[Role of caspase-3, -8, and -9 in apoptosis of copper induced primary cortical neurons].
OBJECTIVE
This study aimed to investigate the mechanism of primary cortical neuron injury induced by high concentrations of copper by observing the effect of aceticum culture medium on apoptosis of rat primary cortical neurons and expression of cleaved caspase 3, caspase 8 and caspase 9.


METHODS
Primary cortical neurons were cultured for 72 hrs and then exposed to different concentrations of aceticum culture medium (20, 40 and 80 microM). The viability of neurons was detected by the MTT method. Apoptosis was observed by Hoechst33258 and flow cytometry Annexin V/PI. Expression of caspase 3, caspase 8 and caspase 9 was measured by Western blot.


RESULTS
Following incubation with aceticum culture medium, apoptosis of neurons was induced. Theviability of neurons was remarkably reduced and the rate of apoptosis was tremendously increased in a concentration dependent manner. Caspase 8 and caspase 9 were activated in 20 microM of copper aceticum culture medium 4 hrs after incubation and peaked at 48 hrs in various concentrations of copper aceticum culture medium, presenting with a time and concentration dependent manner. The activated caspase 3 was observed in 20 microM of copper aceticum culture medium 24 hrs after incubation, which was later than the activated caspase 8 and caspase 9. Caspase 3 expression reached a peak 48 hrs in various concentrations of copper aceticum culture medium, presenting with a time and concentration dependent manner.


CONCLUSIONS
The apoptosis of primary cortical neurons can be induced by copper. Caspase 3, caspase 8 and caspase 9 cascade reaction may involve in the apoptosis of copper induced rat primary cortical neurons.
Jiao
Xian-Ting
XT
Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, Shanghai 200092, China.
Liu
Xiao-Qing
XQ
Huang
Li-Su
LS
Zhang
Yong-Jun
YJ
Han
Lian-Shu
LS
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000502:physiology*; D053181:Caspase 8 / Q000502:physiology*; D053453:Caspase 9 / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects*; D003300:Copper / Q000633:toxicity*; D051381:Rats; D017207:Rats, Sprague-Dawley
D003300:Copper; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9