Home About us Editorial board Search Ahead of print Current issue Archives Submit article Instructions Subscribe Contacts Login 


 
 Table of Contents  
ORIGINAL ARTICLE
Year : 2022  |  Volume : 35  |  Issue : 4  |  Page : 1686-1691

Evaluation of aryl hydrocarbon receptor and cytochrome P450 1A1 gene expressions in patients with vitiligo


1 Department of Dermatology, Andrology and STDs, Faculty of Medicine, Menoufia University, Menoufia, Egypt
2 Department of Dermatology, Andrology and STDs, Shebin Elkom Dermatology and Leprosy Hospital, Shebin Elkom, Egypt
3 Department of Medical Biochemistry and Molecular Biology, Faculty of Medicine, Menoufia University, Menoufia, Egypt

Date of Submission14-Jun-2022
Date of Decision29-Jun-2022
Date of Acceptance03-Jul-2022
Date of Web Publication04-Mar-2023

Correspondence Address:
Sara M. S. Elnaggar
Shebin Elkom, Menoufia
Egypt
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/mmj.mmj_201_22

Rights and Permissions
  Abstract 


Background
Vitiligo represents an acquired depigmented skin disorder. Disruption of aryl hydrocarbon receptor (AhR) and cytochrome P450 1A1 (CYP1A1) is linked to a variety of skin disorders.
Objectives
To evaluate the expression levels of AhR and CYP1A1 in patients with vitiligo and to explore their correlation with disease activity.
Patients and methods
This is a case–control study that included 30 patients with vitiligo and 20 controls. All patients were assessed by vitiligo disease activity (VIDA) score for activity and vitiligo area severity index score for severity. A quantitative real-time PCR approach was used to assess AhR and CYP1A1 mRNA expressions.
Results
Patients with vitiligo displayed significantly decreased AhR and CYP1A1 expressions (P < 0.001) compared with the controls, with a significant positive correlation between AhR and CYP1A1 expressions (r = 0.782, P < 0.001). Moreover, there was a substantial reduction in the expression of AhR and CYP1A1 in progressive vitiligo cases compared with stationary illness cases (P < 0.001 and P = 0.003, respectively) according to the VIDA score. AhR and CYP1A1 had a significant negative connection with the VIDA score (AhR, r=−0.728, P < 0.001, and CYP1A1, r=−0.702, P < 0.001).
Conclusion
Lower mRNA levels of AhR and CYP1A1 might have an active role in vitiligo etiopathogenesis and could be used as predictive indicators for disease activity.

Keywords: aryl hydrocarbon receptor, cytochrome P450 1A1, PCR, vitiligo disease activity score, vitiligo


How to cite this article:
Hagag MM, Elnaggar SM, Habieb MS. Evaluation of aryl hydrocarbon receptor and cytochrome P450 1A1 gene expressions in patients with vitiligo. Menoufia Med J 2022;35:1686-91

How to cite this URL:
Hagag MM, Elnaggar SM, Habieb MS. Evaluation of aryl hydrocarbon receptor and cytochrome P450 1A1 gene expressions in patients with vitiligo. Menoufia Med J [serial online] 2022 [cited 2024 Mar 29];35:1686-91. Available from: http://www.mmj.eg.net/text.asp?2022/35/4/1686/370974




  Introduction Top


Vitiligo is an acquired idiopathic depigmenting skin disorder caused by the selective removal of functioning melanocytes in the epidermis, resulting in the formation of well-defined white macules and patches on the skin. Generally, vitiligo affects 0.5–2% of people [1].

Vitiligo is classified broadly into nonsegmental and segmental vitiligo. Several types of nonsegmental vitiligo have been described, including mucosal, acrofacial, generalized, and universal [2].

Genetic predisposition, environmental factors, metabolic changes, restricted renewal capability, and altered inflammatory as well as altered immunological responses all play a role in the loss of melanocytes [3].

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor found in all types of skin cells, especially melanocytes. It has the ability to maintain redox balance in cells. Its is also involved in mucosal barrier development, melanogenesis, and regulation of the immunological process [4]. Because AhR regulates oxidative stress and can alter melanocyte death, it is reported that AhR is linked to vitiligo activity [5]. It also responds to exogenous and endogenous substances by activating or suppressing genes with toxic or protective effects, such as cytochrome P450 1A1 (CYP1A1), in a variety of species and tissues [6]. It has been shown that AhR regulates the expression of phase I xenobiotic-metabolizing enzymes, such as CYP1A1 [7].

CYP1A1 is thought to be a major downregulator of AhR signaling in skin inflammation as CYP1A1 expression has been found to play a role in psoriasis and atopic dermatitis [8].

The aim of this study was to evaluate the expression levels of AhR and CYP1A1 in patients with vitiligo and to explore their correlation with disease activity.


  Patients and methods Top


This case–control study was conducted at the dermatology outpatient clinic and Biochemistry Department, Faculty of Medicine, Menoufia University. A total of 30 patients with various clinical forms of vitiligo (segmental and nonsegmental) were included in the study, as well as 20 apparently healthy age-matched and sex-matched controls. All of the participants gave their informed consent. The study was approved by the local ethics committee of Faculty of Medicine, Menoufia University. Patients with vitiligo under treatment, patients with systemic disease, patients with dermatological diseases other than vitiligo such as psoriasis, and patients with other inflammatory or autoimmune skin diseases were excluded.

The cases were subjected to history taking and a general examination to rule out any systemic disorders. A dermatologist examined the patient's skin, hair, nails, and oral mucosa to exclude any associated diseases. Examination of vitiligo included site, symmetry, extent, localization, clinical types, activity, and stability. It was determined how severe the disease was by looking at the vitiligo area severity index score [9]. Vitiligo activity was assessed if progressive or stable clinically using the vitiligo disease activity (VIDA) score [10]. A low VIDA score indicates less vitiligo activity and higher scores indicate more recent disease activity. VIDA is a six-point scale that measures disease activity. The following is the rubric for grading: VIDA score +4 indicates the activity lasting 6 weeks or less; VIDA score +3 indicates the activity lasting 6 weeks to 3 months; VIDA score 2 indicates the activity lasting 3–6 months; VIDA score 1 indicates the activity lasting 6–12 months; and VIDA score 0 indicates stability that lasts 1 year or more; and VIDA score −1 indicates stability with spontaneous repigmentation for 1 year or more.

Assessments of AhR and CYP1A1 mRNA expressions in the whole blood were done using a quantitative real-time PCR technique. Under strict aseptic circumstances, 2.5 ml of blood was collected from each individual and deposited in an EDTA-containing tube. The whole blood sample was processed immediately for RNA extraction. The quantitative real-time PCR procedure was carried out as follows: total RNA was extracted using a miRNeasy Mini kit (Qiagen, Hilden, Germany), according to the manufacturer's instructions. The miScript II RT kit (Qiagen), was used. The reverse transcription reactions were kept for the real-time PCR phase at −20°C. Amplification of cDNA was done using Hera plus SYBR Green PCR kit (Willow Fort, Birmingham, UK). In a total volume of 25 μl, the following reagents and volumes were added to each sample: 1 μl of forward and reverse primers, 12.5 μl of SYBR Green Master mix, 5 μl of cDNA template, and 5.5 μl of RNase- free water is one of components of the reaction master mix. The reaction mix was prepared and dispensed into the wells of the PCR plate. Caps were used to close the PCR plate tightly, and the real-time cycler (Applied Biosystem 7500, The Lab World Group , 15 Bonazzoli Ave, Unit 1 Hudson MA 01749 , USA) was programed as follows: initial denaturation step for 15 min at 95°C, 40 cycles of denaturation for 30 s at 94°C, annealing for 60 s at 60°C, extension for 60 s at 72°C, and final extension step for 3 min at 72°C.

The target levels of AhR and CYP1A1 mRNA were normalized to endogenous levels of β-actin, and relative quantification of expression was assessed using the 2ΔΔCt technique with Applied Biosystem 7500 software, version 2.0.1. Each run was completed after melting curve analysis was used to confirm amplification accuracy and the absence of primer dimer.

Version 20.0 of IBM SPSS software (IBM Corp., Armonk, New York, USA) was used to analyze the data. Categorical data were presented as frequency and percentage. Normality of quantitative data was tested by the Kolmogorov–Smirnov test using range (minimum and maximum), median, and interquartile range. The results were deemed significant at 5%. Significant outcomes are those with P values lower than 0.05. χ2 test was used to compare groups of categorical variables. Mann–Whitney U test was used to compare two groups with abnormally distributed quantitative variables. The abbreviation ROC stands for receiver operating characteristics. ROC curve is calculated by charting sensitivity (TP) versus 1-specificity (FP) at various cutoff values. The ROC curve's area indicates the test's diagnostic performance. Correlation of numeric data was done by Pearson's or Spearman correlation (r).


  Results Top


The vitiligo group was made up of 11 (36.7%) men and 19 (63.3%) females, with no statistically significant difference compared with the control group (P = 0.349). The median age of cases was 26 years, with no statistically significant difference compared with the control group (P = 0.263). A total of three (10%) cases of vitiligo had a positive family history. The median duration of vitiligo was 2 years. Generalized vitiligo was the most common (66.7%). In two (6.7%) cases, hair affection was evident, and in three (10%) cases, mucous membrane affection was observed. A total of five (16.7%) cases had a VIDA score of − 1, nine (30%) cases had a VIDA score of 0, seven (23.3%) cases had a VIDA score of +1, four (13.3%) cases had a VIDA score of +2, and five (16.7%) cases had a VIDA score of + 3. From the VIDA score, patients with progressive course accounted for 53.3% of the total, whereas the rest were stable (46.7%). The vitiligo area severity index score had a range between 0.1 and 9 [Table 1].
Table 1: Clinical data for the patient group

Click here to view


Vitiligo cases had substantially lower levels of AhR and CYP1A1 expression than the control people (P < 0.001) [Table 2]. There was a positive correlation between levels of AhR and CYP1A1 expression (r = 0.782, P < 0.001).
Table 2: Comparison of the expression levels of aryl hydrocarbon receptor and cytochrome P450 1A1 in the studied groups

Click here to view


According to the correlation analysis of AhR and CYP1A1 expression levels and VIDA score in patients with vitiligo, AhR and CYP1A1 had an inverse relationship with the VIDA score (AhR; r=−0.728, P < 0.001, CYP1A1; r=−0.702, P < 0.001) [Figure 1]. The level of AhR and CYP1A1 in vitiligo cases with progressive course was significantly lower than that in cases with stationary course (P < 0.001 and P = 0.003, respectively) [Table 3].
Figure 1: (a) Correlation between AhR and VIDA score in patients with vitiligo. (b) Correlation between CYP1A1 and VIDA score in patients with vitiligo. AhR, aryl hydrocarbon receptor; CYP1A1, cytochrome P450 1A1; VIDA, vitiligo disease activity.

Click here to view
Table 3: Relationship between aryl hydrocarbon receptor and cytochrome P450 1A1 expression levels and the progression of vitiligo

Click here to view


AhR and CYP1A1 have diagnostic and prognostic values used to distinguish between patients with vitiligo and controls, as well as between vitiligo cases with a progressive and those with a stationary course. In discriminating between patients with vitiligo and controls, AhR + CYP1A1 had a greater diagnostic accuracy than AhR alone or CYP1A1 alone [area under the curve (AUC) of 0.988, 0.955, and 0.943, respectively]. AhR at a cutoff value of 0.95 had an AUC of 0.871 with a sensitivity of 75%, CYP1A1 at a cutoff value of 1.44 had an AUC of 0.813 with a sensitivity of 81.25%, and AhR + CYP1A1 had an AUC of 0.866 with a sensitivity of 75% in distinguishing between patients with progressive and stationary vitiligo [Table 4].
Table 4: Validity of aryl hydrocarbon receptor and cytochrome P450 1A1 in distinguishing patients with vitiligo from controls, as well as in distinguishing progressive from stationary vitiligo

Click here to view



  Discussion Top


Currently, the AhR expression in patients with vitiligo is substantially reduced than that of the control group. Indeed, AhR has a high discriminative power to identify patients with vitiligo from controls. This was matched with earlier research studies by Schallreuter et al.[11] and Liu et al.[12]. Schallreuter et al.[11] discovered a significant decrease in AhR protein expression as well as proteins encoded by AhR target genes in patients with vitiligo. According to Liu et al.[12], the AhR transcript levels in the vitiligo group were significantly lower than those in the healthy controls.

It is worth noting that AhR is necessary for melanocyte proliferation, differentiation, and melanogenesis control [13]. As a result, a decrease in its level is connected to vitiligo development and faulty melanogenesis [14]. Because AhR drives skin pigmentation, it has been recommended as a potential vitiligo cure [15].

AhR plays a key role in the induction of immunoregulatory factors. As a result of this, it is not surprising that AhR influences autoimmune disorders such as vitiligo [16]. AhR is related to controlling the physiological process of T helper, implicating it in immunity and inflammation. As a result, lack of AhR levels or changes in AhR activity may cause the immune response to be disrupted [17].

Furthermore, Rekik et al.[14] discovered a significant decrease in AhR transcription in the skin of patients with vitiligo, which could be a vitiligo susceptibility factor. However, compared with nonlesional skin, AhR mRNA expression was considerably higher in the lesional skin of patients with vitiligo. They related AhR transcription level to inflammatory cytokine production, which is dependent on illness status. Therefore, this could be a feedback response to combat inflammation.

In contrast, other studies found higher levels of AhR in patients with vitiligo, such as Esser et al.[18] and Behfarjam and Jadali [19]. The discrepancy reported in the study by Behfarjam and Jadali could be linked to the fact that they examined patients with inactive vitiligo. Additionally, this disagreement over AhR expression stems from the fact that AhR is a ligand-activated transcription factor that connects environmental cues to immunological responses. There are several factors that influence the immune system's response to these ligand–receptor interactions. These include the cell type and activation status of cells, the agonistic or antagonistic properties of ligands, T-cell development, and the cytokine milieu [18],[20].

The present study discovered that CYP1A1 expression in patients with vitiligo was significantly lower than that seen in the control group and that the levels of AhR and CYP1A1 expression were positively correlated.

This was in line with the findings of Schallreuter et al.[11], who found that patients with vitiligo have significantly lower CYP1A1 expression than controls and stated that lower epidermal AhR expression in vitiligo in the presence of H2O2 has been linked to reduced COX-2 and CYP1A1 expression. H2O2 affects cell metabolism, resulting in the expression of a variety of cytokines and inflammatory mediators causing vitiligo. Liu et al.[12] also investigated the expression of CYP1A1, which is known to be regulated by AhR. They discovered that low AhR mRNA expression was linked to lower CYP1A1 mRNA expression in the CD4+ T cells of the cases when they compared them to healthy controls.

As discovered by Kyoreva et al.[8], the AhR/CYP1A1 axis is involved in other inflammatory skin diseases such as psoriasis. Reduced CYP1A1 expression in patients with psoriasis suggests a general deregulation of the AhR pathway in this disease, which could decrease the pathway's anti-inflammatory efficacy.

CYP1A1 expression was found to be upregulated in the skin of patients with atopic dermatitis[21] and in the peripheral blood mononuclear cells of patients with systemic lupus erythematosus, suggesting that upregulation occurs during inflammation and autoimmunity is to limit immunopathology [22].

This study also looked into the link between AhR and CYP1A1 levels and vitiligo progression. VIDA score was found to have a significant negative relationship with AhR and CYP1A1. The results indicated that patients with vitiligo with a progressive course had lower levels of AhR and CYP1A1 than those with a stationary course. Additionally, CYP1A1 demonstrated a higher discriminative power between progressive and stationary cases, with a sensitivity of 81.25%. As a result, the AhR/CYP1A1 axis could be employed as a prognostic factor in determining the severity of vitiligo. Similarly, when Liu et al.[12] discovered that lower expression level of AhR is related to unstable progressive vitiligo, which is linked to the increased level of IL17A. Additionally, it is indicated that IL17A is involved in disease progression, as there is a positive link between its level and area of vitiligo lesions [23].

The association of AhR with vitiligo pathology has a physiological basis, as the skin is vulnerable to exogenous and endogenous AhR ligands that regulate the AhR signaling pathways. There is also a close link between AhR and immune regulation, skin homeostasis, pigmentation, and antioxidant activity [24]. Oxidative stress is considered a critical factor in the development and progression of vitiligo, which damages melanocytes. Because AhR regulates oxidative stress and can alter melanocyte death, it is thought that AhR is linked to the start and progression of vitiligo [25]. The data presented by Liu et al.[12] demonstrate that activation of AhR regulates the production of IL17A and IL22 in CD4 + T cells from patients with progressive unstable vitiligo, indicating the potential therapeutic role of AhR in treating vitiligo.


  Conclusion Top


This is the first case–control study showing that lower levels of the mRNA of AhR and CYP1A1 might have an active role in vitiligo etiopathogenesis. Furthermore, significantly lower levels of AhR and CYP1A1 were seen in patients with a progressive course, so they are biomarkers for vitiligo activity and severity. The AhR/CYP1A1 axis can be employed as a prognostic factor in patients with vitiligo. Study limitations include small sample size, so more studies are needed with a larger number of patients to explore more the role of AhR and CYP1A1 in vitiligo progression and their possible application in treatment strategies.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Li D, Liang G, Calderone R, Bellanti JA. Vitiligo and Hashimoto's thyroiditis: autoimmune diseases linked by clinical presentation, biochemical commonality, and autoimmune/oxidative stress-mediated toxicity pathogenesis. Med Hypotheses 2019; 128:69–75.  Back to cited text no. 1
    
2.
Ezzedine K, Eleftheriadou V, Whitton M, van Geel N. Vitiligo. Lancet 2015; 386:74–84.  Back to cited text no. 2
    
3.
Boniface K, Seneschal J, Picardo M, Taïeb A. Vitiligo: focus on clinical aspects, immunopathogenesis, and therapy. Clin Rev Allergy Immunol 2018; 54:52–67.  Back to cited text no. 3
    
4.
Esser C, Rannug A. The aryl hydrocarbon receptor in barrier organ physiology, immunology, and toxicology. Pharmacol Rev 2015; 67:259–279.  Back to cited text no. 4
    
5.
Wang X, Li S, Liu L, Jian Z, Cui T, Yang Y, et al. Role of the aryl hydrocarbon receptor signaling pathway in promoting mitochondrial biogenesis against oxidative damage in human melanocytes. J Dermatol Sci 2019; 96:33–41.  Back to cited text no. 5
    
6.
Napolitano M, Patruno C. Aryl hydrocarbon receptor (AhR) a possible target for the treatment of skin disease. Med Hypotheses 2018; 116:96–100.  Back to cited text no. 6
    
7.
Edamitsu T, Taguchi K, Okuyama R, Yamamoto M. AHR and NRF2 in skin homeostasis and atopic dermatitis. Antioxidants (Basel) 2022; 11:227.  Back to cited text no. 7
    
8.
Kyoreva M, Li Y, Hoosenally M, Hardman-Smart J, Morrison K, Tosi I, et al. CYP1A1 enzymatic activity influences skin inflammation via regulation of the AHR pathway. J Investig Dermatol 2021; 141:1553–1563.  Back to cited text no. 8
    
9.
Hamzavi I, Jain H, McLean D, Shapiro J, Zeng H, 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. 9
    
10.
Njoo MD, Das PK, Bos JD, Westerhof W. Association of the Köbner phenomenon with disease activity and therapeutic responsiveness in vitiligo vulgaris. Arch Dermatol 1999; 135:407–413.  Back to cited text no. 10
    
11.
Schallreuter KU, Salem MA, Gibbons NC, Maitland DJ, Marsch E, Elwary SM, et al. Blunted epidermal L-tryptophan metabolism in vitiligo affects immune response and ROS scavenging by Fenton chemistry, part 2: epidermal H2O2/ONOO–-mediated stress in vitiligo hampers indoleamine 2, 3-dioxygenase and aryl hydrocarbon receptor-mediated immune response signaling. FASEB J 2012; 26:2471–2485.  Back to cited text no. 11
    
12.
Liu B, Xie Y, Mei X, Sun Y, Shi W, Wu Z. Reciprocal regulation of interleukin-17A and interleukin-22 secretion through aryl hydrocarbon receptor activation in CD4+T cells of patients with vitiligo. Exp Ther Med 2021; 21:158.  Back to cited text no. 12
    
13.
Luecke S, Backlund M, Jux B, Esser C, Krutmann J, Rannug A. The aryl hydrocarbon receptor (AHR), a novel regulator of human melanogenesis. Pigment Cell Melanoma Res 2010; 23:828–833.  Back to cited text no. 13
    
14.
Rekik R, Ben Hmid A, Lajnef C, Zamali I, Zaraa I, Ben Ahmed M. Aryl hydrocarbon receptor (AhR) transcription is decreased in skin of vitiligo patients. Int J Dermatol 2017; 56:1509–1512.  Back to cited text no. 14
    
15.
Tsuji G, Hashimoto-Hachiya A, Takemura M, Kanemaru T, Ichihashi M, Furue M. Palladium and platinum nanoparticles activate AHR and NRF2 in human keratinocytes-implications in vitiligo therapy. J Investig Dermatol 2017; 137:1582–1586.  Back to cited text no. 15
    
16.
Shinde R, McGaha TL. The aryl hydrocarbon receptor: connecting immunity to the microenvironment. Trends Immunol 2018; 39:1005–1020.  Back to cited text no. 16
    
17.
Gutiérrez-Vázquez C, Quintana FJ. Regulation of the immune response by the aryl hydrocarbon receptor. Immunity 2018; 48:19–33.  Back to cited text no. 17
    
18.
Esser C, Bargen I, Weighardt H, Haarmann-Stemmann T, Krutmann J. Functions of the aryl hydrocarbon receptor in the skin. Sem Immunopathol 2013; 35:677–691.  Back to cited text no. 18
    
19.
Behfarjam F, Jadali Z. Vitiligo patients show significant up-regulation of aryl hydrocarbon receptor transcription factor. An Bras Dermatol 2018; 93:302–303.  Back to cited text no. 19
    
20.
Cella M, Colonna M. Aryl hydrocarbon receptor: linking environment to immunity. InSeminars in immunology. Academic Press 2015; 27:310–314.  Back to cited text no. 20
    
21.
Hidaka T, Ogawa E, Kobayashi EH, Suzuki T, Funayama R, Nagashima T, et al. The aryl hydrocarbon receptor AhR links atopic dermatitis and air pollution via induction of the neurotrophic factor artemin. Nat Immunol 2017; 18:64–73.  Back to cited text no. 21
    
22.
Shinde R, Hezaveh K, Halaby MJ, Kloetgen A, Chakravarthy A, da Silva Medina T, et al. Apoptotic cell–induced AhR activity is required for immunological tolerance and suppression of systemic lupus erythematosus in mice and humans. Nat Immunol 2018; 19:571–582.  Back to cited text no. 22
    
23.
Basak PY, Adiloglu AK, Ceyhan AM, Tas T, Akkaya VB. The role of helper and regulatory T cells in the pathogenesis of vitiligo. J Am Acad Dermatol 2009; 60:256–260.  Back to cited text no. 23
    
24.
Szelest M, Walczak K, Plech T. A new insight into the potential role of tryptophan-derived AhR ligands in skin physiological and pathological processes. Int J Mol Sci 2021; 22:1104.  Back to cited text no. 24
    
25.
Colucci R, Dragoni F, Moretti S. Oxidative stress and immune system in vitiligo and thyroid diseases. Oxid Med Cell Longev 2015; 2015: 631–1927.  Back to cited text no. 25
    


    Figures

  [Figure 1]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]



 

Top
 
 
  Search
 
Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
Access Statistics
Email Alert *
Add to My List *
* Registration required (free)

 
  In this article
Abstract
Introduction
Patients and methods
Results
Discussion
Conclusion
References
Article Figures
Article Tables

 Article Access Statistics
    Viewed446    
    Printed32    
    Emailed0    
    PDF Downloaded67    
    Comments [Add]    

Recommend this journal


[TAG2]
[TAG3]
[TAG4]