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.

2-(2-Benzofuranyl)-2-imidazoline treatment within 5 hours after cerebral ischemia/reperfusion protects the brain

We previously demonstrated that administering 2-（2-benzofuranyl）-2-imidazolin（2-BFI）, an imidazoline I2 receptor agonist, immediately after ischemia onset can protect the brain from ischemic insult. However, immediate administration after stroke is difficult to realize in the clinic. Thus, the therapeutic time window of 2-BFI should be determined. Sprague-Dawley rats provided by Wenzhou Medical University in China received right middle cerebral artery occlusion for 120 minutes, and were treated with 2-BFI（3 mg/kg） through the caudal vein at 0, 1, 3, 5, 7, and 9 hours after reperfusion. Neurological function was assessed using the Longa＇s method. Infarct volume was measured by 2,3,5-triphenyltetrazolium chloride assay. Morphological changes in the cortical penumbra were observed by hematoxylin-eosin staining under transmission electron microscopy. The apoptosis levels in the ipsilateral cortex were examined with terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling（TUNEL） assay. The protein expression of Bcl-2 and BAX was detected using immunohistochemistry. We found the following： Treatment with 2-BFI within 5 hours after reperfusion obviously improved neurological function. Administering 2-BFI within 9 hours after ischemia/reperfusion decreased infarct volume and alleviated apoptosis. 2-BFI administration at different time points after reperfusion alleviated the pathological damage of the ischemic penumbra and reduced the number of apoptotic neurons, but the protective effect was more obvious when administered within 5 hours. Administration of 2-BFI within 5 hours after reperfusion remarkably increased Bcl-2 expression and decreased BAX expression. To conclude, 2-BFI shows potent neuroprotective effects when administered within 5 hours after reperfusion, seemingly by up-regulating Bcl-2 and down-regulating BAX expression. The time window provided clinical potential for ischemic stroke by 2-BFI.

Protective effect of 4-methyl-2-{[4-(toluene-4-sulfonyl)-thiomorpholine-3-carbonyl]-amino}-pentanoic acid isopropyl ester against cerebral ischemia

To investigate the effects of 4-methyl-2-{[4-(toluene-4-sulfonyl)-thiomorpholine-3-carbonyl]-amino}-pentanoic acid isopropyl ester(HD5-6)against cerebral ischemia in rodents,the models with global and focal ischemia were induced by bilateral common carotid artery occlusion plus hypotension(BCAOH)and permanent cerebral artery occlusion(p-MCAO)in mice(n=10–12 per group in BCAOH;n=8 per group in p-MCAO)and rats(n=10-11 in each group).HD5-6 prolonged lifetime and improved neurological function.Neurological deficits score in HD5-6(30 mg/kg)decreased significantly.Malonaldehyde(MDA)in HD5-6-treated mice with ischemia considerably dropped.The infarction volume of the HD5-6-treated rats with MCAO-induced ischemia decreased significantly in the high dose group(P<0.05,i.g.and P<0.01,i.v.).Immunohistochemistry showed that Brain derived neurotrophic factor(BDNF)in the ipsilateral hemisphere increased and Vascular endothelial growth factor(VEGF)decreased with HD5-6 treatment.HD5-6 has protective effects against experimental cerebral ischemia in rodents and the action mechanism may involve anti-oxidation and neurogenesis.

TNF-α and plasma D(-)-lactate levels in rats after intestinal ischemia and reperfusion

Objective To study the potential role of tumor necrosis factor-α (TNF-α) induction in the development of mucosal barrier dysfunction in rats caused by acute intestinal ischemia-reperfusion injury, and to examine whether pretreatment with monoclonal antibody against TNF-α (TNF-α MoAb) would affect the release of D(-)-lactate after local gut ischemia followed by reperfusion. Methods Anesthetized Sprague-Dawley rats underwent superior mesenteric artery occlusion for 75 min followed by reperfusion for 6 hr. The rats were treated intravenously with either TNF-α MoAb (20 mg/kg) or albumin (20 mg/kg) 30 min prior to the onset of ischemia. Plasma D(-)-lactate levels were measured in both the portal and systemic blood by an enzymatic spectrophotometric assay. Intestinal TNF-αmRNA expression as well as protein levels were also measured at various intervals. In addition, a postmortem examination was performed together with a macropathological evaluation based on a four-grade scoring system.Results Intestinal ischemia resulted in a significant elevation in D(-)-lactate levels in the portal vein blood in both the control and treatment groups ( P ＜0.05). However, animals pretreated with TNF-α MoAb at 6 hr after reperfusion showed significant attenuation of an increase in both portal and systemic D(-)-lactate levels when compared with those only receiving albumin (P ＜ 0.05). In the control animals, a remarkable rise in intestinal TNF-α level was measured at 0.5 hr after clamp release ( P ＜ 0.01); however, prophylactic treatment with TNF-α MoAb completely annulled the increase of local TNF-α levels seen in the control animals. Similarly, after anti-TNF-α MoAb administration, intestinal TNF-α mRNA expression was markedly inhibited, which showed significant differences when compared with the control group at 0.5 hr, 2 hr and 6 hr after the release of occlusion ( P ＜ 0.05-0.01 ). In addition, the pathological examination showed marked intestinal lesions that formed during ischemia, which were much worse upon reperfusion,particularly at the 6 hr time point. These acute injuries were obviously attenuated in animals receiving TNF-α MoAb.Conclusions It appeared that acute intestinal ischemia was associated with failure of the mucosal barrier, resulting in increased plasma D(-)-lactate levels in both portal and systemic blood. These results suggest that TNF-α appears to be involved in the development of local damage associated with intestinal ischemic injury. Moreover, prophylactic treatment with TNF-α MoAb exerts preventive effects on ischemia/ reperfusion-induced circulating D (-)-lactate elevation and gut injury. ( J Geriatr Cardiol 2004;1(2):119-124. )

Puerarin inactivates NLRP3-mediated pyroptotic cell death to alleviate cerebral ischemia/reperfusion(I/R)injury through modulating the LncRNA DUXAP8/miR-223-3p axis

NLRP3 inflammasome-mediated cell pyroptosis aggravates the development of cerebral ischemia/reperfusion(I/R)injury,and the aim of this study is to investigate the potential utilization of the Chinese medicine,Puerarin,in treating this disease.Through conducting in vitro and in vivo experiments,the present study illustrated that Puerarin regulated LncRNA double homeobox A pseudogene 8(DUXAP8)/miR-223-3p axis to inactivate NLRP3-mediated pyroptotic cell death,resulting in the attenuation of I/R injury.Specifically,the cerebral I/R injury in rat models and hypoxia/reoxygenation(H/R)in primary hippocampus neuron(PHN)cells were inducted,which were subsequently exposed to Puerarin treatment.As expected,we validated that Puerarin suppressed cell pyroptosis and rescued cell viability in I/R rat hippocampus tissues and H/R PHN cells.Next,through bioinformatics analysis,we noticed that miR-223-3p targeted both LncRNA DUXAP8 and NLRP3 mRNA,and both LncRNA DUXAP8 ablation and miR-223-3p overexpression inactivate NLRP3-mediated cell pyroptosis to rescue cell viability in H/R PHN cells.Interestingly,we evidenced that Puerarin restrained LncRNA DUXAP8 expressions,but upregulated miR-223-3p in I/R rat tissues and H/R PHN cells,and the protective effects of Puerarin on H/R PHN cells were abrogated by overexpressing LncRNA DUXAP8 and silencing miR-223-3p.Collectively,we concluded that Puerarin regulated LncRNA DUXAP8/miR-223-3p/NLRP3 signaling cascade to attenuate I/R injury.

锌对缺血再灌注大鼠肝脏金属硫蛋白-1基因表达的影响

目的 研究锌对肝脏缺血再灌注损伤 (HIRI)保护作用的分子机制。方法 采用RT- PCR技术 ,检测分析金属硫蛋白 - 1 (MT- 1 )基因在 HIRI大鼠肝组织中的表达和锌对其表达的调节。结果 各组大鼠肝组织均有MT- 1基因表达 ;缺血 30 min、再灌 90 min大鼠肝组织中 MT- 1m RNA较对照组显著降低 (P<0 .0 1 ) ;灌胃补锌后 ,HIRI大鼠肝 MT- 1 m RNA转录水平较对照组明显增高 (P<0 .0 1 ) ,单纯补锌亦可上调其组织中 MT- 1基因表达 (P<0 .0 1 )。结论 在锌对HIRI产生保护作用的分子机制中 ,调节其肝组织中 MT- 1转录水平的表达可能是一条重要途径。

肾缺血-再灌注后大鼠肾组织一氧化氮的代谢研究

目的:探讨肾I-R后大鼠肾组织NO的代谢.方法:建立大鼠肾I-R模型,用硝酸还原酶法测定NO含量,SP试剂盒免疫组织化学法检测肾组织cNOS、iNOS蛋白定位表达,Western blot检测肾组织cNOS、iNOS蛋白半定量表达.结果:(1)肾I-R后大鼠肾脏NO代谢异常,入肾血、出肾血NO2-/NO3-含量增多;同时肾组织NO2-/NO3-含量和尿液NO2-/NO3-排出率也有增高(P<0.05);(2)肾I-R后大鼠肾组织cNOS表达增多,以I-R1 h最明显;iNOS则在I-R5 h表达增多;(3)I-R1 h和I-R5 h大鼠肾小球内cNOS阳性信号均明显增强;而肾小管内cNOS阳性信号I-R1 h显著减弱;I-R1 h可见髓袢升枝和降枝粗段的肾小管有微弱的iNOS表达,而I-R5 h肾小管的iNOS阳性信号明显增强.结论:肾I-R导致了大鼠肾脏cNOS、iNOS蛋白表达改变,从而使NO代谢异常.

瘦素在心肌缺血-再灌注损伤中的作用机制

心肌缺血-再灌注损伤（myocardial ischemia reperfusioninjury,MIRI）是指在阻断冠状动脉一定时间后缺血的心肌在开放循环再灌注期出现的心脏功能、代谢及结构上的损伤。在恢复血流再灌注后,早期缺血心肌会出现损伤加重,引起更多心肌细胞死亡,心肌梗死面积增大,从而进一步损害心功能[1-2]。当前,世界各国都在寻求改进心肌保护的方法,但是心肌缺血-再灌注后的心肌细胞损伤仍无法修复。在众多因素中,瘦素（leptin）与MIRI的关系已经引起广泛关注,大量资料显示。

Investigation of Linarinic acid and one of its derivatives against cerebral ischemia in mice

The study aims to investigate the effects of(-)-Linarinic acid(LA) and one of its derivatives(LAd) on brain injury induced by ischemia. Malonaldehyde(MDA) is determined as an index for lipid peroxidation both in vitro and vivo. Mice were pre-treated with LA and LAd for 3 d.Thereafter, they were induced to have incomplete cerebral ischemia with both bilateral carotid artery occlusion and hypotension(BCAOH). In the first part of the in vivo experiment, mice were divided into four groups: sham(control), ischemia, ischemia + LA(200 mg/kg, i.g.) and ischemia + LAd(200 mg/kg, i.g.). In the second part, the dose-response of LAd was investigated at 100, 200 and 400 mg/kg i.g., respectively. A modified neurological severity score was developed for evaluating behavioral deficits of the mice with ischemia. Brains of the mice were excised in order to determinate MDA after ischemia for 6 h. Survival time, survival rate, neurological injury score and MDA level in brains were observed. Results were: 1) The data in vitro showed that both LA and LAd could inhibit the generation of MDA. IC50 values obtained by Probit analysis were 2.9 mM for LAd and 4.88 mM for LA;2) BCAOH could significantly shorten the survival span, reduce the survival rate and cause neurological deficits,which were associated with high level of lipid hydroperoxide production in cerebral tissues;3) LAd decreased lipid peroxidation and improved the neurological outcome more than LA.It is concluded that LAd offers a better neuroprotection than LA against brain damage caused by cerebral ischemia.

Rac1 relieves neuronal injury induced by oxygen-glucose deprivation and re-oxygenation via regulation of mitochondrial biogenesis and function

Certain microRNAs(miRNAs)can function as neuroprotective factors after reperfusion/ischemia brain injury.miRNA-142-3p can participate in the occurrence and development of tumors and myocardial ischemic injury by negatively regulating the activity of Rac1,but it remains unclear whether miRNA-142-3p also participates in cerebral ischemia/reperfusion injury.In this study,a model of oxygen-glucose deprivation/re-oxygenation in primary cortical neurons was established and the neurons were transfected with miR-142-3p agomirs or miR-142-3p antagomirs.miR-142-3p expression was down-regulated in neurons when exposed to oxygen-glucose deprivation/re-oxygenation.Over-expression of miR-142-3p using its agomir remarkably promoted cell death and apoptosis induced by oxygen-glucose deprivation/re-oxygenation and improved mitochondrial biogenesis and function,including the expression of peroxisome proliferator-activated receptor-γcoactivator-1α,mitochondrial transcription factor A,and nuclear respiratory factor 1.However,the opposite effects were produced if miR-142-3p was inhibited.Luciferase reporter assays verified that Rac Family Small GTPase 1(Rac1)was a target gene of miR-142-3p.Over-expressed miR-142-3p inhibited NOX2 activity and expression of Rac1 and Rac1-GTPase(its activated form).miR-142-3p antagomirs had opposite effects after oxygen-glucose deprivation/re-oxygenation.Our results indicate that miR-142-3p down-regulates the expression and activation of Rac1,regulates mitochondrial biogenesis and function,and inhibits oxygen-glucose deprivation damage,thus exerting a neuroprotective effect.The experiments were approved by the Committee of Experimental Animal Use and Care of Central South University,China(approval No.201703346)on March 7,2017.

Bumetanide promotes neural precursor cell regeneration and dendritic development in the hippocampal dentate gyrus in the chronic stage of cerebral ischemia

Bumetanide has been shown to lessen cerebral edema and reduce the infarct area in the acute stage of cerebral ischemia. Few studies focus on the effects of bumetanide on neuroprotection and neurogenesis in the chronic stage of cerebral ischemia. We established a rat model of cerebral ischemia by injecting endothelin-1 in the left cortical motor area and left corpus striatum. Seven days later, bumetanide 200 μg/kg/day was injected into the lateral ventricle for 21 consecutive days with a mini-osmotic pump. Results demonstrated that the number of neuroblasts cells and the total length of dendrites increased, escape latency reduced, and the number of platform crossings increased in the rat hippocampal dentate gyrus in the chronic stage of cerebral ischemia. These findings suggest that bumetanide promoted neural precursor cell regeneration, dendritic development and the recovery of cognitive function, and protected brain tissue in the chronic stage of ischemia.

醒脑静治疗椎-基底动脉供血不足疗效观察

椎-基底动脉供血不足（vertebrobasilar ischemia,VBI）为临床常见病,是由椎-基底动脉系统短暂脑缺血发作（transient is-chemic attack,TIA）导致。椎-基底动脉系统主要供应脑干、小脑、海马、枕叶、部分颞叶的血流,

尼麦角林治疗椎-基底动脉缺血性眩晕疗效观察

目的：探讨尼麦角林对椎-基底动脉缺血性眩晕的临床疗效。方法：治疗组用尼麦角林6mg加入0．9％生理盐水100ml静脉滴注，每日一次，共用2w。对照组用维脑路通100ml静脉滴注每日一次，共用2w。其他口服药相同。结果：治疗组30例，显效8例，有效21例，无效1例，总有效率为97％；对照组30例，显效8例，有效15例，无效7例，总有效率78％。结论：尼麦角林能显著改善椎一基底动脉缺血性眩晕的症状，其疗效肯定，且未见不良反应，因此，尼麦角林可作为治疗的首选药物之一。

Protective activity of adipose-derived stem cell extracellular vesicles in ischemia and/or reperfusion

Increasing evidence of the significant clinical value of protection against ischemia/reperfusion injury has contributed to the realization of the independent importance of this approach in improving prognosis and reducing cardiovascular mortality.Extracellular vesicles(EVs)derived by adipose mesenchymal stem cells may mediate the paracrine effects of stem cells and provide regenerative and anti-inflammatory properties,which are enhanced byγ-aminobutyric acid.The protective effects on cardiac myocytes may result from the EV embarked by miR-21-5p,which is a target for thioredoxin-interacting protein,regulating the formation of thioredoxin-interacting protein-thioredoxin complexes and subsequently enhancing the antioxidant activity of thioredoxin.It has been found thatγ-aminobutyric acid governs myocardial bioenergetics through suppressing inflammation and supporting mitochondrial structure.Finally,stem cell-based cell-free therapy based on adipose-derived stem cell EVs is considered a promising approach to individualized management of ischemia-induced cardiomyopathy.

NO与肝脏缺血再灌注损伤预处理保护效应

目的 :研究缺血预处理对SD大鼠肝脏缺血再灌注损伤保护机制中NO的作用 ;方法 :建立大鼠肝脏缺血再灌注模型并进行缺血预处理 ,实验分组如下 :假手术组 (Sham -operation ,S组 ) ;缺血再灌注组 (IschemicReperfusion ,I/R组 ) ;缺血预处理组 (IschemicPreconditioning ,PC组 ) ;预处理加左旋硝基精氨酸组 (Precondi tioning +L -NNA ,PC +L组 ) ,各组再灌注后检测血清丙氨酸转氨酶 (ALT)及丙二醛 (MDA)含量 ,肝组织石蜡切片HE染色及电镜观察 ;结果 :PC组ALT及MDA含量明显低于I/R组 (P <0 .0 1) ,肝组织损伤程度明显减轻 ;PC +L组在 0 - 3h阶段ALT和MDA含量与I/R组无显著区别 (P >0 .0 5 ) ,而在 6～ 48h阶段则显著低于I/R组 (P <0 .0 1)但仍高于PC组 (P <0 .0 5 )。结论 :缺血预处理对大鼠肝脏缺血再灌注损伤具有明显保护作用 ,0～ 3h为早期保护效应 。

肢体缺血预处理对止血带性心肌损伤的保护作用

止血带可减少术野出血,保证手术野的清晰而成为骨科常用的设备.但长时间止血带后又必然使肢体产生缺血-再灌注损伤（limbs ischemia reperfusion,LIR）,主要表现细胞因子的释放、中性粒细胞粘附后释放毒素介质及氧自由基的形成.一方面影响了局部组织功能,另一方面产生全身炎症反应,累及心肺等器官[1].特别是心血管系统的损伤最为重要,常表现为应激性心肌缺血和心功能损伤等[2].已有研究发现,肢体缺血预处理能有效启动内源性心肌保护作用,但这些研究主要讨论肢体缺血预处理对心肌缺血-再灌注的保护[3～5].本研究旨在探讨肢体缺血预处理是否能减轻下肢缺血-再灌注所致的心肌损伤.