Intercellular adhesion molecule-1 expression in the hippocampal CA1 region of hyperlipidemic rats with chronic cerebral ischemia

Chronic cerebral ischemia is a pathological process in many cerebrovascular diseases and it is induced by long-term hyperlipidemia,hypertension and diabetes mellitus.After being fed a high-fat diet for 4 weeks,rats were subjected to permanent occlusion of bilateral common carotid arteries to establish rat models of chronic cerebral ischemia with hyperlipidemia.Intercellular adhesion molecule-1 expression in rat hippocampal CA1 region was determined to better understand the mechanism underlying the effects of hyperlipidemia on chronic cerebral ischemia.Water maze test results showed that the cognitive function of rats with hyperlipidemia or chronic cerebral ischemia,particularly in rats with hyperlipidemia combined with chronic cerebral ischemia,gradually decreased between 1 and 4 months after occlusion of the bilateral common carotid arteries.This correlated with pathological changes in the hippocampal CA1 region as detected by hematoxylin-eosin staining.Immunohistochemical staining showed that intercellular adhesion molecule-1 expression in the hippocampal CA1 region was noticeably increased in rats with hyperlipidemia or chronic cerebral ischemia,in particular in rats with hyperlipidemia combined with chronic cerebral ischemia.These findings suggest that hyperlipidemia aggravates chronic cerebral ischemia-induced neurological damage and cognitive impairment in the rat hippocampal CA1 region,which may be mediated,at least in part,by up-regulated expression of intercellular adhesion molecule-1.     

Expression of brain nuclear factor-κB (P50) in mice with traumatic brain injury after hyperbaric oxygen treatment

Objective To investigate the relation between the therapeutic effect of hyperbaric oxygen treatment and nuclear factor-κB (NF-κB) (P50) expression in the brain tissue in mice with traumatic brain injury (TBI). Methods A total of 120 SD rats were randomly divided into sham-operated, TBI model, normobaric oxygen and hyperbaric oxygen (0.2 MPa) groups, and in the latter 3 groups, TBI was induced using Feeney's method. At 6 h and 1, 3, 5, and 7 d following TBI (6 rats at each time point), the rats were sacrificed to observe the pathological changes in the brain tissues under light microscope and detect the expression of NF-κB (P50) using immunohistochemistry. Results The rats in hyperbaric oxygen group showed lessened brain edema as compared with those in TBI model and normobaric oxygen groups. Only trace amount of NF-κB (P50) expression was observed in the sham-operated group, while in the 3 groups with TBI, NF-κB (P50) expression began to increase as early as 6 h after TBI and kept increasing till reaching the peak level at 5 days. At each of the time points for observation, the expression ofNF-κB (PS0) in hyperbaric oxygen group was significantly higher than those in TBI and normobaric oxygen groups (P<0.05). Conclusion Hyperbaric oxygen treatment offers protection of the injured neural cells and promotes their repair in rats following TBI. Increased NF-κB (P50) expression in the brain tissue may serve as one of the pathways mediating the neuroprotective effect of hyperbaric oxygen treatment.     

电针治疗脑缺血：促胰岛素样生长因子1表达的作用

Background Growth factors such as basic fibroblast growth factor (bFGF) have shown neuroprotective effects on cerebral ischemia. Electroacupuncture (EA) could be a potential treatment for cerebral ischemia because it increases local cerebral blood flow and up-regulates the expression levels of bFGF. This study aimed to investigate the neuroprotective effects of EA against cerebral ischemia as well its effect on insulin-like growth factor-1 (IGF-1) expression following middle cerebral artery occlusion (MCAO) in rats. Methods This study compared EA with sham in a sample of 41 male Sprague Dawley rats, weighting between 280-300 g. The animals were randomized into two groups: 12 in the cDNA microarray study and 29 in the immunohistochemical and in situ hybrization study. In the histochemical and in situ hybrization study: normal control (n=1), sham (n=4), Ischemia Ⅰ (9h reperfusion, n=6), Ischemia Ⅰ+EA (9h reperfusion, n=6), Ischemia Ⅱ (24h reperfusion, n=6), Ischemia Ⅱ+EA (24h reperfusion, n=6). Overall changes of gene expression were analyzed with cDNA microarray, and IGF-1mRNA expression was confirmed by in situ hybridization, then the IGF-1 protein expression was detected by immunohistochemistry. EA was given 15 minutes after MCA occlusion and lasted for 1 hour between the Baihui acupoint (GV. 20) and the Renzhong acupoint (GV. 26) using the dense-sparse waveforms, which can be transformed into each other when a dense or sparse wave is terminated. Results After MCA occlusion, IGF-1mRNA and protein expression levels decreased. EA attenuated brain edema, decreased the infarct volume, and led to the upregulation of the decreased IGF-1mRNA and protein expression in the striatum. Conclusions EA is effective to extenuate cerebral ischemia and up-regulate endogenous IGF-1 expression following MCA occlusion, which might be an important mechanism for the neuroprotective effects of EA against cerebral ischemia.     

Expression profiles of microRNAs after focal cerebral ischemia/reperfusion injury in rats

Rat models of focal cerebral ischemia/reperfusion injury were established by occlusion of the middle cerebral artery.Microarray analysis showed that 24 hours after cerebral ischemia,there were nine up-regulated and 27 down-regulated microRNA genes in cortical tissue.Bioinformatic analysis showed that bcl-2 was the target gene of microRNA-384-5p and microRNA-494,and caspase-3 was the target gene of microRNA-129,microRNA-320 and microRNA-326.Real-time PCR and western blot analyses showed that 24 hours after cerebral ischemia,bcl-2 mRNA and protein levels in brain tissue were significantly decreased,while caspase-3 mRNA and protein levels were significantly increased.This suggests that following cerebral ischemia,differentially expressed microRNA-384-5p,microRNA-494,microRNA-320,microRNA-129 and microRNA-326 can regulate bcl-2 and caspase-3 expression in brain tissue.     

microRNA-455-5p alleviates neuroinflammation in cerebral ischemia/reperfusion injury

Neuroinflammation is a major pathophysiological factor that results in the development of brain injury after cerebral ischemia/reperfusion.Downregulation of microRNA(miR)-455-5p after ischemic stroke has been considered a potential biomarker and therapeutic target for neuronal injury after ischemia.However,the role of miR-455-5p in the post-ischemia/reperfusion inflammatory response and the underlying mechanism have not been evaluated.In this study,mouse models of cerebral ischemia/reperfusion injury were established by transient occlusion of the middle cerebral artery for 1 hour followed by reperfusion.Agomir-455-5p,antagomir-455-5p,and their negative controls were injected intracerebroventricularly 2 hours before or 0 and 1 hour after middle cerebral artery occlusion(MCAO).The results showed that cerebral ischemia/reperfusion decreased miR-455-5p expression in the brain tissue and the peripheral blood.Agomir-455-5p pretreatment increased miR-455-5p expression in the brain tissue,reduced the cerebral infarct volume,and improved neurological function.Furthermore,primary cultured microglia were exposed to oxygen-glucose deprivation for 3 hours followed by 21 hours of reoxygenation to mimic cerebral ischemia/reperfusion.miR-455-5p reduced C-C chemokine receptor type 5 mRNA and protein levels,inhibited microglia activation,and reduced the production of the inflammatory factors tumor necrosis factor-αand interleukin-1β.These results suggest that miR-455-5p is a potential biomarker and therapeutic target for the treatment of cerebral ischemia/reperfusion injury and that it alleviates cerebral ischemia/reperfusion injury by inhibiting C-C chemokine receptor type 5 expression and reducing the neuroinflammatory response.     

Hypoxic preconditioning reduces NLRP3 inflammasome expression and protects against cerebral ischemia/reperfusion injury

Hypoxic preconditioning can protect against cerebral ischemia/reperfusion injury. However, the underlying mechanisms that mediate this effect are not completely clear. In this study, mice were pretreated with continuous, intermittent hypoxic preconditioning;1 hour later, cerebral ischemia/reperfusion models were generated by middle cerebral artery occlusion and reperfusion. Compared with control mice, mice with cerebral ischemia/reperfusion injury showed increased Bederson neurological function scores, significantly increased cerebral infarction volume, obvious pathological damage to the hippocampus, significantly increased apoptosis;upregulated interleukin-1β, interleukin-6, and interleukin-8 levels in brain tissue;and increased expression levels of NOD-like receptor family pyrin domain containing 3(NLRP3), NLRP inflammasome-related protein caspase-1, and gasdermin D. However, hypoxic preconditioning significantly inhibited the above phenomena. Taken together, these data suggest that hypoxic preconditioning mitigates cerebral ischemia/reperfusion injury in mice by reducing NLRP3 inflammasome expression. This study was approved by the Medical Ethics Committee of the Fourth Hospital of Baotou, China(approval No. DWLL2019001) in November 2019.     

姜黄素降低短暂性脑缺血再灌注损伤的炎症反应

Inflammatory reactions are important pathophysiological mechanisms of ischemic brain injury.The present study analyzed the anti-inflammatory characteristics of curcumin via myeloperoxidase activity and nitric oxide content after 2-hour ischemia/24-hour reperfusion in Sprague Dawley rats.In addition,expressions of nuclear factor kappa B,tumor necrosis factor-α and interleukin-1β protein were measured.Curcumin significantly reduced myeloperoxidase and nitric oxide synthase activities and suppressed expressions of nuclear factor kappa B,tumor necrosis factor-α,and interleukin-1β in ischemia/reperfusion brain tissue.Results suggested that the neuroprotective effect of curcumin following cerebral ischemia/reperfusion injury could be associated with inhibition of inflammatory reactions.     

Neuroprotective effects of daidzein on focal cerebral ischemia injury in rats

Daidzein, a plant extract, has antioxidant activity. It is hypothesized, in this study, that daidzein exhibits neuroprotective effects on cerebral ischemia. Rat models of middle cerebral artery occlusion were intraperitoneally administered daidzein. Biochemical and immunohistochemical tests showed that superoxide dismutase and nuclear respiratory factor 1 expression levels in the brain tissue decreased after ischemia and they increased obviously after daidzein administration; malondialdehyde level and apoptosis-related cysteine peptidase caspase-3 and caspase-9 immunoreactivity in the brain tissue increased after ischemia and they decreased obviously after daidzein administration. Hematoxylin-eosin staining and luxol fast blue staining results showed that intraperitoneal administration of daidzein markedly alleviated neuronal damage in the ischemic brain tissue. These findings suggest that daidzein exhibits neuroprotective effects on ischemic brain tissue by decreasing oxygen free radical production, which validates the aforementioned hypothesis.     

大鼠脑微血管内皮细胞氧糖剥夺模型的构建及黄体酮的保护作用

目的：体外研究氧糖剥夺对大鼠脑微血管内皮细胞的损伤机制及黄体酮的保护作用。方法体外培养大鼠脑微血管内皮细胞（brain microvascular endothelial cell ,BMEC）,并构建氧糖剥夺（oxygen and glucose deprivation ,OGD）模型。流式细胞技术检测BMEC在OGD后细胞间黏附分子-1（intercellular adhesion molecule-1,ICAM-1）、血管细胞黏附分子-1（vascular cell adhesion molecule-1,VCAM-1）的表达情况。利用Western blot方法检测大鼠BMEC氧糖剥夺后核转录因子-B （nuclear factor-kappa B ,NF-κB）p65亚基含量,染色质免疫沉淀分析（Chromatin immunoprecipitation assay kit ,ChIP）技术检测NF-κB的DNA结合活性。结果 OGD后6 h可诱导BMEC的ICAM-1、VCAM-1和NF-κB表达上调,黄体酮对此有明显抑制作用。氧糖剥夺能显著提高NF-κB的DNA结合活性。黄体酮对此有明显抑制作用。结论 OGD可通过活化NF-κB、上调ICAM-1和VCAM-1的表达、黄体酮能减少OGD对BMEC造成的损伤。     

Resatorvid protects against hypoxic-ischemic brain damage in neonatal rats

Secondary brain damage caused by hyperactivation of autophagy and inflammatory responses in neurons plays an important role in hypoxic-ischemic brain damage(HIBD).Although previous studies have implicated Toll-like receptor 4(TLR4)and nuclear factor kappa-B(NF-κB)in the neuroinflammatory response elicited by brain injury,the role and mechanisms of the TLR4-mediated autophagy signaling pathway in neonatal HIBD are still unclear.We hypothesized that this pathway can regulate brain damage by modulating neuron autophagy and neuroinflammation in neonatal rats with HIBD.Hence,we established a neonatal HIBD rat model using the Rice-Vannucci method,and injected 0.75,1.5,or 3 mg/kg of the TLR4 inhibitor resatorvid(TAK-242)30 minutes after hypoxic ischemia.Our results indicate that administering TAK-242 to neonatal rats after HIBD could significantly reduce the infarct volume and the extent of cerebral edema,alleviate neuronal damage and neurobehavioral impairment,and decrease the expression levels of TLR4,phospho-NF-κB p65,Beclin-1,microtubule-associated protein l light chain 3,tumor necrosis factor-α,and interleukin-1βin the hippocampus.Thus,TAK-242 appears to exert a neuroprotective effect after HIBD by inhibiting activation of autophagy and the release of inflammatory cytokines via inhibition of the TLR4/NF-κB signaling pathway.This study was approved by the Laboratory Animal Ethics Committee of Affiliated Hospital of Yangzhou University,China(approval No.20180114-15)on January 14,2018.     

Mild hypothermia effects on intercellular adhesion molecule-1 and serum interleukin-6 expression in brain tissues of a rat focal ischemia model

BACKGROUND： Previous studies have confirmed the neuroprotective effect of mild hypothermia on ischemic brain injury. OBJECTIVE： To investigate the effects of mild hypothermia on intercellular adhesion molecule-1 expression and serum interleukin-6 levels in ischemic brain tissues of focal brain ischemia rats, and to explore the neuroprotective effects of mild hypothermia on ischemic brain injury. DESIGN, TIME AND SETTING： A randomized, controlled, neurobiological experiment was performed at the Central Laboratory, First Affiliated Hospital, Xinxiang Medical College, China from February to July 2006. MATERIALS： Thirty healthy, adult, Sprague Dawley rats were used to establish middle cerebral artery occlusion models using the suture method, The immunohistochemistry （streptavidin-biotin-peroxidase complex method） kit was purchased from Boster, China. Interleukin-6 radioimmunoassay was supplied by Institute of Radioimmunity, Technology Development Center, General Hospital of Chinese PLA. METHODS： The rats were equally and randomly assigned into mild hypothermia and control groups, and middle cerebral artery occlusion models were established. The rectal temperature was maintained at （37 ±0.5）℃ in the control group. In the mild hypothermia group, the rectal temperature was maintained at （33±1）℃. MAIN OUTCOME MEASURES： At 12 hours after model establishment, the ischemic brain hemispheres were coronally sliced at the level of the optic chiasm. The number of intercellular adhesion molecule-1-positive vessels per high-power field was observed with an optical microscope. Serum interleukin-6 levels were measured by radioimmunoassay. RESULTS： Compared with the control group, intercellular adhesion molecule-1 and serum interleukin-6 expressions were significantly decreased in ischemic brain tissues of the mild hypothermia group （P 〈 0.01）. CONCLUSION： Mild hypothermia exhibits a neuroprotective effect by reducing serum interleukin-6 and intercellular adhesion molecule-1 expression followi     

黄体酮对脑梗死大鼠血脑屏障保护作用的机制研究

Objective To investigate the inhibitory effects of progesterone on the inflammatory response and its influence on the structure of the blood brain barrier (BBB) of rats after cerebral infarction. Methods Progesterone was injected intrapefitoneally following permanent middle cerebral artery occlusion (MCAO) in rats. Western blotting was used to detect the expressions of tumor necrosis factor-alpha (TNF-α) and claudin-5, the major component of tight-junction proteins associated with the BBB. The water content in the injured brain tissue was also examined. Results The expression level of TNF-α increased but claudin-5 expression decreased in the brain tissue of all the rats following MCAO. Progesterone treatment significantly reduced the TNF-α expression in comparison with the vehicle-treated rats, resulting also in ameliorated claudin-5 protein loss and reduced water content in the injured brain tissue. Conclusion Progesterone inhibits the inflammatory response and mitigates the brain damage after cerebral infarction in rats, suggesting the role of this steroid as a neuroprotective agent.     

NF-κB、ICAM-1在实验性SAH后基底动脉壁上的表达变化

目的 探讨免疫炎症反应在蛛网膜下腔出血(subarachnoid hemorrhage,SAH)后脑血管痉挛(cerebral vasospasm,CVS)发病机制中的作用.方法 应用新西兰家兔经枕大池二次注血制成SAH动物模型.在各相应时间点运用数字减影血管造影(digital subtraction angiography,DSA)、HE染色和免疫组化等方法 ,观察兔基底动脉管壁形态以及核转录因子kappaB(nuclear factor-kappaB,NF-κB)、细胞间粘附因子-1(intercellular adhesion molecule,ICAM-1)在管壁上表达的动态变化.结果 在SAH后第3天基底动脉血管腔已开始狭窄,第7天达到高峰,之后狭窄渐减轻,14d时基本接近正常.SAH后3d NF-κB表达增加,5～7d表达持续在一个较高的水平,随后开始下降;ICAM-1表达变化趋势与NF-κB相似,但在表达高峰时相上稍迟于后者.结论 NF-κB、ICAM-1在血管壁上的表达变化与CVS的发生时相密切相关,提示由其参与介导的血管壁炎症反应可能在CVS的发生和发展过程中起了重要的作用.

Abstract：

Objective To evaluate the possible effect of inflammatory reaction in the pathogenesis of CVS after SAH.Methods The "double-hemorrhage" model of vasospasm Was performed by two injections of autologous aterial blood into the cisterna magna of Newzealand rabbits.We oberserved morphological changes and the rule of expression and activity of NF-κB and ICAM-1 in basilar artery(BA)walls by DSA,light microscope and immunohistochemistry.Results The "double-hemorrhage"model of vasospasm was succeeded to be made.The BA vasospasm Was obvious on 3d after autologous aterial blood wa8 first injected into the cisterna magna,and peaked on 7d,and then relieved.On 14d,there was almost no abnormality in the arterial wall.Upregulation of NF-κB expression started on 3d,became stronger on 5d to 7d,and then decreased.The upregulation of ICAM-1 expression in the BA walls Was similar with NF-κB,but was a bit later to tively correlated with the vasospasm.Inflammatory reaction in arterial walls mediated by NF-8κB may play a key mle in the process of CVS after SAH.     

Interleukin-10 inhibits the expression of nuclear factor kappa B in rats with cerebral ischemia

BACKGROUND: Plenty of studies have demonstrated that inflammatory reaction is involved in ischemic cerebral damage, and the expression of inflammatory cytokines can be observed at the initial sites of cerebral damage at early period, including interleukin-6, interleukin-8, etc., which are all the target gene products of nuclear factor kappa B (NF-κB). The process of ischemic damage can be affected by adjusting and controlling NF-κB activity via multi-links. OBJECTIVE: To investigate the inhibitory effect of interleukin-10 on the expression of NF-κB in the ischemic sites of rats with focal cerebral ischemia in rats and its molecular mechanisms. DESIGN: A randomized and controlled animal trial. SETTING: Department of Neurology, the Affiliated Union Hospital of Fujian Medical University. MATERIALS: Thirty-two adult male Sprague-Dawley rats weighing (250±30) g were used. NF-κB p65 (RelA) rabbit anti-rat monoclonal primary antibody was the product of Neomarkers Company; Immunohistochemical kit of the SP two-step method was purchased from Beijing Zhongshan Biotechnology Co., Ltd. METHODS: The experiment was carried out in the Affiliated Union Hospital of Fujian Medical University from August 2005 to April 2006. The rats were randomly assigned into sham-operated group, middle cerebral artery occlusion (MCAO) group, vehicle-treated group and interleukin-10 treated group, 8 rats in each group. Focal cerebral ischemia was induced by occlusion of the middle cerebral artery as previously described. Rats in the MCAO group were anesthetized intraperitoneally, thyroid was bluntly dissected. Right common, external and internal carotid arteries were isolated, the trunk of external carotid artery was ligated and freed, an artery clamp was placed at the internal carotid artery, then a "V" shape incision was made at the free section of external carotid artery, filament was inserted for a depth of (18.5±0.5) mm. The rats in the sham-operated group were given the same treatments with the exception of filament insertion. After the successful model establishment for 1 hour, the rats in the interleukin-10 treated group were injected with human recombinant interleukin-10 (1 μg) via lateral ventricle, whereas those in the vehicle-treated group were injected with 5 mol/L NaP (5 μL). The rectal temperature of rats were kept at about 37 ℃ with heating lamps throughout the operation. Twenty-four hours after MCAO, the rats were examined for neurological deficits. Only those animals that scored at 1-3 points were utilized. The rats were decapitated at 24 hours postoperatively. The expression of NF-κB p65 in peri-infarct core was detected immunohistochemically. The percentage of NF-κB p65 subunit positive cells in 1 000 cells was calculated. MAIN OUTCOME MEASURES: Expression of NF-κB p65 in peri-infarct core; Percentage of NF-κB p65 subunit positive cells. RESULTS: All the 32 rats were involved in the analysis of results. NF-κB p65 expressed in cytoplasm and some nuclei. It was expressed all in cytoplasm in the sham-operated group, and partly expressed in the nucleus after cerebral ischemia. Small amounts of NF-κB p65 positive neurocytes were observed in the sham-operated group[(3.7±0.6)%], those were obviously increased in the MCAO group [(15.4±3.7)%, P < 0.01]. NF-κB p65 positive neurocytes were significantly reduced in the interleukin-10 treated group as compared with those in the vehicle-treated groups [(12.1±2.2)%, (15.5±3.6)%, P < 0.05].     

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.     

Effect of propofol on brain-derived neurotrophic factor and tyrosine kinase receptor B in the hippocampus of aged rats with chronic cerebral ischemia

We intraperitoneally injected 10 and 50 mg/kg of propofol for 7 consecutive days to treat a rat model of chronic cerebral ischemia. A low-dose of propofol promoted the expression of brain-derived neurotrophic factor, tyrosine kinase receptor B, phosphorylated cAMP response element binding protein, and cAMP in the hippocampus of aged rats with chronic cerebral ischemia, but a high-dose of propofol inhibited their expression. Results indicated that the protective effect of propofol against cerebral ischemia in aged rats is related to changes in the expression of brain-derived neurotrophic factor and tyrosine kinase receptor B in the hippocampus, and that the cAMP-cAMP responsive element binding protein pathway is involved in the regulatory effect of propofol on brain-derived neurotrophic factor expression.