Role of tagged SNPs of the AGT gene in causing susceptibility to essential hypertension

Aim: Angiotensinogen (AGT) is one of the candidate genes that has been extensively investigated for association of its variants with essential hypertension. Studies focusing on the contribution of tagged single nucleotide polymorphisms (SNPs) in the AGT gene are limited and lacking from Indian population. Hence, the present study was carried out to examine the role of five tagged SNPs viz., g.6147G>A (rs7539020), g.5978A>G (rs2493134); g.6241T>C (rs1078499), g.7781G>T (rs11122577), and g.5855G>A (rs3789678) in the development of hypertension. Materials and Methods: 202 hypertensives and 222 normotensives were screened for five tagged SNPs using the method of polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP). Results: The present study revealed significant association of g.5855G>A polymorphism with essential hypertension in different logistic regression models wherein protection was conferred by g.5855G>A against developing the condition. The polymorphism led to the creation of new exonic splicing enhancer and destruction of exonic splicing silencer site thereby enhancing the process of mRNA splicing. The haplotypes AGTG and GACG were found to have a significant protective effect. Other polymorphisms did not show any significant association with hypertension. Conclusion: The present study is the first one to report the protective role of g.5855G>A polymorphism in the development of essential hypertension. The results reflect possibility of ethnic variation in the contribution of g.5855G>A polymorphism of the AGT gene to essential hypertension.     

Investigation of T-cell immunoglobulin- and mucin-domain-containing molecule-3 (TIM-3) polymorphisms in essential thrombocythaemia (ET)

Objectives: T-cell immunoglobulin- and mucin-domain-containing molecule-3 (TIM-3) is preferentially expressed on terminally differentiated Th1 cells and inhibits their IFN-γ production. It has been reported that chronic inflammation may be an important driving force for myeloproliferative neoplasms (MPNs). Therefore, we hypothesized that as an important inflammation regulator, TIM-3 may be involved in essential thrombocythaemia (ET). The goal of this study was to investigate whether the ?1516G?>?T, ?574G?>?T and +4259T?>?G single-nucleotide polymorphisms (SNPs) within the TIM-3 gene contribute to the genetic susceptibility of individuals to ET.

Methods: Genotyping of the TIM-3 ?1516G?>?T, ?574G?>?T and?+?4259T?>?G SNPs was performed in 175 patients with ET and in 151 controls via a polymerase chain reaction-restriction fragment length polymorphism assay. We also investigated the relationships between the genotypes of each SNP and the risk factors of ET such as routine blood indexes, age and JAK2 V617F mutation.

Results: The genotype and allele frequencies of the ?1516G?>?T SNP (p?=?0.016 and 0.019, respectively), the ?574G?>?T SNP (p?=?0.035 and 0.038, respectively) and the +4259T?>?G SNP (p?=?0.036 and 0.038, respectively) of the ET patients and the controls were significantly different. A haplotype analysis found that the GGT and TGT haplotypes had significantly different distributions between ET and controls (p?=?0.041 and 0.041, respectively). However, no significant differences were detected between the genotypes of all SNPs and routine blood indexes, age and JAK2V617F mutation.

Conclusion: The ?1516G?>?T, ?574G?>?T and +4259T?>?G SNPs within TIM-3 gene might play an important role as a genetic risk factor in the pathogenesis of ET.     

Molecular Epidemiological Survey of Glucose-6-Phosphate Dehydrogenase Deficiency and Thalassemia in Uygur and Kazak Ethnic Groups in Xinjiang,Northwest China

Glucose-6-phosphate dehydrogenase (G6PD) deficiency and thalassemia occur frequently in tropical and subtropical regions, while the prevalence of relationship between the two diseases in Xinjiang has not been reported. We aimed to determine the prevalence of these diseases and clarify the relationship between genotypes and phenotypes of the two diseases in the Uygur and Kazak ethnic groups in Xinjiang. We measured G6PD activity by G6PD:6PGD (glucose acid-6-phosphate dehydrogenase) ratio, identified the gene variants of G6PD and α- and β-globin genes by polymerase chain reaction (PCR)-DNA sequencing and gap-PCR and compared these variants in different ethnic groups in Xinjiang with those adjacent to it. Of the 149 subjects with molecular analysis of G6PD deficiency conducted, a higher prevalence of the combined mutations c.1311C?>?T/IVSXI?+?93T?>?C and IVSXI?+?93T?>?C, both with normal enzymatic activities, were observed in the Uygur and Kazak subjects. A case of rare mutation HBB: c.135delC [codon 44 (?C) in the heterozygous state], a heterozygous case of HBB: c.68A?>?G [Hb G-Taipei or β22(B4)Glu→Gly] and several common single nucleotide polymorphisms (SNPs) were found on the β-globin gene. In conclusion, G6PD deficiency with pathogenic mutations and three common α-thalassemia (α-thal) [–?–^SEA, ?α^3.7 (rightward), ?α^4.2 (leftward)] deletions and point mutations of the α-globin gene were not detected in the present study. The average incidence of β-thalassemia (β-thal) in Uygurs was 1.45% (2/138) in Xinjiang. The polymorphisms of G6PD and β-globin genes might be useful genetic markers to trace the origin and migration of the Uygur and Kazak in Xinjiang.     

Ten Years of Routine α- and β-Globin Gene Sequencing in UK Hemoglobinopathy Referrals Reveals 60 Novel Mutations

We review and report here the genotypes and phenotypes of 60 novel thalassemia and abnormal hemoglobin (Hb) mutations discovered following the adoption of routine DNA sequencing of both α- and β-globin genes for all UK hemoglobinopathy samples referred for molecular investigation. This screening strategy over the last 10 years has revealed a total of 11 new β chain variants, 15 α chain variants, 19 β-thalassemia (β-thal) mutations and 15 α⁺-thalassemia (α⁺-thal) mutations. The large number of new thalassemia alleles confirms the wide racial heterogeneity of mutations in the UK immigrant population. Eleven of the new variants ran with Hb A on high performance liquid chromatography (HPLC), demonstrating the value of routine sequencing of both α- and β-globin genes for all hemoglobinopathy investigations.

The new β chain variants are: Hb Bury [β22(B4)Glu → Asp (HBB: c.69A?>?T)], Hb Fulwood [β35(C1)Tyr?→?His (HBB: c.106T?>?C)], Hb Little Venice [β42(CD1)Phe?→?Cys (HBB: c.128T?>?G)], Hb Cork [β57(E1)Asn?→?Ser (HBB: c.173A?>?G), Hb Basingstoke [β118(GH1)Phe?→?Ser (HBB: c.356T?>?C)], Hb Howden [β20(B2)Val?→?Ala (HBB: c.62T?>?C)], Hb Wilton [β41(C7)Phe?→?Leu (HBB: c.126C?>?A)], Hb Belsize Park [β120(GH3)Lys?→?Asn (HBB: c.363A?>?T)], Hb Hampstead Heath [β2(NA2)His?→?Gln;β26(B8)Glu?→?Lys (HBB: c.[6C?>?G;79G?>?A])], Hb Grantham [β85(F1)Phe?→?Cys (HBB: c.257T?>?G)] and Hb Calgary [β64(E8)Gly?→?Val (HBB: c.194G?>?T).

The new α chain variants are: Hb Edinburgh [α70(E19)Val?→?Gly (HBA2: c.212T?>?G)], Hb Walsgrave [α116(GH4)Glu?→?Val (HBA2: c.350A?>?T)], Hb Wexham [α117(GH5) and 118(H1) insertion Ser (HBA1: c.354-355insTCA)], Hb Coombe Park [α127(H10)Lys?→?Glu (HBA2: c.382A?>?G)], Hb Oxford [α17(A15)Val?→?Asp (HBA2: c.53T?>?A)], Hb Bridlington [α32(B13)Met?→?Thr (HBA1: c.98T?>?C), Hb Wolverhampton [α81(F2)Ser?→?Tyr (HBA2: c.9245C?>?A)], Hb Little Waltham [α13(A11)Ala?→?Asp (HBA2: c.41C?>?A)], Hb Derby [α61(E10)Lys?→?Arg (HBA1: c.185A?>?G)], Hb Uttoxter [α74(EF3)Tyr?→?Asp (HBA2: c.223G?>?T)], Hb Harehills [α124(H7)Ser?→?Cys (HBA1: c.374C?>?G)], Hb Hekinan II [α27(B8)Glu?→?Asp (HBA1: c.84G?>?T)], Hb Manitoba IV [α102(G9)Ser?→?Arg (HBA1: c.307A?>?C), Hb Witham [α139(HC1)Lys?→?Arg (HBA2: c.419A?>?G) and Hb Farnborough [α9(A7)Asn?→?Asp (HBA1: c.28A?>?G). In addition, 10 more paralogous α-globin chain variants have been discovered.

The novel β-thal alleles are: HBB: c.-138C?>?G, HBB: c.-121C?>?T, HBB: c.-80T?>?G, HBB: c.18_19delTG, HBB: c.219_220insT, HBB: c.315?+?2_315?+?13delTGAGTCTATGGG, HBB: c.316-70C?>?G, HBB: c.345_346insTGTGCTG, HBB: c.354delC, HBB: c.376-381delCCAGTG, HBB: c.393T?>?A, HBB: c.394_395insA, HBB: c.375_376insA, HBB: c.*+95_*+107delTGGATTCTinsC, HBB: c.*?+?111_*+112delAA, HBB: c.*+112A?>?T, HBB: c.394C?>?T, HBB: c.271delG and HBB: c.316-3C?>?T.

The novel α?^+?-thal alleles are: HBA1: c.95+1G?>?C, HBA1: c.315C?>?G [Hb Donnington, α104(G11)Cys?→?Trp], HBA1: c.327delC, HBA1: c.333_345del, HBA1: c.*+96G?>?A, HBA2: c.2T?>?G, HBA2: c.112delC, HBA2: c.143delA, HBA2: c.143_146delACCT, HBA2: c.156_157insG, HBA2: c.220_223delGTGG, HBA2: c.305T?>?C [Hb Bishopstown, α101(G8)Leu?→?His], HBA2: c.169_170delAA, HBA2: c.1A?>?T and HBA2: c.-3delA.     

Association between SNPs in <Emphasis Type="Italic">AdipoQ</Emphasis>, +45 T &gt; G and +276 G &gt; T,and adiponectin levels in the Korean Chinese population in Yanbian,China

This study investigated the association between single nucleotide polymorphisms (SNPs) in AdipoQ, +45 T?>?G and +276 G?>?T, and adiponectin levels in the Korean Chinese population in Yanbian, China. A total of 329 subjects were involved in this study, including 178 female and 151 male individuals. All of them are ethnic Koreans living in Yanbian, aged from 31 to 70, and 58 % of them were diagnosed with type 2 diabetes (T2D). Items tested and calculated include total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), fasting plasma glucose (FPG), body fat percentage (BF%), fasting plasma insulin (FPI), plasma adiponectin (PA), and homeostasis model assessment of insulin resistance (HOMA-IR). SNPs were screened with the method of SNaPshot. P?<?0.05 was defined as the statistically significant threshold. Results include the following: (1) PA levels in the T2D group are much lower than those in the normal group (P?<?0.001); (2) the distribution of genotypes of SNPs +45 T?>?G and +276 G?>?T was determined to be in Hardy-Weinberg equilibrium (P?>?0.05) and complete linkage disequilibrium (|D′|?=?1.0); (3) PA levels in females are much higher than those in males no matter which group they belong to (normal or T2D group) (P?=?0.004 and P?=?0.018); (4) PA levels are higher in the individuals with +45G as a dominant allele than those in individuals with homozygous genotype TT at locus +45 in the normal group (P?=?0.037); and (5) of SNPs +45 T?>?G and +276 G?>?T, no risk factor of genotype or allele was found (P?>?0.05). Three conclusions can be drawn: (1) PA levels are lower in T2D patients than in normal persons; (2) PA levels are higher in females than in males; (3) PA levels are higher for normal individuals with +45G allele than those with homozygous TT genotype at locus +45 in AdipoQ.     

Synergistic effects of gene polymorphisms of the renin–angiotensin–aldosterone system on essential hypertension in Kazakhs in Xinjiang

Objective: To assess the synergistic effects of gene polymorphisms of the renin–angiotensin–aldosterone system (RAAS) on essential hypertension (EH) in Kazakhs in Xinjiang. Methods: A cross-sectional case-control association study was conducted in 52 1 hypertensive and 623 normotensive subjects of Kazakh ethnicity on eight common single nucleotide polymorphisms (SNPs) interspersed over five genes of the RAAS. SNPs were genotyped by polymerase chain reaction-restriction fragment length polymorphism. Interactions among the SNPs were analyzed by the multifactor dimensionality reduction method (MDR). Results: In single-locus analysis, subjects with AGT -6G, ACE D, and CYP11B2 -344C had increased susceptibility to EH (OR: 1.249; 1.425; 1.201). When subgrouped by sex, males with the t allele of REN Taq I had decreased risk for EH (OR: 0.529), and those with AGT -6G and CYP11B2 -344 C had increased risk for EH (OR: 1.498; 1.449). In females, carrying ACE D increased the risk for EH. (OR: 1.327). In six AGT haplotypes, H1 was protective, while H3 increased susceptibility to EH (OR: 0.683; 2.025). Interaction analysis by MDR showed that there was a strong synergistic effect between ACE I/D and CY11B2 (T-344C) and a moderate interaction between both ACE I/D and CY11B2 T-344C and AGT A-6G. Conclusions: There was a strong synergistic effect between ACE I/D and CY11B2 T-344C and a moderate effect between both ACE I/D and CY11B2 T-344C and AGT A-6G. AGT -6G, ACE D, and CY11B2 -344C increased susceptibility to EH. REN Taq I, AGT -6G, CY11B2 -344 C and ACE D were associated with male and female EH, respectively. H1 and H3 of AGT were protective and risk haplotypes, respectively.     

Association between CD14 gene promoter polymorphisms with serum total-IgE and eosinophil levels in atopic and non-atopic asthma patients in a Chinese Han population

Objective: The aim of this study was to investigate the association between CD14 gene promoter SNPs with serum total-IgE and eosinophil levels in atopic asthma and non-atopic asthma in Chinese Han. Methods: A total of 152 patients with asthma were divided into atopic asthma (n?=?100) and non-atopic asthma (n?=?52) groups for this study. Six CD14 gene SNPs were analyzed using PCR and gene sequencing. Serum total-IgE and eosinophil levels were measured. The association between genotype frequencies of the CD14 gene loci with total-IgE and eosinophil levels in atopic asthma and non-atopic asthma was evaluated by the ANOVA test method. Hundred and sixteen healthy subjects constitute the control group. Results: We found that serum total-IgE and eosinophil levels were significantly higher in individuals with atopic asthma when compared to individuals with non-atopic asthma (p?<?0.01). For non-atopic asthma, the total-IgE levels of the heterozygous genotypes were significantly higher than the corresponding levels for the homozygous genotypes in CD14-260C?>?T, CD-651C?>?T, CD-911A?>?C and CD-1247A?>?G (p?<?0.01). In atopic asthma, there was no statistical significance for either serum total-IgE or eosinophil levels among the genotypes of the CD14 gene SNPs. In addition, allele A frequency of CD14-1247A?>?G was significantly different between the atopic asthma and non-atopic asthma groups (p?=?0.025). Conclusions: There was a statistical association between the serum total-IgE level and the CD14 gene promoter SNPs in the non-atopic asthma group. The eosinophil level was not found to be statistically associated with the CD14 gene promoter SNPs in either the atopic asthma or non-atopic asthma groups.     

The Spectrum of β-Thalassemia Mutations in a Population from the Brazilian Amazon

The spectrum of β-thalassemia (β-thal) mutations was investigated for the first time in a cohort of 33 unrelated patients from the Brazilian Amazon attending the Center for Hemotherapy and Hematology of the Pará Foundation (HEMOPA), in Belém, the state capital of Pará, Northern Brazil. Identification of the β-thal mutations was made by direct genomic sequencing of the β-globin gene. Mutations were identified in all patients, corresponding to a spectrum of 10 different point mutations and a total of 37 alleles studied. HBB: c.92?+?5G?>?A [IVS-I-5 (G?>?A)], was the most common β-thal mutation, followed by HBB: c.118C?>?T [codon 39 (C?>?T)], HBB: c.?138C?>?T [?88 (C>T)], HBB: c.92?+?1G?>?A [IVS-I-1 (G?>?A)] and HBB: c.92?+?6T?>?C [IVS-I-6 (T?>?C)] mutations. These five mutations (four Mediterranean origin and one African origin) accounted for 86.5% of the β-thal alleles. The profile of β-thal mutations found in northern Brazil is different from those described in other regions of the country. In the southeast and south, the nonsense mutation HBB: c.118C?>?T is the most prevalent, followed by HBB: c.93-21G?>?A [IVS-I-110 (G?>?A)], whereas in the northeast, HBB: c.92?+?6T?>?C has been identified as the most common mutation, followed by HBB: c.92?+?1G?>?A. This heterogeneous geographical distribution is certainly related to the ancestry of Brazilian populations because they have similar genetic backgrounds (European, African and Amerindian), although with slightly different admixture proportions. Furthermore, the European contribution in the southeast and south was largely made up of immigrants of other nationalities, such as Italian and Spanish, in addition to Portuguese.     

A Study of δ-Globin Gene Mutations in the UK Population: Identification of Three Novel Variants and Development of a Novel DNA Test for Hb A′2

We report here the spectrum of δ-globin gene mutations found in the UK population. Nine different δ chain variants and two δ-thalassemia (δ-thal) mutations were characterized in a study of 127 alleles in patients with either a low Hb A₂ value or a split Hb A₂ peak on high performance liquid chromatography (HPLC). The most common δ chain variant was Hb A′₂ (or Hb B₂) [δ16(A13)Gly?→?Arg; HBD: c.49G?>?C] (77.0%), followed by Hb A₂-Yialousa [δ27(B9)Ala?→?Ser; HBD: c.82G?>?T] (12.0%), Hb A₂-Babinga [δ136(H14)Gly?→?Asp; HBD: c.410G?>?A] (3.0%), Hb A₂-Troodos [δ116(G18)Arg?→?Cys; HBD: c.349C?>?T] (1.0%), Hb A₂-Coburg [δ116(G18)Arg?→?His; HBD: c.350G?>?A] (2.0%) and Hb A₂-Indonesia [δ69(E13)Gly?→?Arg; HBD: c.208G?>?C] (1.0%). Three novel variants were identified: Hb A₂-Calderdale [codon 2 (CAT?>?AAT), His?→?Asn; HBD: c.7C?>?A], Hb A₂-Walsgrave [codon 52 (GAT?>?CAT), Asp?→?His; HBD: c.157G?>?C] and Hb A₂-St. George’s [codon 81 (CTC?>?TTC), Leu?→?Phe; HBD: c.244C?>?T]. In addition, two known δ-thal mutations were observed: ?68 (C?>?T); HBD: c.-118C?>?T and codon 4 (ACT?>?ATT); HBD: c.14C?>?T. Amplification refractory mutation system (ARMS) primers were developed to provide a simple molecular diagnostic test for the most common variant, Hb A′₂. Three of the variants had a characteristic HPLC retention time that can be used for a presumptive diagnosis.     

β-Thalassemia gene mutations in Antalya,Turkey: results from a single centre study

β-Thalassemia (β-thal) is a common autosomal recessive disorder resulting from over 300 different mutations of the β-globin genes. Our aim was to create a mutation map of β-thal in the province of Antalya, Turkey. In this study, mutation analysis of a total 146 of β-thal patients followed at the Thalassemia Center of the Antalya Education and Research Hospital, Antalya, Turkey, were included. Direct DNA sequence analysis was performed for mutation scanning of the β-globin gene. One hundred and forty-six patients with β-thal including all types were analyzed, and 14 different β-thal mutations were detected. The most frequently seen mutation was HBB: c.93???21G?>?A [IVS-I-110 (G?>?A)] (52.7%), followed by HBB: .c.92?+?6T?>?C [IVS-I-6 (T?>?C)] (14.4%), HBB: c.?80T?>?A [–30 (T?>?A)] (8.2%), HBB: c.315?+?1G?>?A [IVS-II-1 (G?>?A)] (8.2%), which made up 83.1% of the observed mutations. Our results indicate the importance of micromapping and epidemiology studies of thalassemia, which will assist in establishing the national prevention and control program in Turkey.     

The Univariation and Multiple Linear Regression Analyses for Seventeen SNPs in Thirteen Cardiovascular Disease-Predisposing Genes and Blood Pressure in Chinese Han Males

Blood pressure (BP) is a complex trait regulated by the interaction among multiple physiologic regulatory systems, likely involving numerous genes that lead to inconsistent findings in genetic studies. One possibility of failure to replicate some single-locus results is that the underlying genetics of hypertension is based on multiple genes with minor effects. To learn the association between 17 single nucleotide polymorphisms (SNPs) in 13 cardiovascular disease-predisposing genes and blood pressure of Han males, the 17 SNPs genotypes of 375 Han males were detected and analyzed with BaiO gene chip. The relationship between the SNPs and blood pressure was analyzed with variance analysis and multiple linear regression analysis. Variance analysis and/or multiple linear regression showed that: systolic blood pressure (SBP) was increasing with the elevation of year; AGT²³⁵M, ApoE^112,158E4, and SerpinA3^rs4934A were relative to the increase of SBP; AGT²³⁵M, ET-2⁹⁸⁵G, ApoC3³²⁰⁶T, and ApoE^112,158E4 may have had some relation with diastolic blood pressure (DBP) elevation; and ApoB^Xba?+?was associated with the increase of pulse pressure (PP). These findings support the multigenic nature of the etiology of essential hypertension and propose a potential gene–gene interactive model for future studies.     

Molecular Scanning of β-Thalassemia in the Southern Region of Central Java,Indonesia; a Step Towards a Local Prevention Program

Abstract

Thalassemia is the most prevalent genetic blood disorder worldwide, and particularly prevalent in Indonesia. The purpose of this study was to determine the spectrum of β-thalassemia (β-thal) mutations found in the southern region of Central Java, Indonesia. The subjects of the study included 209 β-thal Javanese patients from Banyumas Residency, a southwest region of Central Java Province. DNA analysis was performed using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), amplification refractory mutation system (ARMS), and the direct sequencing method. The results showed that 14 alleles were found in the following order: IVS-I-5 (G?>?C) (HBB: c.92?+?5G?>?C) 43.5%, codon 26 (Hb E; HBB: c.79G?>?A) 28.2%, IVS-I-1 (G?>?A) (HBB: c.92?+?1G?>?A) 5.0%, codon 15 (TGG?>?TAG) (HBB: c.47G?>?A) 3.8%, IVS-I-1 (G?>?T) (HBB: c.92?+?1G?>?T) 3.1%, codon 35 (–C) (HBB: c.110delC) 2.4%. The rest, including codons 41/42 (–TTCT) (HBB: c.126_129delCTTT), codons 8/9 (+G) (HBB: c.27_28insG), codon 19 (AAC?>?AGC) (HBB: c.59A?>?G), codon 17 (AAG?>?TAG) (HBB: c.52A?>?T), IVS-I-2 (T?>?C) (HBB: c.92?+?2T?>?C), codons 123/124/125 (–ACCCCACC) (HBB: c.370_378delACCCCACCA), codon 40 (–G) (HBB: c.123delG) and Cap +1 (A?>?C) (HBB: c.-50A?>?C), accounted for up to 1.0% each. The most prevalent alleles would be recommended to be used as part of β-thal screening for the Javanese, one of the major ethnic groups in the country.     

Polymorphisms of tumor necrosis factor-�� promoter region for susceptibility to HLA-B27-positive ankylosing spondylitis in Korean population

This study designed to assess the relationship between tumor necrosis factor (TNF)-?? promoter polymorphisms and disease susceptibility to human leukocyte antigen (HLA)-B27-positive ankylosing spondylitis (AS). One hundred and nineteen HLA-B27⁺ AS patients, 95 HLA-B27⁺ healthy controls, and 135 random healthy controls were enrolled in this study. Six single nucleotide polymorphisms (SNPs) of the TNF-?? promoter at positions ?C1031T/C, ?C863C/A, ?C857C/T, ?C646G/A, ?C308G/A, and ?C238G/A were analyzed. Differences between groups were evaluated using the chi-square test or Fisher??s exact test. Haplotypes from each SNP were constructed, and differences in haplotypic frequencies between groups were evaluated. There were significant differences in the allelic and genotypic frequencies of 1031T/C, ?C863C/A, and ?C857C/T TNF-?? promoters polymorphisms between HLA-B27⁺ AS patients and random controls, but not between patients with AS and HLA-B27⁺ healthy individuals. TNF-?? polymorphisms did not influence the extra-spinal clinical features in patients with AS. The haplotypic sequence ?C1031T/?C863C/?C857C/?C308G increased the risk of susceptibility to AS compared to random controls (P _corr?<?0.001, OR?=?2.756, 95% CI?=?1.894?C4.010), whereas the sequence ?C1031C/?C863A/?C857C/?C308G appeared to be associated with decreased susceptibility to AS compared to random controls (P _corr?=?0.006, OR?=?0.396, 95% CI?=?0.231?C0.679). This study indicates that TNF-?? promoter polymorphism between controls and AS patients with HLA-B27⁺ genetic background is not associated with susceptibility to AS. However, TNF-?? polymorphism, irrespective of HLA-B27, increases risk of susceptibility to AS in general population.     

Multiplex Minisequencing of the HBB Gene: A Rapid Strategy to Confirm the Most Frequent β-Thalassemia Mutations in the Tunisian Population

Abstract

The β hemoglobinopathies [β-thalassemia (β-thal) and structural hemoglobin (Hb) variants such as Hb S (HBB: c.20A?>?T) and Hb E (HBB: c.79G?>?A)] are among the most common inherited diseases worldwide. In Tunisia, due to the high prevalence of consanguineous marriages, the recurrent risk of this disease is high. The average prevalence of hemoglobinopathies is 4.48%, reaching 12.50% in some focus regions. The molecular investigations on thalassemia contributed to establishing the spectrum of mutations in the Tunisian population. The total number of HBB gene mutations identified was 24. The two most frequent mutations, codon 39 (C?>?T) (HBB: c.118C?>?T) and IVS-I-110 (G?>?A) (HBB: c.93–21G?>?A) accounted for 70.0% of the total encountered β-thal cases. These two mutations together with IVS-I-2 (T?>?G) (HBB: c.92?+?2T?>?G) and the Hb S variant account for more than 90.0% of all HBB genetic variants in Tunisia. Thus, developing rapid, inexpensive and reliable mutation-specific molecular diagnostic assays targeting our Tunisian populations is our aim to facilitate routine detection of hemoglobinopathies. In this report, we describe the successful application of the multiplex minisequencing assay as an alternative strategy for genetic diagnosis of HBB gene disorders in Tunisia.     

Association and Interaction of −58C>T and ±9 bp Polymorphisms of BDKRB2 Gene Causing Susceptibility to Essential Hypertension

Introduction. Bradykinin, a vasodilator by nature has been documented to have a protective role against hypertension and cardiovascular complications. Polymorphisms of bradykinin B2 receptor (BDKRB2) gene are reported to be predisposing factors for hypertension. Evaluation of the association between ?58C>T and ±9 bp polymorphisms of BDKRB2 with essential hypertension (EHT) was attempted. Methods. Two hundred and fourteen primary hypertensives and 249 controls were genotyped for the selected markers by polymerase chain reaction, gel electrophoresis (±9 bp), and SSCP (?58C>T). Results. While ?58C>T polymorphism did not reveal any association with EHT, ±9 bp polymorphism showed a significant association with high risk for heterozygotes (+9/?9) when tested against the pooled frequencies of homozygotes (OR [odds ratio] = 1.63, 95% confidence interval [CI] = 1.12–2.38, P = .02), and this risk was 1.7 folds high in males (OR = 1.74, 95% CI = 1.05–2.86, P = .06) and 1.9 folds high in familial cases (OR = 1.96, 95% CI = 1.09–3.53, P = .04). In contrast, significant protective effect was observed for ?9/–9 genotype against EHT when tested under dominant model in general (OR = 0.59, 95% CI = 0.41–0.86, P = .01), in males (OR = 0.49, 95% CI = 0.30–0.82, P = .01), and in familial cases (OR = 0.50, 95% CI = 0.28–0.89, P = .04). Significant risk for +9 bp allele was observed in general (OR = 1.39, 95% CI = 1.05–1.86, P = .04) and in males (OR = 1.65, 95% CI = 1.13–2.41, P = .02). The interaction information analysis revealed a synergistic effect between the two polymorphisms contributing to EHT. +9/+9 genotype of ±9 bp polymorphism when present in combination with CC genotype of ?58C>T polymorphism showed 2.2-fold higher risk for developing EHT. Conclusions. The results suggest that allele +9 bp might be a risk factor for EHT in general and specially in males. Markers ?58C>T and ±9 bp may act synergistically causing susceptibility to EHT.     

Angiotensinogen Promoter Polymorphisms Predict Low Diffusing Capacity in U.S. and Spanish IPF Cohorts

Background

Single nucleotide polymorphisms (SNPs) in angiotensinogen (AGT) at positions ?20 and ?6 are associated with increased severity and progression of various fibrotic diseases. Our earlier work demonstrated that the progression of idiopathic pulmonary fibrosis (IPF) was associated with the A-6 allele. This study examined the hypothesis that the homozygous CC genotype at ?20 and the AA genotype at ?6 would confer worse measures of pulmonary function (measured by pulmonary function tests) in IPF.

Methods

Multiple logistic regression analysis was applied to a NIH Lung Tissue Research Consortium cohort and a Spanish cohort, while also adjusting for covariates to determine the effects of these SNPs on measures of pulmonary function.

Results

Analysis demonstrated that the CC genotype at ?20 was strongly associated with reduced diffusing capacity in males in both cohorts (p = 0.0028 for LTRC and p = 0.017 for the Spanish cohort). In females, the AA genotype was significantly associated with lower FVC (p = 0.0082) and V _alv (p = 0.022). In males, the haplotype CA at ?20 and ?6 in AGT was also strongly associated with reduced diffusing capacity in both cohorts.

Conclusions

This study is the first to demonstrate an association of AGT polymorphisms (?20A > C and ?6G > A) with lower measures of pulmonary function in IPF. It is also the first to relate the effect of gender in lung fibrosis with polymorphisms in AGT.     

Molecular Characterization of δ-Thalassemia in Iran

δ-Thalassemia (δ-thal) (OMIM #142000) resulting from mutations on the HBD gene usually has no clinical consequences. However, it may cause the misdiagnosis of β-thalassemia (β-thal) carriers by lowering the Hb A₂ level to the normal range. Therefore, a study for δ-thal should be considered as a step in the detection of at-risk couple in our region. The aim of the present study was to characterize the mutations of the HBD gene in β-thal carriers with normal Hb A₂ levels, and also in normal individuals with Hb A₂ of less than 2.0%. Four β-thal carriers with normal Hb A₂ and 39 individuals with Hb A₂ of less than 2.0% were enrolled. Genomic DNA was extracted by the salting out method and the HBD gene was investigated by polymerase chain reaction (PCR) and direct DNA sequencing. Hb A₂-Yialousa (HBD: c.82?G?>?T) was the most common variant found in the HBD gene, but the following mutations were also found: Hb A₂-NYU (HBD: c.39?T?>?A), Hb A₂-Coburg (HBD: c.350?G?>?A), Hb A₂-Etolia (HBD: c.257?T?>?C), Hb A₂-Fitzroy (HBD: c.428?C?>?A) and the δ-IVS-I-5 (G?>?T) (HBD: c.92?+?5?G?>?T). One case was a compound heterozygote for δ-IVS-I-5/Hb A₂-Fitzroy. The results of this single center study suggest that the mutations in the HBD gene in the Iranian population are heterogeneous and should be considered in genetic counseling of families.     

Molecular Update of β-Thalassemia Mutations in the Syrian Population: Identification of Rare β-Thalassemia Mutations

β-Thalassemia (β-thal) is an autosomal recessive disorder characterized by variable degrees of anemia, bone marrow hyperplasia, splenomegaly, and complications related to the severity of the anemic state. The β-thalassemias result from mutations in and around the β-globin gene (HBB) located as a cluster on the short arm of chromosome 11. In Syria, β-thal is highly prevalent. The main aim of this study was to identify the frequency of HBB mutations in 189 Syrian β-thal patients and carriers of β-thal. Out of the 189 patients and carriers recruited in this study, 181 patients had at least one HBB mutation and eight patients did not show any mutation. The 10 most frequent ones constituted 77.5% of all HBB mutations. These mutations in order of frequency were: IVS-I-110 (G?>?A) (17.0%), IVS-I-1 (G?>?A) (14.7%), codon 39 (C?>?T) (14.4%), IVS-II-1 (G?>?A) (9.8%), codon 8 (–AA) (6.2%), IVS-I-6 (T?>?C) (5.2%), IVS-I-5 (G?>?C) (4.9%), codon 5 (–C) (3.2%), IVS-I-5 (G?>?A) (3.2%) and codon 37 (G?>?A) (2.2%). Another 21 mutations were less frequent or sporadic. These results provide important tools for adapting a prenatal molecular diagnostic test for the Syrian population.     

Molecular Characterization of β-Thalassemia Intermedia in Southeast Iran

Inheritance of mild mutations within the β-globin gene and coinheritance of α-thalassemia (α-thal) are known as two important genetic modifiers in β-thalassemia (β-thal) intermedia (β-TI). We aimed to evaluate the spectrum of β- and α-thal mutations in β-TI patients in Southeast Iran. Common β- and α-globin gene mutations were detected by amplification refractory mutation system–polymerase chain reaction (ARMS–PCR) and multiplex gap-PCR, respectively. There were 26 male (57.8%) and 19 female (42.2%) patients. HBB: c.92?+?5T?>?C [IVS-I-5 (G?>?C)] and HBB: c.?138C?+?1G?>?A [IVS-II-I (G?>?A)] represented the prevalent alleles with respective frequencies of 60.0 and 10.0%. Other β-globin mutations included HBB: c.-138C?>?T [–88 (C?>?T)], HBB: c.27_28insG [frameshift codons (FSC) 8/9 (+G)], HBB: c.46delT [codon 15 (–T)], HBB: c.93-22_95del (IVS-I, 25 del), and the 619?bp deletion (NG_000007.3: g.71609_72227del619). The predominant genotypic combinations were β⁰/β⁰ (68.9%), β⁰/β^+?(8.9%) and β⁺/β^+?(2.2%). Coinheritance of α-thal was observed in 33.0% of the patients, with the –α^3.7 (rightward) (NG_000006.1: g.34164_37967del3804) as the most common deletion (86.0%). One patient was diagnosed with the –α^4.2 (leftward) (AF221717) and one with the – –^MED (g.24664_41064del16401) deletions, while no patients carried the –(α)^20.5 (g.15164_37864del22701), α^–5?nt (HBA2: c.95?+?2_95_6delTGAGG) or codon 19 (–G) (HBA2: c.56delG) mutations. The alleviating molecular mechanism was not explainable by β^+?or concurrent α-thal in more than half of our β-TI patients. This encourages conducting more studies to identify other contributing factors, especially Hb F-inducing genetic modifiers.     

Mutation in a Highly Conserved COOH-Terminal Residue of Krüppel-Like Factor 1 Associated with Elevated Hb F in a Compound Heterozygous β-Thalassemia Patient with a Nontransfusion-Dependent Thalassemia Phenotype

We present a patient with a compound heterozygosity codon 39 (C?>?T) (β⁰) [or β39(C5)Gln→Stop (G39X); CAG?>?TAG; HBB: c.118C?>?T] and –87 (C?>?T) (β⁺) (HBB: c.?137C?>?T) β-globin mutations, a non transfusion-dependent thalassemia phenotype and 97.0% fetal hemoglobin. A novel heterozygous mutation was identified in a highly conserved residue in the COOH-terminus of the Krüppel-like factor 1, R360H, that likely altered DNA-binding and impaired transactivation.