Soluble ST2 suppresses the effect of interleukin-33 on lung type 2 innate lymphoid cells

Type 2 innate lymphoid cells (ILC2) in lungs produce interleukin (IL)-5 and IL-13 in response to IL-33 and may contribute to the development of allergic diseases such as asthma. However, little is known about negative regulators and effective inhibitors controlling ILC2 function. Here, we show that soluble ST2, a member of the IL-1 receptor family, suppresses the effect of IL-33 on lung ILC2 in vitro. Stimulation with IL-33 to naïve ILC2 induced morphological change and promoted cell proliferation. In addition, IL-33 upregulated expression of cell surface molecules including IL-33 receptor and induced production of IL-5 and IL-13, but not IL-4. Pretreatment with soluble ST2 suppressed IL-33-mediated responses of ILC2. The results suggest that soluble ST2 acts as a decoy receptor for IL-33 and protects ILC2 from IL-33 stimulation.     

TFF3-peptide increases transepithelial resistance in epithelial cells by modulating claudin-1 and -2 expression

TFF3 plays an important role in the protection and repair of the gastrointestinal mucosa. The molecular mechanisms of TFF function, however, are still largely unknown. Increasing evidence indicates that apart from stabilizing mucosal mucins TFF3 induces cellular signals that modulate cell–cell junctions of epithelia. In transfected HT29/B6 and MDCK cells stably expressing FLAG-tagged human TFF3 we have recently shown that TFF3 down-regulates E-cadherin, impairs the function of adherens junctions and thus facilitates cell migration in wounded epithelial cell layers. Here we investigate TFF3-induced effects on the composition and function of tight junctions in these cells. TFF3 increased the cellular level of tightening claudin-1 and decreased the amount of claudin-2 known to form cation-selective channels. Expression of ZO-1, ZO-2 and occludin was not altered. The change in claudin-1 and -2 expression in TFF3-expressing HT29/B6 cells was accompanied by an increase in the transepithelial resistance in confluent monolayers of these cells. These data suggest that TFF3 plays a role in the regulation of intestinal barrier function by altering the claudin composition within tight junctions thus decreasing paracellular permeability of the intestinal mucosa.     

All-trans retinoic acid increases expression of aquaporin-5 and plasma membrane water permeability via transactivation of Sp1 in mouse lung epithelial cells

Aquaporin-5 (AQP5) is a water-selective channel protein that is expressed in lacrimal glands, salivary glands, and distal lung. Several studies using AQP5 knockout mice have revealed that AQP5 plays an important role in maintaining water homeostasis in the lung. We report here that all-trans retinoic acid (atRA) increases plasma membrane water permeability, AQP5 mRNA and protein expression, and AQP5 promoter activity in MLE-12 cells. The promoter activation induced by atRA was diminished by mutation at the Sp1/Sp3 binding element (SBE), suggesting that the SBE mediates the effects of atRA. In addition, atRA increased the binding of Sp1 to the SBE without changing the levels of Sp1 in the nucleus. Taken together, our data indicate that atRA increases AQP5 expression through transactivation of Sp1, leading to an increase in plasma membrane water permeability.     

Oxidative stress reduces histone deacetylase 2 activity and enhances IL-8 gene expression: role of tyrosine nitration

Oxidative stress is a characteristic of chronic inflammatory diseases. The reactive oxygen intermediate hydrogen peroxide (H(2)O(2)) is an important signaling molecule that modulates gene expression. We have demonstrated that H(2)O(2) significantly enhanced cytokine production in BEAS-2B cells, with a maximal effect at 4h. This did not result from enhanced NF-kappaB activation, but through decreased activity of histone deacetylase (HDAC)2. This results in increased inflammatory gene expression following acetylation of specific histone residues. Decreased HDAC2 activity was associated with tyrosine nitration status. Peroxynitrite and SIN-1, a peroxynitrite generator, were also able to reduce HDAC2 activity via tyrosine nitration. Our data suggest that oxidative stress contributes to worsening inflammation via reduction of HDAC2 activity through HDAC2 nitration. This novel mechanism of inflammation may be important in increasing the severity and chronicity of inflammatory diseases.     

Rhinovirus increases human beta-defensin-2 and -3 mRNA expression in cultured bronchial epithelial cells

Human beta-defensins (hBDs) are antimicrobial peptides that play important roles in host defense against infection, inflammation and immunity. Previous studies showed that micro-organisms and proinflammatory mediators regulate the expression of these peptides in airway epithelial cells. The aim of the present study was to investigate the modulation of expression of hBDs in cultured primary bronchial epithelial cells (PBEC) by rhinovirus-16 (RV16), a respiratory virus responsible for the common cold and associated with asthma exacerbations. RV16 was found to induce expression of hBD-2 and -3 mRNA in PBEC, but did not affect hBD-1 mRNA. Viral replication appeared essential for rhinovirus-induced beta-defensin mRNA expression, since UV-inactivated rhinovirus did not increase expression of hBD-2 and hBD-3 mRNA. Exposure to synthetic double-stranded RNA (dsRNA) molecule polyinosinic:polycytidylic acid had a similar effect as RV16 on mRNA expression of these peptides in PBEC. In line with this, PBEC were found to express TLR3, a Toll-like receptor involved in recognition of dsRNA. This study shows that rhinovirus infection of PBEC leads to increased hBD-2 and hBD-3 mRNA expression, which may play a role in both the uncomplicated common cold and in virus-associated exacerbations of asthma.     

Lysophosphatidic acid induces interleukin-13 (IL-13) receptor alpha2 expression and inhibits IL-13 signaling in primary human bronchial epithelial cells

Interleukin-13 (IL-13), a Th2 cytokine, plays a pivotal role in pathogenesis of bronchial asthma via IL-13 receptor alpha1 (IL-13Ralpha1) and IL-4 receptor alpha (IL-4Ralpha). Recent studies show that a decoy receptor for IL-13, namely IL-13Ralpha2, mitigates IL-13 signaling and function. This study provides evidence for regulation of IL-13Ralpha2 production and release and IL-13-dependent signaling by lysophosphatidic acid (LPA) in primary cultures of human bronchial epithelial cells (HBEpCs). LPA treatment of HBEpCs in at imedependent fashion increased IL-13Ralpha2 gene expression without altering the mRNA levels of IL-13Ralpha1 and IL-4Ralpha. Pretreatment with pertussis toxin (100 ng/ml, 4 h) or transfection of c-Jun small interference RNA or an inhibitor of JNK attenuated LPA-induced IL-13Ralpha2 gene expression and secretion of soluble IL-13Ralpha2. Overexpression of catalytically inactive mutants of phospholipase D (PLD) 1 or 2 attenuated LPA-induced IL-13Ralpha2 gene expression and protein secretion as well as phosphorylation of JNK. Pretreatment of HBEpCs with 1 microM LPA for 6 h attenuated IL-13-but not IL-4-induced phosphorylation of STAT6. Transfection of HBEpCs with IL-13Ralpha2 small interference RNA blocked the effect of LPA on IL-13-induced phosphorylation of STAT6. Furthermore, pretreatment with LPA (1 microM, 6 h) attenuated IL-13-induced eotaxin-1 and SOCS-1 gene expression. These results demonstrate that LPA induces IL-13Ralpha2 expression and release via PLD and JNK/AP-1 signal transduction and that pretreatment with LPA down-regulates IL-13 signaling in HBEpCs. Our data suggest a novel mechanism of regulation of IL-13Ralpha2 and IL-13 signaling that may be of physiological relevance to airway inflammation and remodeling.     

Differential expression of cholinephosphotransferase in normal and cancerous human mammary epithelial cells

Membrane phospholipids as well as fatty acid profile of cell membrane phospholipids are altered in tumorigenicity and malignancy. Synthesis of total cellular phosphatidylcholine (PC) can be used as a marker for membrane proliferation in neoplastic mammary gland tissues. Cholinephosphotransferase (CPT), the terminal enzyme in the de novo synthesis of PC, has an important role in regulating the acyl group of PC in mammalian cells. In this study, the effect of neoplasia on CPT was examined. The gene shows an elevated expression in cancerous (11-9-14) breast epithelial cell line when compared to that of normal non-tumorigenic (MCF-12A) breast epithelial cell line. Four nucleotide substitutions are observed in the cancer cell line. Of these, three are null mutations, but the third one shows an interesting serine to tyrosine substitution (at amino acid position 89 of our partial sequence which corresponds to position 323 of the CPT sequence reported as NM_020244 in GenBank) in 11-9-14 cells. The tyrosine is present in the right context of KSELYQDT, which directs tyrosine phosphorylation at the tyrosine site. Biochemical approach also reveals a 1.5-fold stimulation in CPT activity in 11-9-14 cells compared to that of the MCF-12A cells.     

Moderate folate depletion modulates the expression of selected genes involved in cell cycle, intracellular signaling and folate uptake in human colonic epithelial cell lines

Folate deficiency may affect gene expression by disrupting DNA methylation patterns or by inducing base substitution, DNA breaks, gene deletions and gene amplification. Changes in expression may explain the inverse relationship observed between folate status and risk of colorectal cancer. Three cell lines derived from the normal human colon, HCEC, NCM356 and NCM460, were grown for 32–34 days in media containing 25, 50, 75 or 150 nM folic acid, and the expression of genes involved in cell-cycle checkpoints, intracellular signaling, folate uptake and cell adhesion and migration was determined. Expression of Folate Receptor 1 was increased with decreasing media folate in all cell lines, as was p53, p21, p16 and β-catenin. With decreasing folate, the expression of both E-cadherin and SMAD-4 was decreased in NCM356. APC was elevated in NCM356 but unchanged in the other lines. No changes in global methylation were detected. A significant increase in p53 exon 7–8 strand breaks was observed with decreasing folate in NCM460 cells. The changes observed are consistent with DNA damage-induced activation of cell-cycle checkpoints and cellular adaptation to folate depletion. Folate-depletion-induced changes in the Wnt/APC pathway as well as in genes involved in cell adhesion, migration and invasion may underlie observed relationships between folate status and cancer risk.     

The effect of trans-10, cis-12 conjugated linoleic acid on gene expression profiles related to lipid metabolism in human intestinal-like Caco-2 cells

We conducted an in-depth investigation of the effects of conjugated linoleic acid (CLA) on the expression of key metabolic genes and genes of known importance in intestinal lipid metabolism using the Caco-2 cell model. Cells were treated with 80 μmol/L of linoleic acid (control), trans-10, cis-12 CLA or cis-9, trans-11 CLA. RNA was isolated from the cells, labelled and hybridized to the Affymetrix U133 2.0 Plus arrays (n = 3). Data and functional analysis were preformed using Bioconductor. Gene ontology analysis (GO) revealed a significant enrichment (P < 0.0001) for the GO term lipid metabolism with genes up-regulated by trans-10, cis-12 CLA. Trans-10, cis-12 CLA, but not cis-9, trans-11 CLA, altered the expression of a number of genes involved in lipid transport, fatty acid metabolism, lipolysis, β-oxidation, steroid metabolism, cholesterol biosynthesis, membrane lipid metabolism, gluconeogenesis and the citrate cycle. These observations warrant further investigation to understand their potential role in the metabolic syndrome.     

Profiling gene expression of whole cytochrome P450 superfamily in human bronchial and peripheral lung tissues: Differential expression in non-small cell lung cancers

Susceptibility to lung diseases, such as lung cancer and chronic obstructive pulmonary disease, is largely influenced by the metabolic capacity of lung tissues. This capacity is partly determined by the expression profile of the cytochromes P450 (CYPs), a superfamily of enzymes that have relevant catalytic properties toward exogenous and endogenous compounds. Using quantitative real-time RT-PCR, we conducted a comprehensive analysis of the expression profile of the 57 human CYP genes in non-tumoral (bronchial mucosa and pulmonary parenchyma) and tumoral lung tissues of 18 patients with non-small cell lung cancer. This study highlights (i) inter-individual variations in lung expression for some CYPs, (ii) different CYP expression patterns between bronchial mucosa and pulmonary parenchyma, that indicate distinctive susceptibility of these tissues toward the deleterious effects of inhaled chemical toxicants and carcinogens, (iii) high intertumoral variability, that could have major implications on lung tumor response to anti-cancer drugs.