Tetra Amplification Refractory Mutation System-polymerase Chain Reaction Research Articles

Development of T-ARMS-PCR to Detect MYBPC3 Gene Variation in Hypertrophic Cardiomyopathy (HCM) Patients

Hypertrophic cardiomyopathy (HCM) is a common and complex, genetically inherited, cardiovascular disorder. It is typically inherited in an autosomal dominant manner with variable penetrance and mutable expression. Mutations in MYBPC3 gene is one of the genetic causes of HCM. Only 0.2% of general population suffers from HCM. The MYBPC3 gene provides instructions for making cardiac myosin binding protein C, which is imperative for the maintenance and regulation of normal cardiac functions. This study aims to explore the reported SNP rs1052373 from exon 30 of MYBPC3 gene in the population of Punjab, Pakistan. The reported SNP rs1052373 was analysed using Tetra Amplification Refractory Mutation System Polymerase Chain Reaction (T-ARMS-PCR) to find the allelic frequency in the selected population. T-ARMS-PCR is a cost effective, flexible, rapid, and accurate tool for genotyping. The specific sequences of MYBPC3 gene from exons 30 and 31 and introns 29, 30, and 31 were retrieved from NCBI (https://www.ncbi.nlm.nih.gov/). A tetra primer designing tool known as Primer 1 (http://primer1.soton.ac.uk/primer1.html) was used to design the primers for the targeted region of MYBPC3 gene. In this study, the genotyping of previously reported SNP rs1052373 showed variation in the disease group, giving CC, CT, and TT genotypes with the frequency of 0.04. The genotyping analysis of rs1052373 showed that the allelic frequency of homozygous condition T/T was 0.02 and the allelic frequency of heterozygous condition C/T was 0.02 in disease group as compared to the control group. In the latter, the homozygous T/T and heterozygous C/T genotypes were not observed in any individuals. All the individuals in control group carried homozygous C/C genotype. While, the frequency of homozygous C/C genotype was 0.96 in disease group. The findings of this study would help to find novel molecular markers for HCM diagnosis.

Interplay of “leukemia inhibitory factor receptor gene” (rs3099124) polymorphism, leukemia inhibitory factor and ovarian steroids with unexplained infertility

Objective of the study To explore the association of leukemia inhibitory factor receptor (LIFR) gene variant rs3099124, ovarian steroids, and leukemia inhibitory factor with unexplained infertility in Pakistani females. Methodology A case–control investigation in which eighty-one (81) females with unexplained infertility and one hundred and sixty-two (162) fertile counterparts (age and body mass index compared) were recruited between October 2016 and 2018. Ten milliliters of venous blood was collected from all participants. “Genomic DNA” was taken out from lymphocytes in peripheral blood samples. “Tetra Amplification Refractory Mutation System Polymerase Chain Reaction (T-ARMS-PCR)” was constructed through software “Primer-I”. Amplification was carried out by “T-ARMS-PCR” followed by subsequent sequencing for confirmation and extensive consonance. Estradiol, Progesterone and Leukemia Inhibitory Factor (LIF) were measured in serum by ELISA. Results Statistically significant difference was noticed in genotype frequency in “LIFR-gene variant; rs3099124” (χ2 = 28.222, P value < 0.01) between research participants. Although, rs “3099124” “AA” (OR = 0.000; 95%CI = 0–0) and “GA” genotypes (OR = 0.525; 95%CI = 0.226–1.22) showed non-significant safety/protection against unexplained infertility yet minor/risk allele “A” frequency was greater in women with unexplained infertility suggesting a possible explanation of implantation failure. LIF concentration varied between fertile and infertile groups (χ2 = 9.857, P < 0.05) revealing significant threat of unexplained infertility in women with decreased LIF concentration (OR = 2.316, 95%CI = 1.214–4.416). Progesterone was significantly related to unexplained infertility in both study groups (χ2 = 20.347, P < 0.05). High progesterone reduced the possibility of unexplained infertility (OR = 0.306; 95% CI = 0.166–0.567). Conclusion LIFR gene variation (rs3099124) and reduced LIF secretion may cause implantation failure in women with unexplained infertility.

Tetra-ARMS-PCR assay development for genotyping of AGT rs699 T/C polymorphism, its comparison with PCR-RFLP and application in a case-control association study of cardiovascular disease patients

Genetic variations in the AGT gene play a significant role in controlling the plasma concentration of angiotensinogen (precursor protein of bioactive octapeptide angiotensin II) and the efficacy of antihypertensive drugs. In the current study, Tetra–Amplification Refractory Mutation System–Polymerase Chain Reaction (T-ARMS-PCR) was developed for genotyping of AGT rs699 T/C polymorphism and validated through Sanger DNA sequencing. Its efficiency was also tested using 474 human DNA samples [control, n = 181; cardiovascular disease (CVD) patients, n = 293]. Results showed that T-ARMS-PCR is superior to the commonly used PCR-Restriction Fragment Length Polymorphism (PCR-RFLP). Statistical analysis revealed that the AGT rs699 CC genotype is more prevalent in the CVD patient group (37% vs. 28%) and AGT rs699 C allele and CC genotype increased the risk of CVD by 1.4 and 1.9 fold, respectively. In summary, T-ARMS-PCR is the most suitable approach for quick and efficient genotyping of AGT rs699 T/C polymorphism in a large population in resource-limited countries, Furthermore, AGT rs699 T/C polymorphism is associated with the risk of CVD in the Punjabi Pakistani population.

Association of P2X7 receptor genetic polymorphisms and expression with rheumatoid arthritis susceptibility in a sample of the Iranian population: a case-control study.

Rheumatoid arthritis (RA) is a complex inflammatory autoimmune disease with joint eruption, systemic manifestation, and numerous predisposing genetic factors. The P2X7 receptor is an essential ligand-gated channel that contributes to many physiological processes, especially inflammation. However, genetic variations can alter the P2X7 receptor function. Therefore, the present study aimed to explore the impact of P2X7 genetic polymorphisms and expression on susceptibility to RA in a sample of the Iranian population. We enrolled 160 (145 female, 15 male) RA patients and 160 (142 female, 18 male) healthy individuals in this study. Genotyping was performed using tetra amplification refractory mutation system-polymerase chain reaction (TARMS-PCR) for rs1718119, rs2230912, rs2393799, rs28360457, rs35933842, and allele-specific PCR for rs1653624 and rs3751143. Furthermore, 44 new cases of RA and 48 healthy controls were recruited to investigate whether P2X7 mRNA expression is associated with RA susceptibility. The results revealed that the rs2393799 significantly increased the risk of RA in all genetic models (p<0.05), while rs3751143 in codominant (CC vs. AA, OR=0.49, 95% CI=0.26-0.92), dominant (AC+CC, OR=0.59, 95% CI=0.37-0.94), C allele (OR=0.63, 95% CI=0.46-0.88), and rs2230912 in codominant (AG vs. AA, OR=0.56, 95% CI=0.34-0.94), dominant (AG+GG vs. AA, OR=0.59, 95% CI=0.35-0.99), and overdominant (AG vs. AA+GG, OR=0.57, 95% CI=0.33-0.98) significantly decreased the RA risk (p<0.05). Furthermore, the rs1718119 and rs1653624 were not associated with susceptibility of RA (p>0.05), and rs28360457 and rs35933842 were not polymorphic in our study. The mRNA expression level of P2X7 in both groups revealed that the P2X7 gene was significantly upregulated in RA (3.18±0.43) compared to healthy subjects (1.47±0.15, p<0.001). Our results suggest that rs2393799, rs3751143, and rs2230912 variants of the P2X7 gene are associated with RA's susceptibility in a sample of the Iranian population. Also, P2X7 mRNA expression was higher in our new RA patients. The P2X7 receptor has been considered as a potential pharmacologic target in RA. Key Points • P2X7 variants (rs2393799, rs2230912, rs3751143) were associated with RA susceptibility in a sample of the Iranian population. • rs2393799 increases the risk of RA, while rs2230912 and rs3751143 decrease the risk of RA. • P2X7 expression was significantly upregulated in new RA patients compared to controls.

Long non-coding RNA H19 expression and functional polymorphism rs217727 are linked to increased ischemic stroke risk

BackgroundEfforts to identify potential biomarkers for the diagnosis of ischemic stroke (IS) are valuable. The H19 gene plays a functional role in increasing the prevalence of IS risk factors. We evaluated the correlation between H19 rs217727 polymorphism and the expression level of H19 lncRNA with susceptibility to IS among the Iranian population.MethodsBlood samples were collected from IS patients (n = 114) and controls (n = 114). We concentrated on the expression pattern of H19 at different time points (i.e., 0–24, 24–48, and 48–72 h after stroke). The tetra-amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR) method was applied for DNA genotyping. We used the quantitative real-time PCR to evaluate H19 expression levels. We used the receiver operating characteristic (ROC) curve to evaluate the diagnosis and prognosis of IS.ResultsThe rs217727polymorphism of H19 was related with IS susceptibility in the co-dominant (OR = 2.92, 95% CI = 0.91–10.92, P = 0.04) and recessive models (OR = 2.80, 95% CI = 0.96–8.15, P = 0.04). H19 expression was significantly upregulated in IS and remained high for 72 h after stroke. ROC curves showed that H19 expression within the first 24 h from stroke onset might serve as a biomarker for the early diagnosis of IS with 79.49% sensitivity and 80.00% specificity. H19 expression in small vessel occlusion (SVO) and large-artery atherosclerosis (LAA) patients were 3.74 and 3.34 times higher than the undetermined (UD) subtype, respectively [OR = 3.74 95% CL (1.14–12.27) P = 0.030 and OR = 3.34 95% CL (1.13–9.85) P = 0.029].ConclusionThe rs217727 polymorphism of the H19 is correlated with IS susceptibility, and H19 expression levels were higher in SVO and LAA patients. The upregulation of H19 may be considered as a diagnostic biomarker in IS among the Iranian population, but it cannot serve as a useful prognostic marker.

The associations among RARRES2 rs17173608 gene polymorphism, serum chemerin, and non-traditional lipid profile in patients with metabolic syndrome

BackgroundThe adipokine chemerin retinoic acid receptor responder protein 2 (RARRES2) has been associated with insulin resistance, type II diabetes mellitus (T2DM), obesity, and metabolic syndrome (MetS). The impact of RARRES2 rs17173608 gene polymorphism on MetS and chemerin levels is not completely elucidated. This study included 100 patients with MetS and 68 healthy subjects (non-MetS group). The RARRES2 rs17173608 gene variant was analyzed by tetra amplification refractory mutation system polymerase chain reaction (T-ARMS-PCR). Circulating chemerin levels were determined by ELISA. Serum urea, creatinine, fasting blood glucose, glycated hemoglobin, and traditional lipid profile were measured by colorimetric methods. The estimated glomerular filtration rate (eGFR) and non-traditional lipid parameters were calculated.ResultsSerum chemerin levels were significantly higher in MetS than in non-MetS subjects, type II diabetics (T2DM) than non-diabetics, and overweight compared to lean subjects, but it did not differ significantly between patients with and without hypertension. Strikingly, newly diagnosed diabetic patients had significantly higher serum chermerin levels. Correlation and multiple linear regression analysis showed that serum chemerin levels and non-traditional lipid parameters were correlated significantly with the clinical criteria of MetS. Genotyping and allelic frequency distribution of RARRES2 rs17173608 gene polymorphism showed its significant association with MetS. The TT genotype of RARRES2 rs17173608 SNP was more distributed in T2DM in comparison with non-diabetics, and it was associated significantly with higher serum chemerin and higher glycated hemoglobin levels. RARRES2 rs17173608 GG genotype and G allele frequency were less distributed in T2DM patients than in non-diabetic patients.ConclusionsThe RARRES2 rs17173608 SNP might have an impact on chemerin levels and lipid parameters. The GG genotype and G allele may have a protective role towards the risk of T2DM but not for MetS. Serum chemerin and non-traditional lipid profile are significantly associated with MetS.

DETECTIN OF VASPIN RS2236242 AND CHEMERIN RS17173608 GENE POLYMORPHSIMS IN A SAMPLE OF OBESE IRAQI POLYCYCTIC OVARY SYNDROME WOMEN

This study was performed to detect assess Vaspin rs2236242 and Chemerin rs17173608 gene polymorphisms with its association with susceptibility of obese PCOS in Iraqi women .Blood samples from 100 women with obese PCOS and 75 healthy control women matched in age and BMI were collected from AL-Yarmouk Teaching hospital during February2018-July 2018 were enrolled in case control study. Tetra amplification refractory mutation system‐polymerase chain reaction (T-ARMS-PCR) was performed to detect the adipokine SNPs in patients and healthy women under study. Results demonstrated a significant variations in either genotype distribution and alleles frequencies between the obese PCOS and healthy women regarding vaspin rs2236242 polymorphism. The T allele increased the PCOS risk in comparison with the A allele which was the protective against PCOS, as well clear positive correlation between rs17173608 SNP of chemerin and susceptibility of PCOS was identified also, and The T allele had related with elevated PCOS risk as in comparison to the G allele. Data from this work determined for the first time a significant association between vaspin and chemerin SNPs and susceptibility PCOS risk in Iraqi obese women especially T allele .To confirm our findings new studies with large sample size and different ethnicities must be required.

Chemerin rs17173608 Gene Polymorphism is not Associated with Type 2 Diabetes Mellitus: a Cross-sectional Study.

Previous studies have shown that chemerin has important roles in the development of obesity, insulin resistance, metabolic syndrome, polycystic ovary syndrome (PCOS) and T2DM. The main goal of our study was to investigate the role of Chemerin rs17173608 gene polymorphism in T2DM (type 2 diabetes mellitus). 100 patients with T2DM and 50 healthy volunteers were included in the present study. DNA isolation from blood samples was performed with K1820-02 DNA Mini Kit. Chemerin gene polymorphism was detected by Tetra- Amplification Refractory mutation system polymerase chain reaction (T-ARMS-PCR). At the end of T-ARMS-PCR, samples were run using gel electrophoresis. Some samples were validated by sequence analysis. In the genotype analysis, 18.0% of patients had TT genotype and 81.0% of TG genotype was detected. GG genotype was not detected in any patient. Genotype of 1 patient was unidentified. Genotype distribution of healthy control group was 12.0% TT genotype and 88.0% TG genotype. Similar to the T2DM group, the GG genotype was not detected in the control group. There was no statistically significant difference between T2DM group and healthy control group for TG and TT genotypes. To our knowledge, chemerin rs17173608 gene polymorphism has been investigated in T2DM for the first time herein. In the present study, the TT genotype ratios were higher in the T2DM subjects than in healthy subjects. G allele frequency in the T2DM group was lower than that in the control group. However, there was no statistically significant difference between the groups.

Development and application of low-cost T-ARMS-PCR assay for AGT and CYP11B1 gene polymorphisms.

Angiotensin II (Ang II: a truncated octapeptide of angiotensinogen, AGT) and 11-β-hydroxylase influence regulation of blood pressure. Dysregulation of Ang II and 11-β-hydroxylase can lead to hypertension and elevate aldosterone levels. Polymorphisms in AGT (encodes AGT) and CYP11B1 (encodes 11-β-hydroxylase) shift the paradigm from physiological to pathological. Currently, various high-throughput techniques are used to genotype these polymorphisms. These techniques require expensive infrastructure and reagents. However, in developing countries, where cost is the main limiting factor, it is not feasible to use expensive techniques. So, the aim of current study was to develop efficient low-cost method for genotyping of cardiovascular disease and hypertension associated polymorphisms of AGT (rs4762, rs5051) and CYP11B1 (rs6410). For this, tetra amplification-refractory mutation system-polymerase chain reaction (T-ARMS-PCR) method was developed and optimized for aforementioned AGT and CYP11B1 gene polymorphisms. Efficiency of T-ARMS-PCR was tested by genotyping 776 human samples. These T-ARMS-PCR assays were also validated by Sanger DNA sequencing, where 100% concordance was found, allowing the efficient use of these T-ARMS-PCR assays for polymorphism genotyping in AGT and CYP11B1 in resource limited settings. T-ARMS-PCR is low-cost, efficient and reliable assay for genotyping of AGT and CYP11B1 gene polymorphisms.

Association between chemerin rs17173608 and rs4721 gene polymorphisms and gestational diabetes mellitus in Iranian pregnant women

Gestational diabetes mellitus (GDM) is defined as hyperglycemia detected during pregnancy and its risk is increased with obesity. Chemerin, an adipokine, has been proposed as potential mediators of insulin resistance in GDM. This case-control study was designed to assess the relation between chemerin SNPs rs4721 (or rs10278590) and rs17173608 and the development of GDM. One hundred thirty GDM pregnant women with GDM and 160 healthy pregnant women were enrolled in this study. The diagnosis of GDM was based on the International Association of Diabetes and Pregnancy Study Group (IADPSG) criteria. Chemerin rs4721 polymorphism gene was amplified through PCR, and SNP was detected using restriction enzyme AluI. Genotyping for chemerin rs17173608 polymorphism was performed by using tetra-amplification refractory mutation system polymerase chain reaction (T-ARMS-PCR). Blood glucose level was measured by an enzymatic method. Our finding showed that the genotypes frequency of chemerin rs4721 polymorphism was significantly different between GDM and non-GDM groups (χ2 = 7.44, P = 0.02). The genotype of rs4721 was significantly associated with GDM in co-dominant and dominant genotypes (GG vs GT, OR = 2.3, 95%CI = 1.24–4.24, P = 0.008, and GG vs GT + TT, OR = 2.21, 95%CI = 1.23–3.99, P = 0.008, respectively). No significant difference was observed in allele frequency between case and control groups (P = 0.62). Moreover, the genotypes and allele frequencies of chemerin rs17173608 polymorphism did not show significant differences between GDM and non-GDM (P > 0.05). We concluded that the genotype of rs4721 was found to contribute significant risk to GDM while genotype of rs17173608 could not predict the risk of GDM.

Association of miR-146a rs2910164 and miR-222 rs2858060 polymorphisms with the risk of polycystic ovary syndrome in Iranian women: A case–control study

ObjectiveToday, many single nucleotide polymorphisms in microRNA genes are known to alter the microRNA expression levels or processing causing susceptibility of several human diseases. The present study aimed to investigate the association of microRNA-146a (rs2910164) and microRNA-222 (rs2858060) polymorphisms with susceptibility to polycystic ovary syndrome (PCOS) in an Iranian population. Materials and methodsThis case–control study was performed on 205 patients with PCOS and 205 normal women as the control group. After DNA extraction, Tetra-amplification refractory mutation system polymerase chain reaction (T-ARMS-PCR) was used to detect the polymorphisms. The association between genotypes and the risk of PCOS was examined by odds ratios (OR) and 95% of confidence intervals (CIs). ResultsOur results showed that there are significant differences in CG genotype frequencies between case and control groups regarding miR-146a rs2910164 polymorphism (OR = 2.03, CI = 1.3–3, P = 0.001). In a dominant model for the C allele, CC + CG genotypes were associated with PCOS risk (OR = 2, 95% CI = 1.3–2.9, P = 0.001) and the C allele increased the risk of PCOS (OR = 1.6, 95% CI = 1.1–2.1, P = 0.004). Furthermore, a positive association was observed between miR-222 CG genotype and the risk of PCOS (OR = 2.2, 95% CI = 1.1–4.1, P = 0.02). These results were evident after adjustment for age and body mass index. ConclusionThe present results suggest that the miR-146a rs2910164 and miR-222 rs2858060 polymorphisms are associated with an increased risk of PCOS. Therefore, both polymorphisms could play an important role as a genetic risk factor for development of PCOS in the Iranian population.

Obesity risk prediction among women of Upper Egypt: The impact of serum vaspin and vaspin rs2236242 gene polymorphism

BackgroundVaspin is an adipokine that is potentially linking obesity, insulin resistance, metabolic syndrome and type-2 diabetes. AimThe present study aimed to investigate the impact of vaspin rs2236242 gene polymorphism on the risk of obesity, diabetes, their metabolic traits, and serum vaspin levels in a sample of Upper Egyptian women. Subjects and methodsA total of 224 subjects, 112 obese (62 non diabetics, 50 diabetics) and 112 controls were included in this case control study. Vaspin gene rs2236242 polymorphism was performed using tetra-amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR) and serum vaspin levels were estimated by ELISA. ResultsThe minor (A) allele of vaspin rs2236242 gene polymorphism was significantly lower in obese (30.8%) than controls (43.7%) (P=0.005). The protective effect was evident in dominant and recessive inheritance models (TT vs TA+AA, P=0.004 and TT+TA vs AA, P=0.036). After adjusting genotypes for diabetes there were no significant association between vaspin rs2236242 gene polymorphism and obesity but significant association was maintained in the obese diabetics. Vaspin serum levels were found to be lower in minor protective (AA) genotype carriers than the other two genotypes (P<0.001). In the mean-time serum vaspin levels were significantly higher in obese diabetics and non-diabetics than controls (P<0.001 each).There were significant positive correlations between vaspin levels and hs-CRP, cholesterol, LDL-C, fasting glucose, HOMA-IR, insulin, and ALT values (P<0.05 each) and a negative correlation with HDL-C (P<0.01). ConclusionThe minor A allele of vaspin rs2236242 polymorphism plays a protective role against obesity and diabetes but this relation is largely ascribed to its effect on insulin resistance. The serum vaspin concentration was lower in minor protective allele carriers. To the best of our knowledge, this is the first study of vaspin SNP in Upper Egyptian women. The entire understanding of vaspin intimate mechanistic action might enable the development of novel etiology-based treatment strategies for obesity, the complex genetic trait.

Association Between IL12A rs568408, IL12B rs3212227 and IL-12 Receptor rs383483 Polymorphisms and Risk of Pulmonary Tuberculosis

Background: Interleukine-12 (IL-12) induced interferon- (IFN- ) production and development of T cells into Th1 cells, which has a critical role in immunity to intracellular pathogens. Objectives: The current study aimed to evaluate the possible association between IL12A rs568408, IL12B rs3212227 as well as IL-12 receptor (IL-12R) rs383483 polymorphisms and pulmonary tuberculosis (PTB). Methods: This case-control study was conducted on 174 confirmed PTB patients and 177 healthy subjects in a sample of southeast Iranian population. Genotyping was performed by the tetra amplification refractory mutation system- polymerase chain reaction (T-ARMS-PCR) method. Results: The frequencies of GG, GA and AA genotypes of IL12A rs568408 variant in cases and controls were 58.9%, 38.5%, 1.7% and 61.6%, 37.3%, 1.1%, respectively. Regarding rs3212227 variant, the frequencies of AA, AC and CC genotypes in cases and controls were 47.1%, 54.4%, 7.5% and 50.8%, 43.5%, 5.7%, respectively. Concerning IL12R rs383483 polymorphism, the frequencies of AA, AG and GG in cases and controls were 39.1%, 31.4%, 29.5% and 40.2%, 25.3%, 34.5%, respectively. There was no significant difference in allele and genotype frequency of IL12A rs568408, IL12B rs3212227 and IL-12R rs383483 polymorphisms between patients with PTB and control groups (P > 0.05). Conclusions: The findings of the current study show that neither IL12A rs568408 and IL12B rs3212227 nor IL-12R rs383483 polymorphisms are associated with the risk of PTB in our population. Further studies with larger sample sizes and various ethnicities are needed to certify our findings.

INTERLEUKIN-18 GENE POLYMORPHISM AND SOME RISK FACTORS IN IRAQI PATIENTS WITH BREAST CANCER

ABSTRACTObjective: Breast cancer is the most diagnosed cancer in women, which leads to death in a lot of women with breast cancer. The major risk factorsassociated with breast cancer risk related to family history, age, clinical history, lifestyle factors, long-period hormonal exposure, and single nucleotidepolymorphisms in many genes showed possible links with breast cancer incidence risk in different people populations. Our study aimed to figure outthe correlation between smoking, lodging and family history, and other factors with the risk of breast cancer.Methods: Blood sample from female patients with breast cancer and healthy individuals were collected and subjected to tetra-amplification refractorymutation system–polymerase chain reaction (T-ARMS-PCR) technique for −607 C/A mutation of an interleukin (IL-18) gene and SPSS 18 softwareanalyzed the results statically.Results: Results showed no association between lodging and smoking with risk of breast cancer, (p&gt;0.05), while the association between the risk andfamily history were obvious (p&lt;0.05).Conclusion: The results obtained by T-ARMS-PCR technique did not show the association between −607 C/An alternation of IL-18 gene and breastcancer (p&gt;0.05) in the individuals examined in our study.Keywords: Interleukin-18, Gene, Polymorphism, Tetra-amplification refractory mutation system–polymerase chain reaction, Mutations.

TGFβ1 genetic variants are associated with an increased risk of acute brucellosis

Objective: Cytokines play a critical role in the regulation of the immune response against brucellosis infection, and mediate production of many pro- and anti-inflammatory signals. Transforming growth factor-beta 1 (TGFβ1), a powerful suppressive cytokine, inhibits macrophage activation and modulates T-cell function, and plays crucial roles in regulation of microbial replication and host responses to brucella. Methods: The association of three polymorphisms in the TGFβ1 gene (–509 C/T [rs1800469], + 868 C/T [rs1800470], and + 913 G/C [rs1800471]) in promoter, codons 10 and 25, respectively, with brucellosis infection was evaluated. This case-control study was performed on a total of 281 Iranian subjects including 153 patients with active brucellosis and 128 age- and sex-matched healthy individuals as controls. Genotyping for the TGFβ1 –509 C/T and + 868 C/T variants was performed using tetra amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR). Also, the + 913 G/C polymorphism was genotyped using an allele-specific PCR. Results: The results demonstrated that the TGFβ1 + 868 C/T mutant homozygote genotype (TT vs CC), was a risk factor for developing brucellosis in the co-dominant and recessive models (odds ratio (OR) = 2.60, p = 0.023; OR = 2.602, p = 0.014, respectively). Additionally, the diplotype analyses revealed that TGFβ1 codon 10 and 25 diplotype, TT/GG, was associated with an increased risk of brucellosis (OR = 2.49, p = 0.038). Other TGFβ1 variants did not increase the risk of brucellosis infection. Conclusions: Our findings propose that TGFβ1 + 868 TT genotype and TT/GG diplotype may confer increased risk of brucellosis in the examined population.

Apelin rs2235306 polymorphism is not related to metabolic syndrome in Egyptian women

BackgroundApelin is an adipokine that was identified to play a role in the control of glucose homeostasis. Apelin rs2235306 gene polymorphism was linked to insulin resistance and poor glycemic control. Aim of the studyTo assess the relation of apelin rs2235306 polymorphism with metabolic syndrome and its component traits in Egyptian women from Suez Canal area. Subjects and methodsThe study included 100 metabolic syndrome patients and 100 healthy female subjects. The component traits of metabolic syndrome were determined and the genotypes of the polymorphisms were assessed using tetra amplification refractory mutation system polymerase chain reaction (T-ARMS-PCR) technique. ResultsThere was no significant difference in the allele frequencies between the metabolic syndrome and control groups (P=0.841). There was also no association of the different genotypes of this polymorphism with any of the component traits of metabolic syndrome. ConclusionApelin rs2235306 polymorphism is not associated with the incidence of metabolic syndrome in the studied population.

TIRAP rs8177374 gene polymorphism increased the risk of pulmonary tuberculosis in Zahedan, southeast Iran

ObjectiveTo evaluate the possible association between Toll-interleukin 1 receptor (TIR) domain containing adaptor protein (TIRAP; also known as MAL) rs1893352 and rs8177374 (S180L) gene polymorphisms and pulmonary tuberculosis (PTB) in a sample of Iranian population. MethodsThis case-control study was performed on 174 PTB and 177 healthy subjects. Tetra amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR) was used to detect the polymorphisms. ResultsOur finding showed that neither the overall Chi-square comparison of PTB and control subjects nor the logistic regression analysis indicated any association between rs1893352 polymorphism and PTB. Regarding rs8177374 polymorphism, the CT genotype as well as CT+TT increased the risk of PTB in comparison with CC genotype (OR=4.73, 95% CI=2.65–8.45, P<0.0001 and OR=6.47, 95% CI=3.68–11.38, P<0.0001, respectively). The rs8177374 T allele increased the risk of PTB in comparison with C allele (OR=4.21, 95% CI=2.43–7.26, P<0.0001). ConclusionsOur finding indicates that TIRAP rs8177374 polymorphism is associated with PTB in a sample of Iranian population.

Association Between TLR8 and TLR9 Gene Polymorphisms and Pulmonary Tuberculosis

Background: Tuberculosis is still a health problem throughout the world. Both genetic and environmental factors may contribute the susceptibility to tuberculosis. Toll-like receptors play a critical role in the recognition of mycobacterium tuberculosis (TB). Objectives: The aim of this study was to evaluate the possible association between TLR8 rs3764880 and TLR9 rs148805533 polymorphisms and pulmonary tuberculosis (PTB) in a sample of Iranian population. Patients and Methods: In this study, blood samples of 320 subjects including 160 PTB patients and 160 healthy subjects were collected. DNA was extracted and TLR8 rs3764880 polymorphism was analyzed by Tetra Amplification Refractory Mutation System-Polymerase Chain Reaction (TARMS-PCR) and TLR9 rs148805533 polymorphisms was analyzed by allele specific PCR. Results: The allelic and genotypic frequencies of the TLR8 rs3764880 did not differ significantly between PTB and the controls. No significant difference was found between the groups regarding TLR9 rs148805533 polymorphism. Conclusions: TLR8 rs3764880 and TLR9 rs148805533 polymorphisms may not be risk factors for susceptibility to tuberculosis in a sample of Iranian population. Larger studies are required to validate our findings.

Stereological Analysis of Interdental Gingiva in Chronic Periodontitis Patients With Tumor Necrosis Factor-alpha (-308 G/A) Gene Polymorphisms

Background: The association between tumor necrosis factor-alpha (TNF-α) (-308 G/A) gene polymorphisms and level of tissue destruction and periodontal disease progression is not proved. Objectives: The present study investigated the stereological parameters of interdental papilla in chronic periodontitis (CP) patients with TNF-α (-308 G/A) gene polymorphisms. Materials and Methods: Sixty gingiva samples were studied. After determination of TNF-α (-308 G/A) gene polymorphisms using a tetraamplificationrefractory mutation system-polymerase chain reaction (T-ARMS-PCR) technique, 45gum tissues of CP patients (40 GG, 4 GA+1 AA genotype) were considered as the case group. Fifteencontrol samples were also collected from healthy individuals. Interdental gingiva tissues were fixed in Lilli’s solution and after tissue processing were exhaustively sectioned into 4μm thick sections. Ten to 13 sections were sampled by systematic uniform random sampling (SURS) , stained with Masson’s trichrome and then the volume fractions; volume per unit volume (Vv) of the gingival components were estimated using Cavalier’s point counting method. Results: There were statistically significant differences in the Vv of epithelium, connective tissue, collagenous and non-collagenous matrix and blood vessels between the control and CP groups (P < 0.0001) . There were no statistically significant differences in the Vv of epithelium, connective tissue of gingiva, collagenous and non-collagenous matrix and blood vessels between GG, GA, and AA genotypes in CP patients (P > 0.05) . Conclusions: Results of the current study showed that there wasno association between TNF-α (-308 G/A) gene polymorphisms and quantitative parameters of interdental papilla in CP patients.

Association between vaspin rs2236242 gene polymorphism and polycystic ovary syndrome risk

Vaspin, an adipocytokine that has been isolated from the visceral adipose tissue, is a member of the serine protease inhibitor family. In humans, serum vaspin levels are correlated with body mass index (BMI) and obese women with polycystic ovary syndrome (PCOS). The present study is the first investigation to examine the association between vaspin rs2236242 gene polymorphism and risk of PCOS in Iranian patients. This case–control study was performed on 150 patients with PCOS and 150 healthy women. The vaspin genotypes were determined using tetra-amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR). Our finding showed that there are significant differences in genotype frequencies between case and control group regarding vaspin rs2236242 polymorphism (OR=0.59, CI=0.37–0.95, p=0.03). The A allele decreased the risk of PCOS (OR=0.67, CI=0.46–0.96, p=0.03) as compared to the T allele. There was no significant association between vaspin rs2236242 gene polymorphism and PCOS after adjusting genotypes for BMI. In conclusion, our data suggest a significant association between vaspin rs2236242 polymorphism and the PCOS but this relationship is affected by obesity status.