title: Neuroprotective actions of methylene blue and its derivatives.
abstract: Methylene blue (MB), the first lead chemical structure of phenothiazine and other derivatives, is commonly used in diagnostic procedures and as a treatment for methemoglobinemia. We have previously demonstrated that MB could function as an alternative mitochondrial electron transfer carrier, enhance cellular oxygen consumption, and provide protection in vitro and in rodent models of Parkinson's disease and stroke. In the present study, we investigated the structure-activity relationships of MB in vitro using MB and six structurally related compounds. MB reduces mitochondrial superoxide production via alternative electron transfer that bypasses mitochondrial complexes I-III. MB mitigates reactive free radical production and provides neuroprotection in HT-22 cells against glutamate, IAA and rotenone toxicity. Distinctly, MB provides no protection against direct oxidative stress induced by glucose oxidase. Substitution of a side chain at MB's 10-nitrogen rendered a 1000-fold reduction of the protective potency against glutamate neurototoxicity. Compounds without side chains at positions 3 and 7, chlorophenothiazine and phenothiazine, have distinct redox potentials compared to MB and are incapable of enhancing mitochondrial electron transfer, while obtaining direct antioxidant actions against glutamate, IAA, and rotenone insults. Chlorophenothiazine exhibited direct antioxidant actions in mitochondria lysate assay compared to MB, which required reduction by NADH and mitochondria. MB increased complex IV expression and activity, while 2-chlorphenothiazine had no effect. Our study indicated that MB could attenuate superoxide production by functioning as an alternative mitochondrial electron transfer carrier and as a regenerable anti-oxidant in mitochondria.
authors.affiliation: Department of Pharmacology and Neuroscience, Institute for Alzheimer's Disease and Aging Research, University of North Texas Health Science Center at Fort Worth, Fort Worth, Texas, United States of America.
authors.forename: Ethan
authors.initials: E
authors.lastname: Poteet
authors.forename: Ali
authors.initials: A
authors.lastname: Winters
authors.forename: Liang-Jun
authors.initials: LJ
authors.lastname: Yan
authors.forename: Kyle
authors.initials: K
authors.lastname: Shufelt
authors.forename: Kayla N
authors.initials: KN
authors.lastname: Green
authors.forename: James W
authors.initials: JW
authors.lastname: Simpkins
authors.forename: Yi
authors.initials: Y
authors.lastname: Wen
authors.forename: Shao-Hua
authors.initials: SH
authors.lastname: Yang
mesh_terms: D000818:Animals; D002460:Cell Line; D004579:Electron Transport / Q000187:drug effects; D003576:Electron Transport Complex IV / Q000378:metabolism; D016166:Free Radical Scavengers / Q000737:chemistry* / Q000494:pharmacology*; D008751:Methylene Blue / Q000031:analogs & derivatives* / Q000494:pharmacology*; D051379:Mice; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D018696:Neuroprotective Agents / Q000737:chemistry* / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects; D017382:Reactive Oxygen Species / Q000378:metabolism; D013843:Thiazines / Q000494:pharmacology
chemical_list: D016166:Free Radical Scavengers; D018696:Neuroprotective Agents; D017382:Reactive Oxygen Species; D013843:Thiazines; D003576:Electron Transport Complex IV; D008751:Methylene Blue