Mesenteric lymph reperfusion may exacerbate brain injury in a rat model of superior mesenteric artery occlusion shock

BACKGROUND:The intestinal lymphatic pathway and intestinal ischemia/reperfusion are mainly involved in mesenteric lymph duct ligation or drainage; moreover,intervention by reducing the lymph liquid reflux might relieve lung and other organ dysfunction induced by intestinal ischemia/reperfusion; however,research addressing mesenteric lymph reperfusion (MLR) and brain injury has not yet to be reported.OBJECTIVE:To observe the effect of MLR on brain tissue in a rat model of superior mesenteric artery occlusion (SMAO) shock,and to explore the molecular mechanism of MLR.DESIGN,TIME AND SETTING:A randomized,controlled,animal experiment at a neuro-pathophysiology level was performed at the Institute of Microcirculation,Hebei North University; Department of Pathophysiology,Basic Medical College; Department of Pathology,the First Hospital of Hebei North University between December 2007 and March 2009.MATERIALS:Adenosine triphosphate (ATP) standard was provided by the National Institute for the Control of Pharmaceutical and Biological Products; lactic acid (LA),superoxide dismutase (SOD),malonaldehyde (MDA),nitrogen monoxidum (NO),nitric oxide synthase (NOS),myeloperoxidase (MPO) and ATPase assay kits were provided by Nanjing Jiancheng Bioengineering Institute,China.METHODS:A total of 24 male Wistar rats were randomly divided into four groups.In the sham-surgery group (n = 6),both the mesenteric lymph duct and the superior mesenteric artery were not blocked; in the MLR group (n = 6),the mesenteric lymph duct was occluded for 1 hour followed by 2-hour reperfusion; in the SMAO group (n = 6),the superior mesenteric artery was occluded for 1 hour followed by 2-hour reperfusion; in the MLR + SMAO group (n = 6),both the mesenteric lymph duct and superior mesenteric artery were occluded for 1 hour followed by 2-hour reperfusion.MAIN OUTCOME MEASURES:Mean arterial blood pressure prior to and following ischemia/reperfusion; brain tissue morphology levels of LA,MDA,SOD,NO,NOS,MPO,ATPase and ATP following reperfusion.RESULTS:MLR did not cause changes in mean arterial blood pressure,brain tissue morphology,LA,MDA,NO,ATP,SOD,NOS,MPO and ATPase.However,SMAO caused a rapid decrease and gradual increase of mean arterial blood pressure.Neuronal necrosis,degeneration and swelling were observed in brain tissue.Contents of MDA,NO,LA and ATP as well as activities of NOS and MPO were significantly increased (P< 0.05),but activities of SOD and Na+-K+-ATPase were significantly decreased (P < 0.05).MLR aggravated neuronal damage in a rat model of SMAO shock.Following MLR,mean arterial blood pressure was significantly decreased (P < 0.05),contents of MDA and NO as well as activities of NOS and MPO were significantly increased (P <0.05),but activities of Ca2+-ATPase,Mg2+-ATPase and Ca2+-Mg2+-ATPase as well as ATP content were significantly decreased (P< 0.05).CONCLUSION:MLR aggravates brain injury in a rat model of SMAO shock,which correlates with oxygen-derived free radical injury,NO synthesis and release,sequestration of neutrophilic granulocytes,decreasing activity of cell membrane pumps and energy metabolism dysfunction.Pathogenesis of the intestinal lymphatic pathway should be thoroughly investigated to prevent ischemia/reperfusion injury.     

Manometric artefacts suggesting compression of the duodenum by the superior mesenteric artery in healthy humans

Multi-channel manometry offers the opportunity to study intestinal motor activity with high spatiotemporal resolution. We report tonic and phasic intraluminal pressure changes in the mid-portion of the horizontal part of the duodenum. In 10 healthy volunteers, we recorded 2 h of interdigestive duodenal motility using a water-perfused catheter. The assembly incorporated 12 duodenal sideholes at 1.5-cm intervals (D1-D12). Measurement of the antral and duodenal transmucosal potential difference (TMPD) was used to maintain a correct position of the catheter. The incidence of pressure waves (PWs) increased gradually from proximal (D1) to distal (D12) (P < 0.0001), while the mean amplitude of PWs decreased (P < 0.0001). In eight of 10 subjects, the signals recorded from D9 showed tonic pressure elevations with superimposed phasic pressure changes at heart-rate frequency, comprising 13.8% of total recording time. In the other two subjects, this phenomenon occurred in D8 (9.9% of time). D10 showed a lower incidence of PWs compared with neighbouring sideholes (D6-D9/D11-D12) (P < 0.035), with normal amplitudes. Fluoroscopy was performed in three subjects and showed that D9 was located at the midline. In healthy subjects manometric signals recorded from the horizontal part of the duodenum showed localized artefacts, presumably caused by compression by the superior mesenteric artery. In addition, a 'silent' region was present just distal to this site, the origin of which is uncertain.     

Temporary middle cerebral artery occlusion in the rat: consistent protocol for a model of stroke and reperfusion

The intraluminal suture method of middle cerebral artery occlusion (MCAO) in the rat (the suture model) is a model of stroke which readily lends itself to studying the pathophysiology of post-ischaemic reperfusion. Unfortunately, variability of outcome has compromised the potential of the model, but systematic studies might characterize a consistent protocol. Therefore, the clinical and neuropathological outcome of temporary MCAO and reperfusion in the suture model were systematically investigated. Two hours or 4 h of MCAO were employed, measuring the extent of infarction at 24 h with triphenyltetrazolium chloride or at 72 h with histopathological techniques. Outcome was compared in three rat strains. Following 2 h of MCAO, motor function improved during reperfusion in Sprague-Dawley, but not in Wistar or Fischer-344 rats. All Sprague-Dawley and Wistar rats survived the protocol to 72 h, but 33% of Fischer-344 rats died. The extents of infarction and oedema were greater and less variable in Wistar and Fischer-344 than Sprague-Dawley rats, and in all three strains, the extent of infarction increased with reperfusion time. Following 4 h of MCAO, there was no improvement in motor function during reperfusion in Sprague-Dawley rats, and mortality was high at 24 h in Wistar (33%) and Fischer-344 rats (83%). Outcome was only pursued in Sprague-Dawley rats to 72 h, where the extent of infarction was quite variable. It was concluded that the extent and variability of outcome following temporary MCAO in the suture model is strain-dependent, and a consistent protocol with zero mortality was found in Wistar rats using 2 h of MCAO and 70 h of reperfusion.     

白果内酯抑制持续性脑栓塞大鼠水通道蛋白1，4及胶质纤维酸性蛋白的表达

实验结果发现白果内酯预处理可以降低永久性大脑中动脉闭塞大鼠脑组织含水量和梗死面积;下调水通道蛋白1,4 mRNA在水肿脑组织中的表达,继而抑制其合成,特别是在缺血的早期阶段（8 h）;抑制胶质纤维酸性蛋白的表达,减轻反应性胶质增生。说明白果内酯可通过抑制水通道蛋白的表达减轻脑水肿。     

The role of glycogen synthase kinase 3 beta in brain injury induced by myocardial ischemia/reperfusion injury in a rat model of diabetes mellitus

Myocardial ischemia/reperfusion injury can lead to severe brain injury. Glycogen synthase kinase 3 beta is known to be involved in myo-cardial ischemia/reperfusion injury and diabetes mellitus. However, the precise role of glycogen synthase kinase 3 beta in myocardial ischemia/reperfusion injury-induced brain injury is unclear. In this study, we observed the effects of glycogen synthase kinase 3 beta on brain injury induced by myocardial ischemia/reperfusion injury in diabetic rats. Rat models of diabetes mellitus were generated via intraperitoneal injection of streptozotocin. Models of myocardial ischemia/reperfusion injury were generated by occluding the anterior descending branch of the left coronary artery. Post-conditioning comprised three cycles of ischemia/reperfusion. Immunohistochemical staining and western blot assays demonstrated that after 48 hours of reperfusion, the structure of the brain was seriously damaged in the experimental rats compared with normal controls. Expression of Bax, interleukin-6, interleukin-8, terminal deoxynucleotidyl transferase dUTP nick end labeling, and cleaved caspase-3 in the brain was significantly increased, while expression of Bcl-2, interleukin-10, and phospho-glycogen synthase kinase 3 beta was decreased. Diabetes mellitus can aggravate inflammatory reactions and apoptosis. Ischemic post-conditioning with glycogen synthase kinase 3 beta inhibitor lithium chloride can effectively reverse these changes. Our results showed that myocardial ischemic post-conditioning attenuated myocardial ischemia/reperfusion injury-induced brain injury by activating glyco-gen synthase kinase 3 beta. According to these results, glycogen synthase kinase 3 beta appears to be an important factor in brain injury induced by myocardial ischemia/reperfusion injury.     

线粒体钙单向转运体对脑缺血再灌注大鼠脑水肿的影响

实验探讨线粒体钙单向转运体抑制剂钌红及激动剂精胺对缺血再灌注大鼠脑水肿的影响。采用线栓法建立大鼠左侧大脑中动脉闭塞大鼠模型,缺血再灌注24 h后,脑缺血再灌注模型大鼠、钌红及精胺干预的脑缺血再灌注大鼠神经功能评分均显著低于假手术大鼠,脑组织含水量,水通道蛋白4蛋白表达、IgG渗出含量均显著高于假手术大鼠;与脑缺血再灌注模型大鼠和脑缺血再灌注后精胺干预大鼠比较,钌红干预的脑缺血再灌注大鼠神经功能评分明显升高,脑组织含水量,水通道蛋白4蛋白表达及IgG渗出含量明显减少。提示预防性应用线粒体钙单向转运体抑制剂钌红可显著的降低水通道蛋白4和IgG的表达,影响血脑屏障通透性,进而降低脑水肿的程度。结论 线粒体钙单向转运体可能在大鼠脑缺血再灌注损伤中起重要作用,并能影响AQP4的表达和血脑屏障通透性。     

Puerarin protects brain tissue against cerebral ischemia/reperfusion injury by inhibiting the inflammatory response

Puerarin, a traditional Chinese medicine, exerts a powerful neuroprotective effect in cerebral ischemia/reperfusion injury, but its mechanism is unknown. Here, we established rat models of middle cerebral artery ischemia/reperfusion injury using the suture method. Puerarin(100 mg/kg) was administered intraperitoneally 30 minutes before middle cerebral artery occlusion and 8 hours after reperfusion. Twenty-four hours after reperfusion, we found that puerarin significantly improved neurological deficit, reduced infarct size and brain water content, and notably diminished the expression of Toll-like receptor-4, myeloid differentiation factor 88, nuclear factor kappa B and tumor necrosis factor-α in the ischemic region. These data indicate that puerarin exerts an anti-inflammatory protective effect on brain tissue with ischemia/reperfusion damage by downregulating the expression of multiple inflammatory factors.     

高压氧治疗对家兔实验性大脑中动脉闭塞后梗死灶周围巢蛋白表达的影响

目的 探讨高压氧(HBO)治疗对兔大脑中动脉闭塞(MCAO)模型梗死灶周围巢蛋白(nestin)表达的影响.方法 制备兔右侧MCAO模型.将动物分为3组,即MCAO+HBO治疗组(30只):MCAO后立即行HBO治疗,每天1次,连续20 d;MCAO对照组(30只):MCAO后不行HBO治疗;假手术组(10只):仅开颅不闭塞血管.分别于MCAO后10d、20 d观察其行为学、梗死体积变化.并用免疫组化法检测距梗死灶中心3 mm内皮层脑组织内nestin的表达.结果 MCAO后.MCAO+HBO治疗组行为学评分高于MCAO对照组,差异有统计学意义(P<0.05);MCAO+HBO治疗组脑梗死体积小于MCAO对照组,差异有统计学意义(P<0.05);MACO后10 d、20 dMCAO+HBO治疗组nestin免疫阳性细胞数05.88±1.2、20.03±1.6)高于MCAO对照组(6.63±1.6、6.82±0.8),差异有统计学意义(P<0.05).结论 早期、连续20 d HBO治疗对兔MCAO后的神经行为学、梗死体积有明显的治疗效果,并促进兔MCAO后梗死灶周围脑组织内nestin的表达.     

Differential protein expression in spinal cord tissue of a rabbit model of spinal cord ischemia/reperfusion injury

New Zealand rabbits were randomly divided into an ischemia group(occlusion of the abdominal aorta for 60 minutes),an ischemia-reperfusion group(occlusion of the abdominal aorta for 60 minutes followed by 48 hours of reperfusion) and a sham-surgery group.Two-dimensional gel electrophoresis detected 49 differentially expressed proteins in spinal cord tissue from the ischemia and ischemia/reperfusion groups and 23 of them were identified by mass spectrometry.In the ischemia group,the expression of eight proteins was up regulated,and that of the remaining four proteins was down regulated.In the ischemia/reperfusion group,the expression of four proteins was up regulated,and that of two proteins was down regulated.In the sham-surgery group,only one protein was detected.In the ischemia and ischemia/reperfusion groups,four proteins overlapped between groups with the same differential expression,including three that were up regulated and one down regulated.These proteins were related to energy metabolism,cell defense,inflammatory mechanism and cell signaling.     

Hawthorn extract reduces infarct volume and improves neurological score by reducing oxidative stress in rat brain following middle cerebral artery occlusion

In our present investigation the neuroprotective effect of alcoholic extract of Hawthorn (Crataegus oxycantha) was evaluated against middle cerebral artery occlusion induced ischemia/reperfusion injury in rats. Male Sprague–Dawley rats were pretreated with 100 mg/kg body weight of the extract by oral gavage for 15 days. The middle cerebral artery was then occluded for 75 min followed by 24 h of reperfusion. The pretreated rats showed significantly improved neurological behavior with reduced brain infarct when compared to vehicle control rats. The glutathione level in brain was found to be significantly (p < 0.05) low in vehicle control rats after 24 h of reperfusion when compared to sham operated animals. However, in Hawthorn extract pretreated rats the levels were found to be close to that of sham. Malondialdehyde levels in brain of sham and pretreated group were found to be significantly lower than the non-treated vehicle group (p < 0.05). The nitric oxide levels in brain were measured and found to be significantly (p < 0.05) higher in vehicle than in sham or extract treated rats.     

Middle cerebral artery occlusion in the rat: consistent protocol for a model of stroke

Variability in the effects of the intraluminal suture method of middle cerebral artery occlusion (MCAO) in the rat has been a common and disadvantageous finding. Therefore, we systematically investigated the effects of suture type and rat strain on outcome. First, the clinical and neuropathological effects of permanent MCAO with either an uncoated or a silicone-coated nylon suture were studied over 7  days in Sprague–Dawley rats ( n =36 for each type of suture). Outcome was less severe with the uncoated compared with the silicone-coated suture (e.g. total cerebral infarct volume at 24  h before any fatalities was 119.9±79.8  mm³ , cf. 183.0±36.5 mm³ , n =12 for each, P <0.05; and overall mortality rate was 12.5% cf. 33%, respectively), but much more variable (coefficient of variation was 66.6% cf. 19.9%, respectively). Second, being more consistent in its effects, the silicone-coated suture was further studied in Wistar and Fischer-344 rats ( n =12 for each). Seventy-five per cent of the Wistar's died prematurely from gross hemispheric oedema. Motor deficit and extent of infarction in the Fischer-344 rats were both significantly greater compared with Sprague-Dawley rats (e.g. total cerebral infarct volume at 24  h in the former was 253.6±25.4  mm³ , n =11, P <0.05), and more consistent (coefficient of variation was only 10.0%). It was concluded that the silicone-coated suture and the Fischer-344 rats strain produced the most consistent results and their novel combination provides a reliable acute stroke model.     

The brain metabolic activity after resuscitation with liposome-encapsulated hemoglobin in a rat model of hypovolemic shock

We examined the effect of resuscitation with liposome-encapsulated hemoglobin (LEH) on cerebral bioenergetics in a rat model of 45% hypovolemia. The rats were resuscitated with isovolemic LEH or saline after 15 minutes of shock and followed up to 6 hours. Untreated hypovolemic rats received no fluid. The cerebral uptake of F-18-fluorodeoxyglucose (FDG) was measured by PET, and at 6 hours, the brain was collected for various assays. Hypovolemia decreased cellular adenosine triphosphate (ATP), phosphocreatine, nicotinamide adenine dinucleotide (NAD)/NADH ratio, citrate synthase activity, glucose-6-phosphate, and nerve growth factor (NGF), even when FDG uptake remained unchanged. The FDG uptake was reduced by saline, but not by LEH infusion. The reduced FDG uptake in saline group was associated with a decrease in hexokinase I expression. The LEH infusion effectively restored ATP content, NAD/NADH ratio, and NGF expression, and reduced the hypovolemia-induced accumulation of pyruvate and ubiquitinated proteins; in comparison, saline was significantly less effective. The LEH infusion was associated with low pH and high anion gap, indicating anionic gap acidosis. The results suggest that hypovolemic shock perturbs glucose metabolism at the level of pyruvate utilization, resulting in deranged cerebral energy stores. The correction of volume and oxygen deficits by LEH recovers the cerebral metabolism and creates a prosurvival phenotype.     

The expression and activity of brain lipoprotein lipase is increased after acute cerebral ischemia‐reperfusion in rats

Lipoprotein lipase (LPL) is a key enzyme involved in lipid metabolism. Previous studies have shown that the levels of brain LPL mRNA, protein and activity are up‐regulated after brain and nerve injury. The aim of this study was to determine the response of expression and activity of brain LPL following acute cerebral ischemia‐reperfusion. Adult male Sprague‐Dawley rats were subjected to surgical occlusion of the middle cerebral artery. The expression of brain LPL was assessed by immunohistochemical staining and the enzyme activity of brain LPL was evaluated by colorimetric method. Increase of LPL immunopositive cells in the cerebral cortex around the infarction area was observed at 4, 6, 12 h ischemia, 2 h ischemia 2 h reperfusion, and 4 h ischemia 2 h reperfusion. LPL activity was significantly decreased in the ischemic side cortex at 2 h ischemia, and then significantly increased at 4 and 6 h ischemia. Our results showed that LPL immunopositive cells were increased in the cortex around the infarction area, and activity of LPL first decreased and then increased following acute cerebral ischemia‐reperfusion. These results may suggest that LPL plays a potential role in the pathophysiological response of the brain to cerebral ischemia‐reperfusion.     

Neuroprotective effects of willed-movement therapy in a rat model of cerebral ischemia/reperfusion

The present study established rat models of middle cerebral artery ischemia/reperfusion using the thread method.Rats performed willed-movement(climbing a ladder or wall in a box) when induced by food and water.Results revealed that Longa scores of neurological deficits significantly de-creased in the willed-movement group at 15 days after reperfusion,while expression of glial fibrillary acidic protein,neurotrophic factor-3,and growth-associated protein-43 significantly increased at 7 and 15 days after reper...     

Influence of Tanshinone IIa on heat shock protein 70, Bcl-2 and Bax expression in rats with spinal ischemia/reperfusion injury

Tanshinone lla is an effective monomer component of Danshen, which is a traditional Chinese medicine for activating blood circulation to dissipate blood stasis. Tanshinone lla can effectively improve brain tissue ischemia/hypoxia injury. The present study established a rat model of spinal cord ischemia/reperfusion injury and intraperitoneally injected Tanshinone IIa, 0.5 hour prior to model establishment. Results showed that Tanshinone lla promoted heat shock protein 70 and Bcl-2 protein expression, but inhibited Bax protein expression in the injured spinal cord after ischemia/reperfusion injury. Furthermore, Nissl staining indicated a reduction in nerve cell apoptosis and fewer pathological lesions in the presence of Tanshinone lla, compared with positive control Danshen injection.     

Neuronal injury and brain-derived neurotrophic factor expression in a rat model of amygdala kindling seizures Differences in brain regions and kindling courses

BACKGROUND:Studies have demonstrated that brain-derived neurotrophic factor (BDNF) has a dual effect on epilepsy. However, the relationship between epilepsy-induced brain injury and BDNF remains poorly understood.OBJECTIVE:According to ultrastructural and molecular parameters, to detect the degree of neuronal injury and BDNF expression changes at different brain regions and different kindling times to determine the effects of BDNF on epilepsy-induced brain injury.DESIGN, TIME AND SETTING:A randomized, controlled, animal experiment based on neuropathology and molecular biology was performed at the Department of Physiology and Department of Pathology, Basic Medical College of Jilin University in 2003.MATERIALS:UltraSensitiveTM SP kit for immunohistochemistry (Fuzhou Maxim Biotechnology, China), BDNF antibody (concentrated type, Wuhan Boster Biological Technology, China), JEM-1000SX transmission electron microscopy (JEOL, Japan), and BH-2 light microscope (Olympus, Japan) were used in the present study.METHODS:Wistar rats were randomly assigned to control (n = 6), sham-surgery (n = 6), and model (n = 60) groups. The control group rats were not treated; an electrode was embedded into the amygdala in rats from the sham-surgery and model groups; an amygdala kindling epilepsy model was established in the model group.MAIN OUTCOME MEASURES:Pathological changes in the temporal lobe and hippocampus were observed by light and electron microscopy at 1, 3, 7, 14, and 21 days following kindling, and BDNF expression in the various brain regions was determined by immunohistochemistry.RESULTS:In the model group, temporal lobe cortical and hippocampal neurons were swollen and the nuclei were laterally deviated. There were also some apoptotic neurons 3 days after kindling. The nucleoli disappeared and the nuclei appeared broken or lysed, as well as slight microglia hyperplasia, at 7 days. Electron microscopic observation displayed chromatin aggregation in the nuclei and slight mitochondrion swelling 3 days after kindling. Injury changes were aggravated at 7 days, characterized by broken cytoplasmic membrane and pyknosis. With the development of seizure, the number of BDNF-positive neurons in the hippocampus and temporal lobe increased and peaked at 7 days. Moreover, hippocampal and cortical temporal lobe injury continued. Following termination of electrical stimulation after 7 days of kindling, BDNF expression decreased, but continued to be expressed, up to 21 days of kindling. In addition, the number of temporal and hippocampal BDNF-positive neurons was greater than the control group.CONCLUSION:Brain injury and BDNF expression peaked at 7 days after kindling, and hippocampal changes were significant.     

Effect of Fujian tablet on the expression of Nogo-A mRNA in the cervical spinal cord of middle cerebral artery occlusion model rats

BACKGROUND: Inhibiting the expression of Nogo-A in cervical spinal cord by use of interaction of antigen and antibody can help the remodeling of corticospinal projection of focal cerebral ischemia model rats to facilitate neurological recovery, which provides a new possible mechanism for drugs to promote neurological recovery. However, the effects of drugs on the expression of Nogo-A in cervical spinal cord are still unclear. OBJECTIVE: To observe the effect of Fujian tablet on the expression of Nogo-A mRNA in cervical spinal cords of middle cerebral artery occlusion (MCAO) rats, and to investigate the possible regulatory effect of Fujian tablet on the regenerated microenvironment of spinal conduction bundle. DESIGN: A randomized and controlled trial taking Wistar rats as experimental animals. SETTING: Department of Neurology, Affiliated Hospital of Shandong University of Traditional Chinese Medicine. MATERIALS: This experiment was carried out in the laboratory of Shandong Academy of Medical Science between June 2005 and July 2006. A total of 40 healthy male Wistar rats, aged 12 weeks, weighing 250–300 g, were provided by the Experimental Animal Center of Shandong University. Fujian tablets (main components: Heshouwu, Yinyanghuo, etc) were provided by office of Pharmaceutics of Shandong University of traditional Chinese medicine. Nogo-A detection kit was provided by Wuhan Boster Biotechnology Co.,Ltd., and batch number was 040309009. This experiment was approved by Local Animal Ethics Committee. METHODS: Forty male rats were randomly divided into 4 groups, with 10 in each: normal group, sham-operation group, model group and administration group. Rats in the administration group and model group were subjected to MCAO. Rats in the sham-operation group underwent the same craniotomy, and their middle cerebral arteries (MCA) were not occluded. Rats in the normal group were untouched. Rats in administration group were intragastrically administrated with the solution of Fujian tablet at a dose of 9 g/kg and others were given equal dosage of normal saline two days later. The treatments were done once a day and the course totaled 2 weeks. MAIN OUTCOME MEASURES: The expression of Nogo-A mRNA in slices of cervical spinal cords. RESULTS: Forty rats were involved in the final analysis. The expression of Nogo-A mRNA in the cervical spinal cord of rats in the administration group and model group was significantly decreased as compared with that in the normal group (P < 0.01 and P < 0.05, respectively). The expression of Nogo-A mRNA in the administration group was also significantly weaker than that in the model group（P < 0.05）. CONCLUSION: Fujian tablet can inhibit the expression of Nogo-A mRNA in cervical spinal cords of MCAO rats, which facilitates regeneration and remodeling of cervical spinal cords.