title: Administration of the Nrf2-ARE activators sulforaphane and carnosic acid attenuates 4-hydroxy-2-nonenal-induced mitochondrial dysfunction ex vivo.
abstract: The transcription factor NF-E2-related factor 2 (Nrf2) mediates transcription of antioxidant/cytoprotective genes by binding to the antioxidant-response element (ARE) within DNA. Upregulation of these genes constitutes a pleiotropic cytoprotective defense pathway, which has been shown to produce neuroprotection in numerous models by decreasing lipid peroxidation (LP) as measured by the neurotoxic LP by-product 4-hydroxynonenal (4-HNE). As neuronal mitochondria have previously been shown to be susceptible to insult-induced LP-mediated oxidative damage, we sought to mechanistically investigate whether Nrf2-ARE activation in vivo could protect mitochondria from subsequent 4-HNE exposure ex vivo. Young adult male CF-1 mice were administered one of two known Nrf2-ARE activators as single intraperitoneal doses-sulforaphane (SFP; 5.0mg/kg) or carnosic acid (CA; 1.0mg/kg)-or their respective vehicles 48 h before Ficoll isolation of rat cerebral cortical mitochondria. Purified mitochondria were then exposed ex vivo to 4-HNE for 15 min at 37 °C, which we showed to cause a concentration-related inhibition of mitochondrial respiration together with covalent binding of 4-HNE to mitochondrial proteins. We chose a 30 μM concentration of 4-HNE, which produced an approximately 50% inhibition of complex I- or complex II-driven respiration, to assess whether prior in vivo Nrf2-ARE-activating compounds would increase the resistance of the isolated cortical mitochondria to 4-HNE's mitotoxic effects. Administration of either compound significantly increased (p < 0.05) expression of heme oxygenase-1 mRNA in cortical tissue 48 h postadministration, verifying that both compounds were capable of inducing the Nrf2-ARE pathway. Moreover, the prior in vivo administration of SFP and CA significantly (p < 0.05) attenuated 4-HNE-induced inhibition of mitochondrial respiration for complex I, but only carnosic acid acted to protect complex II. Furthermore, both CA and SFP significantly (p < 0.05) reduced the amount of 4-HNE bound to mitochondrial proteins as determined by Western blot. These results demonstrate the capability of in vivo Nrf2-ARE induction to protect from 4-HNE toxicity to cortical mitochondria ex vivo. Ongoing studies will determine the therapeutic efficacy of Nrf2-ARE activators to attenuate traumatic brain injury-induced pathophysiology.
authors.affiliation: Spinal Cord & Brain Injury Research Center, University of Kentucky College of Medicine, Lexington, KY 40536-0509, USA.
authors.forename: Darren M
authors.initials: DM
authors.lastname: Miller
authors.forename: Indrapal N
authors.initials: IN
authors.lastname: Singh
authors.forename: Juan A
authors.initials: JA
authors.lastname: Wang
authors.forename: Edward D
authors.initials: ED
authors.lastname: Hall
mesh_terms: D045784:Abietanes / Q000494:pharmacology*; D000447:Aldehydes / Q000494:pharmacology; D000818:Animals; D016588:Anticarcinogenic Agents / Q000494:pharmacology; D061745:Antioxidant Response Elements / Q000502:physiology*; D000975:Antioxidants / Q000494:pharmacology; D019069:Cell Respiration / Q000187:drug effects; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000378:metabolism; D017879:Isothiocyanates; D008297:Male; D051379:Mice; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism*; D051267:NF-E2-Related Factor 2 / Q000378:metabolism*; D010936:Plant Extracts / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D013454:Sulfoxides; D013861:Thiocyanates / Q000494:pharmacology*
chemical_list: D045784:Abietanes; D000447:Aldehydes; D016588:Anticarcinogenic Agents; D000975:Antioxidants; D015853:Cysteine Proteinase Inhibitors; D017879:Isothiocyanates; D051267:NF-E2-Related Factor 2; C495636:Nfe2l2 protein, mouse; D010936:Plant Extracts; D012333:RNA, Messenger; D013454:Sulfoxides; D013861:Thiocyanates; D051547:Heme Oxygenase-1; C016766:sulforaphane; C027576:4-hydroxy-2-nonenal; C018381:salvin