A Preliminary microRNA Analysis of Non Syndromic Thoracic Aortic Aneurysms.

The development of thoracic aortic aneurysms (TAAs) involves a multifactorial process resulting in alterations of the structure and composition of the extracellular matrix (ECM). Recently, modifications in microRNA (miRNA) expression were implicated in the pathogenesis of TAA. This study presents a preliminary miRNA microarray analysis conducted on pooled ascending aorta RNAs obtained from non familial non syndromic TAA patients (five males and five females) compared to matched control pools. Ninety-nine differentially expressed miRNAs with >1.5-fold-up- or down-regulation in TAAs compared to controls were identified, 16.0% of which were similarly regulated in the two sexes. Genes putatively targeted by differentially expressed miRNAs belonged preferentially to focal adhesion and adherens junction pathways. The results indicate an altered regulation of miRNA-mediated gene expression in the cellular interactions of aneurysmal aortic wall.

Upregulated expression of Toll-like receptor 4 in peripheral blood of ischaemic stroke patients correlates with cyclooxygenase 2 expression.

OBJECTIVES: An inflammatory process following stroke in human brains and systemic inflammatory responses after stroke in humans have been reported by numerous investigators. The aim of the study was to investigate if genes involved in the cyclooxygenase 2 (COX-2) pathway are upregulated at peripheral level in patients after transient ischaemic attack (TIA) and stroke. DESIGN OF STUDY: Blood samples were obtained from two groups of patients undergoing carotid endarterectomy. The first group included 25 patients who presented TIA or ischaemic stroke. The second group included 35 patients who had an asymptomatic internal carotid artery stenosis. Total RNA was isolated and the expression of Toll-like Receptor 4 (TLR4), COX-2, membrane-associated Prostaglandin E synthase (mPGES-1), Prostaglandin E2 receptors (EP3 and EP4) was analysed by real time RT-PCR. RESULTS: Expression of COX-2 and TLR4 were significantly increased in symptomatic patients (p < 0.001). Correlation analysis showed that TLR4 expression significantly correlated with COX-2 expression (R = 0.65; p < 0.01) in ischaemic stroke patients. This correlation was not observed in TIA and asymptomatic patients. CONCLUSIONS: Our results suggest that the peripheral mechanism of inflammatory injury after stroke may be mediated by TLR4 through a COX-2-dependent pathway.

No linkage or association of five polymorphisms in the interleukin-4 receptor alpha gene with atopic asthma in Italian families.

The literature contains conflicting reports on the association of common variants of the interleukin (IL)-4 receptor alpha (IL4RA) gene with atopic asthma. The purpose of the present study was to investigate the linkage and association of several gene polymorphisms with atopic asthma in a large series of well-characterized individuals. Analysis of five polymorphisms (I50V, E375A, C406R, S478P and Q551R) of the IL4RA gene was performed in 823 individuals from 182 families with atopic asthmatic children from north-east Italy. The subjects were tested for clinical asthma, total serum IgE level, skin prick test positivity to common aeroallergens, and bronchial hyperresponsiveness to methacholine. The frequency of the polymorphisms was similar to that reported for other populations. The 375, 406, 478 and 551 polymorphisms were in linkage disequilibrium, as previously reported. No linkage or transmission disequilibrium was observed in the families between any mutation and any of the phenotypes investigated. No multipoint haplotype was associated with any phenotype. In conclusion, the IL4RA gene does not seem to play an important role in genetic predisposition to atopic asthma in the population tested.

Chromosome 14 linkage analysis and mutation study of 2 serpin genes in allergic asthmatic families.

BACKGROUND: Genome and chromosome screens reported DNA markers on chromosome 14 linked to allergic asthma or intermediate phenotypes in several populations. OBJECTIVE: We sought to perform a linkage study on chromosome 14 and a further association study on candidate genes mapped in the region found to be linked to allergic asthma or intermediate phenotypes. METHODS: The study consisted of a sample of 189 families (847 genotyped individuals) from a restricted geographic area in northeastern Italy. The subjects were characterized for the following phenotypes: allergic asthma, total serum IgE levels, skin prick test responses, and bronchial hyperresponsiveness (BHR) to methacholine. Genotyping was done with 14 DNA markers and 4 polymorphisms in the genes encoding alpha(1)-anti-trypsin and alpha(1)-antichymotrypsin (ACT). RESULTS: Multipoint analysis indicated a potential linkage of BHR with marker D14S617 (nonparametric linkage z score = 2.32, P =.01). Transmission disequilibrium of Thr -15Ala in the gene encoding ACT was observed with all the phenotypes investigated: allergic asthma, BHR, total IgE levels, or skin prick test responses (P =.041,.02,.0053, or.026, respectively). CONCLUSION: Chromosome 14 screening and transmission disequilibrium testing on the gene encoding ACT suggest that it or a closely located gene may be involved in susceptibility to allergic asthma in the Italian population.

CFTR and cationic trypsinogen mutations in idiopathic pancreatitis and neonatal hypertrypsinemia.

BACKGROUND/AIMS: The CFTR gene has been shown to be involved in sporadic idiopathic pancreatitis (IP) and neonatal hypertrypsinemia with normal sweat chloride test (NHNST). The cationic trypsinogen gene (Try4) is responsible for hereditary pancreatitis. The aim of the present study was to find a correlation between mutations in the two genes and the two phenotypes. METHODS: Analysis of some known gene mutations and complete gene screening by denaturing gradient gel electrophoresis and DNA sequencing were undertaken. Thirty-two sporadic IP patients were investigated for the CFTR study, while 13 sporadic IP patients plus 4 hereditary pancreatitis families (24 tested individuals) were examined for the Try4 study. Fifty neonates with NHNST were investigated for the study of both genes. RESULTS: CFTR mutations were more frequently observed in sporadic IP cases with a common cystic fibrosis mutation or borderline sweat chloride than in cases with a negative sweat test. Try4 mutations were found in 1 out of the 13 sporadic IP cases tested. CONCLUSIONS: The CFTR gene may be involved in IP and NHNST, while the Try4 gene may be involved in IP, but not in NHNST, in this limited series of observations.

Tumor necrosis factor gene complex in COPD and disseminated bronchiectasis.

BACKGROUND: Tumor necrosis factor (TNF) is a potent proinflammatory cytokine with increased levels in the sputum of COPD subjects. Two biallelic TNF gene complex polymorphisms have been described: LtalphaNcoI, in the first intron of the lymphotoxin alpha (previously referred to as TNF-beta) gene, and TNF-308, in the promoter region of the TNF-alpha gene. Higher levels of TNF production are associated with allele 1 of LtalphaNcoI (LtalphaNcoI*1) and with allele 2 of TNF-308 (TNF-308*2). STUDY OBJECTIVES: To study the frequencies of the two TNF gene complex polymorphisms in patients with COPD and bronchiectasis. DESIGN: Association study. SUBJECTS AND METHODS: We studied the frequencies of these polymorphisms in 66 subjects with COPD and in 23 subjects with disseminated bronchiectasis and compared them to the frequencies in 98 healthy control subjects and 45 subjects with nonobstructive pulmonary disease. Genomic DNA samples were extracted, and TNF-alpha and LtalphaNcoI polymorphisms were detected after polymerase chain reaction by restriction digestion. RESULTS: We found the following frequencies: the TNF-308*2 allele was detected in 11% of COPD individuals, 15% of bronchiectasis patients, 10% of healthy control subjects, and 18% of subjects with nonobstructive pulmonary disease. The LtalphaNcoI*1 allele was detected in 28% of COPD individuals, 30% of bronchiectasis patients, 29% of healthy control subjects, and 29% of subjects with nonobstructive pulmonary disease. We found evidence of linkage disequilibrium between the two loci (Delta = 0.068). CONCLUSIONS: We conclude that the TNF gene complex, at least in Caucasoid individuals and for the considered polymorphisms, does not seem to play a major role as genetic risk factor in COPD and bronchiectasis.

Association of a lymphotoxin alpha gene polymorphism and atopy in Italian families.

Tumour necrosis factor (TNF) is a proinflammatory cytokine that increases human airway tissue responsiveness and is considered a candidate gene for asthma. Two common polymorphisms (LTalphaNcoI and TNFalpha-308) in the TNF gene complex were studied in 600 subjects from 131 Italian families with atopic asthmatic children. Skin prick test (SPT), total IgE levels, atopy (defined as increased IgE levels or SPT positivity or both), bronchial hyperresponsiveness, and clinical asthma were investigated. The observed distribution of the identical by descent alleles at the LTalphaNcoI locus was different from expected for SPT and atopy (p=0.015). The LTalphaNcoI genotype distribution for increased IgE levels was different between males and females (p=0.0011), and an association of the 2.2 genotype with increased IgE levels was observed in females (p=0.0032). The results indicate that the LTalpha gene, or a closely linked locus, is associated with atopy, and suggest a sex difference in the effect of the gene.

Candidate genes and a genome-wide search in Italian families with atopic asthmatic children.

To identify genetic factors for susceptibility to atopy and asthma in childhood, 1,083 subjects were identified, mainly from the Veneto region and Bolzano province in North-east Italy, of whom 817 were from 172 families with at least two affected people, 189 were sporadic cases, and 77 unrelated controls. All the subjects were characterized for clinical asthma (asthma), total serum IgE (IgE), skin prick test (SPT) reactivity to common aeroallergens and bronchial hyperresponsiveness (BHR) to methacoline test. Atopy was defined as SPT positivity and/or increased IgE levels. Several candidate genes were investigated, and genome-wide linkage analysis was been initiated. The high affinity IgE receptor beta chain (FcepsilonRIbeta) locus showed significant allele sharing in affected sib-pairs for BHR and for SPT positivity. Lymphotoxin alpha (Ltalpha) gene Ncol mutation showed a suggestive linkage with atopy, and the LTalphaNcol 2/2 genotype was found to be associated with increased total IgE levels in all females. No evidence for linkage or association of any phenotype to the tumour necrosis factor alpha (TNFalpha) - 308 mutation or to the interleukin 4 receptor alpha (IL-4Ralpha) Q576R mutation was found. BHR, asthma and increased IgE were found to be linked to X and Y long arm pseudoautosomal region (PAR2) markers. Initial data were also collected from linkage analysis with chromosome 12, 14, and 19, DNA markers. Non-parametric multipoint analysis provides preliminary evidence for linkage of asthma with D12S390, of atopy with D19S601, and of BHR with D14S617. These results suggest that several genetic factors contribute to different allergic asthma phenotypes in the population investigated.

Preferential release of high amounts of interleukin-8 by myeloid blasts showing monocytic differentiation.

BACKGROUND: In myeloid blasts, the expression and release of the multifunctional chemokine IL-8 could be expected to be differentiation-associated. METHODS: We investigated the profile of interleukin-8 (IL-8) expression and release by leukemic cells obtained at diagnosis from 42 untreated adult patients with acute myeloid leukemia of various FAB subtypes (2 M0, 7 M1, 6 M2, 6 M3, 10 M4 and 11 M5). IL-8 transcripts were evaluated by Northern blot and densitometric analysis. IL-8 release by myeloid blasts was evaluated by a specific ELISA either in sera at diagnosis or in supernatants (SN) obtained from cultured leukemic cells. RESULTS: In basal conditions, Northern blot analysis revealed detectable IL-8 transcripts in 15/29 cases, eleven of which were classified as M4-M5 and 4 as FAB M0-M3. Densitometric analysis of IL-8 transcript bands showed higher expression in M4-M5 than in M0-M3 cases (mean values +/- SD: 16.5 +/- 21 and 0.77 +/- 1.36 densitometric units, respectively; p = 0.012). Higher IL-8 serum levels were observed in leukemic patients as opposed to normal controls (mean values +/- SD: 0.53 +/- 0.75 vs 0.003 +/- 0.014 ng/mL, respectively; p = 0.006). Furthermore, a trend (though not of statistical significance) towards higher IL-8 serum values was observed in M4-M5 as opposed to M0-M3 subtypes. After 24 hours of culture, the majority of myeloid blasts (95%) spontaneously released detectable amounts of IL-8 into SN. However, M4-M5 released substantially higher amounts of IL-8 than M0-M3 blasts (mean +/- SD: 68 +/- 46 and 8.5 +/- 12 ng/mL, respectively; p < 0.001). This difference between M0-M3 and M4-M5 blasts was already observed after 6 hours of culture and increased over 72 hours. CONCLUSIONS: Our findings confirm and further support the preferential release of high levels of IL-8 by myeloid blasts showing monocytic differentiation.