title: Adaptive responses to the stress induced by hyperthermia or hydrogen peroxide in human fibroblasts.
abstract: Perturbation of oxidant/antioxidant cellular balance, induced by cellular metabolism and by exogenous sources, causes deleterious effects to proteins, lipids, and nucleic acids, leading to a condition named "oxidative stress" that is involved in several diseases, such as cancer, ischemia-reperfusion injury, and neurodegenerative disorders. Among the exogenous agents, both H(2)O(2) and hyperthermia have been implicated in oxidative stress promotion linked with the activation of apoptotic or necrotic mechanisms of cell death. The goal of this work was to better understand the involvement of some stress-related proteins in adaptive responses mounted by human fibroblasts versus the oxidative stress differently induced by 42 degrees C hyperthermia or H(2)O(2.) The research was developed, switching off inducible nitric oxide synthase (iNOS) expression through antisense oligonucleotide transfection by studying the possible coregulation in the expression of HSP32 (also named HO-1), HSP70, and iNOS and their involvement in the induction of DNA damage. Several biochemical parameters, such as cell viability (MTT assay), cell membrane integrity (lactate dehydrogenase release), reactive oxygen species formation, glutathione levels, immunocytochemistry analysis of iNOS, HSP70, and HO-1 levels, genomic DNA fragmentation (HALO/COMET assay), and transmembrane mitochondrial potential (deltaPsi) were examined. Cells were collected immediately at the end of the stress-inducing treatment. The results, confirming the pleiotropic function of i-NOS, indicate that: (i). HO-1/HSP32, HSP70, and iNOS are finely tuned in their expression to contribute all together, in human fibroblasts, in ameliorating the resistance to oxidative stress damage; (ii). ROS exposure, at least in hyperthermia, in human fibroblasts contributes to growth arrest more than to apoptosis activation; and (iii). mitochondrial dysfunction, in presence of iNOS inhibition seems to be clearly involved in apoptotic cell death of human fibroblasts after H(2)O(2) treatment, but not after hyperthermia.
authors.affiliation: Department of Biological Chemistry, Catania 95125, Italy.
authors.forename: S
authors.initials: S
authors.lastname: Grasso
authors.forename: C
authors.initials: C
authors.lastname: Scifo
authors.forename: V
authors.initials: V
authors.lastname: Cardile
authors.forename: R
authors.initials: R
authors.lastname: Gulino
authors.forename: M
authors.initials: M
authors.lastname: Renis
mesh_terms: D000975:Antioxidants / Q000378:metabolism; D016923:Cell Death / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D020552:Comet Assay; D004249:DNA Damage; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006419:Heme Oxygenase (Decyclizing) / Q000378:metabolism*; D051547:Heme Oxygenase-1; D006358:Hot Temperature; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D007150:Immunohistochemistry; D008565:Membrane Proteins; D008928:Mitochondria / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism; D016877:Oxidants / Q000378:metabolism*; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000975:Antioxidants; D018840:HSP70 Heat-Shock Proteins; D008565:Membrane Proteins; D016376:Oligonucleotides, Antisense; D016877:Oxidants; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496541:HMOX1 protein, human; D006419:Heme Oxygenase (Decyclizing); D051547:Heme Oxygenase-1; D005978:Glutathione