title: Evaluation of antioxidant drugs for the treatment of ochronotic alkaptonuria in an in vitro human cell model.
abstract: Alkaptonuria (AKU) is a rare autosomal recessive disease, associated with deficiency of homogentisate 1,2-dioxygenase activity in the liver. This leads to an accumulation of homogentisic acid (HGA) and its oxidized derivatives in polymerized form in connective tissues especially in joints. Currently, AKU lacks an appropriate therapy. Hence, we propose a new treatment for AKU using the antioxidant N-acetylcysteine (NAC) administered in combinations with ascorbic acid (ASC) since it has been proven that NAC counteracts the side-effects of ASC. We established an in vitro cell model using human articular primary chondrocytes challenged with an excess of HGA (0.33 mM). We used this experimental model to undertake pre-clinical testing of potential antioxidative therapies for AKU, evaluating apoptosis, viability, proliferation, and metabolism of chondrocytes exposed to HGA and treated with NAC and ASC administered alone or in combination addition of both. NAC decreased apoptosis induced in chondrocytes by HGA, increased chondrocyte growth reduced by HGA, and partially restored proteoglycan release inhibited by HGA. A significantly improvement in efficacy was found with combined addition of the two antioxidants in comparison with NAC and ASC alone. Our novel in vitro AKU model allowed us to demonstrate the efficacy of the co-administration of NAC and ASC to counteract the negative effects of HGA for the treatment of ochronotic arthropathy.
authors.affiliation: Sezione di Reumatologia, Dipartimento di Medicina Clinica e Scienze Immunologiche, Università degli Studi di Siena, Policlinico Le Scotte, Siena, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Tinti
authors.forename: Adriano
authors.initials: A
authors.lastname: Spreafico
authors.forename: Daniela
authors.initials: D
authors.lastname: Braconi
authors.forename: Lia
authors.initials: L
authors.lastname: Millucci
authors.forename: Giulia
authors.initials: G
authors.lastname: Bernardini
authors.forename: Federico
authors.initials: F
authors.lastname: Chellini
authors.forename: Giovanni
authors.initials: G
authors.lastname: Cavallo
authors.forename: Enrico
authors.initials: E
authors.lastname: Selvi
authors.forename: Mauro
authors.initials: M
authors.lastname: Galeazzi
authors.forename: Roberto
authors.initials: R
authors.lastname: Marcolongo
authors.forename: James A
authors.initials: JA
authors.lastname: Gallagher
authors.forename: Annalisa
authors.initials: A
authors.lastname: Santucci
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology / Q000627:therapeutic use*; D000474:Alkaptonuria / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D000975:Antioxidants / Q000494:pharmacology / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects; D001205:Ascorbic Acid / Q000494:pharmacology / Q000627:therapeutic use*; D002358:Cartilage, Articular / Q000166:cytology; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism; D050560:Homogentisate 1,2-Dioxygenase / Q000235:genetics / Q000378:metabolism; D006713:Homogentisic Acid / Q000378:metabolism; D006801:Humans; D009794:Ochronosis / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D050050:Protein Carbonylation
chemical_list: D000975:Antioxidants; D050560:Homogentisate 1,2-Dioxygenase; D006713:Homogentisic Acid; D001205:Ascorbic Acid; D000111:Acetylcysteine