Brain-derived neurotrophic factors increase the proliferation and differentiation of endogenous neural stem cells in mouse models of cerebral infarction

BACKGROUND: It has been confirmed that brain-derived neurotrophic factor (BDNF) can promote the proliferation of neural stem cells (NSCs) and protect neuron-like cells in vitro. However, its effect on endogenous NSCs in vivo is still unclear. OBJECTIVE: To evaluate whether BDNF can induce the endogenous NSCs to proliferate and differentiate into the neurons in the mice model of cerebral infarction. DESIGN: A synchronal controlled observation. SETTINGS: Department of Neurology, Microbiology Division of the Department of Laboratory, Tianjin First Central Hospital; Howard Florey Institute, Medical College, the University of Melbourne. MATERIALS: Twenty-four pure breed C57BL/6J mice at the age of 10 weeks old (12 males and 12 females) were divided into saline control group and BDNF-treated group, 6 males and 6 females in each group. METHODS: The experiments were performed at the University of Melbourne from July 2004 to February 2005. ① The left middle cerebral artery (MCA) was ligated in both groups to establish models of cerebral infarction and the Matsushita measuring method was used to monitor the blood flow of the lesioned region supplied by MCA. 75% reduction of blood flow should be reached in the lesioned region. ② At 24 hours after infarction, mice in the BDNF-treated group were administrated with BDNF, which was slowly delivered using an ALZET osmium pump design. BDNF was dissolved in saline at the dosage of 500 mg/kg and injected into the pump, which could release the solution consistently in the following 28 days. The mice in the saline control group accepted the same volume of saline at 24 hours after infarction. ③ The Rotarod function test began at 1 week preoperatively, the time stayed on Rotarod was recorded. The mice were tested once a day till the end of the experiment. At 4 weeks post cerebral infarction, double labeling of Nestin and GFAP, BIII tubulin and CNPase immunostaining was performed to observe the differentiation directions of the re-expressed endogenous NSCs, and the percentages of the cells differentiated into astrocytes, neurons and oligodendrocytes were calculated. MAIN OUTCOME MEASURES: ① The differentiation directions of the re-expressed endogenous NSCs, and the percentage of the cells differentiated into astrocytes, neurons and oligodendrocytes. ② Comparison of motor function between the two groups. RESULTS: All the 24 pure C57BL/6J mice were involved in the analysis of results. ① Positively expressed endogenous NSCs appeared in the mice of both groups, and they mainly distributed around the focus of lesion, as well as the contralateral side. The expressed cells in the BDNF-treated group were obviously more than those in the saline control group. ②Activations of endogenous NSCs: At 4 weeks after infarction, re-expressions of endogenous NSCs appeared in both groups. The number of the re-expressed cells in the BDNF-treated group was about 4.2 times higher than that in the saline control group. The percentage of the cells differentiated into neurons in the BDNF-treated group was significantly higher than that in the saline control group (36%, 15%), the percentage of the cells differentiated into astrocytes was lower than that in the saline control group (54%, 77%), whereas the percentage of the cells differentiated into oligodendrocytes was similar to that in the saline control group (10%, 8%). ③ Results of motor functional test: Compared with before cerebral infarction, the mice in both groups manifested as obvious decrease in motor function at 1 week after infarction, whereas the recovery of motor function in the BDNF-treated group was significantly superior to that in the saline control group at 2, 3 and 4 weeks (P < 0.01). CONCLUSION: BDNF can promote the proliferation of endogenous NSCs in the brain of mice with cerebral infarction, it can decrease the differentiation rate of astrocytes, and increase the differentiation rate of neurons. BDNF has small influence on the differentiation of endogenous NSCs into oligodendrocytes, which was not benefit for the recovery of neural axon. Endogenous NSCs may improve the motor function of mice through the above pathways.     

脑源性神经生长因子基因转染对脊髓神经干细胞分化的影响

目的 研究BDNF基因转染小鼠脊髓源性NSCs向神经元分化情况.方法 选取体外培养E14小鼠胚胎脊髓来源NSCs,构建整合有BDNF基因的逆转录病毒载体,感染体外培养的NSCs,诱导其向神经元分化.采用免疫细胞化学方法鉴定,确定NSCs的分化比例.结果 转染后诱导分化24 h后可见部分细胞贴壁分化,48 h后转染细胞大部分贴壁.BDNF转染NSCs分化为神经元比例较未转染NSCs明显增高,差异有统计学意义(P＜0.05).结论 逆转录病毒载体介导BDNF基因转染NSCs可促进细胞分化,且分化多为神经元方向.

Abstract：

Objective To study the differentiation potential of mouse spinal cord derived neural stem cells (NSCs) into neurons after being transfected with BDNF gene in vitro. Methods Spinal cord derived NSCs from the E14 fetus mouse were isolated and cultured in vitro; the retrovirus containing pLXSN-BDNF gene was established and transfected into thc above NSCs, and thea, spinal cord derived NSCs were induced to be differentiated into neuron-like cells. Immunohistochemistry was employed to detect and calculate the ratio of differentiation of NSCs into neurons. Results The NSCs cultured in vitro partly adhered to the wall and differentiated within 24 h of transfection with BDNF gene, and most of the NSCs adhered to the wall differentiated within 48 h of transfection. The level of neurons from spinal cord derived NSCs modified by BDNF gene was markedly increased as compared with that that from normal spinal cord derived NSCs (P＜0.05). Conclusion NSCs transfected by retroviral pLXSN-BDNF can promote the cell differentiation. BDNF gene can increase greatly the percentage of neurons in the course of inducing the differentiation of mouse NSCs.     

Neuroprotective effect of estrogen after chronic spinal cord injury in ovariectomized rats

BACKGROUND: At present, there is still lack of effective drugs for chronic spinal cord injury, whereas it is found recently that estrogen has a neuroprotective effect on brain and spinal cord injuries. OBJECTIVE: To observe the effect of estrogen on the apoptosis of nerve cells after gradual chronic spinal cord injury in ovariectomized rats. DESIGN: A randomized controlled animal trial. SETTING: Institute of Orthopaedics, the Second Hospital of Lanzhou University. MATERIALS: Sixty-five female Wistar rats of common degree, weighing 220–250 g, were provided by the experimental animal center of Lanzhou University. The rats were randomly divided into sham-operated group (n =5), estrogen-treated group (n =30) and saline control group (n =30), and the latter two groups were observed at 1, 3, 7, 14, 28 and 60 days respectively, and 5 rats for each time point. METHODS: All the rats were treated with bilateral oophorectomy 2 weeks before the experiment. T10 vertebral lamina was revolved into using plastic screw. The spinal canal impingement was not induced initially. After that, the original incision was opened to expose the screw every 7–10 days. MAIN OUTCOME MEASURES: The apoptosis and Caspase-3 positive cells in the damaged spinal cord were detected using terminal deoxynucleotidal transferase-mediated dUTP-biotin nick end labeling (TUNEL) method and Caspase-3 immunohistochemical staining at 1, 3, 7, 14, 28 and 60 days after chronic spinal cord injury respectively. RESULTS: Totally 65 rats were used, and the deleted ones during the experiment were supplemented by others. Changes of Caspase-3 expression after spinal cord injury: In the sham-operated group, only a small amount of Caspase-3 proteins were observed in the rat spinal cord, mainly located in motor neurons of spinal cord anterior horn. In the estrogen-treated group and saline control group, positive cells expressed occasionally at 1 day postoperatively, began to increase obviously at 7 days after injury, strongly expressed at 14 and 28 days, but decreased at 60 days, mainly located in the neurons of spinal cord gray matter anterior horn, and they expressed fewer in the motor neurons and white matter of ventral horn, and there were obvious differences between the estrogen-treated group and saline control group at 7, 14, 28 and 60 days (P < 0.05). CONCLUSION: Estrogen can reduce the apoptosis of nerve cells and promote the recovery of neurological function following gradual chronic spinal cord injury.     

Changes in brain-derived neurotrophic factor expression after transplanting microencapsulated sciatic nerve cells of rabbits into injured spinal cord of rats

BACKGROUND: Changes of brain-derived neurotrophic factor (BDNF) expression reflect function of nerve cells; meanwhile, they play a significant role in researching interventions on plerosis of nerve injury. OBJECTIVE: To observe and compare the effects on changes of BDNF expression in rats with spinal cord injury between microencapsulated sciatic nerve cells of rabbits and only transplanting sciatic nerve cells of rabbits. DESIGN: Randomized controlled animal study. SETTING: Medical School of Jiujiang College. MATERIALS: The experiment was carried out in the Medical Science Researching Center, Jiujiang College from May 2004 to May 2006. A total of 90 healthy adult SD rats, weighing 250–300 g, of either gender; and 10 rabbits, weighing 2.0–2.5 kg, of either gender, were provided by Jiangxi Experimental Animal Center. METHODS: Sciatic nerve tissue of rabbits was separated to make cell suspension. After centrifugation, suspension was mixed with 15 g/L alginate saline solution and ejaculated to 20 mmol/L barium chloride saline solution by double-cavity ejaculator. The obtained cell microcapsules were suspended in saline. Rats were randomly divided into microencapsulated group, only suspension group, and only injured group with 30 animals in each group. After anesthesia, T10 spinous process and vertebra lamina of rats in the former two groups were exposed. Spinal cord tissue in 2-mm length was removed from rats by spinal cord right hemi-section. The gelatin sponges with the size of 2 mm × 2 mm × 2 mm were grafted as filing cage, and absorbed 10 μL microencapsulated sciatic nerve cells of rabbit in the microencapsulated group and 10 μL sciatic nerve cells of rabbits in the only suspension group; respectively. No graft was placed in the only injured group. MAIN OUTCOME MEASURES: On the 1st, 3rd, 7th, 14th and 28th days after operation, immunohistochemistry (SABC technique) was used to detect distribution and amount of positive-reactive neurons in BDNF of spinal cord samples which were selected as 2 cm away from the injured surface. RESULTS: All the 90 rats were involved in the final analysis. Masses of brown-yellow particles were found in the microencapsulated group, and most of them were distributed in the spinal cord anterior horn neurons and glial cells. The positive-reactive neuron particles were also found in the white matter and gray matter. On the 3rd, 7th, 14th and 28th days after operation, amount of positive-reactive neurons in BDNF in the microencapsulated group was higher than that in the only injured group (P < 0.01) and only suspension group (P < 0.05). CONCLUSION: After transplanting microencapsulated nerve cell suspension into injured spinal cord of rats, distribution and amount of positive-reactive neurons in BDNF of local samples at injured surface are increased remarkably as compared with those by using tissue cell transplantation.     

Chlorogenic acid protection of neuronal nitric oxide synthase-positive neurons in the hippocampus of mice with impaired learning and memory☆

BACKGROUND： Clinical practice and modern pharmacology have confirmed that chlorogenic acid can ameliorate learning and memory impairments. OBJECTIVE： To observe the effects of chlorogenic acid on neuronal nitric oxide synthase （nNOS）-positive neurons in the mouse hippocampus, and to investigate the mechanisms underlying the beneficial effects of chlorogenic acid on learning and memory. DESIGN, TIME AND SETTING： The present randomized, controlled, neural cell morphological observation was performed at the Institute of Neurobiology, Central South University between January and May 2005. MATERIALS： Forty-eight female, healthy, adult, Kunming mice were included in this study. Learning and memory impairment was induced with an injection of 0.5 uL kainic acid （0.4 mg/mL） into the hippocampus. METHODS： The mice were randomized into three groups （n = 16）： model, control, and chlorogenic acid-treated. At 2 days following learning and memory impairment induction, intragastric administration of physiological saline or chlorogenic acid was performed in the model and chlorogenic acid-treated groups, respectively. The control mice were administered 0.5uL physiological saline into the hippocampus, and 2 days later, they received an intragastfic administration of physiological saline. Each mouse received two intragastric administrations （1 mL solution once） per day, for a total of 35 days. MAIN OUTCOME MEASURES： Detection of changes in hippocampal and cerebral cortical nNOS neurons by immunohistochemistry; determination of spatial learning and memory utilizing the Y-maze device. RESULTS： At day 7 and 35 after intervention, there was no significant difference in the number of nNOS-positive neurons in the cerebral cortex between the model, chlorogenic acid, and control groups （P 〉 0.05）. Compared with the control group, the number of nNOS-positive neurons in the hippocampal CA1-4 region was significantly less in the model group （P 〈 0.05）. However, the control group was not different fro     

高压氧对缺氧缺血脑源性神经干细胞分化的影响

BACKGROUND： It has been previously shown that hyperbaric oxygen may promote proliferation of neural stem cells and reduce death of endogenous neural stem cells （NSCs）.
OBJECTIVE： To explore the effects of hyperbaric oxygen on the differentiation of hypoxic/ischemic brain-derived NSCs into neuron-like cells and compare with high-concentration oxygen and high pressure.
DESIGN, TIME AND SETTING： An in vitro contrast study, performed at Laboratory of Neurology, Central South University between January and May 2006.
MATERIALS： A hyperbaric oxygen chamber （YLC 0.5/1A） was provided by Wuhan Shipping Design Research Institute; mouse anti-rat microtubule-associated protein 2 monoclonal antibody by Jingmei Company, Beijing; mouse anti-rat glial fibrillary acidic protein monoclonal antibody by Neo Markers, USA; mouse anti-rat galactocerebroside monoclonal antibody by Santa Cruz Biotechnology Inc., USA; and goat anti-mouse fluorescein isothiocyanate-labeled secondary antibody by Wuhan Boster Bioengineering Co., Ltd., China.
METHODS： Brain-derived NSCs isolated from brain tissues of neonatal Sprague Dawley rats were cloned and passaged, and assigned into five groups： normal control, model, high-concentration oxygen, high pressure, and hyperbaric oxygen groups. Cells in the four groups, excluding the normal control group, were incubated in serum-containing DMEM/F12 culture medium. Hypoxic/ischemic models of NSCs were established in an incubator comprising 93% N2, 5% 002, and 2% 02. Thereafter, cells were continuously cultured as follows： compressed air （0.2 MPa, 1 hour, once a day） in the high pressure group, compressed air ＋ a minimum of 80% 02 in the hyperbaric oxygen group, and a minimum of 80% Q2 in the high-concentration oxygen group. Cells in the normal control and model groups were cultured as normal.
MAIN OUTCOME MEASURES： At day 7 after culture, glial fibrillary acidic protein, microtubule-associated protein 2, and galactocerebroside immunofluorescence staining were examined to observe differentiation and calculate the percentage of NSCs differentiating into neuron-like cells or neuroglia-like cells.
RESULTS： Neuron-like cells or neuroglia-like cells were visualized in all five groups. There were no significant differences in the percentage of differentiating cells between the hyperbaric oxygen group and the normal control group （P 〉 0.05）. The percentage of NSCs differentiating into neuron-like cells in the hyperbaric oxygen group was significantly greater than model, high-concentration oxygen, and high pressure groups; however, the percentage differentiating into neuroglia-like cells was significantly lower （P 〈 0.01 ）.
CONCLUSION： Hyperbaric oxygen promotes the differentiation of brain-derived neural stem cells into neuron-like cells but inhibits differentiation into neuroglia-like cells. Furthermore, the efficacy of hyperbaric oxygen is superior to high-concentration oxygen and high pressure.     

癫痫细胞悬液对海马干细胞分化的影响

Objective To investigate the effect of suspended cells from the hippocampus of epileptic rats on the differentiation of hippocampal stem cells.Methods Eight groups, including control group,glutamic acid groups (5 and 25 μmol/mL groups),normal hippocampal cells and epileptic cells groups (20 and 40 μmol/mL groups,respectively) were chosen in the experiment.The suspended cells were extracted from the hippocampus of epileptic rats and normal rats,and corresponding dose of them were added into the epileptic cells groups and normal hippocampal cells groups,respectively.Five μmol/mL and 25 μmol/mL glutamic acid were added into the glutamic acid groups,respectively.The vitality of differentiated stem cells was detected and compared with MTT method.Results MTT value (the vitality of differentiated stem cells) was increased following the increased differentiation time.Cell viability in the glutamic acid groups and normal hippocampal cells groups was slightly decreased as compared with that in the control group (P>0.05).The cell viability in the epileptic cells groups was slightly increased as compared with that in the control group (P>0.05).The MTT value in the epileptic cells group and the normal hippocampal cells group was not obviously different (P>0.05).Conclusion The suspended cells from the hippocampus of epileptic rats do not affect the differentiation of the hippocampal stem cells.     

基质细胞衍生因子-1诱导神经干细胞靶向性迁移的实验研究

Objective To investigate the effect ofstromal cell derived factor-1 (SDF-1) on the regulation of neural stem cells (NSCs) migration.Methods NSCs were obtained from the cerebral cortex of embryonic rats and cultured in serum-free medium,and their stem cell properties were assessed by means of induced differentiation in vitro into neurons and astrocytes.After in vitro cell culture,the purity of NSCs and the co-expression rate of CXCR4/nestin were detected by flow cytometry.Blind-well chambers were employed to detect the chemotactic effects of SDF-1 by counting the cells which had crossed a 8 μm pore membrane when confronted with varying concentrations of SDF-1 (0,1,10,50,100,500 and 1000 ng/mL),and the distribution of cells migrated out of the same neurosphere was overviewed by μ-slides in the persistent concentration gradient of SDF-1.Results Neurospheres were formed by persistent proliferation of NSCs, which were capable of differentiating into neurons (β-tubulin+) and astrocytes (GFAP+) in media without mitogens,and flow cytometry analyses showed that most of the cultured cells expressed nestin and the co-expression rate of CXCR4/nestin was nearly 80%.SDF-1 showed great chemotaxis to NSCs,and the amount of cells having migrated through the membrane in 500 ng/ml SDF-1 group was higher than that in other groups (P<0.05).When the cells were confronted with a linear concentration gradient (from 500 to 0 ng/mL),which was generated by diffusion and stable for at least 48 h,the cells migrated out ofa neruosphere could distribute irregularly with more cells locating in the region of higher concentration of SDF-1 and longer migration distance away from the center of the neurosphere than the opposite.Conclusion SDF-1 binding to its specific receptor CXCR4 was capable of inducing NSCs migrating directionally to the source of SDF-1.     

促红细胞生成素与大鼠面神经损伤后运动神经元TNF-a及Bax的表达

This study sought to evaluate the effect of high-dose erythropoietin (EPO; 5 000 IU/kg) on the expression of tumor necrosis factor-alpha (TNF-α) and Bax in the facial nucleus after facial nerve transection in rats. A total of 42 Wistar rats of both genders were used in this study, and 40 rats were randomly divided into 2 groups: EPO group and model group. The EPO group was treated with EPO once a day for 5 days at a dose of 5 000 IU/kg body weight. The model group was treated with saline of the same amount. At day 3 after EPO (or saline) treatment, the right facial nerves of the 40 rats were transected at the level of the stylomastoid foramen, with the left sides untreated. The remaining 2 rats that did not undergo axotomy served as the control group. The surviving motor neurons in operated rats were counted in coronal paraffin sections of the facial nucleus. The expression of TNF-α and Bax in the facial nucleus was detected by immunohistochemical staining at days 3, 7, 14, 21, and 28 after axotomy. At days 14, 21, and 28 after facial nerve axotomy, a significantly greater proportion of facial motor neurons survived in the EPO group than in the model group. After axotomy, the expression of TNF-α and Bax increased in motor neurons in both the EPO and the model groups. TNF-α expression reached its peak level at day 14 after axotomy, while Bax expression reached its peak level at day 21. TNF-α expression was much lower in the EPO group than in the model group at all time points. No significant difference in Bax expression was found between the EPO and the model groups. These results indicate that high-dose EPO treatment attenuates the increase in TNF-α expression in the facial nucleus and reduces the loss of motor neurons after facial nerve transection in rats. However, high-dose EPO treatment has little effect on Bax expression.     

Neuroprotective effect of melatonin against ischemia/reperfusion-induced neuronal apoptosis in mouse cerebellum

BACKGROUND: Some experiments have demonstrated that melatonin (N-aceyl-5-methoxytryptamine, Mel) has antioxidation. However, whether it has neuroprotective effect in the ischemia/reperfusion injury of central nervous system is unclear. OBJECTIVE: To observe the protective effect of Mel on ischemia/reperfusion-induced cerebellar neuronal apoptosis of rats, and the action mechanism. DESIGN: Controlled observation experiment. SETTING: Department of Biochemistry and Molecular Biology, Tongji Medical College, Huazhong University of Science and Technology. MATERIALS: Eight Sprague-Dawley rats aged 7–8 days and weighing 10–12 g were provided by Medical Experimental Animal Center, Tongji Medical College,Huazhong University of Science and Technology. Anti-cytochrome C monoclonal antibody was purchased from R & D Company; 7-dichlorodihydrofluorescein diacetate(DCFH-DA), rhodamine 123 and Mel were purchased from Sigma Company (USA). Lactate dehydrogenase (LDH) kit was purchased from Nanjing Jiancheng Bioengineering Institute. METHODS: This experiment was carried out in the laboratory for Department of Biochemistry and Molecule Biology, Tongji Medical College between October 2002 and March 2004. Cerebellar neurons of rats were cultured in vitro. After oxygen-glucose deprivation (OGD) for 90 minutes, 1×10–4,1×10–6, 1×10–9 mol/L Mel was added, respectively, namely high-, middle-, and low-concentration Mel groups. Cells, which were cultured by OGD, served as model group, and control group, in which OGD intervention was omitted, was set. ①Cytochrome C level of mitochondrial cells in each group was detected by ELISA method. ②LDH activity in the cell culture fluid was measured, and cell membrane permeability change was analyzed. The cells in the Mel group with the lowest LDH activity served as Mel treatment group, i.e. cells were cultured with OGD, and then Mel was added; Meanwhile, Mel prevention group was set, i.e. Mel was added before OGD. Intervention was not changed in the model group and control group. ③ DNA level was analyzed and cell apoptosis was observed by agarose gel electrophoresis(AGE). ④Mitochondrial transmembrane potential of cells, and apoptotic way in each group were analyzed by confocal laser scanning microscopy. MAIN OUTCOME MEASURES: ①Mitochondrial cytochrome C level of cerebellar nerve cells. ②LDH activity of cerebellar nerve cells. ③ DNA AGE results. ④Mitochondrial transmembrane potential change. RESULTS: ①Mitochondrial cytochrome C level of cerebellar nerve cells: cytochrome C was obviously released at 6 hours of OGD-reperfusion. Mel inhibited the release of cytochrome C in dose-dependent manner. ②LDH activity of cerebellar nerve cells: LDH activity (A value) was significantly lower in the high- and middle-concentration Mel groups than in the model group (P < 0.05). LDH activity (A value) in the low-concentration Mel group was 0.415 0±0.012 9, indicating that Mel could decrease LDH activity of OGD-treated cell supernatant and promote membrane stablization in dose-dependent manner. ③AGE results of DNA: 1×10–9 mol/L was considered as the best concentration of melatonin. Cell DNA was extracted for AGE. Results presented typical ladder shape, indicating apoptosis appeared, while apoptosis was lessened in the Mel treatment group and Mel prevention group.④Mitochondrial transmembrane potential change: Experimental results showed that green fluorescein was evenly distributed in cerebellar granule cells cultured normally, and the axons of neurons were very clear. The body of neurons was condensed and the axons disappeared after cerebellar granule cells undergoing OGD injury. Mel could completely reverse the effect of OGD. CONCLUSION: Mel can enhance cerebellar neuronal membrane stabilization of rats in dose-dependent manner, and suppress OGD-induced apoptosis of cerebellar granule cells by preventing against mitochondrial apoptosis.     

Effects of tanshinone Ⅱ sodium sulfonate plus cinepazide maleate on the hemorrheologic indexes and blood lipids in patients with acute cerebral infarction

BACKGROUND: The severity of cerebral infarction is associated with the increase of blood viscosity caused by hyperfibrinogenemia and hyperlipidemia, etc. Thus it has become one of the target for treating cerebral infarction to decrease blood viscosity by integrated Chinese and western medicine. OBJECTIVE: To investigate the influence and clinical therapeutic effects of cinepazide maleate combined with tanshinone Ⅱ A sodium sulfonate on the hemorrheologic indexes and blood lipids of patients with acute cerebral infarction, and compare the results with those of simple cinepazide maleate treatment. DESIGN: A non-randomized case-controlled observation. SETTINGS: Hebei North University; the Second Affiliated Hospitals of Hebei North University; the Third Affiliated Hospitals of Hebei North University. PARTICIPANTS: Eighty-six inpatients with cerebral infarction were selected from the infirmary, the Second and Third Affiliated Hospitals of Hebei North University from September 2004 to October 2006. They were all diagnosed to have acute cerebral infarction by CT or MRI, and accorded with the diagnostic standards for acute cerebral infarction set by the Fourth National Academic Meeting for Cerebrovascular Disease in 1995. Meanwhile, 40 teachers and medical staff of voluntary physical examinees were selected as the control group. Informed contents were obtained from all the patients and their relatives. METHODS: The patients were divided into combined treatment group (n=43) and simple treatment group (n=43). In the combined treatment group, the patients were administrated with 160 mg cinepazide maleate injection (Beijing Four-ring Pharmaceutical, Co.,Ltd, No. H200220125; 80 mg/2 mL) added in 5% glucose, and 40 mg tanshinone Ⅱ sodium sulfonate (Shanghai No.1 Biochemical & Pharmaceutical Co.,Ltd., No. H31022558, 10 mg/2 mL) added in 250 mL normal saline. In the simple treatment group, the patients were only administrated with cinepazide maleate 320 mg added in 5% glucose or 250 mL normal saline. They were treated for 1 or 2 courses, once a day, and 14 days as a course. The patients were detected before treatment and at 14 and 28 days after treatment respectively. ① Determination of hemorrheologic indexes: Whole blood viscosity was determined with LBY-N6B automatic hemorrheologic meter; Plasma viscosity with LBY-F200B automatic plasma viscosity meter; Volume of fibrinogen was determined by the method of 12.5% sodium nitrate depositing biuret reaction. ② Determination of blood lipids: The serum levels of total cholesterol (TC), triglyceride (TG), low density lipoprotein cholesterol (LDL-C) and high density lipoprotein cholesterol (HDL-C) were determined. ③ Severity of neurological deficit: The total score of neurological deficit score (NDS) ranged from 0 to 45 points, 0–15 points was taken as mild, 16–30 points as moderate and 31–45 points as severe. ④ Evaluation of curative effects: Generally cured: NDS decreased by 91%–100%, and disabled severity of grade 0; Significantly improved: NDS decreased by 46%–90%, and disabled severity of grades 1–3; Improved: NDS decreased by 18%–45%; No change: NDS decreased by less than 18%; Aggravated: NDS increased by more than 18%. Generally cured and significant improved were taken as significant effect. ⑤ The adverse events and side effects after medication were observed. MAIN OUTCOME MEASURES: ① Results of hemorrheologic indexes and blood lipids; ② NDS results in the combined treatment group and simple treatment group; ③ Therapeutic effects and adverse events. RESULTS: All the 86 patients with cerebral infarction and 40 healthy controls were involved in the analysis of results. ① Results of hemorrheologic indexes and blood lipids: The hemorrheologic indexes and blood lipids before treatment were manifested as abnormalities to different extents in both the combined treatment group and simple treatment group; The hemorrheologic indexes after treatment were obviously improved in both groups. But the hemorrheologic indexes were improved more obviously in the combined treatment group as compared with those in the simple treatment group (P < 0.05); The levels of TC, TG and LDL-C after treatment in the combined treatment group were obviously lowered (P < 0.05), whereas those in the simple treatment group were not significantly changed (P > 0.05). ② NDS results: The NDS scores at 14 and 28 days after treatment in the combined treatment group [(6.23±2.34), (4.27±1.83) points] were obviously lower than those in the simple treatment group [(8.76±3.41), (6.65±2.49) points, P < 0.05]. ③ Therapeutic effects and side effects: The total significant effective rates in the combined treatment group and simple treatment group were 93% and 81% respectively. In the combined treatment group, 1 case suffered from palpitation, dizziness and agrypnia. In the simple treatment group, 1 case suffered from palpitation, dizziness and agrypnia, 1 case had itch of skin. All the above symptoms disappeared gradually after the transfusing speed was adjusted to be slower. No drug withdrawal occurred in the patients due to the adverse events. CONCLUSION: Cinepazide maleate combined with tanshinon can obviously improve the abnormalities of hemorrheologic indexes and blood lipids and nerve function in patients with acute cerebral infarction, and its curative effect is faster than that of simple cinepazide maleate treatment.     

Effect of intracranial hypertension on cerebral hemorrhage induced autonomic nerve imbalance

BACKGROUND: Cerebral hemorrhage can cause the imbalance of nerve function, whereas its mechanism and main impact factors are still not quite clear. OBJECTIVE: To explore the rules about the changes of intracranial pressure in brainstem hemorrhage and internal capsule hemorrhage, and analyze the role of intracranial hypertension in the changes of nerve function caused by cerebral hemorrhage. DESIGN: A self-controlled trial. SETTING: Department of Physiology, Tianjin Medical University. MATERIALS: Sixty-five healthy male Japanese white rabbits with long ears (1.5–1.8 kg) were supplied and fed by the Department of Animal Experiment of Tianjin Medical University. The RM6240B biological signal collecting and processing system was used. METHODS: The experiments were conducted in the Department of Physiology, Tianjin Medical University from August 2001 to May 2006. ① The rabbits were anesthetized, then fixed onto the brain stereotaxic apparatus, and afterwards fenestration on skull and intubation to lateral ventricle were performed.The dynamic changes of intracranial pressure were monitored continuously. Rabbits were infused with autologous arterial blood (0.3 mL) into midbrain corpora quadrigemina inferior colliculus to induce model of acute brainstem hemorrhage; models of internal capsule hemorrhage were established by infusing autologous arterial blood into internal capsule. ② The dynamic intracranial pressures under the above conditions were recorded continuously with the RM6240B biological signal collecting and processing system. ③ An animal model of persistent intracranial hypertension was established by infusion of physiologic saline into lateral ventricle. ④ The changes of the intensity of autonomic nerve discharge were analyzed, using the biological signal collecting and processing system before and after hemorrhage and under persistent intracranial hypertension. ⑤ Ten animal models of internal capsule hemorrhage and 10 of brainstem hemorrhage were selected respectively, then gross pathological samples were cut open, and the accuracy of hemorrhage models was affirmed. Histological sections in hemorrhage point and around this point were prepared for with hematoxylin and eosin staining, and the pathological changes were observed under light microscope. MAIN OUTCOME MEASURES: ① Changes of intracranial pressures before and after internal capsule hemorrhage and brainstem hemorrhage; ② Changes of the discharge intensity of cervical vagus nerve trunk in animal models of internal capsule hemorrhage, brainstem hemorrhage and persistent intracranial hypertension without hemorrhage; ③ Accuracy of location of internal capsule hemorrhage and brainstem hemorrhage confirmed by gross pathological samples and sections. RESULTS: Totally 65 rabbits were involved in the analysis of results. ① Dynamic state of intracranial pressure: Intracranial pressure increased obviously at 45 minutes after internal capsule hemorrhage and brainstem hemorrhage, the intracranial pressures were (1.31±0.30), (1.82±0.45) kPa, which were obviously higher than those before hemorrhage [(1.04±0.18), ( 1.05±0.19) kPa, P < 0.01]. ② Discharge of vagus nerve: Under intracranial hypertension, the discharge of cervical vagus nerve trunk was enhanced, and the discharge intensity of vagus nerve trunk was significantly different before and after persistent intracranial hypertension [(364.28±78.55), (1252.19±151.75) μV·s, P < 0.01]. The discharges of cervical vagus nerve trunk were significantly enhanced after internal capsule hemorrhage and brainstem hemorrhage (P < 0.01). ③ Validation of hemorrhage sites: The hemorrhage sites were internal capsule and brainstem on histopathological sections. CONCLUSION: Intracranial pressure may play an important role in the pathophysiological process of vagus nerve imbalance caused by cerebral hemorrhage.     

Hyperbaric oxygen therapy for cerebral blood flow and electroencephalogram in patients with acute cerebral infarction ——Choice for therapeutic occasion

BackgroundHyperbaric oxygen (HBO) therapy increases blood oxygen content, changes cerebral blood flow (CBF) and cerebral metabolism. Its therapeutic effects on cerebrovascular disease have been fully confirmed, but the occasion for HBO therapy is still unclear.ObjectiveTo observe the therapeutic effects of HBO therapy at different time on CBF and electroencephalogram (EEG) in patients with acute cerebral infarction (CI).DesignRandomized controlled trial.SettingDepartment of Neurology, Shidong Hospital, Yangpu District of Shanghai.ParticipantsNinety-six inpatients with acute CI, admitted to Department of Neurology, Shidong Hospital, Yangpu District of Shanghai from January 2001 to December 2006, were involved in this experiment. The involved participants met the diagnosis criteria of acute CI and confirmed by skull CT or MRI. They all were patients with moderate CI (16–30 points) according to neurologic deficit score formulated by Chinese Medical Association. Informed consents of detected items and therapeutic regimen were obtained from all the involved participants. They were randomized into two groups with 48 in each: early-stage treatment group and advanced-stage treatment group. Among the 48 patients in the early-stage treatment group, 21 male and 27 female, aged 53–68 years, 22 patients were found with basal ganglia infarction, 10 with brain lobe infarction, 16 with multiple infarction, 27 accompanied with hypertension and 2 accompanied with diabetes mellitus. Among the 48 patients in the advanced-stage treatment group, 23 male and 25 female, aged 52–71 years, 25 patients were found with basal ganglia infarction, 10 with brain lobe infarction, 12 with multiple infarction, 1 with brain stem infarction, 28 accompanied with hypertension and 1 accompanied with diabetes mellitus.MethodsAfter admission, patients of two groups received routine drug treatment.

Patients in the early-stage treatment group and advanced-stage treatment group began to receive HBO therapy within one week of CI and 4 weeks after CI, respectively. The total course of treatment both was 2 weeks. EEG examination was carried out before and after therapy, and CBF was determined with ¹³³Xe inhalation.

Assessment criteria of curative effects: Basically cured: neurologic symptoms and body signs disappeared, could work and do housework; Markedly effective: score of neurologic deficit was decreased by over 21 points, could manage himself/herself partially; Effective: score of neurologic deficit was decreased by 8 to 12 points; Non-effective: Score was increased or decreased less than 8 points, neurologic deficit was worsened, even died. Total effective rate = (number of cured+number of markedly effective+number of effective)/number of total cases×100%.

t test and Chi-square test were used for comparing the difference of measurement data and enumeration data respectively, and Ridit analysis was used for comparing the difference of clinical curative effects.Main outcome measures

Comparison of EEG and CBF of patients from two groups before and after treatment.

Comparison of post-treatment neurologic deficit of patients between two groups.ResultsAll the involved 96 patients with CI participated in the final analysis.

Clinical symptoms of patients from two groups after therapy were significantly improved as compared with those before therapy, and curative effects of early treatment group were better than those of advanced treatment group (U =1.99,P < 0.05).

After treatment, CBF in each region of brains, except for that in parietal lobe of patients in the advanced-stage treatment group, was significantly improved (P < 0.05–0.01); The improvement of CBF of patients in the early-stage treatment group was more obvious than that in the early-stage treatment group (P < 0.05–0.01).

The abnormal rate of EEF of patients from early-stage treatment group and advanced-stage treatment group before treatment was 94% and 96%, respectively. After treatment, improvement rate of EEG of patients in the early-stage treatment group was 95%, which was significantly different from that in the advanced-stage treatment group (82%,χ² =4.32,P < 0.05)ConclusionHBO therapy both at early and advanced stages of CI (within 1 week and 4 weeks after CI attack) can improve CBF and EEG of patients with early CI, especially.     

Correspondence of CT perfusion imaging to pathological manifestations in rabbit models of hyperacute cerebral infarction

BackgroundCould the infarction be diagnosed quickly and accurately at the acute stage by CT perfusion imaging (CTPI) technology? Whether the images of CTPI will correspond with the pathological changes or not? All the questions need to be solved by experimental and clinical studies.ObjectiveTo reveal the rules of perfusion map changes and guide the early diagnosis of hyperacute cerebral infarction by analyzing the correlation of CTPI with pathological manifestations for hyperacute cerebral infarction.DesignA randomized controlled animal experiment.SettingExperimental Center of Medical Radiology, Longgang Central Hospital of Shenzhen City.MaterialsForty-two adult New Zealand rabbits of (2.6±0.5) kg, either male or female, were randomly divided into experimental group (n =36) and control group (n =6). Six rabbits in the experimental group were observed after ischemia for 0.5, 1, 2, 3, 4 and 6 hours respectively, and 1 rabbit in the control group was observed at each corresponding time point.MethodsThe experiments were carried out in the Experimental Center of Medical Radiology, Longgang Central Hospital of Shenzhen City from March 2003 to July 2004. Rabbit models of cerebral infarction were established by modified O'Brein method.

The rabbits in the experimental group were scanned at 0.5, 1, 2, 3, 4 and 6 hours after ischemia respectively. The dynamic CT scan slice was 13 mm from the anterior edge of the frontal cortex, and six fake color functional images were obtained, including cerebral blood flow map (CBF map), cerebral blood volume map (CBV map), peak to enhancement map (PE map), flow without vessels map, time to peak map (TP map), time to start map (TS map). The manifestations and changes of the functional maps in different interval were observed.

Bilateral symmetric ranges of interest (ROI) were drawn separately on the CBF map, CBV map, TP map and TS map. The blood flow parameters of focal and contralateral cerebral tissues could be obtained to calculate relative cerebral blood flow (rCBF, rCBF=focal CBF/contralateral CBF), relative cerebral blood volume (rCBV, rCBV= focal CBV/contralateral CBV), a relative time to peak (rTP, rTP= focal TP–contralateral TP), a relative time to start (rTS, rTS= focal TP–contralateral TP).

The perfusion maps were input into AutoCAD software. The percents of ischemic cores and peri-ischemic areas accounting for contralateral cerebral hemisphere were calculated.

The animals were anesthetized and killed, then the cerebellum and low brain stem were taken out. The brain tissues were cut on coronal plane at 14 mm from the anterior edge of the frontal cortex, a 2-mm piece anterior to the incision, and a 3-mm piece posterior to the incision. The anterior piece was fixed, stained and observed. A 1-mm slice was cut from the front of the posterior piece tissues as electron microscope sample, the remnant was fixed and then taken out, and the location and size of stained “white” areas were observed as the reference for electron microscope sample.

The correlation between CTPI and pathological manifestations was observed.Main outcome measures

Laws of time and spatial changes of ischemic areas;

Pathological changes of the ischemic tissues;

Correspondency between CTPI and pathological manifestations.Results

Laws of time and spatial changes of ischemic areas: Relative ischemic-core areas were consistent in each perfusion map, increased incessantly along with the ischemic times. Relative peri-ischemic areas were inconsistent in each perfusion map, on CBF map from 1 to 6 hours after ischemia, the area of ischemic core increased from (1.503±0.523)% to (7.125±1.054)%, the ascending trend occurred. But the peri-ischemic areas showed a descending trend on CBF map, the areas decreased from (8.960±0.719)% to (5.445±0.884)% from 0.5 to 6 hours; The relative areas were the largest one on TP maps, the average value was (32.796±3.029)% at 0.5 hour after ischemia happening (60.540±1.683)% at 6 hours. The trend of ischemic areas was increased. No obvious change was observed on TS maps.

Pathological changes of the ischemic tissues: Under light microscope, there was no obvious change at 0.5–2 hours after ischemia, edema at 3 hours, karyopycnosis at 4 hours and eosinophilous changes at 6 hours; Under electron microscope, there was edema in ischemic cores within 4 hours after ischemia, whereas karyopycnosis or structure vanished after 4 hours; Edema was observed in peri-ischemic areas.

Correlation between CTPI and pathological manifestations: On CTPI maps, the ischemic core was blue on CBF and CBV maps, black on TP and TS maps. Along with the ischemic times, the rCBF and rCBV decreased, whereas the rTP and rTS prolonged. Hemodynamic parameters were not significantly different within 2 hours of ischemia and 2 hours after ischemia. The rTP and rTS became 0 after 1 and 2 hours respectively. On CTPI maps the peri-ischemic area was red on CBF and CBV maps, red and yellow on TS maps, red on TP maps. Along with the ischemic times, the rCBF decreased, and the lowest level was always at about 20%, whereas the rTP and rTS prolonged.Conclusion

CTPI manifestations corresponded well with pathological findings, and it is a sensitive, stable and reliable technique to diagnose hyperacute cerebral infarction.

TP map was more sensitive than CBF map and TS map in exhibiting the peri-ischemic areas, thus TP maps could be a good choice for observing peri-ischemic areas.     

Hyperglycemia induces protein nonenzymeatic glycosylation in brain neurons of diabetic rats at early stage

BACKGROUND: Protein nonenzymatic glycosylation is supposed to be one of mechanisms for chronic complications development in diabetes mellitus, and therefore, might play an important role in the neuronal degeneration. OBJECTIVE: To study the protein nonenzymatic glycosylation in brain neurons of diabetic rats, and to analyze the pathway of neuronal degeneration at the early stage of hyperglymecia. DESIGN: Randomized controlled animal experiment. SETTING: Department of Endocrinology, First hospital Affiliated to General Hospital of Chinese PLA and Beijing Laboratory for Brain Aging, Xuanwu Hospital Affiliated to Capital Medical University. MATERIALS: Thirty-five male Wistar rats (grade Ⅱ), aged 3 months old, and 11 male purebred Kunming mice (grade Ⅲ) without special pathogen, aged 3 months old, were provided by the Animal Room of Capital Medical University. METHODS: This experiment was carried out in the Beijing Laboratory for Brain Aging, Xuanwu Hospital Affiliated to Capital Medical University in 1998. The rats in the diabetic model group were intraperitoneally injected into 10 g/L STZ according to 60 mg/kg to establish rat models of diabetes mellitus. The blood glucose and body mass of rats in each group were determined respectively at 1, 2 and 3 months after modeling. The antibodies of advanced glycosylation end products (AGEs) of bovine serum albumin (anti-BSA) were self-prepared: ①The antigen of AGEs-BSA was prepared.②Eleven male Kuming mice (grade Ⅱ) of 3 months old without special pathogen were selected to inoculate AGEs-BSA. ③ The animals were immunized. ④Primary purification and detection of poly-antibodies of AGEs: the AGEs were performed immunohistochemical examination at 1 month after diabetic modeling by ELISA method. MAIN OUTCOME MEASURES: ① Detection results of blood glucose and body mass of rats in two groups at different time points. ② Determination of polyclonal antibody titer of AGEs-BSA. ③ Changes in immunohistochemical image of AGEs in brain tissue of rats in two groups. RESULTS: Thirteen rats in the diabetic model group and fifteen rats in the normal model group entered the stage of final analysis. ①Changes of blood glucose and body mass: At 1, 2 and 3 months after modeling, the blood glucose of rats in the diabetic model group were respectively(28.8±2.8),(23.1±5.5),(25.4±5.1) mmol/L, which were significantly higher than those in the normal control group [(6.2±0.9),(6.1±0.8),(6.1±0.7) mmol/L,P < 0.01]; At 1, 2 and 3 months after modeling, the body mass of rats in the diabetic model group were respectively (250.1±52.2),(263.8±50.0),(261.5±42.6) g, which were significantly lower than those in the normal control group [(422.6±36.2),(462.6±39.0),(485.0±28.8) g,P < 0.01].②Determination of antibody titer of immune serum: The mice were treated by AGEs-BSA of different concentrations twice. After that, the titer of AGEs -BSA was determined, and the results of which indicated that a higher absorbance existed at 1∶1 000. ③Determination of antigen concentration: The final titer of antibody in the abdominal dropsy was determined, and the results of which suggested that there was a much higher absorbance in the AGEs-BSA at the concentration of 5–50 mg/L. ④Determination of antibody titer in abdominal dropsy: The antibody titer in abdominal dropsy was detected by ELISA method with antigen at 20 mg/L, which indicated that the maximum absorbance (1.265±0.039) existed at 1∶4 000, and very larger absorbance (0.982±0.067) at 1∶20 000. The polyclonal antibody of AGEs-BSA was successfully prepared. ⑤Immunohistochemical detection results: The immunohistochemical staining of AGEs showed there were positive neurons in the first month in the diabetic model group, whereas it was not significant in the normal control group. The positive substances were found mainly in the cytoplasm. CONCLUSION: Hyperglycemia at the early stage of diabetes mellitus (1 month after modeling) can lead to protein nonenzymeatic glycosylation in brain neurons, and no obvious reactions mentioned above are found in the normal control group. It suggests that the degenerative changes of tissue structure of central nervous system are related with protein nonenzymeatic glycosylation caused by hyperglycemia.     

Spinal cord injury in patients with systemic lupus Erythematosus——Clinical manifestations, imaging characteristics and treatment

BACKGROUND： There are fewer reports on systemic lupus erythematosus （SLE） related myelitis, and definite and uniform therapeutic program is not available. OBJECTIVE： To observe the clinical manifestations, imaging characteristics, results of laboratory examination and treatment of SLE. DESIGN： A retrospective case analysis. SETTING： Department of Neurology, the Second Affiliated Hospital of Sun Yat-sen University. PARTICIPANTS： Totally 1 052 SLE inpatients were selected from the Second Affiliated Hospital of Sun Yat-sen University from January 1995 to May 2005, and they all accorded with the diagnostic standards for SLE set by American Rheumatism Association in 1982. 124 of them were diagnosed to have damage of central nervous system. Inclusive criteria： Patients with one of the focal physical signs, including mental and behavior disorders, headache, seizure and involvement of nervous system. Exclusive criteria： Patients with hypertensive encephalopathy, damage of nervous system due to uremia and infection of central nervous system. Spinal cord lesion occurred in 15 female cases of 23 - 51 years old. Informed consents were obtained from all the participants. METHODS： The physical signs, laboratory examinations, therapeutic program and prognosis were recorded in the 15 patients with symptoms of spinal cord lesions. All the patients underwent MRI scan of brain or lesioned segment of spinal cord, and 8 cases of them underwent lumbar puncture to determine intracranial pressure, routine and biochemical examinations were cerebrospinal fluid were performed. The disease activity of SLE in systems beyond central nervous system was evaluated with modified lupus activity criteria count （LACC）. MAIN OUTCOME MEASURES：① Incidence of SLE related myelitis, attack age distribution and its association with the activity of SLE; ② Comparisons of the clinical characteristics, cranial and spinal cord MRI manifestations, different therapeutic program and prognosis. RESULTS： All the 15 SLE patients were involved in the analysis of results. ① The incidence of SLE related myelitis was low （1%, 15/1 052）. ②SLE related myelitis occurred mostly when the SLE symptoms were active, and only a few occurred at the stable period. ③ Among the SLE patients, MRI displayed abnormal changes in 71% （10/14）, the typical changes appeared abnormal signals at corresponding spinal segments, manifested as prolonged T1 and T2 signals, thickened spinal segments. Lumbar segments were mostly involved. ④ Of the 9 patients treated with hormone impact, 7 cases （78%） had obvious improvements, and the effects were better in those treated with immunosuppressor combined with intravenous immunoglobulin of large dosage. CONCLUSION：① Myelitis is a rare complication of SLE.② MRI serves as a valuable supplementary approach in the diagnosis of SLE related myelitis without specificity. ③ Steroid pulse combined with immunosuppressor and intravenous immunoglobulin of large dosage is effective in the treatment.     

Changes in neuronal apoptosis and apoptosis modulatory factors in cerebral ischemia/reperfusion

BACKGROUND: The high concentration of glutamate release is the main cause for neuronal cell death. The relationship between glutamate level and apoptosis during ischemia/reperfusion injury is still unclear. OBJECTIVE: To observe the neuronal apoptosis at 24 and 72 hours following cerebral ischemia/reperfusion in rats, and analyze the possible influencing factors. DESIGN: A randomized controlled animal experiment. SETTING: School of Medicine, Southern Yangtze University. MATERIALS: Totally 30 male adult Sprague Dawley (SD) rats of clean grade, weighing 240–290 g, were obtained from Shanghai Experimental Animal Center, Chinese Academy of Sciences. The rats were randomly divided into sham-operated group (n=10) and model group (n=20). Each group was observed at 24 and 72 hours after ischemia/reperfusion, 5 rats at each time point in the sham-operated group, whereas 12 at 24 hours and 8 at 72 hours in the model group. Kits for determining apoptosis and Bcl-2 were bought from Wuhan Boster Biological Technology, Co., Ltd.; Kit for calcineurin from Nanjing Jiancheng Bioengineering Institute. METHODS: The experiment was carried out in the Functional Scientific Research Room of Southern Yangtze University from June to October in 2006. ① Right middle cerebral artery was occluded by inserting a thread through internal carotid artery (ICA). The surgical process for the sham-operated rats was the same as that in the model group except a nylon suture inserted the ICA. According to Longa five-degree standard, the neurological deficit evaluation of rats was evaluated after surgery, and grades 1–3 were taken as successful model establishment. The blood was recirculated by withdrawing the nylon filament under anesthesia at 2 hours after ischemia in successful rat models. ②After reperfusion, the brain tissue was quickly removed at 24 or 72 hours and the slices were obtained from optic chiasma to funnel manubrium. The changes of the number of apoptotic cells were observed using the terminal deoxynucleotidyl transferase mediated dUTP-biotin nick-end labeling method. The expressions of Bcl-2 protein were determined with immunohistochemical staining. The activity of calcineurin was determined by the inorganic phosphorus method. The content of excitatory amino acid was detected by high performance liquid chromatography. MAIN OUTCOME MEASURES: ① Glutanate content in brain tissue; ② Conditions of apoptosis; ③ Calcineurin activity in brain tissue; ④ Bcl-2 expression in brain tissue. RESULTS: Totally 30 SD rats were used, 5 died and the other 25 were involved in the analysis of results. ① Changes of apoptosis: There were 0–3 apoptotic cells in the sham-operated group. In the model group, the numbers of apoptotic cells were obviously increased at 24 and 72 hours of reperfusion (P < 0.01), and it was markedly reduced at 72 hours as compared with 24 hours (P < 0.01). ② Changes of glutanate content: The glutamate contents at 24 and 72 hours of reperfusion in the model group were obviously higher than those in the sham-operated group (P < 0.01); In the model group, it was obviously increased at 24 hours as compared with 72 hours (P < 0.01). ③ Changes of Bcl-2 protein: In the model group, the Bcl-2 protein expression had no obvious changes at 24 hours of reperfusion, and it was obviously enhanced at 72 hours, which was obviously different from that in the sham-operated group and that at 24 hours (P < 0.01). ④ Changes of calcinerin activity: In the model group, the activity of calcineurin in brain tissue had no obvious changes at 24 hours of reperfusion; The activity of calcineurin at 72 hours was obviously higher than that in the sham-operated group and that at 24 hours (P < 0.01). CONCLUSION: The brain cyto-apoptosis action at different time points following reperfusion incompletely depends on the glutamate levels, while it depends on the interaction of some apoptosis related factors, such as amino acid, calcineurin, and Bcl-2, etc.     

Neurovascular compression and decompression of the intracranial vagus for blood pressure

BACKGROUND: Previous researches demonstrated that neurovascular decompression could cure hypertension; however, whether it could effectively control refractory hypertension after hypertensive cerebral hemorrhage should be further studied. OBJECTIVE: To observe the effect of neruovascular compression on intracranial vagus for blood pressure of dogs and investigate the effect of neurovascular decompression on blood pressure of patients with hypertensive cerebral hemorrhage. DESIGN: Randomized controlled animal study, clinical effects and retrospective analysis. SETTING: Department of Neurosurgery, Changzheng Hospital Affiliated to the Second Military Medical University of Chinese PLA. MATERIALS: The experiment was carried out in the Department of Neurosurgery, Changzheng Hospital Affiliated to the Second Military Medical University of Chinese PLA from May to October 2006. A total of 15 healthy adult dogs of both genders were randomly divided into experimental group (n =10) and control group (n =5). Clinical observation: A total of 41 patients with hypertensive cerebral hemorrhage were selected from the Department of Neurosurgery, General Hospital of Nanjing Military Area Command of Chinese PLA and the Department of Neurosurgery, Changzheng Hospital Affiliated to the Second Military Medical University of Chinese PLA from October 1999 to October 2006. Among them, one patient had brain stem hemorrhage. There were 27 males and 14 females aged from 41 to 66 years. Inclusion criteria: All patients were diagnosed with CT examination once or several times. Volume of hematoma ranged from 50 to 120 mL and had obviously operative indication. All patients provided consents. In addition, another 281 patients with hypertensive cerebral hemorrhage who received traditionally internal and surgical therapies in our departments of neurosurgery, neurology and emergency room were selected in the control group. METHODS: ① Animal experiments: 20 cm autochthonous great saphenous vein was taken from dogs in the experimental group and coincided with tip of facial artery to form arterial loop so as to oppress left vagus and lateral bulb abdomen. In addition, 20 cm autochthonous great saphenous vein was taken from dogs in the control group and coincided with tip of facial artery to establish arterial loop so as to oppress left cerebellum to observe changes of blood pressure before and at 1, 2, 3 and 4 weeks after operation. ② Clinical observation: Among 41 patients with hypertensive cerebral hemorrhage including one with brain stem hemorrhage, they received microvascular decompression of vagus immediately after getting rid of intracerebral hematoma and stopping bleeding to observe its effect of depressurization. All patients and their relatives provided consents. ③ A total of 281 patients with hypertensive cerebral hemorrhage who discharged after the treatment of traditionally internal and surgical therapies were studied retrospectively to observe changes of blood pressure after routine treatment and compare the results with neurovascular decompression. MAIN OUTCOME MEASURES: ① Changes of blood pressure of experimental dogs; ② effect of vascular decompression of vagus for blood pressure of patients with hypertensive cerebral hemorrhage after clearing intracerebral hematoma; ③ different effects of neurovascular decompression and routinely internal and surgical therapies on hypertension. RESULTS: ① Results of animal experiments: Nine dogs in the experimental group survived. At 1, 2, 3 and 4 weeks after operation, blood pressure of dogs in the experimental group was (139.77±4.06), (149.11±4.90), (148.10±4.16), (147.76±4.15) mm Hg (1 mm Hg=0.133 kPa), which was higher than that of dogs in the control group [(117.20±2.74), (116.65±3.74), (116.26±1.8), (115.81±3.76) mm Hg, P < 0.01]. ② Results of clinical observation: Among 41 patients, 8 (20%) cases died during the operation. In addition, among other 33 (80%) survival patients, 11 (33%) cases had normal blood pressure; blood pressure of 14 (43%) cases was improved or closed to normal value; blood pressure of 8 (24%) cases was not changed obviously as compared with that before operation. ③ The results demonstrated that, by using traditionally internal and surgical therapies, among 281 patients with hypertensive cerebral hemorrhage, blood pressure of about 15% cases was recovered or closed to normal value. Those mentioned above did not have history of hypertension before hemorrhage. However, patients who had history of hypertension before hemorrhage received the traditionally internal or surgical therapies, and the blood pressure was not improved to the normal value after the treatment. CONCLUSION: ① Neurovascular compression in left intracranial vagus can cause obvious increase of blood pressure of dogs, and the increasing volume was 30 mm Hg. ② Vascular decompression of vagus has a great effect on refractory hypertension, and the improvement of blood pressure is superior to traditionally internal and surgical therapies in clinic.     

Changes of serum high sensitive C-reactive protein in patients with acute cerebral infarction

BACKGROUND: Serum high sensitive C-reactive protein (hs-CRP), which regards as a high sensitive mark of systemic inflammatory response syndrome, can provide a lot of valuable information for the treatment and prognosis of cerebrovascular disease. OBJECTIVE: To observe the differences of blood glucose, lipid, homocysteine and previous disease history among patients with acute cerebral infarction at various levels of hs-CRP and compare changes of hs-CRP of patients with various degrees of neurologic impairment. DESIGN: Contrast observation. SETTING: Department of Neurology, Shenzhou Hospital, Shenyang Medical College. PARTICIPANTS: A total of 102 patients with acute cerebral infarction were selected from Department of Neurology, Shenzhou Hospital of Shenyang Medical College from February 2005 to September 2006, including 55 males and 47 females aged from 55 to 86 years. All accepted patients met the diagnostic criteria of cerebral infarction established by the Fourth National Cerebrovascular Disease Academic Meeting and were diagnosed with CT or MRI examination. All patients provided the confirmed consent. Based on clinical criteria of neurologic impairment established by the Fourth National Cerebrovascular Disease Academic Meeting, patients were randomly divided into mild group (0–15 points, n =46), moderate group (16–30 points, n =38) and severe group (31–45 points, n =18). In addition, based on hs-CRP level within 72 hours, patients were divided into normal group (hs-CRP ≤ 3 mg/L, n =53) and increasing group (hs-CRP > 3 mg/L, n =49). METHODS: ① 2 mL venous blood was selected from hospitalized patients in the next morning to separate serum. Quantitative measurement of hs-CRP was dealt with Latex Enhnced Turbidimetric Immunoassay (LETIA). ② Fasting venous blood was colleted from hospitalized patients in the next morning to measure numeration of white blood cells, fibrinogen, blood glucose, total cholesterol (TC), triacylglycerol (TG), high density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol (LDL-C) and homocysteine. ③ Measurement data were compared with t test or analysis of variance. MAIN OUTCOME MEASURES: ① Comparisons of serum biochemical indexes among patients with various levels of hs-CRP; ② comparisons of risk factors among patients with various levels of hs-CRP; ③ comparisons of levels of hs-CRP among patients with various degrees of clinical neurologic impairment. RESULTS: A total of 102 patients were involved in the final analysis. ① Plasma fibrinogen and numeration of leucocytes were more in the increasing group than those in the normal group (t =4.39, 3.54, P < 0.01); while, there were no significant differences of blood glucose, TC, TG, HDL-C, LDL-C and homocysteine between the two groups (P > 0.05). ② Percentage of patients with hypertension and diabetes mellitus (DM) was higher in the increasing group than the normal group (χ2=3.98, 4.23, P < 0.05); while, percentage of patients with smoking in the increasing group was not significantly different from that of patients in the normal group (P > 0.05). ③ Level of hs-CRP of patients with severe neurologic impairment was higher than that of patients with moderate neurologic impairment (t =2.273, P < 0.05); that of patients with moderate neurologic impairment was higher than that of patients with mild neurologic impairment (t =2.586, P < 0.05); that of patients with severe neurologic impairment was obviously higher than that of patients with mild neurologic impairment (t = 4.913, P < 0.01). CONCLUSION: ① With the increase of hs-CRP, plasma fibrinogen and numeration of leucocytes of patients with acute cerebral infarction is increased, especially, they are increased remarkably among patients who have history of diabetes mellitus and hypertension. ② Increase of level of hs-CRP can be regarded as one of marks to evaluate severity of acute stroke.