Increases in lung and brain water following experimental stroke: effect of mannitol and hypertonic saline

OBJECTIVE: Pulmonary edema is a serious condition following brain injury of diverse etiologies, including large hemispheric infarctions. We have previously shown that treatment with hypertonic saline attenuates cerebral edema associated with experimental ischemic stroke. In a well-characterized animal model of large ischemic stroke, we tested the hypotheses that lung water increases following cerebral ischemia and determined the effects of osmotherapy with hypertonic saline and mannitol on total lung water, as well as on cerebral edema. DESIGN: Prospective laboratory animal study. SETTING: Research laboratory in a university teaching hospital. SUBJECTS: Adult male Wistar rats (300-450 g, n = 103). INTERVENTIONS: Under controlled conditions of normoxia, normocarbia, and normothermia, spontaneously breathing, halothane-anesthetized (1.0-1.5%) rats were subjected to permanent middle cerebral artery occlusion by the intraluminal occlusion technique. MEASUREMENTS AND MAIN RESULTS: Cerebral perfusion was monitored by laser-Doppler flowmetry over ipsilateral parietal cortex to ensure adequate vascular occlusion. At 6 hrs following middle cerebral artery occlusion, rats were treated in a blinded randomized fashion with no intravenous fluids (n = 24), a continuous intravenous infusion (0.3 mL/hr) of 0.9% saline (n = 21), 20% mannitol (2 g/Kg) (n = 20), 5% hypertonic saline (n = 20), or 7.5% hypertonic saline (n = 18) as a chloride/acetate mixture (50:50) until the end of the experiment. Brains and lungs were harvested, and tissue water content was estimated by comparing wet-to-dry weight ratios of ipsilateral and contralateral cerebral hemispheres at 48 hrs postischemia. Sham-operated rats served as controls (n = 20). Serum osmolality was determined at the end of the experiment in all animals. Lung water content was increased significantly in rats subjected to middle cerebral artery occlusion and treated with no intravenous fluids (76.7 +/- 0.7%, 317 +/- 7 mOsm/L) (mean +/- sd) and saline (76.8 +/- 1.2%, 311 +/- 10 mOsm/L), compared with sham-operated controls (74.5 +/- 0.9%, 302 +/- 4 mOsm/L). Treatment with 20% mannitol (74.4 +/- 1.2%, 352 +/- 15 mOsm/L), 5% hypertonic saline (75.6 +/- 1.3%, 339 +/- 16 mOsm/L), and 7.5% hypertonic saline (74.9 +/- 0.7%, 360 +/- 23 mOsm/L) significantly attenuated lung water content. Hemispheric brain water content increased both in the ipsilateral ischemic and contralateral hemispheres treated with saline (ipsilateral, 85.1 +/- 1.7%; contralateral, 80.7 +/- 0.7%), compared with sham-operated controls (ipsilateral, 79.6 +/- 0.9%; contralateral, 79.5 +/- 0.9%), as well as in rats that received no fluids (ipsilateral, 84.6 +/- 1.8%; contralateral, 80.4 +/- 0.9%). Treatment with 5% hypertonic saline (ipsilateral, 83.8 +/- 1.0%; contralateral, 79.7 +/- 0.6%) and 7.5% hypertonic saline (ipsilateral, 82.3 +/- 1.3%; contralateral, 78.6 +/- 0.7%) resulted in attenuation of stroke-associated increases in brain water content to a greater extent than mannitol (ipsilateral, 83.6 +/- 1.6%; contralateral, 79.1 +/- 1.0%). CONCLUSIONS: In a well-characterized animal model of large ischemic stroke, total lung water content increases, which is likely neurogenic in origin. Attenuation of stroke-associated increases in lung and brain water content with continuous infusion of hypertonic saline may have therapeutic implication in the treatment of cerebral and pulmonary edema following ischemic stroke.     

Osmotherapy with hypertonic saline attenuates water content in brain and extracerebral organs

OBJECTIVE: Because of their beneficial effects in patients with hemorrhagic shock and multiple-system trauma, hypertonic saline solutions are increasingly being used perioperatively for volume resuscitation. Although the anti-edema effects of hypertonic saline on brain are well documented in a variety of brain injury paradigms, its effects on the water content on other organs has not been studied rigorously. In this study, we tested the hypothesis that a) hypertonic saline when given as an intravenous bolus and continuous infusion attenuates water content of small bowel, lung, and brain in rats without neuro-injury; and b) attenuation of stroke-associated increases in lung water is dependent on achieving a target serum osmolality. DESIGN: Prospective laboratory animal study. SETTING: Research laboratory in a teaching hospital. SUBJECTS: Adult male Wistar rats. INTERVENTIONS: In the first series of experiments, under controlled conditions of normoxia, normocarbia, and normothermia, spontaneously breathing, halothane-anesthetized (1.0-1.5%) adult male Wistar rats (280-320 g) were treated in a blinded randomized fashion with 7.5% hypertonic saline or 0.9% normal saline in a 8-mL/kg intravenous infusion for 3 hrs followed by a continuous intravenous infusion (1 mL/kg/hr) of 5% hypertonic saline or normal saline, respectively (n=10 each), for 48 hrs. A second group of rats were treated with continuous infusion only for 48 hrs of either 7.5% hypertonic saline or normal saline (1 mL/kg/hr) (n=10 each) without an intravenous bolus. Na?ve rats served as controls (n=10). Tissue water content of small bowel, lung, and brain was determined by comparing the wet-to-dry ratios at the end of the experiment. In a second series of experiments, rats (n=94) were subjected to 2 hrs of transient middle cerebral artery occlusion by the intraluminal occlusion technique. At 6 hrs following middle cerebral artery occlusion, rats were treated in a blinded randomized fashion with a continuous intravenous infusion of normal saline, 3% hypertonic saline, or 7.5% hypertonic saline for 24, 48, 72, and 96 hrs. Surgical shams served as controls (n=7). Hypertonic saline was instituted as chloride/acetate mixture (50:50) in all experiments. Serum osmolality was determined at the end of the experiment in all animals. MEASUREMENTS AND MAIN RESULTS: In rats without neuro-injury that received intravenous bolus followed by a continuous infusion, lung water content was significantly reduced with hypertonic saline (73.9+/-1.1%; 359+/-10 mOsm/L) (mean+/-sd) compared with normal saline treatment (76.1+/-0.53%; 298+/-4 mOsm/L) as was water content of small bowel (hypertonic saline, 69.1+/-5.8%; normal saline, 74.7+/-0.71%) and brain (hypertonic saline, 78.1+/-0.87%; normal saline, 79.2+/-0.38%) at 48 hrs. Stroke-associated increases in lung water content were attenuated with 7.5% hypertonic saline at all time points. There was a strong correlation between serum osmolality and attenuation of stroke-associated increases in lung water content (r=-.647) CONCLUSIONS: Bowel, lung, and brain water content is attenuated with hypertonic saline when serum osmolality is >350 mOsm/L without adverse effect on mortality in animals with and without neuro-injury. Attenuation of water content of extracerebral organs with hypertonic saline treatment may have therapeutic implications in perioperative fluid management in patients with and without brain injury.     

The effect of oxygen therapy on brain damage and cerebral pO(2) in transient focal cerebral ischemia in the rat

We examined the effect of hyperbaric oxygen (HBO) and normobaric oxygen (NBO) on neurologic damage and brain oxygenation before and after focal cerebral ischemia in rats. A middle cerebral artery occlusion (MCAO)/reperfusion rat model was used. The rats were sacrificed 22 h after reperfusion, and the infarct volume was evaluated. In study A, HBO (2.0 ATA), NBO (100% oxygen) and normobaric air (NBA) were each administered for 60 min in five different rat groups. The sizes of the infarcts after HBO and NBO applied during ischemia were 8.8 +/- 2.8% and 22.8 +/- 3.7% respectively of the ipsilateral non-occluded hemisphere. The infarct size after HBO applied during ischemia was statistically smaller than for NBO and NBA exposure (p < 0.01). In study B, cerebral pO(2) was measured before and after MCAO and HBO exposure (2.0 ATA for 60 min) in six rats using electron paramagnetic resonance (EPR) oximetry. The pO(2) in the ischemic hemisphere fell markedly following ischemia, while the pO(2) in the contralateral hemisphere remained within the normal range. Measurements of the pO(2) performed minutes after HBO exposure did not show an increase in the ischemic or normal hemispheres. The mean relative infarct size was consistent with the changes observed in study A. These data confirm the neuroprotective effects of HBO in cerebral ischemia and indicate that in vivo EPR oximetry can be an effective method to monitor the cerebral oxygenation after oxygen therapy for ischemic stroke. The ability to measure the pO(2) in several sites provides important information that should help to optimize the design of hyperoxic therapies for stroke.     

An early bolus of hypertonic saline hydroxyethyl starch improves long-term outcome after global cerebral ischemia

OBJECTIVE: The beneficial effect of hypertonic saline solutions in the emergency treatment of shock and traumatic brain injury is well described. The present study determines effects of a single bolus of hypertonic saline on long-term survival, neurologic function, and neuronal survival 10 days after global cerebral ischemia. In addition, we evaluated the therapeutic window for hypertonic saline treatment (early vs. delayed application). DESIGN: Laboratory experiment. SETTING: University laboratory. SUBJECTS: Male Wistar rats weighing 240-330 g. INTERVENTIONS: Rats were submitted to temporal global cerebral ischemia using temporary bilateral carotid occlusion combined with hypobaric hypotension. Animals received 7.5% saline/6% hydroxyethyl starch (HHS) or vehicle (NaCl 0.9%) at either 1.5 mins (early treatment) or 31.5 mins (delayed treatment) of reperfusion. Regional cerebral blood flow (rCBF) and physiologic variables were measured during insult and early reperfusion. Animal survival and neurologic function were evaluated throughout the 10-day observation period. Quantification of brain injury was performed on day 10. MEASUREMENTS AND MAIN RESULTS: Early treatment with HHS resulted in a robust restoration of rCBF after ischemia, reduced postischemic mortality by 77% (9% vs. 39% in vehicle-treated controls), ameliorated neurologic performance (Neuro-Deficit-Score 10 days after insult, 96 +/- 0.7 vs. 85 +/- 1.4, mean +/- se), and almost blunted neuronal cell death (hippocampal CA1, 2150 +/- 191 vs. 884 +/- 141 neurons/mm; cortex, 1746 +/- 91 vs. 1060 +/- 112). In contrast, delayed treatment resulted in no sustained effects. CONCLUSIONS: Timing of HHS treatment is critical after experimental global cerebral ischemia to reduce mortality, improve neurologic function, and neuronal survival. Our results suggest that early application of HHS may be a potential neuroprotective strategy after global cerebral ischemia.     

Regional intraparenchymal pressure differences in experimental intracerebral hemorrhage: effect of hypertonic saline

OBJECTIVE: To study regional intraparenchymal pressures within the cranial cavity during and after formation of intracerebral hemorrhage. We also assessed the effect of hypertonic saline on intraparenchymal pressure in different brain regions and on regional brain distribution of sodium within the brain. DESIGN: Prospective, controlled, laboratory trial. SETTINGS: Animal research laboratory. SUBJECTS: Eight mongrel dogs, weighing 15-25 kg. INTERVENTION: We introduced an intracerebral hematoma in eight mongrel dogs by infusing 6 mL of autologous arterial blood in the deep white matter adjacent to the basal ganglia. Sodium chloride (23.4%, 1.4 mL/kg) then was administered intravenously 6 hrs after introduction of hematoma. MEASUREMENTS AND MAIN RESULTS: Parenchymal pressure monitors were placed in the perihematoma region, both frontal lobes, and the cerebellum to record intraparenchymal pressure during and 6 hrs after intracerebral hematoma formation. Intraparenchymal pressure measurements were recorded for 3 more hours after administration of 23.4% sodium chloride. Regional cerebral perfusion pressure was calculated for each intraparenchymal pressure measurement. Regional sodium distribution was measured in extracts from brain regions by using ion selective electrode technique. A higher elevation in intraparenchymal pressure was recorded in the perihematoma region during the introduction of the hematoma compared with other compartments. After 5 mL of autologous blood was introduced, intraparenchymal pressure (mm Hg +/- SE) was significantly higher in the perihematoma region (42.1 +/- 3.5) than in the ipsilateral (30.0 +/- 4.6, p <.05) and contralateral (27.1 +/- 5.5, p <.01) frontal lobes and cerebellum (29.1 +/- 4.5, p <.05). Four hours after introduction of the hematoma, the cerebral perfusion pressure recorded in the perihematoma region (43.6 +/- 9.7) remained significantly lower than in the ipsilateral (58.6 +/- 9.3, p <.05) but not the contralateral frontal lobes (54.7 +/- 10.1) and cerebellum (51.0 +/- 11.1). Administration of 23.4% sodium chloride immediately reduced intraparenchymal pressure in each compartment. This effect was still observed at 3 hrs in each compartment. Sodium concentration was higher in the perihematoma region than in the frontal lobes, cerebellum, or brain stem. CONCLUSIONS: Prominent differences were observed in intraparenchymal pressure and cerebral perfusion pressure in the perihematoma region and frontal lobes during and after intracerebral hematoma. We speculate that the potential importance of regional intraparenchymal pressure differences in the clinical settings may be under appreciated. In this canine model of intracerebral hematoma, a single dose of hypertonic saline effectively reduces the intraparenchymal pressure in all regions of the brain.     

大鼠脑缺血再灌注区细胞间粘附分子1表达与白细胞浸润的实验研究

目的：观察大鼠脑缺血－再灌注后不同时间缺血区细胞间粘附分子－１（ＩＣＡＭ－１）的表达和中性粒细胞的浸润以及神经细胞的损伤情况。方法：４０只Ｗｉｓｔａｒ大鼠分为对照组、假手术组和缺血２小时再灌注２、４、１２、２４、４８、９６小时组,分别用免疫组织化学和组织切片方法检测大脑中动脉阻塞２小时再灌注不同时间点局部脑组织ＩＣＡＭ－１蛋白表达及中性粒细胞浸润情况。结果：大鼠脑缺血 灌液２小时局部脑组织ＩＣＡＭ     

电针对脑梗死大鼠神经前体细胞增殖水平的影响

目的研究电针治疗对脑梗死病灶周围及海马处神经前体细胞增殖水平的影响。方法采用易卒中型肾血管性高血压大鼠(RHRSP)，电凝法制成大脑中动脉闭塞(MCAO)模型。行神经行为学功能评定、神经前体细胞标记及电针治疗，免疫组化染色观察并计数电针治疗后梗死灶边缘、对侧镜区及双侧海马5-溴脱氧尿核苷(bmmodeoxyuridine，BrdU)标记的细胞。结果MCAO后神经功能评分减低，约5d后恢复正常。电针治疗促使梗死灶边缘、对侧镜区BrdU阳性细胞增多，随着治疗时间增加，细胞增多更明显。结论电针治疗可促使神经前体细胞增殖及迁移，可能是电针促进康复疗效的重要机制之一。     

脑缺血后大鼠神经元和星形胶质细胞p15ink4b表达的差异研究

目的:比较研究大鼠局灶性脑缺血再灌注损伤后细胞周期蛋白依赖性激酶抑制因子p15ink4b在神经元和星形胶质细胞的表达。方法:建立大鼠大脑中动脉阻塞(MCAO)再灌注模型,应用流式细胞术检测各组MCAO再灌注后不同时期神经元和星形胶质细胞中的p15ink4b的表达。结果:缺血侧皮质区星形胶质细胞中的p15ink4b的表达在再灌注3d后开始下调,14d显著下调,与假手术组比较有显著性差异(P<0.05);缺血侧皮质神经元中的p15ink4b在再灌注14d后表达下调,与假手术组比较有显著性差异(P<0.05)。结论:局灶性脑缺血再灌注损伤后,缺血侧皮质区星形胶质细胞和神经元均有p15ink4b不同程度的表达下调,星形胶质细胞中的p15ink4b表达下调比神经元更为显著。     

Effect of ifenprodil, a polyamine site NMDA receptor antagonist, on brain edema formation following asphyxial cardiac arrest in rats

OBJECTIVE: Brain edema occurs in experimental and clinical cardiac arrest (CA) and is predictive of a poor neurological outcome. N-Methyl--aspartate (NMDA) receptors contribute to brain edema elicited by focal cerebral ischemia/reperfusion (I/R). Ifenprodil, a NMDA receptor antagonist, attenuates brain edema and injury size in rats after focal cerebral I/R. We assessed the hypothesis that ifenprodil reduces CA-elicited brain edema. METHODS: Eighteen male Sprague-Dawley rats were assigned to group 1 (normal control, n=6), group 2 (placebo-treated CA, n=6), or group 3 (ifenprodil-treated CA, n=6). CA was induced by 8 min of asphyxiation and the animals were resuscitated with cardiopulmonary resuscitation (CPR), ventilation, epinephrine (adrenaline), and sodium bicarbonate (NaHCO3). Ifenprodil of 10 mg/kg or a placebo vehicle was given intraperitoneally 5 min before CA. Brain edema was determined by brain wet-to-dry weight ratio at 1 h after resuscitation. RESULTS: There were no differences between groups 2 and 3 in all physiological variables at baseline. Time from asphyxiation to CA was 201.5 +/- 7.5 s in group 2 and 160.7 +/- 10.4 s in group 3 (P<0.001). Resuscitation time was 68.2 +/- 13.3 s in group 2 and 92.8 +/- 18.2 s in group 3 (P<0.05). Ifenprodil decreased mean arterial pressure (MAP) before asphyxiation, from 128 +/- 7 in group 2 to 82 +/- 15 mmHg in group 3 (P<0.001), and negated immediate post-resuscitation hypertension. Brain wet-to-dry weight ratio was 5.64 +/- 0.44 in group 1, 7.34 +/- 0.95 in group 2 (P<0.01 versus group 1), and 5.93 +/- 0.40 in group 3 (P<0.05 versus group 2). CONCLUSIONS: Ifenprodil reduces CA-elicited brain edema. In addition, we observed significant hemodynamic changes caused by ifenprodil.     

远程缺血预处理对大鼠局灶性脑缺血/再灌注损伤的保护作用

目的探讨远程缺血预处理(RIPC)对大鼠局灶性脑缺血/再灌注(I/R)损伤的影响。方法SD雄性大鼠70只,随机分组,每组10只。对照组仅行单纯缺血后再灌注;RIPC组按RIPC与脑缺血间隔时间不同分为30min及1、2、12、24和48h组,即反复3次夹闭双侧股动脉造成肢体缺血5min、再灌注5min后,分别间隔30min及1、2、12、24和48h后,行大脑中动脉栓塞(MCAO)120min、再灌注24h。对各组动物进行神经功能缺损评分,然后行氯化三苯四唑(TTC)染色,计算脑梗死容积。结果与对照组比较,RIPC1、2和24h组神经功能缺损评分显著下降,差异有显著性(P均<0.05);而RIPC30min、12h和48h组与对照组比较差异均无显著性(P均>0.05)。脑梗死容积百分比RIPC1h组〔(17.9±7.5)%,P=0.016〕、2h组〔(18.3±11.2)%,P=0.019〕和24h组〔(20.2±11.9)%,P=0.047〕均明显小于对照组〔(30.5±9.8)%〕;而RIPC30min、12h和48h组与对照组比较差异无显著性(P均>0.05)。结论RIPC对大鼠局灶性脑I/R损伤有保护作用,其保护时程为预处理后1～2h,24h后再次出现。     

Effect of AR-R 17477, a potent neuronal nitric oxide synthase inhibitor, on infarction volume resulting from permanent focal ischemia in rats

OBJECTIVE: We tested whether AR-R 17477, a selective inhibitor of neuronal nitric oxide synthase, reduces brain injury in rats subjected to permanent focal ischemia. DESIGN: Randomized within cohort; nonblinded study. SETTING: University basic science laboratory. SUBJECTS: Halothane-anesthetized male Wistar rats (n = 53). INTERVENTIONS: Rats were treated with either intravenous saline (diluent) or AR-R 17477 (1 or 3 mg/kg) 30 mins before or 60 mins after the onset of permanent focal cerebral ischemia. Infarction volume was determined at 18 or 48 hrs of ischemia. MEASUREMENTS AND MAIN RESULTS: Pretreatment with 1 mg/kg AR-R 17477 was associated with a decreased infarct volume (2,3,5-triphenyltetrazolium chloride staining) in the striatum (saline, 81+/-7 mm3; AR-R 17477, 55+/-3 mm3) but not in the cortex at 18 hrs of occlusion (saline, 302+/-29 mm3; AR-R 17477, 237+/-36 mm3). However, this therapeutic effect of AR-R 17477 was no longer evident if the rats were allowed to survive for 48 hrs before analysis of infarction volume. In fact, in this separate cohort of animals, three of eight AR-R 17477-treated and five of eight saline-treated rats died before completing 48 hrs of ischemia. Efficacy of AR-R 17477 was completely absent (even at 18 hrs of ischemia) when drug treatment was delayed until 1 hr after the onset of ischemia. Infarction volume at 18 hrs of ischemia was similar between rats treated with saline, 1 mg/kg (cortex, 229+/-43 mm3; striatum, 67+/-8 mm3) or 3 mg/kg AR-R 17477 (cortex, 284+/-34 mm3; striatum, 75+/-5 mm3). In addition, only one of eight rats treated with 3 mg/kg AR-R 17477 at 1 hr of ischemia survived 48 hrs of occlusion, compared with three of eight rats treated with saline. CONCLUSIONS: Neuronally generated nitric oxide is a mediator of brain injury during permanent focal ischemia in rats. However, severity of the ischemic insult appears to limit the therapeutic efficacy of the specific neuronal nitric oxide synthase inhibitor, AR-R 17477.     

巴曲酶对大鼠局灶性脑缺血再灌注后血小板活化因子和血小板活化因子受体mRNA表达的影响

背景巴曲酶是目前比较公认的治疗缺血性脑血管病的理想药物之一,被广泛地应用于临床,因此对其在脑缺血再灌注损伤中的保护作用进行深入认识很有必要.目的探讨巴曲酶对大鼠局灶性脑缺血再灌注后血小板活化因子(PAF)水平及PAF受体基因(PAF-RmRNA)表达的影响.设计完全随机区组设计.地点和对象实验于2004-03/12在哈尔滨医科大学附属第二医院科研中心完成.选择40只健康Wistar雄性大鼠,体质量200～250 g,随机分为5组,每组8只.Ⅰ组假手术组;Ⅱ组为生理盐水组Ⅱa为缺血6h再灌注6 h组,Ⅱb为缺血6 h组;Ⅲ组为巴曲酶组Ⅲa为缺血6 h再灌注6 h组,Ⅲb为缺血6 h组.方法线栓法建立大鼠大脑中动脉闭塞(MCAO)及再通模型.应用RT-PCR技术检测MCAO及再通后缺血半暗带皮质PAF受体基因表达,同时用ELISA检测对应血浆PAF值.主要观察指标不同时间点各组缺血半暗带皮质PAF mRNA表达及血浆PAF值.结果生理盐水组中再灌组及缺血组PAF值均明显升高,Ⅱa,Ⅱb分别为(1 480±249)和(1 052±199)ng/L,而PAF-RmRNA表达降低,分别为0.44±0.06和0.48±0.05,分别与对应假手术组比较非常显著性意义(P＜0.01).巴曲酶组中再灌注及缺血组PAF值均降低,为(848±80)和(743±105)ng/L,PAF-RmRNA表达增强(0.63±0.08和0.67±0.06),与对应生理盐水组比较差异有显著性意义(P＜0.01).结论巴曲酶可降低脑缺血再灌注后血浆中PAF水平,并且可能对脑缺血再灌注缺血半暗带皮质组织PAF-RmRNA表达有影响,以期为预防性干预提供试验数据.     

Reduced rate of bacterial translocation and improved variables of natural killer cell and T-cell activity in rats surviving controlled hemorrhagic shock and treated with hypertonic saline

OBJECTIVE: To examine the effect of hypertonic saline on bacterial translocation and the number and function of natural killer and T cells in controlled hemorrhagic shock. DESIGN: Randomized controlled study. Duration of follow-up was 24 hrs. SETTING: University research laboratory. SUBJECTS: Adult male Sprague-Dawley rats, weighing 310-390 g. INTERVENTIONS: Controlled hemorrhagic shock was induced by blood withdrawal (mean arterial pressure, 30-40 mm Hg) and maintained for 30 mins. The animals were randomly divided into three groups: group 1 (n = 10) was sham-operated, group 2 (n = 10) was untreated, and group 3 was treated with 5 mL/kg hypertonic saline (n = 10). The rats were killed after 24 hrs. MEASUREMENTS AND MAIN RESULTS: Infusion of hypertonic saline in group 3 was followed by reduced bacterial translocation rate (5.0 +/- 2.2% vs. 18.3 +/- 5.3%, p <.033). The total mass of bacteria isolated from hypertonic saline-treated animals with bacterial translocation was 7.8- to 10.4-fold less than that from untreated rats. Controlled hemorrhagic shock resulted in a low percentage of CD4+ cells in blood (35.2 +/- 3.9%, p <.05) and lymph nodes (44.4 +/- 4.5%, p <.05) and depressed CD4 expression on blood (82 +/- 13 arbitrary units, p <.005) and lymph node (168 +/- 24 arbitrary units, p <.03) cells. A compensatory mobilization of NKR-P1+ cells from lymph nodes (8.6 +/- 2.3%, p <.05) to blood (21.2 +/- 5.2%, p <.01) with down-regulated NKR-P1 expression on blood cells (59 +/- 10 arbitrary units, p <.005) was observed. Natural killer cell-mediated cytotoxicity was decreased (67.9 +/- 9.7%, p <.05). Hypertonic saline treatment greatly stimulated CD4 expression on blood (419 +/- 113 arbitrary units, p <.005) and lymph node (553 +/- 115 arbitrary units, p <.03) cells. Also, normalization of NKR-P1 expression (160 +/- 19 arbitrary units, p <.005) and restoration of natural killer cell-mediated cytotoxicity to near normal values (88.6 +/- 7.4%, p <.05) were demonstrated. CONCLUSION: Controlled hemorrhagic shock was accompanied by CD4+ cells suppression and excessive recruitment of natural killer cells with abnormally low NKR-P1 expression and suppressed cytolytic activity into circulation. Infusion of hypertonic saline reversed these changes and reduced bacterial translocation.     

经颈内动脉冲洗治疗急性脑缺血大鼠的实验研究

目的：观察经颈内动脉冲洗法对急性脑缺血大鼠脑损伤程度的影响。方法：采用改良的MCAO方法造模，MCA梗阻2h后，冲洗组于再灌注超早期经颈内动脉分别注入．3ml生理盐水或中药溶液，对照组于腹腔内注入等体积中药溶液。再灌注12、24和48h，测定大鼠神经功能缺损评分。再灌注48h，测定脑梗死体积。结果：缺血再灌注48h后，和模型组相比，中药冲洗组和腹腔给药组脑梗死体积明显缩小，神经功能缺损明显减轻。中药冲洗组和腹腔给药组相比，两组在脑梗死体积和神经功能缺损评分上有显著差异。结论：(1)经颈内动脉冲洗法减轻急性脑缺血后的脑损伤程度作用要优于腹腔给药法。(2)经颈内动脉冲洗法对于急性脑缺血后的脑损伤保护作用与不同药物有关，祛风通络的中药具有减少急性缺血性大脑损伤程度的作用。     

Oxotremorine-induced cerebral hyperemia does not predict infarction volume in spontaneously hypertensive or stroke-prone rats

OBJECTIVES: We tested the following hypotheses: a) spontaneously hypertensive stroke-prone rats (SHR-SP) have more brain injury than spontaneously hypertensive rats (SHR) and normotensive controls (Wistar-Kyoto rats [WKY]) when exposed to transient focal ischemia; b) infarction size is not correlated with baseline blood pressure; and c) infarction size is inversely related to the cerebral hyperemic response to oxotremorine, a muscarinic agonist that increases cerebral blood flow (CBF) by stimulating endothelial nitric oxide synthase. DESIGN: In vivo study. SETTING: Animal laboratory in a university teaching hospital. SUBJECTS: Adult age-matched male WKY, SHR, and SHR-SP. INTERVENTIONS: Rats were instrumented under halothane anesthesia. Transient focal cerebral ischemia was produced for 2 hrs with the intravascular suture technique. Cerebral perfusion, estimated with laser Doppler flowmetry (LD-CBF), in response to intravenous oxotremorine, was measured in one cohort of rats to estimate endothelial nitric oxide synthase function. Infarction volume was measured at 22 hrs of reperfusion with 2,3,5-triphenyltetrazolium chloride staining. MEASUREMENTS AND MAIN RESULTS: Infarction volume in the striatum of SHR-SP (42+/-4 mm3) was greater than in SHR (29+/-6 mm3) or WKY (1+/-1 mm3) (n = 9 rats/strain). Resting (unanesthetized) mean arterial blood pressure was similar in SHR-SP (177+/-5 mm Hg) and SHR (170+/-5 mm Hg) despite a greater infarction volume in SHR-SP (n = 4) compared with SHR (n = 5). The percentage increase in LD-CBF signal in response to oxotremorine was similar for both groups (SHR, 64%+/-22% [n = 10]; SHR-SP, 69%+/-22% [n = 8]). However, in this cohort, cortical infarction volume was less in SHR (30%+/-4% of ipsilateral cortex) than in SHR-SP (49%+/-2% of ipsilateral cortex). CONCLUSIONS: Although SHR-SP have greater infarction volume than SHR, the mechanism of injury does not appear to be related to a difference in unanesthetized baseline mean arterial blood pressure or to an alteration in endothelium-produced nitric oxide.     

7-硝基吲唑对大鼠脑缺血-再灌注损伤的保护作用

目的　探讨神经元型一氧氮合酶 (nNOS)在脑缺血中的作用。方法　采用栓线法制作大鼠大脑中动脉阻塞再灌注模型 ,观察特异性nNOS抑制剂 7-硝基吲唑 (7-NI)对脑梗死灶、脑水肿和超微结构的影响。结果　与脑缺血 再灌注组相比 ,7-NI能显著减小脑梗死灶和减轻脑水肿 (P <0 0 1,P <0 0 5 ) ,减少脑组织中Na+ 的含量 ,但增加了K+ 的含量 (P <0 0 5 ,P <0 0 5 ) ,并改善缺血神经元的超微结构。结论　nNOS对急性局灶性脑缺血 再灌流损伤起毒性作用 ,nNOS抑制剂有希望用于治疗脑缺血性疾病     

缺血后处理对脑缺血再灌注损伤后ERK1/2和Akt磷酸化及神经细胞凋亡的影响

目的:观察缺血后处理对大鼠局灶性脑缺血再灌注损伤后ERK1/2和Akt及神经细胞凋亡的影响。方法:成年健康SD大鼠72只,随机分为假手术(sham)组、缺血再灌注(I/R)组、缺血后处理(Postcond)组各24只,应用线栓法建立大脑中动脉闭塞(MCAO)再灌注模型。分别于再灌注10min、30min、6h、24h后留取大脑皮质。Western blot检测再灌注10min、30min、6h后ERK1/2和Akt活性变化;原位末端标记(TUNEL)检测再灌注后24h神经细胞凋亡。结果:Postcond组再灌注10min、30min、6h后ERK1/2和Akt活性高于I/R组(P<0.05);脑缺血再灌注24h后,Postcond组与I/R组比较,TUNEL阳性细胞减少(P<0.05)。结论:缺血后处理可提高大鼠脑缺血再灌注后皮质内ERK1/2和Akt活性,减少神经细胞凋亡。     

脑缺血急性期脑细胞脂结合糖和脂结合唾液酸含量的变化

利用大鼠大脑中动脉阻断方法,制备局灶性脑缺血模型;采用鞘糖脂微量分离提取法及地衣酚己糖定量和硫代巴比受酸唾液酸定量方法研究了急性脑缺血期脑细胞脂结合糖及脂结合唾液酸含量的变化。结果发现：在大脑中动脉阻断后3～24h,脂结合糖呈现先降低后升高的变化。与对侧半球相比,大脑中动脉阻断后3、6、12及24h含量的变化率分别为-40.87％、-30.60％、1.52％及35.76％;脂结合唾液酸含量则显著下降,分别较对测半球下降50.15％、38.35％、35.13％及52.27％。结果反映脑缺血损伤急性期神经细胞内源性鞘糖脂代谢有显著异常。     

肌苷对大鼠脑缺血再灌注后血管内皮生长因子表达的影响

背景肌苷参与机体多方面的代谢过程,对缺血性脑损伤具有一定的保护作用,但其机制还没有彻底阐明.目的研究肌苷对大鼠脑缺血再灌注后血管内皮生长因子(vascular endothelial growthfactor,VEGF)表达的影响,探讨肌苷的神经保护作用机制.设计随机对照的实验研究.地点和材料本实验在青岛大学医学院脑血管病研究所和山东省脑血管病防治重点实验室完成.成年健康雌性SD大鼠68只,体质量230～270 g,清洁级,由中国科学院上海实验动物中心提供.干预措施成年健康雌性SD大鼠68只,应用线栓法建立SD大鼠大脑中动脉阻塞(MCAO)再灌注模型,随机分为治疗组32只和对照组32只,每组再随机分为缺血1.5 h再灌流2,6,12,24 h,2,3,7,14 d组(n=4),另外4只作假手术组.应用免疫组织化学方法检测脑缺血再灌流后脑组织VEGF的表达.结果假手术组脑组织未见VEGF阳性表达.对照组在皮层区和纹状体区VEGF在脑缺血再灌注2 h开始表达,12 h达高峰,持续24 h,随即迅速降低.VEGF阳性细胞主要位于Ⅱ,Ⅲ,Ⅳ层神经元和血管内皮细胞,尤其神经细胞核周细胞浆和树突染色最深.肌苷治疗组VEGF表达于缺血再灌注2 h～2 d较对照组显著增高,经统计学处理,差异有非常显著性意义(t=3.78～22.62,P＜0.01).结论肌苷可上调脑缺血再灌注后VEGF的表达,可能是其缺血后神经保护作用的机制之一.     

高渗盐水对脑缺血大鼠Notch信号通路的影响

目的：探讨高渗盐水（ hypertonic saline, HS）对大鼠脑缺血后小胶质细胞Notch信号通路的影响。方法 SPF级-雄性SD大鼠随机（随机数字法）分为假手术组,脑缺血组,生理盐水（ NS）组,10％高渗盐水（10％HS）组。除假手术组外,其他各组采用线栓法复制右侧大脑中动脉栓塞脑缺血模型,缺血2 h后实施再灌注24 h, NS组和10％HS组按0.3 mL／h经尾静脉分别匀速泵入NS和10％HS治疗24 h。然后采用免疫荧光法、 RT-PCR、 Western blot检测各组大鼠脑缺血灶周围Notch1及Notch受体的胞内片段NICD的表达。数据采用单因素方差分析,用LSD法进行组间两两比较,以P＜0.05为差异具有统计学意义。结果免疫荧光表明与假手术组比较,脑缺组和NS组缺血灶周围小胶质细胞Notch1与NICD的表达明显增加;10％HS组与脑缺血组及NS组比较,缺血灶周围小胶质细胞Notch1与NICD的表达明显减少。 RT-PCR表明脑缺血组及NS组与对照组比较, Notch1 mRNA的表达明显增加（对照组：1.000±0.076;脑缺血组：2.203±0.283; NS组：1.616±0.185; P＜0.01）;10％HS组与脑缺血组及NS组比较, Notch1 mRNA的表达均明显减少（脑缺血组：2.203±0.283; NS 组：1.616±0.185; HS 组：1.202±0.177; P ＜0.05）。 Western blot表明脑缺血组和 NS 组与对照组相比,脑缺血灶周围 Notch1蛋白表达明显增加（对照组：0.290±0.079;脑缺血组：0.750±0.029; NS 组：0.765±0.182; P ＜0.01）;10％HS 治疗后, Notch1蛋白表达较脑缺血组和NS组明显减少（脑缺血组：0.750±0.029; NS组：0.765±0.182;HS组：0.390±0.195; P＜0.05）。脑缺血组和NS组与对照组比较,大鼠脑缺血灶周围NICD蛋白表达明显增加（对照组：0.401±0.196;脑缺血组：0.906±0.359; NS组：0.847±0.153; P＜0.01）;10％HS治疗后, NICD蛋白表达较脑缺血组和NS组明显减少（脑缺血组：0.906±0.359;NS组：0.847±0.153; HS组：0.561±0.165; P＜0.05）。结论 HS可抑制大鼠脑缺血后小胶质细胞上Notch信号通路的激活。