title: An autophagic process is activated in HepG2 cells to mediate BDE-100-induced toxicity.
abstract: To reduce flammability and meet regulatory requirements, Brominated Flame Retardants (BFRs) are added to a wide variety of consumer products including furniture, textiles, electronics, and construction materials. Exposure to polybrominated phenyl ethers (PBDEs) adversely affects the human health. Bearing in mind that (i) PBDEs are potentially toxic, (ii) the mechanism of PBDE toxicity is unclear, and (iii) the importance of the autophagy to the field of toxicology is overlooked, this study investigates whether an autophagic process is activated in HepG2 cells (human hepatoblastoma cell line) to mediate BDE-100-induced toxicity. HepG2 cells were exposed with BDE-100 at three concentrations (0.1, 5, and 25μM), selected from preliminary toxicity tests, for 24 and 48h. To assess autophagy, immunocytochemistry was performed after exposure of HepG2 cells to BDE-100. Labeling of HepG2 cells with 100nM LysoTracker Red DND-99 aided examination of lysosome distribution. Proteins that are key to the autophagic process (p62 and LC3) were evaluated by western blotting. DNA was isolated and quantified to assess mitochondrial DNA copy number by qPCR on the basis of the number of DNA copies of a mitochondrial encoded gene normalized against a nuclear encoded gene. Conversion of LC3-I to LC3-II increased in HepG2 cells. Pre-addition of 100nM wortmannin decreased the amount of LC3 in the punctuate form and increased nuclear fragmentation (apoptotic feature). HepG2 cells exposed to BDE-100 presented increased staining with the lysosomal dye and had larger LC3 and p62 content after pre-treatment with ammonium chloride. The mitochondrial DNA copy number decreased, which probably constituted an attempt of the cell to manage mitochondrial damage by selective mitochondrial degradation (mitophagy). In conclusion, an autophagic process is activated in HepG2 cells to mediate BDE-100-induced toxicity.
authors.affiliation: Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Departamento de Análises Clínicas, Toxicológicas e Bromatológicas, Universidade de São Paulo, Av. Bandeirantes, 3900, Bairro Monte Alegre, Ribeirão Preto, CEP:14040901 São Paulo, Brazil. Electronic address: lilianmed2005@yahoo.com.br.
authors.forename: Lilian Cristina
authors.initials: LC
authors.lastname: Pereira
authors.affiliation: Universidade de Coimbra, Departamento de Ciências da Vida, Faculdade de Ciência e Tecnologia, Apartado 3046, 3001401 Coimbra, Portugal.
authors.forename: Filipe Valente
authors.initials: FV
authors.lastname: Duarte
authors.affiliation: Universidade de Coimbra, Departamento de Ciências da Vida, Faculdade de Ciência e Tecnologia, Apartado 3046, 3001401 Coimbra, Portugal.
authors.forename: Ana Teresa Inácio Ferreira
authors.initials: AT
authors.lastname: Varela
authors.affiliation: Universidade de Coimbra, Departamento de Ciências da Vida, Faculdade de Ciência e Tecnologia, Apartado 3046, 3001401 Coimbra, Portugal.
authors.forename: Anabela Pinto
authors.initials: AP
authors.lastname: Rolo
authors.affiliation: Universidade de Coimbra, Departamento de Ciências da Vida, Faculdade de Ciência e Tecnologia, Apartado 3046, 3001401 Coimbra, Portugal.
authors.forename: Carlos Manuel Marques
authors.initials: CM
authors.lastname: Palmeira
authors.affiliation: Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Departamento de Química, Universidade de São Paulo, Av. Bandeirantes, 3900, Bairro Monte Alegre, Ribeirão Preto, CEP:14040901 São Paulo, Brazil.
authors.forename: Daniel Junqueira
authors.initials: DJ
authors.lastname: Dorta
mesh_terms: D001343:Autophagy / Q000187:drug effects* / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D056945:Hep G2 Cells; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000096:biosynthesis; D011075:Polybrominated Biphenyls / Q000633:toxicity*
chemical_list: C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D011075:Polybrominated Biphenyls; C517827:pentabrominated diphenyl ether 100
keywords: Autophagy; BDE-100; HepG2 cells; Mitophagy