Minocycline protects PC12 cells against NMDA-induced injury via inhibiting 5-lipoxygenase activation.
Recently, we have reported that minocycline, a semi-synthetic tetracycline with neuroprotective effects, inhibits the in vitro ischemic-like injury and 5-lipoxygenase (5-LOX) activation in PC12 cells. In the present study, we further determined whether minocycline protects PC12 cells from excitotoxicity via inhibiting 5-LOX activation. We used N-methyl-d-aspartate (NMDA, 200 microM) to induce early (exposure for 6 h) and delayed (exposure for 6 h followed by 24 h recovery) injuries. We found that NMDA receptor antagonist ketamine, 5-LOX inhibitor caffeic acid and minocycline concentration dependently attenuated NMDA-induced early and delayed cell injuries (viability reduction and cell death). However, only ketamine (1 microM) inhibited NMDA-evoked elevation of intracellular calcium. In addition, immunohistochemical analysis showed that NMDA induced 5-LOX translocation to the nuclear membrane after 1- to 6-h exposure which was confirmed by Western blotting, indicating that 5-LOX was activated. Ketamine, caffeic acid and minocycline (each at 1 microM) inhibited 5-LOX translocation after early injury. After delayed injury, PC12 cells were shrunk, and 5-LOX was translocated to the nuclei and nuclear membrane; ketamine, caffeic acid and minocycline inhibited both cell shrinking and 5-LOX translocation. As a control, 12-LOX inhibitor baicalein showed a weak effect on cell viability and death, but no effect on 5-LOX translocation. Therefore, we conclude that the protective effect of minocycline on NMDA-induced injury is partly mediated by inhibiting 5-LOX activation.
Song
Ying
Y
Department of Pharmacology, School of Medicine, Zhejiang University, Hangzhou 310031, PR China.
Wei
Er-Qing
EQ
Zhang
Wei-Ping
WP
Ge
Qiu-Fu
QF
Liu
Jian-Ren
JR
Wang
Meng-Ling
ML
Huang
Xiao-Jia
XJ
Hu
Xin
X
Chen
Zhong
Z
D000704:Analysis of Variance; D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000502:physiology*; D015153:Blotting, Western / Q000379:methods; D002118:Calcium / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018690:Excitatory Amino Acid Agonists / Q000633:toxicity*; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D044950:Flavanones / Q000494:pharmacology; D007150:Immunohistochemistry / Q000379:methods; D007649:Ketamine / Q000494:pharmacology; D008911:Minocycline / Q000494:pharmacology*; D016202:N-Methylaspartate / Q000633:toxicity*; D018696:Neuroprotective Agents / Q000494:pharmacology*; D016716:PC12 Cells / Q000187:drug effects* / Q000473:pathology; D051381:Rats; D013778:Tetrazolium Salts; D013844:Thiazoles; D013997:Time Factors
D004791:Enzyme Inhibitors; D018690:Excitatory Amino Acid Agonists; D018691:Excitatory Amino Acid Antagonists; D044950:Flavanones; D018696:Neuroprotective Agents; D013778:Tetrazolium Salts; D013844:Thiazoles; C006680:baicalein; D016202:N-Methylaspartate; D007649:Ketamine; D001094:Arachidonate 5-Lipoxygenase; C022616:thiazolyl blue; D008911:Minocycline; D002118:Calcium