The Impact Of Therapeutic Hypothermia On ROS, Apoptosis And Autophagy Of Hippocampus Neurocytes In A Rat Model Of Asphyxial Cardiac Arrest

Objective: To observe the impact of therapeutic hypothermia （TH） on the reactiveoxygen species （ROS） and expression of cacpase-3mRNA, LC3B and Parkin ofhippocampus nerve cells in cardiac arrest （CA） rats after cardiopulmonary resuscitation（CPR）.Methods:65healthy male Sprague Dawley（SD）adult rats, being randomly dividedto2groups: blank control group（n=5）and CPR group（n=60）. CA was induced in CPRgroup rats by asphyxia. The surviving rats were randomly divided into2groups:normothermia CPR group （NT） and hypothermia CPR group （HT）. Homeothermia of37℃was maintained on NT group after return of spontaneous circulation （ROSC）,hypothermal intervene of32℃was enforced on HT group for4hours immediately afterROSC. Two CPR groups were then randomly divided to2subgroups （NT-12, NT-24,HT-12, HT-24） according to the time of observation. During observation, the rats werescored by the neurological deficit scores （NDS）, then the bilateral hippocampi wereremoved from rats’ head, and monoplast suspension of fresh hippocampus tissue wasextracted immediately to determine the level of intracellular ROS by flow cytometry.Transmission electron microscope was used to observe the ultramicro changes of cellularnucleus and mitochondria. Reverse transcription-polymerase chain reaction （RT-PCR）was used to determine the expression of caspase-3mRNA and western-blotting （WB）was used to determine the expression of LC3B and Parkin of frozen hippocampus tissue.Measured data were analyzed with paired sample T test and One-Way ANOVA LSD-t.Results: of the60rats that underwent CA,44were successfully resuscitated （73%）and33survived to the end of the experiment （55%）. The NDS of NT and HT group rats were significantly reduced comparing to BC group （F=8.107, P<0.05）, which of HT-12and HT-24group rats were significantly increased comparing to the same time-point of NTgroup respectively （t=9.692, P<0.001; t=14.374, P<0.001）. The ROS in hippocampusnerve cells of NT and HT group rats were significantly increased comparing to BC group（F=16.824, P<0.05）, which of HT-12and HT-24group rats were significantly reducedcomparing to the same time-point of NT group respectively （t=9.836, P<0.001; t=7.499,P<0.001）. The expression of caspase-3mRNA in hippocampus nerve cells of NT and HTgroup rats were significantly increased comparing to BC group （F=24.527, P<0.05）,which of HT group rats were significantly reduced comparing to the same time-point ofNT group respectively （t=6.935, P<0.001; t=4.317, P<0.001）. The expression ofLC3B-II/I in hippocampus nerve cells of NT and HT group rats were significantlyincreased comparing to BC group （F=6.584, P<0.05）, which of HT group rats weresignificantly reduced comparing to the same time-point of NT group respectively（t=10.836, P<0.001; t=2.653, P=0.02）. The expression of Parkin in hippocampus nervecells of NT and HT group rats were significantly increased comparing to BC group（F=4.126, P<0.05）, which of HT group rats were significantly reduced comparing to thesame time-point of NT group respectively （t=4.027, P=0.001; t=5.505,P<0.001）.Ultramicro damages of nucleus and mitochondria in NT group were evidentcomparing to BC group, but the degree of damage of nucleus and mitochondria werereduced in HT group of rats comparing to NT group.Conclusion: TH reduced the injury of nerve cells and improved the neurologicalfunction of rats, likely by reducing ROS level of nerve cells and the cellular apoptosis andmassive autophagy including mitophagy after CPR.