Neuroprotective effect of diazoxide on brain injury induced by cerebral ischemia/reperfusion during deep hypothermia.
OBJECTIVE
The purpose of this study was to determine the effects of diazoxide on apoptosis and the relative mechanisms in a model of brain injury induced by cerebral ischemia/reperfusion (I/R) during deep hypothermia.


METHODS
Three-week-old Sprague-Dawley male rats were randomly and equitably divided into sham-operated group, placebo-treated group and diazoxide-treated group respectively. Specific examination of the regional cerebral blood flow (rCBF) was measured in the three groups continuously during the operation by laser Doppler flowmetry. Terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) was showed DNA fragmentation. The mRNA expressions of cytochrome c and full-length caspase-3 were determined by RT-PCR, while the protein expressions of cytochrome c and cleaved caspase-3 were determined by immunohistochemistry at 1 h, 6 h, 24 h, 72 h and 7 days after I/R, respectively. Cytosolic release of cytochrome c at 24 h after I/R was also confirmed by Western blot.


RESULTS
rCBF was significantly decreased in both of placebo-treated and diazoxide-treated group just after ischemia in the time interval 0-5 min, and had no obvious changes in all the time intervals during the operation. Diazoxide preconditioning significantly decreased the percentage of TUNEL-positive staining cells. The mRNA expressions of cytochrome c and full-length caspase-3 in diazoxide-treated group were significantly decreased. In addition, diazoxide provided a significant reduction in the protein expressions of cytochrome c and cleaved caspase-3.


CONCLUSIONS
These results suggested that the neuroprotective effects of diazoxide against cerebral I/R injury during deep hypothermia correlated with the reduction of DNA fragmentation, prevention of mitochondrial cytochrome c release and inhibition of caspase-3 activation.
He
Xiaomin
X
Department of Cardiothoracic Surgery, Children's Hospital Nanjing, Nanjing Medical, University, Nanjing, 210008, China.
Mo
Xuming
X
Gu
Haitao
H
Chen
Feng
F
Gu
Qun
Q
Peng
Wei
W
Qi
Jirong
J
Shen
Li
L
Sun
Jian
J
Zhang
Rufang
R
Kj
Yin
Y
D000704:Analysis of Variance; D000818:Animals; D001930:Brain Injuries / Q000209:etiology* / Q000503:physiopathology / Q000517:prevention & control*; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D002560:Cerebrovascular Circulation / Q000502:physiology; D045304:Cytochromes c / Q000235:genetics / Q000378:metabolism; D003981:Diazoxide / Q000627:therapeutic use*; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D007036:Hypothermia, Induced / Q000009:adverse effects*; D020287:In Situ Nick-End Labeling / Q000379:methods; D017078:Laser-Doppler Flowmetry / Q000379:methods; D008297:Male; D018696:Neuroprotective Agents / Q000627:therapeutic use*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000628:therapy*; D013997:Time Factors
D018696:Neuroprotective Agents; D045304:Cytochromes c; D053148:Caspase 3; D003981:Diazoxide