大鼠脑创伤后海马CA3区细胞凋亡及相关基因表达研究

目的研究弥漫性脑损伤后不同时间,大鼠海马CA3区细胞凋亡及相关基因Bcl-2、Bax和Caspase-3蛋白的表达情况,探讨脑创伤后神经细胞凋亡的分子生物学机制.方法应用流式细胞仪和免疫组化法,分别检测脑创伤后不同时间海马CA3区细胞凋亡率及Bcl-2,Bax和Caspase-3基因在蛋白质水平的表达情况.结果脑创伤后海马CA3区存在不同程度细胞凋亡,Bcl-2在脑损伤后表达下降,而Bax和Caspase-3在脑创伤后表达升高;Caspase-3表达的峰值时间(72 h)出现在Bax之后(48 h).结论弥漫性脑损伤后,大鼠海马CA3区存在细胞凋亡及Bcl-2,Bax和Caspase-3的表达变化.Bcl-2/Bax表达比值下降早于Caspase-3的上升,Bcl-2/Bax表达比值改变可能与Caspase-3活化有关,进而启动并加重脑损伤后神经细胞凋亡.     

缺氧复氧大鼠海马星形胶质细胞凋亡及Bcl-2和Bax蛋白表达的时间进程变化及异丙酚干预效应

BACKGROUND： Cerebral hippocampal astrocytes are more sensitive.to ischemic injury than neurons. Hypoxic-ischemic brain injury induces profound astrocyte apoptosis, and propofol may protect against astrocyte apoptosis.
OBJECTIVE： To verify the protective effects of propofol against astrocyte apoptosis and to investigate anti-apoptotic Bcl-2 and pro-apoptotic Bax expression in primary cultures of rat hippocampal astrocytes exposed to hypoxia-reoxygenation for different periods of time following propofol treatment.
DESIGN, TIME, AND SETTING： In vitro neural immunocytochemistry was performed at the Central Laboratory of Yunyang Medical College between September 2007 and March 2008.
MATERIALS： A total of 30 Wistar rats, aged 1-3 days, wJth equal numbers of males and females, were included for isolation and culture of .hippocampal astrocytes.
METHODS： Hippocampal astrocytes were purified and cultured for 3 weeks and treated with four culture conditions： 50 μL Hank＇s solution （normal control）; 0.2 mL/L Intralipid; 50 μL Hank＇s solution for 10 minutes followed by hypoxic incubation for 4 hours and normoxic incubation for 12, 24, 36, 48, 60 or 72 hours; propofol （250 μmol/L final） for 10 minutes followed by hypoxic incubation for 4 hours and normoxic incubation for 12, 24, 36, 48, 60 and 72 hours.
MAIN OUTCOME MEASURES： （1） Morphologic changes in hippocampal astrocytes. （2） Levels of astrocyte apoptosis and Bcl-2 and Bax expression.
RESULTS： Hypoxia and reoxygenation increased apoptosis over time, with Bcl-2 expression peaking at 24 hours and decreasing gradually （P 〈 0.01 ）; Bax expression peaked at 72 hours （P 〈 0.01）; the ratio of Bcl-2/Bax was 1.4, 0.8, and 0.6, respectively, at 24, 48 and 72 hours. Non-apoptotic astrocytes showed significant proliferation and swelling. Propofol treatment decreased apoptosis after hypoxia-reoxygenation （P 〈 0.01）, as well as Bct-2 and Bax expression （P 〈 0.05, P 〈 0.01）, with Bcl-2/Bax ratios of 1.6-1.8. Propofol treatmentalso blocked astrocyte proliferation and swelling. No apoptotic cells or Bcl-2/Bax expression was detected in astrocytes cultured in Hank＇s or Intralipid solution.
CONCLUSION： Propofol protects astrocytes against injury caused by hypoxia and reoxygenation via a mechanism that involves maintaining high ratios of Bcl-2/Bax.     

Apoptosis-related protein expression in rabbits with blast brain injury following early hyperbaric oxygen therapy

We treated detonator-explosion-induced craniocerebral injury in rabbits with hyperbaric oxygen 1-24 hours post-injury.Expression of the apoptosis-regulating protein cytochrome c,the pro-apoptotic protein Bax and the apoptosis marker caspase-3 in the tissues surrounding the area of injury was significantly reduced,while that of the anti-apoptotic protein Bcl-2 was significantly increased.Our findings indicate that the curative effects of early hyperbaric oxygen on cortical cell apoptosis is associated with suppression of cytochrome c release from mitochondria.This mechanism underlies the observed reduction in Bax expression and upregulation of Bcl-2 expression.     

大鼠全脑缺血再灌注损伤后调控基因Bcl-2、Bax的表达及与细胞凋亡的关系

目的:本研究旨在探讨Bcl-2及Bax蛋白在大鼠全脑缺血再灌注损伤中的变化及与细胞凋亡的关系。方法:雄性Wistar大鼠56只,随机分为假手术组、缺血15分钟再灌注1、6、12、24、48、72小时组。采用大鼠四条血管阻断方法制备大鼠全脑缺血再灌注模型。采用TUNEL法观察不同再灌注时间组海马CAl区细胞凋亡的变化。采用免疫组化法观察Bcl-2及Bax蛋白表达水平的变化。结果:脑缺血损伤后随再灌注时间延长凋亡细胞逐渐增多,至再灌注48小时达到高峰,72小时后减少。Bcl-2表达至再灌注12小时达高峰,再灌注24~72小时组逐渐减弱。Bax表达至48小时达高峰,再灌注72小时减少。结论:Bcl-2于再灌注早期表达增强,Bax于再灌注中期表达增强,Bcl-2/Bax比例失衡可能是大鼠全脑缺血再灌注后神经细胞凋亡的机制之一。     

葛根素对创伤性脑损伤神经细胞Bcl-2、Bax蛋白表达影响

目的 对比研究苯甲酸雌二醇和葛根素对成年大鼠创伤性脑损伤神经细胞中Bcl-2、Bax 蛋白表达的影响.方法 改良Feeney法建立创伤性脑损伤大鼠模型,苯甲酸雌二醇、葛根素腹腔注射7d,利用病理学镜检、末端脱氧核苷酸转移酶介导的生物素脱氧尿嘧啶核苷酸缺口末端标记法(TUNEL)及免疫组织化学法检测神经细胞凋亡及Bcl-2、Bax蛋白的表达.结果 与损伤对照组相比,两种雌激素均显著改善创伤性脑损伤神经细胞的病理学改变,减轻细胞凋亡程度,明显增加Bcl-2 蛋白的表达、降低Bax/Bcl-2比率,但对Bax蛋白的表达无影响;两种雌激素相比,上述指标差异无统计学意义.结论 苯甲酸雌二醇和葛根素对创伤性脑损伤细胞均有神经保护作用,其作用机制可能与增加Bcl-2蛋白表达、延缓细胞凋亡有关.     

Medium-intensity acute exhaustive exercise induces neural cell apoptosis in the rat hippocampus

The present study assessed the influence of medium-intensity (treadmill at a speed of 19.3 m/min until exhaustion) and high-intensity (treadmill at a speed of 26.8 m/min until exhaustion) acute exhaustive exercise on rat hippocampal neural cell apoptosis. TUNEL staining showed significantly increased neural cell apoptosis in the hippocampal CA1 region of rats after medium- and high-intensity acute exhaustive exercise, particularly the medium-intensity acute exhaustive exercise, when compared with the control. Immunohistochemistry showed significantly increased expression of the antiapoptotic protein Bcl-2 and the proapoptotic protein Bax in the hippocampal CA1 region of rats after medium- and high-intensity acute exhaustive exercise. Additionally, the ratio of Bax to Bcl-2 increased in both exercise groups. In particular, the medium-intensity acute exhaustive exercise group had significantly higher Bax and Bcl-2 protein expression and a higher Bax/Bcl-2 ratio. These findings indicate that acute exhaustive exercise of different intensities can induce neural cell apoptosis in the hippocampus, and that medium-intensity acute exhaustive exercise results in greater damage when compared with high-intensity exercise.     

Temporal characterisation of pro- and anti-apoptotic mechanisms following diffuse traumatic brain injury in rats.

Few studies have characterised apoptosis in a brain injury model that causes a significant degree of diffuse axonal injury. Such characterisation is essential from a clinical viewpoint since diffuse axonal injury is a major component of human head injury. The present study therefore, examines the expression of active and proactive caspase-3, and the bax, bcl-2 and bcl-x members of the bcl-2 family, to characterise the temporal profile of apoptosis in a model of traumatic brain injury in rats that produces significant diffuse axonal injury. Pentobarbital anaesthetised male Sprague-Dawley rats were injured using the 2m impact-acceleration model of diffuse traumatic brain injury. After injury, diffuse trauma resulted in an increased bax expression followed by induction of caspase-3. The increase in caspase-3 was simultaneous with an increase in anti-apoptotic bcl-2 expression. Bcl-x levels were increased after induction of caspase-3 and the increased levels of bcl-x were sustained to the end of the 5-day observation period. Increased active caspase-3 expression was associated with the appearance of TUNEL positive cells. These cells were detected in different brain regions at different times, with some regions showing no apoptotic cells until 3 days after injury. No TUNEL positive cells were detected at 7 and 14 days after injury. DNA electrophoresis confirmed that DNA fragmentation was maximal at 3 days after injury. Increased active caspase-3 levels were also significantly correlated with increased bcl-2 levels (r=0.80; P<0.001) suggesting that the apoptotic cascade after diffuse traumatic brain injury is a carefully controlled cellular homeostatic response. Pharmacological manipulation of this balance may offer a therapeutic approach for preventing cell death and improving outcome after diffuse traumatic brain injury.     

Neuroglobin expression in rats after traumatic brain injury

In this study,we used a rat model of severe closed traumatic brain injury to explore the relationship between neuroglobin,brain injury and neuronal apoptosis.Real-time PCR showed that neuroglobin mRNA expression rapidly increased in the rat cerebral cortex,and peaked at 30 minutes and 48 hours following traumatic brain injury.Immunohistochemical staining demonstrated that neu-roglobin expression increased and remained high 2 hours to 5 days following injury.The rate of in-crease in the apoptosis-related Bax/Bcl-2 ratio greatly decreased between 30 minutes and 1 hour as well as between 48 and 72 hours post injury.Expression of neuroglobin and the anti-apoptotic factor Bcl-2 greatly increased,while that of the proapoptotic factor decreased,in the cerebral cortex post severe closed traumatic brain injury.It suggests that neuroglobin might protect neurons from apoptosis after traumatic injury by regulating Bax/Bcl-2 pathway.     

硫酸镁对大鼠脑弥漫性轴索损伤海马区Bcl-2和Bax蛋白表达影响的研究

目的研究脑弥漫性轴索损伤后神经细胞迟发性死亡的机制,探讨镁离子对大鼠脑弥漫性轴索损伤后神经元的保护作用。方法采用Marmarou方法制备大鼠重度脑弥漫性轴索损伤模型,分为创伤组(n=25)、生理盐水组(n=40)和硫酸镁组(n=40)。硫酸镁组伤后半小时给予25%硫酸镁(750μmol/kg),生理盐水组在相同时间给予等量的生理盐水腹腔注射,于伤后6小时、24小时、3天、5天及7天5个时相点处死。采用HE染色、免疫组织化学技术动态观察大鼠海马区的组织病理改变,Bcl-2和Bax蛋白的表达情况,以及使用硫酸镁干预后对上述表达的影响。结果①Bcl-2蛋白的表达:假手术组大鼠海马区仅见极少量Bcl-2阳性细胞,着色淡。在伤后6小时即有大量的Bcl-2阳性细胞,随时间渐增,24小时达到高峰,3～7天逐渐减少。②Bax蛋白的表达:在DAI后大鼠海马区有大量Bax阳性细胞,在伤后6小时就有所增加,24小时显著增加,3天达到高峰。Bcl-2/Bax值在损伤后随时间逐渐上升。③硫酸镁组中,海马区的Bax表达与对照组相比有所减少,而Bcl-2相应增加,Bcl-2/Bax比值也是上调的,均有统计学意义(P<0.05)。结论大鼠脑弥漫性轴索损伤后,海马区神经元存在有迟发性细胞死亡即凋亡现象。Bax与Bcl-2参与细胞凋亡过程。硫酸镁可通过抑制Bax蛋白,上调Bcl-2蛋白,减少神经细胞凋亡,对促进神经细胞修复和功能重塑有益。     

醒脑静联合丁苯肽对脑缺血再灌注损伤后细胞凋亡的影响

目的 观察醒脑静、丁苯酞及二者联合分别对大鼠脑缺血再灌注损伤后神经细胞凋亡及Bcl-2和Bax表达的影响。方法 60只雄性wistar大鼠(250±20)g采用改良线栓法制作脑缺血再灌注损伤模型(Middle cerebral artery occlusion,MCAO),随机分为4组,即模型组、醒脑静组、丁苯酞组、醒脑静联合丁苯酞(联合用药)组,每组又分为6、24、72 h三个亚组; 通过原位末端转移酶标记技术(TUNEL)检测神经细胞凋亡情况,采用免疫组化法观察大鼠脑缺血再灌注各个时间点Bcl-2、Bax的表达水平。结果(1)模型组手术对侧大脑半球偶见凋亡细胞,病灶区可见大量神经细胞凋亡。丁苯酞用药组、醒脑静用药组凋亡细胞数明显减少,醒脑静联合丁苯酞组凋亡细胞数最少(P<0.05);(2)丁苯酞组及联合用药组Bcl-2阳性表达水平较模型组均有提高,联合用药组Bcl-2阳性表达水平在各时间点均最高(P<0.05); 丁苯酞组及联合用药组Bax阳性表达水平较模型组均有降低,联合用药组Bax阳性表达水平最低(P<0.05)。结论(1)醒脑静、丁苯酞及二者联合均可能通过抑制脑缺血再灌注损伤后神经细胞凋亡来实现神经细胞保护作用,其中二者联合效果最佳;(2)丁苯酞可能通过增加脑缺血再灌注损伤大鼠Bcl-2表达,减少Bax表达的方式来减少神经细胞凋亡,从而减轻脑缺血再灌注损伤;(3)醒脑静本身不能对Bcl-2、Bax的表达水平产生影响,但其可能通过增强丁苯酞作用的方式影响Bcl-2、Bax的表达,从而减轻脑缺血再灌注损伤。     

Pretreatment with scutellaria baicalensis stem-leaf total flavonoid prevents cerebral ischemia- reperfusion injury

Pretreatment with scutellaria baicalensis stem-leaf total flavonoid has protective effects against ischemia and attenuates myocardial ischemia-reperfusion injury. In this study, rats were given scutellaria baicalensis stem-leaf total flavonoid intragastrically at 50, 100, and 200 mg/kg per day for 7 days before focal cerebral ischemia-reperfusion injury models were established using the suture method. We then determined the protective effects of scutellaria baicalensis stem-leaf total flavon- oid pretreatment on focal cerebral ischemia-reperfusion injury. Results showed that neurological deficit scores increased, infarct volumes enlarged, apoptosis increased and Bcl-2 and Bax protein expression were upregulated at 24 hours after reperfusion. Pretreatment with scutellaria baicalensis stem-leaf total flavonoid at any dose lowered the neurological deficit scores, reduced the infarct volume, prevented apoptosis in hippocampal cells, attenuated neuronal and blood-brain barrier damage and upregulated Bcl-2 protein expression but inhibited Bax protein expression. Doses of 100 and 200 mg/kg were the most efficacious. Our findings indicate that pretreatment with scutel- laria baicalensis stem-leaf total flavonoid at 100 and 200 mg/kg can improve the neurological func- tions and have preventive and protective roles after focal cerebral ischemia-reperfusion injury.     

表没食子儿茶素没食子酸酯保护阿米卡星诱导耳蜗螺旋神经节神经元的凋亡

透射电镜观察SD大鼠耳蜗螺旋神经节神经元形态变化,发现阿米卡星可以诱导耳蜗螺旋神经节神经元凋亡。免疫组化染色和RT-PCR检测发现,阿米卡星诱导耳蜗螺旋神经元Bcl-2 蛋白表达下调,Bax、Caspase-3蛋白及Caspase-6 mRNA表达增强。表没食子儿茶素没食子酸酯可以抑制耳蜗螺旋神经元Bax、Caspase-3蛋白及Caspase-6 mRNA的表达,同时增强Bcl-2蛋白表达,从而降低耳蜗螺旋神经元凋亡率。证实表没食子儿茶素没食子酸酯对阿米卡星损伤的耳蜗螺旋神经节具有保护作用。     

盐酸纳洛酮对严重颅脑损伤后神经细胞凋亡及相关基因Bcl-2、Bax表达影响的研究

目的探讨纳洛酮对严重颅脑损伤后神经细胞凋亡及Bcl-2、Bax基因表达的影响及其可能的作用机制。方法采用改进的Feeney自由落体损伤模型,应用分子生物学方法-脱氧核糖核苷酸末端转移酶介导的缺口末端标记法(terminal-deoxynucleotidyltransferasemediatednickendlabeling,TUNEL)以及免疫组织化学方法,对严重颅脑损伤大鼠进行纳洛酮治疗,检测受损脑组织中的凋亡细胞和Bcl-2、Bax免疫反应阳性细胞的数量。结果治疗组大鼠脑组织中凋亡细胞数低于损伤对照组,治疗组与损伤对照组之间Bcl-2的表达无显著差异,但治疗组3d、5d组Bax表达明显低于损伤对照组(P<0.05)。结论纳洛酮通过竞争性拮抗内源性阿片肽受体,可能通过抑制Bax的表达来减少颅脑损伤后神经细胞的凋亡,发挥脑保护作用,其详细机理尚有待进一步研究。     

川芎嗪对大鼠重型颅脑损伤后神经细胞凋亡及Bcl-2、Bax表达的影响

目的探讨川芎嗪对大鼠重型颅脑损伤后神经细胞凋亡及相关基因Bcl-2、Bax表达的影响和对脑神经的保护作用。方法120只SD健康大鼠随机分为假手术组、模型组、治疗组,其中模型组和治疗组采用Feeney自由落体撞击装置制作大鼠重型颅脑损伤模型,治疗组给予盐酸川芎嗪,用TUNEL及免疫组化法检测三组间细胞凋亡及Bcl-2、Bax蛋白的表达情况。结果大鼠脑组织中细胞凋亡率及Bcl-2、Bax表达水平在治疗组和模型明屁高于假手术组（P〈0．05）。在伤后72h、168h,治疗组的细胞凋亡率及Bax表达水平明显低于模型组,而Bcl-2的表达水平则明显高于模型组（P〈0．05）。结论川芎嗪可能通过抑制Bax的表达,上调Bcl-2的表达,减少神经细胞凋亡,减轻重型颅脑损伤后继发脑损害,从而发挥脑神经保护作用。     

亚低温对大鼠全脑缺血-再灌注损伤后神经细胞凋亡的治疗时间窗研究

目的本研究通过观察大鼠全脑缺血-再灌注损伤后不同时间段亚低温对Bcl-2、Bax表达及二者比值的影响,探讨亚低温的保护作用及最佳治疗时间窗。方法 Wistar雄性大鼠随机分为对照组及亚低温组。亚低温组根据亚低温时点不同分为缺血后、再灌注0、6、12、24h 5个亚组。采用四条血管阻断方法(Puisinelli-4VO法)制备大鼠全脑缺血-再灌注模型,流式细胞仪对海马组织的Bcl-2、Bax蛋白的表达进行测定。结果亚低温可使大鼠海马细胞的Bcl-2表达增高、Bax表达降低,以再灌注后0 h亚低温效果最佳,随再灌注时间延长,效果下降。结论亚低温可有效干预大鼠全脑缺血-再灌注损伤后神经细胞的凋亡,且最佳治疗时间窗为再灌注后0h,但再灌注24h也有一定疗效。     

Sex Differences and the Roles of Sex Steroids in Apoptosis of Sexually Dimorphic Nuclei of the Preoptic Area in Postnatal Rats

The brain contains several sexually dimorphic nuclei that exhibit sex differences with respect to cell number. It is likely that the control of cell number by apoptotic cell death in the developing brain contributes to creating sex differences in cell number in sexually dimorphic nuclei, although the mechanisms responsible for this have not been determined completely. The milieu of sex steroids in the developing brain affects sexual differentiation in the brain. The preoptic region of rats has two sexually dimorphic nuclei. The sexually dimorphic nucleus of the preoptic area (SDN-POA) has more neurones in males, whereas the anteroventral periventricular nucleus (AVPV) has a higher cell density in females. Sex differences in apoptotic cell number arise in the SDN-POA and AVPV of rats in the early postnatal period, and an inverse correlation exists between sex differences in apoptotic cell number and the number of living cells in the mature period. The SDN-POA of postnatal male rats exhibits a higher expression of anti-apoptotic Bcl-2 and lower expression of pro-apoptotic Bax compared to that in females and, as a potential result, apoptotic cell death via caspase-3 activation more frequently occurs in the SDN-POA of females. The patterns of expression of Bcl-2 and Bax in the SDN-POA of postnatal female rats are changed to male-typical ones by treatment with oestrogen, which is normally synthesised from testicular androgen and affects the developing brain in males. In the AVPV of postnatal rats, apoptotic regulation also differs between the sexes, although Bcl-2 expression is increased and Bax expression and caspase-3 activity are decreased in females. The mechanisms of apoptosis possibly contributing to the creation of sex differences in cell number and the roles of sex steroids in apoptosis are discussed.     

Evaluation of the apoptosis-related proteins of the BCL-2 family In the traumatic penumbra area of the rat model of cerebral contusion, treated by hyperbaric oxygen therapy: a quantitative immunohistochemical study

The growth and progression of traumatic brain injury (TBI) lesions depend significantly on developments in the traumatic penumbra area, perilesional region, where delayed neuronal death occurs. Recent data supports the important role of apoptosis in delayed cell death in TBI. Previously we demonstrated a significant reduction of apoptosis in traumatic penumbra in animals treated by hyperbaric oxygen (HBO).In this study we evaluate the expression of apoptosis-related proteins of the Bcl-2 family (Bcl-2, Bax and Bcl-xL) in the traumatic penumbra area in correlation with the extent of apoptosis in the rat model of focal cerebral contusion, treated by HBO. Sprague-Dawley rats underwent cortical dynamic deformation, some with subsequent hypoxemia. A group of both hypoxemic and non-hypoxemic animals was treated by HBO. The pathological study was based on immunohistochemical staining of the brain sections for Bcl-2, Bax and Bcl-xL with quantitative evaluation of staining by image analysis. The expression of Bcl-2 in hypoxemic animals was lower than in non-hypoxemic animals, but a significant increase in Bcl-2 expression was seen in both groups after HBO treatment. Bcl-xL also demonstrated an increase after HBO treatment but less significant. Staining for Bax protein did not demonstrate significant change after treatment. These data correlate well with the reduction of TUNEL-positive cells in traumatic penumbra after HBO treatment. We concluded that the apoptotic mechanisms are important in delayed cell death in TBI and that post-traumatic hypoxemia increases the intensity of apoptosis, probably through a decrease in Bcl-2 and Bcl-xL expression which normally repress apoptosis. The beneficial effect of HBO treatment in our model of brain contusion correlates well with the increased expression of anti-apoptotic proteins (Bcl-2 and Bcl-xL) following treatment and the appropriate decrease in the extent of apoptosis. In light of these results, the usage of HBO is justified as neuroprotective treament in TBI.     

Age-dependent cyclooxygenase-2 induction and neuronal damage after status epilepticus in the postnatal rat hippocampus

PURPOSE: Epileptic seizures lead to age-dependent neuronal damage in the developing brain, particularly in the hippocampus, but the mechanisms involved have remained poorly elucidated. In this study, we investigated the contribution of apoptosis and inflammatory processes to neuronal damage after status epilepticus (SE) in postnatal rats. METHODS: SE was induced by an intraperitoneal injection of kainic acid (KA) in 21- and 9-day-old (P21 and P9) rats. The expression of Bax, Bcl-2 and caspase-3, markers for apoptosis, and cyclooxygenase-2 (COX-2), an indicator for activation of inflammatory processes, were studied from 6 h up to 1 week after SE by Western blotting and immunocytochemistry. Neuronal damage was verified by Fluoro-Jade B staining. RESULTS: In P21 rats, SE resulted in neuronal damage in the CA1 neurons of the hippocampus. COX-2 expression was extensively, but transiently, increased and its immunoreactivity pronouncedly enhanced in several hippocampal subregions, amygdala, and piriform cortex by 24 h after SE. The expression of Bax and caspase-3 remained unchanged, whereas the antiapoptotic factor Bcl-2 transiently decreased by 24 h. Single caspase-3 positive neurons appeared in the CA1 region of both control and KA-treated rats. In P9 rats, no neuronal death was detected, and COX-2 expression and immunoreactivity remained at the control level. DISCUSSION: Our results suggest that SE provokes age-specific effects on COX-2 expression. This together with the activation of putative inflammatory processes may contribute to neuronal cell death in the hippocampus of postnatal rats, whereas caspase-dependent apoptosis seems not to be involved in the death process.     

The effect of ischemic post-conditioning on hippocampal cell apoptosis following global brain ischemia in rats

We evaluated the effect of brain ischemic post-conditioning on cell apoptosis in the hippocampus following global brain ischemia in rats. Adult male Sprague-Dawley rats were randomly divided into three groups (n=15/group): sham operation, ischemia/reperfusion (I/R) and ischemic post-conditioning (I PostC). Global brain ischemia was induced by four-vessel occlusion. Ischemic post-conditioning consisted of six cycles of 10s/10s reperfusion/reocclusion at the onset of reperfusion. All rats were sacrificed 24 hours or 72 hours after reperfusion. The hippocampal CA1 regions were analysed using the terminal deoxynucleotidyl transferase-mediated biotinylated deoxyuridine triphosphate nick end-labelling (Tunel) staining technique for determining cell apoptosis. Levels of caspase-3 and Bcl-2 were measured by Western blotting. After 72 hours, fewer Tunel-positive brain cells were observed in rats from the I PostC group than in rats from the I/R group (10.3 ± 2.7% versus 40.8 ± 6.2%, p<0.01). After reperfusion at 24 hours and 72 hours, expression of caspase-3 in the I PostC group was significantly decreased (p<0.01) and expression of Bcl-2 in the I PostC group was significantly increased (p<0.01) compared with the I/R group. We conclude that down-regulation of caspase-3 and up-regulation of Bcl-2 by ischemic post-conditioning may underlie the protective effects of post-conditioning.     

Neuroprotective effects of the immunodepressant FTY720 on caspase-3 expression and neural apoptosis in a rat model of acute spinal cord injury

BACKGROUND:Studies on the immunodepressant FTY720 have primarily focused on organ transplantation and autoimmune disease therapy.However,the effects on caspase-3 expression and neural apoptosis following acute spinal cord injury remain uncertain.OBJECTIVE:To elucidate the underlying mechanism of the immunodepressant FTY720 to alleviate spinal cord injury by inhibiting expression of caspase-3 and neural apoptosis.DESIGN,TIME AND SETTING:A randomized,controlled,animal experiment was performed at Central Laboratory of the Second Affiliated Hospital of Dalian Medical University from April to July 2009.MATERIALS:FTY720 was provided by Wuhan Yuancheng Technology Developing,China.METHODS:A total of 120 Sprague Dawley rats were randomly assigned to sham-surgery,model,and FTY720 groups.Spinal cord injury at the T9-10 segment was induced in model groups using the free-fall method.Following establishment of spinal cord injury at the T9-10 segment in the FTY720 group,rats were treated with an intragastric injection of 0.3 mL saline-diluted FTY720 (3 mg/kg).MAIN OUTCOME MEASURES:At 6,12,24,48,and 72 hours following spinal cord injury,caspase-3 expression was detected using streptavidin-peroxidase immunohistochemistry,and neural apoptosis was detected using the TUNEL method.RESULTS:Positive caspase-3 expression and neural apoptosis was not observed in the sham-surgery group at the various time points.The number of apoptotic cells increased with time after acute spinal cord injury,peaked at 24 hours following injury,and then gradually reduced.However,neural apoptosis remained at a high level.Caspase-3 expression positively correlated with neural apoptosis (r= 0.864,P< 0.05).Caspase-3 expression and neural apoptosis significantly decreased following FTY720 therapy (P< 0.05).CONCLUSION:FTY720 significantly reduced caspase-3 expression and neural apoptosis in a rat model of acute spinal cord injury.