Human melanoma cells differentially express RNASEL/RNase-L and miR-146a-5p under sex hormonal stimulation

Polymorphisms in the ribonuclease L (RNASEL) coding gene and hsa-miR-146a-5p (miR-146a) have been associated with melanoma in a sex-specific manner. We hypothesized that RNASEL and miR-146a expression could be influenced by sex hormones playing a role in the female advantages observed in melanoma incidence and survival. Thus, we explored the effects of testosterone and 17β-estradiol on RNASEL and miR-146a expression in LM-20 and A375 melanoma cell lines. Direct targeting of miR-146a to the 3' untranslated region (3'UTR) of RNASEL was examined using a luciferase reporter system. Our results indicate that RNASEL is a direct target of miR-146a in both melanoma cell lines. Trough qPCR and western blot analyses, we explored the effect of miR-146a mimic transfection in the presence of each hormone either on RNASEL mRNA level or on protein expression of RNase-L, the enzyme codified by RNASEL gene. In the presence of testosterone or 17β-estradiol, miR-146a overexpression did not influence RNASEL transcript level in LM-20 cell line, but it slightly induced RNASEL mRNA level in A375 cells. Remarkably, miR-146a overexpression was able to repress the protein level of RNase-L in both LM-20 and A375 cells in the presence of each hormone, as well as to elicit high expression levels of the activated form of the extracellular signal-regulated kinases (ERK)1/2, hence confirming the pro-tumorigenic role of miR-146a overexpression in melanoma. Thereafter, we assessed if the administration of each hormone could affect the endogenous expression of RNASEL and miR-146a genes in LM-20 and A375 cell lines. Testosterone exerted no significant effect on RNASEL gene expression in both cell lines, while 17β-estradiol enhanced RNASEL transcript level at least in LM-20 melanoma cells. Conversely, miR-146a transcript augmented only in the presence of testosterone in either melanoma cell line. Importantly, each hormone acted quite the opposite regarding the RNase-L protein expression, i.e., testosterone significantly decreased RNase-L expression, whereas 17β-estradiol increased it. Overall, the data show that, in melanoma cells treated with 17β-estradiol, RNase-L expression increased likely by transcriptional induction of its gene. Testosterone, instead, decreased RNase-L expression in melanoma cell lines with a post-transcriptional mechanism in which miR-146a could play a role. In conclusion, the pro-tumor activity of androgen hormone in melanoma cells could be exacerbated by both miR-146a increase and RNase-L downregulation. These events may contribute to the worse outcome in male melanoma patients

Haplotype block structure study of the CFTR gene. Most variants are associated with the M470 allele in several European populations

An average of about 1700 CFTR (cystic fibrosis transmembrane conductance regulator) alleles from normal individuals from different European populations were extensively screened for DNA sequence variation. A total of 80 variants were observed: 61 coding SNSs (results already published), 13 noncoding SNSs, three STRs, two short deletions, and one nucleotide insertion. Eight DNA variants were classified as non-CF causing due to their high frequency of occurrence. Through this survey the CFTR has become the most exhaustively studied gene for its coding sequence variability and, though to a lesser extent, for its noncoding sequence variability as well. Interestingly, most variation was associated with the M470 allele, while the V470 allele showed an 'extended haplotype homozygosity' (EHH). These findings make us suggest a role for selection acting either on the M470V itself or through an hitchhiking mechanism involving a second site. The possible ancient origin of the V allele in an 'out of Africa' time frame is discussed

Case-control association analysis of candidate genes in asthma, rhinitis and COPD: A preliminary report

This study aims to determine the genetic involvement in the susceptibility to asthma, rhinitis and COPD, by candidate gene association analysis, in a large and accurately de\ufb01ned series of Italian subjects, even considering exposure to some environmental contexts and life-styles

IFRD1 gene polymorphisms are associated with nasal polyposis in cystic fibrosis patients

Nasal polyposis (NP) is an inflammatory disease of the upper nasal airways frequently present in CF patients. Interferon-Related Developmental Regulator 1 (IFRD1) gene was reported as a possible modifier of CF lung disease severity. Three IFRD1 SNPs were analyzed to investigate a possible effect on the development of NP in CF patients. The DNA of 143 patients with CF (40 with and 103 without NP) was purified from peripheral blood samples. IFRD1 SNPs (rs7817, rs3807213, rs6968084) were genotyped by restriction enzyme analysis. The T allele of the common polymorphism rs7817 and the rs7817-rs3807213 haplotype were associated with NP(p=0.002 and 0.004 respectively). These results showed the association of the IFRD1-rs7817 polymorphism with NP in CF patients

The association among inflammation-related genes, fractional exhaled nitric oxide (FeNO), and symptom severity in adult asthma: a structural equation model (SEM).

Background. Airway inflammation is a key component of asthma that depends on the interplay of multiple genes. However, the association between health outcomes and genetic factors is usually evaluated by testing each candidate gene variant individually. Aim. To simultaneously assess the association of symptom severity with tag-SNPs located in inflammation-related genes and the FeNO level. Methods. In Verona (Italy), 327 asthmatics (aged 20-64) were identified from the general population in the GEIRD study (2008/2010). Individuals underwent a clinical interview, blood sampling for genotyping, and the measurement of FeNO (200 ml/s). SNPs in the CHI3L1, RAD50/IL4/IL5/IL13, IL33, and MS4A2 gene regions were genotyped by a custom GoldenGate Genotyping Assay. A symptom severity score (SSS) was computed by a multiple correspondence analysis on the basis of asthma-like symptoms, asthma attacks and hospitalization in the past year. The association of the SSS with SNPs and FeNO was assessed by a SEM. SNPs were summarised by a latent variable; atopy, gender, smoking habits and ICS use in the past year were considered as potential confounders. Results. The SEM is described in figure 1. Conclusions. This preliminary analysis suggests that inflammation-related genes could have a direct effect (not mediated by FeNO and atopy) on symptom severity in adult asthmatics

ASSOCIATION ANALYSIS OF CANDIDATE GENE POLYMORPHISMS WITH ASTHMA SEVERITY: RESULTS FROM THE GEIRD STUDY

Background: Many genes are associated with asthma severity1. However, studies on the association of genes with a measure of asthma severity enabling to catch the heterogeneity of the phenotypes are needed. Aims: To assess the single nucleotide polymorphisms (SNPs) associated with an overall severity score in adult subjects with active or non-active asthma. Methods: We evaluated 334 asthmatic subjects (aged 20-64) from the general population in Verona, who were identified in the clinical stage of an Italian multi-centre (multi)casecontrol study (GEIRD; 2008-2010)2. A panel of 384 Tag-SNPs (representative of 69 candidate genes) was genotyped by a custom GoldenGate Genotyping Assay. A symptom score [range: from 0 (non-active asthma and no respiratory symptoms in the past 12 months) to 8.5 (active asthma and maximum frequency of respiratory symptoms in the past 12 months)] was computed by a nonlinear principal component analysis (NLPC). The overall severity score was computed as a function of the symptom score, treatment (0=no treatment, 1=relievers only, 2=inhaled corticosteroids), and lung function (LF) [0=normal, 1=borderline, 2=airflow obstruction (AO) with FEV1 6580% predicted, 3=AO with FEV1 <80% predicted] by a second NLPC. For each SNP, the association with the overall severity score and the symptom score was tested by quasi-gamma models, whereas the association with treatment and LF was tested by ordered logistic models, adjusting for gender, BMI, and smoking habits. Simes multiple testing procedure was used for controlling the false discovery rate (FDR)3. Results: A SNP in the IL13 gene region was associated with the overall severity score (uncorrected p=0.0001; FDR-corrected p=0.036), whereas a second SNP in linkage disequilibrium (LD; r2=0.94) did not reach the statistical significance after the correction for multiple comparisons (uncorrected p=0.0006; FDR-corrected p=0.09). Both the SNPs were strongly associated with the symptom score (uncorrected p 640.00004; FDR-corrected p 640.006), but they were not associated with treatment and LF. A SNP in the SMAD3 gene region was associated with LF only (uncorrected p=0.00007; FDRcorrected p=0.021). Conclusions: Our results suggest that IL13 and SMAD3 gene regions (or genes in LD) play a role in asthma severity in adults