title: Essential role of intracellular glutathione in controlling ascorbic acid transporter expression and function in rat hepatocytes and hepatoma cells.
abstract: Although there is in vivo evidence suggesting a role for glutathione in the metabolism and tissue distribution of vitamin C, no connection with the vitamin C transport systems has been reported. We show here that disruption of glutathione metabolism with buthionine-(S,R)-sulfoximine (BSO) produced a sustained blockade of ascorbic acid transport in rat hepatocytes and rat hepatoma cells. Rat hepatocytes expressed the Na(+)-coupled ascorbic acid transporter-1 (SVCT1), while hepatoma cells expressed the transporters SVCT1 and SVCT2. BSO-treated rat hepatoma cells showed a two order of magnitude decrease in SVCT1 and SVCT2 mRNA levels, undetectable SVCT1 and SVCT2 protein expression, and lacked the capacity to transport ascorbic acid, effects that were fully reversible on glutathione repletion. Interestingly, although SVCT1 mRNA levels remained unchanged in rat hepatocytes made glutathione deficient by in vivo BSO treatment, SVCT1 protein was absent from the plasma membrane and the cells lacked the capacity to transport ascorbic acid. The specificity of the BSO treatment was indicated by the finding that transport of oxidized vitamin C (dehydroascorbic acid) and glucose transporter expression were unaffected by BSO treatment. Moreover, glutathione depletion failed to affect ascorbic acid transport, and SVCT1 and SVCT2 expression in human hepatoma cells. Therefore, our data indicate an essential role for glutathione in controlling vitamin C metabolism in rat hepatocytes and rat hepatoma cells, two cell types capable of synthesizing ascorbic acid, by regulating the expression and subcellular localization of the transporters involved in the acquisition of ascorbic acid from extracellular sources, an effect not observed in human cells incapable of synthesizing ascorbic acid.
authors.affiliation: Departamento de Fisiopatología, Facultad de Ciencias Biológicas, Universidad de Concepción, Casilla 160C, Concepción, Chile.
authors.forename: Lorena
authors.initials: L
authors.lastname: Mardones
authors.forename: Felipe A
authors.initials: FA
authors.lastname: Zúñiga
authors.forename: Marcelo
authors.initials: M
authors.lastname: Villagrán
authors.forename: Kirsty
authors.initials: K
authors.lastname: Sotomayor
authors.forename: Pamela
authors.initials: P
authors.lastname: Mendoza
authors.forename: David
authors.initials: D
authors.lastname: Escobar
authors.forename: Mauricio
authors.initials: M
authors.lastname: González
authors.forename: Valeska
authors.initials: V
authors.lastname: Ormazabal
authors.forename: Mafalda
authors.initials: M
authors.lastname: Maldonado
authors.forename: Gloria
authors.initials: G
authors.lastname: Oñate
authors.forename: Constanza
authors.initials: C
authors.lastname: Angulo
authors.forename: Ilona I
authors.initials: II
authors.lastname: Concha
authors.forename: Alejandro M
authors.initials: AM
authors.lastname: Reyes
authors.forename: Juan G
authors.initials: JG
authors.lastname: Cárcamo
authors.forename: Valeria
authors.initials: V
authors.lastname: Barra
authors.forename: Juan Carlos
authors.initials: JC
authors.lastname: Vera
authors.forename: Coralia I
authors.initials: CI
authors.lastname: Rivas
mesh_terms: D000818:Animals; D001205:Ascorbic Acid / Q000008:administration & dosage; D001483:Base Sequence; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular / Q000378:metabolism* / Q000473:pathology; D017931:DNA Primers; D005978:Glutathione / Q000037:antagonists & inhibitors / Q000378:metabolism*; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D008113:Liver Neoplasms / Q000378:metabolism* / Q000473:pathology; D051381:Rats; D017207:Rats, Sprague-Dawley; D060347:Sodium-Coupled Vitamin C Transporters / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D060347:Sodium-Coupled Vitamin C Transporters; D019328:Buthionine Sulfoximine; D005978:Glutathione; D001205:Ascorbic Acid