title: Peroxynitrite-mediated glyoxalase I epigenetic inhibition drives apoptosis in airway epithelial cells exposed to crystalline silica via a novel mechanism involving argpyrimidine-modified Hsp70, JNK, and NF-κB.
abstract: Glyoxalase I (Glo1) is a cellular defense enzyme involved in the detoxification of methylglyoxal (MG), a cytotoxic by-product of glycolysis, and MG-derived advanced glycation end products (AGEs). Argpyrimidine (AP), one of the major AGEs coming from MG modification of protein arginines, is a proapoptotic agent. Crystalline silica is a well-known occupational health hazard, responsible for a relevant number of pulmonary diseases. Exposure of cells to crystalline silica results in a number of complex biological responses, including apoptosis. The present study was aimed at investigating whether, and through which mechanism, Glo1 was involved in Min-U-Sil 5 crystalline silica-induced apoptosis. Apoptosis, by TdT-mediated dUTP nick-end labeling assay, and transcript and protein levels or enzymatic activity, by quantitative real-time PCR, Western blot, and spectrophotometric methods, respectively, were evaluated in human bronchial BEAS-2B cells exposed or not (control) to crystalline silica and also in experiments with appropriate inhibitors. Reactive oxygen species were evaluated by coumarin-7-boronic acid or Amplex red hydrogen peroxide/peroxidase methods for peroxynitrite (ONOO(-)) or hydrogen peroxide (H2O2) measurements, respectively. Our results showed that Min-U-Sil 5 crystalline silica induced a dramatic ONOO(-)-mediated inhibition of Glo1, leading to AP-modified Hsp70 protein accumulation that, in a mechanism involving JNK and NF-κB, triggered an apoptotic mitochondrial pathway. Inhibition of Glo1 occurred at both functional and transcriptional levels, the latter occurring via ERK1/2 MAPK and miRNA 101 involvement. Taken together, our data demonstrate that Glo1 is involved in the Min-U-Sil 5 crystalline silica-induced BEAS-2B cell mitochondrial apoptotic pathway via a novel mechanism involving Hsp70, JNK, and NF-κB. Because maintenance of an intact respiratory epithelium is a critically important determinant of normal respiratory function, the knowledge of the mechanisms underlying its disruption may provide insight into the genesis, and possibly the prevention, of a number of pathological conditions commonly occurring in silica dust occupational exposure.
authors.affiliation: Department of Experimental Medicine University of Perugia, 06129 Perugia, Italy. Electronic address: cinzia.antognelli@unipg.it.
authors.forename: Cinzia
authors.initials: C
authors.lastname: Antognelli
authors.affiliation: Department of Medicine, School of Medicine, University of Perugia, 06129 Perugia, Italy.
authors.forename: Angela
authors.initials: A
authors.lastname: Gambelunghe
authors.affiliation: Department of Medicine, School of Medicine, University of Perugia, 06129 Perugia, Italy.
authors.forename: Giacomo
authors.initials: G
authors.lastname: Muzi
authors.affiliation: Department of Experimental Medicine University of Perugia, 06129 Perugia, Italy.
authors.forename: Vincenzo Nicola
authors.initials: VN
authors.lastname: Talesa
mesh_terms: D017209:Apoptosis; D002460:Cell Line; D004785:Environmental Pollutants / Q000633:toxicity*; D004794:Enzyme Repression / Q000187:drug effects; D044127:Epigenesis, Genetic; D004847:Epithelial Cells; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D017127:Glycation End Products, Advanced / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D007791:Lactoylglutathione Lyase / Q000235:genetics* / Q000378:metabolism; D035683:MicroRNAs / Q000502:physiology; D016328:NF-kappa B / Q000378:metabolism; D018384:Oxidative Stress; D030421:Peroxynitrous Acid / Q000494:pharmacology*; D020545:Respiratory Mucosa / Q000201:enzymology; D012822:Silicon Dioxide / Q000633:toxicity*
chemical_list: D004785:Environmental Pollutants; D017127:Glycation End Products, Advanced; D018840:HSP70 Heat-Shock Proteins; C534072:MIRN101 microRNA, human; D035683:MicroRNAs; D016328:NF-kappa B; D030421:Peroxynitrous Acid; D012822:Silicon Dioxide; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; C000597196:GLO1 protein, human; D007791:Lactoylglutathione Lyase
keywords: Apoptosis; Argpyrimidine-modified Hsp70; Crystalline silica Min-U-Sil 5; ERK1/2 MAPK; Free radicals; Glyoxalase I; Human bronchial BEAS-2B cells; JNK; MiR-101; NF-κB