V-set domain containing T-cell activation inhibitor-1 (VTCN1): A potential target for the treatment of autoimmune diseases.

Autoimmunity eventuates when the immune system attacks self-molecules as a result of the breakdown in immune tolerance. Targeting autoimmune diseases via immunomodulation has become an essential strategy in today's era. A B7 superfamily member immune checkpoint, the V-set domain containing T-cell activation inhibitor-1 (VTCN1), also known as B7-H4, B7S1, and B7x, is involved in negatively regulating T-cell activation. VTCN1 transcript has been reported in various lymphoid and non-lymphoid tissues, but its protein expression is restricted, indicating its translational regulation. Dysregulation of VTCN1 has resulted in the exacerbation of various autoimmune diseases. Moreover, increased soluble form of VTCN1 in the patient's sera positively correlates with the disease progression and severity. The current review summarizes all the reports till date, unfolding the role of VTCN1 in various autoimmune diseases and its therapeutic potential.

Altered Levels of Negative Costimulatory Molecule V-Set Domain-Containing T-Cell Activation Inhibitor-1 (VTCN1) and Metalloprotease Nardilysin (NRD1) are Associated with Generalized Active Vitiligo.

BACKGROUND: Vitiligo is characterized by depigmented macules on the skin caused due to autoimmune destruction of melanocytes. V-set domain-containing T-cell activation inhibitor-1 (VTCN1) is a negative costimulatory molecule that plays a vital role in suppressing autoimmunity and tuning immune response. Nardilysin (NRD1), a metalloproteinase, cleaves membrane-tethered VTCN1 resulting in the shedding of soluble-VTCN1 (sVTCN1). However, the role of VTCN1 and NRD1 in vitiligo pathogenesis is unexplored. OBJECTIVES AND METHODS: This study was aimed to (i) Investigate the association of VTCN1 intronic polymorphisms (rs10923223 T/C and rs12046117 C/T) with vitiligo susceptibility in Gujarat population by using Polymerase Chain Reaction- Restriction Fragment Length Polymorphism (PCR-RFLP) (ii) Estimate VTCN1 & NRD1 transcript levels from peripheral blood mononuclear cells (PBMCs) and skin samples of vitiligo patients by real-time PCR, (iii) Estimate sVTCN1 and NRD1 protein levels from plasma by ELISA and (iv) Estimate VTCN1 protein levels in the skin samples of vitiligo patients by immunofluorescence. RESULTS: The analysis revealed increased VTCN1 and NRD1 transcript levels in the skin (p = .039, p = .021 respectively), increased sVTCN1 and NRD1 levels (p = .026, p = .015 respectively) in the plasma, and decreased VTCN1 protein levels (p = .0002) in the skin of vitiligo patients as compared to healthy controls. The genetic analysis revealed no significant association of VTCN1 intronic polymorphisms rs10923223 T/C and rs12046117 C/T with vitiligo susceptibility in Gujarat population (p = .359, p = .937, respectively). CONCLUSIONS: The present study revealed altered VTCN1 and NRD1 expressions in the blood and skin of vitiligo patients, suggesting their potential role in the development and progression of Vitiligo.

Autoantigen Discovery in the Hair Loss Disorder, Alopecia Areata: Implication of Post-Translational Modifications.

Alopecia areata (AA) is a chronic, multifactorial, polygenic, and heterogeneous disorder affecting growing hair follicles in susceptible individuals, which results in a non-scarring and reversible hair loss with a highly unpredictable course. Despite very considerable research effort, the nature of the precipitating factor(s) responsible for initiating AA in any given hair follicle remains unclear, due largely to significant gaps in our knowledge of the precise sequence of the etiopathogenic events in this dermatosis. However, disease-related changes in the immune-competence of the lower growing hair follicle, together with an active immune response (humoral and cellular) to hair follicle-associated antigens, are key associated phenomena. Confirmation of the hair follicle antigen(s) implicated in AA disease onset has remained stubbornly elusive. While it may be considered somewhat philosophical by some, it is also unclear whether immune-mediated hair loss in AA results from a) an ectopic (i.e., in an abnormal location) immune response to native (unmodified) self-antigens expressed by the healthy hair follicle, b) a normal immune response against modified self-antigens (or neoantigens), or c) a normal immune response against self-antigens (modified/non-modified) that were not previously visible to the immune system (because they were conformationally-hidden or sequestered) but become exposed and presentable in an MHC-I/-II molecule-restricted manner. While some candidate hair follicle antigen target(s) in AA are beginning to emerge, with a potential role for trichohyalin, it is not yet clear whether this represents the initial and immunodominant antigenic focus in AA or is simply one of an expanding repertoire of exposed hair follicle tissue damage-associated antigens that are secondary to the disease. Confirmation of autoantigen identity is essential for our understanding of AA etiopathogenesis, and consequently for developing a more informed therapeutic strategy. Major strides have been made in autoantigen discovery in other autoimmune conditions. In particular, some of these conditions may provide insights into how post-translational modifications (e.g., citrullination, deamidation, etc.) of hair follicle-restricted proteins may increase their antigenicity and so help drive the anti-hair follicle immune attack in AA.

Investigation of the Role of Interleukin 6 in Vitiligo Pathogenesis.

Interleukin-6 (IL6) is involved in pathogenesis of several autoimmune disorders including vitiligo. Hence, we aimed to investigate the association of IL6 -174 G/C and -572 G/C polymorphisms and its transcript levels with vitiligo; to evaluate the effect of IL-6 on normal human melanocyte (NHM) viability and expression of IL6R, MITF and TYR. IL6 -174 G/C and -572 G/C polymorphisms were genotyped by ARMS-PCR and PCR-RFLP respectively in 343 controls and 322 vitiligo patients. IL6 transcript levels were estimated from PBMCs (96 controls and 77 patients) and skin samples (15 controls and 15 patients) by qPCR. NHM viability was assessed by MTT; IL6R, MITF and TYR transcript and protein levels were monitored by qPCR and ICC respectively. Genetic analyses revealed no association of IL6 -174 G/C polymorphism (p> .05) with vitiligo. Analysis of IL6 -572 G/C revealed reduced risk of vitiligo in individuals with GC/CC genotypes compared to GG genotype (p = .010). IL6 expression was significantly increased (p = .0197) in PBMCs of patients. Further, IL6 expression was significantly higher in non-lesional skin compared to controls (p = .009). In-vitro NHM viability was decreased upon IL-6 exposure (10-50 ng/ml; p< .05), with significantly increased IL6R transcript (p = .042) and protein levels (p = .003) however, MITF transcript (p = .0003) and protein levels (p = .016), and TYR transcript levels (p = .001) were significantly decreased. The results suggest that IL6 -572 G/C polymorphism might be associated with vitiligo susceptibility in Gujarat population. Moreover, increased IL6 expression in vitiligo patients and its effect on NHM suggest a potential role in melanocyte biology. CONCLUSION: The results suggest that IL6 - 572 G/C polymorphism might be associated with vitiligo susceptibility in Gujarat population. Moreover, increased IL6 expression in vitiligo patients and its effect on NHM suggest a potential role in melanocyte biology.

Elevated X-Box Binding Protein1 Splicing and Interleukin-17A Expression Are Associated With Active Generalized Vitiligo in Gujarat Population.

Vitiligo is an autoimmune skin disorder defined by the destruction of functional epidermal melanocytes. It is a multifactorial and polygenic disorder caused due to oxidative stress, endoplasmic reticulum (ER) stress, and autoimmunity, among other factors. In the present study, we aimed to investigate the association of X-box Binding Protein 1 (XBP1) and Interleukin-17A (IL-17A) polymorphisms and monitor their systemic as well as skin expression levels in vitiligo patients from Gujarat population in India. XBP1 rs2269577 G/C, IL17A rs2275913 G/A and IL17A rs8193036 C/T polymorphisms were genotyped by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) method in 312 controls and 276 vitiligo patients. Transcript levels of spliced (sXBP1), unspliced XBP1 (uXBP1) and IL17A from peripheral blood mononuclear cells (PBMCs) as well as spliced and unspliced XBP1 from skin samples were analyzed by qPCR. IL-17A protein levels in suction-induced blister fluid (SBF) from the skin of study subjects were estimated by ELISA. The results revealed that genotype (p=0.010) and allele (p=0.014) frequencies of XBP1 rs2269577 G/C polymorphism were significantly different, however, no significant difference was observed in frequencies of IL17A rs2275913 G/A and IL17A rs8193036 C/T polymorphisms in control and patient population. Gene expression analysis revealed that sXBP1 and IL17A levels were significantly higher in PBMCs of generalized (p=0.030 and p=0.039, respectively) and active (p=0.024 and p=0.017, respectively) vitiligo patients. Moreover, we observed a significantly elevated sXBP1 expression (p=0.037) as well as IL-17A protein levels (p=0.009) in perilesional skin of vitiligo patients as compared to controls. Overall, these findings suggest XBP1 and IL17A play an important role in vitiligo and further substantiate the involvement of ER stress in exacerbating immune-mediated vitiligo pathogenesis.

Publisher Correction: A genetic analysis identifies a haplotype at adiponectin locus: Association with obesity and type 2 diabetes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

A genetic analysis identifies a haplotype at adiponectin locus: Association with obesity and type 2 diabetes.

Adiponectin is a prime determinant of the status of insulin resistance. Association studies between adiponectin (ADIPOQ) gene single nucleotide polymorphisms (SNPs) and metabolic diseases have been reported earlier. However, results are ambiguous due to apparent contradictions. Hence, we investigated (1) the association between ADIPOQ SNPs: -11377C/G, +10211T/G, +45T/G and +276G/T for the risk towards type 2 diabetes (T2D) and, (2) genotype-phenotype association of these SNPs with various biochemical parameters in two cohorts. Genomic DNA of diabetic patients and controls from Gujarat and, Jammu and Kashmir (J&K) were genotyped using PCR-RFLP, TaqMan assay and MassArray. Transcript levels of ADIPOQ were assessed in visceral adipose tissue samples, and plasma adiponectin levels were estimated by qPCR and ELISA respectively. Results suggest: (i) reduced HMW adiponectin/total adiponectin ratio in Gujarat patients and its association with +10211T/G and +276G/T, and reduced ADIPOQ transcript levels in T2D, (ii) association of the above SNPs with increased FBG, BMI, TG, TC in Gujarat patients and (iii) increased GGTG haplotype in obese patients of Gujarat population and, (iv) association of -11377C/G with T2D in J&K population. Reduced HMW adiponectin, in the backdrop of obesity and ADIPOQ genetic variants might alter metabolic profile posing risk towards T2D.

A Concise Review on the Role of Endoplasmic Reticulum Stress in the Development of Autoimmunity in Vitiligo Pathogenesis.

Vitiligo is characterized by circumscribed depigmented macules in the skin resulting due to the autoimmune destruction of melanocytes from the epidermis. Both humoral as well as cell-mediated autoimmune responses are involved in melanocyte destruction. Several studies including ours have established that oxidative stress is involved in vitiligo onset, while autoimmunity contributes to the disease progression. However, the underlying mechanism involved in programing the onset and progression of the disease remains a conundrum. Based on several direct and indirect evidences, we suggested that endoplasmic reticulum (ER) stress might act as a connecting link between oxidative stress and autoimmunity in vitiligo pathogenesis. Oxidative stress disrupts cellular redox potential that extends to the ER causing the accumulation of misfolded proteins, which activates the unfolded protein response (UPR). The primary aim of UPR is to resolve the stress and restore cellular homeostasis for cell survival. Growing evidences suggest a vital role of UPR in immune regulation. Moreover, defective UPR has been implicated in the development of autoimmunity in several autoimmune disorders. ER stress-activated UPR plays an essential role in the regulation and maintenance of innate as well as adaptive immunity, and a defective UPR may result in systemic/tissue level/organ-specific autoimmunity. This review emphasizes on understanding the role of ER stress-induced UPR in the development of systemic and tissue level autoimmunity in vitiligo pathogenesis and its therapeutics.

Circulatory Omentin-1 levels but not genetic variants influence the pathophysiology of Type 2 diabetes.

OBJECTIVE: Omentin-1, an anti-inflammatory protein, is secreted by the visceral adipose tissue. Altered levels of Omentin-1 are associated with obesity and Type 2 Diabetes (T2D). Although Omentin-1 is implicated in the insulin signaling pathway, the relationship between the genetic variants of Omentin-1 and T2D is not yet explored. The current study evaluates the association of Omentin-1 polymorphisms (rs2274907 A/T and rs1333062 G/T), its transcript and protein levels, and genotype-phenotype correlation with metabolic parameters and T2D susceptibility. METHODS: Plasma and Peripheral Blood Mononuclear Cells (PBMCs) were separated from venous blood taken from 250 controls and 250 T2D patients recruited from Gujarat, India. Genomic DNA was isolated from PBMCs and genotyping of Omentin-1 variants was performed by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP). RNA was isolated from Visceral Adipose Tissue (VAT) samples of 12 controls and 10 patients, and transcript levels of Omentin-1 were assessed by qPCR. Plasma Omentin-1 levels were estimated by ELISA. Fasting Blood Glucose, Body Mass Index (BMI) and plasma lipid profile were considered for the genotype-phenotype correlation analysis. RESULTS: Our study revealed no association of Omentin-1 genetic variants with T2D risk (p > 0.05). However, the AT genotype of Omentin-1 rs2274907 A/T polymorphism was associated with increased BMI (p = 0.0247). Plasma Omentin-1 levels were significantly decreased (p < 0.0001) however, increased VAT Omentin-1 transcript levels (p = 0.0127) were observed in T2D patients. CONCLUSION: Our findings suggest that decreased circulatory Omentin-1 levels could pose a risk towards T2D susceptibility.

Cytokines: the yin and yang of vitiligo pathogenesis.

INTRODUCTION: Dysregulation of melanocyte function is associated with vitiligo, an idiopathic autoimmune hypopigmentary skin disorder, caused by the selective destruction of melanocytes. Cytokines, the key mediators of immune response, which are pivotal in maintaining immune homeostasis, are crucial in vitiligo pathogenesis. Several studies indicate that there is an imbalance between pro- and anti-inflammatory cytokines in the skin and serum of vitiligo patients. Areas covered: In this comprehensive review, we have summarized the correlation of cytokine imbalance and vitiligo pathogenesis, its role in melanocyte biology, and its impact on vitiligo treatment. We have integrated various published reports on the levels of major cytokines from skin and serum samples of vitiligo patients. We have also discussed the role of endoplasmic reticulum and oxidative stress on cytokine imbalance and vice versa leading to destruction of melanocytes. Expert commentary: The review reflects that dysregulation of cytokines is multifactorial, ranging from genetic predisposition to altered protein expression relevant to vitiligo pathogenesis. We emphasize that cytokine imbalance in systemic and skin microenvironment plays a crucial role in vitiligo pathogenesis and has promising potential as therapeutic targets for vitiligo.

Association of interleukin 1 receptor antagonist intron 2 variable number of tandem repeats polymorphism with vitiligo susceptibility in Gujarat population.

BACKGROUND: Vitiligo is a multifactorial, polygenic, autoimmune skin disorder caused by selective destruction of melanocytes. Interleukin 1 receptor antagonist intron 2 polymorphism was found to be associated with various autoimmune disorders. AIMS: We aimed to investigate the association of interleukin 1 receptor antagonist intron 2 variable number of tandem repeats polymorphism (rs2234663) with vitiligo to assess interleukin 1 receptor antagonist transcript levels and to perform possible genotype-phenotype correlation. METHODS: Three hundred and seven vitiligo patients and 316 controls were enrolled in the study, genotyping of interleukin 1 receptor antagonist rs2234663 was performed by polymerase chain reaction, and relative gene expression of interleukin 1 receptor antagonist was carried out in peripheral blood mononuclear cells from patients (n = 36) and controls (n = 36) by real-time-PCR. RESULTS: A significant difference was observed in the frequency of interleukin 1 receptor antagonist *A (1/2) genotype among patients with active and stable vitiligo (P = 0.0172). Interleukin 1 receptor antagonist*A (2/2) genotype and allele frequencies were significantly different between SV patients and controls (P = 0.0246 and P = 0.0046, respectively). Significant difference was also observed for interleukin 1 receptor antagonist*A2 (allele) in active and stable vitiligo patients (P = 0.0060). However, other comparisons did not show any significant difference in genotype and allele frequencies. Moreover, interleukin 1 receptor antagonist*A (3/2) genotype was observed only in patients whereas interleukin 1 receptor antagonist*A (5/2) was observed only in controls. Gene expression analysis showed no significant difference in interleukin 1 receptor antagonist transcript levels in patients compared to controls (P = 0.5962). Interestingly, genotype-phenotype correlation analysis revealed that individuals with IL1RN*A (2/2) exhibited higher interleukin 1 receptor antagonist expression compared to other major genotypes interleukin 1 receptor antagonist*A (1/2) (P = 0.01) and interleukin 1 receptor antagonist*A (1/1) (P = 0.03). LIMITATIONS: More case-control studies on interleukin 1 receptor antagonist rs2234663 polymorphism and gene expression from different ethnic populations are required to explore the impact of interleukin 1 receptor antagonist in vitiligo susceptibility. CONCLUSION: Interleukin 1 receptor antagonist*A2 might be a risk factor for progressive vitiligo.

Association of elevated homocysteine levels and Methylenetetrahydrofolate reductase (MTHFR) 1298 A > C polymorphism with Vitiligo susceptibility in Gujarat.

BACKGROUND: Several studies have reported hyperhomocysteinemia in vitiligo patients, suggesting the potential role of elevated homocysteine levels in precipitating vitiligo. OBJECTIVES: We aimed to estimate homocysteine and vitamin B12 levels, and to investigate the role of MTHFR 677 C > T and 1298 A > C polymorphisms in vitiligo susceptibility in Gujarat population. METHODS: Homocysteine and vitamin B12 levels were estimated in plasma of 55 vitiligo patients and 60 controls by Electrochemiluminescence immunoassay (ECLIA). Polymerase chain reaction- restriction fragment length polymorphism (PCR-RFLP) and amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) techniques were used to genotype MTHFR 677 C > T and 1298 A > C polymorphisms in 520 vitiligo patients and 558 controls. RESULTS: Our results showed significantly elevated homocysteine levels (p = 0.0003) as well as significant decrease in vitamin B12 levels (p = 0.0102) in vitiligo patients, as compared to controls. No significant difference in genotype and allele frequencies of MTHFR 677 C > T polymorphism was observed among patients and controls, however, the frequency of 'CC' genotype of MTHFR 1298 A > Cpolymorphism was significantly increased in patients as compared to controls (p = 0.0151). Analysis based on the type of vitiligo revealed a significant increase in 'C' allele of MTHFR 1298 A > C polymorphism in patients with generalized (p = 0.003) and active (p = 0.007) vitiligo as compared to controls. Both the polymorphisms of MTHFR were in low linkage disequilibrium (LD) and susceptible 'TC' haplotype was more frequently observed (p = 0.008) in vitiligo patients. Interestingly, elevated homocysteine levels were also positively correlated with MTHFR 1298 A > C polymorphism in vitiligo patients. Structure based in silico prediction revealed structural perturbations in MTHFR protein due to Ala222Val and Glu429Ala amino acid substitution. CONCLUSIONS: The present findings suggest that MTHFR 1298 A > C polymorphism and, altered homocysteine and vitamin B12 levels might play a vital role in the precipitation of vitiligo.

A case-control study on association of proteasome subunit beta 8 (PSMB8) and transporter associated with antigen processing 1 (TAP1) polymorphisms and their transcript levels in vitiligo from Gujarat.

BACKGROUND: Autoimmunity has been implicated in the destruction of melanocytes from vitiligo skin. Major histocompatibility complex (MHC) class-II linked genes proteasome subunit beta 8 (PSMB8) and transporter associated with antigen processing 1 (TAP1), involved in antigen processing and presentation have been reported to be associated with several autoimmune diseases including vitiligo. OBJECTIVES: To explore PSMB8 rs2071464 and TAP1 rs1135216 single nucleotide polymorphisms and to estimate the expression of PSMB8 and TAP1 in patients with vitiligo and unaffected controls from Gujarat. METHODS: PSMB8 rs2071464 polymorphism was genotyped using polymerase chain reaction- restriction fragment length polymorphism (PCR-RFLP) and TAP1 rs1135216 polymorphism was genotyped by amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) in 378 patients with vitiligo and 509 controls. Transcript levels of PSMB8 and TAP1 were measured in the PBMCs of 91 patients and 96 controls by using qPCR. Protein levels of PSMB8 were also determined by Western blot analysis. RESULTS: The frequency of 'TT' genotype of PSMB8 polymorphism was significantly lowered in patients with generalized and active vitiligo (p = 0.019 and p = 0.005) as compared to controls suggesting its association with the activity of the disease. However, TAP1 polymorphism was not associated with vitiligo susceptibility. A significant decrease in expression of PSMB8 at both transcript level (p = 0.002) as well as protein level (p = 0.0460) was observed in vitiligo patients as compared to controls. No significant difference was observed between patients and controls for TAP1 transcripts (p = 0.553). Interestingly, individuals with the susceptible CC genotype of PSMB8 polymorphism showed significantly reduced PSMB8 transcript level as compared to that of CT and TT genotypes (p = 0.009 and p = 0.003 respectively). CONCLUSIONS: PSMB8 rs2071464 was associated with generalized and active vitiligo from Gujarat whereas TAP1 rs1135216 showed no association. The down-regulation of PSMB8 in patients with risk genotype 'CC' advocates the vital role of PSMB8 in the autoimmune basis of vitiligo.