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Serum 25-hydroxy vitamin D3 level in Egyptian patients with alopecia areata

El-Mongy, Naglaa N.a; El-Nabarawy, Emana; Hassaan, Shahenda A.c; Younis, Eman R.d; Shaker, Olfatb

Journal of the Egyptian Women's Dermatologic Society: January 2013 - Volume 10 - Issue 1 - p 37–41
doi: 10.1097/01.EWX.0000419612.74665.2b
Original articles
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Background Vitamin D has multiple effects on innate and adaptive immune responses. The relation between vitamin D levels and alopecia areata (AA), a tissue-specific autoimmune disease, represents an attractive area of research.

Objective The objective of the study was to estimate the serum level of 25-hydroxy vitamin D (25-OH Vit D) in AA patients and explore a probable relation.

Patients and methods A total of 140 individuals from the Egyptian population were recruited into a case–control study, of whom 70 were patients with AA of various types and 70 were healthy controls. Serum 25-OH Vit D levels were measured. Correlations with disease duration and severity, positive family history, age and sex were made.

Results AA patients comprised 37 male and 33 female patients (13–50 years). The healthy controls comprised 44 male and 26 female individuals (12–50 years). Serum 25-OH Vit D levels were significantly decreased in AA patients than in controls (P=0.023). The female participants in both groups (patients and controls) showed significantly lower vitamin D levels compared with male participants (P=0.005). There was no significant relationship between serum 25-OH Vit D levels and age, duration, severity or positive family history.

Conclusion AA is associated with low levels of 25-OH Vit D, especially in female participants. More studies are required to prove its possible role in AA pathogenesis.

Departments of aDermatology

bMedical Biochemistry, Cairo University

Departments of cDermatology

dBiochemistry, National Research Centre, Cairo, Egypt

Correspondence to Naglaa N. El-Mongy, 35 Dokki Street, Apt. 111, 12311 Giza, Egypt Tel: +20 100 171 3196; fax: +20 223 687 673; e-mail: nnradw@yahoo.com

Received May 21, 2012

Accepted July 25, 2012

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Introduction

Alopecia areata (AA) is a tissue-restricted autoimmune disease mediated by T lymphocytes with a Th1 cytokine profile 1. AA occurs mostly in genetically predisposed individuals triggered by environmental factors 2. There is evidence for loss of immune privilege coupled with T-cell-mediated attack of hair follicle autoantigens, as well as for the role of autoantibodies in the pathogenesis of AA 1.

Interferon-γ, interleukins (IL) and tumor necrosis factor-α are cytokines that are known to have a major role in the pathogenesis of the disease 3. CD8+ T cells are considered to act as effector cells with help from CD4+ T cells 4. The analysis of the skin of mice with chronic AA and of mice receiving a transplant of AA-affected skin suggests that autoaggression is due to a defect in regulatory T cells 5. Many autoimmune conditions have been associated with reduced vitamin D levels 6. However; little is known about the association between AA and reduced vitamin D levels.

Vitamin D is a steroid hormone synthesized in epidermal keratinocytes under the influence of UV-B light or acquired from diet and dietary supplements 7. The active form of vitamin D, 1,25-dihydroxyvitamin D3 [1,25(OH)2 Vit D3], has multiple effects on innate and adaptive immune responses through its varied effects on T and B lymphocytes, macrophages and dendritic cells (DCs), all of which express vitamin D receptors (VDRs) 6. As such, the impact of vitamin D on human physiology and disease is broad and there is wide interest in the role of this hormone in many areas of medicine 8. Determination of vitamin D status is not based on measurement of serum 1,25(OH)2 Vit D3 concentrations. It is assessed by measuring the prohormone 25-hydroxy vitamin D (25-OH Vit D), which is an indicator of supply rather than function. The most stable and plentiful metabolite of vitamin D in human serum is 25-OH Vit D, which has a half-life of about 3 weeks, making it the most suitable indicator of the vitamin D status 9.

The study aimed at the evaluation of serum 25-OH Vit D levels in patients with AA in an attempt to identify any possible relation in the pathogenesis of the disease.

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Patients and methods

The present study included 140 individuals from the Egyptian population: 70 AA patients and 70 healthy controls. All patients of AA were recruited from the Outpatient Clinic of Dermatology, Kasr Al Aini University Hospital and National Research Centre, Egypt, during a period of 6 months (1 January 2011 to 30 June 2011). The study protocol was approved by the Dermatology Research Ethical Committee (Derma REC) of the Faculty of Medicine, Cairo University. Written informed consent was signed by patients and by one parent in the case of minors (below 18 years). The following patients were excluded: any patient known to have a state of vitamin D deficiency or a disease that could affect its level (e.g. renal diseases, cancers, Systemic Lupus Erythematosus or psoriasis) and those who were experiencing spontaneous regrowth of terminal hair at the time of presentation. Furthermore, any patient who was taking vitamin D supplementation, oral or topical, for the last 2 months or receiving Psoralen+Ultra Violet A (PUVA) treatment was excluded. History of the patients, including age, sex, duration, history of adequacy of sun exposure and family history of similar condition, was recorded carefully. Adequacy of sun exposure was considered as unprotected, noon time exposure for 5 min daily in fair skinned individuals or 10 min daily in dark skinned individuals of more than 5% of the body surface area (the arms and legs, or the hands, arms and face, two or three times per week) 10.

The extent of scalp hair loss was determined by dividing the scalp into four quadrants, followed by visually determining the percentage of scalp hair loss in each quadrant and adding the numbers together, with a maximum score of 100%. This was determined according to the severity of alopecia severity in alopecia tool (SALT) score 11. Scalp hair loss (S) was classified as follows: S0=no hair loss; S1<25% hair loss; S2=25–49% hair loss; S3=50–74% hair loss; S4=75–99% hair loss, a=75–95% hair loss, b=96–99% hair loss; and S5=100% hair loss. Patients were classified according to the severity of AA into: mild AA – patients who were S1; moderate AA – patients who were S2; and severe AA – patients who were S3, S4 and S5. Patients were also classified according to the pattern of AA into patchy AA or extensive AA [alopecia totalis/alopecia universalis (AT/AU)].

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Quantitation of 25-hydroxy vitamin D in serum

In the present study, the serum 25-OH Vit D levels were measured by a direct enzyme-linked immunosorbent assay with the 25-Hydroxy Vitamin D Direct EIA kit purchased from Immunodiagnostic Systems Ltd (Stubenwald-Allee, Bensheim, Germany).

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Assessment of vitamin D status

Vitamin D status was classified according to serum 25-OH Vit D concentrations into:

  • vitamin D deficiency: 25-OH Vit D concentration less than 30 nmol/l;
  • vitamin D insufficiency: 25-OH Vit D concentration 30–75 nmol/l;
  • vitamin D sufficiency: 25-OH Vit D concentration 75 nmol/l or more 9,12.
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Statistical analysis

All statistical calculations were carried out using computer programs statistical package for the social science (SPSS; SPSS Inc., Chicago, Illinois, USA) version 15, for Microsoft Windows. Comparisons of numerical variables between the study groups were made using the Student t-test for independent samples when comparing two groups and the one-way analysis of variance test when comparing more than two groups. For comparing categorical data, the χ2-test was performed. The exact test was used when the expected frequency was less than 5. Accuracy was represented using the terms sensitivity and specificity. Receiver operator characteristic analysis was used to determine the optimum cutoff value. Correlation between various variables was made using the Pearson moment correlation equation for linear relation. P-values less than or equal to 0.05 were considered statistically significant.

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Results

This case–control study was conducted on 70 patients with AA and on 70 normal controls. The demographic and clinical data of the studied groups are presented in Table 1.

Table 1

Table 1

A significant difference was found in serum 25-OH Vit D levels between patients and controls (P<0.05) (Table 2). In contrast, vitamin D status showed a statistically border-line significant difference (P=0.05). Vitamin D deficiency and insufficiency were more common in patients than in controls (Table 3).

Table 2

Table 2

Table 3

Table 3

To determine the cutoff point for 25-OH Vit D level with the best sensitivity and specificity, we performed an receiver operator characteristic analysis, which revealed a level of 65.63 nmol/l as a cutoff point with a sensitivity of 58.6% and a specificity of 55.7% (CI: 0.480 to 0.671, P=0.123), as shown in Fig. 1.

Figure 1

Figure 1

There was a highly significant difference in serum 25-OH Vit D levels between male and female participants, among both patients and controls. Serum 25-OH Vit D levels were much lower in female participants than in male participants in both groups (P=0.005, 0.005) (Table 4). Vitamin D deficiency and vitamin D insufficiency were more common in female than in male participants (P<0.0005) (Fig. 2).

Table 4

Table 4

Figure 2

Figure 2

The participants under study were divided into four age groups: group 1 – from 10 to 20 years; group 2 – from 21 to 30 years; group 3 – from 31 to 40 years; and group 4 – from 41 to 50 years. By comparing the serum 25-OH Vit D levels of patients in all age groups, it was found that the serum 25-OH Vit D level in the third group was the highest and that the older age group (from 41 to 50 years) showed the lowest level. However, this was not statistically significant (P=0.178). In the control group, the serum 25-OH Vit D levels in the first group were the highest and the older age group (from 41 to 50 years) again showed the lowest levels. The difference between these groups also was not statistically significant (P=0.350).

There was no statistically significant relationship between serum 25-OH Vit D level and adequacy of sun exposure in patients, whereas in controls the relationship was highly significant (P<0.0005). There was no positive correlation between serum 25-OH Vit D levels and duration of disease, type, severity of AA or family history (P=0.674, 0.140, 0.176 and 0.589, respectively).

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Discussion

The present study showed a statistically significant decrease in the mean serum 25-OH Vit D levels in patients with AA. To our knowledge, this is the first study to determine the serum 25-OH Vit D levels in patients with AA.

The active form of vitamin D has a role in immunoregulation through inhibition of the maturation and differentiation of DCs 13. It inhibits the production of IL-12 (an immunostimulatory cytokine secreted by DCs, which is crucial for polarizing the immune system towards the Th1 phenotype). Furthermore, vitamin D increases the production of IL-10 by DCs, which counteracts the function of IL-12 14. It inhibits T-cell proliferation and the secretion of some Th1 cytokines, primarily interferon-γ and IL-2 15. It also increases the secretion of Th2 cytokines (IL-4, IL-5 and IL-10) 16. Vitamin D also causes suppression of autoimmune diseases by enhancing the production and function of regulatory T cells 17, which is vital for preserving peripheral self-tolerance 14. It also causes inhibition of Th17 cell functions and differentiation 17. Th17 cells were shown to play a significant role in autoimmune reactions 18. Although vitamin D is thought to play a role in hair production, most of the evidence for this hypothesis comes from studies on mice lacking the VDRs and on humans with VDR mutations 19.

In the patient group, the relation between serum 25-OH Vit D level and sun exposure was not significant, whereas in the control group it was significant. This controversy suggests the presence of factors in patients with AA that may affect the cutaneous photosynthesis of vitamin D or its further activation in the liver by 25-hydroxylase enzyme. This result also revealed that sun exposure alone cannot alter the vitamin D status in patients with AA and that vitamin D supplementation is required for the treatment of vitamin D deficiency in those patients.

More than 90% of the vitamin D requirement for most people comes from casual exposure to sunlight 20. However, the exact amount of sun exposure required for optimal vitamin D levels is affected by several factors such as time of day, season, latitude, skin color, body fat, excessive use of sunscreens, old age 10 and air pollution 21.

In the present study, the serum 25-OH Vit D level – the best clinical index of vitamin D status – was significantly much lower in female participants than in male participants, among both patients and controls. Only two out of 33 female patients had the optimal level of vitamin D. Many studies have shown the prevalence of vitamin D deficiency in the female population 22–24. Age, season, physical activity and hormonal status are important predictors of vitamin D status in women 25. The groups that are at greatest risk include: house-bound, dark-skinned people who are modestly dressed and those who regularly avoid sun exposure or work indoors 26. In our study, social and cultural factors such as the conservative dress of Egyptian women block exposure to sunlight. In addition, the reduction in outdoor leisure time and the increase in office-based work have led to an increased lack of sunlight exposure.

There was an insignificant tendency for a lower serum 25-OH Vit D level in the older age group (from 41 to 50 years) among both patients and controls. This may be because of an age-related decline in skin 7-dehydrocholesterol content, decrease in sun exposure and decrease in the intake of vitamin D 27.

There was no significant linear correlation between serum 25-OH Vit D levels and the duration or severity of AA. Therefore, a low serum 25-OH Vit D level may play a role only in the incidence of the disease but not in the subsequent course.

A nonsignificant decrease in the serum 25-OH Vit D level was noted in patients with extensive AA (AT/AU). This may be because of the limited number of patients with AT/AU. Furthermore, no statistically significant relation was found between serum 25-OH Vit D level and family history of AA.

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Conclusion

AA is associated with low serum 25-OH Vit D, especially in the female population. More studies are needed to determine its possible role as a cause of the disease.

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Acknowledgements

The authors express their gratitude to Professor Wedad Mostafa (Dermatology Department, Cairo University) and Professor Hanaa Emam (National Research Centre) whose help, stimulating suggestions and encouragement helped at the time of research involved in writing this paper.

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Conflicts of interest

There are no conflicts of interest.

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Keywords:

adaptive immunity; alopecia areata; autoimmunity; innate immunity; vitamin D

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