17-β雌二醇抑制过氧化氢诱导鼠脊髓星形胶质细胞凋亡

目的 探讨17-β雌二醇治疗脊髓损伤的可能机制,以期为临床急性脊髓损伤的治疗提供理论依据.方法 建立原代培养的星形胶质细胞凋亡模型,将培养的细胞按随机排序的方法分为5组:对照组、过氧化氢组、l7-β雌二醇+过氧化氢组、17-β雌二醇组、二甲基亚砜溶剂组.利用四甲基偶氮唑蓝比色法检测细胞活性,蛋白免疫印迹检测第10号染色体缺失的磷酸酶与张力蛋白同源物基因(PTEN)、磷酸化蛋白激酶B(p-Akt)、天冬氨酸特异性半胱氨酸蛋白酶3(caspase-3)和B细胞淋巴瘤/白血病-2( Bcl-2)蛋白的表达.结果 与对照组(0.401±0.043)比较,过氧化氢处理后(0.172 ±0.018)吸光度(A)值明显降低(q’=-1.450,P =0.001),可反映细胞生存率、活力降低,而17-β雌二醇预处理后(0.312±0.023),A值增加(q’=7.025,P=0.0025),提示细胞生存率增加及细胞活力上升,各组间比较A值差异也存在统计学意义;PTEN蛋白表达降低、Bcl-2表达上调;caspase-3表达降低、p-Akt表达上调.结论 17-β雌二醇有可能通过下调PTEN蛋白,促进p-Akt、Bcl-2蛋白的表达,抑制过氧化氢诱导星形胶质细胞凋亡.因此,脊髓损伤时早期应用17-β雌二醇有助于减轻脊髓损伤的程度,促进脊髓神经功能的恢复.     

罗格列酮抑制同型半胱氨酸诱导内皮细胞凋亡

目的 探讨罗格列酮(Rosiglitazone,Ros)对同型半胱氨酸(homocysteine,Hcy)所致的人脐静脉内皮细胞(HUVECs)凋亡的影响.方法 将原代培养的HUVECs分为对照组、Hcy刺激组和Ros干预组,MTT检测细胞存活率.免疫细胞化学方法检测Bax,Bcl-2,Caspase-3蛋白表达水平.采用Annexin V/PI染色加流式细胞仪检测细胞凋亡.比色法检测细胞培养上清液MDA和SOD含量.Western blot检测PPARγ蛋白表达水平.结果 与对照组比较,Hcy组细胞存活率下降;Bax和Caspase-3表达上调,Bcl-2 和PPARγ表达下调,MDA含量增加和SOD活性降低(P<0.05),而罗格列酮逆转上述变化(P<0.05).结论 罗格列酮可抑制Hcy所致的HUVECs凋亡.     

化疗性痛动物模型的建立及发病机制探讨

目的利用长春新碱(Vincristine)或紫杉醇(Paclitaxel)建立化疗性痛大鼠模型并探讨该模型的发病机制。方法利用微量释放泵连续给与SD大鼠一定量的长春新碱(Vincristine)或紫杉醇(Paclitaxel),而对照组给与相当量的溶剂。在注射前和注射后的连续时间点观察大鼠的痛觉行为学变化;当痛觉阈值降到最低点时,对处理组和对照组大鼠脊髓背角进行针对星形胶质细胞胶质纤维酸性蛋白(GFAP)和小胶质细胞OX42的免疫荧光染色,并鞘内给药抑制胶质细胞观察镇痛作用,同时取出两组大鼠的新鲜脊髓背角组织进行Western blot分析。结果长春新碱组注射后第6 d出现了明显的机械和热痛觉过敏,到注射后第8 d痛觉阈值降到最低点,以后基本维持在此水平;而紫杉醇组第4 d出现了明显的机械和热痛觉过敏,第12 d痛觉阈值降到最低点,以后基本维持在此水平。与对照组相比,处理组脊髓背角GFAP和OX42的染色密度出现了显著增强,鞘内给予针对星形胶质细胞的特异性抑制剂L-α-氨基己二酸(LAA)和小胶质细胞的特异性抑制剂米诺环素(minocycline)能起到有效镇痛作用。另外处理组脊髓细胞因子白介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)和磷酸化丝裂原活化蛋白激酶(MAPK)类分子磷酸化丝裂原活化蛋白激酶p38(p-p38)的表达显著上调。结论我们成功建立了化疗性痛模型,脊髓背角胶质细胞的激活有可能参与了化疗性痛的形成。     

小续命汤有效成分对慢性脑缺血大鼠氧化应激损伤及细胞凋亡的影响

目的 探讨小续命汤有效成分对慢性脑缺血大鼠氧化应激损伤及神经细胞凋亡的影响.方法 采用永久性结扎双侧颈总动脉法建立慢性脑缺血大鼠模型,将造模成功的大鼠按随机数字表法分为模型组、银杏叶提取物组和小续命汤有效成分低、中、高剂量组;另设假手术组作为对照.采用分光光度法检测各组大鼠脑组织超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)、过氧化氢酶(CAT)活性和丙二醛(MDA)含量的变化,TUNEL法检测神经细胞凋亡情况,Western blotting检测细胞凋亡相关蛋白caspase-3、Bcl-2、Bax的表达变化.结果 与模型组相比,低剂量和中剂量小续命汤有效成分可显著抑制慢性脑缺血大鼠长期缺血缺氧导致的氧化应激反应,表现为降低脂质过氧化水平及增强脑组织抗氧化酶GSH-Px活性,差异有统计学意义(P＜0.05),但对SOD和CAT活性无显著影响.与模型组相比,中剂量小续命汤有效成分可显著减少慢性脑缺血大鼠神经细胞的凋亡率,同时低剂量和中剂量小续命汤有效成分还可显著升高脑组织Bcl-2/Bax比值,抑制下游Caspase凋亡级联反应,差异均有统计学意义(P＜0.05).结论 小续命汤有效成分可显著抑制慢性脑缺血导致的氧化应激损伤,减少神经细胞凋亡.     

RNAi抑制星形胶质细胞Cx43对缺氧/再复氧后细胞凋亡的影响

目的探讨RNA干扰技术对星形胶质细胞Cx43蛋白的抑制作用以及对细胞缺氧/再复氧后凋亡的影响。方法化学合成针对大鼠Cx43基因的小干扰RNA(Small interfering RNA,siRNA)及其阴性RNA,通过脂质体转染体外培养原代星形胶质细胞,Western blot检测Cx43蛋白表达情况,MTT法检测siRNA转染对细胞的毒性作用;通过流式细胞术观察抑制Cx43蛋白对星形胶质细胞缺氧/再复氧后细胞凋亡情况的影响。结果 Western blot结果可见,转染siRNA后12h,Cx43蛋白表达开始抑制,48h抑制最明显,MTT结果显示siRNA组和脂质体组的细胞活性明显低于未转染组(P0.05),siRNA组与脂质体组未见明显区别(P0.05);流式细胞术示缺氧12h复氧不同时间,凋亡率逐渐增加,复氧48h凋亡率最高,siRNA干扰组明显低于正常对照组(P0.05)。结论 RNAi技术是研究Cx43功能的一种有效方法,siRNA转染能有效抑制星形胶质细胞Cx43蛋白表达,siRNA转染时,细胞毒性来自脂质体;siRNA抑制Cx43蛋白能够明显减轻星形胶质细胞缺氧/再复氧后的细胞凋亡。     

Sev对低氧缺糖诱导的脑神经元细胞损伤的影响

目的 研究七氟醚(Sev)调控Toll样受体4 (TLR4)/核转录因子κB (NF-κB)信号通路抑制低氧缺糖对脑神经元细胞损伤的保护作用。方法 将大鼠皮质神经元细胞分为对照组、模型组、1.0MAC和2.0 MAC七氟醚组;细胞计数试剂盒(CCK-8)法、流式细胞术检测细胞增殖和凋亡。试剂盒检测丙二醛(MDA)含量以及超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性。酶联免疫吸附测定法(ELISA)检测肿瘤坏死因子(TNF-α)、白细胞介素-6 (IL-6)、白细胞介素-8 (IL-8)水平;蛋白印迹法(Western blot)检测Bcl-2相关X蛋白(Bax)、B细胞淋巴瘤/白血病-2 (Bcl-2)、TLR4、磷酸化核因子κB抑制蛋白α (p-IκBα)和核因子κB抑制蛋白α (IκBα)蛋白表达。结果 与对照组比较,模型组细胞存活率、SOD、CAT活性降低(^均P<0.05),凋亡率、MDA含量、TNF-α、IL-6、IL-8水平升高(P<0.05),TLR4、p-IκBα蛋白表达升高;与模型组比较,Sev显著升高细胞存活率、SOD、CAT活...     

Gap19介导Cx43半通道抑制JAK2/STAT3通路活化对急性脑梗死大鼠的保护作用

目的探究Gap19通过介导Cx43半通道活性和JAK2/STAT3通路对急性脑梗死(ACI)大鼠的影响。方法 24只SD大鼠随机分为3组:对照组、MCAO组和MCAO+Gap19组,每组8只。建立ACI大鼠模型(MCAO)和氧葡萄糖剥夺(OGD)体外模型。TTC染色检测大鼠脑梗死面积;mNSS法检测大鼠神经功能缺损;CCK-8检测星形胶质细胞活力;Western blot检测NVU模型Cx43、Cleaved caspase-3、Bcl、Bax和JAK2/STAT3通路相关基因的蛋白表达;溴化乙锭摄取实验检测星形胶质细胞中半通道活性;免疫荧光法检测星形胶质细胞中pSTAT3和Cx43的共定位。结果 MCAO处理导致大鼠脑梗死面积和神经功能缺损均显著上调(P 0.05),Gap19处理显著下调大鼠脑梗死面积和神经功能缺损(P 0.05)。OGD处理导致NVU模型Cx43的蛋白表达和半通道活性显著上调(P 0.05),细胞活力显著下调(P 0.05),促进细胞凋亡;Gap19处理显著上调OGD处理后NVU模型细胞活力(P 0.05),显著下调Cx43的蛋白表达和星形胶质细胞的半通道活性(P 0.05),抑制细胞凋亡,促进JAK2/STAT3通路活化,其中Cx43与pSTAT3存在共定位关系。结论 Gap19通过抑制Cx43半通道和活化JAK2/STAT3通路发挥对ACI大鼠的保护作用。     

实验性变态反应性脑脊髓炎大鼠星形胶质细胞的变化

目的观察实验性变态反应性脑脊髓炎(EAE)大鼠脊髓中星形胶质细胞的变化,探讨EAE大鼠的发病相关生物学机制。方法采用免疫组化法,对豚鼠全脊髓匀浆诱导的Wistar大鼠EAE的过程中,脊髓内星形胶质细胞变化情况进行研究。结果EAE大鼠症状高峰期时星形胶质细胞开始激活,恢复期时激活达到高峰,而且活化的星形胶质细胞未见表达主要组织相容性抗原(MHC)。结论活化的星形胶质细胞可能与EAE大鼠的恢复有关。     

睫状神经营养因子对坐骨神经切断吻合后大鼠脊髓前角胶质纤维酸性蛋白

背景：睫状神经营养因子具有多种生物活性,在神经系统发育、分化和损伤修复中具有重要意义。 目的：观察睫状神经营养因子对坐骨神经切断吻合后大鼠相应脊髓节段前角星形胶质细胞的特异标记物胶质纤维酸性蛋白表达的影响。 方法：将SD大鼠随机分为对照组、模型组、生理盐水组及药物组。除对照组外,对所有大鼠实施双侧坐骨神经切断吻合术,药物组手术区局部注射睫状神经营养因子100 ng/kg,1次/d,生理盐水组局部注射等量生理盐水。术后1,3,7,14,21,28 d取相应脊髓节段,免疫组织化学染色观察胶质纤维酸性蛋白的表达,苏木精-伊红染色、TUNEL染色对脊髓前角神经元进行计数。 结果与结论：大鼠坐骨神经切断吻合后相应脊髓节段星形胶质细胞胞体大,突起分枝多且粗大,神经元数目逐渐减少,凋亡神经元增多,胶质纤维酸性蛋白表达增高。与模型组和生理盐水组比较,药物组神经元存活数目增多,凋亡减少,胶质纤维酸性蛋白表达明显增加(P < 0.05或P < 0.01)。同时,药物组大鼠的运动功能障碍较轻,恢复较快。说明睫状神经营养因子可以通过促进大鼠脊髓前角胶质纤维酸性蛋白的表达起到神经保护作用。 关键词：胶质纤维酸性蛋白;睫状神经营养因子;星形胶质细胞;神经元凋亡;周围神经损伤     

吡格列酮对大鼠脑出血后脑组织细胞凋亡及自由基水平的影响

目的:探讨吡格列酮对大鼠脑出血的保护作用及可能作用机制。方法:雄性SD大鼠分为6组,干预组1造模前6 h给药(吡格列酮15 mg.kg-1.d-1,灌胃),造模后维持原剂量;干预组2造模前6 h给药,造模后停用;干预组3造模前不给药,造模后给药;干预组4造模前后均不给药;另设假手术组和正常大鼠组。各组大鼠术后72 h处死,检测脑组织中超氧化物歧化酶(SOD)及丙二醛(MDA)浓度,Western blot测定过氧化小体增殖剂激活型受体γ(PPARγ)及半胱氨酸蛋白-3(caspase-3)表达;苏木精-伊红染色及TUNEL染色观察脑水肿及细胞凋亡。结果:吡格列酮能显著减轻脑出血大鼠72 h后的脑组织水肿程度,减少凋亡细胞数量,增加SOD活性和PPARγ蛋白表达,抑制MDA水平及caspase-3蛋白表达,且脑出血前预防性给药的效果较出血后治疗的效果好。结论:吡格列酮可减轻大鼠脑出血性脑损伤,可能作用机制是增加脑组织中PPARγ蛋白表达及SOD活性,抑制自由基水平和细胞凋亡。     

Denervation study of synapses of organ of corti of old world monkeys

Fine structural characteristics of synapses in the spiral organ of Corti were examined, with reference to differences between inner and outer haircell systems, and to location of neurons of origin of efferent axons. Surgical interruption of crossed olivocochlear bundle, of vestibular nerve, of facial nerve, and excision of superior cervical ganglia were used to determine the pathways of efferent axons. Interruption of the vestibular nerve near the brainstem results in degeneration of all efferent terminals on outer hair cells. Mid-line lesions at, and caudal to, the facial colliculus result in degeneration of about half of these efferent terminals. Efferent synaptic bulbs to the inner hair-cell system are small, of the order of one micron, and form type 2 junctions with afferent dendrites. They tend to have more large dense-core vesicles (about 80 nm) than the large efferent terminals of the outer hair-cell system, and appear to be the terminals of axons in the habenula perforata, which exhibit varicosities laden with large dense core vesicles. The varicosities are unaffected by excision of the superior cervical ganglia. So far as our material can reveal, it appears that the varicosities in the habenula perforata do not survive vestibular root interruption, nor do the efferent processes in the internal spiral bundle or at the base of inner hair cells. Most interestingly, the afferent processes of the inner hair-cell system, as identified for example by their relation to pre-synaptic bodies in the inner hair cells, are subject to a trans-synaptic reaction after severance of the vestibular root. They undergo a dramatic cytological transformation, characterized by increase of volume, engorgement with microtubules, microfilaments, microvesicles of various sizes, and clusters of lysosomes. Thus, both the efferent and afferent terminals of the inner hair-cell system show marked cytological differences from the corresponding terminals of the outer hair cell system.     

Mechanism of action of tubocurarine on nicotinic cholinoreceptors of neurons of the sympathetic ganglia of rats

Tubocurarine (Tc) effect on membrane currents elicited by acetylcholine (ACh) was studied in isolated superior cervical ganglion neurons of rat using patch-clamp method in the whole-cell recording mode. The "use-dependent" block of ACh current by Tc was revealed in the experiments with ACh applications, indicating that Tc blocked the channels opened by ACh. Mean lifetime of Tc-open channel complex, tau, was found to be 9.8 +/- 0.5 s (n = 7) at -50 mV and 20-24 degrees C. tau exponentially increased with membrane hyperpolarization (e-fold change in tau corresponded to the membrane potential shift by 61 mV). Inhibition of the ACh-induced current by Tc (3-30 microM/1) was completely abolished by membrane depolarization to the level of 80-100 mV. Inhibition of ACh-induced current was augmented at increased ACh doses. It is concluded that the open channel block produced by Tc is likely to be the only mechanism for Tc action on nicotinic acetylcholine receptors in superior cervical ganglion neurons of rat.     

The effect of age of onset of PD on risk of dementia

Background Dementia occurs in the majority of patients with Parkinson’s disease (PD). Late onset of PD has been reported to be associated with a higher risk for dementia. However, age at onset (AAO) and age at baseline assessment are often correlated. The aim of this study was to explore whether AAO of PD symptoms is a risk factor for dementia independent of the general effect of age. Methods Two community-based studies of PD in New York (n = 281) and Rogaland county, Norway (n = 227) and two population-based groups of healthy elderly from New York (n = 180) and Odense, Denmark (n = 2414) were followed prospectively for 3–4 years and assessed for dementia according to DSM-IIIR. All PD and control cases underwent neurological examination and were followed with neurological and neuropsychological assessments. We used Cox proportional hazards regression based on three different time scales to explore the effect of AAO of PD on risk of dementia, adjusting for age at baseline and other demographic and clinical variables. Findings In both PD groups and in the pooled analyses, there was a significant effect of age at baseline assessment on the time to develop dementia, but there was no effect of AAO independent of age itself. Consistent with these results, there was no increased relative effect of age on the time to develop dementia in PD cases compared with controls. Interpretation This study shows that it is the general effect of age, rather than AAO that is associated with incident dementia in subjects with PD. Received in revised form: 22 December 2005     

Limits of deinstitutionalization--perspectives of relatives of psychiatric patients

After a hopeful beginning, the social process of the reintegration of those with severe mental illness has come to a standstill. I am led to wonder whether "the community" really wants to live together with people suffering from severe mental illness, and if so, how closely? As long as the medical treatment of mental illness provided by the general practitioners is fundamentally deficient, as they are not able to prescribe the necessary interventions--such as out-patient psychiatric nursing, and service providers in the out-patient sector are content with offering increasingly intensive forms of care for the less seriously ill at the cost of the Social Welfare System--the reintegration of those with serious mental illness remains an illusion--which is mainly to the benefit of providers of residential care in homes and hostels.