TaqI polymorphism T/t genotypes at the vitamin D receptor gene (VDR) are associated with increased serum vitamin D levels in mild and moderate psoriasis vulgaris: A pilot study

Several types of polymorphisms in vitamin D receptor (VDR) have been found in psoriasis.


| INTRODUCTION
Psoriasis is a repetitive, immune-mediated inflammatory condition defined by clearly delineated, erythematous papules and plaques enveloped in silvery-white scales. 1 Psoriasis affects 2-4% of the world's population. People of both genders suffer equally, while the disorder primarily affects individuals between the ages of 20 and 30, and 50 and 60. 2 Vitamin D3 (cholecalciferol) in its natural state is mainly synthesized in the skin and acquired through nutrition. 3,4 Its bio-active form, 1,25-dihydroxyvitamin D [1,25 (OH) 2D3], which acts through a specialized vitamin D receptor (VDR), is implicated in various operations, such as transcriptional regulation of various genes to facilitate their genomic activity on calcium homeostasis, aging, immune function, immune modulation, cell growth, proliferation and differentiation. [5][6][7][8][9][10][11][12] The VDR, a member of the nuclear steroid receptor subgroup, has many polymorphisms: BsmI, TaqI, ApaI, and FokI, and they correspond to VDR gene polymorphisms and vulnerability to atopic dermatitis, asthma and psoriasis. [13][14][15][16][17][18] To the best of our knowledge, no research on the distribution of VDR gene polymorphisms in psoriasis or changes in serum vitamin 25(OH)D levels (sVDLs) has been conducted in Saudi Arabia. Therefore, researchers are interested in conducting this study to determine whether polymorphisms in the vitamin D receptor gene play a key role, especially TaqI, in the risk of psoriasis. Therefore, this study investigated the role of the TaqI polymorphism in the VDR gene as a factor in changing sVDLs in psoriasis patients and evaluated its possible relationship with clinical outcomes.

| Subjects
This comparative case-control study was carried out in the Dermatology Outpatient Department for a year. It was carried out in accordance with the Helsinki Declaration guidelines and was authorized by the Local Ethical Committee. Before enrollment, all subjects provided written informed consent after being briefed on the purpose and nature of the study.

| Inclusion criteria
Adult male and female patients of any age with a clinically established diagnosis of psoriasis but with no systemic involvement were eligible.

| Exclusion criteria
Patients on vitamin D therapy, pregnant, lactating, or with malignancies, active liver disease or renal disorders were excluded.
In terms of subject count, we conducted a pilot study and used a small number of subjects as a "feasibility" study. It is a small-scale preliminary study conducted prior to large-scale quantitative research to assess the potential for a future study evaluating the TaqI polymorphism and its association with psoriasis severity. This has allowed us to forecast an adequate sample size, budget for it, and strengthen the study design before embarking on a full-scale project.
A total of 118 patients (64 males and 54 females) were enrolled, and the diagnosis was performed through clinical and physical examinations. Patients were divided into three groups based on the body surface area affected: severe psoriasis vulgaris involving more than 10% of the body surface; moderate psoriasis vulgaris involving 5-10% of the body surface; and mild psoriasis vulgaris involving less than 5% of the body surface. 19 The remaining 94 subjects were healthy participants who were gender and age aligned to the psoriasis group (58 males and 36 females); they had no clinical evidence of psoriasis or any other autoimmune condition. Patients were recruited from the clinic when searching for medical advice. Controls were selected from close associates of the cases, such as friends and relatives who were attending with the patients or healthy escorts of dermatology patients attending the dermatology outpatient clinic.
Both groups underwent comprehensive physical and clinical assessments, genetic analysis and vitamin D estimations.

| Genomic DNA extraction
Anticoagulant Na 2 EDTA was used to gather samples of blood. The QIAamp ® DNA BloodMini Kit was used to purify genetic DNA from 200 L of whole blood in accordance with the Blood guidelines.

| Detection of TaqI polymorphism using PCR-RFLP
To genotype the TaqI polymorphism, a genomic DNA fragment was amplified using the polymerase chain reaction (PCR) technique and a couple of oligonucleotide primers: the upstream primer sequence was

| Serum vitamin D concentration measurements
Patients' baseline sVDLs were determined. Blood samples were obtained from veins and analysed using the Roche Cobas e411 in less than 24 h (Roche Diagnostics System, Switzerland). According to the Food and Nutrition Board of the Institute of Medicine, sVDLs were classified as adequate (>20 ng/ml), inadequate (12-20 ng/ml) and deficient (12 ng/ml). 21

| Statistical analysis
The Statistical Package for Social Sciences, version 20.0, was used to perform statistical analysis (SPSS Inc., Chicago, Illinois, USA). To assess the mean and percentage differences, an analysis of variance (ANOVA) testing was performed between more than two means. t-Tests and chi-squares were used; p > 0.05 was regarded as insignificant. Additionally, quantitative data are represented as a mean ± standard deviation (SD), whereas qualitative data are articulated as the percentage and frequency.

| RESULTS
The TaqI gene polymorphism was investigated in 118 psoriasis patients (64 males and 54 females) and 94 control healthy individuals (58 males and 36 females). There was no clinical evidence or family history of psoriasis or an autoimmune condition in the healthy individuals. Table 1  was the most prevalent, followed by t/t (28.8%), and then T/T (25.4%). In control healthy individuals, the most common genotype was t/t (44.6%), while the frequencies of T/t and T/T genotypes were 29.9 and 25.5%, respectively ( Table 2). The allele frequency did not differ significantly between patients and control healthy individuals.
The T allele was found to be more prevalent in patients than in healthy subjects (48.3 vs. 40.4%), with no significant differences (p = 0.421). Furthermore, there were no significant differences in the frequency of the t allele p = 0.883 between patients and healthy individuals (Table 3). Serum vitamin D levels in T/t patients were statistically significantly higher (p = 0.004; Table 4).
When serum vitamin D levels were compared, the different clinical types of psoriasis, serum vitamin D levels were found to be higher in patients with moderate psoriasis than in patients with mild and severe psoriasis vulgaris patients (p = 0.032 Table 5). The T/T genotypes were compared between clinical types, finding that serum vitamin D levels do not differ by p = 0.723. When comparing t/t genotypes between clinical types, the same results were observed; p = 0.004. Table 6 shows that there are significant differences between the T/t genotypes and the variable clinical types.
Only 12 of the 38 patients with severe psoriasis vulgaris had T/T genotypes, whereas 13 of the 32 patients with moderate psoriasis vulgaris had T/t genotypes; t/t genotypes were found in 19 of the 48 moderate patients. Vitamin D levels were higher in patients with moderate psoriasis vulgaris than in patients with severe and mild psoriasis vulgaris (p ≤ 0.001; Table 7A). When we compared serum vitamin D levels in six patients with severe psoriasis vulgaris t/t, 13 moderate patients with T/t and eight mild patients with T/T, we discovered that T/t in moderate patients had higher levels of serum vitamin D than severe t/t and mild T/T (p ≤ 0.001; Table 7B). There was no significant variation noted when we compared severe psoriasis vulgaris patients with T/t genotypes with T/T moderate patients and T A B L E 1 General and laboratory characteristics of psoriatic patients and healthy subjects t/t mild patients (Table 7C). The same results were shown between the severe psoriasis vulgaris patients with T/t genotypes, moderate patients with t/t and mild patients with T/T (Table 7D).
Serum vitamin D levels were significantly higher in moderate and mild psoriasis vulgaris patients with T/t compared with the other genotypes (T/T and t/t) in the same clinical type p ≤ 0.001 (Table 8).

| DISCUSSION
Because psoriasis is multidimensional, genetic factors are thought to be important predictors of these illnesses, and researchers have been  [22][23][24] In this study, we discovered that the TaqI polymorphism is associated with psoriasis.
T/t and t/t findings were significantly higher in patients than in controls. The allele frequency did not differ significantly between patients and healthy subjects. The results showed that the T allele was more prevalent in patients than in healthy subjects. This is consistent with the results of Acikbas et al., in a study conducted on 204 participants. 18 Our findings contradict the findings of Halsall et al, 22 who reported that the T allele and T/T genotype are linked with the vitamin D3 reaction. It is well recognized that societies' ethnic composition and genetic background significantly impact the VDR genotype distribution.
In contrast to our findings, the T/T genotype frequency was greater in patients than in healthy subjects in our previous research on Japanese psoriasis patients 29 and Asian and Turkish populations. 32 There are two possible explanations for the obvious disparity between these previous findings and ours. Initially, the size of the healthy subject population in our study was quite large. Second, Saudi Arabia has geographically distinct areas with variable climatic, ideological, social, economic and genetic variations, and the country's population diversity complicates genetic analysis.
Our results indicate that T/t genotype patients have statistically Vitamin D levels were greater in patients with moderate psoriasis vulgaris than in those with severe and mild psoriasis vulgaris (p = 0.001). We discovered that Tt in moderate patients has higher serum vitamin D levels than severe T/T and mild T/T (p = 0.001).
When we compared the T/t genotypes in patients with severe psoriasis vulgaris with the T/T genotypes in moderate patients and the tt genotypes in mild patients, there was no discernible difference.
The same results were discovered between severe psoriasis vulgaris with T/t genotypes, moderate patients with Tt and mild patients with T/T. Despite the few patients with psoriasis studied, this is the first study to show that the T allele is more prevalent in Saudi patients with psoriasis than in healthy subjects. We hypothesized that the alleles of the polymorphic regions of the VDR gene were closely This was consistent with the findings of Liu's meta-analysis study. 35 Finally, this study found that VDR TaqI polymorphism increases psoriasis vulnerability in the Saudi population. More research into the connections between VDR polymorphisms and psoriasis vulnerability is needed, based on these findings. To explore the specific role of VDR gene polymorphisms in psoriasis pathogenesis, larger scale studies in larger populations are necessary.

| CONCLUSION
The research was based on the difference in vitamin D levels between patients and healthy subjects; the study also discovered a difference in vitamin D levels with different TaqI genotypes. Furthermore, moderate patients had higher increases in vitamin D levels than mild and severe patients. T/t genotypes are also linked to higher vitamin D levels in moderate and mild patients.

| Limitations
The limitations of this study are that, first, the small number of patients does not cover all regions of Saudi Arabia, and second, we recommend investigating other VDR polymorphism genotypes.

FINANCIAL SUPPORT AND SPONSORSHIP
Nil.

CONFLICTS OF INTEREST
There are no conflicts of interest.

DATA AVAILABILITY STATEMENT
The data that support the findings of this study are available from the corresponding author upon reasonable request.