title: Creatine and antioxidant treatment prevent the inhibition of creatine kinase activity and the morphological alterations of C6 glioma cells induced by the branched-chain alpha-keto acids accumulating in maple syrup urine disease.
abstract: Accumulation of the branched-chain alpha-keto acids (BCKA), alpha-ketoisocaproic acid (KIC), alpha-keto-beta-methylvaleric acid (KMV), and alpha-ketoisovaleric acid (KIV) and their respective branched-chain alpha-amino acids (BCAA) in tissues and biological fluids is the biochemical hallmark of patients affected by the neurometabolic disorder known as maple syrup urine disease (MSUD). Considering that brain energy metabolism is possibly altered in MSUD, the objective of this study was to determine creatine kinase (CK) activity, a key enzyme of energy homeostasis, in C6 glioma cells exposed to BCKA. The cells were incubated with 1, 5, or 10 mM BCKA for 3 h and the CK activity measured afterwards. The results indicated that the BCKA significantly inhibited CK activity at all tested concentrations. Furthermore, the inhibition caused by the BCKA on CK activity was totally prevented by preincubation with the energetic substrate creatine and by coincubation with the N-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthase inhibitor, indicating that deficit of energy and nitric oxide (NO) are involved in these effects. In contrast, other antioxidants such as glutathione (GSH) and trolox (soluble Vitamin E) were not able to prevent CK inhibition. In addition, we observed that the C6 cells changed their usual rounded morphology when exposed for 3 h to 10 mM BCKA and that creatine and L-NAME prevented these morphological alterations. Considering the importance of CK for brain metabolism homeostasis, it is conceivable that inhibition of this enzyme by increased levels of BCKA may contribute to the neurodegeneration of MSUD patients.
authors.affiliation: Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Rua Ramiro Barcelos 2600 anexo, 90035-003, Porto Alegre, RS, Brazil. csfunchal@yahoo.com.br
authors.forename: Cláudia
authors.initials: C
authors.lastname: Funchal
authors.forename: Patrícia Fernanda
authors.initials: PF
authors.lastname: Schuck
authors.forename: André Quincozes Dos
authors.initials: AQ
authors.lastname: Santos
authors.forename: Maria Caroline
authors.initials: MC
authors.lastname: Jacques-Silva
authors.forename: Carmem
authors.initials: C
authors.lastname: Gottfried
authors.forename: Regina
authors.initials: R
authors.lastname: Pessoa-Pureur
authors.forename: Moacir
authors.initials: M
authors.lastname: Wajner
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000378:metabolism / Q000627:therapeutic use*; D045744:Cell Line, Tumor; D048430:Cell Shape / Q000187:drug effects*; D003401:Creatine / Q000378:metabolism / Q000627:therapeutic use*; D003402:Creatine Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005910:Glioma / Q000378:metabolism; D045782:Hemiterpenes; D006801:Humans; D007651:Keto Acids / Q000378:metabolism / Q000494:pharmacology; D008375:Maple Syrup Urine Disease / Q000188:drug therapy / Q000378:metabolism; D051381:Rats
chemical_list: D000975:Antioxidants; D045782:Hemiterpenes; D007651:Keto Acids; C016211:alpha-keto-beta-methylvaleric acid; C001505:alpha-ketoisovalerate; C013082:alpha-ketoisocaproic acid; D003402:Creatine Kinase; D003401:Creatine