Histopathology and Molecular Pathology of Vitiligo

*Amit Kumar Yadav*

## **Abstract**

Vitiligo is a common skin disorder that manifests as whitish macules. There is no special geographic or sex predilection. Vitiligo is a multifactorial disorder. The various theories proposed include neutral theory, autoimmune theory, zincα2-glycoprotein theory, viral infection, intrinsic theory and melanocytorrhagy theory. However, the currently favored opinion is that there is a convergence of various theories known as the convergence theory. The basic defect is the absence of functional melanocytes from the epidermal melanin unit. This absence can be demonstrated by using special stains like Fontana-Masson, immunohistochemistry like HMB-45 and Melan-A and electron microscopy. Margins of lesions especially early lesions show inflammatory cells principally CD4+ and CD8+ T cells. The cornerstone of management in vitiligo is correct categorization of a case into stable and unstable vitiligo. This distinction is based mainly on clinical criteria. It is recommended that while evaluating biopsies, histopathological examination should be primarily concentrated on evaluating five histopathological variables—spongiosis, epidermal lymphocytes, basal cell vacuolation, dermal lymphocytes and melanophages. These parameters are then scored using a scoring system, and the recommended diagnoses based on these scores are given. Adoption of a systematic reporting system brings more consistency and objectivity in the diagnosis.

**Keywords:** vitiligo, multifactorial, convergence theory, melanocytes, histological scoring

#### **1. Introduction**

Vitiligo is a common acquired, idiopathic, progressive disorder which is characterized by the development of depigmented milky white macules of variable sizes. These often enlarge and coalesce to form extensive areas of leukoderma [1–3]. It equally affects both sexes with a worldwide prevalence of 0.1–2% [4]. It is a psychologically devastating and frequently resistant to treatment [5, 6]. The basic defect in vitiligo is a selective destruction of functional melanocytes [7].

The role of histopathology in the diagnosis of vitiligo is not yet fully established. So much so that routinely in these cases biopsy is not performed. The diagnosis is made primarily on clinical grounds.

#### **2. Pathogenesis**

Vitiligo is a multifactorial disorder [8, 9]. In its genesis both genetic and nongenetic factors are believed to play a role. It is observed that clinically no two patients of vitiligo are alike. This suggests that etiology also varies among different patients. Due to the observed variation in clinical manifestations of the disease, it seems likely that etiology of vitiligo may differ among patients [10]. These several theories have been combined into the convergence theory [11] which is currently the most accepted theory.

Briefly in the earliest theory, it was proposed by Lerner that vitiligo was neural in origin [12]. This theory could explain the segmental form of vitiligo which follows dermatomal distribution and is associated with hyperhidrosis and emotional disturbances. In another study the role of sympathetic nervous system in vitiligo was studied [13]. It was observed that the cutaneous blood flow in the lesional skin was three times higher than the normal skin in cases of segmental vitiligo. However, in other cases of non-segmental vitiligo, this was not observed.

Studies on the expression of neural proteins like neuropeptide Y (NPY), calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP) and polyclonal general neuronal marker (PGP) have shown variable results. In one study NPY expression was found to be increased in cases of segmental vitiligo [14]. It is proposed that precipitating factors like stress lead to increased NPY expression [15].

But this theory failed to explain the other forms of vitiligo. For that matter generalized or non-segmental vitiligo is better explained by autoimmune hypothesis. In previously done studies, antibodies against various targets like tyrosine hydroxylase, melanin-concentrating hormone receptor-1 (MCHR1), tyrosinase [16] and pigment cell surface antigens [17] have been demonstrated. In a study carried out to evaluate the various immunoglobulins, it was observed that 80% of active vitiligo patients showed the presence of IgG and IgM against melanocytes [17]. Other studies have shown the presence of anti-thyroglobulin antibodies, antithyroid antibodies, anti-thyroperoxidase and anti-smooth muscle antibody in these cases [18, 19].

Besides humoral immunity, cell-mediated immunity may also play an important role. Immunohistochemical examination of perilesional skin in vitiligo patients showed increased CD8:CD4 ratio and HLA-DR production along suprabasal and basal keratinocytes. Macrophages were found to be quite numerous [20]. However, not only the immune cells and antibodies but expression of various cytokines is also increased. Chief among these which have been studied are tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ) and IL-10 [21, 22]. IL-17 has also been shown to be significantly increased in cases of vitiligo [23].

The redox (reduction–oxidation) state of vitiliginous patients has been studied by many authors. These studies have shown increased serum levels of selenium, superoxide dismutase (SOD) and malondialdehyde (MDA) [24, 25]. Increased levels of these substances indicate the presence of oxidative stress in vitiligo. Increased levels of tetrahydrobiopterin [26] and xanthine oxidase [27] leading to increased levels of H2O2 may also be contributory.

Few authors have pointed towards the role of zinc-α2-glycoprotein (ZAG) in the pathogenesis of vitiligo. They hypothesize that lack of ZAG causes impaired melanocytic adhesion to other cells in the epidermis [28, 29]. The efficacy of zinc in the treatment of vitiligo may be due to its ability to precipitate ZAG at the site of vitiligo [30].

The role of viral infection in vitiligo has been proposed by certain authors. The potential candidates include hepatitis C virus (HCV) [31], cytomegalovirus (CMV) [32], Epstein–Barr virus (EBV), hepatitis e virus, herpes virus and HIV [33]. However, the evidence available is scant and not conclusive enough to attribute a significant role for viral agents in vitiligo.

**23**

**3. Histopathology**

**Figure 1.**

*Pathogenesis of vitiligo.*

*Histopathology and Molecular Pathology of Vitiligo DOI: http://dx.doi.org/10.5772/intechopen.84258*

The intrinsic theory states that in vitiligo there is loss of melanocytes due to various abnormalities which lead to increased apoptosis [34] and accelerated cell senescence [35, 36]. Studies done previously have shown various abnormalities in the melanocytes including cytoplasmic vacuolization, DNA marginalization, dendrite loss and detachment [36, 37]. The evidence in favor of increased apoptosis in vitiligo includes reduced expression levels of the antiapoptotic proteins Bcl-2 and FLIP in vitiliginous skin as compared to normal skin [34]. On the other hand, marked increase in the expression of proapoptotic factors such as Bax and p53 along

The melanocytorrhagy theory states that in vitiligo there is chronic melanocyte detachment and loss caused by trauma and other stressors which include catechol-

However, the consensus opinion of majority of experts is that vitiligo occurs due to convergence of these various pathways [39]. These are also depicted in (**Figure 1**). The author also is in agreement with this view; however, it is likely that in various subtypes of vitiligo the relative contribution of these pathways may vary. For example, in segmental vitiligo the neural theory may be more relevant than the

In order to understand the histopathology of vitiligo, it is essential to first understand the concept of *epidermal melanin unit* [40]. Melanocytes are neural crest derivatives, and they reach their final destination of basal layer of the epidermis and hair follicles via a process of migration. Each melanocyte then transfers its melanosomes to approximately 36 keratinocytes via a unique mechanism known as the shedding vesicle system. In the normal skin in the basal layer of the epidermis for every five basal keratinocytes, there is a presence of a single melanocyte [41]. The basic histopathological finding in vitiligo is the absence of functional melanocytes in the basal layer of the epidermis (**Figure 2**) [42–44]. This absence can also be demonstrated by using special stains like Fontana-Masson (**Figure 3**) [45].

other theories, whereas the same may not hold true for vitiligo vulgaris.

with the various caspases has also been observed [34].

amines, free radicals or autoimmune elements [38].

#### *Histopathology and Molecular Pathology of Vitiligo DOI: http://dx.doi.org/10.5772/intechopen.84258*

**Figure 1.** *Pathogenesis of vitiligo.*

*Depigmentation*

**2. Pathogenesis**

the most accepted theory.

Vitiligo is a multifactorial disorder [8, 9]. In its genesis both genetic and nongenetic factors are believed to play a role. It is observed that clinically no two patients of vitiligo are alike. This suggests that etiology also varies among different patients. Due to the observed variation in clinical manifestations of the disease, it seems likely that etiology of vitiligo may differ among patients [10]. These several theories have been combined into the convergence theory [11] which is currently

Briefly in the earliest theory, it was proposed by Lerner that vitiligo was neural in origin [12]. This theory could explain the segmental form of vitiligo which follows dermatomal distribution and is associated with hyperhidrosis and emotional disturbances. In another study the role of sympathetic nervous system in vitiligo was studied [13]. It was observed that the cutaneous blood flow in the lesional skin was three times higher than the normal skin in cases of segmental vitiligo. However,

Studies on the expression of neural proteins like neuropeptide Y (NPY), calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP) and polyclonal general neuronal marker (PGP) have shown variable results. In one study NPY expression was found to be increased in cases of segmental vitiligo [14]. It is proposed that precipitating factors like stress lead to increased NPY expression [15]. But this theory failed to explain the other forms of vitiligo. For that matter generalized or non-segmental vitiligo is better explained by autoimmune hypothesis. In previously done studies, antibodies against various targets like tyrosine hydroxylase, melanin-concentrating hormone receptor-1 (MCHR1), tyrosinase [16] and pigment cell surface antigens [17] have been demonstrated. In a study carried out to evaluate the various immunoglobulins, it was observed that 80% of active vitiligo patients showed the presence of IgG and IgM against melanocytes [17]. Other studies have shown the presence of anti-thyroglobulin antibodies, antithyroid antibodies, anti-thyroperoxidase and anti-smooth muscle antibody in these cases [18, 19]. Besides humoral immunity, cell-mediated immunity may also play an important

role. Immunohistochemical examination of perilesional skin in vitiligo patients showed increased CD8:CD4 ratio and HLA-DR production along suprabasal and basal keratinocytes. Macrophages were found to be quite numerous [20]. However, not only the immune cells and antibodies but expression of various cytokines is also increased. Chief among these which have been studied are tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ) and IL-10 [21, 22]. IL-17 has also been shown to be

The redox (reduction–oxidation) state of vitiliginous patients has been studied by many authors. These studies have shown increased serum levels of selenium, superoxide dismutase (SOD) and malondialdehyde (MDA) [24, 25]. Increased levels of these substances indicate the presence of oxidative stress in vitiligo. Increased levels of tetrahydrobiopterin [26] and xanthine oxidase [27] leading to increased

Few authors have pointed towards the role of zinc-α2-glycoprotein (ZAG) in the pathogenesis of vitiligo. They hypothesize that lack of ZAG causes impaired melanocytic adhesion to other cells in the epidermis [28, 29]. The efficacy of zinc in the treatment of vitiligo may be due to its ability to precipitate ZAG at the site of vitiligo [30]. The role of viral infection in vitiligo has been proposed by certain authors. The potential candidates include hepatitis C virus (HCV) [31], cytomegalovirus (CMV) [32], Epstein–Barr virus (EBV), hepatitis e virus, herpes virus and HIV [33]. However, the evidence available is scant and not conclusive enough to attribute a

significantly increased in cases of vitiligo [23].

levels of H2O2 may also be contributory.

significant role for viral agents in vitiligo.

in other cases of non-segmental vitiligo, this was not observed.

**22**

The intrinsic theory states that in vitiligo there is loss of melanocytes due to various abnormalities which lead to increased apoptosis [34] and accelerated cell senescence [35, 36]. Studies done previously have shown various abnormalities in the melanocytes including cytoplasmic vacuolization, DNA marginalization, dendrite loss and detachment [36, 37]. The evidence in favor of increased apoptosis in vitiligo includes reduced expression levels of the antiapoptotic proteins Bcl-2 and FLIP in vitiliginous skin as compared to normal skin [34]. On the other hand, marked increase in the expression of proapoptotic factors such as Bax and p53 along with the various caspases has also been observed [34].

The melanocytorrhagy theory states that in vitiligo there is chronic melanocyte detachment and loss caused by trauma and other stressors which include catecholamines, free radicals or autoimmune elements [38].

However, the consensus opinion of majority of experts is that vitiligo occurs due to convergence of these various pathways [39]. These are also depicted in (**Figure 1**). The author also is in agreement with this view; however, it is likely that in various subtypes of vitiligo the relative contribution of these pathways may vary. For example, in segmental vitiligo the neural theory may be more relevant than the other theories, whereas the same may not hold true for vitiligo vulgaris.
