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Background & Aims: A genome-wide significant association between anti–Helicobacter pylori (H pylori) IgG titers and Toll-like receptor (TLR1/6/10) locus on 4p14 was demonstrated for individuals of European ancestry, but not uniformly replicated. We re-investigated this association in an updated genome-wide association study (GWAS) meta-analysis for populations with low gastric cancer incidence, address potential causes of cohort heterogeneity, and explore functional implications of genetic variation at the TLR1/6/10 locus. Methods: The dichotomous GWAS (25% individuals exhibiting highest anti–H pylori IgG titers vs remaining 75%) included discovery and replication sampls of, respectively, n = 15,685 and n = 9676, all of European ancestry. Longitudinal analysis of serologic data was performed on H pylori–eradicated subjects (n = 132) and patients under surveillance for premalignant gastric lesions (n = 107). TLR1/6/10 surface expression, TLR1 mRNA, and cytokine levels were measured in leukocyte subsets of healthy subjects (n = 26) genotyped for TLR1/6/10 variants. Results: The association of the TLR1/6/10 locus with anti–H pylori IgG titers (rs12233670; β = −0.267 ± SE 0.034; P = 4.42 × 10−15) presented with high heterogeneity and failed replication. Anti–H pylori IgG titers declined within 2–4 years after eradication treatment (P = 0.004), and decreased over time in patients with premalignant gastric lesions (P < 0.001). Variation at the TLR1/6/10 locus affected TLR1-mediated cytokine production and TLR1 surface expression on monocytes (P = 0.016) and neutrophils (P = 0.030), but not mRNA levels. Conclusions: The association between anti–H pylori IgG titers and TLR1/6/10 locus was not replicated across cohorts, possibly owing to dependency of anti–H pylori IgG titers on therapy, clearance, and antibody decay. H pylori–mediated immune cell activation is partly mediated via TLR1 signaling, which in turn is affected by genetic variation.

The severity of COVID-19 disease is partly determined by host genetic factors that have been reported by GWAS. We evaluated nine previously reported genome-wide significant associations regardless of the disease severity in a representative sample from the population of Latvia. Our cohort consisted of 475 SARS-CoV-2 positive cases, from which 146 were hospitalized individuals and 2217 controls. We found three variants from Neanderthal introgression event at the 3p21.31 region to be significantly associated with increased risk of SARS-CoV-2 infection and hospitalization status. The strongest association was displayed by rs71325088 with Bonferroni adjusted P=0.007, OR=1.46 [95% CI 1.17-1.81]. We performed fine-mapping by exploring 1 Mb region at 3p21.31 locus and identified 9 SNPs with even lower p-values with the strongest association estimated for rs2191031 P=5e-05, OR = 1.40[CI 95% 1.19-1.64] located in the LZTFL1. We show clear replication of 3p.21.31 locus in an independent cohort which favors further functional investigation of leading variants.

ObjectiveCirculating miRNAs are found in bodily fluids including plasma and can serve as biomarkers for diseases. The aim of this study was to provide the first insight into the landscape of circulating miRNAs in close proximity to the adrenocorticotropic hormone (ACTH) secreting PitNET. To achieve this objective next-generation sequencing of miRNAs in plasma from bilateral inferior petrosal sinus sampling (BIPSS) - a gold standard in diagnosing ACTH-secreting PitNETs, was carried out.MethodsSinistral (left) and dextral (right) BIPSS blood samples of the patient were collected in three time points: before the administration of corticotropin-releasing hormone, 5 and 15 minutes after stimulation. Peripheral venous blood samples were also collected 24 hours before and after BIPSS and before the resection of PitNET and 24 hours after. In differential expression analysis sinistral plasma was compared with dextral.ResultsBIPSS concluded that the highest amount of ACTH was released in the sinistral side at the 5th minute mark indicating a presence of tumor. The highest amount of differentially expressed miRNAs was observed 5 minutes after stimulation (20 upregulated, 14 downregulated). At the 5th minute mark in sinistral plasma, two miRNAs were identified: hsa-miR-7-5p and hsa-miR-375-3p that were highly upregulated compared to other BIPSS samples and peripheral plasma samples. Clustering analysis showed that BIPSS plasma differs from peripheral plasma in miRNA expression patterns.Conclusionsdata indicates that ACTH-secreting PitNET actively releases two circulating miRNAs upon stimulation with CRH (hsa-mir-7-5p, hsa-mir-375-3p) alongside with ACTH implying further studies of these miRNA as diagnostic markers are needed.

Simple Summary Pituitary neuroendocrine tumors are non-cancerous tumors of the pituitary gland, that may overproduce hormones leading to serious health conditions or due to tumor size cause chronic headache, vertigo or visual impairment. In recent years pituitary neuroendocrine tumors are studied with the latest molecular biology methods that simultaneously investigate a large number of factors to understand the mechanisms of how these tumors develop and how they could be diagnosed or treated. In this review article, we have studied literature reports, compiled information and described molecular factors that could affect the development and clinical characteristics of pituitary neuroendocrine tumors, discovered factors that overlap between several studies using large scale molecular analysis and interpreted the potential involvement of these factors in pituitary tumor development. Overall, this study provides a valuable resource for understanding the biology of pituitary neuroendocrine tumors. Abstract Pituitary neuroendocrine tumors (PitNETs) are non-metastatic neoplasms of the pituitary, which overproduce hormones leading to systemic disorders, or tumor mass effects causing headaches, vertigo or visual impairment. Recently, PitNETs have been investigated in large scale (exome and genome) molecular analyses (transcriptome microarrays and sequencing), to uncover novel markers. We performed a literature analysis on these studies to summarize the research data and extrapolate overlapping gene candidates, biomarkers, and molecular mechanisms. We observed a tendency in samples with driver mutations (GNAS, USP8) to have a smaller overall mutational rate, suggesting driver-promoted tumorigenesis, potentially changing transcriptome profiles in tumors. However, direct links from drivers to signaling pathways altered in PitNETs (Notch, Wnt, TGF-β, and cell cycle regulators) require further investigation. Modern technologies have also identified circulating nucleic acids, and pinpointed these as novel PitNET markers, i.e., miR-143-3p, miR-16-5p, miR-145-5p, and let-7g-5p, therefore these molecules must be investigated in the future translational studies. Overall, large-scale molecular studies have provided key insight into the molecular mechanisms behind PitNET pathogenesis, highlighting previously reported molecular markers, bringing new candidates into the research field, and reapplying traditional perspectives to newly discovered molecular mechanisms.

Background Pituitary adenomas (PA) have an increased potential for relapse in one to 5 years after resection. In this study, we investigated the genetic differences in genomic DNA of primary and rapidly recurrent tumours in the same patient to explain the causality mechanisms of PA recurrence. Case presentation The patient was a 69-year-old female with non-functional pituitary macroadenoma with extension into the left cavernous sinus (Knosp grade 2) who underwent craniotomy and partial resection in August 2010. Two years later, the patient had prolonged tumour growth with an essential suprasellar extension (Knosp grade 2), and a second craniotomy with partial tumour resection was performed in September 2012. In both tumours, the KI-67 level was below 1.5%. Exome sequencing via semiconductor sequencing of patient germline DNA and somatic DNA from both tumours was performed. Tmap alignment and Platypus variant calling were performed followed by variant filtering and manual review with IGV software. We observed an increased load of missense variants in the recurrent PA tumour when compared to the original tumour. The number of detected variants increased from ten to 26 and potential clonal expansion of four variants was observed. Additionally, targeted SNP analysis revealed five rare missense SNPs with a potential impact on the function of the encoded proteins. Conclusions In this case study, an SNP located in HRAS is the most likely candidate inducing rapid PA progression. The relapsed PA tumour had a higher variation load and fast tumour recurrence in this patient could be caused by clonal expansion of the leftover tumour tissue.

Objective Although pituitary adenomas (PAs) affect a significant proportion of the population, only a fraction have the potential to become clinically relevant during an individual’s lifetime, causing hormonal imbalance or complications due to mass effect. The overwhelming majority of cases are sporadic and without a clear familial history, and the genotype–phenotype correlation in PA patients is poorly understood. Our aim was to investigate the involvement of genes known for their role in familial cases on drug response and tumor suppression in the development and pathology of PAs in a patient group from Latvia. Design The study included 143 cases and 354 controls, we investigated the role of single-nucleotide polymorphisms (SNPs) in seven genes (SSTR2, SSTR5, DRD2, MEN1, AIP, GNAS, and PRKAR1A) associated with pituitary tumor occurrence, phenotype, and clinical symptoms. Methods Genotyping of 96 tag and nonsynonymous SNPs was performed in the genomic regions of interest. Results We discovered a significant association (OR=17.8, CI 0.95=2.18–145.5, P=0.0002) between a rare MEN1 mutation (rs2959656) and clinically active adenoma in our patients. Additionally, rs7131056 at DRD2 was associated with a higher occurrence of extrasellar growth in patients with prolactinoma and somatotropinoma (OR=2.79, CI 0.95=1.58–4.95, P=0.0004). Conclusions rs2959656, a nonsynonymous variant in MEN1, is associated with the development of clinically active PA. Furthermore, rs7131056 in DRD2 contributes to either faster growth of the adenoma or reduced symptomatic presentation, allowing PAs to become larger before detection.

Background and objective: North Carolina macular dystrophy (NCMD) is a very rare autosomal dominant hereditary disease. Up to date there are three types of NCMD described and consequently named macular dystrophy, retinal: MCDR1, MCDR2 and MCDR3. The aim of this study was to perform linkage and copy number variation analysis for the family affected by NCMD followed by the selected candidate gene sequencing. Materials and methods: This study concerned a 3-generation, non-consanguineous Latvian family with NCMD. Genome-wide scan, copy number variation and non-parametric linkage analysis was performed. Analysis resolved the locus of interest to the 5p15.33 region. Two of the genes, iroquois homeobox 2 (IRX2) and iroquois homeobox 4 (IRX4), were selected and sanger sequencing was performed. Results: Linkage analysis indicated a region on chromosome 5 for the analyzed family, corresponding to a genetic locus previously described for MCDR3 (5p15-p13). Chromosomal aberrations were not identified in the affected family members. An upstream intron variant (NM_001278634: c.-139G &gt, A (rs6876836)) in IRX4 gene segregated with NCMD phenotype in the analyzed family. Conclusions: It is unlikely to be the causative mutation of NCMD due to its high minor allele frequency 0.3532. Therefore, the role of IRX2 and IRX4 genes in the pathogenesis of NCMD has not been proved. Considerable variability in visual acuity between individuals of the same age group in all the families examined was noted. No overlap between NCMD grade and family generation was seen in the family described in the present study.

We read with interest the article by Passamonti et al ,1 reporting the performance of two different faecal immunochemical tests (FITs) highlighting the importance of standardisation and validation of screening methodologies. Conventionally, laboratory-based FIT is the preferred approach in testing for occult blood in faeces, which includes colorectal cancer screening programmes.2–4 The potential of preserving stable faecal samples in a widely used FIT buffer for microbiome research would enable prospective microbiome studies in generally healthy subjects undergoing colorectal cancer screening. For this purpose, we evaluated faecal sample stability in the commonly used OC-Sensor (Eiken Chemical, Tokyo, Japan) under various storage conditions. Faecal samples from five healthy adult individuals were used for the analysis and exposed to 16 different conditions: immediately frozen at −86°C (with FIT (wFIT)/without FIT (woFIT)); immediately frozen at −20°C (wFIT/woFIT); wFIT stored at 4°C for 1, 2, 7, 14 days; stored at 20°C for 1, 2, 7, 14 days; stored at 30°C for 1, 2, 7 days and lastly stored at 4°C for 2 days and at 20°C for additional 2 days. Shotgun metagenomic analysis was performed by Illumina 2500, while taxonomic compositions were determined by 16S rRNA analysis employing Illumina MiSeq and Ion Torrent PGM. Our first attempt to extract DNA from wFIT samples failed due to low amount of DNA obtained. Thus, lyophilisation for all of the wFIT samples, except for woFIT samples, was applied, increasing the yield of DNA up to 30 times. As shown by the figure 1, the Shannon index in both platforms displayed a clear pattern of …

Purpose In this report, we summarise data on BRCA1 gene analysis in Latvia to characterise criteria of genetic testing for breast and ovarian cancer susceptibility. Material/methods Analysis by SSCP/HD, MALDI-TOF mass spectrometry or DNA sequencing was used for mutation detection. Mutations identified were confirmed by direct DNA sequencing. Results Out of 1068 breast and 231 ovarian cancer patients from different families: 58 carried the c.5266dupC and 43 carried the c.4035delA mutations. Every 4th patient in our study did not report cancer in the family. The breast cancer was diagnosed earlier in carriers of the c.5266dupC than in carriers of the c.4035delA ( p = 0.003). The incidence of breast or ovarian cancer does not differ among the 2 mutation carriers in our patient group. The nature of the c.5266dupC mutation might be more deleterious. Conclusions We recommend the screening of 4 founder BRCA1 mutations in all breast and ovarian cancer patients in Latvia at diagnosis of disease regardless of family history or age. The BRCA1 screening can be carried out efficiently using the MALDI-TOF mass spectrometry mutation detection method developed in the Biomedical Research and Study Centre (Riga, Latvia).

Melanocortin 4 receptor (MC4R) is an important regulator of food intake and number of studies report genetic variations influencing the risk of obesity. Here we explored the role of common genetic variation from MC4R locus comparing with SNPs from gene FTO locus, as well as the frequency and functionality of rare MC4R mutations in cohort of 380 severely obese individuals (BMI39 kg/m(2)) and 380 lean subjects from the Genome Database of Latvian Population (LGDB). We found correlation for two SNPs--rs11642015 and rs62048402 in the fat mass and obesity-associated protein (FTO) with obesity but no association was detected for rs17782313 located in the MC4R locus in these severely obese individuals. We sequenced the whole gene MC4R coding region in all study subjects and found five previously known heterozygous non-synonymous substitutions V103I, I121T, S127L, V166I and I251L. Expression in mammalian cells showed that the S127L, V166I and double V103I/S127L mutant receptors had significantly decreased quantity at the cell surface compared to the wild type MC4R. We carried out detailed functional analysis of V166I that demonstrated that, despite low abundance in plasma membrane, the V166I variant has lower EC50 value upon αMSH activation than the wild type receptor, while the level of AGRP inhibition was decreased, implying that V166I cause hyperactive satiety signalling. Overall, this study suggest that S127L may be the most frequent functional MC4R mutation leading to the severe obesity in general population and provides new insight into the functionality of population based variants of the MC4R.

Variations in the FTO gene and near the TMEM18 gene are risk factors for common form of obesity, but have also been linked with type 2 diabetes (T2D). Our aim was to investigate the contribution of these variants to risk of T2D in a population in Latvia. Four single nucleotide polymorphisms (SNP) in the first and fourth intronic regions of FTO and one close to TMEM18 were genotyped in 987 patients with T2D and 1080 controls selected from the Latvian Genome Data Base (LGDB). We confirmed association of SNPs in the first intron (rs11642015, rs62048402 and rs9939609) of FTO and rs7561317 representing the TMEM18 locus with T2D. Association between SNP in FTO and T2D remained significant after correction for body mass index (BMI). The rs57103849 located in the fourth intron of FTO and rs7561317 in TMEM18 showed BMI independent association with younger age at diagnosis of T2D. Our results add to the evidence that BMI related variants in and near FTO and TMEM18 may increase the risk for T2D not only through secondary effects of obesity. The influence of variants in the fourth intron of the FTO gene on development of T2D may be mediated by mechanisms other than those manifested by SNPs in the first intron of the same gene.

Pituitary adenomas are one of the most common endocrine and intracranial neoplasms. Although they are theoretically monoclonal in origin, several studies have shown that they contain different multipotent cell types that are thought to play an important role in tumor initiation, maintenance, and recurrence after therapy. In the present study, we isolated and characterized cell populations from seven pituitary somatotroph, nonhormonal, and lactotroph adenomas. The obtained cells showed characteristics of multipotent mesenchymal stromal cells as observed by cell morphology, cell surface marker CD90, CD105, CD44, and vimentin expression, as well as differentiation to osteogenic and adipogenic lineages. They are capable of growth and passaging under standard laboratory cell culture conditions and do not manifest any hormonal cell characteristics. Multipotent mesenchymal stromal cells are present in pituitary adenomas regardless of their clinical manifestation and show no considerable expression of somatostatin 1–5 and dopamine 2 receptors. Most likely obtained cells are a part of tissue-supportive cells in pituitary adenoma microenvironment.

The purinergic 1 receptor (P2RY1) has been implicated in development of heart disease and in individual pharmacodynamic response to anticoagulant therapies. However, the association of polymorphisms in the P2RY1 gene with myocardial infarction (MI), and its associated conditions, has yet to be reported in the literature. We evaluated seven known SNPs in P2RY1 for association with MI in a Latvian population. Seven independent parameters that are related to MI [body mass index (BMI), type 2 diabetes (T2D), angina pectoris, hypertension, hyperlipidemia, atrial fibrillation and heart failure] were investigated. No significant association with MI was observed for any of the polymorphisms. Those SNPs for which the P value was close to significance were located in coding or promoter regions. Intriguingly, carriers of the minor allele in the P2RY1 gene locus showed a tendency towards higher onset age for MI, suggesting a possible protective effect of these SNPs against MI or their contribution in progression as opposed to onset. Finally, a linkage disequilibrium (LD) plot was generated for these polymorphisms in the Latvian population. The results of this study suggest that the role of P2RY1 in individuals from Latvian population is likely to be principally involved in platelet aggregation and thromboembolic diseases, and not as a significant contributing factor to the global metabolic syndrome.

Analysis of Polymorphisms at the Adiponectin Gene Locus in Association with Type 2 Diabetes, Body Mass Index and Cardiovascular Traits in Latvian PopulationDespite the number of recently conducted studies seeking to determine the association between genetic variants of adiponectin gene and susceptibility to type 2 diabetes (T2D) and increased body mass index (BMI), the results obtained are often inconsistent. To determine the impact of common polymorphisms in promoter and coding regions of adiponectin gene on these conditions in Latvian population, we selected ten SNPs (rs2241767, rs1501299, rs3777261, rs16861210, rs2241766, rs822396, rs182052, rs17300539, rs16861194, rs266729) based on haploblock structure and previously reported association studies. The selected SNPs were screened in a study group of 835 participants from the Genome Data Base of Latvian Population and mainly consisted of patients with T2D and coronary heart disease. None of the individual polymorphisms were significantly associated with T2D status or BMI when analysed using logistic or linear regression and adjusted for gender, age and other significant covariates. Frequency of rs2241766 T allele homozygotes however was significantly increased in T2D patients compared to controls (uncorrected P = 0.007). When analysed with other traits, the rs182052 G allele was found to be less frequent in patients suffering from myocardial infarction (P = 0.02; OR = 0.76, CI95% [0.61-0.92]) compared to others. Haplotype analysis revealed significant association of one haplotype with atrial fibrillation (uncorrected P = 0.01). In summary, we conclude that SNPs in adiponectin gene are unlikely to represent the risk for T2D, but may be involved in pathogenesis of CHD in the Latvian population.

Polymorphisms in the gene coding for transcription factor 7 like 2 (TCF7L2) are recognized as the strongest common genetic risk factors for type 2 diabetes (T2D) across multiple ethnicities. This study was conducted to evaluate an association between TCF7L2 variants and diabetes susceptibility in the population of Latvia. We genotyped 4 single nucleotide polymorphisms (SNP) rs7901695, rs7903146, rs11196205 and rs12255372 in 1 093 controls and 1 043 diabetic subjects. Association with T2D was found for 3 SNPs rs7901695, rs7903146 and rs12255372 in the whole sample (under an additive genetic model, the adjusted odds ratios (OR) were 1.26, 95% CI [1.08-1.48], P=0.003; OR=1.32, 95% CI [1.12-1.55], P=0.001 and OR=1.35, 95% CI [1.15-1.60], P=0.0004 respectively). In addition observed effects on T2D susceptibility for analysed SNPs were higher among subjects with BMI under 30 kg/m². The impact of TCF7L2 variation on T2D risk in Latvian population is compatible with that demonstrated by a range of studies conducted in various ethnic groups.

Introduction Deep vein thrombosis (DVT) has a strong inherited predisposition that is partly explained by the strong genetic risk factors such as mutations in factor V, prothrombin, antithrombin III, protein C and S genes. Only recently the first GWAS have been performed on DVT resulting in discovery of novel genetic variants, however, the information on the common polymorphisms predisposing to the risk of DVT is still scarce. Materials and Methods Here we selected six SNPs (rs5361 in SELE, rs2066865 in FGG, rs2227589 in SERPINC1, rs1613662 in GP6, rs13146272 in CYP4V2, rs2289252 in F11) reported to be associated with venous thrombosis conditions and studied the association of these common variants in selected case (n = 177) and control (n = 235) groups from population of Latvia. Genotyping was performed using TaqMan hybridization probe SNP genotyping assay. Results Patients with DVT had a significantly higher frequency of F11 rs2289252 polymorphism (p = 0.001; OR [95%CI] = 1.61 [1.20-2.14]). When stratified by recurrence of DVT the tendency was observed that the same SNP had higher OR value in group of DVT patients with repeated episodes of DVT compared to patients with single DVT episode (p = 0.009; OR [95%CI] = 2.27[1.22-4.21] and p = 0.009; OR [95%CI] = 1.52[1.11-2.08] respectively), but due to limited group of cases this finding should be replicated. Conclusion We conclude that F11 gene variant rs2289252 contribute to inherited forms of DVT incidence and correlation of other analysed SNPs should be explored in populations with greater sample size and associated with various thrombosis related traits.

Background and objective: Additional loading doses and higher maintenance doses (MDs) have been used to overcome hyporesponsiveness of clopidogrel. We aimed to investigate whether genetic polymorphisms of two cytochromes (CYP2C19 and CYP2C9) and ABCB1 modify effect of such dose-adjustment strategy.Materials and methods: We enrolled 118 patients undergoing elective or acute percutaneous coronary intervention (PCI) with drug eluting stent (DES). Platelet reactivity index (PRI) was measured using the vasodilator-stimulated phosphoprotein (VASP) index and a cut-off value of ≥60% was defined as hyporesponsiveness. Polymorphism of two cytochromes (CYP2C19, CYP2C9) and gene ABCB1 were determined. In patients hyporesponsive to the initial LD the dose-adjustment was performed using up to 3 additional 600 mg LDs in order to achieve PRI &lt, 60%, and both 150 mg and 75 mg MD were tested at the follow-up.Results: Patients with at least one CYP2C19*2 allele had higher baseline PRI after the initial LD (78.2 ± 13.1 vs. 65.3 ± 19.5, P = 0.005). The PRI reduction with additional LD was significantly smaller in carriers of the CYP2C19*2 (25.2 ± 15.6 vs. 35.5 ± 16.8, P = 0.025) and similar trend was observed with subsequent additional LDs. Both MDs were less effective in presence of CYP2C19*2. Target PRI was, however, more frequently achieved with higher MD even in presence of CYP2C19*2 (in 70.6% vs. 23.5% of hyporesponders, P = 0.008). No such differences were observed for other polymorphisms.Conclusions: In patients hyporesponsive to a routine clopidogrel doses the potency of additional LD and higher MD of clopidogrel is compromised by presence of CYP2C19*2 allele. The dose-adjustment strategy is not affected by ABCB1 C3435T or CYP2C9 genotypes.

The glyoxalase system and its main enzyme, glyoxalase 1 (GLO1), protect cells from advanced glycation end products (AGEs), such as methylglyoxal (MG) and other reactive dicarbonyls, the formation of which is increased in diabetes patients as a result of excessive glycolysis. MG is partly responsible for harmful protein alterations in living cells, notably in neurons, leading to their dysfunction, and recent studies have shown a negative correlation between GLO1 expression and tissue damage. Neuronal dysfunction is a common diabetes complication due to elevated blood sugar levels, leading to high levels of AGEs. The aim of our study was to determine whether single nucleotide polymorphisms (SNPs) in the GLO1 gene influence activity of the enzyme. In total, 125 healthy controls, 101 type 1 diabetes, and 100 type 2 diabetes patients were genotyped for three common SNPs, rs2736654 (A111E), rs1130534 (G124G), and rs1049346 (5'-UTR), in GLO1. GLO1 activity was determined in whole blood lysates for all participants of the study. Our results showed a significant association between the minor alleles rs1130534 and rs1049346 and decreased enzyme activity (P=0.001 and P=2.61×10(-5), respectively). Increased allelic counts of the risk alleles were strongly associated with decreased GLO1 activity (standardised β=-0.24, P=2.15×10(-5)), indicating independent actions of these variants on GLO1 activity, as supported by the haplotype analysis. We showed for the first time an association between genetic variants with GLO1 enzyme activity in humans. SNPs in GLO1 can be used to predict enzyme activity and detoxifying capabilities, but further studies are needed to link these SNPs with common complications in diabetes.

ObjectiveThe aim of this study was to characterize the genetic variance of somatostatin receptor 5 (SSTR5) and investigate the possible correlation of such variants with acromegaly risk and different disease characteristics.Design and methodsThe SSTR5 gene coding region and 2000 bp upstream region was sequenced in 48 patients with acromegaly and 96 control subjects. Further, three single nucleotide polymorphisms (SNPs) were analyzed in the same group of acromegaly patients and in an additional group of 475 age- and sex-matched controls.ResultsIn total, 19 SNPs were identified in the SSTR5 gene locus by direct sequencing. Three SNPs (rs34037914, rs169068, and rs642249) were significantly associated with the presence of acromegaly using the initial controls. The allele frequencies were significantly (PP=0.00016 and OR=2.41; 95% CI, 1.41–4.13; P=0.0014 respectively). Haplotype reconstruction revealed two possible risk haplotypes determined by rs34037914 (633T) and rs642249 (1044A) alleles. Both haplotypes were found in significantly higher frequency in acromegaly patients compared with controls (Pβ=−10.4; P=0.002), increased body mass index (β=4.1; P=0.004), higher number of adenoma resection (PP=0.014).ConclusionsOur results demonstrate a previously undetected strong association of two SSTR5 SNPs with acromegaly. The data also suggest a possible involvement of SSTR5 variants in decreased suppression of GH production and increased tumor proliferation.

Adenosine plays an important part in the cardiac response to ischemia and reperfusion. The human adenosine receptor A3 (A3R), along with other adenosine receptors, is involved in mediation of those effects. The aim of the study was to ascertain whether the nonsynonymous single-nucleotide polymorphism (SNP) I248L (reference SNP ID: rs35511654) located in the A3R gene is associated with coronary heart disease (CHD). DNA samples from 683 individuals with CHD and from 826 control subjects selected from the Latvian Genome Database were successfully screened for rs35511654 using the TaqMan SNP Genotyping Assay. We observed a significantly decreased frequency of the rs35511654 C allele in a group of CHD patients compared with that in controls (p = 0.009). The association remained significant after adjustment for age, sex, and other nongenetic factors (p = 0.02). These results suggest that A allele of rs35511654 may predispose to CHD.

The adrenocorticotropic hormone (ACTH) receptor has highly specific membrane expression that is limited to adrenal cells; in other cell types the polypeptide fails to be transported to the cell surface. Unlike other evolutionarily related members of the melanocortin receptor family (MC1R-MC5R) that recognize different melanocortin peptides, ACTHR (MC2R) binds only ACTH. We used a mutagenesis approach involving systematic construction of chimeric ACTHR/MC4R receptors to identify the domains determining the selectivity of ACTHR membrane transport and ACTH binding. In total 15 chimeric receptors were created by replacement of selected domains of human ACTHR with the corresponding regions of human MC4R. We developed an analytical method to accurately quantify cell-membrane localization of recombinant receptors fused with enhanced green fluorescent protein by confocal fluorescence microscopy. The chimeric receptors were also tested for their ability to bind ACTH (1-24) and the melanocyte-stimulating hormone (MSH) analog, Nle4, DPhe7-alpha-MSH, and to induce a cAMP response. Our results indicate that substitution of the MC4R N-terminal segment with the homologous segment of ACTHR significantly decreased membrane transport. We also identified another signal localized in the third and fourth transmembrane regions as the main determinant of ACTHR intracellular retention. In addition, we found that the fourth and fifth transmembrane domains of the ACTHR are involved in ACTH binding selectivity. We discuss the mechanisms involved in bypassing these arrest signals via an interaction with melanocortin 2 receptor accessory protein (MRAP) and the possible mechanisms that determine the high ligand-binding specificity of ACTHR.

Objective(s) High variability in clinical response to metformin is often observed in type 2 diabetes (T2D) patients, and it highlights the need for identification of genetic components affecting the efficiency of metformin therapy. Aim of this observational study is to evaluate the role of tagSNPs (tagging single nucleotide polymorphisms) from genomic regions coding for six metformin transporter genes with respect to the short-term efficiency. Design 102 tagSNPs in 6 genes coding for metformin transporters were genotyped in the group of 102 T2D patients treated with metformin for 3 months. Methods Most significant hits were analyzed in the group of 131 T2D patients from Slovakia. Pharmacokinetic study in 25 healthy nondiabetic volunteers was conducted to investigate the effects of identified polymorphisms. Results In the discovery group of 102 patients, minor alleles of rs3119309, rs7757336 and rs2481030 were significantly nominally associated with metformin inefficiency (P = 1.9 × 10−6 to 8.1 × 10−6). Effects of rs2481030 and rs7757336 did not replicate in the group of 131 T2DM patients from Slovakia alone, whereas rs7757336 was significantly associated with a reduced metformin response in combined group. In pharmacokinetic study, group of individuals harboring risk alleles of rs7757336 and rs2481030 displayed significantly reduced AUC∞ of metformin in plasma. Conclusions For the first time, we have identified an association between the lack of metformin response and SNPs rs3119309 and rs7757336 located in the 5′ flanking region of the genes coding for Organic cation transporter 2 and rs2481030 located in the 5′ flanking region of Organic cation transporter 3 that was supported by the results of a pharmacokinetic study on 25 healthy volunteers.

Background Type 2 diabetes complications cause a serious emotional and economical burden to patients and healthcare systems globally. Management of both acute and chronic complications of diabetes, which dramatically impair the quality of patients' life, is still an unsolved issue in diabetes care, suggesting a need for early identification of individuals with high risk for developing diabetes complications. Methods We performed a genome-wide association study in 601 type 2 diabetes patients after stratifying them according to the presence or absence of four types of diabetes complications: diabetic neuropathy, diabetic nephropathy, macrovascular complications, and ophthalmic complications. Results The analysis revealed ten novel associations showing genome-wide significance, including rs1132787 (GYPA, OR = 2.71; 95% CI = 2.02–3.64) and diabetic neuropathy, rs2477088 (PDE4DIP, OR = 2.50; 95% CI = 1.87–3.34), rs4852954 (NAT8, OR = 2.27; 95% CI = 2.71–3.01), rs6032 (F5, OR = 2.12; 95% CI = 1.63–2.77), rs6935464 (RPS6KA2, OR = 2.25; 95% CI = 6.69–3.01) and macrovascular complications, rs3095447 (CCDC146, OR = 2.18; 95% CI = 1.66–2.87) and ophthalmic complications. By applying the targeted approach of previously reported susceptibility loci we managed to replicate three associations: MAPK14 (rs3761980, rs80028505) and diabetic neuropathy, APOL1 (rs136161) and diabetic nephropathy. Conclusions Together these results provide further evidence for the implication of genetic factors in the development of type 2 diabetes complications and highlight several potential key loci, able to modify the risk of developing these conditions. Moreover, the candidate variant approach proves a strong and consistent effect for multiple variants across different populations.

Background Metformin is a widely prescribed antihyperglycemic agent that has been also associated with multiple therapeutic effects in various diseases, including several types of malignancies. There is growing evidence regarding the contribution of the epigenetic mechanisms in reaching metformin’s therapeutic goals; however, the effect of metformin on human cells in vivo is not comprehensively studied. The aim of our study was to examine metformin-induced alterations of DNA methylation profiles in white blood cells of healthy volunteers, employing a longitudinal study design. Results Twelve healthy metformin-naïve individuals where enrolled in the study. Genome-wide DNA methylation pattern was estimated at baseline, 10 h and 7 days after the start of metformin administration. The whole-genome DNA methylation analysis in total revealed 125 differentially methylated CpGs, of which 11 CpGs and their associated genes with the most consistent changes in the DNA methylation profile were selected: POFUT2, CAMKK1, EML3, KIAA1614, UPF1, MUC4, LOC727982, SIX3, ADAM8, SNORD12B, VPS8, and several differentially methylated regions as novel potential epigenetic targets of metformin. The main functions of the majority of top-ranked differentially methylated loci and their representative cell signaling pathways were linked to the well-known metformin therapy targets: regulatory processes of energy homeostasis, inflammatory responses, tumorigenesis, and neurodegenerative diseases. Conclusions Here we demonstrate for the first time the immediate effect of short-term metformin administration at therapeutic doses on epigenetic regulation in human white blood cells. These findings suggest the DNA methylation process as one of the mechanisms involved in the action of metformin, thereby revealing novel targets and directions of the molecular mechanisms underlying the various beneficial effects of metformin. Trial registration EU Clinical Trials Register, 2016-001092-74. Registered 23 March 2017, https://www.clinicaltrialsregister.eu/ctr-search/trial/2016-001092-74/LV. Electronic supplementary material The online version of this article (10.1186/s13148-018-0593-x) contains supplementary material, which is available to authorized users.