The protective effect of PI3K/AKt/GSK3β signaling pathway on brain during resuscitation with neck cooling / 中华急诊医学杂志

ABSTRACT

Objective To study the changes of PI3K/Akt/GSK3β signaling pathway during resuscitation with neck cooling in order to explore the relationship between the protective effect of neck cooling and the phosphorylation of PI3K/Akt and GSK3β.Methods Thirty rabbits were randomly(random number) divided into five groups, and models of cadiac arrest were induced by ventricular fibrillation(VF, the positive electrode in the right ventricle and negative pole on the apex of heart) for 4 min.In sham group,a electrode was placed into right ventricle without electric current conducted, and CA was not induced.The rabbits were sacrificed and specimens were taken at 24 hours after modeling.In normothermia treat group(NT group),resuscitation was carried out to restoration of spontaneous circulation(ROSC),and the rabbits were sacrificed and specimens were taken at 24 hours after modeling.In intra-arrest therapeutic hypothermia group (IATH group), rapid neck cooling was initiated at the same time with CPR,and the target brain temperature was set at 34 ℃ maintained for 4 hours after ROSC.Rabbits were sacrificed and specimens were taken at 24 hours after modeling.In recovery period cooling + LY294002 group(PATH+LY294002 group), LY294002 was injected intra-ventricularly at 20 minutes before resuscitation.Rapid neck cooling was started at the same time with CPR,and the target brain temperature was set at 34 ℃ maintained for 4 hours after ROSC.The rabbits were sacrificed and specimens were taken at 24 hours after modeling.In post-arrest therapeutic hypothermia group (PATH group), rapid neck cooling was begun after CPR for 1 hour,and the target brain temperature was set at 34 ℃ maintained for 4 hours after ROSC.The rabbits were sacrificed and specimens were taken at 24 hours after modeling.Animals were sacrificed by using overdose anesthetic drug.Western blot was used to detect the level of Akt p-Akt GSK-3β p-GSK-3β (ser9) protein, and TUNEL was used to observe apoptosis of tissues in each group.Multiple comparisons were performed with one-way analysis of variance (ANOVA).Results Compared with Sham group, Akt (Thr-308) phosphorylation (P-AKT) and P-GSK-3β levels in the brain neuron cytoplasm in 24 hours after CPR resuscitation in NT group was significantly reduced, and showed a gradual reduction trend (P<0.05);the P-AKT and P-GSK-3β levels in the brain neuron cytoplasm in 24 hours after CPR resuscitation in IATH group were significantly enhanced compared with NT group (P<0.05);the levels of these two kinds of protein at one hour after resuscitation in PATH group were significantly enhanced compared with NT group (P<0.05), but lower in IATH group.Intra-ventricularly injection of LY294002 made the effect of hypothermia lost, indicating that LY294002 inhibited the phosphorylation of Akt.Apoptosis cells were significantly reduced in IATH group and normothermia theatment group compared with PATH group and LY294002 group(P<0.05).Conclusions Neck cooling can reduce apoptosis in rabbit brain cells after recovery, and the protective effect on brain is best in intra-arrest therapeutic hypothermia group.LY294002 specifically block the PI3K/Akt pathway, and the protective effect of cooling on the brain can be abolished,indicating hypothermia protects the neurological function via activation of PI3K/Akt pathway.Neck cooling protects the neurological function by activating PI3K/Akt/GSK-3β, promoting the Akt activation, and increasing the expression of P-GSK3β.Specific Akt inhibitor LY294002 inhibits Akt phosphorylation of brain tissue recovery and further inhibit the phosphorylation of GSK-3β, thus abolishing protective effect of cooling on neurological function.