6-Hydroxydopamine induces cystatin C-mediated cysteine protease suppression and cathepsin D activation.
Alteration in the lysosomal system (LS) may represent a central mechanism in neurodegeneration. 6-Hydroxydopamine (6-OHDA) induces oxidative stress and cell death in catecholaminergic cells. The LS and caspases participate in apoptosis, although the mechanism(s) that is involved is not completely understood. Here, we show that Pheochromocytoma (PC12) cells exposed to 6-OHDA results in lysosomal dysregulation, caspase activation and cell death. Cells exposed to 6-OHDA increased expression and release of cystatin C (CC) and suppressed cathepsin B (CB). CB activity significantly declined 24h following exposure to 6-OHDA, however neutralization of CC restored CB activity. Cathepsin D (CD) and caspase-3 activity also increased following exposure to 6-OHDA. Inhibition of CD and caspase-3 with pepstatin A (PA) and DEVD-Cho, respectively, attenuated the 6-OHDA induced cell death at 48 and 72 h. However, the CB inhibitor CA-074 Me failed to protect cells. Additionally, poly-ADP-ribose polymerase (PARP) cleavage was evaluated after exposure to 6-OHDA and PA, CA-074 Me, and DEVD-Cho. Only DEVD-Cho significantly decreased PARP cleavage following exposure to 6-OHDA. Hence, caspase-3 mediated PARP cleavage following exposure to 6-OHDA appears independent of CB and CD alterations. These studies suggest alternate pathways and potential therapeutic targets of cell death associated with oxidative stress, CC, and lysosomal dysregulation.
Lee
Daniel C
DC
College of Pharmacy and Pharmaceutical Sciences, Florida A&M University, Tallahassee, FL 32307, USA.
Womble
Tracy A
TA
Mason
Ceceile W
CW
Jackson
Inneke M
IM
Lamango
Nazarius S
NS
Severs
Walter B
WB
Palm
Donald E
DE
D000818:Animals; D053148:Caspase 3 / Q000187:drug effects / Q000378:metabolism; D002401:Cathepsin B / Q000187:drug effects / Q000378:metabolism; D002402:Cathepsin D / Q000378:metabolism*; D016923:Cell Death / Q000187:drug effects / Q000502:physiology; D055316:Cystatin C; D015891:Cystatins / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004298:Dopamine / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008247:Lysosomes / Q000187:drug effects / Q000201:enzymology; D009474:Neurons / Q000187:drug effects / Q000201:enzymology*; D018696:Neuroprotective Agents / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D016627:Oxidopamine / Q000633:toxicity*; D016716:PC12 Cells; D010300:Parkinson Disease / Q000201:enzymology / Q000503:physiopathology; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D013565:Sympatholytics / Q000633:toxicity
C525031:Cst3 protein, mouse; C525032:Cst3 protein, rat; D055316:Cystatin C; D015891:Cystatins; D004791:Enzyme Inhibitors; D018696:Neuroprotective Agents; D013565:Sympatholytics; D016627:Oxidopamine; C499321:Parp1 protein, mouse; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D053148:Caspase 3; D003546:Cysteine Endopeptidases; D002401:Cathepsin B; D002402:Cathepsin D; D004298:Dopamine