Acapsular induces IL-8 production by BEAS-2B cells in a dose, time, and temperature dependent manner

<p><b>Copyright information:</b></p><p>Taken from "induces IL-8 secretion and CXCL1 expression by human bronchial epithelial cells"</p><p>http://respiratory-research.com/content/9/1/9</p><p>Respiratory Research 2008;9(1):9-9.</p><p>Published online 22 Jan 2008</p><p>PMCID:PMC2254606.</p><p></p> (A) BEAS-2B cells were unstimulated (NS, white box) or stimulated (black box) for 24 h with various MOI (100, 50, 20, 10, 1) of B3501 cultured at RT; LPS (1 μg/ml) was used as a positive stimulus. B) BEAS-2B cells were untstimulated (NS, white box) or stimulated for 24 h with various MOI (50, 20, 10, 5 and 1) of acapsular . C) BEAS-2B cells were stimulated with acapsular (MOI of 20) for 6 h and 24 h. (D) BEAS-2B cells were stimulated for 24 h with acapsular (MOI of 20) grown at 37°C or RT. The cell supernatants were collected and ELISA was used to measure IL-8 concentrations. All results are expressed as the mean ± S.D. of triplicate measurements and are representative of three independent experiments; *≤ 0.05, ** ≤ 0.01, *** ≤ 0.001 relative to NS

NHBE cells induce chemokine expression and are susceptible to damage following stimulation with

<p><b>Copyright information:</b></p><p>Taken from "induces IL-8 secretion and CXCL1 expression by human bronchial epithelial cells"</p><p>http://respiratory-research.com/content/9/1/9</p><p>Respiratory Research 2008;9(1):9-9.</p><p>Published online 22 Jan 2008</p><p>PMCID:PMC2254606.</p><p></p> (A) NHBE cells were left unstimulated (NS, white box) or stimulated with a MOI = 20 of capsular or acapsular cultured at RT or 37°C, as indicated. Twenty-four hours later, supernatants were collected and ELISA was used to measure IL-8 concentrations. Results are expressed as the mean ± S.D. of triplicate measurements and are representative of three independent experiments. (B) Supernatants were also assayed for LDH release using colorimetry. Data are expressed as a percentage of LDH release from unstimulated cells treated with a detergent lysis solution (triton) and represent the mean ± S.D. of triplicate measurements from two independent experiments. (C, D) Relative quantification of CXCL1 and CEBP/β expression was determined by real time PCR. Results are representative of the mean ± S.D. of one experiment performed in triplicate; δ ≤ 0.05 vs. NS, * ≤ 0.05, ** ≤ 0.01, *** ≤ 0.001

BEAS-2B cells induce chemokine gene expression in response to stimulation with acapsular

<p><b>Copyright information:</b></p><p>Taken from "induces IL-8 secretion and CXCL1 expression by human bronchial epithelial cells"</p><p>http://respiratory-research.com/content/9/1/9</p><p>Respiratory Research 2008;9(1):9-9.</p><p>Published online 22 Jan 2008</p><p>PMCID:PMC2254606.</p><p></p> (A) BEAS-2B cells were left unstimulated (NS) (left panel) or stimulated (right panel) with acapsular MOI = 20 cultured at 37°C. Twenty-four hours later, RNA was extracted and analyzed by microarray for expression of inflammatory cytokines and their receptors. A box indicates the location of endogenous control (housekeeping) genes; glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and beta-2 microglobulin (B2M) were used for normalization. (B) Summary of relative gene expression following stimulation with . Data shown is representative of three independent experiments. *Not determined due to undetectable IL-8 expression in untreated cells. (C) Expression of CCL2, CXCL1, and CEBP/β was analyzed by RT-PCR; β-actin was used as an endogenous control. (D) Relative quantification of CXCL1, CCL2, and CEBP/β expression in untreated (white columns) and stimulated (black columns) BEAS-2B cells was determined by real time PCR. Results are representative of three independent experiments

Table_1_SLC26A9 Gene Is Associated With Lung Function Response to Ivacaftor in Patients With Cystic Fibrosis.DOC

<p>Ivacaftor is a drug used to treat cystic fibrosis (CF) patients carrying specific gating CFTR mutations. Interpatient variability in the lung response has been shown to be partly explained by rs7512462 in the Solute Carrier Family 26 Member 9 (SLC26A9) gene. In an independent and larger cohort, we aimed to evaluate whether SLC26A9 variants contribute to the variability of the lung phenotype and if they influence the lung response to ivacaftor. We genotyped the French CF Gene Modifier Study cohort (n = 4,840) to investigate whether SLC26A9 variants were involved in the lung phenotype heterogeneity. Their influence in the response to ivacaftor was tested in the 30 treated patients who met the inclusion criteria: older than 6 years of age, percent-predicted forced expiratory volume measured in 1 s (FEV_1pp) in the 3 months before treatment initiation ranging between 40 and 90%. Response to treatment was determined by the change in FEV_1pp from baseline, averaged in 15–75 days, and the 1st-year post-treatment. We observed that SLC26A9 variants were not associated with lung function variability in untreated patients and that gain of lung function in patients treated with ivacaftor was similar to clinical trials. We confirmed that rs7512462 was associated with variability in ivacaftor-lung response, with a significant reduction in lung function improvement for patients with the C allele. Other SLC26A9 SNPs also contributed to the ivacaftor-response. Interindividual variability in lung response to ivacaftor is associated with SLC26A9 variants in French CF patients. Pharmacogenomics and personalized medicine will soon be part of CF patient care.</p

Table_2_SLC26A9 Gene Is Associated With Lung Function Response to Ivacaftor in Patients With Cystic Fibrosis.docx

<p>Ivacaftor is a drug used to treat cystic fibrosis (CF) patients carrying specific gating CFTR mutations. Interpatient variability in the lung response has been shown to be partly explained by rs7512462 in the Solute Carrier Family 26 Member 9 (SLC26A9) gene. In an independent and larger cohort, we aimed to evaluate whether SLC26A9 variants contribute to the variability of the lung phenotype and if they influence the lung response to ivacaftor. We genotyped the French CF Gene Modifier Study cohort (n = 4,840) to investigate whether SLC26A9 variants were involved in the lung phenotype heterogeneity. Their influence in the response to ivacaftor was tested in the 30 treated patients who met the inclusion criteria: older than 6 years of age, percent-predicted forced expiratory volume measured in 1 s (FEV_1pp) in the 3 months before treatment initiation ranging between 40 and 90%. Response to treatment was determined by the change in FEV_1pp from baseline, averaged in 15–75 days, and the 1st-year post-treatment. We observed that SLC26A9 variants were not associated with lung function variability in untreated patients and that gain of lung function in patients treated with ivacaftor was similar to clinical trials. We confirmed that rs7512462 was associated with variability in ivacaftor-lung response, with a significant reduction in lung function improvement for patients with the C allele. Other SLC26A9 SNPs also contributed to the ivacaftor-response. Interindividual variability in lung response to ivacaftor is associated with SLC26A9 variants in French CF patients. Pharmacogenomics and personalized medicine will soon be part of CF patient care.</p

Table_3_SLC26A9 Gene Is Associated With Lung Function Response to Ivacaftor in Patients With Cystic Fibrosis.doc

<p>Ivacaftor is a drug used to treat cystic fibrosis (CF) patients carrying specific gating CFTR mutations. Interpatient variability in the lung response has been shown to be partly explained by rs7512462 in the Solute Carrier Family 26 Member 9 (SLC26A9) gene. In an independent and larger cohort, we aimed to evaluate whether SLC26A9 variants contribute to the variability of the lung phenotype and if they influence the lung response to ivacaftor. We genotyped the French CF Gene Modifier Study cohort (n = 4,840) to investigate whether SLC26A9 variants were involved in the lung phenotype heterogeneity. Their influence in the response to ivacaftor was tested in the 30 treated patients who met the inclusion criteria: older than 6 years of age, percent-predicted forced expiratory volume measured in 1 s (FEV_1pp) in the 3 months before treatment initiation ranging between 40 and 90%. Response to treatment was determined by the change in FEV_1pp from baseline, averaged in 15–75 days, and the 1st-year post-treatment. We observed that SLC26A9 variants were not associated with lung function variability in untreated patients and that gain of lung function in patients treated with ivacaftor was similar to clinical trials. We confirmed that rs7512462 was associated with variability in ivacaftor-lung response, with a significant reduction in lung function improvement for patients with the C allele. Other SLC26A9 SNPs also contributed to the ivacaftor-response. Interindividual variability in lung response to ivacaftor is associated with SLC26A9 variants in French CF patients. Pharmacogenomics and personalized medicine will soon be part of CF patient care.</p

List of genes in the protein-binding gene-ontology category that were most upregulated (FC ≥ 3) in CTRL cells 6 hours after <i>P</i>. <i>aeruginosa</i> infection.

<p>Genes were classified according their molecular function, using the PANTHER Classification System (<a href="http://www.pantherdb.org/" target="_blank">http://www.pantherdb.org/</a>). 0 h, nonstimulated; 6 h, 6 hours postinfection; FC, fold change; adjpBH, adjusted <i>P</i> value to which the α cutoff was applied.</p><p>List of genes in the protein-binding gene-ontology category that were most upregulated (FC ≥ 3) in CTRL cells 6 hours after <i>P</i>. <i>aeruginosa</i> infection.</p

Gene-ontology molecular-function categories upregulated in infected cystic-fibrosis cells compared to CTRL cells.

<p>Gene-ontology molecular-function categories upregulated in infected cystic-fibrosis cells compared to CTRL cells.</p

Genes in the catalytic-activity (A) and in the protein-binding (B) gene-ontology category: five downregulated genes with the greatest difference in expression level between cystic-fibrosis and CTRL cells after infection.

<p>FC, fold change; h, hours; adjpBH, adjusted <i>P</i> value to which the α cutoff was applied.</p><p>Genes in the catalytic-activity (A) and in the protein-binding (B) gene-ontology category: five downregulated genes with the greatest difference in expression level between cystic-fibrosis and CTRL cells after infection.</p

Overview of gene expression profiles.

<p>A- The heat-map of the mean expression levels of all the genes in each condition revealed two distinct clusters that separated cystic-fibrosis (CF) cells from CTRL cells. B and C- Venn diagram of differentially expressed genes between cystic-fibrosis and CTRL cells upon <i>P</i>. <i>aeruginosa</i> infection. Circles: Differentially expressed genes upregulated (B) or downregulated (C) at a single postinfection time point or at two or three consecutive postinfection time points (thus, 0h was not considered). We selected the genes whose fold-change in expression level was ≥2 in the event of upregulation and ≤0.5 in the event of downregulation. Squares: Differentially expressed genes upregulated (B) or downregulated (C) at two, three, or four consecutive time points (thus, 0h was considered). We selected the genes for which the ratio of FC in infected cells over FC in noninfected cells was ≥1.5 in the event of upregulation or ≤0.6 in the event of downregulation.</p