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Cells phenotypically adapt to alterations in their intra- and extracellular environment via organised alterations to gene and protein expression. Many chemical and physical stimuli are known to drive such responses, including the induction of oxidative stress and heat shock. Increasing use of mobile telephones in our society, has brought focus on the potential for radio frequency (microwave) electromagnetic radiation to elicit biological stress responses, in association with potentially detrimental effects of this to human health. Here we review evidence suggesting altered gene and protein expression in response to such emissions, with particular focus on heat shock proteins. Non-thermal induction of heat shock proteins has been claimed by a number of investigations in in vitro cellular systems, and appears pleiotropic for many other regulatory events. However, many of these studies are flawed by inconsistencies in exposure models, cell types used and the independent reproducibility of the findings. Further, the paucity of evidence from in vivo experimentation is largely contradictory. Therefore, the validity of these effects in human health risk assessment remain unsubstantiated. Where possible, suggestions for further experimental clarification have been provided.

Intracellular oxidative stress is a dynamic situation characterized by the accumulation of reactive oxygen metabolites, such as hydrogen peroxide. This is traditionally associated with both macromolecular damage and adaptive changes in gene expression, aimed at preventing cellular demise. However, the overall extent of such genetic changes is not well characterized. Here we present a comprehensive analysis of altered mRNA profiles in human A549 type II lung epithelial cells in response to hydrogen peroxide, at concentrations failing to induce necrotic toxicity. The results of an Affymetrix-based screen of the steady-state levels of mRNAs for several thousand genes revealed a complex pattern of transcriptional and/or posttranscriptional response to oxidative stress, which can be functionally related to both the oxidation and repair of damaged DNA, the induction and permanency of cell cycle arrest, and caspase-3 activation. Many of the genetic events can be related to activation of the p53/p21 pathway, but many other novel inductions and suppressions were detected, revealing the intricacy of the response. The data also disclosed a potential interaction between hydrogen peroxide treatment and increased sensitivity to cell killing by TRAIL, which could be functionally confirmed at the level of induction of caspase-3 activity.

Redox modification of proteins is proposed to play a central role in regulating cellular function. However, high-throughput techniques for the analysis of the redox status of individual proteins in complex mixtures are lacking. The aim was thus to develop a suitable technique to rapidly identify proteins undergoing oxidation of critical thiols by S-glutathionylation. The method is based on the specific reduction of mixed disulfides by glutaredoxin, their reaction with N-ethylmaleimide-biotin, affinity purification of tagged proteins, and identification by proteomic analysis. The method unequivocally identified 43 mostly novel cellular protein substrates for S-glutathionylation. These include protein chaperones, cytoskeletal proteins, cell cycle regulators, and enzymes of intermediate metabolism. Comparisons of the patterns of S-glutathionylated proteins extracted from cells undergoing diamide-induced oxidative stress and during constitutive metabolism reveal both common protein substrates and substrates failing to undergo enhanced S-glutathionylation during oxidative stress. The ability to chemically tag, select, and identify S-glutathionylated proteins, particularly during constitutive metabolism, will greatly enhance efforts to establish posttranslational redox modification of cellular proteins as an important biochemical control mechanism in coordinating cellular function.

Human A549 lung epithelial cells were challenged with 18 O-labeled hydrogen peroxide ([ 18 O]-H 2 O 2 ), the total RNA and DNA extracted in parallel, and analyzed for 18 O-labeled 8-oxo-7,8-dihydroguanosine ([ 18 O]-8-oxoGuo) and 8-oxo-7,8-dihydro-2′-deoxyguanosine ([ 18 O]-8-oxodGuo) respectively, using high-performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC-MS/MS). [ 18 O]-H 2 O 2 exposure resulted in dose-response formation of both [ 18 O]-8-oxoGuo and [ 18 O]-8-oxodGuo and 18 O-labeling of guanine in RNA was 14–25 times more common than in DNA. Kinetics of formation and subsequent removal of oxidized nucleic acids adducts were also monitored up to 24 h. The A549 showed slow turnover rates of adducts in RNA and DNA giving half-lives of approximately 12.5 h for [ 18 O]-8-oxoGuo in RNA and 20.7 h for [ 18 O]-8-oxodGuo in DNA, respectively. [ABSTRACT FROM AUTHOR]

During situations of oxidative stress phenotypic adaptation to altered redox state is achieved by changes in expression of selected genes. The mechanisms regulating this may involve reversible S-glutathionylation of cellular proteins. In this study we compared and contrasted changes in gene expression patterns in human type II lung epithelial A549 cells and human endothelial ECV304 cells in correlation to glutathione oxidation and the formation of glutathione–protein mixed disulphides, after exposure to subtoxic levels of hydrogen peroxide, formed in the medium by addition of glucose oxidase, or the thiol oxidant diamide. Both the number of specific mRNAs and their levels of induction were grossly correlated to the degree of S-glutathionylation of cellular protein. Thus, diamide induced the expression of a variety of protein and DNA chaperones and transcriptional regulators, particularly in ECV304 cells. On the other hand, the peroxide failed to induce many of these species, in association with only minimal disturbances to glutathione homeostasis. The induction of the chaperone responses at the level of mRNA was clearly shown to translate into a more resistant morphological phenotype in response to both heat shock and oxidative stress induced by the DNA-damaging pro-oxidant potassium bromate. [Copyright &y& Elsevier]