeny. These considerations call for a concerted research effort to dissect the (neuro-)endocrinology of HFeSCs much more systematically than before.

Hair follicles (HFs) enjoy a relative immune privilege (IP) that is characterized by downregulation of major histocompatibility complex (MHC) class I and local expression of potent immunosuppressants. Normally, natural killer (NK) cells attack cells with absent/low MHC class I expression. However, because few perifollicular NK cells are found around healthy human anagen HFs, we asked how HFs escape from NK cell attack. This study suggests that this happens via an active NK cell suppression. Alopecia areata (AA), an organ-specific autoimmune disease thought to result from a collapse of HF-IP, in contrast, shows striking defects in NK cell inhibition/containment. We show that the NK cell inhibitor macrophage migration inhibitory factor is strongly expressed by the HF epithelium, and very few CD56(+)/NKG2D(+) NK cells are observed in and around normal anagen HFs compared to AA with prominent aggregations of CD56(+)/NKG2D(+) NK around AA-HFs. By flow cytometry, many fewer NK function-activating receptors (NKG2D, NKG2C) and significantly more killer cell Ig-like receptors-2D2/2D3 were found to be expressed on peripheral blood CD56(+) NK cells of healthy controls than on those of AA patients. In addition, only weak immunoreactivity for MHC class I chain-related A gene was observed in normal anagen HFs compared to AA. To our knowledge, this defect is previously unreported and must be taken into account in AA pathogenesis and its management.

Objective: To improve the method of obtaining purified and viable human hair follicle stem cells (HFSCs) from bulge cells (BCs)., Methods: Firstly, the BCs were isolated from human hair follicles by microdissection. Secondly, the CD200+ HFSCs were selected from BCs using magnetic cell sorting method. The viability of these purified HFSCs was detected under light microscope. The purification rate was analyzed by flow cytometry. The pre- and post-purification cells were compared by immunofluorescence staining., Results: The adherent BCs displayed a typical cobblestone morphology on day 6. The BCs expressed K19 strongly. The viability rate of pre-purification cells was 95.0% +/- 0.6% while that of post- purification cells was 94.2% +/- 1.0%. There was no significant difference (P < 0.05). By flow cytometry and immunofluorescence staining examination, the CD200+ cell rate was 8.31% before cell sorting purification while that was 82.31% after cell sorting purification., Conclusion: Highly purified and viable HFSCs could be obtained by micromani pulation and magnetic cell sorting assay.

This chapter summarizes the evidence that defined compartments of the hair follicle (HF) and nail epithelium maintain an area of relative immune privilege (IP). HF and nail IP is chiefly characterized by absent or very low level of expression of major histocompatibility complex class Ia antigens, complemented by a number of factors, such as the local production of potent immunosuppressive agents, dysfunction of professional antigen-presenting cells and inhibition of natural killer cell activities. In the hair bulb, IP is seen only in the anagen stage of HF cycling, while the nail apparatus continuously maintains an IP site in its proximal nail matrix, since the nail apparatus does not cycle. Possibly, the (non-cycling) bulge area of human scalp HFs also enjoys some relative, stably maintained IP, even though it is not as pronounced as the IP of the anagen hair bulb. A collapse of HF and nail IP likely plays a key role in the pathogenesis of one of the most common organ-specific autoimmune diseases, alopecia areata. Therefore, the therapeutic restoration of IP collapse promises to be a particularly effective future strategy for the treatment of alopecia areata.

Erythema Toxicum, a rash frequently present in the healthy newborn infant is an innate, immune response to the first commensal micro flora. Flushing and urtication are seen in this manifestation suggesting mast cell (MC) activation and MC derived mediator release. It has recently become evident that MCs participate in the protective, innate immune response against microbes also by secreting products toxic to pathogens such as cathelicidin peptide antibiotics. We hypothesized that MCs contribute to the process of inflammation in Erythema Toxicum and that skin MCs of human newborns express the cathelicidin peptide antibiotic LL-37. Skin sections were immunostained for MC tryptase. Double immunofluorescence was performed by staining LL-37 in combination with tryptase. We studied ultra structure of skin MCs with transmission (TEM) and immunoelectron microscopy (IEM). Seven infants with and six infants without the rash, as well as three adults were included. We found numerous tryptase-expressing MCs recruited around the hair follicles in the lesions of Erythema Toxicum. TEM analysis of MCs exhibited signs of degranulation in the lesion. Neither skin MCs from newborns nor adults did express LL-37 as judged by confocal and IEM. MCs participate in the inflammatory responses of Erythema Toxicum by taking an active part in the immune system of the hair follicle. However, their immunological activity is not linked to the expression of the cathelicidin antimicrobial peptide LL-37. A pivotal role of MCs in the innate, inflammatory response at the site of pathogen invasion during the critical time of perinatal colonization is suggested.

An attractive approach to immunization is to apply DNA vaccine topically onto the skin. However, it is important to ensure that a strong immune response is induced without disrupting the skin stratum corneum. The hair follicles have been shown to be the major portal of entry for DNA applied onto the skin, and it has been reported that the transfection of hair follicle cells occurs mainly at the onset of a new growing stage of the hair cycle. Using an anthrax protective antigen (PA) protein-encoding plasmid in mice, we demonstrated that the anti-PA immune responses were significantly stronger when the hair follicles in the application area were induced into anagen-onset stage than when in telogen stage. The anti-PA antibodies enabled the immunized mice to survive a lethal dose of anthrax lethal toxin challenge. The enhanced immune responses can be partially attributed to the enhanced antigen gene expression and plasmid DNA uptake in the skin area wherein the hair follicles were induced into anagen-onset stage. Moreover, the moderate dermal inflammation associated with the anagen induction may also have contributed to the enhancement of the resultant immune response. This represents a novel approach to enhancing the immune response induced by a topically applied DNA vaccine.

For many years, the role of T lymphocytes was hypothesized as being a major factor responsible for hair loss in alopecia areata (a.a.). While numerous lymphocytic populations were found around hair bulbs, changes in their distribution patterns in the skin during the course and recovery of the disease remain unknown. In the present study, distribution of CD8 lymphocytes in biopsies obtained from a.a. patients was measured before and during treatment using diphencyprone (DCP). Results show about a 600% increase in the number of CD8 lymphocytes under the epithelium and about a 250% increase around hair bulbs and other epidermal appendages during the study. These results were more significant in a group, which had good clinical response to the treatment. No change in the quantity of CD8 lymphocytes was observed around the blood vessels. Since CD8 lymphocytes are considered to be directly involved in the hair destruction process in a.a., their increased number around hair bulbs followed by hair regrowth may suggest that during DCP treatment they regain normal reactivity to hair antigens.

Many lessons in autoimmunity - particularly relating to the role of immune privilege and the interplay between genetics and neuroimmunology - can be learned from the study of alopecia areata, the most common cause of inflammation-induced hair loss. Alopecia areata is now understood to represent an organ-restricted, T cell-mediated autoimmune disease of hair follicles. Disease induction is associated with collapse of hair follicle immune privilege in both humans and in animal models. Here, the role of HLA associations, other immunogenetic factors, and neuroendocrine parameters in alopecia areata pathogenesis are reviewed. This instructive and clinically significant model disease deserves more widespread interest in the immunology community.

Alopecia areata (AA) is an autoimmune disorder of the hair follicle characterized by inflammatory cell infiltrates around actively growing (anagen) hair follicles. Substance P (SP) plays a critical role in the cutaneous neuroimmune network and influences immune cell functions through the neurokinin-1 receptor (NK-1R). To better understand the role of SP as an immunomodulatory neuropeptide in AA, we studied its expression and effects on immune cells in a C3H/HeJ mouse model for AA. During early stages of AA development, the number of SP-immunoreactive nerve fibers in skin is increased, compared to non-affected mice. However, during advanced stages of AA, the number of SP-immunoreactive nerves and SP protein levels in skin are decreased, whereas the expression of the SP-degrading enzyme neutral endopeptidase (NEP) is increased, compared to control skin. In AA, NK-1R is expressed on CD8+ lymphocytes and macrophages accumulating around affected hair follicles. Additional SP supply to the skin of AA-affected mice leads to a significant increase of mast cell degranulation and to accelerated hair follicle regression (catagen), accompanied by an increase of CD8+ cells-expressing granzyme B. These data suggest that SP, NEP, and NK-1R serve as important regulators in the molecular signaling network modulating inflammatory response in autoimmune hair loss.

Apoptosis signal-regulating kinase 1 (ASK1) is a member of the mitogen-activated protein 3-kinase family that activates both c-Jun NH(2)-terminal kinase and p38 pathways in response to inflammatory cytokines and physicochemical stress. We report that ASK1 deficiency in mice results in dramatic retardation of wounding-induced hair regrowth in skin. Oligonucleotide microarray analysis revealed that expression of several chemotactic and activating factors for macrophages, as well as several macrophage-specific marker genes, was reduced in the skin wound area of ASK1-deficient mice. Intracutaneous transplantation of cytokine-activated bone marrow-derived macrophages strongly induced hair growth in both wild-type and ASK1-deficient mice. These findings indicate that ASK1 is required for wounding-induced infiltration and activation of macrophages, which play central roles in inflammation-dependent hair regrowth in skin.

Expression of a tumor suppressor gene, programmed cell death 4 (PDCD4), was investigated at the protein level in the human skin. Immunohistochemically, PDCD4 protein expressed mainly in suprabasal layers, while PDCD4-positive and -negative areas were observed discontinuously in the basal cell layer of the epidermis. In hair follicles, the suprabulbar area including the hair and inner root sheath was immunoreactive, while the bulbar area, containing germinative cells which were strongly proliferating cell nuclear antigen (PCNA)-positive, was not or less. PDCD4 therefore appears to be important in the differentiation of hair follicles. PDCD4-positive cells were localized in the inside layers while PCNA-positive cells were located in the basal layer in the outer root sheath of hair follicles. The cells of sebaceous glands and sweat glands also were PDCD4-positive. The PDCD4 protein was localized mostly in nuclei of cutaneous cells. PDCD4 expression was found to be suppressed in the epidermis overlying an adult T-cell lymphoma (ATL), possibly reflecting a paracrine effect of factors produced by ATL cells. PDCD4 expression was suppressed in the keratinocyte cell line HaCaT by exposure of cultures to epidermal growth factor, transforming growth factor-beta1 or hepatocyte growth factor. Immunohistochemically, various skin cancers tended to show less PDCD4 expression than normal skin. Promotion of expression might prove useful in preventing or treating certain skin cancers.

Previous studies of experimental Bacillus anthracis cutaneous infections in mice have implicated hair follicles as a likely entry site. Hairless HRS/J mice were used to investigate this possibility because of their non-functional hair follicles that lack penetrating hair shafts. These mice also have diminished macrophage function, increased susceptibility to Listeria, and enhanced neutrophil responses. HRS/J and Balb/c mice were found to be resistant to epicutaneous inoculation with Bacillus anthracis (Sterne) spores onto abraded skin when compared with DBA/2 mice or leucopenic C57BL/6 mice. The HRS/J mice also resisted spore injections that bypassed hair follicles. Haired HRS/J heterozygote mice demonstrated similar reduced susceptibility to B. anthracis spores. Hairless HRS/J mice that were made leucopenic did become susceptible to the epicutaneous spore inoculations. Histologically, the hairless and haired HRS/J mice showed markedly reduced numbers of organisms in hair follicles and the interfollicular dermis when compared even with the resistant Balb/c mice; inflammatory cell infiltrates in the superficial dermis were increased in the HRS/J mice compared with more sensitive strains. Therefore, resistance in the HRS/J mice was apparent at the initial site of epicutaneous inoculation and seemed related to an accumulation of dermal neutrophils rather than to a lack of functional hair follicles.

Background: Outer root sheath melanocytes (ORSM) are not yet routinely cultured and their biology is not known in detail because of their relatively low numbers in the hair follicle and their limited proliferative capacity in in vitro culture in routine media., Objectives: To develop a method for culturing ORSM more easily and to investigate the length of telomeres and antigenic characteristics of ORSM compared with epidermal melanocytes (EM)., Methods: Hair follicles were obtained from three Korean individuals during hair transplantation surgery. Single-cell suspensions of the outer root sheath were made and cultured in melanocyte growth medium with stem cell factor. After 21 days, second-passage outer root sheath keratinocytes (ORSK) (2 x 10(4) mL(-1) MGM) were added into the culture plates. We studied the proliferation pattern, morphological and antigenic characteristics of ORSM for each passage of cultured cells, and observed ORSM telomere length., Results: We established an ORSM culture method using ORSK. Two morphologically different ORSM types were obtained in the primary cultures. At the end of primary culture, ORSM appeared as whitish-cream pellets. The proliferation pattern of ORSM showed a sigmoidal shape, the accumulated numbers of population doublings showed a plateau after approximately 5 months, and senescence occurred at approximately 33 +/- 5 accumulated population doublings. The length of ORSM telomeres continued to shorten as the cells proliferated. In contrast, EM showed a marked proliferation from the early proliferation period which formed a plateau pattern towards the later period, and the number of accumulated population doublings was estimated to be 18 +/- 5 after 2 months. ORSM in the primary culture reacted variably with l-dihydroxyphenylalanine (DOPA): some cells were DOPA negative, some DOPA positive. There were some different antigenic expressions of microphthalmia-associated transcription factor (MITF) showing cytoplasmic expression in ORSM and nuclear expression in EM. By nuclear extraction and Western blotting, we showed that MITF expression of ORSM was marked in the cytoplasm and minimal in the nucleus. Antigenic expression of MITF and Bcl-2 gradually decreased with increasing passage number, whereas tyrosinase-related protein-1 expression did not change., Conclusions: Culture of ORSM requires ORSK or ORSK-related factors; ORSM have greater proliferation potential and show different MITF antigenic expression compared with EM; and the length of ORSM telomeres shortens with repeated proliferation.

Alopecia areata (AA) is a T cell-mediated autoimmune disease that can be treated with the contact sensitizer diphenylcyclopropenone (DCP). Peripheral blood leukocytes from AA patients are relatively resistant to apoptosis which might be due to decreased Fas Ligand (FasL) expression, or to an increase in CD44v7 expression. Moreover it has been suggested in a murine model of AA that contact allergen treatment might interfere with the emigration of Langerhans cells into the draining lymph node, thus hampering autoreactive T-cell activation. To assess whether and which of these mechanisms is of clinical relevance, immunohistochemistry was performed in scalp biopsies of successfully DCP-treated AA patients in the early phase of hair regrowth. In line with recent studies in a murine model of AA, there was no evidence that DCP treatment would interfere with extravasation and skin homing of activated leukocytes. Perifollicular infiltrates of DCP-treated as compared to untreated AA patients actually showed an increased number of perifollicular CD8(+) and CD1a(+) cells. Furthermore, the expression of CD44 and CD49d, which are of major importance in leukocyte extravasation, was even increased in DCP-treated as compared to AA patient infiltrates. The same accounted for the skin homing receptor CD44v10. When we evaluated the leukocyte subpopulations in DCP-treated as compared to untreated AA patients' skin biopsies, there was an undue increase in CD1a(+) cells, that could well be indicative of hampering of the emigration of antigen presenting cells (APC) by allergen treatment. Most importantly, the number of perifollicular TUNEL- and FasL-positive cells was strikingly increased, whereas the number of CD44v7(+) cells remained unaltered. Taken together, this study provides strong evidence that long term treatment with a contact sensitizer allows for the recovery of hair follicle by driving autoreactive T cells into activation-induced cell death. In addition the replacement with newly activated autoreactive T-cells might be impaired due to a DCP-mediated hindrance of APC emigration.

Background: Alopecia areata (AA) is a T-cell mediated putative autoimmune disease of hair follicles, which can be transferred by CD4(+) T cells. However, whether T-helper (Th) 1 or Th2 cytokines are predominant has not yet been defined., Objectives: To elucidate the importance of Th1 cells in the pathogenesis of AA we investigated the functional role of interferon (IFN)-gamma in the experimental induction of AA., Methods: AA was experimentally induced by grafting full-thickness skin from AA-affected C3H/HeJ mice on to C3H/HeJ mice with a targeted deletion of the Th1 cytokine IFN-gamma gene (IFNgamma(-/-)) and on to wild-type mice (IFNgamma(+/+))., Results: While 90% of wild-type mice developed AA, none of the IFNgamma(-/-) mice exhibited hair loss. Immunohistochemistry of skin sections revealed a dense perifollicular and intrafollicular infiltrate of CD4(+) and CD8(+) T cells in controls, while in IFNgamma(-/-) mice skin-infiltrating CD8(+) T cells were absent and the number of CD4(+) cells was significantly reduced. Aberrant expression of major histocompatibility complex class I and II molecules in the putative immune-privileged infrainfundibular site of the hair follicle was found to be weaker in AA-resistant IFNgamma(-/-) mice than in control mice with AA. Flow cytometry revealed that leucocytes of IFNgamma(-/-) mice did not respond to the transfer of AA-affected skin. As distinct from IFNgamma(+/+) mice, neither T-cell activation markers nor Th1 cytokines were upregulated in draining lymph node cells or skin-infiltrating leucocytes of AA-resistant IFNgamma(-/-) mice. However, there was no evidence for a shift towards a Th2 cytokine profile, nor for upregulation of regulatory T cells in IFNgamma(-/-) mice., Conclusions: IFNgamma(-/-) mice fail to activate Th1 cells in response to the transplanted (auto)antigens, which suggests an essential requirement for IFN-gamma-mediated Th1 activation in the induction of AA.

Alopecia areata (AA) is generally regarded as an organ-specific autoimmune disease. Although it has been hypothesized that the autoimmunity is mediated by T cells and that hair follicle melanocyte is one of the targets, definitive evidence is lacking. We here demonstrate that AA-like lesions can be induced in mice by inducing CD8(+) T-cell-mediated immunity to hair follicle melanocytes. We found that hair loss was induced in mice-bearing interleukin-12-producing B16 melanoma cells by the depletion of CD4(+) T cells, accompanied by vitiligo-like coat color change. The alopecic lesions varied in size from pachy to extensive. In many instances, hair loss developed and was followed by the regrowth of white hairs. Histological analysis revealed that mononuclear cells infiltrated in and around the bulb region of hair follicles. Furthermore, immunohistochemical examination clearly showed the intra-follicular infiltration of CD8(+) T cells. Neither the vitiligo-like coat color nor AA-like lesions were induced when CD8(+) T cells were codepleted. These observations indicate that the induction of CD8(+) T-cell-mediated immunity against hair follicle melanocytes causes alopecia. It is thought that there are many types of AA with different mechanisms, targets etc. Although hair follicle melanocytes have long been thought to be one of the targets of AA, evidence to support the hypothesis is sparse. Therefore, we believe that our observation is significant to support the hypothesis.

Background: CD1d belongs to a family of antigen-presenting molecules structurally related to the classical major histocompatibility complex class I proteins., Objectives: To examine the expression pattern of CD1d protein in normal human skin with ageing., Methods: Twenty normal human skin biopsy specimens were obtained from 20 healthy individuals. The latter were divided into three age groups: children (5-20 years), adults (21-50 years) and the elderly (51-81 years). The intensity of CD1d protein production was examined in human skin using immunofluorescent and immunoalkalinephosphatase staining methods., Results: In the epidermis, CD1d protein production was strong in the skin of the children and declined gradually with age, being moderate in adults and weak in the elderly. As compared with values in children, there was a statistically significant decrease (P<0.05) in CD1d protein production in the elderly. In the dermis, CD1d protein production was strong in the fibroblasts, sweat glands, sebaceous glands, blood vessels and hair follicles regardless of age., Conclusions: Our study reports a decreased CD1d protein production in normal human skin with ageing. The clinical ramifications of these observations mandate further investigations.

Long-lasting allergen treatment is the most efficient therapy in alopecia areata (AA). The underlying mechanism is unknown. We here asked whether treatment with a contact sensitizer influences leukocyte migration such that dendritic cell (DC) migration or the recruitment of activated T-cells towards the skin become hampered. Allergen treatment of AA mice was not accompanied by a decrease in skin-infiltrating leukocytes or draining lymph node cells (LNC). However, the distribution of leukocyte subsets was changed with a dominance of monocytes in the skin and a reduced percentage of DCs in draining nodes. Chemokine and chemokine receptor expression in skin and draining nodes was strikingly increased and LNC from untreated and allergen-treated AA mice showed high migratory activity in vitro and readily homed in draining nodes and skin after intravenous injection. However, FITC labelling of the skin and subcutaneous transfer of dye-labelled DC revealed that allergen treatment created a chemokine milieu severely hampering DC migration from the skin towards the draining node. An allergic eczema-induced reduction in DC migration and antigen transfer could well contribute to insufficient T-cell activation and the recovery of hair follicle in AA and possibly be of relevance for other skin-related autoimmune diseases.

Although conventional vaccines have generated major successes in the control of infectious diseases, several obstacles remain in their development against chronic diseases (HIV, tuberculosis), against which no current candidate vaccines yet ensure protection. The transcutaneous route of vaccine administration appears to be a promising approach of targeting vaccines toward antigen-presenting cells (APCs) and thus improving immune responses. We investigated the suitability of nanoparticles in this approach. We found a high density of Langerhans cells (LCs) around hair follicles that, when sorted, readily internalized all size particles. However, flow cytometry after transcutaneous application of 40, 750, or 1,500 nm nanoparticles on human skin samples revealed that only 40 nm particles entered epidermal LC. Fluorescence and laser scan microscopies, which were carried out to identify the penetration pathway of transcutaneously applied nanoparticles, revealed that only 40 nm particles deeply penetrate into vellus hair openings and through the follicular epithelium. We conclude that 40 nm nanoparticles, but not 750 or 1,500 nm nanoparticles, may be efficiently used to transcutaneously deliver vaccine compounds via the hair follicle into cutaneous APCs.

The goal of this review is to introduce the immunologic community to alopecia areata as a model system for the study of tissue directed autoimmune disease. Alopecia areata is marked by autoimmune assault on the hair follicle resulting in hair loss. It is linked to HLA-DQ3 and evidence suggests it is mediated by T-lymphocytes with a TH1 cytokine profile. Hair follicles are an immune protected site with deficient MHC expression. Evidence is presented suggesting that alopecia areata results from loss of immune privilege with presentation of autoantigens. Alopecia areata is one of the most common human autoimmune conditions, with a lifetime risk of approximately 1.7%. Study of alopecia areata in humans is facilitated by the accessibility of scalp for biopsy. It is possible to transfer the condition with lesional human lymphocytes in a human scalp graft/SCID mouse model. There are also spontaneous animal models which share the features of the human condition. For these reasons, alopecia areata is a powerful model for study of the induction and pathogenesis of tissue directed autoimmune disease.

The skin offers an ideally suited, clinically relevant model for studying the crossroads between peripheral and systemic responses to stress. A 'brain-skin connection' with local neuroimmunoendocrine circuitry underlies the pathogenesis of allergic and inflammatory skin diseases, triggered or aggravated by stress. In stressed mice, corticotropin-releasing hormone, nerve growth factor, neurotensin, substance P and mast cells are recruited hierarchically to induce neurogenic skin inflammation, which inhibits hair growth. The hair follicle is both a target and a source for immunomodulatory stress mediators, and has an equivalent of the hypothalamus-pituitary-adrenal axis. Thus, the skin and its appendages enable the study of complex neuroimmunoendocrine responses that peripheral tissues launch upon stress exposure, as a basis for identifying new targets for therapeutic stress intervention.

The human hair follicle bulge is an important niche for keratinocyte stem cells (KSCs). Elucidation of human bulge cell biology could be facilitated by analysis of global gene expression profiles and identification of unique cell-surface markers. The lack of distinctive bulge morphology in human hair follicles has hampered studies of bulge cells and KSCs. In this study, we determined the distribution of label-retaining cells to define the human anagen bulge. Using navigated laser capture microdissection, bulge cells and outer root sheath cells from other follicle regions were obtained and analyzed with cDNA microarrays. Gene transcripts encoding inhibitors of WNT and activin/bone morphogenic protein signaling were overrepresented in the bulge, while genes responsible for cell proliferation were underrepresented, consistent with the existence of quiescent noncycling KSCs in anagen follicles. Positive markers for bulge cells included CD200, PHLDA1, follistatin, and frizzled homolog 1, while CD24, CD34, CD71, and CD146 were preferentially expressed by non-bulge keratinocytes. Importantly, CD200+ cells (CD200hiCD24loCD34loCD71loCD146lo) obtained from hair follicle suspensions demonstrated high colony-forming efficiency in clonogenic assays, indicating successful enrichment of living human bulge stem cells. The stem cell behavior of enriched bulge cells and their utility for gene therapy and hair regeneration will need to be assessed in in vivo assays.

Background: CD1d belongs to a family of antigen presenting molecules that are structurally and distantly related to the classic major histocompatibility complex class I (MHC I) proteins. However, unlike MHC I molecules, which bind protein antigens, CD1d binds to lipid and glycolipid antigens. CD1d is expressed by cells of lymphoid and myeloid origin, and by cells outside of the lymphoid and myeloid lineages, such as human keratinocytes of psoriatic skin., Aims: To investigate whether CD1d is also expressed in sun exposed skin and in the immuno-privileged anagen hair follicle., Materials/methods: CD1d immunoreactivity was studied in human scalp skin and hair follicles of healthy women in situ by immunofluorescent and light microscopic immunohistology. Skin biopsies were obtained from normal human scalp containing mainly anagen VI hair follicles from women (age, 53-57 years) undergoing elective plastic surgery., Results: CD1d showed strong immunostaining in human scalp skin epidermis, pilosebaceous units, and eccrine glands. In the epidermis, CD1d was strongly expressed by basal and granular keratinocytes. In hair follicles, CD1d was expressed in the epithelial compartment and showed hair cycle related alterations, with an increase in the anagen and a reduction in the catagen and telogen phases., Conclusions: These results suggest that CD1d plays a role in human scalp skin immunology and protection against lipid antigen rich infectious microbes. They also raise the question of whether keratinocytes of the immuno-privileged anagen hair follicle can present lipid antigens to natural killer T cells. These data could help provide new strategies for the manipulation of hair related disorders.

Background: Lichen planopilaris (LPP) is the prototype of scarring alopecias that mainly target the infundibuloisthmic (bulge) region of hair follicle. Hair follicle stem cells have been shown to reside in the bulge., Methods: We carried out this study to better define the possible pathogenetic role of the bulge in LPP. Thirty-five cases of LPP were studied. Multiple serial sections of biopsy specimens stained with hematoxylin and eosin, periodic acid Schiff-diastase, and Elastic van Gieson. The following immunostains were applied: CD3, CD4, CD8, CD1a, and Ki-67. Uninvolved follicles and normal scalp biopsy specimens served as normal controls., Results: All cases showed a lichenoid lymphocytic infiltrate at the bulge region. The bulb area was spared. CD8(+) T cells were increased compared with CD4(+) T-cell population. Langerhans' cells were decreased. Proliferating stem cells, highlighted by Ki-67, showed a marked decrease in the bulge compared with uninvolved follicles., Conclusion: Our study supports the finding that in LPP, the inflammatory infiltrate mainly involves the bulge region, where the stem cells reside. Once this area is damaged, the hair loses its potential of regrowth with resulting scarring alopecia. This is in contrast with inflammatory non-scarring alopecias such as alopecia areata, where the bulb region is targeted, sparing the stem cells.

Alopecia is a common dermatological condition in humans and other mammals. Here, we present two similar but histologically distinct mouse models of scarring alopecia. Both mutant lines were generated using random genome-wide N-ethyl-N-nitrosourea mutagenesis, and both harbor dominant mutations on chromosome 11. In both mutants, there is an early onset of alopecia that progresses to nearly complete pelage hair loss in both males and females by 20 weeks of age. Histologically, there is an increased dermal cellularity due to inflammatory cell infiltration at 7-10 days of age. By 3 weeks of age, the epidermis is acanthotic and the dermis is approximately twice as thick as in control mice due to a substantial, mostly mononuclear, inflammatory cell infiltrate. This infiltrate becomes more perifollicular by 4-5 weeks of age but is localized differently in the two mutants. In alopecia 1 (Alo-1), the perifollicular infiltrate is confined to the portion of the follicle within the dermis, whereas in Alo-2, the infiltrate extends the full length of the follicle. Expression of major histocompatibility complex (MHC) class I on the follicular epithelium in the two mutants is much greater than that in non-mutants. Furthermore, MHC class I expression is localized differently in the two mutant lines and mirrors the pattern of the inflammatory infiltrate. Despite these differences, the clinical progression of alopecia is identical in both mutants. The early onset of the disease, predictable progression, and differences in inflammatory cell localization between the two mutants make these mice particularly useful models for inflammatory hair loss and autoimmune diseases in general.

Understanding how selected tissue sites establish immune privilege (IP) is of interest to both basic immunology and clinical medicine: it provides novel insights into autoimmunity, fetal and allotransplant rejection and tumor escape from immunosurveillance. Here, we review why the hair follicle can serve as a uniquely accessible, widely available and instructive model for studying the establishment, maintenance, collapse and restoration of IP. The hair follicle epithelium rhythmically generates, maintains and deconstructs an area of relative IP, characterized by very low expression of MHC Ia and suppressed MHC II-dependent antigen presentation, accompanied by the local production of potent immunosuppressants capable of downregulating MHC I (e.g. transforming growth factor-beta1, alpha-melanocyte-stimulating hormone). We discuss the physiological functions of hair follicle IP, illustrate its clinical and therapeutic relevance by focusing on alopecia areata, an autoimmune hair loss disorder, and outline important unanswered questions for future research into one of nature's most intriguing and abundant, yet commonly ignored, sites of IP.

CD200 (OX-2) is a transmembrane glycoprotein that transmits an immunoregulatory signal through the CD200 receptor (CD200R) to attenuate inflammatory reactions and promote immune tolerance. CD200 expression in the skin has not been described previously. We now report that freshly isolated cells of the murine epidermis contain a subpopulation of major histocompatibility complex (MHC) class II-negative, CD3-negative keratinocytes that are CD200-positive. CD200 expression was accentuated in keratinocytes comprising the outer root sheath of the murine hair follicle (HF). When syngeneic skin grafts were exchanged between gender-matched wild-type (WT) and CD200-deficient C57BL/6 mice, significant perifollicular and intrafollicular inflammation was observed, eventually leading to the destruction of virtually all HF (alopecia) without significant loss of the CD200-negative grafts. Minimal and transient inflammation was observed in WT grafts, which persisted long term with hair. There was a 2-fold increase in graft-infiltrating T cells in CD200-deficient skin at 14 d. Alopecia and skin lesions were induced in CD200-deficient hosts by adoptive transfer of splenocytes from WT mice previously grafted with CD200-negative skin, but not from mice grafted with WT skin. Collectively, these results suggest that the expression of CD200 in follicular epithelium attenuates inflammatory reactions and may play a role in maintaining immune tolerance to HF-associated autoantigens.

The four isoforms of desmosomal cadherin desmogleins (Dsg1-4) are expressed in epithelial tissues in a differentiation-specific manner. Extensive sequencing of the human genome has revealed only one copy of the Dsg1 gene. However, we recently cloned two novel additional mouse Dsg1 genes, Dsg1-beta and -gamma, which flank the original Dsg1-alpha on chromosome 18. Sequence conservation between the Dsg1 isoforms diverged significantly at exon 11, particularly in the region that encodes for the extracellular anchoring (EA) domains. Computational analysis revealed very low hydrophilic potential of the Dsg1-gamma EA compared with the corresponding sequences of Dsg1-alpha and -beta, suggesting that the Dsg1-gamma EA domain may have a stronger affinity to the cell membrane. We generated antibodies using synthetic peptides or recombinant proteins localized within the EA domains. These antibodies were tested for their specificity and were then used to demonstrate expression of Dsg1 isoforms in various tissues. In the epidermis, all Dsg1 isoforms were differentially expressed in the differentiating cell layers. In the hair follicle, all Dsg1 isoforms were present throughout the entire process of its development and cycling but the expression of Dsg1 isoforms is subject to significant hair cycle-dependent changes. Dsg1-beta and -gamma, but not Dsg1-alpha, were detected in the sebaceous gland epithelium and the stratified epithelium of the stomach. Finally, Dsg1-alpha and Dsg1-beta, but not Dsg1-gamma, are proteolytically cleaved by exfoliative toxin A. These results suggest that the developmental complexity of mouse tissues, including skin and hair, may play a significant role in the evolutionary driving force to maintain multiple Dsg1 genes in mouse.

The collapse of major histocompatibility complex (MHC) class-I-dependent immune privilege can lead to autoimmune disease or fetal rejection. Pragmatic and instructive models are needed to clarify the as yet obscure controls of MHC class I down-regulation in situ, to dissect the principles of immune privilege generation, maintenance, and collapse as well as to develop more effective strategies for immune privilege restoration. Here, we propose that human scalp hair follicles, which are abundantly available and easily studied, are ideally suited for this purpose: interferon-gamma induces ectopic MHC class I expression in the constitutively MHC class-I-negative hair matrix epithelium of organ-cultured anagen hair bulbs, likely via interferon regulatory factor-1, along with up-regulation of the MHC class I pathway molecules beta(2)microglobulin and transporter associated with antigen processing (TAP-2). In the first report to identify natural immunomodulators capable of down-regulating MHC class I expression in situ in a normal, neuroectoderm-derived human tissue, we show that ectopic MHC class I expression in human anagen hair bulbs can be normalized by treatment with alpha-MSH, IGF-1, or TGF-beta1, all of which are locally generated, as well as by FK506. These agents are promising candidates for immune privilege restoration and for suppressing MHC class I expression where this is clinically desired (eg, in alopecia areata, multiple sclerosis, autoimmune uveitis, mumps orchitis, and fetal or allograft rejection).

Background: Alopecia areata (AA) is suspected to be an autoimmune disease directed preferentially against hair follicles (HF) affecting both humans and various mammalian species. Recently, two rodent models of AA were described, namely the ageing C3H/HeJ mouse and the DEBR rat. Despite several case reports of canine AA in the literature, there has been no systematic assessment of the disease in these companion animals, and it is also not known whether dogs with AA could be useful as an outbred homologue of this disease in humans., Objectives: To evaluate the clinical, histopathological and immunopathological features of 25 dogs with AA and compare these data with those found in the human disease., Patients/methods: Twenty-five client-owned dogs exhibiting macroscopic alopecia with peri- or intrabulbar lymphocytic infiltrates were selected for study. Biopsies and sera were obtained and assessed by histopathology, direct immunofluorescence of immunoreactant deposition, immunohistochemistry for lymphocyte markers, indirect immunofluorescence and immunoblotting analysis of circulating serum IgG, selective immunoprecipitation of HF proteins by serum IgG, and passive transfer of purified canine IgG into naïve C57BL/10 mice., Results: Clinical signs including alopecia, skin hyperpigmentation and leucotrichia usually developed during adulthood and were first seen on the face, followed by the forehead, ears and legs. Spontaneous remission of alopecia occurred in 60% of dogs and regrowing hair shafts were often non-pigmented. Histological examination of skin biopsy specimens revealed peri- and intrabulbar mononuclear cell infiltrates affecting almost exclusively anagen HF. Direct immunofluorescence analysis detected HF-specific IgG in 73% of dogs, while indirect immunofluorescence revealed circulating IgG autoantibodies to the HF inner and outer root sheaths, matrix and precortex. Immunoblotting analysis revealed IgG reactivity to proteins in the 45-60 kDa molecular weight range and with a 200-220 kDa doublet. The latter was identified as trichohyalin by selective immunoprecipitation. Purified HF-reactive IgG, pooled from AA-affected dogs, was injected intradermally to the anagen skin of naïve mice where it was associated with the local retention of HFs in an extended telogen phase in AA-treated skin compared with that seen in controls., Conclusions: These findings are very similar to those reported for human AA patients; therefore, they support the consideration of dogs with AA as a useful homologue for the study of the pathogenesis of this common autoimmune disease of humans.

Acne a disease of the pilosebaceous unit is characterised by hypercornification and hyperkeratosis of outer root sheath (ORS) and sebaceous duct and perilesional infiltrate. Lesions may be characterised as "non"-inflammatory versus inflammatory. Hypercornification of the distal ORS and the pilosebaceous duct in concert with increased sebum production and abnormalities of the microbial flora are considered to be major factors in the pathogenesis of acne vulgaris. However, the basic mechanisms involved in the development of inflammation during acne vulgaris remain unclear. We have investigated the expression patterns of two antimicrobial peptides, human beta-defensin 1 (hBD1) and human beta-defensin 2 (hBD2) in healthy human hair follicles as well as in peri- and intralesional skin of acne vulgaris lesions such as comedones, papules and pustules. Strong hBD1 and hBD2 immunoreactivity was found in all suprabasal layers of the epidermis, and all permanent compartments of the hair follicle including the distal ORS of the hair follicle and the pilosebaceous duct. Moreover, marked hBD1 and hBD2 expression was also detected in the hair follicle stem cell compartment. In contrast, the proximal follicle bulb which undergoes apoptotic regression and is also able to regenerate following injury did not express hBD1 or hBD2. The majority of acne biopsies displayed a marked upregulation of hBD2 IR in the lesional and perilesional epithelium; in particular in pustules, and a less marked upregulation of hBD1 IR. The upregulation of beta-defensins expression in acne vulgaris lesions when compared to controls suggests that beta-defensins may be involved in the pathogenesis of acne vulgaris.

There is strong evidence indicating that alopecia areata is a tissue-specific, autoimmune disease. Hair loss is associated with a perifollicular lymphocytic infiltrate made up primarily of CD4+ cells, along with a CD8+ intrafollicular infiltrate. Evidence of immune activation includes expression of HLA-DR; HLA-A,B,C; and ICAM-1 on the follicular epithelium. It is likely that the follicular expression of HLA-DR and ICAM-1 is induced by interferon-gamma produced by T cells. Antibodies to follicular epithelium are often present, but their significance is not known. Lesional scalp from alopecia areata patients grafted onto nude mice regrows hair coincident with a loss of infiltrating lymphocytes from the graft. Hair loss can be transferred to human scalp explants on SCID mice by injection of lesional T cells. It is necessary to activate the T cells by culture with follicular autoantigens. Melanocyte-associated antigens are also capable of activating T cells to induce hair loss, suggesting that they are capable of functioning as autoantigens for alopecia areata. Parallel evidence in rodent models of spontaneous alopecia areata also strongly supports a role for T cells in the pathogenesis of this condition.

It has been recognized for over a century that the eye is endowed with remarkable properties that permit the long-term survival of foreign tumor and tissue grafts that are normally rejected at extraocular sites. This ocular immune privilege was originally attributed to a putative sequestration of antigens in the eye as a result of the conspicuous absence of intraocular lymphatic drainage channels. In the last 30 years, a sizeable body of information indicates that ocular immune privilege is a product of multiple anatomical, physiological, and immunoregulatory processes. Ocular tissues and fluids express a wide variety of anti-inflammatory and immunosuppressive molecules, including CD95L (FasL), transforming growth factor-beta, macrophage migration inhibitory factor, alpha-melanocyte-stimulating hormone, calcitonin gene-related peptide, somatostatin, and complement regulatory proteins. Moreover, antigens entering the anterior chamber of the eye evoke a unique form of immune deviation that culminates in the antigen-specific suppression of TH1 immune responses. Finally, the intraocular milieu contains both cell membrane and soluble factors that inhibit both the adaptive and innate immune systems. The hair follicle is also recognized for its immune privilege. Like the anterior chamber of the eye, it produces anti-inflammatory and immunosuppressive cytokines, such as transforming growth factor-beta and adrenocorticotrophic hormone. The cells of the hair follicle display limited expression of class Ia MHC molecules and, like cells that line the anterior chamber of the eye, are protected against CD8+ cytotoxic T lymphocyte attack. Gaining a better understanding of the immune privilege of the hair follicle may provide insights into the regulation and pathogenesis of immune-mediated diseases of the skin.

Alopecia areata is a common disfiguring hair loss disorder that primarily affects the hair follicle as it enters the prolonged growth phase called anagen. The last few years have yielded an explosion of more rigorously obtained data on the etiology and pathogenesis of this disorder. While a consensus is rapidly building in support of an autoimmune pathogenesis, there are still several enigmatic issues to be resolved. These include the possibility that alopecia areata is really a multientity disorder with causes that are multifactorial. This will have important implications for the research scientist's search for the jigsaw puzzle's largest missing piece--the identification of the target autoantigen(s). There is now much evidence that autoimmune diseases with both T and B cell components have shared target autoantigens/epitopes. It is likely that alopecia areata is similar, as there is now very strong evidence for the generation of autoantibodies as well as autoreactive T cells to hair follicles in the pathogenesis of this disease. The following brief review outlines the progress we have made over the last five to ten years in the characterization of hair follicle antigens targeted by antibodies in alopecia areata. Results of these studies now show that the elicitation of antibodies to hair follicle-specific proteins is a highly conserved phenomenon in all affected species studied to date. Candidate autoantigens that have been identified include the 44/46 kDa hair-specific keratin (expressed in the precortical zone of anagen hair follicles) and trichohyalin (an important intermediate filament-associated protein) expressed in the inner root sheath of the growing hair follicle. Moreover, there is evidence that anti-hair follicle antibodies are modulated during the disease process, can occur before clinically detectable hair loss, and may be reduced in titer during successful treatment. Preliminary data from passive transfer experiments suggest that in some species these antibodies may disrupt hair cycling. We are currently applying a more molecular approach (e.g., cDNA library screening) to identify hair follicle antigens truly associated with the onset of the disorder.

This essay reviews the available evidence that the proximal hair follicle epithelium generates and maintains an area of relative immune privilege during a defined segment of the hair cycle (i.e., during anagen). This immune privilege is chiefly characterized by a very low level of expression of MHC class Ia antigens and by the local production of potent immunosuppressive agents, such as alpha-MSH and TGF-beta1. We discuss the putative functions of immune privilige of the anagen hair bulb, favoring the view that immune privilege serves mainly to sequester anagen- and/or melanogenesis-associated autoantigens from immune recognition by autoreactive CD8+ T cells. On this basis, we develop how the "immune privilege collapse model" of alopecia areata pathogenesis was conceived. In our discussion of the clinical implications of immune privilege, we outline the currently available evidence in support of this still hypothetical scenario to explain the initiation, progression, and termination of alopecia areata lesions. We review the most recent evidence from our laboratory that alpha-MSH, IGF-1, and TGF-beta1 can downregulate IFN-gamma-induced ectopic MHC class I expression in human anagen hair bulbs in vitro. Finally, we suggest that hair follicle-derived alpha-MSH, IGF-gamma, and TGF-beta1 form part of a constitutively active "IP restoration machinery" of the anagen hair bulb, which we propose to be recruited whenever the hair follicle suffers immune injury. Finally, we sketch some particularly promising avenues for future investigation into the far too long ignored hair follicle immune privilege.

Alopecia areata is a T-cell mediated autoimmune disease directed against an unknown auto antigen of the hair follicle. There is a genetic predisposition to develop alopecia areata, whereas environmental triggers have so far not been identified. The diagnosis can be established by characteristic clinical features of alopecia areata including its severe forms alopecia areata totalis and universalis. Nail changes may help confirm the diagnosis. On rare occasions a histopathological examination may be necessary, whereas other laboratory investigations are unnecessary. Because of the high rate of spontaneous remission, the efficacy of a rational treatment of alopecia areata has to be proven in controlled studies and it should be associated with only minor side effects. According to the rules of evidence-based medicine, treatment with a contact sensitizer is at present the most effective treatment of alopecia areata showing only mild side effects. However, it is time-consuming and in some cases ineffective, making it desirable to develop new, more specific forms of treatment.

The earliest subclinical acne "lesion" is a microcomedone, of which hyperproliferation of the follicular epithelium is a characteristic feature. Inflammatory cells have been observed at the periphery of these "lesions". This study investigated whether inflammatory events occur pre or post hyperproliferative changes. Cellular, vascular, and proliferative markers were examined by immunohistochemical techniques on biopsies of clinically normal follicles from uninvolved skin and early inflamed lesions from acne patients. Control follicles were obtained from non-acne subjects. Follicles from uninvolved skin exhibited no microcomedonal features. Proliferation in the epithelium was comparable to controls and was significantly lower than in inflamed lesions. Numbers of CD3+, CD4+ T cells were elevated in the perifollicular and papillary dermis although levels were not equivalent to those in papules. The number of macrophages was also greatly increased and similar to those in papules. There were no changes in blood vessel numbers or vascular intercellular adhesion molecule 1 expression but E-selectin expression was increased to levels found in papules and vascular adhesion molecule 1 levels were upregulated. Levels of the pro-inflammatory cytokine interleukin-1 were also upregulated perifollicularly. Moreover, aberrant integrin expression was demonstrated in the epidermis around these uninvolved follicles and inflamed lesions whereas the basement membrane was still intact. These results provide novel evidence for vascular endothelial cell activation and involvement of inflammatory responses in the very earliest stages of acne lesion development.

Exponentially growing Meth-A cells expressing H-2K(d).D (d) antigen were found to induce alopecia when injected intraperitoneally into normal C57BL/6 mice, which express the H-2K(b).D (b) antigen. However, the capacity to induce alopecia disappeared when Meth-A cells were treated with K252a, which inhibits protein kinases. Histologically, skin in affected areas showed dense mononuclear cell infiltration and a focal foreign-body giant-cell reaction in hair follicles. The subtyping of lymphocytes in peripheral blood demonstrated a significant difference between normal mice and Meth-A cell-injected mice. To further examine the mechanism by which the alloantigen induces alopecia, lymphocytes isolated from the peripheral blood of normal C57BL/6 mice were cultured in medium containing Meth-A cell homogenate, phytohemagglutinin (PHA) and recombinant mouse interleukin-2 (rm IL-2), and intravenously injected into normal C57BL/6 mice. The adoptive transfer of the lymphocytes induced alopecia in a similar way. These findings suggest that the protein kinase-modulated alloantigen induces alopecia by disturbing the immunological homeostasis, and that lymphokine-activated killer cells play an important role in induction of alopecia by cross-reacting with hair follicles.

Background: Folliculotropic mycosis fungoides (MF) is a rare variant of cutaneous T-cell lymphoma, MF type, characterized by atypical lymphocytes preferentially infiltrating the hair-follicle epithelium relative to the epidermis., Observations: We describe a rare case of folliculotropic MF involving the central nervous system. This is also the first case in which laser capture microdissection was used to show that the atypical lymphocytes within the hair-follicle epithelium were part of the same tumor clone present in other tissue compartments., Conclusions: In reviewing the literature describing atypical lymphocytes infiltrating hair-follicle epithelium relative to the epidermis, we encourage the use of the term folliculotropic mycosis fungoides. Our case also supports previous findings that central nervous system involvement can occur in advanced MF. The successful procurement and analysis of atypical lymphocytes from hair-follicle epithelium by laser capture microscopy ushers in a new era in molecular diagnostics.