Objective To establish the electric controlled cortical impact (eCCI)-induced traumatic brain injury (TBI) model in rats with different severity in degree,which may serve as a suitable platform to provide experimental evidence for the pathophysiological following TBI.Methods A total of 40 male Wistar rats were randomly divided into 3 experimental groups and sham group.TBI rats (n=10/group) were positioned beneath the controlled cortical impactor device (eCCI) and subjected to impact injury at 2 mm depth of penetration,for a sustained depression of 200 ms,at 4 m/s,5 m/s,6 m/s velocity for mild,moderate,and severe TBI,respectively.Sham-operated rats (n=10) underwent identical surgical procedures,including craniotomy,without receiving the cortical impact.Neurological function and regional cerebral flow (24 h after CCI),contusion volume,histopathological,and ultrastructural changes (48 h after CCI) were measured,respectively.Results The severity of the pathological changes in rats was increased as the injury aggravated.The eCCI device impacted the brain at 4 m/s,5 m/s,6 m/s velocity for mild,moderate,and severe TBI,respectively.TBI groups showed impaired neurological function,and decreased rCBF lower than that of sham-operated group (all P＜0.01).Furthermore,neuronal pathological abnormalities in TBI groups,including neuron shrinking,perineuronal vacuole,and structural abnormalities of mitochondria.Increased severity of injury was apparent following the increased level of the impacted velocity,and significant differences were observed between TBI groups (P＜0.05).Conclusion The TBI animal model with mild,moderate,and severe brain injury can be established successfully by 4 m/s,5 m/s,and 6 m/s of impact velocity respectively with the eCCI-6.3 device.The novel eCCI-induced TBI model in rats possibly serves as a novel useful approach in the development of TBI models.

ObjectiveTo investigate the roles of 11 β-hydroxysteroid dehydrogenase type 1 ( 11 β-HSD1 )on hippocampus of rat with chronic unpredictable mild stress (CUMS).MethodsTwenty-four male SpragueDawley rats were randomly divided into control group and depressive model group. Chronic unpredictable mild stress (CUMS) was used to make up depressive animal model.Behavioral changes were recorded by body weight measuring,sucrose consumption test (SCT) and open field test (OFT),respectively.The mRNA transcription of 11β-HSD1 in hippocampus tissues of the rats were detected by real-time RT-PCR,and the protein expression of 11β-HSD1 were detected by western blot and immunofluorescence.ResultsBcforc starting CUMS protocol,the rats exhibited equivalent weight and sucrose consumption.Twenty-eight days after CUMS protocol,behavior parameters such as body weight,sucrose consumption,nunber of crossing,and number of rearing were significantly decreased in rats exposed to CUMS group compared with control group (P ＜ 0.05,P ＜ 0.01 ).Correspondingly,realtime RT-PCR assays showed the mRNA expression of 11 β-HSD1 in the hippocampus of CUMS group,which was (31 ±9) ％ lower than that of control group.Meanwhile,the protein expression of it in CUMS group was lower than that of control group (P ＜ 0.05 ).Inmunofluorescence revealed that the number of positive 11 3-HSD1 cells was high (223 ± 13) in the control group,while the number was decreased prominently (92 ± 11 ) in the CUMS group (P ＜ 0.01 ).ConclusionDepressive behavior of rats is induced and the expression of 11 β-HSD1 in the hippocampus is decreased prominently by CUMS,the mechanism of which is at least related to the low expression of 11β-HSD1 and disturbance of glucocorticoid metabolism caused by CUMS.

Objective To study the effect of brain-derived neurotrophic factor (BDNF) on the environmental nutrition and neural differentiation of the transplanted stem cells under hypothermia.Methods The BDNF gene mediated by liposome was transfected into 293T cell line, and ELISA assay was applied to find the peak time of BDNF expression. When BDNF was highly expressed, the supernatant was collected for establishment of SD rat models of brain injury. The rats were divided into Group A (stem cell transplantation group) and Group B (stem cell transplantation and BDNF group). Rats in both groups were under hypothermia treatment for five days. Four and eight days later ( three days from rewarming), rat brain tissues were obtained to detect the expressions of proliferating cell nuclear antigen (PCNA), nestin, neuron-specific enolase (NSE) and glial fibrillary acidic protein (GFAP) by immunohistochemical method and to detect the apoptosis by in situ hybridization. Finally, the nerve function scores were obtained for evaluation of the nerve function. Results The ELISA showed that the high level of BDNF expression was at 48 to 60 hours after gene transfection. PCNA and nestin were highly expressed, while NES and GFAP showed nil or low level of expression in both groups at the fourth day after hypothermia, with little apoptotic cells especially in the Group B (P ＜0.05). The expressions of PCNA and nestin were decreased, but the expressions of NSE and GFAP were increased at the third day after rewarming. The positive rate of NSE expression in the Group B was much higher and the apoptotic cells were much less compared with the Group A ( P ＜ 0. 05 ). A better nerve score was obtained in the Group B. Conclusion BDNF can enhance the survival rate of the transplanted stem cells and induce their differentiation into neurons under hypothermia.

ObjectiveTo investigate the dynamic changes in microvascular ultrastructure in the cortex after the acute cerebral ischemia-reperfusion.Methods40 male rats were randomly divided into two groups（n=20 for sham operation group and cerebral ischemia-reperfusion group）. Cerebral ischemia-reperfusion model was produced using suture middle cerebral artery occlusion. Rats were sacrificed and the brain samples were adopted 1,3,12,72 h after ischemia-reperfusion, methyl methacrylate composite brain microvascular casting. The production of brain microvascular specimens, scanning electron microscopy of normal rat cerebral cortex microvessels and cerebral cortex of acute brain injury morphological changes in microvascular.ResultsCompared with the sham-operated group, cerebral ischemia-reperfusion in the cortex after the signs of vascular damage, then, vascular casting was to "bean" shape or even had a completely broken "tears candles" stump-like vascular casting, finally, to further the formation of a vascular zone cortex. ConclusionThe structural changes of brain microvascular in the cortex after acute cerebral ischemia-reperfusion is an important cause of cerebral microcirculation in rats.

Objective To estimate whether the ligation of bilateral internal carotid artery in combination with one vertebral artery can lead to chronic cerebral hypoperfusion. Methods The sham operation, 2?VO and 3?VO rat models were subjected to the matching operation. Four weeks after operation,the cortical blood flow was determined. The learning and memory abilities were measured with Morris water maze test eight weeks later,then the rats were sacrificed to observe the morphological change of hippocampal CA1 region. Results Compared with the sham operation group((47±8.797)ml·min-1·100 g-1),the cerebral blood flow of 2?VO((24.30±8.999)ml ·min-1·100 g-1) and 3?VO((9.870±2.208)ml·min-1·100 g-1) were significantly decreased (P<0.01). Compared with the sham operation group((8.33±4.88)s),escape latencies of Morris water maze of 2?VO group ((14.78±7.84)s) and 3?VO group((14.86±7.96)s) in the fifth days also presented significantly increased (P<0.01),but rare difference between the two groups. Compared with the sham operation group[ (37.20±9.21) s, (10.01.±2.91)times],the target quadrant swimming time and crossing times of 2?VO group((20.13±5.80)s, (6.60±3.19)times) and 3?VO group((20.05±5.76)s,(6.55±2.59)times) in the fifth days also presented signifi?cantly decreased (P<0.01). There were distinct pathomorphology changes in hippocampal CA1 region of the two groups. Conclusion The ligation of bilateral internal carotid artery in combination with one vertebral artery can lead to chronic cerebral hypoperfusion,and can make the similar ethology representation with the 2?VO models.

Objective To investigate the possible mechanism of aggregation and activation of neu-trophils(polymorphonuclear neutrophils,PMN)in mice with chlamydial pneumonitis. Methods C57BL/ 6 mice were inoculated intranasally with 3×103 inclusion-forming units(IFU)of Chlamydia muridarum(Cm) to induce the murine model of chlamydial pneumonitis. Samples of lung tissues collected at different time points after infection were stained by hematoxylin and eosin for histopathological assessment of inflammation. The levels of myelo-peroxidase(MPO)were detected for the evaluation of PMN aggregation. The mononu-clear cells were isolated from lung tissues. The inflammatory cells were counted with Giemsaˊs staining. CD11b+Gr1+ cell population and CD11b expression in lung mononuclear cells were analyzed by flow cytome-try. The expression of chemokines(MIP-2,LIX,KC and MCP-1)in lung tissues at mRNA level was meas-ured by RT-PCR. Results Chlamydial pneumonitis was induced in mice by intranasal inoculation of 3×103 IFU of Cm. Compared with the mice from control group,large amounts of inflammatory cells including PMN, monocytes and lymphocytes were induced in lung tissues of mice with Cm infection. PMN responded earlier than monocytes to the infection. The levels of MPO were significantly increased in mice with Cm infection and reached the highest level on the 7th day after infection. A decline in MPO levels was observed on the 14th day but the levels were still higher than those on day 0. The percentages and total numbers of CD11b+Gr1+ cells were significantly increased after Cm infection. Moreover,an increased expression of PMN CD11b was also detected by flow cytometry. The expression of chemokines(MIP-2,LIX,KC and MCP-1)was in-creased in lung tissues of mice after Cm infection. The results of the study indicated that Cm infection in-duced the expression of PMN chemoattractants,resulting in the recruitment of PMN. Conclusion The infil-tration and activation of PMN in lung tissues of mice were induced by Cm infection through increasing the ex-pression of chemokines. PMN played an important role in immune responses against Cm infection.

Objective To investigate the effect of silence regulatory protein 1 (Sirt1) on axonal outgrowth. Methods The hippocampal neurons was first isolated in vitro from rat embryos. The distribution and expression of Sirt1 were then detected 72 h later. The down-regulation of Sirt1 was induced by RNAi technology and up-regulation of Sirt1 was in-duced by overexpression of Sirt1 and resveratrol (RES). Immunofluorescence staining was used to examine the axon length. Results Immunofluorescence staining showed that Sirt 1 was located in neuronal cell body and neurite, especially in the distal axons. Down-regulation of Sirt1 significantly decreased axonal length compared with siRNA control group [(178.3 ± 3.2) μm vs. (110.2 ± 18.30) μm, P< 0.01 ]; Overexpression of Sirt1 significantly increased axonal length com-pared with eGFP control group [(178.3±3.2)μm vs (310.6±39.5)μm, P<0.01 ];Activation of Sirt1 by RES treatment al-so significantly increased axonal length compared with vehicle control group (DMSO treated group) [(291.7±13.2)μm vs. (525.1±49.1)μm, P<0.01 ]. Conclusions Sirt1 plays a key role in axonal growth which may be used as a potential thera-peutic target of axon regeneration.

Objective To investigate the effect of temperature sensitive umbilical cord mesenchymal stem cells (tsUCMSCs) transplantation under mild hypothermia environment on cognitive function of rats after severe traumatic brain injury (TBI).Methods Thirty-six male SD rats weighing (300 ± 20)g were subjected to severe TBI by fluid percussion device (2.5 atm) and managed with tsUCMSCs combined with mild hypothermia.Rats were divided into three groups:sham operation group,TBI group,and TBI + tsUCMSCs + mild hypothermia group (TBI + intervention group) according the random number table,with 12 rats per group.Latency to the platform and the time in targeted quadrant were recorded to evaluate the learning and memory with Morris water maze.Population spike (PS) and excitatory postsynaptic potential (EPSP) of long-term potential (LTP) were examined to evaluate synaptic plasticity of hippocampus.Phosphorylated Tau at Thr231 and phosphorylated GSK3 at Ser9 were detected by Western blot.Results Latency to the platform [(15.9 ±3.2) s vs (8.08 ±2.69) s,P＜0.01]was prolonged and time in targeted quadrant [(14.4 ± 3.0) s vs (36.3 ± 5.7) s,P ＜ 0.01] was shortened in TBI group at day 28 compared with sham operation group.Whereas,latency to the platform [(10.7 ± 2.8) s,at Ser9 were detected by Western blot.Results Latency to the platform [(15.9 ± 3.2) s vs (8.08 ± 2.69) s,P ＜ 0.01] was prolonged and time in targeted quadrant [(14.4 ± 3.0) svs (36.3 ± 5.7) s,P ＜0.01] was shortened in TBI group at day 28 compared with sham operation group.Whereas,latency to the platform [(10.7 ± 2.8) s,P ＜ 0.01] and time in targeted quadrant [(29.4 ± 4.4) s,P ＜ 0.05 |were reversed in TBI + intervention group.PS and EPSP of LTP were elevated in TBI group,but the elevation was suppressed in TBI + intervention group.Meanwhile,Tau hyperphosphorylation (0.80 ± 1.00vs 1.24 ±0.13,P＜0.05) and GSK3 deactivation (3.01 ±0.41 vs 1.27 ±0.22,P ＜0.01) were significantly reversed in TBI + intervention group compared with TBI group.Conclusion Combination of tsUCMSCs and mild hypothermia therapy can improve the TBI-induced cognitive deficit.

Objective To investigate the effect of mild hypothermia on proliferation and differentiation of neural stem cells (NGCs) in hippocampal subgranular zone after traumatic brain injury (TBI) and the underlying mechanism.Methods SD rats were divided into sham-injured group (only left dura mater exposed),hypothermia group (sham injury + mild hypothermia therapy for 72 hours),TBI group (unilateral fluid percussion was used to generate severe TBI),and TBI + hypothermia group (TBI + mild hypothermia therapy for 72 hours) according to the random number table,with 8 rats per group.Hippocampal homogenates or brain tissues were harvested after BrdU (100 mg/kg) was intraperitoneally administered to rats once a day for 7 days postTBI.Expressions of BrdU and double cortin in hippocampal subgranular zone were respectively detected by immunohistochemical or immunofiuorescence staining.Level of Sirt1 (silence information regulatory proteins,Sirt1) in hippocampus was detected by Western blot.Results BrdU-and double cortin-positive cells in rat hippocampal subgranular zone greatly increased at 7 days after TBI in comparison with sham-injured group (P ＜ 0.01).Moreover,BrdU and double cortin in rat hippocampal subgranular zone in TBI + hypothermia group was significantly higherthan that in TBI group [(257.4 ± 34.3) vs (196.4 ± 23.8) ; (346.4 ± 42.2) vs (245.7 ± 33.2),P ＜0.01].Moreover,mild hypothermia reversed TBI-induced over-expression of Sirt1 [(0.62 ± 0.075) vs(1.18 ± 0.11),P ＜ 0.01].Conclusion Mild hypothermia therapy can promote proliferation andneuronal differentiation of NSCs in hippocampal subgranular zone after TBI and the possible mechanismmay relate to the inhibition of over-expression of Sia1.

Objective To determine the effect of moderate hypothermia on coagulation in patients with severe traumatic brain injury (sTBI) and investigate the clinical significance of thrombelastogram (TEG) monitoring.Methods Seventy-five patients with sTBI were randomly assigned to hypothermia group (conventional treatment + moderate hypothermia within 24 hours posttrauma,n =38) and control group (conventional treatment alone,n =38).TEG aided in monitoring coagulation function by measuring clot reaction time (R),clot formation time (K),clotting rate (α),maximal amplitude (MA),and percent fibrinolysis at 30 minutes after MA (LY30).Meantime,the intracranial pressure,vital signs,blood gas values,and blood electrolytes were also measured.Outcome was evaluated by using Glasgow outcome scale (GOS).Results The two groups were similar on admission with respect to R,K,α,MA,and LY30 (P ＞ 0.05),but the coagulation index in hypothermia group was significantly different from that in control group at days 1,2,3 and 7 posttreatment (P ＜ 0.05).Moreover,moderate hypothermia therapy demonstrated decrease of intraeranial pressure (P ＜ 0.01),with no severe complications,low mortality and improved outcome in comparison with control group.Conclusion Moderate hypothermia improves the hypercoagulability in patients with sTBI without increasing the risk of hyperfibrinolysis and protects brain tissue by decreasing intracranial pressure.