Total serum immunoglobulin E in patients with alopecia areata.

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.

INTRODUCTION

Alopecia areata (AA) is a common form of localized, non-scarring hair loss that occurs on any hair bearing skin.[1] The etiopathogenesis of the disease is still unclear. It is suspected to be an autoimmune disease having a genetic predisposition, influenced by environmental and ethnic factors.[2]

Although, the pathogenesis of AA is poorly understood, evidence which suggests that T cells and cytokines play an important role is accumulating. Previous mouse data suggested that the initiation phase of AA is a heavily Th1-based immune response.[3] However, the maintenance of destruction of the hair follicles by cytotoxic cells was suggested to be due to a shift from a Th1 response to a more chronic Th2 immune profile.[4] Thus, AA is a cell-mediated autoimmune disease with late, possibly secondary, humoral response.[5]

Serum IgE concentrations are high in atopy,[6] parasitosis,[7] human immunodeficiency virus infection,[8] systemic lupus erythematosus,[9] and certain types of cancer.[10] The production of IgE is usually Th2 cell determined. The Th2 cytokines, IL (interleukin)-4 and IL-13, are required signals for IgE synthesis. Keratinocytes do not produce IL-4 or IL-13, but are involved in IL-4 or IL-13 induced biological effects.[11] Different studies have measured IgE in AA patients with controversial results.[121314]

The aim of this work was to evaluate total serum IgE in Egyptian AA patients to investigate the role of Th2 cytokines, which are known to stimulate IgE production, in AA pathogenesis.

MATERIALS AND METHODS

Ethics

Written consent forms approved by the Committee of Human Rights in Research in Menoufiya University were obtained from studied cases and control subjects. Those were in accordance with the Helsinki Declaration of 1975 (revised in 2000).

Patients

This study included 50 cases with AA and 50 healthy, age- and sex-matched subjects as a control group. Cases were selected from the Dermatology Outpatient Clinic, Menoufiya University Hospital between February 2012 and December 2012.

All studied patients were subjected to complete history taking, general, and dermatological examination. Demographic data (age and gender) and clinical variables (site of lesions, disease severity, disease duration, and family history of AA in first degree relatives of every case) were all documented. Disease duration was calculated from disease onset to time of the first visit.

The severity of AA was graded according to Kavak et al.[15] as follows:

Mild: The presence of three or less patches of alopecia with a widest diameter of 3 cm or less, or the disease is limited to the eyelashes and eyebrows

Moderate: Existence of more than three patches of alopecia, or a patch greater than 3 cm at the widest diameter without alopecia totalis or alopecia universalis

Severe: Alopecia totalis or alopecia universalis

Ophiasis: Snake shaped plaques extending to the scalp border or loss of hair in the shape of a wave at the circumference of the head.

Patients with one or more of the following were excluded:

Atopy or any form of allergy, including first degree relatives. This was carried out by:

History taking (past or current history of atopic dermatitis, asthma and/or allergic rhinoconjunctivitis in cases and/or relatives)

Clinical examination (dermatological, ophthalmic, chest and ear, nose, and throat examination)

Skin prick test to a panel of allergens.

Bacterial, viral infection and/or parasitic infestations

Smokers

Use of any form of topical or systemic treatment

Sessions of Psoralen + UVA (PUVA), for at least 6 months before this study

Hyper IgE syndrome and acute phase of acute coronary syndromes

Past or present history of malignancy.

Measurement of serum total IgE

Cases and control subjects were subjected to determination of total serum IgE level by enzyme-linked immunosorbent assay using the enzyme immunoassay for the quantitative determination of IgE concentration in human serum, Immunospec Corporation, Canoga Park CA.

Skin biopsy taking

Skin biopsy samples were taken under local anesthesia from every case from an active area of hair loss. All specimens were fixed in 10% neutral-buffered formalin and subjected to routine tissue processing that ended with paraffin-embedded blocks ready for sectioning.

Statistical procedure

Results were collected, tabulated, statistically analyzed using statistical package SPSS version 11. Data were statistically described in terms of range, mean ± standard deviation (±SD), frequencies (number of cases) and relative frequencies (percentages) when appropriate. Kruskal-Wallis test was used for comparison between three or more groups not normally distributed having quantitative variables. Comparison of quantitative variables was carried out using Mann-Whitney test for independent samples. Associations between inflammatory cell infiltration and laboratory values were analyzed using the Pearson's correlation test. A P < 0.05 was considered statistically significant.

RESULTS

Studied population

Cases included 28 males (56%) and 22 females (44%). Patients’ ages ranged from 10 years to 53 years with a mean ± SD age of 28.38 ± 11.52 years. Disease duration ranged from 1 month to 26 months with a mean ±SD duration of 4.39 ± 4.74 months [Table 1].

Table 1

Demographic and clinical data of studied patients

Control group included 25 males (50%) and 25 females (50%). Their ages ranged from 12 years to 50 years with a mean ±SD ageof 27.54 ± 10.36 years.

Thirty cases (60%) had scalp AA, 10 cases (20%) had beard AA and 10 cases (20%) had alopecia universalis [Table 1].

Regarding disease severity, 17 cases (34%) had mild AA, 18 cases (36%) had moderate AA, 10 cases (20%) had severe AA and 5 cases (10%) had ophiasis. Family history was positive in 12 (24%) cases [Table 1].

Histopathological examination

Examination of hematoxylin and eosin-stained sections sections showed increased proportion of catagen and telogen hair follicles, and perifollicular mononuclear inflammatory infiltration. Lesional mononuclear cell count around hair follicles has a mean ± SD value of 25.38 ± 12.69 cell. Slight perivascular inflammatory cell infiltration comprised mainly of mononuclear cells was also noted. Eosinophilic infiltration around lower hair follicles was found in 25 patients (50%) (5 cases with mild, 8 cases with moderate, 10 cases with severe AA and 2 cases with ophiasis). Lesional eosinophilic count around hair follicles has a mean ± SD value of 2.15 ± 1.68.

Serum IgE in studied population

Serum IgE level in studied cases ranged from 13.5 IU/ml to 780 IU/ml with a mean ± SD value of 187.92 ± 242.51 (IU/ml). IgE was elevated in 27 cases (54%). There was a statistically significant difference between cases and controls regarding mean value of total IgE (P < 0.05) [Table 2].

Table 2

Comparison between cases and control group regarding mean values of serum IgE

There were insignificant differences in mean values of IgE among AA cases regarding disease severity (P > 0.05), Patients’ ages (P > 0.05), patients gender (P > 0.05), disease duration (P > 0.05), and site of affection and positive family history of AA [Table 3].

Table 3

Association between IgE levels and clinical parameters of AA cases

Serum IgE level was not correlated with the perifollicular mononuclear cell infiltrate or esinophilic count (P > 0.05) [Table 4].

Table 4

Pearson's correlation between IgE values and histopathological changes in AA cases

DISCUSSION

Immunoglobulin E plays a crucial role in the pathogenesis of allergic disorders. The process involved in IgE production by B cells include germline ε transcript expression, IgE class switching, clonal expansion of B cells, and differentiation into IgE-secreting plasma cells.[16]

It is believed that AA is an organ-specific autoimmune disease, targeting anagen-stage follicles which leads to disruption of hair follicle growth.[17]

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]

The aim of this work was to evaluate total serum IgE in Egyptian AA patients to investigate the role of Th2 cytokines, which are known to stimulate IgE production, in AA pathogenesis.

In the current study, total serum IgE was elevated in 27 (54%) cases with significant difference in its mean values between cases and controls.

The association of serum IgE levels and AA has been previously investigated with varying results. Przybilla et al.[21] found elevated total IgE in 19.7% of studied AA patients. Later on, O’Loughlin et al.[22] found elevated total serum IgE in 30% of AA patients. Kasumagić-Halilović and Prohić[14] also found elevated total IgE in 37% of AA patients. Attia et al.[5] found elevated total IgE in 48.3% of Egyptian AA cases. Zuel Fakkar et al.[23] found elevated total IgE in 50% of studied Egyptian cases. Zhao et al.[24] demonstrated significant elevation of IgE in patchy AA than healthy control. Bork et al.[25] also recorded a significant increase in serum IgE (32%) in children with AA.

However, in contrast to these results, the total serum IgE levels were not elevated in AA patients than normal population in other previous similar studies.[262728] This discrepancy could be attributed the different numbers of patients involved, and to the different environmental factors, which may affect IgE level.

The mechanism of IgE elevation in AA is not exactly known. A variety of cytokines control IgE production. IL-4, IL-6, IL-7, IL-9 and IL-13, enhance IgE production. In contrast, IFN-γ and IL-10 inhibit IgE synthesis.[11]

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.

Another possible explanation can be introduced based on the observation of Katagiri et al.[20] who reported lower levels of IFN-γ, and TGF-β1 mRNA in non-atopic patients with AA than those in healthy controls. Decreased levels of IFN-γ and Transforming growth factor β1 (TGF-β1) were also shown in patients with atopic dermatitis. These results indicated a similarity between atopic dermatitis and AA based on their cytokine profile. Decreased levels of these cytokines lead to enhanced IgE production. Going with that, experiments carried out by Deeths et al.[30] showed that antigen specific T-cells from AA patients appear to have some intrinsic defect towards the production of IFN-γ, possibly suggesting a state of partial tolerance in the skin of these patients.

Contrary to these results, Teraki et al.[19] reported that serum levels of IFN-γ were significantly elevated in patients with extensive AA. However, Gregoriou et al.[31] concluded that, the elevated serum levels of IFN-γ in AA patients may reflect the state of inflammation, especially in the extensive forms of the disease, and the measurement of serum IFN-γ may be useful in discriminating those who are likely to develop alopecia universalis from the remaining local disease, or as a prognostic indicator.

Elevated serum IgE in AA cases may also due to increased tumor necrosis factor (TNF)- α that is known to play a key role in the pathogenesis of AA. TNF-α is synthesized in epidermal keratinocytes along with several other cytokines[32] and is a very potent inhibitor of proliferation.[33] TNF-α may enhance IgE levels by creating a micro-environment rich in the Th2 cytokines, IL-4 and IL-13, stimulating IgE class switching.[34]

In addition, the sera of patients with AA have been found to contain extremely high levels of B cell activating factor (BAFF) that belongs to the TNF family, produced by myeloid lineage cells.[35] It is considered that the production of BAFF is stimulated by IFN-γ that is known to be increased in severe AA patients.[3637]

Previous studies reported higher serum levels of IL-4 in AA cases than in normal subjects.[520] IL-4 is a stimulant of IgE production.[11]

Another possible mechanism of IgE elevation in AA patients may be also due to over-expression of CD40. It is a member of a family of surface molecules with homology to the nerve growth factor receptor and is expressed on B cells.[38] Zhang et al.[39] suggested that CD40 stimulation alone could enhance IgE production from in vivo - driven, IgE-producing cells. CD40 was also found to be expressed in the hair structures including the dermal papilla of AA lesions confirming a possible role in elevated serum IgE in some AA patients.[40]

Another possible explanation for elevated IgE in AA is supported by genetic linkage. Many genetic loci and defined genes have been associated with the etiology of allergic disorders.[41] The atopic disorders have been reported to occur with an incidence that varies from 1% to 52% in patients with AA.[42] It has also been observed that AA pursues a severe course in the presence of atopy and has an early onset and longer duration, with poor response to treatment.[43]

Association between atopy and 11q13 genetic locus was discovered by Young et al.[44] Subsequent research revealed that 11q13 genetic locus contains among other genes the FCεRIβ gene, encoding for the high affinity IgE receptor β chain (FCεRIβ).[45] Genes for Ig heavy (Gm) and light (Km) chain genotypes, discovered on human chromosome 14, have also been implicated in AA susceptibility.[46] Attia et al.[5] reported that elevated serum IgE in patients with AA may be related to their HLA profile. Elevated IL-4 and IgE were observed in patients with DRB1*07 and DRB1*11.

With respect to histopathological changes, our study showed perifollicular esinophilic infiltration in 50% of examined cases. Zhao et al.[24] reported the presence of esinophilic infiltration around hair follicles in patchy AA. Elston et al.[47] detected eosinophils in all stages of AA. Peckham et al.[48] detected the presence of eosinophils in fibrous tracts and in peribulbar infiltrates in 44% of examined AA biopsies and concluded that eosinophilic infiltration in fibrous tracts and near hair bulbs is a helpful diagnostic feature of AA, especially when biopsy specimens lack the peribulbar lymphocytic infiltrate.

It was postulated that activated eosinophils may underlie an increased total serum IgE level in patients with eosinophilic pneumonia.[49] Further studies are required to confirm whether this is true for AA or not.

In the present study, there was insignificant difference in mean IgE levels among AA cases with different disease severities. This was in line with Kasumagić-Halilović and Prohić[14] and Zuel Fakkar et al.[23] However, contrary to our finding, Attia et al.[5] reported significant elevation of IgE in alopecia universalis than patchy AA and alopecia totalis. This conflict may be due to different proportions of severe and localized cases in the two studies.

In the present work, there was insignificant variation in mean IgE levels among AA cases with with different disease durations. This was at variance with the report of Zuel Fakkar et al.[23] and Attia et al.[5] who found a significant association between elevated IgE and long disease duration. Higher values were reported among patients having disease for more than one year. This controversy may be attributed to different clinical characteristics of studied cases in each study.

Insignificant differences in IgE levels among cases, regarding their ages and gender in our study were similar to the findings of Zuel Fakkar et al.[23]

In the current work, no correlation was detected between IgE values and lesional esinophilic count. Similar result was obtained by Zhao et al.[24] in their studied population of patchy AA.

In summary, IgE elevation in AA cases suggests a shift from a Th1 response in early AA to a more chronic Th2 immune profile, with secondary B-cell stimulation and possible IgE class switching. However, esinophilic infiltration and/or genetic factors may be also responsible for such elevation. Further studies on a larger scale and on different AA presentations are required for firmer conclusions.