title: A selective cysteine protease inhibitor is non-toxic and cerebroprotective in rats undergoing transient middle cerebral artery ischemia.
abstract: Ischemic neuronal injury mediated by cysteine proteases such as calpains and caspases has been demonstrated in various experimental models. Cathepsins B and L are also cysteine proteases which may contribute to neuronal death after ischemia. The authors measured in vitro and in vivo toxicity and post-ischemic cytoprotective effects of a cysteine protease inhibitor which does not block calpain or caspase but, rather, is relatively selective for cathepsins B and L. The compound belongs to the peptidyl-diazomethane family (cysteine protease inhibitor 1, termed CP-1). In vitro toxicity was measured using an assay of cell viability, and in vivo toxicity was measured by histological tissue analysis after infusion of CP-1 in rats. Two hours of middle cerebral artery (MCA) occlusion in rats was performed by the intravascular suture method. Immediately following reperfusion, intravenous infusion of CP-1 or vehicle was performed for 4 h at 0.9 ml/h. After a 7-day survival, the infarct volumes were measured. CP-1 was non-toxic to cultured glial cells to a local concentration of 200 microM, and relatively non-toxic to cultured endothelial cells at concentrations of 100-200 microM. No animal exhibited toxic effects at any of the doses used. Histologic comparisons revealed no signs of tissue toxicity. CP-1 significantly reduced hemispheric infarct volume compared to control (37+/-8.2%) at concentrations of 10, 50, and 250 microM [22+/-15%, P=0.008; 20+/-13%, P=0.002; 23+/-15%, P=0.022, respectively (mean+/-standard deviation; N=7-10 per group)]. CP-1, at the concentration of 50 microM, improved the functional score of the animals, but did not significantly alter cerebral blood flow. This study supports the hypothesis that the lysosomal cathepsins B and/or L contribute to cerebral injury after focal ischemia with reperfusion. Cysteine protease inhibitors which are relatively selective for cathepsins B and L, but not the calpains or caspases, are effective at reducing infarct volume after intravenous post-ischemic administration.
authors.affiliation: Department of Neurosurgery, Henry Ford Hospital, 2799 West Grand Boulevard, Detroit, MI 48202, USA. nsdos@neuro.hfh.edu
authors.forename: D M
authors.initials: DM
authors.lastname: Seyfried
authors.forename: R
authors.initials: R
authors.lastname: Veyna
authors.forename: Y
authors.initials: Y
authors.lastname: Han
authors.forename: K
authors.initials: K
authors.lastname: Li
authors.forename: N
authors.initials: N
authors.lastname: Tang
authors.forename: R L
authors.initials: RL
authors.lastname: Betts
authors.forename: S
authors.initials: S
authors.lastname: Weinsheimer
authors.forename: M
authors.initials: M
authors.lastname: Chopp
authors.forename: J
authors.initials: J
authors.lastname: Anagli
mesh_terms: D000818:Animals; D001786:Blood Glucose / Q000187:drug effects; D001921:Brain / Q000187:drug effects / Q000378:metabolism / Q000503:physiopathology; D002545:Brain Ischemia / Q000188:drug therapy* / Q000378:metabolism / Q000503:physiopathology; D002320:Cardiovascular Physiological Phenomena / Q000187:drug effects; D002401:Cathepsin B / Q000187:drug effects / Q000378:metabolism; D056668:Cathepsin L; D002403:Cathepsins / Q000187:drug effects / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects* / Q000502:physiology; D002560:Cerebrovascular Circulation / Q000187:drug effects / Q000502:physiology; D003546:Cysteine Endopeptidases; D015853:Cysteine Proteinase Inhibitors / Q000633:toxicity*; D003978:Diazomethane / Q000031:analogs & derivatives / Q000494:pharmacology; D004151:Dipeptides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D010450:Endopeptidases; D020244:Infarction, Middle Cerebral Artery / Q000188:drug therapy / Q000378:metabolism / Q000503:physiopathology; D007275:Injections, Intravenous; D008297:Male; D009410:Nerve Degeneration / Q000188:drug therapy* / Q000378:metabolism / Q000503:physiopathology; D018696:Neuroprotective Agents / Q000494:pharmacology*; D009498:Neurotoxins / Q000633:toxicity; D051381:Rats; D017208:Rats, Wistar; D015427:Reperfusion Injury / Q000188:drug therapy* / Q000378:metabolism / Q000503:physiopathology; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000378:metabolism
chemical_list: D001786:Blood Glucose; D015853:Cysteine Proteinase Inhibitors; D004151:Dipeptides; D018696:Neuroprotective Agents; D009498:Neurotoxins; C431444:carbobenoxy-phenylalanyl-serine(O-benzyl)diazomethane; D003978:Diazomethane; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; D002401:Cathepsin B; D056668:Cathepsin L; C534280:Ctsl protein, rat