Selective anti-CXCR2 receptor blockade by AZD5069 inhibits CXCL8-mediated pro-tumorigenic activity in human thyroid cancer cells in vitro.

BACKGROUND: Thyroid cancer is the most common endocrine malignancy. Current therapies are successful, however some patients progress to therapeutically refractive disease. The immunotherapeutic potential of the CXCL8-chemokine/CXCR2-chemokine-receptor system is currently being explored in numerous human cancers. This study aimed to evaluate if the targeting of CXCR2 by its selective antagonist, AZD5069, could modulate CXCL8-mediated pro-tumorigenic effects in thyroid-cancer (TC) cells in vitro. METHODS: Normal human primary thyroid cells (NHT) and TC cell lines TPC-1 (RET/PTC), BCPAP, 8505C and 8305C (BRAFV600e) were treated with AZD5069 (100 pM-10 µM) over a time-course. Viability and proliferation were assessed by WST-1 and crystal violet assays. CXCL8 and CXCR2 mRNA were evaluated by RT-PCR. CXCL8-protein concentrations were measured in cell culture supernatants by ELISA. CXCR2 on cell surface was evaluated by flow-cytometry. Cell-migration was assessed by trans-well-migration chamber-system. RESULTS: AZD5069 exerted negligible effects on cell proliferation or viability. AZD5069 significantly reduced CXCR2, (but not CXCL8) mRNAs in all cell types. CXCR2 was reduced on the membrane of some TC cell lines. A significant reduction of the CXCL8 secretion was found in TPC-1 cells (basal-secretion) and NHT (TNFα-induced secretion). AZD5069 significantly reduced basal and CXCL8-induced migration in NHT and different TC cells. CONCLUSIONS: Our findings confirm the involvement of the CXCL8/CXCR2-axis in promoting pro-tumorigenic effects in TC cells, further demonstrating its immunotherapeutic significance in human cancer.

Alpha-1 deficiency in severe asthma patients.

Alpha-1 antitrypsin (AAT) deficiency, an autosomal co-dominant condition, decreases protein concentration and activity at both serum and tissue levels. Few studies investigated whether the type of SERPINA1 gene phenotype in patients with severe asthma can influence symptoms and disease control during follow-up.To assess whether the presence of a non-MM genotype of SERPINA1 in patients with severe asthma is associated with disease control, systemic and airway inflammation, lung function and comorbidities prevalence compared to severe asthma patients with a homozygous genotype (MM).Asthmatic patients belonging to Global Initiative for Asthma (GINA) step 5 were retrospectively analysed in an Italian reference asthma clinic. We collected clinical, biological and functional variables at baseline and for the three following years.Out of 73 patients enrolled, 14 (19.18%) were non-MM and 59 (80.8%) were MM. Asthmatics with non-MM genotype had lower serum AAT concentration (P = 0.004) and higher emphysema prevalence than the MM group (P = 0.003) at baseline. During follow up, only MM patients showed a significant improvement of both ACQ-6 score (P < 0.0001) and eosinophilic systemic inflammation (P < 0.0001).Our findings emphasise the importance of a screening for AAT deficiency in severe asthma, as alleles mutation may influence patient's follow-up..

Why lipid transfer protein allergy is not a pollen-food syndrome: novel data and literature review.

Summary: Background.Based on the cross-reactivity between pollen lipid transfer proteins (LTPs) and the peach LTP, Pru p 3, it has been suggested that the pollen might initiate the LTP sensitization process. Objective. To establish whether LTP allergy can be considered as a pollen-food syndrome. Methods. The literature was reviewed and new data of component-resolved diagnosis from Italy obtained by both ISAC immunoassay and ImmunoCAP on large populations of LTP hypersensitive patients were provided and analyzed. Results. Among Pru p 3 reactors, patients positive for Art v 3 and Pla a 3 largely exceeded those sensitized to the respective major pollen allergens, Art v 1 and Pla a 1/Pla a 2. Pru p 3 reactivity remained stable around 80-90% at all ages, whereas Art v 3 and Ole e 7 recognition was missing in younger patients. Pru p 3 IgE exceeded IgE specific for pollen LTP at all ages. Inhibition studies carried out on LTP reactors showed that commercial extracts of mugwort and plane pollen were unable to inhibit significantly Pru p 3 IgE reactivity. In follow-up studies, baseline Pru p 3 IgE levels exceeded Art v 3 IgE levels in 84% of those sensitized to both allergens, and all patients positive to only one LTP allergen at baseline were sensitized to Pru p 3. Further, most of the patients who did not show any LTP reactivity at baseline became exclusive Pru p 3 reactors. On ImmunoCAP singleplex Pru p 3 IgE levels exceeded Art v 3 IgE levels in 89% of cases (p less than 0.0001). Most literature data were in keeping with these new observations. Conclusions. The evidence for LTP syndrome being a pollen-food syndrome is presently very thin. Our data do not rule out the possible sensitization to the protein, via the airways or the skin.

Minimal clinically important difference of the 6-min walking test in patients with asthma.

BACKGROUND: The 6-min walking test (6MWT) is responsive to physiological changes and pulmonary rehabilitation (PR) in patients with asthma. The minimal clinically important difference (MCID) has not been established yet.OBJECTIVE: To determine the MCID of 6MWT in patients with asthma.METHODS: Using the perceived change in walking ability and the modified Medical Research Council (mMRC) score as anchors, receiver operating characteristic curves and quantile regression, we evaluated 6MWT before and after PR in these patients. The St George Respiratory Questionnaire (SGRQ), the COPD assessment test (CAT) and other outcome measures were also assessed.RESULTS: Of 142 patients with asthma, 37 were enrolled. After PR, 6MWT increased from 453.4 m ± 88.8 to 493.0 m ± 97.2 (P = 0.0001); other outcome measures also increased. There was a slight correlation between baseline 6MWT and SGRQ, CAT and mMRC. No significant correlations were found between post-PR changes in 6MWT and in other outcome measures. Comparing different methods of assessment, the MCID ranged from 26 m to 27 m.CONCLUSION: The most conservative estimate of the MCID of 6MWT after PR was 26 m in patients with asthma. This estimate may be useful in clinical interpretation of data, particularly in response to intervention studies.

The new generation PFAS C6O4 does not produce adverse effects on thyroid cells in vitro.

PURPOSE: Per- and poly-fluoroalkyl-substances (PFASs) are synthetic compounds that raised concern due to their potential adverse effects on human health. Long-chain PFAS were banned by government rules in many states, and thus, new emerging PFAS were recently introduced as substitutes. Among these, Perfluoro{acetic acid, 2-[(5-methoxy-1,3-dioxolan-4-yl)oxy]}, ammonium salt (C6O4) was recently introduced to produce a range of food contact articles and literature data about this compound are scanty. The aim of this study was to evaluate the in vitro effects of exposure to C6O4, compared with PFOA and PFOS on thyroid cells. METHODS: FRTL5 rat-thyroid cell lines and normal human thyroid cells (NHT) were incubated with increasing concentrations of C6O4 for 24, 48, 72, and 144 h to assess cell viability by WST-1. Cell viability was confirmed by AnnexinV/PI staining. Long-chain PFAS (PFOA and PFOS) were used at same concentrations as positive controls. The proliferation of cells exposed to C6O4, PFOA, and PFOS was measured by staining with crystal violet and evaluation of optical density after incubation with SDS. Changes in ROS production by FRTL5 and NHT after exposure to C6O4 at short (10, 20, and 30 min) and long-time points (24 h) were evaluated by cytofluorimetry. RESULTS: C6O4 exposure did not modify FRTL5 and NHT cell viability at any concentration and/or time points with no induction of necrosis/apoptosis. At difference, PFOS exposure reduced cell viability of FRTL5 while and NHT, while PFOA only in FRTL5. FRTL5 and NHT cell proliferation was reduced by incubation with by PFOA and PFOS, but not with C6O4. ROS production by NHT and FRTL5 cells was not modified after C6O4 exposure, at any time/concentration tested. CONCLUSIONS: The present in vitro study constitutes the first evaluation of the potential adverse effects of the new emerging PFAS C6O4 in cultured rat and human thyroid cells, suggesting its safety for thyroid cells in vitro.

Relationship between diabetes and respiratory diseases-Clinical and therapeutic aspects.

Diabetes is a common metabolic disorder affecting the entire body with high morbidity and mortality worldwide. The major complications related to diabetes are mostly due to the macrovascular and microvascular bed impairment due to metabolic, hemodynamic and inflammatory factors. However, studies over the past decades have added also the lung as a target organ in both type 1 and type 2 diabetes. Diabetes has always been addressed as a major comorbidity conditioning the disease behaviour and the natural history of several respiratory diseases. Increased interest has recently focused on the pathophysiology of the metabolic glycaemic disorder and the respiratory diseases suggesting a similar background shared by the two conditions. The true relationship between pulmonary diseases and diabetes mellitus has not been clarified, this review aims to summarize the link between diabetes and coexisting respiratory diseases such as asthma, chronic obstructive pulmonary disease, respiratory infections, cystic fibrosis, lung cancer and obstructive sleep apnea from a pathogenetic and therapeutic point of view.

An atlas of IgE sensitization patterns in different Italian areas. A multicenter, cross-sectional study.

Summary: Background. The development of recombinant technology supported the allergy diagnostic work-up in the daily clinical practice, representing a useful tool for epidemiological studies. Methods. An atlas of the IgE sensitization profiles found throughout Italy was prepared from a nationwide, multicenter, cross-sectional study. Results. 6052 unselected consecutive individuals, belonging to North-West [NW], North-East [NE], Centre [C], South [S], and Islands subset [Is] were evaluated by means of the ImmunoCAP ISAC test. The top-ranked sensitizations found were Cup a 1 in [C] (58.1%) and [S] (53.6%), Phl p 1 in the North (from 46.1% to 49%), and Cyn d 1 in [Is] (44.2%). High frequency of house dust mite group 2 molecules sensitization was found in [C] (36.9%) and [S] Italy (40.8%), whilst low level of reactivity was recorded in [NW] (20%). Pellitory hypersensitivity was mainly found in [C], [S], and [Is], whilst ragweed Amb a 1 sensitivity was particularly found in [NW] Italy. IgE recognition of PR-10, Profilin, and nsLTP was mutually exclusive in 69.1% of cases, PR-10 reactivity mostly occurring in [NE], Profilin in [NW], and nsLTP molecules recognition mainly recorded in [C] and [S]. Conclusions. Divergent IgE sensitization patterns were found along Italy, possibly linked to the distinct geographical locations, indicating multiplex system IgE analysis as a reliable approach for epidemiological evaluation even in small geographical areas.

The AMPK-activator AICAR in thyroid cancer: effects on CXCL8 secretion and on CXCL8-induced neoplastic cell migration.

PURPOSE: The AMPK-activator AICAR recently raised great interest for its anti-cancer properties. With specific regard to thyroid cancer, AICAR reduces cancer cell growth, invasion and metastasis. CXCL8, a chemokine with several recognized tumorigenic effects, is abundantly secreted in thyroid cancer microenvironment. The aim of this study was to investigate if AICAR could inhibit the basal and the TNFα-induced CXCL8 secretion in normal human thyroid cells (NHT) and in thyroid cancer cell lines TPC-1 and BCPAP (RET/PTC and BRAFV600e mutated, respectively). METHODS: The effect of AICAR on basal and CXCL8-induced cell migration was assessed. Cells were incubated with AICAR (0.05, 0.5, 1, 2 mM) alone or in combination with TNF-α (10 ng/ml) for 24 h. CXCL8 concentrations were measured in cell supernatants. Transwell migration assays were performed in NHT, TPC-1 and BCPAP, basally and after treatment with AICAR (2 mM) and rh-CXCL8 (50 ng/ml) alone or in combination. RESULTS: AICAR dose dependently inhibited the basal secretion of CXCL8 in TPC-1 (F = 4.26; p < 0.007) and BCPAP (F = 6.75; p < 0.0001) but not in NHT. TNFα-induced CXCL8 secretion was dose dependently reduced by AICAR in NHT (F = 9.99; p < 0.0001), TPC-1 (F = 9.25; p < 0.0001) and BCPAP (F = 6.82; p < 0.0001). AICAR significantly reduced the basal migration of TPC-1 and BCPAP but not of NHT. CONCLUSIONS: CXCL8-induced cell migration was inhibited in NHT, TPC-1 and BCPAP. This is the first demonstration of the inhibition of CXCL8 secretion exerted by AICAR in TPC-1 and BCPAP indicating that the anti-cancer properties of AICAR are, at least in part, mediated by its ability to reduce the pro-tumorigenic effects of CXCL8.

Can pet keeping be considered the only criterion of exposure to cat/dog allergens in the first year of life?

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

BACKGROUND: 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. METHODS AND 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. RESULTS: The T allele of the common polymorphisms rs7817 and the rs7817-rs3807213 haplotype were associated with NP (p = 0.002 and 0.004, respectively). CONCLUSIONS: These results showed the association of the IFRD1-rs7817 polymorphism with NP in CF patients.

EAACI Position Paper on assessment of cough in the workplace.

Cough is a nonspecific and relatively common symptom that can present difficulties in diagnosis and management, particularly when it is reported to be associated with the workplace. The present consensus document, prepared by a taskforce of the Interest Group on Occupational Allergy of the European Academy of Allergy and Clinical Immunology by means of a nonsystematic review of the current literature, is intended to provide a definition and classification of work-related chronic cough (WRCC) to assist the daily practice of physicians facing with this symptom. The review demonstrates that several upper and lower airway work-related diseases may present with chronic cough; hence, the possible link with the workplace should always be considered. Due to the broad spectrum of underlying diseases, a multidisciplinary approach is necessary to achieve a definite diagnosis. Nevertheless, more epidemiological studies are necessary to estimate the real prevalence and risk factors for WRCC, the role of exposure to environmental and occupational sensitizers and irritants in its pathogenesis and the interaction with both upper and lower airways. Finally, the best management option should be evaluated in order to achieve the best outcome without adverse social and financial consequences for the worker.

PPARG2 Pro12Ala and ADAMTS9 rs4607103 as "insulin resistance loci" and "insulin secretion loci" in Italian individuals. The GENFIEV study and the Verona Newly Diagnosed Type 2 Diabetes Study (VNDS) 4.

We investigated cross-sectionally whether the type 2 diabetes (T2DM) risk alleles of rs1801282 (PPARG2) and rs4607103 (ADAMTS9) were associated with T2DM and/or insulin sensitivity (IS) and beta cell function (ßF) in Italians without and with newly diagnosed T2DM. In 676 nondiabetic subjects (336 NGR and 340 IGR) from the GENFIEV study and in 597 patients from the Verona Newly Diagnosed Type 2 Diabetes Study (VNDS), we (1) genotyped rs1801282 and rs4607103, (2) assessed ßF by C-peptide/glucose modeling after OGTT, and (3) assessed IS by HOMA-IR in both studies and by euglycemic insulin clamp in VNDS only. Logistic, linear, and two-stage least squares regression analyses were used to test (a) genetic associations with T2DM and with pathophysiological phenotypes, (b) causal relationships of the latter ones with T2DM by a Mendelian randomization design. Both SNPs were associated with T2DM. The rs4607103 risk allele was associated to impaired ßF (p < 0.01) in the GENFIEV study and in both cohorts combined. The rs1801282 genotype was associated with IS both in the GENFIEV study (p < 0.03) and in the VNDS (p < 0.03), whereas rs4607103 did so in the VNDS only (p = 0.01). In a Mendelian randomization design, both HOMA-IR (instrumental variables: rs1801282, rs4607103) and ßF (instrumental variable: rs4607103) were related to T2DM (p < 0.03-0.01 and p < 0.03, respectively). PPARG2 and ADAMTS9 variants are both associated with T2DM and with insulin resistance, whereas only ADAMTS9 may be related to ßF. Thus, at least in Italians, they may be considered bona fide "insulin resistance genes".

Recommendations for the classification of diseases as CFTR-related disorders.

Several diseases have been clinically or genetically related to cystic fibrosis (CF), but a consensus definition is lacking. Here, we present a proposal for consensus guidelines on cystic fibrosis transmembrane conductance regulator (CFTR)-related disorders (CFTR-RDs), reached after expert discussion and two dedicated workshops. A CFTR-RD may be defined as "a clinical entity associated with CFTR dysfunction that does not fulfil diagnostic criteria for CF". The utility of sweat testing, mutation analysis, nasal potential difference, and/or intestinal current measurement for the differential diagnosis of CF and CFTR-RD is discussed. Algorithms which use genetic and functional diagnostic tests to distinguish CF and CFTR-RDs are presented. According to present knowledge, congenital bilateral absence of vas deferens (CBAVD), acute recurrent or chronic pancreatitis and disseminated bronchiectasis, all with CFTR dysfunction, are CFTR-RDs.

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.

Variants and haplotypes of TCF7L2 are associated with ß-cell function in patients with newly diagnosed type 2 diabetes: the Verona Newly Diagnosed Type 2 Diabetes Study (VNDS) 1.

CONTEXT: Intronic variants of TCF7L2 are confirmed genetic risk factors for type 2 diabetes and are associated to alterations in beta cell function in nondiabetic individuals. OBJECTIVE: The objective of the study was to test whether TCF7L2 variability may affect ß-cell function also in patients with type 2 diabetes. DESIGN: This was a cross-sectional association study. SETTING: The study was conducted at a university hospital referral center for diabetes. PATIENTS: Patients included 464 (315 males and 149 females) glutamic acid decarboxylase-negative patients [age: median 59 yr (interquartile range: 52-65); body mass index: 29.3 kg/m(2) (26.5-32.9); fasting plasma glucose: 7.0 mmol/liter (6.1-8.0)] with newly diagnosed type 2 diabetes. INTERVENTION(S): Interventions included frequently sampled oral glucose tolerance test and euglycemic insulin clamp. MAIN OUTCOME MEASURE(S): ß-Cell function (derivative control and proportional control); insulin sensitivity; genotypes of the following TCF7L2 single-nucleotide polymorphisms: rs7901695, rs7903146, rs11196205, and rs12255372. RESULTS: Both rs7901695 and rs7903146 diabetes risk alleles were associated with reduced proportional control of ß-cell function (P = 0.019 and P = 0.022, respectively). Two low-frequency haplotypes were associated with extreme (best and worst) phenotypes of ß-cell function (P < 0.01). No associations between TCF7L2 genotypes and insulin sensitivity were detected. CONCLUSIONS: TCF7L2 diabetes risk variants, either as single-nucleotide polymorphisms or as haplotypes, detrimentally influence ß-cell function and might play a role in determining the metabolic phenotype of patients with newly diagnosed type 2 diabetes.

[Professional eosinophilic bronchitis: considerations and new diagnostic methods in a clinical case]. / La bronchite eosinofila professionale: considerazioni e nuovi strumenti diagnostici a margine di un caso clinico.

Nonasthmatic eosinophilic bronchitis (NAEB) is a condition characterized by corticosteroid-responsive chronic cough, sputum eosinophilia and absence of symptoms or objective evidence of variable airflow obstruction and airway hyper-responsiveness. Like asthma, NAEB can be associated with exposure to occupational sensitizers and can be considered as being a variant of occupational asthma when it develops as a consequence of work exposure. Few case reports of NAEB caused by workplace exposure have been reported. Bakers are at high risk of developing occupational respiratory disorders and three cases of occupational NAEB have been described. We describe the first case of occupational NAEB due to storage mites in a baker in which the offending agent was identified by means of the basophil activation test (BAT), a new tool which has never been proposed in diagnostic procedures of occupational respiratory allergy. BAT's results allowed the recognition of the offending agent, that is mandatory for diagnosis.