Lithium enhances TRAIL-induced apoptosis in human lung carcinoma A549 cells

Non-small cell lung cancer (NSCLC) A549 cells are resistant to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis. Therefore, combination therapy using sensitizing agents to overcome TRAIL resistance may provide new strategies for treatment of NSCLC. Here, we investigated whether lithium chloride (LiCl), a drug for mental illness, could sensitize A549 cells to TRAIL-induced apoptosis. We observed that LiCl significantly enhanced A549 cells apoptosis through up-regulation of death receptors DR4 and DR5 and activation of caspase cascades. In addition, G2/M arrest induced by LiCl also contributed to TRAIL-induced apoptosis. Concomitantly, LiCl strongly inhibited the activity of c-Jun N-terminal kinases (JNKs), and the inhibition of JNKs by SP600125 also induced G2/M arrest and augmented cell death caused by TRAIL or TRAIL plus LiCl. However, glycogen synthase kinase-3β (GSK3β) inhibition was not involved in TRAIL sensitization induced by LiCl. Collectively, these findings indicated that LiCl sensitized A549 cells to TRAIL-induced apoptosis through caspases-dependent apoptotic pathway via death receptors signaling and G2/M arrest induced by inhibition of JNK activation, but independent of GSK3β.     

LEDGF binds to heat shock and stress-related element to activate the expression of stress-related genes

We have investigated the mechanism by which LEDGF protects cells against environmental stress. Our earlier report showed that a low level of LEDGF was present in the nucleus of most cell types and significant elevation of LEDGF level was induced by heat and oxidative stress. The cells overexpressing LEDGF-activated expression of heat shock proteins and enhanced survival of many cell types. Here we show that LEDGF binds to heat shock element (HSE) and stress-related regulatory element (STRE) to activate the expression of stress-related genes (Hsp27 and alphaB-crystallin). Apparently, HSE and STRE are present in promoters of many stress-related genes. Elevation of many stress-related proteins (STRPs) induced by LEDGF may protect cells against environmental stress. In yeast, it has been demonstrated that a single stress can activate the expression of multiple STRPs. This is known as "cross-protection," and now similar mechanism has been found in mammalian cells and LEDGF plays a vital role in it.     

Halothane affects focal adhesion proteins in the A 549 cells

Halothane is a volatile anaesthetic, which is known to induce alterations in cellular plasma membranes, modulating the physical state of the membrane lipids and/or interacting directly with membrane-bound proteins, such as integrin receptors. Integrin-mediated cell adhesion is a general property of eukaryotic cells, which is closely related to cell viability. Our previous investigations showed that halothane is toxic for A 549 lung carcinoma cells when applied at physiologically relevant concentrations and causes inhibition of adhesion to collagen IV. The present study is focused on the mechanisms underlying halothane toxicity. Our results imply that physiologically relevant concentrations of halothane disrupt focal adhesion contacts in A 549 cells, which is accompanied with suppression of focal adhesion kinase activity and paxillin phosphorylation, and not with proteolytic changes or inhibition of vinculin and paxillin expression. We suggest that at least one of the toxic effects of halothane is due to a decreased phosphorylation of the focal contact proteins.     

Hypoxia decreases proteins involved in epithelial electrolyte transport in A549 cells and rat lung

Fluid reabsorption from alveolar space is driven by active Na reabsorption via epithelial Na channels (ENaCs) and Na-K-ATPase. Both are inhibited by hypoxia. Here we tested whether hypoxia decreases Na transport by decreasing the number of copies of transporters in alveolar epithelial cells and in lungs of hypoxic rats. Membrane fractions were prepared from A549 cells exposed to hypoxia (3% O(2)) as well as from whole lung tissue and alveolar type II cells from rats exposed to hypoxia. Transport proteins were measured by Western blot analysis. In A549 cells, alpha(1)- and beta(1)-Na-K-ATPase, Na/K/2Cl cotransport, and ENaC proteins decreased during hypoxia. In whole lung tissue, alpha(1)-Na-K-ATPase and Na/K/2Cl cotransport decreased. alpha- and beta-ENaC mRNAs also decreased in hypoxic lungs. Similar results were seen in alveolar type II cells from hypoxic rats. These results indicate a slow decrease in the amount of Na-transporting proteins in alveolar epithelial cells during exposure to hypoxia that also occurs in vivo in lungs from hypoxic animals. The reduced number of transporters might account for the decreased transport activity and impaired edema clearance in hypoxic lungs.     

Characterization and expression pattern of IbPRP1 and IbPRP2 stress-related genes from sweetpotato

Two putative stress-related genes were isolated from sweet-potato and were designated as IbPRP1 and IbPRP2 (Ipomoea batatas proline-rich proteins 1 and 2). The deduced amino acid aligment of IbPRP1 and IbPRP2 shows that these two genes belong to the AAI_LTSS superfamily. Proteins in this family are known to play primary roles including defending plants from pathogens and insects, lipid transport between intracellular membranes, and nutrient storage. The mRNA expression of IbPRP1 and IbPRP2 were investigated and the results demonstrate that IbPRP2 has tissue-specific expression. Moreover, IbPRP1 and IbPRP2 may be involved in response to various stresses including drought, pathogen, and oxidative stress. In addition, when leaf disc test was performed, the IbPRP1 transgenic tobacco plants showed increase in tolerance to salt (100 mM, 200 mM, and 300 mM). Moreover, IbPRP1 and IbPRP2 may have some roles of transmembrane protein in sweetpotato when checked through GFP fusion cell localization and transmembrane analysis. All of these results indicate that IbPRP1 and IbPRP2 might play an important role in plant stress responses.     

Retroposons of Alu family from cis-regulatory modules of DNAse II and CAML genes effect gene expression in A549 and HEK293 cells

Different fragments of promoters of deoxyribonuclease II (DNAse II) and calcium-modulating cyclophilin ligand (CAML) associated with Alu family repeats have been inserted into a luciferase reporter vector. These constructions were introduced into A549 and HEK293 cell lines and after transient transfection we lysed cells and analysed luciferase activities in these lysates. It has been shown that Alu repeats localized in constructions influence expression of luciferase. Therefore, Alu copies which are associated with cis-regulatory modules of protein-coding genes have biological activity.     

Developmental expression of polyubiquitin genes and distribution of ubiquitinated proteins in generative and sperm cells

Polyubiquitin-encoding cDNA clones were isolated from the generative cells of lily (Lilium longiflorum) and the sperm cells of Plumbago zeylanica. The described genes encode identical amino acid sequences, with no homology outside the coding regions. This gene participates in ubiquitination of proteins, presumably enhancing protein turnover in the germline during male reproductive differentiation. In this paper we show that the gene encoding polyubiquitin is highly up-regulated in both Lilium generative cells and one of the Plumbago sperm cell types in particular.     

Contaminant loading in remote Arctic lakes affects cellular stress-related proteins expression in feral charr

The remote Arctic lakes on Bj?rn?ya Island, Norway, offer a unique opportunity to study possible affect of lifelong contaminant exposure in wild populations of landlocked Arctic charr (Salvelinus alpinus). This is because Lake Ellasj?en has persistent organic pollutant (POP) levels that are significantly greater than in the nearby Lake ?yangen. We examined whether this differential contaminant loading was reflected in the expression of protein markers of exposure and effect in the native fish. We assessed the expressions of cellular stress markers, including cytochrome P4501A (Cyp1A), heat shock protein 70 (hsp70), and glucocorticoid receptor (GR) in feral charr from the two lakes. The average polychlorinated biphenyl (PCB) load in the charr liver from Ellasj?en was approximately 25-fold higher than in individuals from ?yangen. Liver Cyp1A protein expression was significantly higher in individuals from Ellasj?en compared with ?yangen, confirming differential PCB exposure. There was no significant difference in hsp70 protein expression in charr liver between the two lakes. However, brain hsp70 protein expression was significantly elevated in charr from Ellasj?en compared with ?yangen. Also, liver GR protein expression was significantly higher in the Ellasj?en charr compared with ?yangen charr. Taken together, our results suggest changes to cellular stress-related protein expression as a possible adaptation to chronic-contaminant exposure in feral charr in the Norwegian high-Arctic.     

Gene expression profile in response to chromium-induced cell stress in A549 cells

Chromium (Cr) is a trace element required for life. Biological activities of Cr are complicated and remain to be fully investigated. It is known that the valence of Cr plays an important role in the biological activities of Cr. For example, Cr (VI) is classified as a metal carcinogen, but Cr (III) is widely used as a nutritional supplement. Establishment of a gene expression profile for Cr-induced cellular response is necessary to facilitate investigation of the biological activities of Cr. In the present study, a large-scale gene expression analysis was conducted using RNA of human lung epithelial cells after in vitro exposure to Cr (VI). Utilizing high-density oligonucleotide arrays representing 2400 genes, we observed that expression of 150 genes was up-regulated, and that of 70 genes were down-regulated by Cr (VI). Functional analysis of these responsive genes led to an outline of potential biological activities of Cr in six aspects. The gene expression profile reveals that Cr may involves in redox stress, calcium mobilization, energy metabolism, protein synthesis, cell cycle regulation and carcinogenesis in the cell. The results provide a critical clue for understanding molecular mechanisms of the biological activities of Cr.     

Effects of alginate oligomer on the expression of cell cycle- and stress-related genes in Chlamydomonas reinhardtii

Enzymatically prepared alginate oligomer (AO) promoted the growth of Chlamydomonas reinhardtii in a concentration-dependent manner. AO at 2.5 mg/mL induced increase in expression levels of cyclin A, cyclin B, and cyclin D in C. reinhardtii. CuSO₄ at 100 μM suppressed the growth of C. reinhardtiin, and AO at 2.5 mg/mL significantly alleviated the toxicity of CuSO₄. Increased intracellular reactive oxygen species level in C. reinhardtii induced by CuSO₄ was reduced by AO. After cultivation with CuSO₄ at 100 μM, expression levels of ascorbate peroxidase and superoxide dismutase in C. reinhardtii were increased, and AO reduced the increased levels of these enzymes. These results suggest that AO exhibits beneficial effects on C. reinhardtii through influencing the expression of various genes not only at normal growth condition but also under CuSO₄ stress.     

Propynylated phosphodiester oligonucleotides inhibit ICAM-1 expression in A549 cells on electroporation

Oligodeoxynucleotides (ODN) are used largely as either primers, antisense, or triplex-forming units. Phosphodiester ODN (PO-ODN), which are very rapidly degraded by exonucleases, must be protected at their ends. Even so, their life span inside cells is quite short. Phosphorothioate ODN (PS-ODN) are less sensitive to nucleases and are extensively used as antisense. Unfortunately, unlike PO-ODN, they interact with a number of molecules, including proteins, in addition to their specific nucleic acid targets. Their affinity for their target is lower than that of PO-ODN. PS-ODN containing propyne groups on C5 of pyrimidine have been shown to have a higher affinity toward their nucleic acid target. Here, we show that propynylated PO-ODN are more stable and much more efficient than their propyne-free counterparts. They are not efficient when they are used as lipoplexes, but they act as specific antisense on electroporation.     

布地奈德对A549细胞胸腺基质淋巴细胞生成素表达作用的研究

目的:观察细胞因子刺激气道上皮细胞胸腺基质淋巴细胞生成素(TSLP)表达是否涉及核因子κB(NF-κB),并探讨糖皮质激素布地奈德对气道上皮细胞TSLP表达和NF-κB核转位的影响.方法:A549细胞与细胞因子白介素1β(IL-1β)、白介素4(IL-4)和布地奈德共同孵育,以不加任何细胞因子或布地奈德培养的A549细胞为对照组,采用RT-PCR方法测定TSLP mRNA表达,细胞免疫荧光方法检测TSLP和NF-κB的表达情况.结果:与对照组比较,IL-1β(10 ng/ml)及IL-4(10 ng/ml)显著刺激A549细胞TSLP mRNA表达,且NF-κB(p65)核转住增加(均P＜0.05).布地奈德干预后TSLP mRNA的表达和NF-κB(p65)的核转位显著减少(P＜0.05).结论:细胞因子促进气道上皮细胞诱导性表达TSLP与NF-κB激活有关,抑制TSLP表达和NF-κB激活可能是布地奈德治疗哮喘的重要机制.     

Genotoxicity of hydroquinone in A549 cells

Hydroquinone (HQ) is found in natural and anthropogenic sources including food, cosmetics, cigarette smoke, and industrial products. In addition to ingestion and dermal absorption, human exposure to HQ may also occur by inhaling cigarette smoke or polluted air. The adverse effects of HQ on respiratory systems have been studied, but genotoxicity HQ on human lung cells is unclear. The aim of this study was to investigate the cytotoxicity and genotoxicity of HQ in human lung alveolar epithelial cells (A549). We found that HQ induced a dose response in cell growth inhibition and DNA damage which was associated with an increase in oxidative stress. Cytotoxicity results demonstrated that HQ was most toxic after 24 h (LC₅₀?=?33 μM) and less toxic after 1 h exposure (LC₅₀?=?59 μM). Genotoxicity of HQ was measured using the Comet assay, H2AX phosphorylation, and chromosome aberration formation. Results from the comet assay revealed that DNA damage was highest during the earlier hours of exposure (1 and 6 h) and thereafter was reduced. A similar pattern was observed for H2AX phosphorylation suggesting that damage DNA may be repaired in later exposure hours. An increase in chromosomal aberration corresponded with maximal DNA damage which further confirmed the genotoxic effects of HQ. To investigate whether oxidative stress was involved in the cytotoxic and genotoxic effects of HQ, cellular glutathione and 8-Oxo-deoguanisone (8-Oxo-dG) formation were measured. A decrease in the reduced glutathione (GSH) and an increase oxidized glutathione (GSSG) was observed during the early hours of exposure which corresponded with elevated 8-Oxo-dG adducts. Together these results demonstrate that HQ exerts its cytotoxic and genotoxic effects in A549 lung cells, probably through DNA damage via oxidative stress.     

The effect of temperature and pH gradients on Lactobacillus rhamnosus gene expression of stress-related genes

In this study, Lactobacillus rhamnosus, a renowned probiotic, was cultivated in fluctuating environment. Base gradients caused by a pH control in an industrial process and temperature gradients caused by uneven heating were simulated with a scale-down method. A pH gradient was created in a plug flow reactor (PFR). Expression of pH stress-related genes (atpA, aldB, cfa, groEL, hrcA and pstS) were studied as a relative gene expression study using ldhD as a reference gene. Expression measurements were carried out with the TRAC method. The responses of groEL, hrcA and atpA genes to temperature and pH changes were observed. The expression of phosphate uptake system-related pstS gene was induced almost linearly in the chemostat cultivation experiments when the base gradient in the PFR was increased. Correlations between the results from gene expression studies and freeze stability or acid stress survival were studied. However, by measuring the expression of these genes, we were not able to predict eventual freeze stability or survival from the acid stress test.     

Target genes involved in antiproliferative effect of modified ginseng extracts in lung cancer A549 cells

Lung cancer is the most common cancer and the leading cause of cancerrelated deaths. Panax ginseng has long been used to treat cancer and other diseases worldwide. Most of the pharmacological actions of ginseng are attribu ted to a variety of ginsenosides, which are often metabo lized by intestinal bacteria into more effective forms. In this study, we found that the antiproliferative activity of ginseng was increased after enzymatic processing of ginseng saponin （50% inhibitory concentration, 〉70μg/ml）. To elucidate the mechanism by which modified ginseng extract （MGX） induced cell death in human lung cancer cells, the gene expression profiles ofA549 cells regulated by MGX were assayed using Agilent PrimeView Human Gene Expression Arrays. The expression of 17 genes involved in the regulation of cell signaling, cell metabolism, transport, and cytoskeletonregulation was upregulated, whereas the expression of 16 genes implicated in invasion and metastasis and cellular metabolism was downregulated in MGX treated A549 cells. Moreover, nuclear staining with 4：6dia midino2phenyHndole revealed that MGX clearly caused nuclear condensation and fragmentation which are observed in apoptosis cell. These results elucidate crucial antieancer mechanisms of MGX and provide potential new targets for the assessment of anticancer activity of MGX.