The Experimental Study On Neuro-protection Mechanism Of Hypothermia In Brain Injury Induced By Status Epilepticus

Objective Intraperitoneal injection of poliocarpine in young rats to induce ststus epilepticus, to study the neuro-protection of hypothermia on hippocampus neurons.Methods Sixty 3 weeks old male Wister rats were randomly devided into 3 groups. Normal saline (NS) group, SE30min with diazepam (SE30minD)group, SE30min with diazepam+hypothermia(SE30minDH) group,20 rats each group. SE30minD group was given poliocarpine 380mg/kg by intraperitoneal injection to induce SE 30min and diazepam 5mg/kg by intraperitoneal injection to end SE. SE30minDH group was given diazepam 5mg/kg by intraperitoneal injection and core temperature was controlled between 32-34℃for 2 hours by hypothermy with physicalmeasure. NS group was given normal saline by intraperitoneal injection. According to hour 3, 6,24 and 72 after SE, each group was divided into 4 subgroups. The necrosis and apoptosis in hippocampal CA1 and CA3 regions were appraised by HE and TUNEL staining.Results 1.Necrosis in hippocampal CA1 and CA3 regions (HE staining). Neurons in the NS group were arranged densely and clear,the neuron morphology of each time quantum was normal. In SE30minD group, cell necrosis was seen at 3h and become more obvious at 6h and 24h. At 72h, normal neuron decreased and the arrangement of neuron was loose.The nectotic neurons in CA3 region was more than that in in CA1 region. In SE30minDH group, cell necrosis was also seen at 3h but not become obvious at 6h and 24h. At 72h, the arrangement of neuron was slightly loose. from 6h,the percentage of necrotic neuron in CA1 and CA3 regions singanificantly decreased compared with that of SE30minDgroup. (2) Apoptosis in hippocampal CA1 and CA3 regions. The positive neurons in TUNEL staining sections were apoptotic neurons, in which the cell nucleus were presented brown-yellow fine particle shape, were occasionally found in in hippocampal CA1 and CA3 regions in NS group. From 6h in SE30minD group, TUNEL positive neurons started increasing, and at 72h obviously increased. It showed chromatin condensation,round or irregular apoptotic bodies, nuclear pkynosis. The apoptotic neurons in CA1 region was more than that in in CA3 region. From 6h, TUNEL positive neurons singanificantly increased compared with that of NS group P<0.05. In SE30minDH group, from 24h,TUNEL positive neurons singanificantly decreased compared with that of SE30minDgroup P<0.05.Conclusions 1.After SE, necrotic neurons were obvious from 3h, and apoptotic neurons were obvious from 6h. With time going by, necrotic and apoptotic neurons increased gruadually.2. After SE, mild hypothermia could protect against cell necrosis and apoptosis.Objective To study the mechanism of mild hypothermia protection on pathogenesis of brain injury,and to provide experimental data for clinical application of mild hypothermia for brain injury of following recurrent seizures in children. Methods A total of sixty 3 week old male Wister rats were randomly assigned into 3 groups according to different intervention methods and each group was divided into 4 subgroups according to different time point after SE. NMDAR1 and C-JUN protein expression were determined by immunohistochemistry,and NMDAR1 and C-JUN mRNA expressions were determined by real-time PCR.Results 1.Change of NMDARl:In NS group, NMDAR1-immunoreactivity positive neurons were mainly located in CA1 and CA3 pyramidal layer, and NMDAR1 positive products were also found in Dendrites. The expression of NMDAR1 protein was stable. In SE30minD group, from 6h, the expression of NMDAR1 protein started increasing, and at 72h obviously increased which were significant (P<0.05) compared with NS group. In SE30minDH group, from 24h, the expression of NMDAR1 protein singanificantly decreased compared with that of SE30minDgroup P<0.05. The expression of NMDAR1 mRNA was also stable in NS group. In SE30minD group, it increased and peaked at 3h,and slightly declined since 6h and maintained high level till 72h. In SE30minDH group, it rapidly declined at 24h and fell down to lower level at 72h which was significant (P<0.05)compared with SE30minD group.2.Change of C-JUN:In NS group, C-JUN-immunoreactivity positive neurons,in which the cell nucleus and cytoplasm were presented brown-yellow particle shape, were occasionally found in in hippocampal CA1 and CA3 regions. In SE30minD group, from 6h, C-JUN positive neurons started increasing, at 24h obviously increased, and at 72h slightly declined. From 24h, C-JUN positive neurons singanificantly increased compared with that of NS group P<0.05. In SE30minDH group, at 6h,C-JUN positive neurons singanificantly increased compared with that of SE30minD group P<0.05. From 24h, C-JUN positive neurons singanificantly declined compared with that of SE30minD group P<0.05. The expression of C-JUNmRNA had a basic level in NS group. In SE30minD group, it increased since 3h, slightly declined at 6h, increased again at 24h, slightly declined and remained at higher level at 72h, forming double peaks. In SE30minDH group, it obviously increased at 3h, declined t 6h and further declined at 24h to a lower level at 72h. there was significant difference (P<0.05) compared with SE group at 3 time points(3h,24h and 72h).Conclusions NMDARl and C-JUN were participated in of SE induced brain injury, mild hypothermia reduced neuron necrosis and apoptosis through down-regulateing expression of NMDAR1 gene and protein, up-regulateing early expression of C-JUN gene and protein, down-regulateing late expression of C-JUN gene and protein.