|Year : 2019 | Volume
| Issue : 1 | Page : 14-20
Decreased serum levels of immunoglobulin M and increased complement 3 in Egyptian vitiligo patients
Dina M Kadry1, Heba M. Abdel Raheem1, Heba N Baz2, Rania I Sallam1
1 Department of Dermatology, Faculty of Medicine, Cairo University, Cairo, Egypt
2 Department of Clinical and Chemical Pathology, Faculty of Medicine, Cairo University, Cairo, Egypt
|Date of Submission||23-Apr-2018|
|Date of Acceptance||23-Oct-2018|
|Date of Web Publication||28-Jan-2019|
Heba M. Abdel Raheem
52, Mohamed Mandour Street, Nasr City, PO 11759, Cairo, Egypt
Source of Support: None, Conflict of Interest: None
Background Vitiligo is a common, acquired, depigmenting disorder that results from loss of functional melanocytes in the skin and mucous membranes. It is occasionally associated with autoantibodies against the melanocyte-specific proteins such as tyrosine-related protein-1 tyrosine-related protein-2. These antibodies could induce melanocyte damage in vitro by a complement-mediated mechanism and antibody-dependent cellular cytotoxicity might play an active role in the stimulation and inappropriate expression of HLA-DR and induction of intercellular adhesion molecule-1 on melanocytes.
Objective Our aim was to assess the serum levels of different immunoglobulins (Ig) in addition to C3 and C4 in vitiligo patients and to determine their correlation with different clinical aspects of the disease.
Patients and methods Serum levels of IgG, IgA, and IgM as well as C3 and C4 were estimated in 24 vitiligo patients and 24 healthy controls using the Radial Immuno-Diffusion (immunoprecipitation) technique.
Results Serum IgM levels were found to be significantly decreased and C3 levels were significantly increased among patients compared with the controls. No significant difference was found in the serum levels of IgA, IgG, or C4 between both groups. Also, a significant positive correlation was found between serum IgA levels and the age of the patients, as well as between C3 and C4 and a family history of vitiligo.
Conclusion Changes in Ig serum levels might be a cause or an effect in the pathogenesis of vitiligo, as well as alterations in the complement system. These ffindings suggest that altered immunity is to be considered in the therapeutic management of these patients.
Keywords: complement, immunoglobulins, vitiligo
|How to cite this article:|
Kadry DM, Abdel Raheem HM, Baz HN, Sallam RI. Decreased serum levels of immunoglobulin M and increased complement 3 in Egyptian vitiligo patients. Egypt J Dermatol Venerol 2019;39:14-20
|How to cite this URL:|
Kadry DM, Abdel Raheem HM, Baz HN, Sallam RI. Decreased serum levels of immunoglobulin M and increased complement 3 in Egyptian vitiligo patients. Egypt J Dermatol Venerol [serial online] 2019 [cited 2019 Jul 18];39:14-20. Available from: http://www.ejdv.eg.net/text.asp?2019/39/1/14/250814
| Introduction|| |
Vitiligo is a common, acquired, depigmenting disorder that results from loss of functional melanocytes in the skin and mucous membranes. It affects about 0.1–2% of the worldwide population . Vitiligo usually presents with milky white macules or patches . Vitiligo is frequently associated with disorders of autoimmune origin . Research hypotheses for its pathogenesis go in three main directions: an inherent defect in the melanocyte, an alteration in the development of the peripheral nervous system damaging the neural crest-derived melanocytes, and a dysregulation of the immune response .
Vitiligo is occasionally associated with autoantibodies against the melanocyte-specific proteins such as autoantibodies against tyrosinase  tyrosine-related protein-1  and tyrosine-related protein-2 . These antibodies could induce melanocyte damage in vitro by a complement-mediated mechanism and antibody-dependent cellular cytotoxicity might play an active role in the stimulation and inappropriate expression of HLA-DR and induction of intercellular adhesion molecule-1 on melanocytes . Ruiz-Argüelles et al.  found antibodies to a 75 kDa protein in serum samples that induced apoptosis in cultured melanocytes.
For the complement system, heterozygous C4 deficiency has been linked to an increased risk for vitiligo . Complement-activating anti-melanocyte antibodies have been implicated in the pathogenesis of vitiligo. A plasmatic protein called mannose-binding lectin activates the classic complement pathway through an associated serine protease . Aberrant expression of complement-regulatory proteins leaves melanocytes sensitive to complement-mediated lysis ,.
| Patients and methods|| |
Twenty-four patients with vitiligo (and no history of any other autoimmune disease) were included in this case–control study. They were recruited from the Dermatology Outpatient Clinic at the Faculty of Medicine, Cairo University, during the period from December 2013 to March 2014. Twenty-four age-matched and sex-matched healthy volunteers were included as a control group. Informed written consent and the approval of the dermatology Research Ethics Committee were obtained from every participant before enrollment in this study.
All patients were subjected to a complete assessment of history including the onset, course, and duration of the disease, history of any precipitating or aggravating factors, and personal, family, and past history. A six-point Vitiligo Disease Activity (VIDA) score was calculated for all patients to determine disease activity . In this score, grading is performed on the basis of disease activity and time period. It is graded as follows: VIDA score +4 (activity lasting 6 weeks or less), score +3 (activity lasting 6 weeks to 3 months), score 2 (activity lasting 3–6 months), score 1 (activity lasting 6–12 months), score 0 (stable for 1 year or more), and score −1 (stable with spontaneous repigmentation for 1 year or more). A low VIDA score indicates less vitiligo activity. A clinical examination was performed to determine the type and extent of vitiligo.
The immunoglobulin (Ig)G, IgM, IgA, C3, and C4 were assayed using the Radial Immuno-Diffusion (immunoprecipitation) technique. Three centimeters of venous blood was obtained by aseptic venipuncture. All samples were collected on serum separator tubes that were separated by centrifugation. The serum was then harvested and stored in an Eppendorf 1.5 ml tube, frozen at −20°C. The procedure is based on the principle of immunoprecipitation in agarose between an antigen and its homologous antibody. This is performed by incorporating one of the two immune reactants uniformly throughout a layer of agarose gel and then introducing the other reactants into wells duly punched in the gel. Antigen diffuses radially out of the well into the surrounding gel–antibody mixture and a visible ring of precipitation forms where the antigen and antibody reacts. A quantitative relationship exists between ring diameter and antigen concentration. On reaction completion, the ratio between the ring square diameter and the antigen concentration shows a linear ratio.
Reading of results
The end point of diffusion was indicated by a sharp precipitation ring. This is achieved when the incubation period is complete. The precipitation rings were measured with 0.1 mm precision using a ruler. The measurement was then compared with a reference table to yield the exact concentration of the studied immunoglobulins and complement factors.
Data were described statistically in terms of mean±SD, median, and range, or frequencies (number of cases) and percentages when appropriate. Comparison of numerical variables between the study groups was performed using the Mann–Whitney U test for independent samples. For comparison of categorical data, the χ2 test was performed. Fisher’s exact test was used when the expected frequency was less than 5. The correlation between various variables was determined using the Pearson product–moment correlation coefficient for linear relations in normally distributed variables and Spearman’s rank correlation coefficient for non-normal variables. P values less than 0.05 were considered statistically significant. All statistical calculations were carried out using the computer program statistical package for the social science (SPSS Inc., Chicago, Illinois, USA) release 15 for Microsoft Windows (2006).
| Results|| |
This study was carried out on 24 patients with vitiligo, nine (37.5%) men and 15 (62.5%) women, whose ages ranged from 18 to 60 years, with a mean age of 34.04±13.741. Twenty-four age-matched and sex-matched healthy individuals were included as controls, 13 (54.2%) men and 11 (45.8%) women, whose ages ranged from 18 to 45 years, with a mean age of 27.92±7.058. There was no statistically significant difference between the age or the sex of the patients and the controls (P=0.223 and 0.247, respectively) ([Table 1]). Twenty-one (87.5%) patients had generalized vitiligo and three (12.5%) patients had other types of vitiligo. The duration of the disease ranged from 1 to 31 years, with a mean of 9.13±8.53 years. The extent of body involvement ranged from 2 to 70%, with a mean of 33.17±20.09. Six (25%) patients had a positive family history and 18 (75%) patients had a negative family history. The VIDA score ranged from 0 to 4, with a median of 4. Seventeen patients had VIDA score 4, five patients had VIDA score 3, one patient had VIDA score 2, and one patient had VIDA score 0 ([Table 2]).
In terms of the serum levels of Ig between patients and controls, the IgA serum level among the patients ranged from 46 to 374 mg/dl (180.83±101.02), whereas among the controls, it ranged from 20 to 413 mg/dl (141.72±99.76). Serum IgG level among the patients ranged from 185.8 to 2057.9 mg/dl (1162.38±489.62), and among the controls, it ranged from 469.1 to 1786.3 mg/dl (1039.55±360.19). There was no statistically significant difference between serum levels of IgA and IgG among patients and controls, with P values of 0.131 and 0.396, respectively. However, there was a statistically significant difference between the serum level of IgM among patients, which ranged from 13.4 to 142.3 mg/dl (57.85±33.85), and controls, which ranged from 9.8 to 187.9 mg/dl (84.79±46.68), with a P value of 0.031 ([Figure 1]).
|Figure 1 Mean values of serum immunoglobulins among patients and controls.|
Click here to view
The serum level of C3 among the patients ranged from 19 to 176 mg/dl (99.73±38.76) and among the controls, it ranged from 19 to 148 mg/dl (74.71±30.29), with a statistically significant difference (P=0.026). No significant difference was found between serum levels of C4 among patients, which ranged from 3 to 53 mg/dl (22.46±14.11), and controls, which ranged from 3 to 34 mg/dl (17.73±8.2) (P=0.278) ([Figure 2]).
A statistically significant positive correlation was found between serum IgA level and the age of the patients (r=0.388, P=0.006) ([Figure 3]). However, serum IgA level did not correlate with the patients’ sex, disease duration, disease extent, family history, or VIDA score. Also, a statistically significant positive correlation was found between serum IgG level and the duration of the disease (r=0.474, P=0.019) ([Figure 4]), but it did not correlate with the patients’ age, sex, disease extent, family history, or the VIDA score. There was no statistical significant correlation between the serum IgM level and any of the measured clinical parameters of the disease.
|Figure 3 Correlation between serum immunoglobulin A (IgA) level and the age of the patients.|
Click here to view
|Figure 4 Correlation between serum immunoglobulin G (IgG) level and duration of the disease.|
Click here to view
A statistically significant positive correlation was found between serum C3 level and family history (r=0.552, P=0.005). However, it did not correlate with the patients’ age, sex, disease extent, disease duration, or the VIDA score. Similarly, a statistically significant positive correlation was found between serum C4 level and family history (r=0.515, P=0.010), with no other significant correlations.
A significant positive correlation was found between the mean serum level of C3 and C4 (r=0.644, P=0.001) among the patients.
| Discussion|| |
Vitiligo is an acquired, common, depigmenting skin disorder, characterized by progressive, well-circumscribed milky macules . The pathogenesis of vitiligo is unclear. Different theories have been put forward in an attempt to explain the cause of vitiligo, including genetic, autoimmune, oxidant-antioxidant, auto-cytotoxic, neural, viral, and ultraviolet radiation theory, for the destruction of epidermal melanocytes .
An elevated ratio of CD4+/CD8+ T cells was found in vitiligo patients, which was considered a sign of an imbalanced lymphocyte immune response in vitiligo patients, but no evidence for a pathological distribution of B cells in peripheral blood was found among these patients . Different circulating antibodies to melanocytes have been found in the sera of vitiligo patients .
The aim of this study was to further investigate the role of humoral immunity and the complement system in the pathogenesis of vitiligo. The study included 24 vitiligo patients and 24 age-matched and sex-matched healthy control participants.
In our study, the serum Ig profile showed no significant difference in serum IgA or IgG among the patients and the controls (P=0.131 and 0.396), respectively. However, the serum level of IgM was significantly decreased in patients compared with the controls (P=0.031). These findings might suggest that IgM anti-melanocyte antibodies are involved in the pathogenesis of vitiligo. In contrast to our results, Ali et al.  and Singh et al.  detected significantly lower serum IgG and IgA levels in patients compared with controls, whereas the serum IgM level showed a nonsignificant difference among both groups. Gul et al.  detected significantly decreased serum IgA levels in vitiligo patients, whereas IgG and IgM were in the normal range. The discrepancy in Ig profile levels among the different studies could be attributed to several factors, including the different numbers of patients in each study, the varying degree of disease activity among the studied population, and whether or not patients in the study had received any kind of immunomodulatory therapy. Also, racial or ethnic differences in the studied populations might be a contributing factor, where the baseline level of different Igs might be variable. Furthermore, different autoantigens are probably involved in the pathogenesis of vitiligo and, accordingly, the changes in Ig levels might not be consistent.
In the present study, we found a significant increase in the serum level of C3 in patients compared with controls (P=0.026), but no significant difference in the C4 serum level among both groups (P=0.278) was found. This finding supports the role of complement-mediated lysis of melanocytes in the pathogenesis of vitiligo. Singh et al.  found a significant decrease in C3 and C4 levels in patients compared with controls. Similarly, a study carried out on a sample of Iranian patients with vitiligo reported significantly decreased serum C3 and C4 levels in 25.5% of patients . However, Gul et al.  detected no significant difference in the serum levels of C3 or C4 among patients and controls. These results, given their diversity, support the findings of an increased frequency of heterozygous genes encoding the complement system in vitiligo patients, leading to abnormalities in the complement system .
Furthermore, we found a significant positive correlation between serum IgA level and the age of the patients (r=0.388, P=0.006). Similar to our results, Beharka et al.  found that IgA levels in the serum increased significantly with age. The immunological basis of the age-dependent changes in IgA levels could be explained partly by the age-related changes in cytokine production. T-helper 2 secretions, especially IL-5, are responsible for IgA production .
In terms of serum C3 and C4 levels, there was a statistically significant positive correlation with family history (r=0.552, P=0.005 and r=0.515, P=0.01, respectively). As mentioned before, this might be because of the increased frequency of heterozygous genes among family members, encoding complement system abnormalities . We also found a significant positive correlation between the serum levels of C3 and C4 among patients (r=0.644, P=0.001), indicating that they are both involved in the pathogenesis of vitiligo by the activation of anti-melanocyte antibodies.
| Conclusion|| |
Autoantibodies are clearly involved in the pathogenesis of vitiligo, but the exact mechanism of this involvement has yet to be fully understood. Changes in Ig levels can either be a cause of vitiligo or an effect of the disease itself. These findings should direct our attention toward considering altered immunity during the treatment of vitiligo patients. Further studies on a larger number of patients are recommended to establish the relation between serum Igs and complement levels in the pathogenesis of vitiligo, with possible determination of the target antigens; this might direct us towards more specific therapeutic modalities for vitiligo.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Alkhateeb A, Fain PR, Thody A, Bennett DC, Spritz RA. Epidemiology of vitiligo and associated autoimmune diseases in Caucasian probands and their families. Pigment Cell Res 2003; 16:208–214.
Schallreuter KU, Lemke R, Brandt O. Vitiligo and other diseases: coexistence or true association? Dermatology 1994; 188:269–275.
Yang Y, Lin X, Fu W, Luo X, Kang K. An approach to the correlation between vitiligo and autoimmune thyroiditis in Chinese children. Clin Exp Dermatol 2009; 23:121–125.
Forschener T, Buchholtz S, Stockfleth E. Current state of vitiligo therapy-evidence-based analysis of the literature. J Dtsch Dermatol Ges 2007; 5:467–475.
Song YH, Connor E, Li Y. The role of tyrosinase in autoimmune vitiligo. Lancet 1994; 344:1049–1052.
Kemp EH, Waterman EA, Gawkrodger DJ, Watson PF, Weetman AP. Autoantibodies to tyrosinase-related protein-1 detected in the sera of vitiligo patients using a quantitative radiobinding assay. Br J Dermatol 1998; 139:798–805.
Okamoto T, Irie RF, Fujii S, Huang SK. Anti-tyrosinase-related protein-2 immune response in vitiligo patients and melanoma patients receiving active-specific immunotherapy. J Invest Dermatol 1998; 111:1034–1039.
Li YL, Yu CL, Yu HS. IgG anti-melanocyte antibodies purified from patients with active vitiligo induce HLA-DR and intercellular adhesion molecule-1 expression and an increase in interleukin-8 release by melanocytes. J Invest Dermatol 2000; 115:969–973.
Ruiz-Argüelles A, Brito GJ, Reyes-Izquierdo P, Pérez-Romano B, Sánchez-Sosa S. Apoptosis of melanocytes in vitiligo results from antibody penetration. J Autoimmun 2007; 29:281–286.
Venneker GT, Westerhof W, De Vries IJ. Molecular heterogeneity of the fourth component of complement (C4) and its genes in vitiligo. J Invest Dermatol 1992; 99:853–858.
Onay H, Pehlivan M, Alper S. Might there be a link between mannose binding lectin and vitiligo? Eur J Dermatol 2007; 17:146–148.
Venneker GT, Vodegel RM, Okada N, Westerhof W, Bos JD, Ashgar SS. Relative contributions of decay acceleration factor (DAF), membrane cofactor protein (MCP) and CD59 in the protection of melanocytes from homologous complement. Immunobiology 1998; 198:476–484.
Van den Wijngaard RMJGJ, Wankowicz-Kalinska A, Pals S, Weening J, Das PK. Autoimmune melanocyte destruction in vitiligo. Lab Invest 2001; 81:1061–1067.
Njoo MD, Westerhoff W, Bos JD, Bossuyt PM. The development of guidelines for the treatment of vitiligo. Arch Dermatol 1999; 135:407–413.
Wolff K, Goldsmith LA, Katz SI, Gilchrest BA, Paller AS, Leffell DJ. Fitzpatrick’s dermatology in general medicine. 7th ed. New York, NY: McGraw-Hill; 2007. 616–621.
Halder RM, Taliaferro SJ. Vitiligo. In Wolff K, Goldsmith L, Katz S, Gilchrest B, Paller A, Lefell D, eds. Fitzpatrick’s dermatology in general medicine. New York, NY: McGraw-Hill; 2008. 72.
Pichler R, Sfetsos K, Badics B, Gutenbrunner S, Berg J, Auböck J. Lymphocyte imbalance in vitiligo patients indicated by elevated CD4+/CD8+ T-cell ratio. Wien Med Wochenschr 2009; 159:337–341.
Abu Tahir M, Pramod K, Ansari SH, Ali J. Current remedies for vitiligo. Autoimmun Rev 2010; 9:516–520.
Ali R, Ahsan MS, Azad MA, Ullah MA, Bari W, Islam SN et al.
Immunoglobulin levels of vitiligo patients. Pak J Pharm Sci 2010; 23:97–102.
Singh U, Singh S, Pandey SS. Study of serum immunoglobulin levels: complements C3 and C4 in vitiligo patients in northern India. In Sabu A, Augustine A, eds. Prospects in Bioscience: Addressing the Issues. India: Springer; 2012. 111–115.
Gul U, Soylu S, Demiriz M. Colocalization of lichen planus and vitiligo associated with selective IgA deficiency. Skin Med Dermatol Clin 2007; 6:202–203.
Farsangi MH, Farokhi SA, Nouhpisheh MK, Tahmasbi R. The role of humoral immune system in the pathogenesis of vitiligo. Iran S Med J 2003; 5:124–128.
Beharka A, Paiva S, Leka SL, Ribaya-Mercado DJ, Robert M, Russell RM, Meydani SN. Effect of age on the gastrointestinal-associated mucosal immune response of humans. J Gerontol A Biol Sci Med Sci 2001; 56:218–223.
Jafarzadeh A, Sadeghi M, Karam GA, Vazirinejad R. Salivary IgA and IgE levels in healthy subjects: relation to age and gender. Braz Oral Res 2010; 24:21–27.
[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2]