Association between genetic variant rs2267716 of CRHR2 gene with colorectal cancer

Colorectal cancer (CRC) is the third most common cancer and one of the main causes of death around the world. Multiple lines of evidence have suggested the role of the corticotropin-releasing hormone (CRH) family in CRC induction, including the low expression of corticotropin-releasing hormone receptor 2 (CRHR2), which is an angiogenesis inhibitor and inflammatory modulator. Previous research suggests that CRHR2 expression in colonic intestinal cells can regulate migration, proliferation and apoptosis through the modulation of several pathways. The aim of this study was to analyze the association of the rs10250835, rs2267716 and rs2267717 variants of CRHR2 gene with CRC in the Mexican population in order to consider its predictive value in CRC. This cross-sectional study included a group of 187 unrelated patients with sporadic CRC and a control group of 191 healthy blood donors. DNA extraction from peripheral blood was carried out using the Miller method. Identification of the rs10250835 variant was performed using PCR-restriction fragment length polymorphism (RFLP) and the rs2267716 and rs2267717 variants using TaqMan allelic discrimination assay. The minor allele homozygous CC of the rs2267716 variant of CRHR2 showed significant difference between CRC and control group (p=0.025), as well as the GCA haplotype (p=0.007), corresponding to the rs10250835, rs2267716 and rs2267717 variants, respectively. Our results suggest that the rs2267716 variant and GCA haplotype of CRHR2 represent a risk factor for CRC development in Mexican patients.

VDR Polymorphisms in Autoimmune Connective Tissue Diseases: Focus on Italian Population

Vitamin D is an important hormone involved in various physiologic processes, and its activity is linked to binding with vitamin D receptor (VDR). Genetic polymorphisms in the VDR gene could modulate the expression or function of the receptor and, consequently, alter the effects of vitamin D. Variants in VDR gene have been associated with susceptibility to many illnesses sensitive to vitamin D administration and to autoimmune disorders, but no data are available regarding autoimmune connective tissue diseases in Italian population. We analyzed three VDR polymorphisms in 695 Italian patients with autoimmune connective tissue diseases (308 with systemic lupus erythematosus (SLE), 195 with primary Sjogren’s syndrome (pSS), and 192 with rheumatoid arthritis (RA)) and in 246 healthy controls with the aim to evaluate a possible association of VDR SNPs with susceptibility to these diseases in the Italian population. Genotyping of rs2228570, rs7975232, and rs731236 in VDR gene was performed by an allelic discrimination assay. A case/control association study and a genotype/phenotype correlation analysis have been performed. We observed a higher risk to develop SLE for rs2228570 TT genotype ( P = 0.029 , OR = 1.79 ). No association was observed between susceptibility to pSS or RA and this SNP, although this variant is significantly less present in RA patients producing autoantibodies. For rs7975232 SNP, we observed a significant association of the variant homozygous genotype with SLE ( P = 0.009 , OR = 1.82 ), pSS ( P = 0.046 , OR = 1.66 ), and RA ( P = 0.028 , OR = 1.75 ) susceptibility. Moreover, we reported associations of this genotype with clinical phenotypes of SLE and pSS. Lastly, the GG genotype of rs731236 was associated with a lower RA susceptibility ( P = 0.045 , OR = 0.55 ). Our results show that the explored VDR polymorphisms are significantly associated with autoimmune connective tissue disorders and support the hypothesis that the genetic variability of VDR gene may be involved in susceptibility to these diseases in Italian population.

Beclometasone but not fluticasone modulates PDGFD expression in the H295R adrenal cell line.

BackgroundAdrenal suppression is a clinically concerning side effect of inhaled corticosteroid (ICS) treatment in patients with asthma. Increased susceptibility to ICS-induced adrenal suppression has previously been identified in those with the rs591118 polymorphism in Platelet Derived Growth Factor D (PDGFD). The mechanism underpinning this relationship is not known.MethodsH295R cells were genotyped for rs591118 using a validated Taqman PCR allelic discrimination assay. H295R cell viability was determined after treatment with beclometasone and fluticasone (range 0-330 μM). Cortisol was measured in cell culture medium using competitive enzyme immunoassay.ResultsPDGFD protein expression in H295R cells was confirmed using Western blotting. When ACTH and forskolin were added to H295R cells, a reduction in PDGFD expression was seen which was then restored by incubation with prochloraz, a known inhibitor of steroidogenesis.A dose-dependent, decrease in PDGFD expression was observed with beclometasone (over a 24 h incubation period) but not with beclometasone incubations beyond 24 hour nor with fluticasone (at 24 or 48 hours).ConclusionsH295R cells express PDGFD protein which can be modulated by incubation with steroidogenesis agonists and antagonists and additionally with exogenous beclometasone.

Experience of using the molecular genetic studies to assess the risk of suicide

In the recent decades, there has been widespread the opinion that genetic markers of the suicidal behavior (suicide, suicidal attempts, suicidal thoughts) can be used to predict the suicidal behavior.The purpose of the study was to determine the possibility of using the method of molecular genetic research to assess the risk of suicide in men of 18‒27 years.The study used the case-control method. The control group included 100 men of 18‒27 years who never had mental disorders. The suicide group included the persons who committed highly traumatic methods of self-harm and were motivated to commit suicide (30 persons). DNA isolation was performed using a NucleoSpin Blood kit (Macherey‒Nagel, Germany) according to the manufacturer’s protocol. Each DNA sample was analyzed for polymorphism by allelic discrimination using the real-time polymerase chain reaction (PCR).The frequencies of occurrence of genotypes and alleles of the following genes were analyzed: HTR1A, rs6295 (G/C); BDNF, rs6265 (G/A); COMT, rs4680 (G/A); SKA2, rs7208505 (C/T); SLC6A4 (5HTT), rs25531 (T/C); 5HTR2A, rs6313 (G/A); TPH2, rs4570625 (G/T); TPH1, rs1800532 (G/T).A statistically significant difference was found for the frequency of occurrence of genotypes and alleles of the rs25531 polymorphism of the SLC6A4 (5HTT) gene. The chance of being in the suicide group with a heterozygous genotype (T/C) carriage was 2.346 times higher.The significance of the rs25531 polymorphism of the SLC6A4 (5HTT) gene for the formation of the suicidal behavior was confirmed.

Involvement of lncRNA IL21-AS1 in interleukin-2 and T follicular regulatory cell activation in systemic lupus erythematosus

Background The single nucleotide polymorphism (SNP) rs62324212, located in IL21 antisense RNA 1 (IL21-AS1), has been identified as a genetic risk variant associated with systemic lupus erythematosus (SLE). We aimed to probe the characteristics of IL21-AS1 and explore its clinical relevance focusing on T helper subsets and disease activity in patients with SLE. Methods rs62324212 genotyping was determined using allelic discrimination by quantitative PCR. Gene expression in peripheral blood mononuclear cells and cell surface markers in CD4+ T cells were analyzed using PCR and flow cytometry. The association among IL21-AS1, CD4+ T cell subsets, and SLE disease activity was accessed. Results Ensembl Genome Browser analysis revealed that rs62324212 (C>A) was located in the predicting enhancer region of IL21-AS1. IL21-AS1 was expressed in the nucleus of CD4+ T and B cells, but its expression was decreased in patients with SLE. IL21-AS1 expression was positively correlated with mRNA levels of IL-2 but not IL-21, and it was associated with the proportion of activated T follicular regulatory (Tfr) cells. Furthermore, we observed a significant negative correlation between IL21-AS1 expression and disease activity in patients with SLE (n = 53, p < 0.05). Conclusion IL21-AS1 has an effect on disease activity through an involvement of IL-2-mediated activation of Tfr cells in SLE. Thus, targeting the IL21-AS1 may provide therapeutic approaches for SLE.

Decreased Risk of Ovarian Cancer Associated With rs9898876 Sex Hormone-binding Globulin Gene Variant

Background. Ovarian cancer (OC) is one of the most common gynecologic cancers,with significant morbidity and mortality. The risk of OCis influenced by hormone status, of which sex hormone-binding globulin (SHBG), whichinfluences the serum availability of steroid sex hormones, is implicated in the pathogenesis and evolution of OC. The aim of this study is to evaluate the involvement of common SHBG gene variants in OC susceptibility and evolution. Materials. A case control study including 71 OC patients and 74 cancer-free controls, who were genotyped for rs9898876, rs13894, rs1799941 and rs6257 SHBG SNP. Genotyping was done by the allelic discrimination method, using VIC- and FAM-labeled primers.Results. The minor allele frequencies of rs9898876, rs13894, rs1799941 and rs6257 SHBG SNP was comparable between OC cases and control women, implying no significant associations of the tested variants and overall OC risk. Taking homozygous wild-type genotype as reference (OR=1.00), heterozygous rs9898876 (G/T), and minor allele-carrying genotypes [G/T+T/T] were associated with reduced risk of OC. Whilers9898876 heterozygosity (G/T) was predictive of OC occurrence, no significant association of the remaining three tested SNPs was noted with altered risk of OC. Irrespective of FIGO staging, the four tested SHBG SNPs were not associated with the clinical progression of OC.Conclusion. In conclusion, SHBG rs9898876 is associated with a decreased risk of OC, and thus constitutes a potential diagnostic biomarker of OC.

Comparative Analysis of COL9A1 Genotyping in Oral Squamous Cell Carcinoma Diagnosis: A Pilot Study

The epidemiology of OSCC continues to increase despite the progress that has been made. More than ever, the diagnostic approach process needs to focus on genetic and epigenetic alterations. The aim of our study was to identify and correlate the presence of COL9A1 gene variants in two types of samples from OSCC patients. Methods: Our pilot study included 32 subjects diagnosed with OSCC. Fresh tumour tissue and peripheral blood samples were used in order to identify the genotypes of the COL9A1 gene. Variables, such as age, gender and tobacco and alcohol use, were also taken into consideration. The DNA analysis of the samples was based on a tagged SNP (rs550675) for the allelic discrimination. Results: The statistical significance and correlation of the COL9A1 genotypes within the two categories of samples was statistically significant (p < 0.001) for the C/T and T/T genotypes, providing an important perspective on the potential identification in blood samples of the gene mutation encountered in OSCC. Conclusions: This is the first study that focused on providing preliminary results using blood samples via the identification of COL9A1 gene variants in OSCC patients. The possibility of introducing a liquid biomarker by targeting this genetic variant is an appealing perspective for screening and diagnosis.

Carriage of mutations R462Q (rs 486907) and D541E (rs 627928) of the RNASEL gene and risk factors in patients with prostate cancer in Burkina Faso.

Background Prostate cancer (Pca) is a public health problem that affects men, usually of middle age or older. It is the second most common cancer diagnosed in men and the fifth leading cause of death. The RNASEL gene located in 1q25 and identified as a susceptibility gene to hereditary prostate cancer, has never been studied in relation to prostate cancer in Burkina Faso. The aim of this study was to analyze the carriage of RNASEL R462Q and D541E mutations and risks factors in patients with prostate cancer in the Burkina Faso. Methods This case-control study included of 38 histologically diagnosed prostate cancer cases and 53 controls (cases without prostate abnormalities). Real-time PCR genotyping of R462Q and D541E variants using the TaqMan® allelic discrimination technique was used. Correlations between different genotypes and combined genotypes were investigated. Results The R462Q variant was present in 5.3% of cases and 7.5% of controls. The D541E variant was present in 50.0% of cases and 35% of controls. There is no association between R462Q variants (OR= 0.60; 95%IC, 0.10 - 3.51; p= 0.686) and D541E variants (OR=2.46; 95%IC, 0.78 - 7.80; p= 0.121) and genotypes combined with prostate cancer. However, there is a statistically significant difference in the distribution of cases according to the PSA rate at diagnosis (p ˂ 0.001). For the Gleason score distribution, only 13.2% of cases have a Gleason score greater than 7. There is a statistically significant difference in the Gleason score distribution of cases (p ˂ 0.001) Conclusions These variants, considered in isolation or in combination, are not associated with the risk of prostate cancer.

Duplex RTqPCRs for detection and relative quantification of SARS-CoV-2 variants of concern (VOC)

During the evolution of the SARS-CoV-2 pandemic, new variants of the virus have emerged and spread worldwide. The increased transmissibility and proclivity of some of these variants to cause more serious disease threatens public health responses against the virus, and they are classified as variants of concern (Variants of Concern, VOCs). While Next-Generation-Sequencing (NGS) is the gold standard to identify VOC, it cannot always be rapidly implemented in some settings to provide information as an early warning tool. Duplex quantitative real time RTqPCR assays offer a sensitive and easy-to-use tool to detect, discriminate, and estimate relative proportions of SARS-CoV-2 variants containing VOC-specific signature mutations from variants lacking it, using allelic discrimination probes. We developed three multiplexed RTqPCR assays that can detect Beta (B.1.351), Gamma (P.1) and Delta (B.1.617.2) VOCs by targeting 22281_22289DelCTTTACTTG, 28262_28263InsAACA and 22029_22034DelAGTTCA deletions/insertions in their genomes, respectively. Beta and Delta deletions are mapped to the S gene (residues 241/243 and 157/158, respectively), while Gamma insertion is located between the end of ORF8 and the beginning of N gene. Our specific duplex RTqPCR assays have been adapted from a previously designed duplex RTqPCR assay used to estimate the relative proportion of genomes containing 21765-21770DelTACATG mutation affecting residues HV69/70, a signature mutation of Alpha VOC (Carcereny et al., 2021). All duplex RTqPCR assays targeting signature mutations of VOCs may be used as a complementary tool to NGS for rapid variant tracking and surveillance in wastewater-based epidemiology.

Evaluation of seven gene signature for predicting HCV recurrence post-liver transplantation

Background Orthotropic liver transplantation (OLT) offers a therapeutic choice for hepatocellular carcinoma (HCC) patients. The poor outcome of liver transplantation is HCV recurrence. Several genome-wide associated studies (GWAS) have reported many genetic variants to be associated with HCV recurrence. Seven gene polymorphisms formed a cirrhosis risk score (CRS) signature that could be used to distinguish chronic HCV patients at high risk from those at low risk for cirrhosis in non-transplant patients. This study aims to examine the association of CRS score and other clinical parameters with the probability for HCC emergence and/or the rate of HCV recurrence following liver transplantation. Results Seven gene polymorphisms, forming the CRS, were genotyped by real-time PCR using allelic discrimination protocol in 199 end-stage liver disease patients (79 child A, 43 child B, and 77child C), comprising 106 patients who encountered liver transplantation. Recipient CRS scores were correlated with HCV recurrence (HCV-Rec) at the end of the third year after OLT. Around 81% (39) recipients with low steatosis (LS; < 3.5%) donor percentage revealed no HCV recurrence (non-Rec) (p<0.001). CRS score could distinguish between child A, child B, and child C only at the low-risk group. Among the HCV Rec group 27% (8/30), 40% (12/30), and 33% (10/30) fell into the high, moderate, and low CRS risk groups, respectively. Stepwise logistic regression evinced two features more likely to be seen in HCV-Rec patients: abnormal ALT [OR, 1.1; 95% CI, 1.02–1.2] and donor steatosis >3.5% [OR, 46.07; 95% CI, 1.5–1407.8]. Conclusions Accordingly, the CRS score seems to be less useful to predict HCV recurrence after OLT. ALT and donor steatosis (exceed 3.5%) can significantly promote the HCV recurrence post-OLT. Moreover, the combination of MMF and CNI positively heightens HCV recurrence.