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Objective To investigate the effects of umbilical cord blood neural stem cells (UCBNSCs) via stromal cell-derived factor-1 (SDF-1)/CXC chemokine receptor 4 (CXCR4) signaling on neural recovery in rat models ofintracerebral hemorrhage.Methods (1) In migration assay in vitro,UCBNSCs were distributed in the upper wells of Transwell plates,and SDF-1 at concentrations of 30,60 and 120 ng/mL was placed in the lower wells.(2) Sixty rat models of intracerebral hemorrhage were randomly divided into UCBNSCs-transplanted group and phosphate buffer (PBS)-transplanted group (n=30);two d after modeling,10 L UCBNSCs suspension and same amount of PBS were,respectively,transplanted into the two groups,and intraperitoneal injection of deoxyuridine (BrdU) labeled endogenous neural stem cells was performed;neurological functions were assessed with modified Neurological Severity Scale (mNSS) one d,and one and two weeks after cell transplantation;the expressions of SDF-1,vascular endothelial growth factor (VEGF),glial fibrillar acidic protein (GFAP),and doublecortin (DCX) were detected by immunofluorescence when the rats were sacrificed two weeks after cell transplantation;cell apoptosis was detected by TUNEL.Results (1) In the in vitro experiment,CXCR4 expression could be detected in UCBNSCs;60 ng/mL SDF-1 had the greatest migration effect on UCBNSCs,and this effect showed statistically significant difference as compared with that at other concentrations (P＜0.05).(2) In the in vivo experiment,two weeks after transplantation,the UCBNSCs-transplanted group had significantly increased number of BrdU-labeled cells in the subventricular zone,and significantly larger number of BrdU/DCX and BrdU/GFAP cells than the PBS-transplanted group (P＜0.05);the VEGF expression in the brain injury area of the UCBNSCs-transplanted group ([88.30±7.21]/field) was significantly higher than that of PBS-transplanted group ([53.20±4.45]/field,t=4.144,P=0.000);the number of apoptotic cells in the brain injury area of the UCBNSCs-transplanted group ([34.30 ±2.44] /field) was significantly smaller than that of PBS-transplanted group ([47.70±1.98] /field,t=4.266,P=0.001);two weeks after transplantation,the mNSS scores of UCBNSCs-transplanted group (6.40±0.163) were significantly lower than those of the PBS-transplanted group (7.50±0.17,t=4.714,P=0.002).Conclusion SDF-1/CXCR4 can reach the injured area of cerebral hemorrhage after chemotactic transplantation of UCBNSCs and promote the recovery of nerve function in rats,whose mechanism may be that it can promote neurogenesis and VEGF secretion and inhibit apoptosis.
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Objective To explore the effect of R-spondin3 on the proliferation of neural stem cells (NSCs) and its mechanism in mice.Methods The mouse NSCs derived from the subventricular zone of E 14-15d CD 1 mice were confirmed by immunofluorescence assay.The NSCs after 3 passages of culture were chosen and randomly divided into 2 groups (V=1 mL).In the experimental group,0.8 μL of R-spondin3 with an initial concentration of 50 μg/mL was added (final concentration:40 ng/mL) while in the control group an equal amount of culture fluid was added.The proliferation of the cells in the 2 groups was detected by 5-Bromo-2-deoxy Uridine (BrdU) kits after the cells were treated by R-spondin3 for 6 hours.The protein expression of [β-catenin was measured by western blotting after the cells were treated by R-spondin3 for 4 and 8 hours.Results Under optical microscopy,the round and bright cells grew in culture medium and easily accumulated to become neurospheres.Immunofluorescence assay showed that over 90％ of the cells expressed Nestin and SOX2 and that some of them expressed NeuN or GFAP after induced differentiation.Brdu proliferation test showed that the proliferation rate of Brdu+/DAPI+ for the experimental group (1.56±0.03) was significantly higher than that for the control group (1.04±0.04) (P＜0.05).Western blotting showed that the expression levels of [β-catenin were increased at both 4h and 8h after treatment for the experimental group (1.09±0.10 and 1.20±0.13),significantly higher than those for the control group (0.56±0.05 and 0.83±0.04) (P＜0.05).Conclusions R-spondin3 can promote in vitro proliferation of NSCs in mice,which may be associated with activated Wnt/β-catenin signal pathways.
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Objective To investigate the effect ofmiR-184 on proliferation of neural stem cells (NSCs) and its mechanisms in mice.Methods The pHBLV-U6-GFP-miR-184 over-expression plasmid and pHBLV-U6-GFP-miR-184 inhibitor plasmid were used to construct recombinant lentivirus.And the NSCs derived fiom subventricular zone of E14d CD1 mouse were confirmed by immunofluorescence assay.There were four groups that contain a miR-184 overexpression group,a miR-184 inhibitor group and two control groups.The NSCs which infected with lentiviral vectors were selected for puromycin resistance for 5-7 days,and then surviving cells were cultivated to three generations.The expression level ofmiR-184 was detected by real time-quantitative PCR (RT-qPCR).And the target genes ofmiR-184 were predicted through TargetScan,IRTarBase and MiRanda,and were confirmed by Western blotting and RT-qPCR.The cells in the four groups were culttared under proliferating conditions incorporated bromodeoxyuridine (BrdU) in cell proliferation analyses.The protein expressions of Hesl and Hes5,the target proteins of Notch signaling pathways,and their mRNA expressions were detected by Western blotting and RT-qPCR.Results There were 90％ of cells in each group expressing both Nestin and Sox2.The miR-184 level in the miR-184 overexpression group was 67.63±7.53 times of that of the control group,with significant difference (P＜0.05).The percent of BrdU+/DAPI+ of the miR-184 overexpression group was 1.47±0.05 times of that in the control group,with significant difference (P＜0.05);and the percent of BrdU+/DAPI+ of the miR-184 inhibitor group was 0.84±0.03 times of that in the inhibitor control group,with significant difference (P＜0.05).Numbl was a target gene ofmiR-184 indicated by IRTarBase and MiRanda.The miR-184 could inhibit Numbl protein expression;the Numbl protein expression level in the miR-184 overexpression group was 0.73±0.07times of that in the control group,and the Numbl protein expression level in the miR-184 inhibitor group was 1.30±0.05 times of that in the control group,with significant difference (P＜0.05);but miR-184 did not change the Numbl mRNA level.The miR-184 could activate Notch signaling pathway through inhibiting the Numbl protein expression,and increase the Hes1 and Hes5 protein and mRNA expression levels (P＜0.05).Conclusion The miR-184 promotes the NSCs proliferation through inhibiting the Numbl protein translation and further activating the Notch signaling pathway.
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Objective To investigate the effects of growth differentiation factor 11 (GDF11) on proliferation of mouse neural stem cells (NSCs) and expression levels of transforming growth factor (TGF)-β/Smads and Wnt/β-Catenin signal key proteins.Methods NSCs,derived from the subventricular zone of E14 d CD1 mice,were cultured and induced differentiation;specific proteins nestin and SOX2 were confirmed by immunofluorescence assay.Neuron marker nucleus antigen (NeuN)and astrocyte marker glial fibrillary acidic protein (GFAP) were identified by immunofluorescent staining.The cells of third generation in their exponential phase were chosen and randomly divided into experimental group (adding GDF11 to make the final concentration as 40 ng/mL) and control group (adding equal amount of culture fluid).The proliferation of the cells in the two groups was detected by 5-ethynyl-2'-deoxyuridine (EdU) kits and protein expressions of Smad2/3,phosphorylated (p)-Smad2/3,Smad4 and β-Catenin were measured by Western blotting one and 6 h after treatment.Results Round and bright cells suspended in culture medium were observed through optical microscope.Immunofluorescence assay showed that over 90％ cells expressed both nestin and SOX2,and some of them expressed NeuN or GFAP.EdU proliferation test showed that the percentage of EdU positive cells in the experimental group (0.34±0.08) was significantly higher than that in the control group (0.24±0.03,P＜0.05).Western blotting showed that the expression levels ofp-Smad2/3,Smad4 and β-Catenin were significantly increased one and 6 h after treatment as compared with those in the control group (P＜0.05).Conclusion GDF11 can promote the proliferation of NSCs in vitro and probably is on account of activating TGF-β/Smads and Wnt/β-Catenin signal pathways.
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Objective To investigate the effect of galangin on proliferation and apoptosis of glioma cells in vitro.Methods (1) The glioma cells U87 and U25 1were divided into blank control group,DMSO group,100,200,300 and 400 μmol/L galangin treatment groups.MTT was used to study the effects of drugs on the proliferation of U251 and U87 cells.(2) Hoechest staining was used to observe cell apoptosis in the presence of different concentrations of galangin (0,100 and 200 μmol/L).(3) Flow cytometry was employed to detect the apoptosis of U251 and U87 cells in the presence of different concentrations of galangin (1 00 and 200 μmol/L).(4) Western blotting was employed to detect the expressions of apoptosis-related protein 3-Catenin,B-cell lymphoma-2 (Bcl-2),Bcl-2 related protein gene (Bax),cleaved-caspase-3,cleaved-caspase-9 and poly (ADP-ribose) polymerase (PARP) in the presence of different concentrations of galangin.Results (1) The proliferation of U251 and U87 cells was obviously inhibited atter 100,200,300 and 400 μmol/L galangin treatments,and dose-effect relation was noted.The concentrations of galangin at half rate of inhibition (IC50) were 281,321,276 and 229 μmol/L in U251 cells,and 289.4,261.1,247.4 and 225.3 μ mol/L in the U87 cells after 100,200,300 and 400μmol/L galangin treatments for 24 h.(2) Under the action of galangin,corresponding increase in apoptosis rates of U251 and U87 cells was noted following the increase of galangin concentrations (0,100 and 200 μmol/L),with significant differences (P＜0.05).(3) The detection of cell apoptosis by flow cytometry found similar changes.(4) Western blotting results indicated that galangin at the concentration of 0,100 and 200 μmol/L could significantly decrease the expressions of apoptosis-related protein 3-Catenin and Bcl-2,and increase the Bax,cleaved-caspase-3 and cleaved-caspase-9,and cleaved-PARP expressions;significant differences were noted between each two concentrations (P＜0.05).Conclusion Galangin can inhibit proliferation of glioma cells U251 and U87,and induce mitochondrial pathway of apoptosis via Wnt/β-Catenin signaling.
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Metastatic encephaloma from lung carcinoma is one of the nost common intracranial tumors,which is paid to more and more attentions in Neurosurgery.Patients with metastatic neoplasm suffer great physiological and psychological pain for short survival time and survival of poor quality.In terms of treatment,surgery is the major method for metastatic encephaloma of lung cancer.But the subject range of surgery is limited to many factors,therefore many patients can not be treated with surgery.In order to solve the issue of applicability restriction,diversified adjuvant treatment strategies appear successively,such as chemotherapy,targeted therapy,immunotherapy.It has been verified by a number of clinical evidences that these therapies can prolong lifetime and improve living quality of patients.But at the present stage,there is no an uniform standard for the treatment of metastatic encephaloma from lung carcinoma.To give full play the role of each treatment,it is important to understand the advantages and disadvantages.Only in this way,can the patient's interests be optimized.
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Objective To review systematically the clinical curative effects and time window of hyperbaric oxygen treatment of persistent vegetative state (PVS).Methods All the clinical research articles about hyperbaric oxygen for PVS from January 1990 to December 2013 were retrieved from China national knowledge internet,Wanfang Database and Vip Database (the Chinese key words:vegetative state,the vegetable,or hyperbaric oxygen),and then,a Meta-analysis was conducted.Results Search terms yielded 165 pieces of articles and 19 were included in the final analysis for treatment efficacy;the treatment group included 939 patients and the control group included 659 patients.Totally,17 pieces of articles of time window were included;the patients with onset＜60 d group included 700 patients and the patients with onset＞ 60 d group included 330 patients.The Jadad scores of included articles were not high in general,and the highest scores were two points.Comparable baseline data were demonstrated in all of the articles.Data from articles were pooled and analyzed,and the results showed that the effective rate of treatment group and control group were 67.51％ and 34.45％,with significant difference (Z=12.16,P=0.000,odd ratio=0.25,95％ confidence interval=0.20-0.31).The effective rate of patients with onset＜60 d group and patients with onset≥60 d group was 22.73％ and 63.29％ (Z=9.72,P=0.000,odd ratio=5.21,95％ confidence interval=3.74-7.27) Conclusion Hyperbaric oxygen treatment enjoys better treatment efficacy than conventional treatment;patients with onset＜60 d have better prognosis than patients with onset≥ 60 d.
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Objective To investigate the calcium/calmodulin-dependent serine protein kinase (CASK) expression in the hippocampus of epileptic rat models and its mechanism.Methods Thirty healthy adult male Sprague-Dawley rats were randomly divided into 6 groups (n=5):control group and experimental groups of 1,3,7,14 and 30 d after kindling by lithium chloride-pilocarpine;CASK expressions in the hippocampus tissues were measured by immunohistochemistry,immunoflurescence and Western blotting.Another 21 healthy adult male Sprague-Dawley rats were divided into 3 groups (n=7):transfected group,empty adenoviral group and control group,and after anesthesia with chloral hydrate,the same volume of CASK-RNAi-LV,LV-scrRNAi and normal saline was given,respectively;the behavior changes of rats in the three groups were observed within one h of kindling;the expression of N-methel-D-aspartate receptor subunit 2b (NMDAR2b,the downstream of CASK complex),in the transfected group and control group was further examined by Western blotting.Results CASK expressed only in neurons not gliacyte;and the number of CASK positive cells in experimental groups was larger than that in the control group.CASK expression in the hippocampus reduced to minimum level one d after kindling,and maintained at increased levels until 30 days.Both seizure frequency and seizure grade of the epilepsy in the transfected group were significantly decreased as compared with those in the empty adenoviral group and control group (P＜0.05).NMDAR2b expression in the hippocampus of control group increased at the first 24 h of kindling and maintained for 3 days,and then,it decreased 10 days after kindling;NMDAR2b expression in the hippocampus of transfected group was significantly reduced as compared with that in the control group at all time points of kindling (P＜0.05).Conclusions The CASK expression of epilepsy rat models is increased,which influences the occurrence and development of epilepsy.CASK is linked to epilepsy via its action on synaptic NMDAR2b,which may be the potential target of anti-epileptic drugs.
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BACKGROUND:With the medical development, prognostic outcomes of spinal cord injuries have not been improved significantly, and most patients also suffer from severe complications. Nowadays, lots of laboratories and clinical researches have suggested that celltherapy has a great potential, especial y the application of umbilical cord blood stem cells in nervous system diseases. OBJECTIVE:To explore the feasibility and clinical effect of umbilical cord blood neural stem cells transplantation for patients with obsolete spinal cord injury. METHODS:Umbilical cord blood was harvested from newborns under aseptic condition, and differentiated into neural stem cells in vitro that were prepared into cellsuspension at a concentration of 109/L. The cellsuspension (3 mL) was injected via the L 3-4 or L 4-5 into the subarachnoid space. American Spinal Injury Association (ASIA) scores and the residual urine were assessed before and 3 months after transplantation. RESULTS AND CONCLUSION:After transplantation, al the patients showed a stable life indication. Three months later, ASIA scores were increased and the residual urine decreased, which significantly differed from those before transplantation (P<0.05). These findings indicate that umbilical cord blood neural stem cells transplantation is a new treatment that can improve the limb function and life quality of patients with obsolete spinal cord injury.
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Objective To evaluate the clinical effect of functional neuronavigation combined with intra-operative magnetic resonance imaging (iMRI) in resection of symptomatic occipital lobe epilepsy.Methods Sixteen patients with symptomatic occipital lobe epilepsy,admitted to our hospital from January 2011 to April 2012,were involved in present study.All patients underwent surgical operation assisted by functional neuronavigation.Before the operation,MRI scanning,imaging fusing,scope of lesion excision describing,and operative approach designing were performed successively.Three-D reconstructed navigation plan was integrated into neuronavigation system and used by combining with functional neuronavigation under microscope during the surgical procedure.And,the iMRI was performed in five of them; the operative plan of the residuary lesion was redesigned accordingly.After the operation,the regular clinical and iconography follow-up were in progress.Results The functional neuronavigation was successfully completed.The lesions,posterior central gyrus and pyramidal tracts were projected into operation microscope.The lesions were precisely excised,and the posterior central gyrus and pyramidal tracts were protected effectively.The last iMRI indicated that the range of resection was coincided with the pro-operative plan.During the follow-up of 12-24 months,11 patients were found to be completely seizure-free or with only aura (Engel I),and 5 patients had only rare seizure (Engel Ⅱ,fewer than three seizures per year).Nine patients appeared visual field defect without hemiparalysis or hemianesthesia.Conclusion Functional neuronavigation combined with iMRI is a safe and reliable technique for perfect resection of symptomatic occipital lobe epilepsy and reduction of normal brain functions by precisely locating lesions and important functional structures.
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Objective To investigate the hemostatic effect and influence on coagulation function of hemecongulase during neurosurgical operation.Methods Sixty patients with neurosurgical trauma at American statistical association(ASA) Ⅰ-Ⅱ were randomly divided into hemocoagnlase treatment group (n =30) and control group(n =30).Both two group were injected Baquting 2U at the day before the operation,30 min before the operation,every two day after the operation and end up 3 d.Treatment group were pedormed with Baqyting 4 U + physiological saline 10 ml topical spraying.The intelligibility of operating region,the volume of intraoperative,the volume of bleeding during the operation,transfusion of blood,postoperative drainage,and drainage tube exelcymosis time were recorded in all the patients.Prothrombin time (PT),activated partial thromboplastin time (APTT),fibrinogen level(FIB),fibrinogen degradation product(FDP) and the two D-dimer and platelet count(PLT) before and after the surgery were also determined.All the patients were postoperatively followed up.Results The intelligibility of operating region was 70.0％ (21/30) in the hemocoagnlase treatment group,higher than that in control group (0％,P ＜0.05).The volume of bleeding during the operation in the hemecoagulase treatment group was (680.00 ± 95.22) ml,significantly fewer than that in the control group((790.00 ±47.00) ml,P =0.034).PLT significantly decreased after the surgery in both of the groups compare to that in preoperation (P ＜ 0.05 or P ＜ 0.01) and no significant difference was seen between two groups (P ＞ 0.05).No severe adverse events were found in both groups.Conclusion Hemocoagulase treatment during the operation can improve the intelligibility of operating region,reduce the volume of bleeding and transfusion of blood,and do not affect the coagulation function in the patients.Therefore,hemocoagulase is a safe and effective hemostatic and through local application during the operation it can improve curative effect.
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Objective To explore the method and management of pre-hospital care and raise the level of traffic injuries in pre-hospital care by summarizing the clinical features of death patients with severe tragic accident trauma Methods The basic data of 62 death patients with severe traffic accident trauma was analyzed according to death report statistics of severe traffic accident trauma from January 1st,2005 to December 31th,2008 Results Brain injury death accounted for most of traffic accident trauma death.The mortality rate of brain injury in the wounded wag 8.28%(13/157),but of asphyxia and hemorrhagic shock was 2.55%(4/157),3.18%(5/157)respectively in 2005.With the development of treatment and rescued in time, the mortality rate reduced to 6.11%(11/180),0,0.56%(1/180)in 2008.Conclesions It should be trying to shorten the time of pre-hospital care for pafients with trsffic accident trauma,especially in patients combined with hemorrhagic shock,asphyxia,severe brain injury.It is concluded that rapid and effective pre-hospital care can significantly reduce death rate and self-help or each other rescue training would also be effective to reduce mortality.
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Objective For decreasing the infected rate,the prevention and cure methods of intracranial infections following posterior fossa craniotomy were study. Methods Twenty-eight patients with the intracranial infections following posterior fossa craniotomy were examined by lumbar puncture,and analyzed cerebrospinal fluid with routine examination and reference to the bacteriological data and drug sensitive tests. All the patients were treated with high dosage sensitive antibiotics, and draining continually the infected cerebrospinal fluid by lumbar puncture catheterization and injected small dosages of antibiotics into intraspinal for most cases. Results Twenty-eight patients had intracranial hypertension by lumbar puncture examination, outcome of cerebrospinal fluid culture indicated that 17 cases had bacteria growth and 11 cases had no bacteria. The intracranial infection was controlled effectively,and 96.4%(27 cases) were cured, 1 case dead of systemic failure. Conclusions Strict aseptic techniques,reduce operative time,decrease intracranial place of foreign matters, such as gelfoam, hemostatic gauze and artificial implants, could reduce the possibilities of intracranial infections. Appropriate antibiotics selection,lumbar puncture catheterization and intraspinal administration of antibiotics can cure intracranial infections effectively.
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BACKGROUND: Vascular restenosis alter carotid endarteractomy (CEA) is an important factor affecting curative affect ofoperation.OBJECTIVE: To explore the role of matrix metalloproteinase-9 (MMP-9) mRNA dynamic expression in the development of early vascular restenosis after carotid endarterectomy.DESIGN, TIME AND SETTING: A random grouping contrast observation was completed in the General Hospital of Beijing Military Area Command of Chinese PLA from February 2006 to December 2007. MATERIALS: Forty-one healthy male New Zealand rabbit, weighing about 3.0 kg, with 36 ones used for preparing carotid atherosclerotic stenosis (CASS) models. experimental group, each 6 of the CASS rabbit models (n =36) were selected at the time points of hour 4, day 1, 3, 7, 30, and 90 following CEA respectively. Then they were fixed with 40 g/L polyoxymethylene perfusion and stained with hematoxylin-eosin to observe their morphologic changes.MAIN OUTCOME MEASURES: The expression changes of MMP-9 mRNA were observed dudng the development of early vascular restenosis by the quantitative real-time polymerase chain reaction technique preoperatively as well as at day 1, 3 and 7 following CEA.RESULTS: Several stages could be seen in the reparative process of neointima after CEA, including the thrombosis, the inflammatory reaction, the repair of endothelium, the proliferation of vascular smooth muscle call, the formation and accumulation of extracellular matrix. MMP-9 mRNA was expressed since day 1, reached a peak at day 3 and then decreased significantly at day 7 postoperatively.CONCLUSION: MMP-9 plays an important role in the proliferation, migration and reconstruction of vascular smooth muscle calls, the mediated reconstruction of local blood vessels, as well as the development of vascular restenosis.
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To investigate the feasibility of in vitro inducing differentiation of bone marrow stromal cells (BMSCs) into neural stem cells and mature neural cells, and to offer reference for the application of BMSCs in the field of neuroscience, BMSCs were acquired from the marrow of dogs. Basic fibroblast growth factor (bFGF), all trans retinoic acid (RA) and glial cell line derived neurotrophic factor (GDNF) etc were used as proliferation or differentiation factors. Immunocytochemistry was employed to identify the cells at every culture stage. When BMSCs were proliferatively cultured for 24～72h, cleavage phase and cluster like clones appeared. On the 3rd day, some of the BMSCs derived cells started to express neuron specific enolase (NSE) or glial fibrillary acidic protein (GFAP). The same stage cells could be cloned, that is one of the characters of stem cells, when they were cultured in the proliferation medium. The neural cell modality like cells appeared on the 10th day after adding inducing factors into culture medium, which were proved by testing the express of NSE and GFAP. BMSCs can differentiate into neural stem cells and mature neural cells in vitro, and can be used as "seed cells" in the field of neuroscience.