普伐他汀对脑缺血大鼠的神经保护及神经发生的作用(英文)

目的研究中枢神经系统缺血损伤后,普伐他汀的神经保护和促进神经发生作用。方法采用线栓法造成大鼠大脑中动脉的暂时性缺血,在以下时间点给予普伐他汀:伤后6 h,伤后每天直至伤后14 天。用神经学评分、平衡实验和旋转实验评价伤后神经学恢复情况。检测血清胆固醇和甘油三酯的含量,计算脑梗塞面积。通过三染色法(BrdU, DCX, NeuN染色)研究普伐他汀对神经发生的作用。结果各组间血清胆固醇和甘油三酯无显著性差异;与对照组相比,实验组动物术后旋转实验评分显著性增加,梗塞面积减小;普伐他汀显著增加了齿状回和脑室下区的BrdU阳性细胞数,并增加了齿状回、脑室下区和纹状体中的BrdU/DCX阳性细胞数。结论中枢神经系统损伤早期重复使用低剂量的普伐他汀是相对安全的,并能够显著改善伤后的神经功能恢复,减少梗死面积。普伐他汀能够诱导大鼠齿状回及脑室下区的神经发生并增加纹状体中迁移神经元的数量,这与普伐他汀的降脂作用无关。     

普伐他汀对脑缺血大鼠的神经保护及神经发生的作用

目的研究中枢神经系统缺血损伤后,普伐他汀的神经保护和促进神经发生作用。方法采用线栓法造成大鼠大脑中动脉的暂时性缺血,在以下时间点给予普伐他汀：伤后6h,伤后每天直至伤后14天。用神经学评分、平衡实验和旋转实验评价伤后神经学恢复情况。检测血清胆固醇和甘油三酯的含量,计算脑梗塞面积。通过三染色法（BrdU, DCX, NeuN染色）研究普伐他汀对神经发生的作用。结果各组间血清胆固醇和甘油三酯无显著性差异;与对照组相比,实验组动物术后旋转实验评分显著性增加,梗塞面积减小;普伐他汀显著增加了齿状回和脑室下区的BrdU阳性细胞数,并增加了齿状回、脑室下区和纹状体中的BrdU/DCX阳性细胞数。结论中枢神经系统损伤早期重复使用低剂量的普伐他汀是相对安全的,并能够显著改善伤后的神经功能恢复,减少梗死面积。普伐他汀能够诱导大鼠齿状回及脑室下区的神经发生并增加纹状体中迁移神经元的数量,这与普伐他汀的降脂作用无关。     

Leptin induces neuroprotection neurogenesis and angiogenesis after stroke

Leptin is a potent AMP kinase (AMPK) inhibitor that is central to cell survival. Hence, we explored the effects of leptin on neurogenesis and angiogenesis after stroke. Neural stem cells (NSC) were grown as neurospheres in culture and treated with vehicle or leptin and neurosphere size and terminal differentiation were then determined. We then explored the effects of leptin on endogenous repair mechanisms in vivo. Sabra mice underwent photothrombotic stroke, were given vehicle or leptin and newborn cells were labeled with Bromo-deoxy-Uridine. Functional outcome was studied with the neurological severity score for 90 days post stroke and the brains were then evaluated with immunohistochemistry. In a subset of animals the brains were also evaluated for changes in the expression of leptin receptor and AMPK. In vitro, leptin led to a 2-fold increase in neurosphere size but did not change the differentiation of newborn cells. Following stroke, exogenous leptin led to a 4-fold increase in the number of NSC in the cortex abutting the lesion. There was a 1.5-fold increase in the number of newborn neurons and glia in leptin treated animals. Leptin also significantly increased the number of blood vessels in the peri-lesioned cortex. Leptin treated mice had increased expression of leptin receptor and increased phosphorylated AMPK concentration. Animals treated with leptin also had significantly better functional states. In conclusion, leptin induces neurogenesis and angiogenesis after stroke and leads to increased leptin receptor and pAMPK concentrations. This may explain at least in part the better functional outcome observed in leptin treated animals after stroke.     

变构促红细胞生成素对小鼠脑缺血后神经新生和血管新生的影响

目的 探讨一种氨基酸突变后无促红细胞生成能力的变构促红细胞生成素(mEPO)对大脑中动脉闭塞小鼠神经新生和血管新生的作用.方法 将30只成年雄性C57BL/B6小鼠随机分为Sham组、I/R+ Veh组和I/R+ mEPO组,每组10只.制作小鼠大脑中动脉闭塞缺血(MCAO)模型,I/R+mEPO组小鼠在再灌注即刻腹腔注入mEPO(5 000 IU/kg),I/R+ Veh组小鼠在制模后注入等体积的生理盐水.通过BrdU检测细胞增殖情况,采用转棒试验对小鼠神经功能进行评估.在脑缺血后第14天,检测3组小鼠脑组织丢失比例、神经新生及血管新生情况.结果 用mEPO处理后,I/R+mEPO组小鼠脑组织丢失比例(14.62±5.80)％显著低于I/R+ Veh组小鼠(29.81±7.75)％,差异有统计学意义(P＜0.05);I/R+mEPO组小鼠脑缺血后第3,7天神经功能均有明显改善,第7天即恢复到正常水平,差异有统计学意义(P＜0.05);I/R+mEPO组小鼠脑缺血后第14天BrdU+/NeuN+双阳性细胞数量(36.25±10.53)和BrdU+/Laminin+双阳性细胞数量(25.25±6.34)显著高于Sham组小鼠和I/R+ Veh组小鼠,差异有统计学意义(P＜0.05).结论 mEPO促进了小鼠脑缺血周边区神经新生及血管新生,从而减轻脑组织的损伤程度并提高神经功能.这种无促红细胞生成作用的mEPO可能会成为临床脑血管病的治疗药物.     

补阳还五汤对局灶性脑缺血大鼠神经元再生的影响

目的探讨补阳还五汤对局灶性脑缺血大鼠神经元再生的影响。方法Wistar大鼠随机分为假手术组、模型组、补阳还五汤组，每组32只，模型组、补阳还五汤组采用大脑中动脉阻塞法复制脑缺血模型，假手术组仅切开皮肤、分离右侧颈总动脉即缝合，补阳还五汤组在造模后予以补阳还五汤灌服，每日一次：另设正常对照组8只．不做任何处理。大鼠于实验开始后第一周每日腹腔给予5-溴脱氧核苷一次，此后每周两次。分别于l、7、14、28d时各处死8只大鼠（正常对照组为2只），观察大鼠神经功能，采用免疫组织化学法检测新生神经元数。结果补阳还五汤能显著减轻神经功能缺损；缺血后每组大鼠脑内均有少量新生神经元，补阳还五汤组数量较多，在7、14、28d与模型组比较，差异有显著性（P〈0．05或P〈0．01）。结论缺血后大鼠脑内存在有限的神经再生，补阳还五汤能增强此增生。     

Platelet microparticles induce angiogenesis and neurogenesis after cerebral ischemia

Activated platelets shed microparticles, which contain a variety of growth factors central to angiogenesis and neurogenesis. The aim of this study was to explore whether platelet derived microparticles (PMP) can boost endogenous neural stem cells dependent repair mechanisms following stroke in a rat model. To examine the effects of PMP therapy in-vivo, we delivered PMP or vehicle via a biodegradable polymer to the brain surface after permanent middle cerebral artery occlusion (PMCAO) in rats. Rats were tested with the neurological severity score and infarct volumes were measured at 90 days post-ischemia. Immunohistochemistry was used to determine the fate of newborn cells and to count blood vessels in the ischemic brain. The results show that PMP led to a dose dependent increase in cell proliferation, neurogenesis and angiogenesis at the infarct boundary zone and significantly improved behavioral deficits.     

Ligustilide provides neuroprotection by promoting angiogenesis after cerebral ischemia

ABSTRACT

Objectives

Following stroke, angiogenic strategy has been proposed to alleviate ischemia-induced injury by promoting angiogenesis and improving cerebrovascular function in the ischemic regions. Ligustilide (LGSL) is known to have neuroprotection against ischemic stroke-induced injury. But the effect of LGSL on promoting angiogenesis after ischemic stroke is unknown. The purpose of this study was to determine the effects of LGSL on neuroprotection and angiogenesis after stroke.     

细胞周期抑制剂对大鼠局灶性脑缺血后胶质细胞增殖和瘢痕形成的影响及对神经元的保护作用

目的 观察选择性细胞周期抑制剂olomoucine对胶质细胞增殖和瘢痕形成及神经元凋亡的影响.方法 建立光化学法诱导大鼠局灶性脑缺血模型,随机分为假手术组、对照组和干预组,采用MRI显示梗死灶并计算其梗死灶体积;应用免疫荧光化学法检测胶质纤维酸性蛋白(GFAP)的表达及通过TUNEL方法检测神经元凋亡;免疫印迹法观察光照侧皮质GFAP、增殖细胞核抗原(proliferation cell nuclear antigen,PCNA)、周期素蛋白A和周期素蛋白B1蛋白的表达.结果 缺血后3、7、30 d对照组(5.10%±0.35%,4.60%±0.26%,3.96%±0.28%)梗死灶体积占全脑体积百分比值的平均值明显大于干预组(2.27%±0.28%,1.87%±0.19%,1.08%±0.18%,P＜0.05);缺血后各时间点组GFAP表达明显增强,对照组明显强于干预组,并且7、30 d对照组梗死灶周边可见明显的胶质瘢痕形成,以30 d最为显著;缺血后3 d梗死灶周围可见大量TUNEL阳性染色细胞,对照组[(41±11)个/高倍视野]数量明显多于干预组[(26±8)个/高倍视野,P＜0.05];干预组大鼠(22.44%±2.17%)NeuN+TUNEL双标阳性表达明显弱于对照组大鼠(34.41%±3.10%,P＜0.05);3、7、30 d的GFAP、PCNA、周期素蛋白A和周期素蛋白B1蛋白量表达,对照组明显高于干预组.结论 通过对细胞周期的调控,可部分抑制胶质细胞的活化增殖及瘢痕形成,同时减小脑梗死体积及减少神经元凋亡.     

Growth factors improve neurogenesis and outcome after focal cerebral ischemia

Stem cells have been proposed as a new form of cell-based therapy in a variety of disorders, including acute and degenerative brain diseases. Endogenous neural stem cells (eNSC) reside in the subventricular zone and in the subgranular zone of the hippocampus. eNSC are capable of self-renewal and differentiation into functional glia and neurons. Unfortunately, spontaneous brain regeneration is inefficient for clinically significant improvement following brain injury. However, eNSC responses may be augmented considerably by perturbing the pathways governing cell proliferation, migration and differentiation by application of exogenous growth factors. Importantly, current evidence suggests that such perturbations may lead to better functional outcome after stroke. This article summarizes the progress made in this field.     

Behavioral and histological neuroprotection by tamoxifen after reversible focal cerebral ischemia

We have previously shown that tamoxifen can reduce infarct sizes measured by 2,3,5,-triphenyltetrazolium chloride (TTC) staining at 72 h after 2 h of reversible middle cerebral artery occlusion (rMCAo) in rats. In this study, we tested whether improvement is found in both behavioral measures of protection and by histological measures of infarcted tissue at 7 and 14 days after 2 h rMCAo. Tamoxifen (10 mg/kg) was given once by intravenous injection 1 h after reperfusion, i.e. 3 h after initiation of rMCAo. Neurobehavioral deficits were evaluated daily for 1 week or 2 weeks followed by infarct volumes measurements by hematoxylin-eosin (HE) staining. Tamoxifen-treated rats had significantly improved neurobehavioral deficit scores when evaluated daily throughout the 1 week or 2 week periods and showed significantly reduced median infarct volumes measured after 1 week and 2 weeks. Median infarct values were 149 mm3 (interquartile range, IQR: 92 to 258) and 124 mm3 (IQR: 69 to 174) for the 1 and 2 week vehicle groups, respectively, compared with 5 mm3 (IQR: 3 to 16) and 4 mm3 (IQR: 0 to 48) for the comparable treated groups (both P < 0.05, Mann-Whitney test), giving a reduction of more than 90% in both cases. Thus, a single administration of tamoxifen given 3 h after initiation of rMCAo is extremely effective in producing long-term neuroprotection as assessed by neurobehavioral measures and histopathology in experimental stroke in rats. If these results are extrapolatable to human stroke, these data indicate that tamoxifen may be a useful neuroprotectant.     

Erythropoietin enhances long-term neuroprotection and neurogenesis in neonatal stroke

Neonatal stroke leads to mortality and severe morbidity, but there is no effective treatment currently available. Erythropoietin (EPO) has been shown to promote cytoprotection and neurogenesis and decrease subventricular zone morphologic changes following brain injury. The long-term cellular response to EPO has not been defined, and local changes in cell fate decision may play a role in functional improvement. We performed middle cerebral artery occlusion in P10 rats. EPO treatment (5 U/g i.p.) significantly preserved hemispheric brain volume 6 weeks after injury. Furthermore, EPO increased the percentage of newly generated neurons while decreasing newly generated astrocytes following brain injury, without demonstrating long-term differences in the subventricular zone. These results suggest that EPO may neuroprotect and direct cell fate toward neurogenesis and away from gliogenesis in neonatal stroke.     

TRPC1沉默对脑缺血大鼠神经干细胞迁移的影响

目的探讨瞬时受体电位通道1(TRPC1)沉默对脑缺血大鼠内源性神经干细胞增殖、迁移和分化的影响。方法 SD大鼠分为假手术组、脑缺血组、TRPC1沉默组(脑缺血+TRPC1沉默)和阴性对照组,以脑室内注射siRNA沉默TRPC1,以线栓法制作大鼠大脑中动脉脑缺血(MCAO)模型,脑缺血模型制作成功后,腹腔内注射Brdu标记内源性神经干细胞,分别于48 h、4 w后处死大鼠,行Brdu免疫组化染色、免疫荧光双标染色(Brdu/GFAP、Brdu/Neun)观察NSC的增殖、迁移和分化情况。结果脑缺血后48 h,脑缺血组和TRPC1沉默组SVZ区Brdu阳性表达均较假手术组明显增多(P0.01),但TRPC1沉默组Brdu阳性细胞数较缺血组低(P0.01)。脑缺血4 w后,缺血组与假手术组相比,皮质区具有更多的Brdu阳性细胞(P0.01),同样TRPC1沉默组Brdu阳性细胞数多于假手术组,但明显低于脑缺血组(P0.01);免疫荧光双标染色发现,脑缺血各组Brdu/GFAP、Brdu/Neun双阳性细胞的数量均比假手术组明显增高,但TRPC沉默组双阳性细胞显著少于脑缺血组和阴性对照组(P0.01)。结论 TRPC1沉默显著影响脑缺血大鼠SVZ区NSC的增殖、向脑缺血区迁移及向成熟细胞的分化。     

Thrombolysis and neuroprotection in cerebral ischemia

Stroke is a major cause of death and disability worldwide. The resulting burden on society grows with the increase in the incidence of stroke. The term brain attack was introduced to describe the acute presentation of stroke and emphasize the need for urgent action to remedy the situation. Though a large number of therapeutic agents, like thrombolytics, NMDA receptor antagonists, calcium channel blockers and antioxidants, have been used or are being evaluated, there is still a large gap between the benefits of these agents and the properties of an ideal drug for stroke. So far, only thrombolysis with rtPA within a 3-hour time window has been shown to improve the outcome of patients with ischemic stroke. Understanding the mechanisms of injury and neuroprotection in these diseases is important to target news sites for treating ischemia. Better evaluation of the drugs and increased similarity between the results of animal experimentation and in the clinical setting requires critical assessment of the selection of animal models and the parameters to be evaluated. Our laboratory has employed a rat embolic stroke model to investigate the combination of rtPA with citicoline as compared to monotherapy alone and investigated whether neuroprotection should be provided before or after thrombolysis in order to achieve a greater reduction of ischemic brain damage.     

Acute neuroprotection by pioglitazone after mild brain ischemia without effect on long-term outcome

Peroxisome proliferator-activated receptor-gamma (PPAR-γ) agonists (thiazolidinediones) have anti-inflammatory effects and improve endothelium function. Here, we analyzed the effects of pioglitazone on short- and longer-term outcome after mild transient brain ischemia. 129/SV mice were subjected to 30 min filamentous middle cerebral artery occlusion (MCAo), followed by reperfusion. Post event, animals were treated with daily intraperitoneal (i.p.) pioglitazone (20 mg/kg body weight) or vehicle. Pioglitazone given acutely after transient brain ischemia/reperfusion reduced lesion size and the number of Iba1-expressing microglia in the ischemic striatum at three days. In vitro, pioglitazone attenuated migration and proliferation of primary mouse microglia. However, analysis at 6 weeks after MCAo/reperfusion no longer yielded an effect of pioglitazone on either lesion size or Iba1+ cell counts. Regarding functional longer-term outcome, we also did not detect a beneficial effect of pioglitazone on motor function measured either on the pole test or the wire hanging test or on learning and memory in the Morris water maze. Our study thus underscores the importance of extending experimental stroke studies to an analysis of longer-term outcome.