Caspase inhibitor z-DEVD-fmk attenuates calpain and necrotic cell death in vitro and after traumatic brain injury.
In studies designed to evaluate the therapeutic window for treatment of traumatic brain injury, the caspase 3 inhibitor z-DEVD-fmk improved neurologic function and reduced lesion volumes when administered at 1 but not at 4, 8, or 24 hours after injury. Moreover, neither caspase 3 nor PARP, a caspase 3 substrate, were cleaved in injured, untreated cortex from 1 to 72 hours after injury. Few cortical neurons expressed active caspase 3 or were TUNEL positive from 6 to 24 hours after injury, and TUNEL staining was primarily Type I (necrotic). Nissl staining revealed extensive neuronal necrosis in the injured cortex from 6 to 24 hours after impact. Considered together, these data suggested that z-DEVD-fmk may reduce neuronal necrosis, so we used an in vitro model of necrotic cell death induced by maitotoxin to test this further and explore the potential mechanism(s) involved. Z-DEVD-fmk (1 nM-100 microM) significantly attenuated maitotoxin induced neuronal cell death and markedly reduced expression of the 145 kD calpain-mediated alpha-spectrin breakdown product after maitotoxin injury. Neither the 120 kD caspase-mediated alpha-spectrin cleavage product nor cathepsin B were expressed after maitotoxin injury. In a cell free assay, z-DEVD-fmk reduced hydrolysis of casein by purified calpain I. Finally, z-DEVD-fmk reduced expression of the 145 kD calpain-mediated alpha-spectrin cleavage fragment after traumatic brain injury in vivo. These data suggest that neuroprotection by z-DEVD-fmk may, in part, reflect inhibition of calpain-related necrotic cell death.
Knoblach
Susan M
SM
Department of Neuroscience, Georgetown University Medical Center, Washington, DC 20057, USA. sknobl01@georgetown.edu
Alroy
Daniel A
DA
Nikolaeva
Maria
M
Cernak
Ibolja
I
Stoica
Bogdan A
BA
Faden
Alan I
AI
D000818:Animals; D001930:Brain Injuries / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D002154:Calpain / Q000037:antagonists & inhibitors* / Q000378:metabolism; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002474:Cell-Free System; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009336:Necrosis; D009410:Nerve Degeneration / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D009842:Oligopeptides / Q000494:pharmacology*
D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D009842:Oligopeptides; C110772:benzoylcarbonyl-aspartyl-glutamyl-valyl-aspartyl-fluoromethyl ketone; D002154:Calpain; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases