title: Branched-chain amino acids accumulating in maple syrup urine disease induce morphological alterations in C6 glioma cells probably through reactive species.
abstract: In the present study, we investigated the effects of the branched-chain amino acids (BCAA) leucine (Leu), isoleucine (Ile) and valine (Val), which accumulate in maple syrup urine disease (MSUD), on C6 glioma cell morphology and cytoskeletal reorganization by exposing the cultured cells to 1 and 5 mM BCAA. We observed that cells showed a fusiform shape with processes after 3 h treatment. Cell death was also observed when cells were incubated in the presence of the BCAA for 3 and 24 h. Val-treated cells presented the most dramatic morphological alterations. Immunocytochemistry with anti-actin and anti-GFAP antibodies revealed that all BCAA induced reorganization of actin and GFAP cytoskeleton. Although phosphorylation regulates intermediate filament (IF) assembly/disassembly, we verified that the BCAA did not change the in vitro phosphorylation of IF proteins either in C6 cells or in slices of cerebral cortex of rats during development (9-, 12-, 17- and 21-day-old). Furthermore, we observed that 3 h cell exposure to 5 mM of each BCAA resulted in a marked reduction of reduced glutathione (GSH) levels and significantly increased nitric oxide production. Finally, we observed that the morphological features caused by the BCAA on C6 cells were prevented by the use of the antioxidants GSH (1 mM) and N(omega)-nitro-L-arginine methyl ester (L-NAME, 0.5 mM). On the basis of the present results, we conclude that free radical attack might be involved in the cell morphological alterations, as well as, in the cytoskeletal reorganization elicited by the BCAA. It is therefore presumed that these findings could be involved in the neuropathological features observed in patients affected by MSUD.
authors.affiliation: Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS, Brazil.
authors.forename: Priscila
authors.initials: P
authors.lastname: de Lima Pelaez
authors.forename: Cláudia
authors.initials: C
authors.lastname: Funchal
authors.forename: Samanta Oliveira
authors.initials: SO
authors.lastname: Loureiro
authors.forename: Luana
authors.initials: L
authors.lastname: Heimfarth
authors.forename: Ariane
authors.initials: A
authors.lastname: Zamoner
authors.forename: Carmem
authors.initials: C
authors.lastname: Gottfried
authors.forename: Alexandra
authors.initials: A
authors.lastname: Latini
authors.forename: Moacir
authors.initials: M
authors.lastname: Wajner
authors.forename: Regina
authors.initials: R
authors.lastname: Pessoa-Pureur
mesh_terms: D000199:Actins / Q000378:metabolism; D000367:Age Factors; D000597:Amino Acids, Branched-Chain / Q000378:metabolism* / Q000494:pharmacology; D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D045744:Cell Line, Tumor; D002531:Cerebellum / Q000166:cytology / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression / Q000187:drug effects; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D005910:Glioma / Q000378:metabolism* / Q000473:pathology*; D005978:Glutathione / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors
chemical_list: D000199:Actins; D000597:Amino Acids, Branched-Chain; D005904:Glial Fibrillary Acidic Protein; D009569:Nitric Oxide; D005978:Glutathione