A potential oral microbiome signature associated with coronary artery disease in Tunisia.

The coronary artery disease (CAD) is a chronic inflammatory disease involving genetic as well as environmental factors. Recent evidence suggests that the oral microbiome has a significant role in triggering atherosclerosis. The present study assessed the oral microbiome composition variation between coronary patients and healthy subjects in order to identify a potential pathogenic signature associated with CAD. We performed metagenomic profiling of salivary microbiomes by 16S ribosomal RNA (rRNA) next-generation sequencing. Oral microbiota profiling was performed for 30 individuals including 20 patients with CAD and ten healthy individuals without carotid plaques or previous stroke or myocardial infarction. We found that oral microbial communities in patients and healthy controls are represented by similar global core oral microbiome. The predominant taxa belonged to Firmicutes (genus Streptococcus, Veillonella, Granulicatella, Selenomonas), Proteobacteria (genus Neisseria, Haemophilus), Actinobacteria (genus Rothia), Bacteroidetes (genus Prevotella, Porphyromonas), and Fusobacteria (genus Fusobacterium, Leptotrichia). More than 60% relative abundance of each sample for both CAD patients and controls is represented by three major genera including Streptococcus (24.97 and 26.33%), Veillonella (21.43 and 19.91%), and Neisseria (14.23 and 15.33%). Using penalized regression analysis, the bacterial genus Eikenella was involved as the major discriminant genus for both status and Syntax score of CAD. We also reported a significant negative correlation between Syntax score and Eikenella abundance in coronary patients' group (Spearman rho = -0.68, P=0.00094). In conclusion, the abundance of Eikenella in oral coronary patient samples compared with controls could be a prominent pathological indicator for the development of CAD.

Leukocyte mitochondrial DNA copy number is a potential non-invasive biomarker for psoriasis.

Abnormalities in the mitochondria have been linked to psoriasis, a chronic immune-mediated inflammatory skin disease. The mitochondrial DNA (mtDNA) is present in thousands of copies per cell and altered mtDNA copy number (mtDNA-CN), a common indicator of mitochondrial function, has been proposed as a biomarker for several diseases including autoimmune diseases. In this case-control study, we investigated whether the mtDNA-CN is related to psoriasis, correlates with the disease duration and severity, and can serve as a disease biomarker. Relative mtDNA-CN as compared with nuclear DNA was measured by a quantitative real-time polymerase chain reaction in peripheral blood buffy coat samples from 56 patients with psoriasis and 44 healthy controls. The receiver operating characteristic (ROC) curve analysis was performed to evaluate the value of mtDNA-CN as a biomarker. We found that the mtDNA-CN was significantly decreased in patients with psoriasis compared to healthy controls (93.6±5.3 vs. 205±71; P = 0.04). Sub-group analyses with stratification of patients based on disease duration under or over 10 years and disease severity indicated that the mtDNA-CN was significantly lower in patients with longer disease duration (74±4.3 in disease duration >10 years vs. 79±8.3 in disease duration <10 years, P = 0.009), and higher disease severity (72±4.3 in moderate-to-severe index vs. 88.3 ± 6 in mild index, P = 0.017). Moreover, the mtDNA-CN was negatively correlated with the disease duration and disease severity (r = -0.36, P = 0.006; r = -0.41, P = 0.003 respectively). The ROC analysis of mtDNA-CN showed an area under the curve (AUC) of 0.84 (95% confidence interval: 0.69-0.98; P = 0.002) for differentiating patients from healthy controls. Our study suggests that low mtDNA-CN may be an early abnormality in psoriasis and associates with the disease progression. Our study also suggests that mtDNA-CN may be a novel blood-based biomarker for the early detection of psoriasis.

Next-generation sequencing of the whole mitochondrial genome identifies novel and common variants in patients with psoriasis, type 2 diabetes mellitus and psoriasis with comorbid type 2 diabetes mellitus.

Recent studies have shown the role of mitochondrial DNA (mtDNA) variants in the pathogenesis of both psoriasis (Ps) and type 2 diabetes (T2D) amongst different ethnicities. However, no studies have investigated the mtDNA variants present in patients with Ps, T2D, and both Ps and T2D (Ps-T2D) in the Arab population. The entire mitochondrial genomes of Kuwaiti subjects with Ps, T2D, Ps-T2D and healthy controls were sequenced using Ion Torrent next-generation sequencing. A total of 36 novel mutations and 51 previously reported mutations were identified in the patient groups that were absent in the controls. Amongst the novel mutations, eight were non-synonymous and exhibited amino acid changes. Of these, two missense mutations (G5262A and A12397G) in the ND genes were detected in the Ps group and a C15735T missense mutation in the CYB gene was detected in Ps-T2D. Other known sequence variations were seen more frequently in all or certain patient groups compared with the controls (P<0.05). Additionally, the A8701G missense mutation in the ATPase 6 gene missense mutation was also observed in a higher frequency in the Ps group compared with the control. The present study is the first to perform a complete mitochondrial genome sequence analysis of Kuwaiti subjects with Ps, T2D and Ps-T2D, and both novel and known mtDNA variants were discovered. The patient-specific novel non-synonymous mutations may be co-responsible in the determination of these diseases. The higher frequency of certain mtDNA variants in the patients compared with the controls may suggest a role in predisposing patients to these diseases. Further functional analyses are required to reveal the role of the identified mutations in these disease conditions.

Mitochondrial Haplogroup Reveals the Genetic Basis of Diabetes Mellitus Type 2 Comorbidity in Psoriasis.

OBJECTIVE: Published data show a clear link between psoriasis (Ps) and the increasing prevalence of comorbid conditions, such as diabetes mellitus type 2 (DM2). The role of the mitochondrial genomic haplogroup in the potential coexistence of Ps and DM2 comorbidity is the subject of this study. MATERIAL AND METHODS: Ninety-eight Kuwaiti individuals were recruited in 4 cohorts (20 healthy controls, 15 with DM2, 34 with Ps, and 29 with Ps and diabetes mellitus). An Ion Torrent S5XL was used to sequence mitochondrial DNA (mtDNA). χ2 test was used to assess differences in the distribution of each haplogroup between cases and controls (p < 0.05). The Bonferroni correction was applied (p < 0.004). The mtDNA haplogroups were analyzed by HaploGrep. RESULTS: Haplogroups R0, U, J, T, N, L3, M, H, X, HV, R, and K were detected in the studied population. Haplogroup M had a high risk for Ps (odds ratio (OR) 4.0, p = 0.003). Haplogroup R0 and J had decreased the risk of DM2 (OR 0.28, p = 0.007). CONCLUSION: Our results indicated that mtDNA haplogroups have a potential contribution to the pathogenesis of Ps and DM2 comorbidity. We show for the first time that the comorbidity of diabetes in Ps may be related to mitochondrial dysfunction.

The Effects of HFE Polymorphisms on Biochemical Parameters of Iron Status in Arab Beta-Thalassemia Patients.

In this study, the potential effect of three HFE gene polymorphisms (C282Y, H63D and S65C) and the SLC40A1 A77D polymorphism on iron balance was investigated in 234 subjects (91 Arab beta-thalassemia major (BTM) patients, 34 beta-thalassemia trait (BTT) individuals and 109 health controls). Genotyping was done using restriction-fragment-length polymorphism and direct-sequencing. Serum-iron, total iron binding capacity, transferrin and ferritin were estimated in all BTT and BTM, and in 65 healthy controls. H63D was the only polymorphism detected in our cohort. Allele frequency was 13% in both BTM and BTT and 10% in controls with no significant difference. Serum iron, ferritin and transferrin saturation were significantly higher in normal males heterozygous for H63D as compared to homozygous wild-type males. Ferritin was significantly higher in BTT males with or without H63D polymorphism when compared to the healthy males with H/H genotype. No such difference was observed between H/H versus H/D BTT subgroups. We conclude that H63D is the only significant hemochromatosis-associated polymorphism in the Arabian Gulf region. The heterozygous state of H63D may significantly alter iron parameters in normal males. In BTT, it appears that the beta-thalassemia allele has an overriding influence on ferritin values, and this generally manifest in males.

Discordance between Lifestyle-Related Health Practices and Beliefs of People Living in Kuwait: A Community-Based Study.

OBJECTIVE: To examine the concordance between lifestyle practices and beliefs of people living in Kuwait, and between their lifestyle practices and established evidence-informed recommendations for health. SUBJECTS AND METHODS: A cross-sectional interview questionnaire study was conducted using a convenience sample of 100 adults living in Kuwait (age range 19-75 years). The interview included sections on demographics, and lifestyle-related practices and beliefs related to smoking, diet/nutrition, physical activity/exercise, sleep, and stress. Diet/nutrition and physical activity/exercise benchmarks were based on international standards. Analyses included descriptive statistics and the χ2 test. RESULTS: Beliefs about the importance of nutrition in lifestyle-related conditions were limited, and this was apparent in participants' dietary habits, e.g., low consumption of fruit/vegetables and multigrains: 16 (16%) and 9 (9%) met the recommended guidelines, respectively. Ninety-nine (99%) believed physical activity/exercise affects health overall, and 44 (44%) exercised regularly. Of the sample of 100, 20 (20%) exercised in accordance with evidence-based recommendations for maximal health. Compared with beliefs about other lifestyle-related behaviors/attributes, respondents believed nutrition contributed more than stress to heart disease, cancer, and stroke, and stress contributed more than nutrition to hypertension and diabetes. CONCLUSION: In this study, our findings showed a discrepancy between lifestyle-related practices and beliefs, and between each of these and evidence-based recommendations for maximal health, i.e., not smoking, several servings of fruit and vegetables and whole-grain foods daily, healthy weight, restorative sleep, and low-to-moderate stress levels.

De-regulation of diabetic regulatory genes in psoriasis: Deciphering the unsolved riddle.

The purpose of our study was to identify the currently lacking molecular mechanism that accounts for the co-occurrence of two seemingly disparate diseases: psoriasis and type II diabetes. We aimed to investigate a panel of 84 genes related to the diabetic regulatory network in psoriasis (Ps), psoriasis type II diabetes (Ps-T2D), type II diabetes (T2D) and healthy control (HC). We hypothesize that such attempts would provide novel diagnostic markers and/or insights into pathogenesis of the disease. A quantitative Real Time-PCR Human Diabetes RT(2) Profiler PCR Array was chosen to explore the expression profile 84 diabetic genes in study subjects. Statistical analysis was carried out using appropriate software. The analysis revealed three candidate genes GSK3B, PTPN1, STX4 that are differentially expressed in study subjects. GSK3B was highly significant in Ps-T2D (P=0.00018, FR=-26.6), followed by Ps (P=0.0028, FR=-14.5) and T2D groups (P=0.032, FR=-5.9). PTPN1 showed significant association only with PS-T2D (P=0.00027, FR=-8.5). STX4 showed significant association with both Ps (P=0.0002, FR=-20) and Ps-T2D (P=0.0016, FR=-11.2). ACE represents an additional marker that showed suggestive association with Ps (P=0.0079, FR=-9.37). Our study highlights the complex genetics of Ps-T2D and present biomarkers for the development of T2D in Ps cases.

Influence of MX1 promoter rs2071430 G/T polymorphism on susceptibility to systemic lupus erythematosus.

The expression of interferon inducible genes are reported to be heightened in systemic lupus erythematosus (SLE); nevertheless, not much is known regarding the genetic variants underlying these genes and their role in the pathogenesis of disease. Herein, we aim to explore the potential association and contribution of polymorphisms in MX1 gene (i) promoter with part of exon 1 (ii) intron 6, and (iii) their resulting haplotypes, with susceptibility to SLE. A total of 306 subjects, 152 SLE and 154 healthy controls (HC), were screened by direct sequencing method. Statistical analysis was carried out using appropriate software. The screening region of interest in MX1 revealed the existence of promoter (-123C/A, -88G/T, -20 A/C) and intron 6 (+9862G/A, +10190G/A, +9901C/G, +9920C/A, +9959C/T, +10047A/G) variants in SLE and HC. A significant association was observed between MX1 -88G/T SNP and susceptibility to SLE (χ (2) = 4.18, p = 0.04, OR = 1.89, 95 % CI 1.03-3.5). Haplotype analysis also revealed increased risk of SLE among individuals carrying CTA haplotype (-123 C, -88 T, -20 A) (χ (2) = 5.74, p = 0.017, OR = 4.28, 95 % CI 1.30-14.06). None of the other tested variants showed any significant association with SLE. The present study is the first to reveal the influence of genetic variation in MX1 gene in susceptibility to SLE.

Th-17 related regulatory network in the pathogenesis of Arab patients with systemic lupus erythematosus and lupus nephritis.

AIM: The present study aimed to identify the genes involved in the pathogenesis of systemic lupus erythematosus (SLE) in Arabs by investigating a panel of 84 genes related to the t helper (Th)17-related regulatory network and to further explore the expression levels of serum interleukin (IL)-17A and IL-17F in a studied cohort. A comparative analysis of gene expression profile in SLE and lupus nephritis (LN) patients against that of healthy controls (HC) was performed. METHOD: A quantitative real-time polymerase chain reaction (PCR) (Th17 autoimmunity and inflammation) array analysis was performed in peripheral white blood cells of 66 SLE patients under specific medical treatment and 30 age/gender/ethnically matched healthy controls. Statistical analysis was carried out using the RT(2) Profiler TM PCR Data Analysis tool. RESULTS: The analysis of Th17 pathway revealed 14 genes (IL-17A, IL-17C, IL-17D, IL-17F, IL-18, IL-12RB2, IL-23R, CCL2, CCL20, CXCL5, MMP3, RORC, STAT4 and TRAF6) that are differentially expressed in SLE and HC (fold change [FC] < 2, P < 0.0006). No significant difference in expression profiles was observed between SLE and LN. A significant difference in serum concentration of IL-17A (P = 0.002) and IL-17F (P = 0.002) was observed between SLE (13.91 ± 4.25) and LN (18.26 ± 4.24). CONCLUSION: Our study is the first to investigate a panel of 84 genes related to Th17 regulatory pathway in Arab SLE subjects and the first to explore the effect of current immunosuppression regimens on Th17 regulatory pathway. It paves the way for understanding the etiology of SLE and autoimmune diseases in general.

Genetic structure of the Kuwaiti population revealed by paternal lineages.

OBJECTIVE: We analyzed the Y-chromosome haplogroup diversity in the Kuwaiti population to gain a more complete overview of its genetic landscape. METHOD: A sample of 117 males from the Kuwaiti population was studied through the analysis of 22 Y-SNPs. The results were then interpreted in conjunction with those of other populations from the Middle East, South Asia, North and East Africa, and East Europe. RESULTS: The analyzed markers allowed the discrimination of 19 different haplogroups with a diversity of 0.7713. J-M304 was the most frequent haplogroup in the Kuwaiti population (55.5%) followed by E-M96 (18%). They revealed a genetic homogeneity between the Kuwaiti population and those of the Middle East (FST = 6.1%, P-value < 0.0001), although a significant correlation between genetic and geographic distances was found (r = 0.41, P-value = 0.009). Moreover, the nonsignificant pairwise FST genetic distances between the Kuwait population on the one hand and the Arabs of Iran and those of Sudan on the other, corroborate the hypothesis of bidirectional gene flow between Arabia and both Iran and Sudan. CONCLUSION: Overall, we have revealed that the Kuwaiti population has experienced significant gene flow from neighboring populations like Saudi Arabia, Iran, and East Africa. Therefore, we have confirmed that the population of Kuwait is genetically coextensive with those of the Middle East.

Genome-wide differential expression reveals candidate genes involved in the pathogenesis of lupus and lupus nephritis.

BACKGROUND: Systemic lupus erythematosus (lupus) is an autoimmune disease characterized by multiorgan pathology, accelerated apoptosis and hyper-autoantibody production against self-components. The root cause of lupus remains unknown, although multiple susceptibility factors have been reported in different ethnic group. OBJECTIVE: We aimed to explore the genome-wide differential expression spectrum of lupus and its severe form lupus nephritis (LN) in Arab females. METHODS: A total of 98 subjects: 40 lupus, 18 LN and 40 age/gender/ethnically matched healthy controls (HC) were recruited. Carefully chosen subjects (n = 11) were employed for whole human-genome expression profiling using high-density Human Exon 1.0.ST arrays (Affymetrix) and statistical analysis was carried out using appropriate software. Validation cohorts (n = 98) were investigated to quantify the expression of the nine selected candidate genes relative to GAPDH as endogenous control. RESULTS: Genome-wide differential analysis revealed seven candidate genes in lupus and 36 in LN, when individually compared to HC (anova Welch t-test, P ≤ 0.005, Tukey's honestly post hoc analysis). Analysis of differentially expressed genes with a fold change of 2, revealed 16 Gene Ontology terms satisfying a P ≤ 0.05. We further detected five distinct inflammatory and metabolic pathways such as TWEAK, osteopontin, endochondral ossification, fluropyrimidine activity and urea cycle and metabolism of amino groups that significantly contribute to the pathogenesis of lupus (P < 0.05). Validation of selected candidate genes (IRF9, ABCA1, APOBEC3, CEACAM3, OSCAR, TNFA1P6, MMP9, SLC4A1) revealed significant differences in expression, indicating their promissory role in the pathogenesis of lupus. CONCLUSION: Our study provides central gene regulators of therapeutic potential, indicating the future prospects of the study.

Genome-wide peripheral blood transcriptome analysis of Arab female lupus and lupus nephritis.

Systemic lupus erythematosus (lupus) is a genetically heterogeneous autoimmune disorder with an obscure etiology. With 92-94% of human genes exhibiting alternative splicing, gaining insights to such events may lead to better diagnostics. Herein, we explored the genome-wide peripheral blood transcriptome of lupus and its severe form lupus-nephritis (LN) compared to healthy controls (HC). Age/gender/ethnically-matched Arab females were tested using high-density arrays and statistical analysis was carried out using appropriate software. Analysis revealed 15 splice variants that are differentially expressed between lupus/HC and 99 variants between LN/HC (p ≤ 0.05, SI> or ≤ 0.5, Benjamin Hochberg-False discovery rate correction). Comparison between LN/lupus revealed 7 variants that significantly differed in expression. Pathway analysis of differentially spliced-genes postulated 11 significant pathways in lupus and 12 in LN (p<0.05). Analysis of peripheral blood transcriptome possibly revealed signature causative genes that are alternatively spliced, signifying their clinical relevance. Present study is the first to reveal the significance of alternative variants in lupus and LN.

Violating the theory of single gene-single disorder: inhibitor development in hemophilia.

Hemophilia is clinically and genetically heterogeneous blood disorder with several known gene defects accounting for the diversity of disease phenotype and inhibitor production. Although increasing number of causative mutations have been reported, not much is known regarding the root cause of inhibitor development against infused blood clotting factors, which represents a major challenge in the treatment of disease. The variations in the severity and frequency of bleeding in hemophiliacs with same molecular defect, indicates the role of modifier genes in the pathogenesis of disease. Herein, we aim to review and summarise the literature over the past decade, to gain insight into what is critical for the development of inhibitors in hemophilia. Aside from potential mutations in factor VIII and IX, polymorphisms in various genes such as human leukocyte antigen-I (HLA-I), HLA-II, tumor necrosis factor-alpha, interleukin-10 and cytotoxic T-lymphocyte associated antigen-4, also tends to contribute to the development of inhibitors. Violating the theory of single gene-single disorder, new research indicates that inhibitor arise from a complex interplay of multiple genetic, immunological and environmental factors. With the revolutionary advances in whole genome sequencing, we propose a detailed genome wide study to identify the spectrum of genetic markers involved in the development of inhibitors for better diagnostics and therapeutics.

Association of interferon regulatory factor 5 (IRF5) markers with an increased risk of lupus and overlapping autoimmunity in a Kuwaiti population.

BACKGROUND: Variations in IRF5 have been associated with an increased risk of developing lupus (SLE). AIM: To explore the association of IRF5 markers (rs2004640, rs10954213) with susceptibility to SLE and to the development of more than one autoimmune disease (OP) and to analyse the influence of rs2004640 on IRF5 isoform expression. METHODS: A total of 300 age/gender/ethnicity-matched Kuwaitis were screened using the TaqMan®SNP Genotyping Assay. Data analysis was conducted in three groups: patients with high/low anti-dsDNA antibody titre, lupus nephritis/non-nephritis and disease onset age ≤28 years/>28 years. cDNA samples with defined rs2004640 genotypes were amplified using specific primers and transcripts associated with each of the exon1 variants were detected on polyacrylamide-gel. RESULTS: A significant association was observed between the IRF5 markers (rs2004640, rs10954213) and susceptibility to SLE (p < 0.0001) and OP (p < 0.025). Stratified analysis showed a significant association of the rs2004640 T-allele with a high titre of anti-dsDNA antibody (p = 0.0035, OR = 1.92) and a low disease onset age ≤28 years (p = 0.0026, OR = 1.94). Additionally, the influential role of the rs2004640 T allele on IRF5 expression was observed. CONCLUSIONS: This study is the first to investigate the association of IRF5 markers with OP and the very first to explore the association of these markers with SLE in Arabs. The novelty of this study is the association of the rs2004640 T-allele with increased anti-dsDNA antibody production and SLE in a younger age group.

Influence of HumDN1 VNTR polymorphism on DNASE1 expression in systemic lupus erythematosus and rheumatoid arthritis.

The purpose of this study was to analyze the effect of the HumDN1 VNTR polymorphism on DNASE1 mRNA expression and enzyme activity in lupus (SLE) and rheumatoid arthritis (RA) compared to healthy control (HC). Kuwait subjects (n = 500) matched by age/gender/ethnicity were genotyped by fragment-analysis. DNASE1 expression was analysed using quantitative Real-Time-PCR and sera from subjects were screened for DNase1 reduction activity by ELISA. Allele and genotype distribution of HumDN1 VNTR revealed a significant association with susceptibility to SLE and RA (p < 0.05, OR > 1). Relative expression analysis revealed a significant increase in DNASE1 mRNA in SLE (p = 0.0001) and RA (p = 0.002) compared to HC. Stratification of subjects revealed, increased DNASE1 expression in SLE with 5/5 (p = 0.0001), 3/4 (p = 0.0001) and 3/5 genotype (p = 0.01). A reduction in DNASE1 expression was specifically observed in SLE with 4,4 genotype (p = 0.0004). RA patients with 3/4 genotype (p = 0.02) showed a significant increase in DNASE1 expression. Similarly a significant association was observed between DNase1 reduction activity and SLE (p = 0.0001). SLE patients with 3,4 (p = 0.0001) and 5,5 genotype (p = 0.0001) showed increased DNase1 reduction activity, while a lack of association was observed with RA. The present study is the first to reveal the effect of HumDN1 VNTR on DNASE1 expression in SLE and RA.

Differential expression of alternative splice variants of CTLA4 in Kuwaiti autoimmune disease patients.

Cytotoxic T lymphocyte associated antigen4 (CTLA4) is a candidate susceptibility gene for the study of autoimmune diseases. The present study sought to explore the expression profile of the CTLA4 gene in autoimmune patients, such as rheumatoid arthritis (RA), systemic lupus erythematous (SLE) and Hashimoto's thyroiditis (HT), compared to healthy controls (HCs). A total of 88 (22 RA, 22 SLE 22 HT, 22 HCs) age-, gender- and ethnicity-matched individuals were recruited. The hypersensitive capillary electrophoresis method was employed to detect the CTLA4 splice variants. PCRs of the patient's cDNA using CTLA4-specific primers followed by cloning and sequencing were used to distinguish the various splice variants. The biochemical properties of all known CTLA4 variants were analysed using the ExPASy and ESEfinder programmes. Six alternatively spliced variants of the CTLA4 gene were detected in this study. These included mCTLA4-672, sCTLA4-562, N-CTLA4-292, L-CTLA4-277, ssCTLA4-214 and K-CTLA4-142 bp. K-CTLA4-142 bp and N-CTLA4-292 bp represented two novel splice variants of the CTLA4 gene. A reduction in the frequency of mCTLA4-672 bp and sCTLA4-562 bp was observed in SLE and RA patients compared to healthy controls. The shortest splice variant, K-CTLA4-142 bp, was predominantly detected in all of the tested cohorts,while the decreased expression of the N-CTLA4-292 bp variant was observed in the autoimmune subjects. The exonic splice enhancer motifs of the SRp40 protein were found exactly at the splice junction of wCTLA4 (-ACAGAGC-, 2.7) and K-CTLA4 (-TGAAAAG-, 3.37), and that of the SRp55 protein was found at the splice junction of L-CTLA4 (-TGTGTG-, 2.82). Our study highlights the discrepancies in the expression spectrum of the CTLA4 gene in autoimmune patients and healthy subjects. The abnormal expression pattern of the CTLA4 gene in autoimmune patients suggests that in addition to allelic variation, the expression pattern of CTLA4 could contribute to autoimmunity.

Genetic analysis of interleukin-1 receptor antagonist and interleukin-1ß single-nucleotide polymorphisms C-511T and C+3953T in alopecia areata: susceptibility and severity association.

The present study was aimed to explore the effect of two selected polymorphisms from interleukin-1ß (IL-1ß) gene [SNPs -511 and +3953] and a variable number of tandem repeat (VNTR) from interleukin-1 receptor antagonist (IL-1RN) on the susceptibility and severity of alopecia areata (AA) in Kuwaiti subjects. IL-1ß SNPs C-511T, C+3953T, and IL-1RN VNTR were screened in 96 alopecia patients classified clinically, according to the disease severity as patchy (P), semiuniversalis (SU), and universalis (U), and in 100 ethnically matched healthy controls. Polymerase chain reaction followed by restriction fragment length polymorphism and direct DNA sequencing were employed for genotyping. Comparing the stratified AA cases based on severity, IL-ß SNP C-511T showed a significant association (genotype and allelotype levels p = 0.03 and p = 0.028, respectively). Genotype CC was 50 % more frequent in U cases than in P or SU. When P and SU were grouped and tested against U, a significant difference was observed (genotype and allelotype levels p = 0.006 and p = 0.008, respectively). Compared to genotype CT, carriers of IL-1ß -511 CC genotype showed an increased risk to develop severe AA (p = 0.004, OR = 4.14, 95 % CI = 1.61-10.69). Four alleles and genotypes (1/1, 1/3, 1/4, and 2/2) of IL-1RN VNTR were detected in AA patients while only two (1/1 and 1/3) in controls. IL-1RN VNTR showed genotype and allelotype association with AA (p = 0.05 and p = 0.025, respectively). Our results showed that IL-1ß and IL-1RN VNTR are significantly associated with the susceptibility to alopecia areata. Allele C of the IL-ß C-511T SNP is linked to the severe form of AA.

The effect of UGT1A1 promoter polymorphism in the development of hyperbilirubinemia and cholelithiasis in hemoglobinopathy patients.

Present study was aimed to explore the effect of (TA)n UGT1A1 gene promoter polymorphism on bilirubin metabolism, bilirubinaemia, predisposition to cholelithiasis and subsequent cholecystectomy, in Sickle-Cell Anemia (SCA) and beta-Thalasemia major (bTH) in Kuwaiti subjects compared to other population. This polymorphism was analyzed and correlated to total bilirubin and cholelithiasis in 270 age, gender, ethnically matched subjects (92 bTH, 116 SCA and 62 Controls) using PCR, dHPLC, fragment analysis and direct sequencing. Four genotypes of UGT1A1 were detected in this study (TA6/6, TA6/7, TA6/8 and TA7/7). (TA)6/8 was found only in four individuals; hence it was not included in the analysis. There was a statistically significant association of genotypes with serum total bilirubin levels in both bTH and SCA groups (p<0.001). Subjects with (TA)7/7 had the highest total serum bilirubin level (178.7 ± 3.5 µmole/l). A significant association was observed between allele (TA)7 and cholelithiasis development (p = 0.0001). The 40%, 67.5% and 100% of SCA with (TA)6/6, (TA)6/7 and (TA)7/7 respectively developed cholelithiasis and were subsequently cholecystectomized. Our results confirm UGT1A1 (TA)7 allele as one of the factors accounting for the hyperbilirubinemia and cholelithiasis observed in SCA and bTH.

Association analysis of nitric oxide synthases: NOS1, NOS2A and NOS3 genes, with multiple sclerosis.

BACKGROUND: Multiple sclerosis (MS) is a chronic inflammatory autoimmune disorder of the central nervous system. AIM: To explore the genetic basis of three nitric oxide synthase (NOS) genes: NOS1, NOS2A and NOS3, with susceptibility to MS. SUBJECTS AND METHODS: A total of 122 MS patients and 118 healthy controls screened for NOS1 (rs2682826, rs41279104), NOS2A (CCTTT)n/(TAAA)n and NOS3 (rs1800783, rs1800779, rs2070744, 27bpVNTR) markers, using TaqMan®SNP Genotyping Assays and fragment analysis were enrolled in this study. QRT-PCR and ELISA were used to analyse the expression of NOS3 mRNA and Nitric Oxide (NO) levels. RESULTS: Two NOS3 markers were associated with susceptibility to MS and early disease development. The NOS3 rs1800779 G-allele (p = 0.04) and GG-genotype (p = 0.02) showed association with susceptibility to MS. Short NOS2 (CCTTT)n (p = 0.03) and short/long repeat (p = 0.04) genotypes also showed associations with MS. These associations were intensified by sub-division of patients into Kuwaiti Arabs and Persians (p < 0.05). The NOS3-27 bp-VNTR a-allele was associated with early MS disease onset ≤26 years (p = 0.04). The NOS3-27 bp-VNTR a/b-genotype resulted in 23% lower NO production and the NOS3-rs1800779 AA-genotype resulted in lower NOS3 expression. Haplotypes obtained from NOS2A and NOS3 showed increased susceptibility to MS. NOS1 showed no significant association with MS. CONCLUSION: This study provides evidence for the association between selected NOS2 and NOS3 markers and MS susceptibility.