Phosphorylation In A549 Cells Research Articles

IL-1β and TNF-α induce increased expression of CCL28 by airway epithelial cells via an NFκB-dependent pathway

CCL28 is a mucosal chemokine that attracts eosinophils and T cells via the receptors CCR3 and CCR10. Consequently, it is a candidate mediator of the pathology associated with asthma. This study examined constitutive and induced expression of CCL28 by A549 human airway epithelial-like cells. Real-time RT-PCR and ELISA of cultured cells and supernatants revealed constitutive levels of CCL28 expression to be low, whereas IL-1β and TNF-α, induced significantly increased expression. Observations from induced sputum and human airway biopsies supported this. Signal transduction studies revealed that IL-1β and TNF-α stimulation induced NFκB phosphorylation in A549 cells, but antagonist inhibition of NFκB p50–p65 phosphorylation correlated with marked reduction of IL-1β or TNF-α induced CCL28 expression. Together these studies imply a role for CCL28 in the orchestration of airway inflammation, and suggest that CCL28 is one link between microbial insult and the exacerbation of pathologies such as asthma, through an NFκB-dependent mechanism.

Human 8-oxoguanine DNA glycosylase increases resistance to hyperoxic cytotoxicity in lung epithelial cells and involvement with altered MAPK activity

It is unknown whether base excision DNA repair (BER) proteins interact with mitogen-activated protein kinases (MAPK) under oxidation. Here, we explored roles of BER proteins in signaling transduction involving MAPK during hyperoxia. We demonstrated that ERK1/2 phosphorylation in A549 cells was increased in 95% O2. p38 activity in A549 cells was also increased by exposure to 95% O2. To evaluate regulatory roles of MAPK, we have transduced A549 cells and primary alveolar epithelial type II cells (AECII) to overexpress 8-oxoguanine DNA glycosylase (hOgg1). Overexpression of hOgg1 reduced hyperoxic toxicity in A549 and AECII cells. Furthermore, protection by BER against hyperoxia appeared to involve an upregulation of ERK1/2 and downregulation of p38. These observations demonstrate, for the first time, that reduction of hyperoxic toxicity by BER proteins may be involved with MAPK activity, thereby impacting cell survival. Furthermore, our studies suggest that modulation of MAPK may be used in combination with BER proteins to counteract hyperoxic toxicity.

Sulindac sulfide and caffeic acid phenethyl ester suppress the motility of lung adenocarcinoma cells promoted by transforming growth factor-beta through Akt inhibition.

Cell migration is essential for invasive and metastatic phenotypes of cancer cells. Potential chemopreventive agents of cancer-sulindac sulfide, caffeic acid phenethyl ester (CAPE), curcumin, and (+)-catechin-have been reported to interfere with several types of intracellular signaling. In this study, we examined the effects of these agents on transforming growth factor-beta(TGF-beta)-induced motility and Akt phosphorylation in A549 cells. Judged by gold particle phagokinesis assay, sulindac sulfide, CAPE, and curcumin suppressed the motility of A549 cells promoted by TGF-beta. LY294002, a specific inhibitor of phosphatidylinositol 3-kinase(PI3K)/Akt signaling, also suppressed TGF-beta-induced motility and Akt phosphorylation. Sulindac sulfide and CAPE, but not curcumin, suppressed TGF-beta-induced Akt phosphorylation. We conclude that sulindac sulfide and CAPE suppress the motility promoted by TGF-beta in lung adenocarcinoma cells through the suppression of Akt. Our observations raise the possibility that these agents, except for (+)-catechin, can be applied not only as chemopreventive agents but also as anti-metastatic therapy.

Analysis of HSF-1 phosphorylation in A549 cells treated with a variety of stresses.

In the absence of stress, heat shock transcription factor-1 (HSF-1) exists as a monomer. After the treatment of cells with variety of stresses, HSF-1 forms a trimer and binds to the heat shock element (HSE), a motif consisting of three consecutive NGAAN sequences located in an inverted orientation upstream of the heat shock genes. HSF-1 is then phosphorylated causing transactivation of heat shock mRNAs. Treatment of cells with some of the stresses has been shown to increase HSF binding to HSE without detectably increasing the synthesis of heat shock mRNAs. Here we used antibody specific to HSF-1 to detect its phosphorylation status following exposure of A549, a human lung carcinoma cell line to a variety of stresses in order to correlate HSF-1 phosphorylation with its transactivation ability. Our studies show that HSF-1 is phosphorylated following heat shock (43 degrees C for 1 h), hypoxia (5 h exposure to 0.02% oxygen), 8% ethanol (1 h exposure at 37 degrees C), or 200 microM sodium arsenite (1 h exposure at 37 degrees C). All such stresses have previously been shown to increase the synthesis of heat shock proteins (hsps). However, there are no detectable increases in HSF-1 phosphorylation after the treatment of cells with X-irradiation (2-8 Gy) or 100 microM canavanine, an amino acid analogue (1 h exposure at 37 degrees C). Treatment of cells with X-irradiation increases HSF binding to HSE without increasing the synthesis of hsps, while treatment of cells with canavanine has been shown to increase the synthesis of hsps.(ABSTRACT TRUNCATED AT 250 WORDS)