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 Table of Contents  
ORIGINAL ARTICLE
Year : 2020  |  Volume : 33  |  Issue : 3  |  Page : 1031-1035

The role of chemokine CXC-motif receptor 3 in vitiligo: a clinical and immunohistochemical study


1 Department of Dermatology, Andrology and STDs, Faculty of Medicine, Menoufia University, Menoufia, Egypt
2 Department of Pathology, Faculty of Medicine, Menoufia University, Menoufia, Egypt

Date of Submission06-Feb-2019
Date of Decision26-Mar-2019
Date of Acceptance30-Mar-2019
Date of Web Publication30-Sep-2020

Correspondence Address:
Noura N. N. Shehata
Shibin El-Kom, Menoufia
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/mmj.mmj_38_19

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  Abstract 


Objectives
The present study aimed at evaluating the immunohistochemical expression of CXCR3 in the involved skin in vitiligo patients and to correlate its evaluated expression with the available clinicopathological parameters in those patients.
Background
Vitiligo is an autoimmune skin disorder in which the loss of melanocytes is mainly attributed to defective autoimmune mechanisms, cytotoxic mechanisms, an intrinsic defect of melanocytes, oxidant–antioxidant mechanisms, and neural mechanisms. The assessment of maintenance of vitiligo could be related to CXCR3 expression. The treatment of vitiligo remains a challenge.
Patients and methods
This case–control study was carried out on 35 patients with variable degrees of vitiligo severity. They were recruited from the Dermatology Outpatient Clinic, Faculty of Medicine, Menoufia University Hospital from May 2016 to March 2017. Normal skin biopsies were obtained from the control group which included 20 persons of age, sex, Fitzpatrick skin phototype, and occupation-matched healthy participants attending the Plastic Surgery Department.
Results
The results were collected, tabulated, and statistically analyzed by an IBM-compatible personal computer with SPSS statistical package, version 20 (SPSS Inc. Released 2011, IBM SPSS statistics for Windows, version 20.0). The relative expression of CXCR3 chemokine in vitiligo patients showed that the percent of CXCR3 expression in the epidermis was significantly higher in vitiligo cases (median, 80) than controls (median, 20) (P = 0.001). H score was significantly higher in vitiligo cases (median, 100) than controls (median, 30) (P = 0.003).
Conclusion
CXCR3 may have an active role in the pathogenesis of vitiligo.

Keywords: CXCR3, Fitzpatrick skin phototype, vitiligo


How to cite this article:
Farag AG, Al-Sharaky DR, Hassan RA, Elshafey EN, Shehata NN. The role of chemokine CXC-motif receptor 3 in vitiligo: a clinical and immunohistochemical study. Menoufia Med J 2020;33:1031-5

How to cite this URL:
Farag AG, Al-Sharaky DR, Hassan RA, Elshafey EN, Shehata NN. The role of chemokine CXC-motif receptor 3 in vitiligo: a clinical and immunohistochemical study. Menoufia Med J [serial online] 2020 [cited 2020 Oct 22];33:1031-5. Available from: http://www.mmj.eg.net/text.asp?2020/33/3/1031/296677




  Introduction Top


Vitiligo is a common, noncontagious disorder, characterized by progressive patchy loss of skin pigmentation. It affects 0.1–2% of the world population with no sexual or racial preference and appears at any age, but has been most frequently observed in the first two decades, with a significant decrease in the quality of life of affected cases [1].

Vitiligo is a multifactorial polygenic disorder with a complex pathogenesis. Several theories have been proposed about the pathogenesis of vitiligo including autoimmune mechanisms [2]. However, the precise cause behind the destruction of melanocytes remains unknown [3].

The chemokine receptor CXCR3 is a G protein-coupled receptor found predominantly on T cells that are activated by three ligands as follows: CXCL9 (Mig), CXCL10 (IP-10), and CXCL11 (I-TAC) [4].

CXC-motif chemokine receptor 3 (CXCR3) is linked to the Th1 pattern and have been suggested as one of the most relevant chemokine axes that promote T-cell migration in different autoimmune and inflammatory processes [5].

CXCR3 is increased in a number of autoimmune diseases including autoimmune thyroiditis, alopecia areata, and rheumatoid arthritis [6].

CXCR3 expression on T cells require activation by triggering a nonpersistent T-cell receptor in the presence of interferon gamma (IFNγ) [7]. IFNγ-induced CXCL10-CXCR3 chemokine pathway plays a key role in CD8 T-cell skin accumulation and melanocyte destruction [8],[9]. CD8+ T-cell participates in melanocyte-specific autoimmune response, leading to depigmentation of the skin.

A significantly enhanced number of CXCR3-expressing cells in vitiligo lesions (n = 80) was reported compared with healthy controls (n = 40) [10]. Moreover, it was found that the mRNA expression of CXCR3 was significantly increased in both vitiligo (n = 10) and halo nevus (n = 10) compared with healthy controls (n = 10) [11].

The current study aimed at evaluating the immunohistochemical expression of CXCR3 in the involved skin in vitiligo patients and to correlate its evaluated expression with the available clinicopathological parameters in those patients.


  Patients and Methods Top


This prospective case–control study was carried out on 35 patients with variable degrees of vitiligo severity, which is calculated by the study sample equation:



They were recruited from the Dermatology Outpatient Clinic, Faculty of Medicine, Menoufia University Hospital from May 2016 to March 2017 (group 1). Additionally, 20 persons of age, sex, Fitzpatrick skin phototype, and occupation-matched healthy participants attending the Plastic Surgery Department were included as controls (group 2). A written consent form approved by the Committee of Human Rights in Research in Menoufia University was obtained from every participant before the study initiation.

Inclusion and exclusions criteria

Vitiligo patients from both sex without treatment of their vitiligo in the last 1 month were included. However, those who are under treatment were excluded from the study.

Each of the vitiligo patients was subjected to full history taking and clinical examination to identify any excluding factor. The dermatological examination was performed including the vitiligo area severity index score to assess the severity of vitiligo [12].

Skin biopsies

Punch skin biopsies from the involved skin of vitiligo patients and normal healthy skin of controls were taken under local anesthesia. Each specimen was submitted to routine tissue processing to be embedded in paraffin blocks. For each specimen, two sections of 4 μm thickness were cut.

Histopathological evaluation

One section was cut for hematoxylin and eosin staining to be examined by light microscope and to assess the pathological changes.

Immunohistochemical staining

Another section was cut on poly-L-lysine coated slides for immunohistochemical staining using purified rabbit polyclonal antibody (Cat. # YPA1366) raised against CXCR3 which was received as concentrated 0.1 ml from Abcam Company (http://www.abcam.com). The procedure of immunohistochemical staining was done according to the received datasheet of the used antibodies.

Immunohistochemically, CXCR3 expression is confirmed by cytoplasmic and/or membranous staining according to the supplier's datasheet and was evaluated in both the epidermis and dermis. In case of positively expressed cells, the percentage of the positive cell was assessed at a ×200 magnification field [13]. The intensity of the stain was graded as mild, moderate, or strong. Histo-score (H score) was calculated (H score = 1×% of mildly stained cells +2×% moderately stained cells +3×% of strongly stained cells) [14]. CXCR3 stain distribution patterns were categorized according to the layers of the epidermis.

Ethical consideration

The study was approved by the Ethics Committee of Menoufia Faculty of Medicine and informed consent was obtained from all participants before the start of the study.

Statistical analysis of data

Data were collected, tabulated, and statistically analyzed using a personal computer with the Statistical Package for Social Sciences (SPSS), version 20 (IBM Corp., Armonk, New York, USA).

Descriptive statistics were expressed as percentage (%), mean, and SD. Analytic statistics included χ2 to compare between qualitative data; Fisher's exact test was used in the analysis of 2 × 2 contingency tables when at least 25% of cells have an expected number of less than 5; t test was used in comparison between two normally distributed groups; Mann–Whitney (U test) was used in comparison between two non-normally distributed groups. The Wilcoxon signed-rank test was used when comparing two related samples, matched samples, or repeated measurements on a single sample to assess whether their population means ranks differ. McNemar's test of significance was used on paired (e.g. pre and post) qualitative data. Marginal homogeneity test of significance was used on paired (e.g. pre and post) qualitative data when a category of the sample is more than two. Kruskal–Wallis test (K) was used for comparison between more than two not normally distributed groups. Spearman's correlation (r) was used to measure the association between two not normally distributed quantitative variables or one quantitative and one qualitative ordinal variable. A P value of less than 0.05 was considered statistically significant.


  Results Top


Personal and clinical data of the studied groups: This study included 35 patients with vitiligo, 17 (48.6%) men and 18 (51.4%) women; their age ranged from 15 to 50 years with a mean ± SD of 35.4 ± 11.3 years and a median value of 35 years. Regarding Fitzpatrick skin phototype: 10 (28.6%) cases were of type II, 15 (42.9%) cases were of type III, and 10 (28.6%) cases were of type IV. Additionally, 16 (45.7%) cases had outdoor occupations and 19 (54.3%) cases had indoor ones.

The control group included 20 apparently healthy volunteers, seven (35%) men and 13 (65%) women, their age ranged from 15 to 54 years with a mean ± SD of 33.2 ± 12.4 years and a median value of 33 years. Regarding Fitzpatrick skin phototype: five (25%) patients were of type II, eight (40%) patients were of type III, and the other seven (35%) patients were of type IV. Additionally, 11 (55%) patients had outdoor occupations and nine (45%) patients had indoor occupations.

There were insignificant differences between vitiligo patients and controls regarding their age, sex, skin phototypes, and occupation (P > 0.05 for all) [Table 1].
Table 1: Comparison between the studied groups regarding personal data

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CXCR3 immunohistochemical profile in the control group: however, the normal skin of controls showed that CXCR3 expression was positive in 17 (85%) normal control sections. Regarding CXCR3 intensity, 13 (76.5%) sections were mild and four (23.5%) were moderate. The percent of epidermal CXCR3 immunoreactivity ranged from 0 to 40 with a mean ± SD of 22.7 ± 16.1 and a median value of 20. H score ranged from 5 to 30 with a mean ± SD of 32.9 ± 25.6 and median value of 30 [Figure 1].
Figure 1: CXCR3 immunostaining in normal skin showing mild cytoplasmic expression of CXCR3 in the epidermis mainly confined to the stratum basalis (red arrow) (immunoperoxidase ×400).

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CXCR3 immunohistochemical staining in the studied group: the investigated baseline of epidermal CXCR3 immunohistochemical staining in vitiligo patients showed that CXCR3 expression was positive in 33 (94.3%) patients. The intensity of CXCR3 immunoreactivity was mild in nine (27.3%) cases, moderate in 14 (42.4%) cases, and strong in 10 (30.3) cases. The percent of epidermal CXCR3 immunoreactivity ranged from 10 to 90 with a mean ± SD of 61.2 ± 26.1 and a median of 80. H score ranged from 10 to 240 with a mean ± SD of 91.4 ± 57.3 and a median of 100 [Figure 2].
Figure 2: CXCR3 immunostaining in vitiliginous skin showing strong diffuse (yellow brackets) membranous expression (immunoperoxidase ×400).

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Comparison between CXCR3 immunohistochemical staining in studied cases and controls: the investigated epidermal CXCR3 immunohistochemical staining showed that the percent of CXCR3 expression in the epidermis was significantly higher in vitiligo cases (median, 80) than controls (median, 20) (P = 0.001). H score was significantly higher in vitiligo cases (median, 100) than controls (median, 30) (P = 0.003) [Table 2].
Table 2: Comparison between CXCR3 immunohistochemical staining in studied cases and controls

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  Discussion Top


CXCR3 is a chemokine receptor that is highly expressed on effector T cells and plays an important role in T-cell function. CXCR3 is activated by three IFN-inducible ligands CXCL9 (MIG), CXCL10 (IP-10), and CXCL11 (l-TAC) [4].

CXCR3 plays a role in the trafficking of Th1 and CD8 T cells to peripheral sites of Th1-type inflammation and the establishment of a Th1 amplification loop mediated by IFNγ and IFNγ-inducible CXCR3 ligands. CXCR3 plays a role in the migration of T cells in the microenvironment of the peripheral tissue and lymphoid compartment, facilitating the interaction of T cells with the antigen presenting cells leading to the generation of effector and memory cells [15].

Gregg et al. [8] and Harris et al. [9] found that the IFNγ-induced CXCL10-CXCR3 chemokine pathway plays a key role in CD8+ T-cell skin accumulation and melanocyte destruction. CD8+ T-cell participates in melanocyte-specific autoimmune response, leading to depigmentation of the skin.

Confirming these findings, the present study demonstrated that CXCR3 immunoreactivity was observed in 85% of examined sections of the control group.

Herein, we observed a significant difference between vitiligo patients and control group regarding CXCR3 immunohistochemical staining. And the result of the current study showed a statistically significant up-regulation of the percent of epidermal expressions of CXCR3 expression and H score of epidermal CXCR3 expression in vitiligo cases than their matched peers. In agreement of this result, Wang and Xiang [10] showed a significantly enhanced number of CXCR3-expressing cells in vitiligo lesions (n = 80) compared with healthy controls (n = 40). Also, Yang et al. [11] reported that the mRNA expression of CXCR3 and its ligand CXCL10 were significantly increased in both vitiligo (n = 10) and halo nevus (n = 10) compared with healthy controls (n = 10). This difference could be attributed to the small sample size in our study.

Additionally, our results showed CXCR3 expression in 94.3% of vitiligo patients especially expression in the epidermis. In agreement with our results, Yang et al. [11] found significantly increased expressions of the chemokine receptor CXCR3 and its ligands, especially the accumulated CXCl10 in skin lesions of vitiligo and halo nevus. Moreover, Rashighi et al. [16] found that the levels of the chemokines CXCL9, CXCL10, and CXCL11, all of which are ligands of CXCR3, were significantly upregulated in vitiligo skin.


  Conclusion Top


CXCR3 may play a role in the development or maintenance of vitiligo.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Kumar CM, Sathisha UV, Dharmesh S, Rao AG, Singh SA. Interaction of sesamol (3,4- ethylenedioxyphenol) with tyrosinase and its effect on melanin synthesis. Biochimie 2011; 93w:562–569.  Back to cited text no. 1
    
2.
Farag AG, Haggag MM, Muharram NM, Mahfouz R, Elnaidany NF, Abd El Ghany HM. Is vitamin D a participant in narrow-band ultraviolet B-induced pigmentation in patients with vitiligo?. J Egypt Women's Dermatol Soc 2018; 15:30–34.  Back to cited text no. 2
    
3.
Gauthier Y, Cario Andre M, Taïeb A. A critical appraisal of vitiligo etiologic theories. Is melanocyte loss a melanocytorrhagy?. Pigment Cell Res 2003; 16:322–332.  Back to cited text no. 3
    
4.
Colvin RA, Campanella GS, Sun J, Luster AD. Intracellular domains of CXCR3 that mediate CXCL9, CXCL10, and CXCL11 function. J Biol Chem 2004; 279:30219–30227.  Back to cited text no. 4
    
5.
Lonsdorf AS, Hwang ST, Enk AH. Chemokine receptors in T-cell-mediated diseases of the skin. J Invest Dermatol 2009; 129:2552–2566.  Back to cited text no. 5
    
6.
Ruffilli I, Ferrari SM, Colaci M, Ferri C, Fallahi P, Antonelli A. IP-10 in autoimmune thyroiditis. Horm Metab Res 2014; 46:597–602.  Back to cited text no. 6
    
7.
Lacotte S, Brun S, Muller S, Dumortier H. CXCR3, inflammation. And autoimmune disease. Ann N Y Acad Sci 2009; 1173:310–137.  Back to cited text no. 7
    
8.
GGregg RK, Nichols L, Chen Y, Lu B, Engelhard VH. Mechanisms of spatial and temporal development of autoimmune vitiligo in tyrosinase-specific TCR transgenic mice. J Immunology 2010; 184:1909–17.  Back to cited text no. 8
    
9.
Harris JE, Harris TH, Weninger W, Wherry EJ, Hunter CA, Turka LA. A mouse model of vitiligo with focused epidermal depigmentation requires IFN-gamma for autoreactive CD8(+) T-cell accumulation in the skin. J Invest Dermatol 2012; 132:1869–1876.  Back to cited text no. 9
    
10.
Wang X, Xiang L. Increased expression of CXCR3 and its ligands in vitiligo patients and its role as potential clinical markers for vitiligo. J Dermatol Sci 2016; 84:e101.  Back to cited text no. 10
    
11.
Yang Y, Li S, Zhu G, Zhang Q, Wang G, Gao T, et al. A similar local immune and oxidative stress phenotype in vitiligo and halo nevus. J Dermatol Sci 2017; 87:50–59.  Back to cited text no. 11
    
12.
Hamzavi H, Jain D, McLean J, Shapiro H, Zeng, Lui H. Parametric modeling of narrowband UV-B phototherapy for vitiligo, using a novel quantitative tool: the vitiligo area scoring index. Arch Dermatol 2004; 140:677–683.  Back to cited text no. 12
    
13.
Bahnassy AA, Zekri AR, El-Houssini S, El-Shehaby AM, Mahmoud MR, Abdallah S, et al. Cyclin A and cyclin D1 as significant prognostic markers in colorectal cancer patients. BMC Gastroenterol 2004; 23:4–22.  Back to cited text no. 13
    
14.
Smyth JF, Gourley C, Walker G, Mackean MJ, Stevenson A, Williams AR, et al. Antiestrogen therapy is active in selected ovarian cancer: the use of letrozole in estrogen receptor-positive patients. Clin Cancer Res 2007; 13:3617–3622.  Back to cited text no. 14
    
15.
Groom JR, Luster AD. CXCR3 in T cell function. Exp Cell Res 2011; 317:620–631.  Back to cited text no. 15
    
16.
Rashighi M, Agarwal P, Richmond JM, Harris TH, Dresser K, Su MW, et al. CXCL10 is critical for the progression and maintenance of depigmentation in a mouse model of vitiligo. Sci Transl Med 2014; 6:223ra23.  Back to cited text no. 16
    


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