title: Probenecid and N-Acetylcysteine Prevent Loss of Intracellular Glutathione and Inhibit Neuronal Death after Mechanical Stretch Injury In Vitro.
abstract: Probenecid and N-acetylcysteine (NAC) can preserve intracellular levels of the vital antioxidant glutathione (GSH) via two distinct biochemical pathways. Probenecid inhibits transporter-mediated GSH efflux and NAC serves as a cysteine donor for GSH synthesis. We hypothesized that probenecid and NAC alone would maintain intracellular GSH concentrations and inhibit neuronal death after traumatic stretch injury, and that the drugs in combination would produce additive effects. Sex-segregated rat primary cortical neurons were treated with probenecid (100 μM) and NAC (50 μM), alone and in combination (Pro-NAC), then subjected to mechanical stretch (10s-1strain rate, 50% membrane deformation). At 24 h, both probenecid and NAC inhibited trauma-induced intracellular GSH depletion, lactate dehydrogenase (LDH) release, and propidium iodide (PI) uptake in both XY- and XX-neurons. Combined Pro-NAC treatment was superior to probenecid or NAC alone in maintenance of intracellular GSH and neuronal death assessed by PI uptake. Interestingly, caspase 3 activity 24 h after mechanical trauma was more prominent in XX-neurons, and treatment effects (probenecid, NAC, and Pro-NAC) were observed in XX- but not XY-neurons; however, XY-neurons were ultimately more vulnerable to mechanical stretch-induced injury than their XX counterparts, as was evidenced by more neuronal death detected by LDH release and PI uptake. In addition, after stretch injury in HT22 hippocampal cells, both NAC and probenecid were highly effective at reducing oxidative stress detected by dichlorofluorescein fluorescence. These in vitro data support further testing of this drug combination in models of traumatic neuronal injury in vivo.
authors.affiliation: 1 Department of Critical Care Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.
authors.forename: Lina
authors.initials: L
authors.lastname: Du
authors.affiliation: 2 The Clinical and Translational Science Institute, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|3 Department of Pharmacy and Therapeutics, School of Pharmacy, University of Pittsburgh , Pittsburgh, Pennsylvania.
authors.forename: Philip E
authors.initials: PE
authors.lastname: Empey
authors.affiliation: 1 Department of Critical Care Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|4 Department of Neurosurgery, The First Affiliated Hospital of Nanjing Medical University , Nanjing, China .
authors.forename: Jing
authors.initials: J
authors.lastname: Ji
authors.affiliation: 1 Department of Critical Care Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.
authors.forename: Honglu
authors.initials: H
authors.lastname: Chao
authors.affiliation: 1 Department of Critical Care Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|5 Department of Pediatrics, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|6 Department of Anesthesiology, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|7 The Safar Center for Resuscitation Research and the Children's Hospital of Pittsburgh of UPMC , Pittsburgh, Pennsylvania.
authors.forename: Patrick M
authors.initials: PM
authors.lastname: Kochanek
authors.affiliation: 1 Department of Critical Care Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|5 Department of Pediatrics, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|7 The Safar Center for Resuscitation Research and the Children's Hospital of Pittsburgh of UPMC , Pittsburgh, Pennsylvania.|8 Department of Environmental and Occupational Health, Center for Free Radical and Antioxidant Health, University of Pittsburgh , Pittsburgh, Pennsylvania.
authors.forename: Hülya
authors.initials: H
authors.lastname: Bayır
authors.affiliation: 1 Department of Critical Care Medicine, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|3 Department of Pharmacy and Therapeutics, School of Pharmacy, University of Pittsburgh , Pittsburgh, Pennsylvania.|5 Department of Pediatrics, University of Pittsburgh School of Medicine , Pittsburgh, Pennsylvania.|7 The Safar Center for Resuscitation Research and the Children's Hospital of Pittsburgh of UPMC , Pittsburgh, Pennsylvania.
authors.forename: Robert S B
authors.initials: RS
authors.lastname: Clark
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology*; D000818:Animals; D000070642:Brain Injuries, Traumatic / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D005260:Female; D005978:Glutathione / Q000378:metabolism; D008297:Male; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D018696:Neuroprotective Agents / Q000494:pharmacology*; D011339:Probenecid / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013314:Stress, Mechanical
chemical_list: D018696:Neuroprotective Agents; D005978:Glutathione; D011339:Probenecid; D000111:Acetylcysteine
keywords: adenosine triphosphate binding cassette transporter C1; gender; multidrug resistance associated protein 1; primary cortical neurons; spinal cord injury; traumatic brain injury
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