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Article
Cytokine mRNA Levels in Alopecia Areata Before and After Treatment with the Contact Allergen Diphenylcyclopropenone
- October 1994
- Journal of Investigative Dermatology 103(4):530-533
Abstract
Although the nature of the noxious signal and the anatomical target in alopecia areas (AA) are still unknown, it has been assumed that CD4+ T lymphocytes surrounding and infiltrating the hair bulb might trigger the hair loss. As these T lymphocytes do not promote cytotoxic activity we hypothesize that AA is triggered by cytokines. Topical immunotherapy with diphenylcyclopropenone (DCP) is at present the most effective approach. If it is true that AA results from a distinct cytokine pattern, we can hypothesize that the beneficial effect of DCP should be mediated by locally secreted cytokines during the contact allergy. Using semiquantitative reverse transcription-polymerase chain reaction with RNA extracted from scalp biopsies from patients with AA before and after successful treatment with DCP, and from healthy controls we detected a T-cell response with increased steady state mRNA levels for interferon (IFN)-, interleukin (IL)-1, and IL-2 in untreated AA of the totalis type. After DCP treatment, the IFN- expression was reduced but still above the constitutive level found in controls, whereas mRNA expression of IL-2, IL-8, IL- 10, and tumor necrosis factor- was increased. Our results point towards cytokines involved in the pathogenesis in AA. A TH1 type cytokine pattern is present in untreated AA, and this is modified by cytokines secreted during DCP treatment. IL-10 has recently been described as an immunomodulator of the TH1 response and, therefore, we hypothesize that basal keratinocytes or lesional T cells secrete bioactive IL-10 after DCP application, resulting in an inhibitory effect on lesional T lymphocytes. This hypothesis would explain the effectiveness of DCP and implies the theoretical possibility of a response to topical or intralesional application of recombinant IL-10.Keywords: alopecia areata, diphenylcyclopropenone
... The expression of HLA-DR/HLA-ABC molecules and intercellular adhesion molecule-1 in the aVected hair follicles in AA suggests local release of IFN- [11,3,10]. As a matter of fact, increased levels of Th1 type cytokine (IFN-and IL-2), and IL-1 mRNA in the lesional skin have been reported [6]. On the other hand, AA is frequently associated with atopic diseases [12], which are thought to be supported by the Th2 type immune response. ...
... On the other hand, AA is frequently associated with atopic diseases [12], which are thought to be supported by the Th2 type immune response. Besides, the intralesional over-expression of IL-10 and TGF-1 mRNA [6], which are known to be suppressive cytokines, has been reported after successful treatment with contact allergen, diphenylcyclopropenone in patients with AA. ...
... IFN-has been considered to be a key cytokine in pathogenesis of AA [10,3,11,6]. Elevated levels of serum IFN- [2] and expanded population of IFNexpressing cells in PBMC of patients with AA [15] thus suggest that the peripheral blood might reXect the role of IFN-in the pathogenesis of AA. ...
Although infiltrating lymphocytes in the lesion of alopecia areata are considered to play an important role in the pathogenesis, specific functions of these lymphocytes in the disease have been rarely studied.We studied spontaneous expression of IL-4 and IFN- mRNA in peripheral blood mononuclear cells, which might reflect characteristics of lymphocytes in the lesional skin, from eighteen patients and eight healthy individuals using semi-quantitative RT-PCR method. Amplification signals for IL-4 were slightly higher, while those for IFN- were slightly lower in the patients than in healthy controls. Among the patients, the intensities of amplification signals for either IL-4 or IFN- in those with severe symptoms were much stronger than those with mild symptoms. These results suggested that lymphocytes in the patients with severe symptoms were activated in vivo. Besides, different levels of the expression of IL-4 and IFN- mRNA, Th2 and Th1 cytokine respectively, in the severe condition, suggested the heterogeneous pathogenesis of alopecia areata.
... Lymphocytic infiltrates including both CD4+ and CD8+ T-cells exist in and around the hair follicles [1,3]. T helper 1 (Th1) cytokines like IL-2 and IFN-í µí»¾ play an important role in the disease process [2,4]. The most popularly accepted mechanism for this disease is loss of immune privilege in the hair bulbs. ...
... Several studies have suggested that cytokines may play an important role in the disease process. A Th1 cytokine profile with elevated levels of TNF-í µí»¼ [18], IL1, IL-2 [4], and IFN-í µí»¾ [2,4] and low levels of TGF í µí»½1 due to improper function of regulatory T-(T reg) cells [19] has been reported in this disease. IFN-í µí»¾ may play an important role in the pathogenesis of AA, and IFN-í µí»¾ level may be an indicator of disease activity [2]. ...
... Several studies have suggested that cytokines may play an important role in the disease process. A Th1 cytokine profile with elevated levels of TNF-í µí»¼ [18], IL1, IL-2 [4], and IFN-í µí»¾ [2,4] and low levels of TGF í µí»½1 due to improper function of regulatory T-(T reg) cells [19] has been reported in this disease. IFN-í µí»¾ may play an important role in the pathogenesis of AA, and IFN-í µí»¾ level may be an indicator of disease activity [2]. ...
Background. Alopecia areata (AA) is a chronic inflammatory disease of the hair follicle. The exact pathogenesis of AA remains unknown, although recent studies support a T-cell mediated autoimmune process. On the other hand, some studies have proposed that the renin-angiotensin-aldosterone system (RAAS) may play a role in autoimmunity. Therefore, we assessed serum activity of angiotensin converting enzyme (ACE), a component of this system, in AA. Methods. ACE activity was measured in the sera of 19 patients with AA and 16 healthy control subjects. In addition, the relationship between severity and duration of the disease and ACE activity was evaluated. Results. Serum ACE activity was higher in the patient group (55.81 U/L) compared to the control group (46.41 U/L), but the difference was not statistically significant (P = 0.085). Also, there was no correlation between ACE activity and severity (P = 0.13) and duration of disease (P = 0.25) in the patient group. Conclusion. The increased serum ACE activity found in this study may demonstrate local involvement of the RAAS in the pathogenesis of AA. Assessment of ACE in a study with a larger sample size as well as in tissue samples is recommended in order to further evaluate the possible role of RAAS in AA.
... Increases in number of CD1a + dendritic cells means that the emigration of the antigen presenting cell is disturbed by DPCP application 22 . DPCP treatment affects not only T cells but also the compositions of cytokines around hair follicles 23 . In a study of AA patients treated with DPCP for at least 6 months, no differences were found between the levels of IL-12 and IFN-gamma in peripheral blood mononuclear cells in patients who responded to the treatment as com-pared with healthy controls, suggesting a loss of upregulation of the activation markers. ...
... In a study using scalp biopsies from patients with alopecia totalis before and after successful DPCP treatment, T helper 1 cytokines, such as, IFN-gamma, IL-1 beta, and IL-2 were found to be elevated in untreated patients, but after DPCP treatment IFN-gamma decreased and IL-2, IL-8, IL-10, and TNF-alpha levels increased. Hoffmann et al. 23 proposed that the T helper 1 cytokine pattern was observed in untreated patients and suggested that IL-10 might be effective at inhibiting T helper 1 response. IL-10 is known to act as an anti-inflammatory cytokine in association with Treg and T helper 2 cells 10 . ...
... IL-10 is known to act as an anti-inflammatory cytokine in association with Treg and T helper 2 cells 10 . After DPCP treatment, IL-10 was reported to be increased in scalp tissues 23 , and a continuous increase in the expression of IL-10 in peripheral blood mononuclear cells was observed than controls, whereas this was not observed for IL-12 or IFN-gamma 12 . However, some consider AA relatively restrained in IL-10 knockout mice, which suggests IL-10 acts to both contain and promote sensitivity to AA 25 . ...
Background
Contact immune modulating therapy with diphenylcyclopropenone (DPCP) is a topical treatment option for extensive alopecia areata (AA). Because the response to DPCP treatment varies according to the patient, and it takes several months to evaluate the clinical effectiveness of the treatment, it is necessary to identify the factors that can predict the prognosis of the disease while treating with topical DPCP.
Objective
In this study, cytokine levels in the scales of alopecic patches were investigated to identify whether they could predict response to DPCP during the early treatment period.
Methods
Scale samples were taken from the alopecic patches in eight AA patients at 1 week, 2 months, and 4 months after DPCP sensitization. The patients were divided into responders and non-responders according to the clinical responses of DPCP treatment. Interferon (IFN)-gamma, interleukin (IL)-2, IL-12 and IL-10 levels of the subjects were compared in several perspectives.
Results
Cytokine levels after 1 week of DPCP sensitization showed no statistically significant difference between two groups. After 4 months of treatment, IFN-gamma levels were significantly lower in responders than in non-responders.
Conclusion
The results of this study show IFN-gamma levels in the scales of alopecic patches might possibly reflect the clinical response in AA patients treated with DPCP. However, initial cytokine levels could not predict the treatment response.
... Arps and Kφlsch[29] reported that IL-10 deficient mice showed elevated ratios of CD4+ : CD8+ T cells, indicating a higher capacity to provide B cell activation, which results in a strongly elevated IgE response. In accordance with this observation, intralesional under-expression of IL-10 mRNA, was demonstrated in patients with AA.[18] Thus, elevated serum IgE in AA patients may reflect IL-10 deficiency-associated B cell stimulation. Further studies on a larger scale are required to prove this observation. ...
... Although, increased levels of Th1 cytokines (interferon-γ (IFN-γ) and IL-2) in lesional AA skin have been reported,[18] Th2 immune response was also incriminated in the pathogenesis of AA.[1920] ...
Context:Alopecia areata (AA) is a common form of localized, non-scarring hair loss. The pathogenesis of the disease is unknown. Previous evidence suggested the involvement of Th2 cytokines in disease pathogenesis.Aim:To determine serum level of total IgE, this is mainly influenced by Th2 cytokines, in Egyptian patients with AA.Materials and Methods:Fifty subjects with AA (28 males and 22 females) were selected from Dermatology Outpatient Clinic, Menoufiya University Hospital from February 2012 to December 2012. Subjects with other conditions that might elevate serum IgE were excluded from the study. Fifty age- and sex-matched healthy subjects were selected as a control group. Venous blood samples were taken from cases and controls for measurement of total serum IgE by enzyme-linked immunosorbent assay. Skin biopsy was taken from every case from an active area of hair loss.Results:Total serum IgE was elevated in 27 (54%) cases. Its values among patients ranged from 13.5 IU/ml to 780 IU/ml. There was a statistically significant difference between cases and controls with regard to mean value of serum IgE (P < 0.05). Mean value of IgE did not vary significantly with disease severity, patients’ age, patients’ gender, disease duration, site of lesions, and positive family history of AA. No correlation was found between serum IgE levels and histopathological changes detected in examined cases.Conclusions:Total serum IgE is elevated in AA. This elevation is not related to age, gender, disease duration, disease severity, site of affection or family history of AA.
... Of note, TLR1 polymorphisms may be related to AA susceptibility. [34] Type II interferon IFNγ plays an important role in AA [35][36][37] ; ...
Alopecia areata (AA) is thought to be an autoimmune process. In other autoimmune diseases, the innate immune system and Toll‐like receptors (TLRs) can play a significant role. Expression of TLR7, TLR9, and associated inducible genes were evaluated by quantitative PCR in peripheral blood mononuclear cells (PBMCs) from 10 healthy individuals and 19 AA patients, categorized according to disease duration, activity, and hair loss extent. Microdissected scalp biopsies from 5 patients and 4 controls were also assessed by quantitative PCR and immunohistology. TLR9 was significantly upregulated 2.37 fold in AA PBMCs. Notably, TLR9 was most significantly up regulated in patients with active AA, as shown by a positive hair pull test, compared to stable AA patients. In hair follicle bulbs from AA patients, IFNG and TLR7 exhibited statistically significant 3.85 and 2.70 fold increases in mRNA respectively. Immunohistology revealed TLR7 present in lesional follicles, while TLR9 positive cells were primarily observed peri‐bulbar to AA affected hair follicles. The increased expression of TLR7 and TLR9 suggest components of the innate immune system may be active in AA pathogenesis.
... In these treatments, Japanese guideline strongly recommends contact immunotherapy and local injection of corticosteroid in the hair loss lesions as B level [28]. Although the mechanism of contact immunotherapy is still remained to be elucidated, Th1/Th2 cytokine balance might be affected as downregulation of IFN-γ and up-regulation of IL-4 [35]. Therefore, it can be speculated that the chemotactic activity of CXCR3 + CD4 + Th1 cells and CXCR3 + CD8 + Tc1 cells might be down-modulated by contact immunotherapy although there is no in vivo/in vitro evidence of suppressive effect in contact immunotherapy. ...
Alopecia areata (AA) is an organ-specific and cell-mediated autoimmune disease. Recent studies have suggested the most important effector cell of AA is NKG2D+CD8+T cells, and outer root sheath (ORS) cells highly express NKG2D ligands, such as MICA, in AA lesions. T lymphocytes densely surround lesional hair bulbs, which is histologically referred to as “swarm of bees”. Immunohistochemical and real-time RT-PCR studies reveal that hair follicles of acute-phase AA expressed a high level of Th1-associated chemokine CXCL10. In the skin lesions of acute-phase AA, CXCR3+CD4+ and CXCR3+CD8+ T cells infiltrated in the juxta-follicular area. In chronic-phase AA, CXCR3+CD8+ T cells dominated the infiltrate around hair bulbs, possibly contributing to the prolonged state of hair loss. Lymphocytes obtained from a lesional skin of acute-phase AA contained CXCR3+CD4+ and CXCR3+CD8+ T cells at higher percentages than those of PBMCs, suggesting preferential emigration from the blood. Furthermore, freshly isolated PBMCs from acute-phase AA patients had a strong velocity of chemotaxis toward CXCL10 with increased expression of F-actin. Antihistaminic drugs have been used in Japan, and these have possibility to downregulate chemotactic activity in AA. Olopatadine shows suppressive effects of chemotactic activity in the AA patients’ CD4+ and CD8+ T cells towards CXCL10 by reducing CXCR3 expression, F-actin polymerization, and Ca++ influx. In conclusion, the increased production of CXCL10 from hair follicles induces preferential infiltrates of Th1 and Tc1 cells in the acute phase of AA, and Tc1 infiltration remains prolonged in the chronic phase. Therefore, inhibitory treatment of chemotactic activity might be novel target for the treatment of AA.
... Furthermore, DPCP alters the cytokine profile in treated alopecic scalp, in particular increasing IL-2 and IL-10 expression. (Hoffmann et al., 1994) This increased IL-10 expression has been hypothesized to inhibit the lesional T-cells of alopecia areata, and our data confirm an upregulation of IL-10 expression, as well as increases in CD8+ T-cells, in normal human skin treated with DPCP. ...
Diphencyprone (DPCP) is a hapten that induces delayed-type hypersensitivity (DTH) reactions. It is used as an immune modulating therapeutic, but its molecular effects in human skin are largely unknown. We studied cellular and molecular characteristics of a recall response to 0.04% DPCP at 3 day (peak) and 14 day (resolution) timepoints using immune markers, RT-PCR and gene array approaches. A peak response showed modulation of ~7,500 mRNA transcripts, with high expression of cytokines that define all major effector T-cell subsets. Concomitant increases in T-cell and CD11c+ dendritic cell (DC) infiltrates were measured. The resolution reaction was characterized by unexpectedly high levels of T-cells and mature (DC-LAMP+) DCs, but with marked decreases in expression of IL-2, IFNγ, and other T-cell derived cytokines. However, negative immune regulators such as IDO1 that were high in peak reactions, continued to have high expression in resolution reactions. In the resolution reaction, ~1,500 mRNA transcripts were significantly different from placebo-treated skin. These data suggest the response to DPCP evolves from an inflammatory/effector peak at day 3 to a more regulated immune response after 14 days. This model system could be useful for further dissection of mechanisms of immune activation or negative immune regulation in human skin.Journal of Investigative Dermatology accepted article peview online, 21 April 2014. doi:10.1038/jid.2014.196.
... Adverse effects of topical immunomodulatory therapy include local irritation, blister formation, persistent dermatitis, lymphadenopathy, generalized eczema, and urticarial reaction. Mechanism of contact immunotherapy is suggested to modulate cytokine gene expression balance, and interferon-γ expression was reduced while interleukin-2 (IL-2), IL-8, IL-10 and tumor necrosis factor-α levels were increased in the lesional skin [1]. ...
Squaric acid dibutylester (SADBE) is frequently used for the treatment of alopecia, but sometimes unwanted side effects occur. Herein we report a case which developed vitiliginous lesions induced by topical SADBE application in a patient with autoimmune thyroiditis. A 60-year-old female visited our department, complaining of diffuse alopecia of the scalp. She was suffering from chronic autoimmune thyroiditis over several years, and taking thyradin (90mg per day).
... However, in human AA, the CD4 : CD8 ratio changed from 4 : 1 before therapy to 1 : 1 after therapy. Hoffmann et al. [25] demonstrated that after treatment with a contact sensitizer, the mRNA-expression of IFN-g is reduced, whereas the expression of IL-10 is increased. Whether this is due to a Th1-Th2 shift or whether it is caused by the introduction of regulatory T cells with a type 2 cytokine profile is unclear. ...
Background Alopecia areata (AA) is a highly unpredictable, autoimmune skin disease affecting ∼1.7% of the population worldwide. Treatment for AA is generally unsatisfactory. A number of treatments can induce hair growth but none have been shown to alter the course of the disease. Objective The aim of this study was to compare the efficacy of narrowband UVB (NB-UVB) phototherapy with that of topically applied khellin followed by UVA irradiation (KUVA) in the treatment of AA. Patients and methods This was a comparative study involving 38 patients. The patients were divided into two groups, group I and group II. Patients of group I (n=19) were treated with KUVA on the scalp and those of group II (n=19) were treated with NB-UVB irradiation on the scalp. In both groups, irradiation was administered three times weekly for 24 weeks or until complete terminal hair regrowth. Results The patient response to KUVA therapy (57.89%) was statistically higher than that to NB-UVB therapy (10.52%, P<0.05) at the end of the treatment course (24 weeks). Conclusion KUVA therapy is better than NB-UVB therapy in the treatment of AA resistant to other treatment modalities.
... In fact, in the absence of iNOS activity, macrophages may acquire a nonclassical activated phenotype that may have profibrotic characteristics [48,49] and then may account for the increased collagen deposition in the implants from iNOS-deficient mice. In addition, it is worth mentioning that although IL-10 is understood as an antifibrotic cytokine, it has been suggested that IL-10 cooperates with Th1 cytokines, such as IFN-(and even TNF-), to suppress collagen deposition [50,51]. Interestingly, here, there were no changes in IFNlevels, and TNF-levels were even decreased, indicating a possible lack of the regulatory pathway for collagen deposition and, consequently, a supportive microenvironment for increasing fibrogenesis in the implants is endorsed in the absence of iNOS activity. ...
There is considerable interest in implantation techniques and scaffolds for tissue engineering and, for safety and biocompatibility reasons, inflammation, angiogenesis, and fibrosis need to be determined. The contribution of inducible nitric oxide synthase (iNOS) in the regulation of the foreign body reaction induced by subcutaneous implantation of a synthetic matrix was never investigated. Here, we examined the role of iNOS in angiogenesis, inflammation, and collagen deposition induced by polyether-polyurethane synthetic implants, using mice with targeted disruption of the iNOS gene (iNOS −/−) and wild-type (WT) mice. The hemoglobin content and number of vessels were decreased in the implants of iNOS −/− mice compared to WT mice 14 days after implantation. VEGF levels were also reduced in the implants of iNOS −/− mice. In contrast, the iNOS −/− implants exhibited an increased neutrophil and macrophage infiltration. However, no alterations were observed in levels of CXCL1 and CCL2, chemokines related to neutrophil and macrophage migration, respectively. Furthermore, the implants of iNOS −/− mice showed boosted collagen deposition. These data suggest that iNOS activity controls inflammation, angiogenesis, and fibrogenesis in polyether-polyurethane synthetic implants and that lack of iNOS expression increases foreign body reaction to implants in mice.
... Increased or unbalanced activation of Th1 or Th2 subgroups, not only compromise normal immune system function and haemostasis but also promote and support the development of AA. However, there is controversy in the literature about AA being a Th1 or Th2 autoimmune phenomenon, or both [3][4][5][6]. Thus, identifying the molecular switches that regulate differentiation of CD4+ T cells into Th1 and Th2 effector cells in diseased subjects is the keystone for the characterization of the errors in the function of the immune system. ...
Alopecia areata represents an autoimmune pathological process driven primarily by cellular aberrations contained within the immune system, which activates various humoral and cellular elements of the immune response. The aim of this study was to determine the mRNA expression levels of T-bet and GATA-3 as potential inducers of T helper (Th)1 and Th2 differentiation, respectively, as well as Th1(IFN-γ) and Th2(IL-4) cytokine mRNA expression in patients with alopecia areata. Using real-time reverse transcriptase PCR (RT-PCR), the relative amounts of T-bet, GATA-3, IFN-γ, and IL-4 mRNA transcripts were determined in PBMCs from 20 Iranian patients with alopecia areata and compared with those of 20 healthy control subjects. In comparison with the normal group, T-bet and IFN-γ mRNA expression levels were significantly up-regulated in the alopecia areata patients, while GATA-3 and IL-4 mRNA expression levels were down-regulated. Notably, positive correlation (P < 0.05) was found between IFN-γ and T-bet levels in patients and controls. In addition, significant positive correlations existed between GATA-3 and IL-4 (P < 0.05). These results indicate that a Th1/Th2 imbalance exists in alopecia areata, and it may be implicated in the pathogenesis of disease.
... There is also some other studies, which introduced CD8 + T cells and mast cells as the responsible players for AA development instead Th1 cells [5][6][7]. Heffler et al. [8] found that the DPCD contact allergy effects may be associated with the increase in matrix metalloproteinase 9. Hoffmann et al. [9] analysed the levels of some cytokines before and after DPCD therapy in AA patients. It was found that IFN-c level was decreased. ...
Alopecia areata (AA) remained a disease without any licensed treatment. Although it is not a life-threating disease, it significantly affects the quality of life of patients. Lack of effective treatment for this disease may be due to little knowledge about its exact cellular mechanism. Several studies have shown that interferon-gamma (IFN-γ) is increased in AA, which could induce major histocompatibility complex (MHC) I expression. This article is protected by copyright. All rights reserved.
... PRL is an important immunomodulator, possibly with overall pro-inflammatory effects in response to the immunosuppressive effects of stress [68]; consistent with the finding that IFNc increases PRL expression in the HF. In terms of HF pathophysiology , lesional alopecia areata skin has been shown to have increased levels of IFNc mRNA [69] and IFNc has potent catagen (HF regression) promoting effects [39]. Intriguingly, PRL can exert the same effects [1], raising the possibility that IFNc may partially exert its effects via alterations in intrafollicular PRL expression. ...
Human scalp skin and hair follicles (HFs) are extra-pituitary sources of prolactin (PRL). However, the intracutaneous regulation of PRL remains poorly understood. Therefore we investigated whether well-recognized regulators of pituitary PRL expression, which also impact on human skin physiology and pathology, regulate expression of PRL and its receptor (PRLR) in situ. This was studied in serum-free organ cultures of microdissected human scalp HFs and skin, i.e. excluding pituitary, neural and vascular inputs. Prolactin expression was confirmed at the gene and protein level in human truncal skin, where its expression significantly increased (p = 0.049) during organ culture. There was, however, no evidence of PRL secretion into the culture medium as measured by ELISA. PRL immunoreactivity (IR) in female human epidermis was decreased by substance P (p = 0.009), while neither the classical pituitary PRL inhibitor, dopamine, nor corticotropin-releasing hormone significantly modulated PRL IR in HFs or skin respectively. Interferon (IFN) γ increased PRL IR in the epithelium of human HFs (p = 0.044) while tumour necrosis factor (TNF) α decreased both PRL and PRLR IR. This study identifies substance P, TNFα and IFNγ as novel modulators of PRL and PRLR expression in human skin, and suggests that intracutaneous PRL expression is not under dopaminergic control. Given the importance of PRL in human hair growth regulation and its possible role in the pathogenesis of several common skin diseases, targeting intracutaneous PRL production via these newly identified regulatory pathways may point towards novel therapeutic options for inflammatory dermatoses.
... IL-6 is an important proinflammatory mediator produced by mast cells, which inhibits hair shaft elongation and suppresses proliferation of matrix cells in cultured human hair follicles 26,27 . IL-1β is also known to inhibit hair follicle growth, and is highy expressed in patients with active alopecia areata 28 . Interestingly, both inflammatory cytokines, IL-6 and IL-1β are suppressed by cortisol 29 . ...
Background
Stress is a known cause of hair loss in many species.
Objective
In this study, we investigated the role of acute stress on hair growth using a rat model.
Methods
Rats were immobilized for 24 hours and blood samples, and skin biopsies were taken. The effect of stress-serum on the in vitro proliferation of rat and human dermal papilla cells (hDPCs), as well as serum cortisol and corticotropin-releasing hormone levels, were measured. Mast cell staining was performed on the biopsied tissue. In addition, Western blot and quantitative real time polymerase chain reaction were used to assess mast cell tryptase and cytokine expression, respectively in rat skin biopsies.
Results
Stress-serum treatment reduced significantly the number of viable hDPCs and arrested the cell cycle in the G1 phase, compared to serum from unrestrained rats (p<0.05, respectively). Moreover, restrained rats had significantly higher levels of cortisol in serum than unrestrained rats (p<0.01). Acute stress serum increased mast cell numbers and mast cell tryptase expression, as well as inducing interleukin (IL)-6 and IL-1β up-regulation.
Conclusion
These results suggest that acute stress also has an inhibitory effect on hair growth via cortisol release in addition to substance P-mast cell pathway.
... The exact mechanism of action of the immune-modulating effect in DPCP is unclear, but to date it is generally assumed that it acts on different pathways to inhibit inflammation in alopecic areas which is achieved mainly by suppression of inflammatory cells, such as CD4+ and CD8+ T cells, restoration of equilibrium between Th1 and Th2 responses, or involvement of regulatory T cells. [36][37][38][39][40][41][42] The functional significance of elevated IL-4 in AA is unclear, but it suggests there is also a role for Th2 type cytokines in AA pathogenesis in at least some patients, as indicated by other research. [42] Increased serum IL-5 and IL-17 levels were also found both in responder and non-responder groups after treatment when compared to pre-treatment levels, as well as to controls; possibly a response caused by the DPCP induced contact dermatitis. ...
Background
This study investigated predictors of response to topical diphenylyclopropenone (DPCP) immunotherapy in patients with alopecia areata (AA).
Objective
To identify predictors of response, or resistance, to treatment for AA through clinical observations and serum tests.
Methods
84 AA patients were treated with DPCP. Serum cytokine levels were measured in 33 AA patients pre‐ and post‐treatment, and in 18 healthy controls, using ELISA assays.
Results
56.1% of patients responded to DPCP with satisfactory hair regrowth; the response rate was negatively correlated with hair loss extent. Before DPCP treatment, higher serum IFN‐γ and IL‐12 cytokine levels were observed in AA patients compared to healthy controls. Non‐responders to DPCP had significantly elevated serum IL‐4 pre‐treatment (3.07 fold higher) and lower IL‐12 levels compared with responders. After DPCP treatment, non‐responders had persistently high IL‐4, increased IL‐12, negligible decrease in IFN‐γ and decreased IL‐10. Post‐treatment DPCP responders exhibited significantly decreased IFN‐γ and IL‐12, and increased IL‐4 and IL‐10. Development of adverse side‐effects was significantly associated with higher pre‐treatment serum IgE levels.
Limitations
a small number of subjects were evaluated.
Conclusions
Potentially, elevated pre‐treatment serum levels of IL‐4 and IL‐12 can be used as unfavorable and favorable predictors of DPCP therapeutic effect, respectively. In addition, pre‐treatment elevated serum total IgE may predict increased risk for severe adverse side‐effects to DPCP application. Whether serum cytokine expression levels can be used as predictors of response to other forms of treatment is unknown, but it may warrant investigation in the development of personalized treatments for AA.
This article is protected by copyright. All rights reserved.
... Diphenylcyclopropenone or squaric acid dibutyl ester are topical sensitizers that cause contact dermatitis through hapten binding. 4 Their exact mechanism for hair regrowth in AA is not clearly elucidated, although many therapeutic mechanisms have been suggested, including immunomodula-tory effect, 8 changes in the CD4 + /CD8 + lymphocyte ratio, 9 lesional T-lymphocyte inhibition induced by T H 2-related cytokine, lymphocyte apoptosis, 10 and antigenic competition. 11 The therapeutic hair regrowth benefits of contact immunotherapy have been reported to be variable in the literature. ...
Importance
Contact immunotherapy with diphenylcyclopropenone or squaric acid dibutyl ester is a preferred treatment for severe alopecia areata; however, the defined criteria for therapeutic hair regrowth and regrowth rate have been highly heterogeneous across studies.
Objective
To summarize the clinical outcomes of contact immunotherapy for alopecia areata according to standardized criteria for therapeutic hair regrowth and several prognostic factors.
Data Source
A database search of MEDLINE, Embase, and Cochrane Library was performed for articles published before November 20, 2017, using the search terms areata, totalis, universalis, sensitizer, sensitization, immunotherapy, DPCP, diphenylcyclopropenone, diphencyprone, SADBE, and squaric.
Study Selection
Clinical trials or observational studies that investigated contact immunotherapy for alopecia areata and subgrouped the disease into patchy alopecia or alopecia totalis/universalis and reported their hair regrowth rates were included, whereas studies that investigated combination therapy or nonconventional protocol and case series or reviews were excluded.
Data Extraction and Synthesis
The following data were extracted from each of the studies included in this meta-analysis: study year and setting, sensitizer type, study population, study population composition by disease subtype, defined criteria for therapeutic hair regrowth, and regrowth rate of contact immunotherapy. The incidence of adverse effects and recurrence rate were also recorded. A random effects model was used for data synthesis because of the expected high heterogeneity of the included studies.
Main Outcomes and Measures
The main outcome was therapeutic hair regrowth rate according to the 4-grade criteria for therapeutic regrowth. Secondary outcomes included incidence of treatment-related adverse effects and recurrence rate.
Results
Forty-five studies comprising 2227 patients were analyzed. The overall rate of any hair regrowth was 65.5% among patients with alopecia areata (74.6% in the patchy alopecia and 54.5% in the alopecia totalis/universalis subgroups). However, the complete regrowth rate was 32.3% (24.9% in the patchy alopecia and 32.3% in the alopecia totalis/universalis subgroups). Disease extent of 50% or greater (odds ratio [OR], 3.05; 95% CI, 2.26-4.12), atopic history (OR, 1.61; 95% CI, 1.03-2.50), and nail involvement (OR, 2.06; 95% CI. 1.26-3.36) were associated with poorer therapeutic outcome. Recurrence rates were 38.3% among patients receiving maintenance treatment and 49.0% among those not receiving maintenance treatment.
Conclusions and Relevance
Various factors were associated with the clinical outcomes of contact immunotherapy for alopecia areata, with significant differences in hair regrowth rates according to the level of expected therapeutic regrowth. Quantitative summarization may improve patient education and lead to better therapeutic adherence and outcomes.
... Several clinical and experimental findings point towards IL-2 as a crucial mediator inducing immunologic reaction against human follicle in AA [15,16]. By using semiquantitative reverse transcription-polymerase chain reaction with RNA extracted from scalp biopsies from patients with AA, Hoffmann et al. [17] reported that IL-2 mRNA levels were increased in untreated AT. ...
Background:
Alopecia areata (AA) is a disease characterized by focally, nonscarring hair loss on the scalp or any hair-bearing surface. The etiology is unknown, although the evidence suggests that AA is an immunologically mediated disease. In the pathogenesis of AA, Th1 immune response is predominant. A special cytokine profile is created by Th1 cells, which disturbs the natural balance of the cytokine networks and leads to inflammatory reaction and follicle damage.
Objective:
The aim of our study was to evaluate serum concentrations of IL-2 in patients with AA and healthy subjects. We also examined a possible association between serum levels of IL-2, disease severity, and duration of AA.
Methods:
Sixty patients with AA and 20 healthy controls were enrolled in the study. Serum concentrations of IL-2 were measured using enzyme-linked immunoassay techniques.
Results:
Comparison of mean values of IL-2 has showed that serum concentrations of this cytokine are significantly higher in serum samples of AA patients in relation to the control group (22.2 ± 1.19 vs. 21.1 ± 2.68 pg/mL, respectively; p = 0.0142). No correlations were found between clinical type, duration of the disease, and serum levels of IL-2.
Conclusion:
Our findings support the evidence that elevation of serum IL-2 is associated with AA. The exact role of serum IL-2 in AA should be additionally investigated in future studies.
In this work we studied the reactivity of isopropylphenylcyclopropenone towards some nitrogen nucleophiles whose reactions with methylphenylcyclopropenone and diphenylcyclopropenone were previously studied. The electrochemical behavior of these cyclopropenones was evaluated for the first time, and a correlation between electrochemical parameters and reactivity of this class of compounds was done.
Autoimmune skin diseases are complex processes in which autoreactive cells must navigate through the skin tissue to find their targets. Regulatory T cells in the skin help to mitigate autoimmune inflammation and may in fact be responsible for the patchy nature of these conditions. In this review, we will discuss chemokines that are important for global recruitment of T cell populations to the skin during disease, as well as signals that fine‐tune their localization and function. We will describe prototypical disease responses and chemokine families that mediate these responses. Lastly, we will include an overview of chemokine‐targeting drugs that have been tested as new treatment strategies for autoimmune skin diseases.
Die Alopecia areata ist eine T-Zell-vermittelte Autoimmunerkrankung, die sich gegen ein bisher unbekanntes Antigen des Haarfollikels richtet. Es besteht eine genetische Prädisposition für die Erkrankung; auslösende Umweltfaktoren ließen sich bisher nicht nachweisen. Das typische klinische Bild des kreisrunden Haarausfalls einschließlich seiner Maximalformen, der Alopecia areata totalis und universalis, ermöglicht in den meisten Fällen die Diagnose, die durch das Vorliegen von Nagelveränderungen untermauert werden kann. Selten wird eine Histologie erforderlich; alle anderen Laboruntersuchungen sind überflüssig. Wegen der hohen Spontanremissionsrate muss die Wirksamkeit einer rationalen Therapie der Alopecia areata in kontrollierten Studien nachgewiesen werden und im Rahmen der Nutzen-Risiko-Abwägung ein geringes Nebenwirkungsprofil aufweisen. Nach den Regeln der evidenzbasierten Medizin ist die Behandlung mit einem Kontaktallergen derzeit die effektivste und nebenwirkungsärmste Therapie der Alopecia areata; da sie aber sehr aufwändig ist und nicht in allen Fällen wirkt, ist die Entwicklung neuer, spezifischerer Therapieformen notwendig.
Alopecia areata (AA) is an autoimmune disease resulting in the premature arrest of the follicular growth cycle clinically resulting in patchy, non-scarring hair loss. The presence of a dense follicular T cell infiltrate and variations in cytokines have led to the hypothesis that T cell activation and alterations in inflammatory mediators are crucial participants in the etiopathogenesis of the disease. Various studies suggest that the AA pathogenesis has a dominant TH1-mediated component, with potential involvement of the TH17 pathway. However, a fully integrated view of intersecting cytokine networks that support the autoimmune response in AA is lacking. A more precise understanding of cytokine pathways in disease is required to rationally explore cytokine targeted treatment strategies. Here, we comprehensively and critically review the current literature to provide a working framework regarding the complexity of T cell-cytokine interactions in AA, emphasizing the areas necessitating further research, particularly for the development of novel therapeutic options.
Topical immunotherapy with diphenylcyclopropenone (DPCP) is a treatment that can be used in patients with alopecia areata (AA) with more than 50% involvement of the scalp. The aim of this study is to assess the response of our patients with AA treated with topical immunotherapy with DPCP at the American University of Beirut-Medical Center (AUB-MC) and to characterize the favorable prognostic factors that predict response to treatment. This is a retrospective study of all patients diagnosed with AA at AUB-MC and treated with topical immunotherapy with DPCP over a period of 10 years. A total of 34 cases were included for analysis (19 males and 15 females). The majority of patients had limited AA (58.8%) with a mean of 39% of scalp involvement. The remaining patients had alopecia universalis (29.4%) and alopecia totalis (11.8%). The percentage of patients that responded to DPCP therapy in our series was 79.4% (n = 27). Ten patients achieved a maximal grade of 3 following treatment, six patients only achieved a grade of 1, and six patients achieved a grade of 2. Only five of the patients who responded to therapy achieved a grade of 4. Of the patients who responded, 10 relapsed (29.4%), and the mean time to relapse was 74.6 weeks from the initiation of treatment. No specific favorable prognostic factors were identified to predict response to treatment; however, a negative family history of atopy was found to be protective against relapse (P = 0.020). The most common side effect of therapy was itching (85.3%), followed by contact dermatitis (58.8%), blistering (17.6%), and cervical lymphadenopathy (17.6%). Limitations of this study were the retrospective nature of the study and the limited number of patients. This is, to the best of our knowledge, the first study on topical immunotherapy with DPCP in patients with extensive AA from a Middle Eastern population. This modality of treatment is effective in inducing a response in patients with extensive AA, although the response is partial in the majority of the cases. Benefits should be weighed against the high side-effect profile of therapy before initiation of treatment.
Background
Alopecia areata (AA) describes a sudden localized patchy alopecia. The cause of AA is not completely clear and its incidence may be related to genetic, autoimmune, and environmental factors.
Aim
To explore the possible mechanisms of AA and to provide a basis for the early diagnosis and treatment of AA.
Methods
Gene microarray data from 122 scalp skin biopsy tissue samples from patients with AA or healthy controls from the Gene-Cloud of Biotechnology Information database were analyzed using bioinformatics analysis methods. Molecular network analysis of the differentially expressed genes (DEGs) was conducted by Cytocluster using the Molecular Complex Detection (MCODE) algorithm.
Results
The gene expression profile of skin lesions from patients with AA was significantly altered, with 111 DEGs found in the skin lesions of AA, compared with that of the healthy skin. The DEGs were mainly related to biological processes such as the development of the epidermis and inflammatory reaction. The protein–protein interaction network analysis of DEGs revealed bone morphogenetic protein 2 (BMP2) as a core protein interaction network. BMP2 acted not only via the inflammatory response but also via the signaling pathways in epithelial cell development and epidermal cell differentiation to affect the epidermal development. MCODE analysis further showed that keratins (KRTs) and keratin-associated proteins (KRTAPs) can affect the epidermal development via the epidermal development pathway.
Conclusions
The abnormal development of the epidermis and inflammatory reactions in skin tissue play important roles in the pathogenesis of AA and are closely related to BMP2, KRTs, and KRTAPs genes.
Limitations
Our study was limited by experimental verification.
Interleukin (IL)-1 has been shown to be a potent inhibitor of hair growth in vitro. We hypothesized that this cytokine might be a decisive factor causing hair loss during the lymphocytic attack in alopecia areata. Neither the intracellular pathways involved in hair growth inhibition mediated by IL-1β nor the signal transduction processes within hair follicles in general are known. We therefore investigated the intracellular signals involved in human hair growth in vitro. Hair follicles were isolated from scalp biopsies by microdissection and hair growth was measured daily by image analysis. We assessed intracellular signal transducing elements using specific inhibitors or activators either alone or in combination with IL-1β. The calcium ionophore A 23187 induced a rapid and complete arrest of hair growth and phorbol-12-myristate-13-acetate (PMA), genistein, or IL-1β decreased hair growth by approximately 60%- 80%. IL-1β- elicited hair growth arrest was not antagonized by calphostin C, a specific inhibitor of protein kinase C. In contrast, coincubation of IL-1β with pertussis toxin or H 1004 neutralized the effect of IL-1β and dibutyryl-cAMP and cholera toxin, an activator of adenylate cyclase, inhibited hair growth. These data suggest that cAMP acts as a second messenger for IL-1β-induced inhibition of hair growth. Moreover, our data indicate that in vitro hair growth is dependent on intracellular Ca22+ levels and activation of tyrosine kinase as well as protein kinase C. We were unable to detect a signal transducing element responsible for enhanced hair growth in vitro.
Cytokines have long been postulated to play a role in alopecia areata (AA), and antigen-specific scalp T cells from patients with extensive AA have been found to have an intrinsic defect in the production of TH1 cytokines such as interferon gamma and interleukin 2 (IL-2).1,2 In the interesting study presented by Castela et al,3 5 patients with extensive AA were treated with low-dose IL-2, and 4 of the 5 patients attained considerable hair regrowth. This was accompanied by an increase in regulatory T (Treg) cells; cells defined by the expression of CD4, CD25, and transcription factor forkhead box P3 (FOXP3) in skin biopsy specimens; and no significant change in circulating Treg cells. Adverse effects in this study were reported to be minimal. The investigators in the Department of Dermatology at University Hospital in Nice, France, continue to enroll research participants and list this study as a clinical trial on ClinicalTrials.gov (NCT01840046).
The objective of this article is to discuss some facts of modern immunomodulators that might be useful for clinical dermatology. Moreover, it aims to dispel myths that might have a negative impact on the use of such drugs by clinicians. The primary focus is on immunomodulators that stimulate and may enhance the normal response of immunocompetent cells. Therefore, several aspects associated to immune system regulation, and regulatory pathways of immune cells are also mentioned. Furthermore, aberrant regulation is discussed in the context of immunomodulator use and the impact this has on the immune system. This review also examines the class of immunosuppressive drugs and their well-established function. Several drugs were not mentioned since the article focuses on well accepted facts or myths that new scientific evidences have changed. With that in mind, it is likely that there is a considerable amount of similarity in concepts, given that they describe immunomodulating drugs. In this context, the intention to provide important insight into how the immune system can be modulated by theses drugs surpasses this problem.
Hair loss is accompanied by keratinocyte apoptosis-regression during catagen and prolonged telogen. Angelica sinensis was reported to promote hair growth in vitro. Based on previous studies, we explored the hair growth effect and the mechanism of A. sinensis related to keratinocyte apoptosis-regression during catagen in mice. The 70% Ethanol extract of A. sinensis was applied topically at doses of 1 and 100 mg/mL to the dorsa of C57BL/6 mice for 2 weeks. The A. sinensis-treated group showed noticeable hair regrowth. Treatment with A. sinensis restored the lengths of hair shafts and size of hair follicles. In addition, mice treated with A. sinensis showed notably decreased apoptotic cells, along with a significant change in the expression of cleaved caspase-3 and the ratio of a pair of apoptosis-associated proteins: Bcl-2 and Bax. Also, A. sinensis inhibited the nuclear translocation of NF-κB, the phosphorylation of IκB-α, the phosphorylation of three mitogen-activated protein MAP kinases, and the activation of c-Jun with decreased TNF-α. These findings reveal a role of A. sinensis as an alternative treatment for hair loss that acts through hair cycle pathways associated with apoptosis regression during catagen.
The etiology of alopecia areata (AA) is still not fully understood. However, recent clinical and experimental studies have provided insights into the pathomechanisms of AA and revealed that it is an organ-specific and cell-mediated autoimmune disease. Some triggers, such as viral infections, trauma, hormones, and emotional/physical stressors, may cause activation of autoreactive T cells that target hair follicle (HF) autoantigens. In these immunological responses, cytokines and chemokines are regarded as key players that mediate the autoimmune inflammation. This results in the collapse of HF immune privilege, which is central to the pathogenesis of AA. This essay will focus on how cytokines and chemokines contribute to the immunological aspects of AA. The management of AA often remains difficult in a number of cases. Our review suggests that novel therapies for AA may involve targeting cytokines and chemokines.This article is protected by copyright. All rights reserved.
La alopecia areata es una afección frecuente en niños y en adultos, con gran repercusión psicosocial. Se trata de una alopecia no cicatrizal, que suele aparecer en placas, y en raras ocasiones afecta a todo el cuero cabelludo o incluso a todo el sistema piloso; también puede alterar las uñas; a menudo es un proceso recidivante, que evoluciona por episodios de ritmo y velocidad impredecibles. El rompecabezas fisiopatológico todavía está incompleto; las investigaciones se dirigen hacia una enfermedad autoinmunitaria con especificidad tisular y desaparición de un privilegio inmunitario, que aparece en un contexto genético predisponente. La asociación con otras enfermedades autoinmunitarias no es infrecuente. A nivel local existe un infiltrado constituido por linfocitos CD4+ y CD8+ en el bulbo piloso y alrededor del mismo. El inicio durante la infancia tiene peor pronóstico. No existe un tratamiento de eficacia uniforme, sobre todo en las formas extensas y en los niños. Tras la anamnesis y la valoración clínica se intenta instaurar un tratamiento completo con una estrategia terapéutica razonada y explicaciones simples sobre la enfermedad y su tratamiento. Las diversas vías de investigación fisiopatológica y terapéutica, así como un mayor rigor en los ensayos terapéuticos, permiten esperar progresos en los próximos años.
CD4⁺Foxp3⁺ regulatory T cells (Tregs) are suppressors of immune activation and play a crucial role in the maintenance of peripheral tolerance. Mutations of Foxp3 result in fatal autoimmunity in multiple organs, including the skin, in both humans and mice. Many studies have demonstrated the altered frequency and functions of Tregs, changes in cytokine and chemokine levels related to Tregs, and the differences in genetic background regarding Tregs in autoimmune skin disorders. Recent studies have extended our knowledge of certain properties of Tregs, especially skin‐resident Tregs. In addition, some novel therapies have been performed by modulating the number and the function of Tregs. This review focuses on the role of Tregs in some autoimmune skin disorders, including alopecia areata, vitiligo, pemphigoid and pemphigus, and systemic sclerosis, and discusses questions that remain to be addressed.
This article is protected by copyright. All rights reserved.
Background: Alopecia areata (AA) is an autoimmune non-scarring hair loss disease, in which B cell stimulation is suggested. Immunoglobulin E (IgE) is a member of Ig family, produced by B lymphocytes. The mechanism by which IgE might interact in the pathogenesis of AA is unknown. Total serum IgE was measured in previous studies with controversial results. Objective: to compare total serum levels of IgE between patients with different clinical forms of AA, and healthy subjects, to find out possible role of it in the etio-pathogenesis of this disease.
• Alopecia areata (AA) is a reversible, initially patchy hair loss most commonly involving the scalp although other regions of the head, including eyelashes and beard, may also be affected. The disease may sometimes lead to complete baldness of the scalp (alopecia areata totalis) or of the entire body (alopecia areata universalis).
• The course of AA is usually characterized by phases of acute hair loss followed by spontaneous hair regrowth and waxing and waning of the lesions. However, in severe forms hair loss can persist for many years or even life.
• Typical nail changes of AA are pitting, transversal grooves, red spotted lunulae or trachyonychia.
• Histopathological features of AA include perifollicular and intrafollicular lymphocytic infiltrates involving only anagen hair follicles with subsequent miniaturization of these structures.
• Alopecia areata is regarded as a T-cell-mediated autoimmune disease of the hair follicle that is mediated by CD4+ and CD8+ T-lymphocytes. As with other autoimmune diseases, AA most likely has a polygenic character, where susceptibility is dictated by several major genes and the phenotype may be modified by numerous minor genes.
• The most effective treatment for severe AA is the application of a contact sensitizer, while limited AA can be treated by intralesional corticosteroids or clobetasol propionate 0.05% ointment under occlusion. Pulse therapy with systemic corticosteroids or psoralen UV A (PUVA) is often used but neither has yet been proven in controlled studies to be effective in AA.
Formulation of topical treatmentTopical treatments used in the management of skin disease
Alopecia areata (AA) is a non-scarring hair loss condition that affects individuals of all ages and ethnic backgrounds. It is thought to be autoimmune in nature, but the exact cause is not yet known. This condition is usually asymptomatic and the patches are discovered incidentally in most patients. Scalp is the site most commonly affected by AA. AA patients have slightly higher chances of developing some other autoimmune conditions. There are no FDA-approved treatments for AA. Our treatment options aim to control the disease but none is curative. Despite the fact the treatment options for AA remained without major breakthroughs over the last few decades, recent understanding of the genetic structure of AA may carry newer management ideas. Addressing the psychological impact of the disease is of paramount importance in the management. This chapter will review the clinical picture and management of AA paying special attention to the pediatric age group.
Alopecia areata (AA) is a widely prevalent non-scarring hair loss disease. AA typically presents as asymptomatic well-defined round or oval patches of alopecia. Alopecia is classified based on the extent or pattern of the hair loss. Alopecia totalis (AT) is the loss of all hair on the scalp. Alopecia universalis (AU) is a 100% loss of all scalp and body hair. Alopecia ophiasis is the band like hair loss in the parieto-temporo-occipital area. Evidence suggests that AA is the organ-specific autoimmune disease that have certain genetic background. Hitopathologically, a peribulbar mononuclear cell infiltrate (“swarm of bees”) is characteristic in the acute phase, suggesting that chemokines play pivotal roles in the disease manifestation by the recruitment of these cells. In a study examining serum chemokine levels in 85 AA patients including 22 patients with “mono AA” (patients with one or two lesions of AA), 37 patients with “poly AA” (patients with three or more lesions of AA), and 26 patients with “AT/AU” (patients who experienced complete baldness in the past or in the 3-month observation period after taking blood samples), serum IP-10, MCP- 1, MIP-1α and MIP-1β levels were not significantly elevated in patients with AA compared with normal controls, while serum MIG, RANTES, IL-8 and eotaxin levels were significantly increased in patients with AA compared with normal controls. Additonally, serum MIG and RANTES levels were higher when the disease was active. Another study investigated chemokine mRNA expressions in biopsy samples from AA patients using in situ hybridization and demonstrated the strong expression of MIG, a moderate expression of MCP-1 and a weak expression of IP-10. In AA-affected mice, CCL2, CCL5 (RANTES), CXCL10, CCL17 and CCL20 expressions were upregulated in lymph node cells compared with normal mice. Also, CCL2, CCL5 and CCL17 expressions were elevated also in the AA-affected dermis in comparison to normal controls. Collectively, these findings demonstrate the importance of chemokines and their receptors in the pathogenesis of AA and suggest that they may be therapeutic targets in the disease.
Die Diagnostik und Behandlung von Haarerkrankungen ist ein wesentlicher Bestandteil dermatologischer Tätigkeit. Im folgenden werden drei in der Praxis häufige und oft schwer behandelbare Störungen der Kopfhaut und Haarfollikel abgehandelt — die vernarbenden Alopezien, die Alopecia areata und die androgenetische Alopezie.
Background: Various kinds of sensitizers are administrated for alopecia areata treatment. The aim of this study was to evaluate treatment response to Dinitrochlorobenzene (DNCB) in alopecia areata patients. Method: In this study, 117 patients were treated with DNCB under a specific checklist. All patients were sensitized with a 2% DNCB and then treated with ascending DNCB concentrations (0.001%-2%). Response to treatment was categorized as none, mild, moderate and marked improvement. Result: Thirty three (27.5%) patients showed no response, 49 (40.8%) had relapse 6 months after improvement, 29 (24.2%) had no relapse 6 months after the treatment and 6 patients were excluded because they did not return for follow-up visits. Response to treatment in patients without eyelash and eyebrow involvement increased significantly (P=0.01). We did not observe any side effects except for localized dermatitis seen in 5% of the patients. Conclusion: With respect to the suitable response to DNCB application and its availability, the authors suggest that DNCB be reconsidered in alopecia areata. However, attention must be paid to its mutagenicity.
Dermatologists have the good fortune to work on the most accessible organ of the body. Many inflammatory and neoplastic conditions can be effectively managed using the wide range of locally applied physical or pharmacological modalities that are available. The latter are the subject of this chapter, which reviews the pharmacological treatments used topically, i.e. by application to the surface of the skin. Some of these are time-honoured treatments that have been used for a century or more, whilst others belong to the ever-expanding range of newer and increasingly potent agents constantly being developed and formulated for topical use. Topical treatment offers the potential to achieve high concentrations of a drug in the skin with minimal exposure of other organs. This can greatly increase efficacy and also safety relative to systemic administration. When side effects do occur, they are most likely to take the form of localized reactions.
Topische Immuntherapie mit obligaten Kontaktallergenen bei Alopecia areata (AA): Die derzeit nach wie vor wirksamste Methode zur Therapie der ausgedehnteren Alopecia areata wird ausführlich, auch anhand aktueller Literatur vorgestellt und die praktische Durchfühung erläutert.
Although the loss of scalp hair is distressing and many medical treatments focus on its restoration, the removal of body hair has been adopted since ancient times. Beauty standards, which r eflect the culture of each society, have been presenting the depilated body as absolutely desirable. Through the ages various methods of hair removal have been used depending on the requirements of the individuals. In recent years, Laser and Intense Pulse Light devices have been considered as the most promising solution for excess hair growth, without excluding the efficacy of other methods to induce satisfactory epilatory results. The enzyme-based hair removal method has received little recognition even though experimental and clinical data support its efficacy to provide long term or even permanent epilation. The present review presents these data and examines the likelihood of considering the aforementioned method as ideal.
Diphencyprone (DPCP) is a hapten that causes delayed-type hypersensitivity (DTH) reactions in human skin, and is used as a topical therapeutic for alopecia areata, warts, and cutaneous melanoma metastases. We examined peak DTH reactions induced by DPCP (3 days post-challenge) by comprehensive gene expression and histological analysis. To better understand how these DTH reactions naturally resolve, we compared our DPCP biopsies to those from patients with psoriasis vulgaris, a chronic inflammatory disease that does not resolve. By both microarray and qRT-PCR, we found that psoriasis lesional skin has significantly lower expression of many negative immune regulators compared to peak DPCP reactions. These regulators include: interleukin-10, cytotoxic T lymphocyte-associated 4 (CTLA4), programmed cell death 1 (PD1), programmed cell death 1 ligand 1 (PDL1), programmed cell death 1 ligand 2 (PDL2), and indoleamine 2,3-dioxygenase (IDO1). Their decreased expression was confirmed at the protein level by immunohistochemistry. To more completely determine the balance of positive vs. negative immune regulators in both DPCP reactions and psoriasis, we developed one comprehensive gene list for positive regulatory (inflammatory) genes, and another for negative regulatory (immunosuppressive) genes, through Gene Ontology terms and literature review. With this approach, we found that DPCP reactions have a higher ratio of negative to positive regulatory genes (both in terms of quantity and expression levels) than psoriasis lesional skin. These data suggest that the disease chronicity that distinguishes psoriasis from transient DTH reactions may be related to absence of negative immune regulatory pathways, and induction of these is therefore of therapeutic interest. Further study of these negative regulatory mechanisms that are present in DPCP reactions, but not in psoriasis, could reveal novel players in the pathogenesis of chronic inflammation. The DPCP system used here thus provides a tractable model for primary discovery of pathways potentially involved in immune regulation in peripheral tissues.
Squaric acid dibutylester (SADBE) is frequently used for the treatment of alopecia; however, unwanted side effects sometimes occur. We herein report two cases of severe pustular lesions induced by topical SADBE application.
Background:
Alopecia areata (AA) is an organ-specific autoimmune disease with T-cell-mediated attack of hair follicle autoantigens. As T-helper 17 (Th17) cells and T regulatory (Treg) cells are crucially involved in the pathogenesis, the role of Th17 and Treg cytokines has not been studied yet.
Objective:
To determine whether AA is associated with alterations in lesional and serum Th17 and Treg cytokines, and studied whether they were associated with clinical type.
Methods:
Scalp skin samples from 45 patients and 8 normal controls were obtained for PCR specific for IFN-γ, TNF-α, TGF-β, IL-1, IL-2, IL-4, IL-10, IL-12A, IL-13, IL-17, IL-22 and IL-23. Serum cytokines were measured from 55 patients and 15 normal controls using ELISA.
Results:
Lesional IL-17 and IL-22 were significantly increased in patients group. Moreover, positive correlations were shown between lesional IL-17, IL-22 and disease severity. Serum IL-1, IL-17, TNF-α, and TGF-β were significantly increased, and positive correlation was shown between serum IL-17 and disease severity.
Conclusion:
These results showed significantly high Th17 cytokines in both lesion and serum in AA patients, which may highlight a functional role of these cytokines in the pathogenesis of AA. This article is protected by copyright. All rights reserved.
Keratinocytes play a pivotal role in the regulation of immune responses but the impact of antigen-presentation by these cells is still poorly understood, particularly in a situation where the antigen will be presented only in adult life. Here, we generated a transgenic mouse model in which keratinocytes exclusively present a myelin basic protein (MBP) peptide covalently linked to the MHC class II β-chain, solely under inflammatory conditions. In these mice, inflammation caused by epicutaneous contact sensitizer treatment resulted in keratinocyte-mediated expansion of MBP-specific CD4(+) T cells in the skin. Moreover, repeated contact sensitizer application preceding a systemic MBP immunization reduced the reactivity of the respective CD4(+) T cells and lowered the symptoms of the resulting experimental autoimmune encephalomyelitis. This down-regulation was CD4(+) T cell mediated and dependent on the presence of the immune modulator Dickkopf-3. Thus, presentation of a neo self-antigen by keratinocytes in the inflamed, adult skin can modulate CD4(+) T cell auto-aggression at a distal organ.In parallel to the DPCP skin treatments, 0.5 mg of the anti-Dkk3 4.22 mAb (Papatriantafyllou et al., 2012) in 200 μl PBS was applied i.p., 3 times, each one week apart.Journal of Investigative Dermatology accepted article preview online, 02 April 2015. doi:10.1038/jid.2015.130.
Some patients with chronic extensive alopecia areata (AA) may be refractory to topical immunotherapy. Combination therapy is recommended for such patients. Efficacy and safety of a combination therapy with diphenylcyclopropenone (DPCP) and anthralin in chronic extensive AA is unknown.
We sought to determine whether the combination therapy of DPCP and anthralin is superior to DPCP alone in chronic extensive AA.
We retrospectively analyzed the efficacy, side effects, and relapse rates of DPCP (alone or with anthralin) in chronic extensive AA.
A total of 47 patients (22 were treated only with DPCP, and 25 with DPCP and anthralin for at least 30 weeks) were evaluated. Complete hair regrowth was observed in 36.4% and 72% of the patients who received DPCP and combination therapy, respectively (P = .01). Hair regrowth duration was shorter with combination therapy (P = .01). Regrowth rates of the eyebrows, eyelashes, and beard in patients on combination therapy were higher than those in patients on DPCP (P = .01). Side effects such as folliculitis, hyperpigmentation, and staining of skin, hair, and clothes were more common in combination therapy group.
The retrospective design and small number of patients are limitations.
Combination therapy with DPCP and anthralin is superior to DPCP alone in chronic extensive AA.
Copyright © 2015 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.
Alopecia areata (AA) is a chronic inflammatory disease mediated by an array of cells and cytokines. Immunohistochemistry (IHC) of histological sections with antibodies to mast cell tryptase, CD4, CD8, CD1a and semi-quantitative real-time PCR analysis of Th1- and Th2-type cytokines were performed in 55 patients to investigate the infiltration features of mast cells (MCs), T lymphocytes and Langerhans cells (LCs) in scalp lesions of patients with AA. In AA patients, increased MCs mainly infiltrated the peri-follicular and peri-vascular areas, and correlated positively with numbers of CD8(+) T lymphocytes in deep peri-follicular areas (P = 0.04), but negatively with CD4(+) T lymphocytes in deep peri-vascular areas (P = 0.031). In patients with active hair loss, LCs in epidermis, deep dermis and peri-vascular were elevated (Ps < 0.05). Infiltration of LCs in upper peri-vascular areas and CD8(+) T cell infiltration in deep peri-follicular areas were positively correlated (R = 0.618, P = 0.011), as well as LCs in deep peri-vascular areas with CD8(+) T cells in upper peri-follicular areas (R = 0.570, P = 0.017). In patients with active hair loss, Th1-type cytokine (IL-2, IL-8, TNF-α) mRNA expression in deep dermis were higher than in upper dermis (Ps < 0.05). However, in patients with non-active hair loss, Th2-type cytokine (IL-5, IL-10) mRNA expression in deep dermis was higher than that in the upper dermis (Ps < 0.05). Positive correlations were found existing between MCs and CD8(+) T cells, as well as between LCs and CD8(+) T cells. In conclusion, findings in this study allow us to propose a close relationship between mast cells and CD8(+) T cells, as well as between LCs and CD8(+) T cells in AA, as well as allergy may interfere with infiltrating T lymphocytes in AA lesional regions. Also, Th1-type cytokine are related to disease activity of alopecia areata, whereas Th2-type cytokines may be associated with persistence of AA.
Contact sensitizers are defined as reactive molecules (electrophilic) which have the ability to modify skin proteins to form an antigen (hapten). In addition to the haptenation mechanism, danger signals, leading to the activation of dendritic cells, are described to be crucial for the effective induction of an hapten-specific T cell immune response. In the context of the 7th amendment to the Cosmetic Directive, the cosmetic industry is concerned by the challenge of finding non-animal approaches to assess the sensitizing potential of chemicals. While danger signals induced by sensitizers in steady-state conditions have already been analyzed, we chose to investigate the impact of sensitizers on the course of an inflammatory response. For this purpose we used the U937 cell line differentiated with PMA and activated with LPS. In these conditions, cells produce a large amount of inflammatory mediators (IL-β, TNF-α, IL-6, IL-10, IL-8, PGE2, PGD2, TxB2) through the activation of pathways leading to the activation of the transcription factors NF-κB and Nrf2 and through AA metabolism by the cPLA2/COX-2 cascade. Interestingly, we showed that 6 contact sensitizers with various potential (DNCB, PPD, HQ, PG, CIN, EUG) significally and specifically decrease the production of prostanoïds and in particular of PGE2 induced by PMA/LPS. We further demonstrated that there is no unique inhibition profile of the sensitizers even if the majority (except for DNCB) of the effects applies on COX-2 (i.e. inhibition of the expression and/or activity). For DNCB, inhibition mechanism appears to be dependant of its capacity to react with thiols residues and in particular to deplete intracellular glutathione possibly leading to the inactivation of the PG-synthases. In parallel, we assess a statistical analysis on 160 molecules that allow us to define the test parameters (a molecule is a sensitizer if the PGE2 inhibition at 24h is more than 60%) and to calculate the test performance toward LLNA (78%). Moreover we demonstrated that the PGE2 test could be complementary to other already existing in vitro tests like MUSST or Nrf2-HTS. In summary, we add here a new insight into the multiple biochemical effects described so far for sensitizers. Even if the underlying biological relevance remains unclear, the parameter “PGE2 inhibition” is good test for skin sensitization evaluation. Further studies will precise how this parameter could be implemented into an alternative testing strategy for the evaluation of skin sensitization.
The patchy form of alopecia areata has a high rate of spontaneous remission. In severe forms, the chance of a spontaneous regrowth is much lower but still present. This fact must be taken into account when different therapeutic modalities are evaluated. For topical therapeutics, efficacy must be established by treating only parts of the scalp, the nontreated areas serving as a control. Systemic drugs must be evaluated in placebo-controlled studies. Although alopecia areata is a disease that may cause major psychosocial problems, it is at any rate not life threatening. For this reason, the obtained results must be weighed against possible undesired effects. Maintenance treatment should be suitable for a long period of time, and possible long-term side effects must be taken into consideration.
The term “alopecia” {άγωπεκíα = fox’s disease) was first used by Hippocrates in the 35th chapter of his book On Diseases. He wrote that the so-called blemishes include “the foxes”, by which he meant that the hair “gets foxy” at particular spots (“areae”). The term was probably coined because “no grass grows where the fox urinates” (Ebstein), but this interpretation was rejected by Richter (1928), who claimed that the Indian source allegedly quoted by Ebstein did not exist. The term “area” is known from Celsus, a contemporary of Virgil. In the 4th chapter of his 6th book, Celsus wrote “arearum quorum duo genera sunt”, i.e., there are two types of baldness: “… the form which leads to slight fattiness and complete baldness of the skin is worse. The form termed “alopekia” may take any shape. It originates in areas covered with scalp and beard hair. But the type which is called “ophiasis” (ἤ#x03D5;ιξ = snake), because of its similarity to the windings of a snake develops from the back of the head, more than two fingers in width, and its two ends creep on towards the ears and the forehead, where they meet. The former may occur at any age, while the latter is usually seen in children only. The former hardly ever heals without treatment, the latter frequently heals spontaneously…” (Celsus). According to Montgomery (1931), the term “area” was used by Virgil to mean a circular place similar to that where the grain was threshed. On the other hand, the chaff remaining on the threshing-floor makes one think of “favus” rather than the smooth, candlewax-white spot characteristic of alopecia areata. It is assumed that Celsus himself was only the translator of a medical work written by the Greek Menekrates, a personal physician of Tiberius.
In the present study we demonstrate that human monocytes activated by lipopolysaccharides (LPS) were able to produce high levels of interleukin 10 (IL-10), previously designated cytokine synthesis inhibitory factor (CSIF), in a dose dependent fashion. IL-10 was detectable 7 h after activation of the monocytes and maximal levels of IL-10 production were observed after 24-48 h. These kinetics indicated that the production of IL-10 by human monocytes was relatively late as compared to the production of IL-1α, IL-1β, IL-6, IL-8, tumor necrosis factor Oi(TNFα), and granulocyte colony-stimulating factor (G-CSF), which were all secreted at high levels 4-8 h after activation. The production of IL-10 by LPS activated monocytes was, similar to that of IL-1α, IL-1β, IL-6, IL-8, TNFα, granulocyte-macrophage colony-stimulating factor (GM-CSF), and G-CSF, inhibited by IL-4. Furthermore we demonstrate here that IL-10, added to monocytes, activated by interferon γ(IFN-γ), LPS, or combinations of LPS and IFN-γ at the onset of the cultures, strongly inhibited the production of IL-1α, IL-1β, IL-6, IL-8, TNFα, GM-CSF, and G-CSF at the transcriptional level. Viral-IL-10, which has similar biological activities on human cells, also inhibited the production of TNFα and GM-CSF by monocytes following LPS activation. Activation of monocytes by LPS in the presence of neutralizing anti-IL-10 monoclonal antibodies resulted in the production of higher amounts of cytokines relative to LPS treatment alone, indicating that endogenously produced IL-10 inhibited the production of IL-1α, IL-1β, IL-6, IL-8, TNFα, GM-CSF, and G-CSF. In addition, IL-10 had autoregulatory effects since it strongly inhibited IL-10 mRNA synthesis in LPS activated monocytes. Furthermore, endogenously produced IL-10 was found to be responsible for the reduction in class II major histocompatibility complex (MHC) expression following activation of monocytes with LPS. Taken together our results indicate that IL-10 has important regulatory effects on immunological and inflammatory responses because of its capacity to downregulate class II MHC expression and to inhibit the production of proinflammatory cytokines by monocytes.
The Journal of Investigative Dermatology publishes basic and clinical research in cutaneous biology and skin disease.
43 patients with alopecia areata were treated with weekly applications of dinitrochlorobenzene (D.N.C.B.), dissolved in acetone, on one side of the head, with the other side serving as control region. The therapeutic aim was a mild contact dermatitis. A significant difference of hair growth between the treated and untreated sides was observed in 33 patients. 21 patients showed regrowth of hair exclusively on the treated side, and in 12 patients regrowth was considerably faster and more dense on the treated side. In the majority of patients the difference was noted within 3 months.
Within a group of 139 patients previously studied during treatment for alopecia areata with diphenylcyclopropenone (DCP), hair growth was re-evaluated after a period of 19 months following completion of our previous study. Fifty-four patients treated with DCP had total and 6 had partial but cosmetically acceptable regrowth. Twenty-five patients with total regrowth had stopped DCP treatment for a mean period of 15 months and had not relapsed. Nineteen of 28 patients who still applied DCP were in the process of stepwise discontinuation of treatment. Fifteen patients had subsequently been treated with squaric acid dibutylester (SADBE) after having acquired 'tolerance' to DCP; at the time of re-evaluation 3 of these patients had complete regrowth, and 4 patients had partial but cosmetically acceptable regrowth. Topical immunotherapy with DCP and SADBE had resulted in total regrowth in 57/139 patients (41.0%) and in partial but cosmetically satisfactory regrowth in 10/139 patients (7.2%). The type of involvement and duration of alopecia areata were factors of prognostic significance.
It has previously been demonstrated that the epidermis is a rich source of proinflammatory cytokines and growth factors and that complex interactions between these factors may affect inflammatory responses in skin. To investigate whether IL-10 (cytokine synthesis inhibitory factor) is part of this complex process, RNA was extracted from normal epidermis at various times after application of various chemicals to murine skin and mRNA signals for IL-10 were sought using a quantitative reverse-transcriptase-polymerase chain reaction technique. IL-10 signal strength was normalized to that of beta-actin in each sample. IL-10 mRNA signals were occasionally identified in normal epidermis but were uniformly enhanced 4 h after hapten application, and were maximal after 12 h. Contact allergens induced IL-10 mRNA signals whereas vehicles and irritants did not. Depletion of Langerhans cells, Thy-1+ dendritic epidermal cells, and T lymphocytes demonstrated that keratinocytes were the main source of IL-10 mRNA. IL-10 signals were also detected in mRNA derived from PAM 212 (spontaneously transformed keratinocyte) cells. IL-10 protein could be detected by immunoprecipitation, with an IL-10 mAB, of supernatants obtained 16 h after cultured epidermal cells were coupled with hapten. This study demonstrates that murine keratinocytes are capable of producing IL-10 mRNA and protein, and that signal strength of IL-10 mRNA is enhanced by hapten application.
To understand the molecular events which are important in leucocyte trafficking in cutaneous inflammation, poison ivy/oak extract was applied topically to the skin, and the simultaneous assessment of a variety of clinical and immunopathological parameters performed. The clinical response of subjects was divided into three main groups: I, 2-24h after application, before the onset of erythema; II, 48 h-1 week after application during maximal clinical changes; III, 2-3 weeks after application when the inflammation had subsided. Six different biopsies per subject were evaluated over the study period and the density of dermal cellular infiltrate, and the distribution of intercellular adhesion molecule-1, (ICAM-1), endothelial leucocyte adhesion molecule-1, (ELAM-1), vascular cell adhesion molecule-1, (VCAM-1), interleukin 8 (IL-8) and tumour necrosis factor-alpha (TNF-alpha), determined. Eight hours after exposure, before lymphocytes and monocytes had entered the dermal interstitium or epidermis, the keratinocytes expressed TNF-alpha and ICAM-1, whilst the endothelial cells expressed ELAM-1, VCAM-1 and ICAM-1. Group II biopsies revealed increasing keratinocyte expression of TNF-alpha and ICAM-1 with the appearance of IL-8, which correlated with the onset of epidermal T-cell trafficking. The endothelium was strongly positive for ELAM-1 and VCAM-1, but there was no influx of neutrophils. Group III biopsies showed a decrease in the expression of ICAM-1, VCAM-1 and ELAM-1 by both keratinocytes and endothelium with a reduction in epidermal/dermal inflammation, although the endothelial cell staining of VCAM-1 and ELAM-1 did not completely disappear. These results suggest that on exposure to poison ivy/oak, keratinocytes rapidly produce TNF-alpha which leads to an early autoinduction of ICAM-1, and later IL-8. There is also a paracrinemediated induction and augmentation of underlying endothelial cell ELAM-1, VCAM-1 and ICAM-1.
Murine IL-10 (cytokine synthesis inhibitory factor) inhibits cytokine production by Th1 cell clones when they are activated under conditions requiring the presence of APC. By preincubating APC with IL-10, we demonstrate that IL-10 acts principally on APC to inhibit IFN-gamma production by Th1 clones. Moreover, IL-10 is not active when Th1 cells are stimulated with glutaraldehyde-fixed APC, which also indicates that its action involves regulation of APC function. Furthermore, IL-10 inhibits cytokine synthesis by Th1 cells stimulated with the super-antigen Staphylococcus enterotoxin B, which does not appear to require processing. Flow microfluorimetry purified splenic or peritoneal B cells and macrophages, and B cell and macrophage cell lines can present Ag to Th1 clones. However, IL-10 acts only on sorted macrophages and the macrophage cell line to suppress IFN-gamma production by Th1 clones. IL-10 does not show this effect when B cells are used as APC. In contrast, IL-10 does not impair the ability of APC to stimulate cytokine production by Th2 cells. IL-10 does not decrease IFN-gamma-induced I-Ad levels on a macrophage cell line. Inasmuch as IL-10 also inhibits IL-2-induced IFN-gamma production by Th1 cells in an Ag-free system requiring only the presence of accessory cells, these data suggest that IL-10 may inhibit macrophage accessory cell function which is independent of TCR-class II MHC interactions.
We examined the role of cytokines in the cutaneous response to the application of trinitrochlorobenzene (TNCB) in both nonsensitized and sensitized mice, i.e., in the irritant reaction (IR) and contact hypersensitivity reactions (CH). When administered immediately before challenge, anti-tumor necrosis factor (TNF) antibody abrogated the ear swelling response in CH; antibody directed against interferon gamma or antibodies to both granulocyte/macrophage colony-stimulating factor and interleukin 3 (IL-3) had a partial inhibitory effect; anti-IL-2 receptor antibody had no effect. Anti-TNF prevented the various features of the CH, as seen on histological sections, e.g., leukocyte infiltration and hemorrhages within the dermis and keratinocytes necrosis. Anti-TNF antibody also prevented the IR. The presence of TNF mRNA was evaluated on Northern blots; TNF-alpha mRNA was detectable in an untreated ear, increased after the application of TNCB in nonsensitized mice, and was highest in sensitized mice. TNF mRNA accumulation, which was evident 0.5 h after hapten application and lasted greater than 72 h, was abolished by treatment with anti-TNF antibody, thus suggesting an auto-amplification of TNF production. The cellular origin of TNF mRNA was explored by in situ hybridization; basal keratinocytes showed the highest labeling, but TNF mRNA was also detectable in cells of the dermal infiltrate. After hapten (TNCB) application at sites susceptible (the ear) or resistant (the foot pad) to CH or IR, a close correlation was observed between TNF mRNA accumulation and the intensity of the inflammatory reaction. The major role played by TNF in both the CH and the IR explains the histologically similar aspects of these reactions and the extreme variability of these reactions at various anatomical sites.
One hundred thirty-nine patients with severe alopecia areata (the majority with the subtotal, total, or universalis type) were treated with topical immunotherapy (diphenylcyclopropenone). Patients were initially treated unilaterally; the other side of the scalp served as a control. In 50.4% of the patients the response was either excellent (total regrowth) or satisfactory (subtotal regrowth with only a few remaining bald patches). The most frequent side effects were eczematous reactions with blistering, spreading of the induced contact eczema, and sleep disturbances.
The Journal of Investigative Dermatology publishes basic and clinical research in cutaneous biology and skin disease.
Hybridomas were produced from a rat that was immunized with partially purified proteins from supernatants of induced Th2 cells. These preparations were enriched for cytokine synthesis inhibitory factor (CSIF, IL-10). The mAb in the supernatants were screened by a solid phase radioimmunoadsorbent assay using 35S-methionine-labeled secreted proteins from a lectin-stimulated Th2 clone. A total of 18 anticytokine mAb were isolated, comprising 6 anti-CSIF, 1 anti-IL-4, 1 anti-IL-5, and 10 anti-IL-6 mAb. The anti-CSIF mAb were separable into three groups. mAb in groups A and B neutralized and depleted bioactivity, and bound to overlapping but nonidentical subpopulations of CSIF molecules. The 2 mAb in group C did not neutralize CSIF activity, and bound to CSIF molecules not recognized by mAb from groups A or B. A two-site sandwich ELISA for CSIF could be established with the group A antibody, SXC1, combined with any of the three group B antibodies. The sensitivity of this assay was equivalent to that of the CSIF bioassay. These antibodies have been used to show that CSIF is responsible for most or all of the ability of Th2 supernatants to inhibit cytokine synthesis by Th1 cells. In addition, the ELISA has been used to confirm that CSIF is produced by Th2 but not Th1 clones.
A panel of antigen-specific mouse helper T cell clones was characterized according to patterns of lymphokine activity production, and two types of T cell were distinguished. Type 1 T helper cells (TH1) produced IL 2, interferon-gamma, GM-CSF, and IL 3 in response to antigen + presenting cells or to Con A, whereas type 2 helper T cells (TH2) produced IL 3, BSF1, and two other activities unique to the TH2 subset, a mast cell growth factor distinct from IL 3 and a T cell growth factor distinct from IL 2. Clones representing each type of T cell were characterized, and the pattern of lymphokine activities was consistent within each set. The secreted proteins induced by Con A were analyzed by biosynthetic labeling and SDS gel electrophoresis, and significant differences were seen between the two groups of T cell line. Both types of T cell grew in response to alternating cycles of antigen stimulation, followed by growth in IL 2-containing medium. Examples of both types of T cell were also specific for or restricted by the I region of the MHC, and the surface marker phenotype of the majority of both types was Ly-1+, Lyt-2-, L3T4+, Both types of helper T cell could provide help for B cells, but the nature of the help differed. TH1 cells were found among examples of T cell clones specific for chicken RBC and mouse alloantigens. TH2 cells were found among clones specific for mouse alloantigens, fowl gamma-globulin, and KLH. The relationship between these two types of T cells and previously described subsets of T helper cells is discussed.
There is evidence suggesting that alopecia areata (AA) may have an autoimmune pathogenesis, and it was recently reported that keratinocytes in the bulb of some hair follicles affected by this condition express class II HLA (HLA-DR) antigens, which are not present on the same cells in normal tissue. Since it has been proposed that an analogous ectopic HLA-DR expression by epithelial cells in other organs might be an early event leading to organ-specific autoimmunity, we have investigated the sequence in which perifollicular mononuclear cell (MNC) infiltration and ectopic HLA-DR expression on keratinocytes appear in recent-onset and long-standing cases of AA by immunostainings of affected and unaffected areas with monoclonal antibodies against leukocyte and HLA-DR antigens. In recent-onset AA lesions, ectopic HLA-DR expression on hair follicle keratinocytes was found only occasionally (in 3 out of 247 follicles examined) and was restricted to biopsies from the affected areas. This prevalence was significantly lower than the prevalence of hair follicles showing perifollicular MNC infiltrates in the same biopsies, and was also significantly lower than the prevalence of hair follicles showing ectopic HLA-DR expression on keratinocytes in the affected areas of longstanding cases. These findings suggest that in AA lesions the perifollicular MNC infiltration precedes the ectopic HLA-DR expression on hair follicle keratinocytes, and therefore argue against the notion of a primary role for that ectopic HLA-DR expression on epithelial cells in triggering the putative autoimmune response in AA.
One type of Th cell clone, Th1, causes delayed-type hypersensitivity (DTH) when injected with the appropriate Ag into the footpads of naive mice. Because IFN-gamma, one of the Th1-specific cytokines, has been reported to produce some of the manifestations of DTH, we have investigated the role of IFN-gamma in the DTH reaction induced by Th1 cells by using a mAb which neutralizes the biologic activity of IFN-gamma. Anti IFN-gamma inhibited up to 55% of the swelling response induced by seven Th1 clones in BALB/c and CBA/J mice suggesting that IFN-gamma is an important mediator of Th1-induced DTH. This inhibition was consistently observed during the peak phase of the DTH response and could be partly but not entirely explained by decreases in vascular leakage. The DTH responses induced by two Th1 clones in C57BL/6 mice were not inhibited by anti IFN-gamma. Taken together, these data suggest that other inflammatory mediators also play a role in Th1-induced DTH responses. Based on the gross description and kinetics of the response and the numbers of polymorphonuclear neutrophilic granulocytes in the cellular infiltrate, Th1-induced DTH appears to be similar to the Jones-Mote type of hypersensitivity.
A cytokine synthesis inhibitory factor (CSIF) is secreted by Th2 clones in response to Con A or antigen stimulation, but is absent in supernatants from Con A-induced Th1 clones. CSIF can inhibit the production of IL-2, IL-3, lymphotoxin (LT)/TNF, IFN-gamma, and granulocyte-macrophage CSF (GM-CSF) by Th1 cells responding to antigen and APC, but Th2 cytokine synthesis is not significantly affected. Transforming growth factor beta (TGF-beta) also inhibits IFN-gamma production, although less effectively than CSIF, whereas IL-2 and IL-4 partially antagonize the activity of CSIF. CSIF inhibition of cytokine synthesis is not complete, since early cytokine synthesis (before 8 h) is not significantly affected, whereas later synthesis is strongly inhibited. In the presence of CSIF, IFN-gamma mRNA levels are reduced slightly at 8, and strongly at 12 h after stimulation. Inhibition of cytokine expression by CSIF is not due to a general reduction in Th1 cell viability, since actin mRNA levels were not reduced, and proliferation of antigen-stimulated cells in response to IL-2, was unaffected. Biochemical characterization, mAbs, and recombinant or purified cytokines showed that CSIF is distinct from IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IFN-gamma, GM-CSF, TGF-beta, TNF, LT, and P40. The potential role of CSIF in crossregulation of Th1 and Th2 responses is discussed.
It has previously been shown that, in patients with untreated progressive alopecia areata (AA), the peribulbar T4/T8 ratio is about 4:1. In the present study, the immunohistochemical findings obtained in untreated AA patients were compared to those obtained in patients who had received topical immunotherapy with diphencyprone. The untreated group consisted of 5 patients with progressive AA and 5 patients with inactive AA. The treated group consisted of 5 patients with a good response to diphencyprone and 5 patients with little or no hair regrowth after treatment. In untreated patients with progressive AA, the mean peribulbar T4/T8 ratio was 4:1, whereas in untreated patients with stable AA, the ratio was 2:1. In the treated patients with a good response to diphencyprone, the mean T4/T8 ratio was 1:1, while in the patients with poor or no response to treatment, the ratio was 0.7. In conclusion, topical immunotherapy considerably alters the peribulbar T4/T8 ratio in AA. The results are consistent with, but do not prove, the concept of topical immunomodulation.
We have previously shown that at least two types of Lyt-1+, Lyt-2-, L3T4+ helper T cell clones can be distinguished in vitro by different patterns of lymphokine secretion and by different forms of B cell help. Evidence is presented here to show that one type of helper T cell clone (TH1) causes delayed-type hypersensitivity (DTH) when injected with the appropriate antigen into the footpads of naive mice. The antigen-specific, major histocompatability complex (MHC)-restricted footpad swelling reaction peaked at approximately 24 hr. Footpad swelling was induced by all TH1 clones tested so far, including clones specific for soluble, particulate, or allogeneic antigens. In contrast, local transfer of TH2 cells and antigen did not produce a DTH reaction, even when supplemented with syngeneic spleen accessory cells. Similarly, local transfer of an alloreactive cytotoxic T lymphocyte clone into appropriate recipients did not produce DTH. The requirements for the DTH reaction induced by TH1 cells were investigated further by using TH1 clones with dual specificity for both foreign antigens and M1s antigens. Although these clones responded in vitro to either antigen + syngeneic presenting cells, or M1s disparate spleen cells, they responded in vivo only to antigen + MHC and did not cause footpad swelling in an M1s-disparate mouse in the absence of antigen. Moreover, in vitro preactivation of TH1 or TH2 cells with the lectin concanavalin A was insufficient to induce DTH reactions upon subsequent injection into footpads. From these results, we conclude that the lack of DTH given by TH2 clones in vivo could be due to the inability of the TH2 cells to produce the correct mediators of DTH, or to a lack of stimulation of TH2 clones in the footpad environment.
A biphasic dose-response curve was observed when the IL-1-dependent HTL clone D10 was exposed to IL-1 plus supernatants from some activated T cell clones but not others. The active component that inhibited proliferation at high concentrations of these supernatants appeared to be IFN-gamma based on the following findings: 1) the biphasic pattern of responsiveness correlated with the presence of IFN-gamma in the supernatants; 2) an anti-IFN-gamma mAb augmented the proliferation of D10 cells to these supernatants; 3) rIFN-gamma inhibited profoundly the response of D10 cells stimulated with rIL-1 plus supernatant from activated D10 cells or with rIL-1 plus rIL-4; 4) the response of D10 cells to rIL-1 plus rIL-2 also was inhibited by rIFN-gamma, although to a lesser extent. The proliferation of an additional Th2 clone stimulated with rIL-1 plus rIL-4 or rIL-2 also was inhibited by rIFN-gamma, implicating IFN-gamma as an inhibitory lymphokine for Th2 cells in general. rIFN-gamma did not affect the proliferation of two Th1 clones, nor did it affect the proliferation of an unconventional HTL clone which produces both IL-4 and IFN-gamma and proliferates in response to IL-2 or IL-4 in an IL-1-independent fashion. The proliferation of D10 cells stimulated by Ag or by immobilized anti-CD3 antibody also was blocked by rIFN-gamma, whereas IL-4 production in response to these stimuli was unaffected, indicating that proliferation and not general cell function was specifically inhibited. Collectively, these data implicate IFN-gamma as a suppressive factor for the proliferation of the subset of HTL designated Th2, and suggest that the relative amounts of the various lymphokines present during an immune response may direct which T cell types increase in number.
In situ lymphocyte subsets in 12 patients with patchy alopecia areata or alopecia universalis were estimated using monoclonal antibodies and immunoperoxidase technique. The majority of the inflammatory cells around the hair bulbs and follicles where leu 4+ T-cells with the subphenotypes of either leu 2a+ (suppressor/cytotoxic T-cells) or leu 3a+ (helper/inducer T-cells). Many of the T-cells expressed HLA-DR class II antigen. In about half of the patients, the hair bulbs and follicles were also infiltrated with T-cells.
Eight of the 12 patients were treated with dinitroch-loro-benzene. Six of these patients were biopsied during treatment. The number of T-cells around the hair bulbs and follicles increased during; treatment and niam T-cells were seen in the hair bulbs and follicles. In five of the six patients treated with dinitrochlorobenzene, an increase in the percentage of leu 2a+ cells around the bulbs and follicles was noticed. The increase in leu 2a+ cells may be of importance for the DNCB-induced re-growth of hair.
Fifty-eight scalp biopsies were immunohistologically investigated with monoclonal antibodies against HLA-ABC, HLA-DR, and T6 antigens. The following 3 groups were compared: control biopsies obtained from healthy volunteers (n = 5) or patients with unrelated scalp diseases (n = 6); biopsies from untreated alopecia areata (AA), obtained either from untreated patients (n = 19) or from the untreated side in patients receiving unilateral treatment with the contact allergen diphencyprone (DCP) (n = 13); biopsies obtained from the treated side in patients receiving unilateral treatment with DCP (n = 13). While HLA-ABC antigens were strongly expressed by epidermal keratinocytes and the infundibular epithelium of hair follicles in all biopsies, these antigens were either not detectable or only faintly expressed on the subinfundibular epithelium and the hair matrix in the control series. By contrast, 30 out of 32 biopsies from untreated AA showed expression of HLA-ABC antigens on hair matrix epithelium, and the subinfundibular epithelium was HLA-ABC-positive in 15 out of 32 cases. In the biopsies from treated AA, HLA-ABC antigens were expressed on hair matrix epithelium in 9 out of 13 cases, and on the subinfundibular epithelium in 1 case. In the controls and untreated AA, HLA-DR expression was confined to dendritic cells in the epidermis and the follicular infundibulum. Its expression on hair matrix epithelium was found in 15 out of 32 biopsies from untreated AA and in 4 out of 13 biopsies from treated AA. In the control series, intrabulbar T6+ dendritic cells were either absent or present in low numbers. High numbers of intrabulbar T6+ cells were present in 7 out of 32 biopsies from untreated AA and in 0 out of 13 biopsies from treated AA. The data show that abnormal expression of class I major histocompatibility (MHC) antigens on hair matrix epithelium is a constant feature in AA, whereas class II MHC antigens are less frequently expressed. Topical immunotherapy with DCP, which induced expression of HLA-DR in epidermal keratinocytes in 6 out of 13 cases, reduced the abnormal expression of both HLA-ABC and -DR antigens in the epithelium of lower hair follicles in AA.
A histopathological study was performed in 17 patients with alopecia areata to elucidate the changes in hair cycle dynamics. The findings confirm the view that the initial event in alopecia areata is the premature entry of anagen follicles into telogen, although some follicles survive for a time in a dystrophic anagen state. However, after re-entry into anagen takes place, growth appears to be halted in anagen III rather than anagen IV, as has previously been suggested. Follicles then return prematurely to telogen and these truncated cycles are repeated until the disease activity subsides. A new pathogenic hypothesis is presented which relates alterations in hair cycle dynamics to pathological changes within the anagen follicle and also provides an explanation for the formation of exclamation mark hairs and the non-destructive nature of the disease.
In 11 patients with untreated alopecia areata in the progressive stage of the disease, an in situ analysis of the inflammatory infiltrate of the hair bulbs was performed by means of different monoclonal antibodies. Most of the peribulbar cells reacted with the pan T-cell antibodies OKT 3 and Lyt 3. Staining for T-cell subsets revealed that the proportion of OKT 4+ cells was about fourfold higher than that of OKT 8+ cells. Almost all of the T cells were OKIa1+, indicating that they were in an activated state. In four of the 11 cases, both subsets of T lymphocytes were also found to infiltrate the hair matrix itself. These results would appear to be consistent with the assumption that alopecia areata is caused by a T cell mediated autoimmune mechanism.
Alopecia areata can be treated effectively by topical application of potent contact allergens. To explain the response, the following hypothesis is presented. Alopecia areata is considered an autoimmune disease. The characteristic peribulbar round cell infiltrates probably reflect a cell-mediated immune reaction to some hair-associated antigen. With the elicitation of contact allergy, a second antigen is introduced at the same site. The infiltrates of the allergic contact dermatitis contain suppressor T cells and suppressor macrophages which, in terms of local immunoregulation, exert a nonspecific inhibitory effect on the immune response against hair follicles. Regrowth of hair would be due to a change in the local balance between helper and suppressor cells. In conclusion, the phenomenon of antigenic competition is proposed as a therapeutic concept.
Squaric acid dibutylester (SADBE), a potent contact allergen, was tested for mutagenicity in the bacterial plate incorporation assay (Ames test), in the presence and absence of mammalian microsomes. In contrast to dinitrochlorobenzene which is mutagenic in this test, SADBE was found not to be mutagenic. In 53 patients with extensive or total alopecia areata, SADBE dissolved in acetone was applied weekly to one side of the head, the other side serving as control. In 46 patients (87%), hair regrew either exclusively on the treated side, or considerably faster and denser on this side. In some patients, continuous treatment failed to maintain the response. Persistent response was observed in 37 patients (70%). These results are essentially the same as those obtained with DNCB. Therefore, contact allergy is proposed as a therapeutic concept for alopecia areata.
Topical diphenylcyclopropenone (DPCP) and minoxidil have been used in the treatment of alopecia areata with variable results.
This study was designed to evaluate the efficacy of DPCP alone or in combination with topical 5% minoxidil for the treatment of chronic severe alopecia areata. The effect of therapy on cutaneous T-cell and Langerhans cell subpopulations and intercellular adhesion molecule-1 (ICAM-1) expression was also examined.
Fifteen patients with chronic (more than 2 years), severe (more than 50% scalp involvement) alopecia areata participated in a 24-week trial. Half of the scalp was treated with DPCP once weekly and with either 5% minoxidil solution or a vehicle solution twice daily in a randomized double-blind design. Skin biopsy specimens from each half of the scalp were obtained before therapy and after 12 and 24 weeks of therapy for histologic and immunophenotypic analysis.
Thirteen patients completed the study. Five of 13 patients (38%) showed marked regrowth of coarse terminal hair after 24 weeks of treatment with DPCP. The addition of topical 5% minoxidil did not produce any significant clinical benefit in this 24-week trial. Immunophenotypic analysis showed no differences between responders and nonresponders at baseline. During treatment, Leu-4, Leu-2, Leu-3, and keratinocyte ICAM-1 expression were significantly reduced in biopsy specimens of responders versus nonresponders.
DPCP treatment showed a 38% success rate in producing cosmetically acceptable regrowth in patients with chronic severe alopecia areata.
IL-10 is a product of activated keratinocytes and is released during the induction phase of contact sensitivity. As IL-10 effects have been described as being mediated by APC, we investigated effects of IL-10 on epidermal Langerhans cells (LC), the resident APC in the epidermis. Initial studies failed to demonstrate effects of IL-10 on MHC class II Ag expression by LC or anti-CD3 mAb- or alloantigen-induced LC-dependent T cell proliferation. However, production of IFN-gamma and IL-2, (but not IL-6) was markedly reduced in these assays. When the soluble-protein Ag specific T cell clones AE7 (Th1) and D10.G4 (Th2) were substituted for unprimed T cells, differential effects of IL-10 on T-cell proliferation were observed. Whereas IL-10-pretreated and untreated LC supported Th2 cell proliferation equally well, IL-10-pretreated LC were essentially unable to induce Th1 cell proliferation in response to native protein or peptide Ag. The inhibitory influence of IL-10 on Th1 cells was observed when fresh or 1 day cultured LC were used; 2- or 3-day cultured LC were affected to a much lesser extent by IL-10 pretreatment. Further, coculture experiments using IL-10-pretreated or untreated LC of a different haplotype suggest that IL-10 negatively regulates a costimulatory signal required for induction of Th1 cell proliferation. To assess whether T cells incubated with Ag and IL-10-pretreated LC were responsive to further stimulation, T cells were rescued after 1 day of coculture with IL-10-pretreated LC and restimulated, either immediately or after 1 to 5 days of rest, with untreated LC in the presence of Ag. T cells incubated with IL-10-pretreated LC were found to be anergic, whereas T cells incubated with untreated LC proliferated normally after further stimulation. However, anergic T cells responded vigorously to IL-2. These data indicate that although IL-10-pretreated LC are effective APC for Th2 cells, they fail to induce Th1 cell proliferation and rather induce clonal anergy in these cells.
A novel and reliable high-performance liquid chromatography (HPLC) method is described for the purification and quantification of double-stranded DNA. The nucleic acids may be obtained by polymerase chain reaction (PCR) or as restriction fragments from enzymatic cleavage; the separated products are devoid of contaminating material like agarose, ethidium bromide or non-specific DNA sequences. Because of the non-destructive nature of this HPLC procedure, the purified DNA is optimally suited for cloning experiments. The DNA separation by HPLC has major advantages when combined with reverse transcription (RT)-PCR. This is exemplified by analysis of the TNF-alpha mRNA obtained from endotoxin-elicited rat liver macrophages. If the standard procedure of Northern blotting is compared with the combination of RT-PCR and quantification of the PCR products by HPLC, it is obvious that the dynamic changes of tumor necrosis factor (TNF)-alpha mRNA synthesis are at least as precisely reflected with the RT-PCR/HPLC combination. The latter method is presented as a reliable and powerful tool for quantitative studies on gene expression.
We determined the lymphokines involved in the immune response to epicutaneously applied dinitrofluorobenzene (DNFB), a sensitizer, and dinitrothiocyanobenzene (DNTB), a tolerogen. Hapten-dependent T-cell proliferation and production of interleukin-2, interleukin-3 and interleukin-4 by lymph node cells (LNC) in mice painted with these cross-reactive haptens were measured by specific lymphokine assays. Proliferation of LNC in tolerized animals was lower than in sensitized mice. LNC from DNTB-treated mice produced lower amounts of interleukin-2, interleukin-3 and interleukin-4 than cells from DNFB-painted mice. These results may explain hapten-specific tolerance induced by DNTB which results in deficient production of both type 1 T-helper cell (Th1) and type 2 T-helper cell (Th2) lymphokines in response to hapten re-exposure. Deficient interleukin-4 production by cells from tolerized mice was corrected by the addition of exogenous interleukin-2. The suppressor function of adoptively transferred T cells from animals tolerized with dinitrothiocyanobenzene may be related to a shift in the balance of Th1 and Th2 lymphokines in favour of the latter, since recipient T cells might provide the source of interleukin-2 that induces interleukin-4 production by donor T cells.
