β-淀粉样蛋白25~35和人参皂苷Rb1对神经干细胞分化过程中tau蛋白磷酸化水平的影响

背景：通过调节神经干细胞分化过程中的tau蛋白磷酸化水平可提高神经干细胞移植治疗阿尔茨海默病的效果。 目的：观察β-淀粉样蛋白25~35和人参皂苷Rb1对神经干细胞分化过程中tau蛋白磷酸化水平的影响。 方法：分离、培养新生大鼠海马神经干细胞,诱导第3代神经干细胞分化1周后,分别采用免疫荧光细胞化学和Western-blot检测Tau[pS396] 、Tau[pS262] 及GSK-3β[pTyr279,216]表达情况。 结果与结论：正常神经干细胞分化过程中有Tau[pS396] 和Tau[pS262]的表达,β-淀粉样蛋白25~35可以使Tau[pS396]、Tau[pS262]和GSK-3β[pT279/216]表达上调,而人参皂苷Rb1可以逆转β-淀粉样蛋白25~35的作用。提示β-淀粉样蛋白25~35和人参皂苷Rb1通过调节GSK-3β的活性对tau蛋白的磷酸化水平产生调控作用。     

β-淀粉样蛋白诱导骨髓间充质干细胞向神经元样细胞分化过程中Tau蛋白的过度磷酸化★

背景：大多数学者认为β-淀粉样蛋白是阿尔茨海默病发病的始动因素，而Tau蛋白过度磷酸化可能是阿尔茨海默病最重要的分子病理变化之一。 目的：观察β-淀粉样蛋白25~35对大鼠骨髓间充质干细胞分化的神经元样细胞Tau蛋白磷酸化的影响。 方法：体外分离纯化大鼠骨髓间充质干细胞，将第4代骨髓间充质干细胞分为两组：实验组中加入预诱导培养基和20 μmol/L β-淀粉样蛋白25~35，24 h后用含2%二甲基亚砜和200 μmol/L丁羟茴醚的DMEM诱导其向神经细胞分化，5 h后收取细胞。对照组诱导方法同实验组，但在诱导过程中未加入β-淀粉样蛋白25~35。 结果与结论：光镜下可见纺锤状的骨髓间充质干细胞诱导后出现长突起，呈现神经细胞样形态，但实验组突起数量和长度均少于对照组；免疫细胞化学法检测两组诱导后的神经元样细胞为神经元特异性烯醇化酶阳性；免疫蛋白印记法证明实验组的GSK-3β、Tau[pSer262]和Tau[pSer396]表达均显著高于对照组。结果提示β-淀粉样蛋白25~35能够通过GSK-3β途径诱导骨髓间充质干细胞分化的神经元样细胞Tau蛋白过度磷酸化。     

Aβ诱导骨髓间充质干细胞对其分化的神经细胞Tau蛋白过度磷酸化的影响

目的研究β-淀粉样蛋白25-35(Aβ25-35)对大鼠骨髓间充质干细胞(BMSCs)分化的神经细胞Tau蛋白磷酸化的影响。方法体外分离纯化大鼠骨髓MSCs,将P4 MSCs分为两组:(1)Aβ25-35实验组中加入预诱导培养基(含10μg/L bFGF,10%FBS的DMEM)和20μmol/L Aβ25-35,24h后用含2%DMSO和200μmol/L BHA的DMEM诱导其向神经细胞分化,5h后收取细胞;(2)对照组诱导方法同Aβ实验组,但在诱导过程中未加入Aβ25-35。光镜下观察细胞形态变化,免疫细胞化学法检测神经神经元特异性烯醇化酶(NSE),western blotting检测GSK-3β、Tau[pSer262]和Tau[pSer396]。结果光镜下可见纺锤状的MSCs诱导后出现长突起,呈现神经细胞样形态,但Aβ25-35实验组突起数量和长度均少于对照组;免疫细胞化学法检测两组诱导后的神经元样细胞为NSE阳性;免疫蛋白印记法证明Aβ25-35实验组的GSK-3β、Tau[pSer262]和Tau[pSer396]表达均显著高于对照组。结论 Aβ25-35能够通过GSK-3β途径诱导骨髓间充质...     

亚低温抑制重型颅脑创伤诱导Tau蛋白过度磷酸化的实验研究

目的 观察亚低温对重型颅脑创伤诱导的Tau蛋白磷酸化的影响.方法 体外研究通过细胞液压冲击仪建立SK-N-SH细胞损伤模型,观察冲击1h、6h和12h后Tau蛋白的磷酸化水平.体内研究通过小动物液压冲击仪建立大鼠重型颅脑损伤模型(2.4 atm),接受或不接受亚低温治疗(33℃,4h).观察液压冲击6h、24 h和72 h后Tau蛋白的磷酸化水平,以及亚低温对此变化的影响.结果 SK-N-SH细胞在液压冲击后,Tau蛋白在Thr231和Ser396位点的磷酸化水平从1h开始升高,到6h左右达到高峰.SD大鼠在液压冲击后,Tau蛋白在Thr231和Ser396位点的磷酸化水平从6h开始升高,到24h左右达到高峰,到72 h仍处于高水平;免疫组化的结果也显示,大鼠脑部海马CA1区和皮质区的Tau蛋白磷酸化水平在Thr231位点也明显增高.亚低温可显著逆转液压冲击诱导的SD大鼠Tau蛋白过度磷酸化(P＜0.01).结论 亚低温可抑制重型颅脑创伤诱导的Tau蛋白过度磷酸化.     

人羊膜间充质干细胞移植对阿尔茨海默病转基因小鼠的行为学和β-淀粉样蛋白的影响

背景：研究发现APP基因与阿尔茨海默病发病密切相关。β-淀粉样蛋白是阿尔茨海默病患者脑内老年斑的主要成分，基因突变和外界环境的影响能破坏β-淀粉样蛋白的动态平衡，从而引发或加速阿尔茨海默病的产生和发展。 目的：探讨人羊膜间充质干细胞尾静脉移植对阿尔茨海默病转基因小鼠学习记忆能力及脑组织β-淀粉样蛋白表达的影响。 设计、时间及地点：随机对照动物实验，于2008-05/10在郑州大学微生物学与免疫学教研室和河南省中医药治疗研究院完成。 材料：健康剖宫产产妇志愿捐献的羊膜，由郑州大学第一附属医院产科提供。APP转基因鼠29只，PCR技术鉴定转APP-基因小鼠9只，作为正常组；转APP+基因小鼠20只，随机分为细胞移植组、对照组，10只/组。 方法：无菌条件下体外分离培养人羊膜间充质干细胞，传至第3代将细胞浓度调整为1×109 L-1，经尾静脉注入0.5 mL至细胞移植组小鼠体内；对照组经尾静脉注入同体积的生理盐水；正常组小鼠不给予任何干预措施。 主要观察指标：采用Morris水迷宫测定小鼠逃避潜伏期、穿越平台次数及在平台象限的时间，刚果红染色观察小鼠脑组织内β-淀粉蛋白的表达。 结果：定位航行试验中，移植前与正常组比较，细胞移植组、对照组小鼠逃避潜伏期差异均有显著性意义（P < 0.05）；移植后2周细胞移植组小鼠逃避潜伏期与正常组基本相似（P > 0.05），但明显短于对照组（P < 0.05）。空间探索试验中，移植前后小鼠穿越平台次数及其在平台象限的时间3组间比较差异均无显著性意义（P > 0.05）。移植后1个月，正常组未见或仅见极少量的β-淀粉样蛋白沉积，细胞移植组淀粉样蛋白的沉积明显少于对照组。 结论：人羊膜间充质干细胞尾静脉移植能促进阿尔茨海默病转基因小鼠空间定位及学习记忆能力的提高，且减少小鼠脑内β-淀粉样蛋白的沉积。     

人羊膜间充质干细胞静脉移植治疗转基因APP+小鼠的机制***◆

背景：研究发现APP基因与阿尔茨海默病发病密切相关。课题组前期实验发现人羊膜间充质干细胞静脉移植可以促进阿尔茨海默病APP+转基因小鼠的学习,记忆能力改善。 目的：探讨人羊膜间充质干细胞尾静脉移植后,能否归巢到小鼠脑组织中,并分化为相关的中枢神经细胞,治疗阿尔茨海默病。 方法：无菌条件下体外分离培养人羊膜间充质干细胞,传至第3代将细胞浓度调整为1×109 L-1,经尾静脉注入0.5 mL至细胞移植组转APP+基因小鼠体内;对照组经尾静脉注入同体积的生理盐水;正常组转APP-基因小鼠不给予任何干预措施。采用免疫组化方法检测、测定Brdu标记的人第3代羊膜间充质干细胞在小鼠脑组织中的表达,各组小鼠脑组织中GFAP、Nestin和NSE的表达。 结果与结论：光镜下观察可见,移植组小鼠脑组织中大部分细胞核被染成蓝色,但有一些细胞核被染成棕色或褐色,Brdu呈阳性。与对照组相比,移植组小鼠脑组织中的GFAP增加了近4倍,表达显著增加,甚至超过正常组的表达(P < 0.05);Nestin的表达则显著升高,增加了近10%,但是还比正常组低了近20%(P < 0.05)。NSE的表达降低了近1/3,但仍高于正常组(P < 0.05)。表明人羊膜间充质干细胞尾静脉移植治疗阿尔茨海默病可能是通过羊膜间充质干细胞归巢到转基因APP+小鼠脑组织中,并分化成中枢神经细胞的途径实现的。 关键词：人羊膜间充质干细胞;转基因APP+小鼠;尾静脉移植;胶质纤维蛋白;巢蛋白;神经元特异性烯醇化酶     

厄贝沙坦对Alzheimer病大鼠海马Tau蛋白磷酸化的影响

目的研究厄贝沙坦对Alzheimer病(AD)大鼠海马Tau蛋白磷酸化的影响。方法侧脑室注射寡聚体形式的β淀粉样蛋白(Aβ)1-42制作AD大鼠模型;自术前1周至术后2周(手术当天暂停给药1次),厄贝沙坦干预组给予厄贝沙坦30mg/(kg.d)灌胃,AD模型组给予等量生理盐水灌胃。用Morris水迷宫试验检测学习记忆能力,Westernblot法检测海马Tau蛋白及其丝氨酸199/202位点、396位点磷酸化水平;分析软件测定Westernblot图像条带光密度(OD)值。并与正常对照组和假手术组比较。结果AD模型组Morris水迷宫试验逃避潜伏期比正常对照组和假手术组长(均P<0.05),厄贝沙坦干预组与正常对照组和假手术组差异无统计学意义。4组间海马Tau蛋白总量差异无统计学意义;与AD模型组比较,厄贝沙坦干预组丝氨酸199/202和396位点磷酸化Tau蛋白水平明显降低(均P<0.05);而与正常对照组和假手术组差异无统计学意义。结论厄贝沙坦能减轻AD大鼠海马Tau蛋白磷酸化水平。     

人羊膜间充质干细胞静脉移植治疗阿尔茨海默病APP+转基因鼠的

目的：评估人羊膜间充质干细胞静脉移植治疗阿尔茨海默病APP转基因鼠的有效性和安全性。 方法：饲养转基因APP+胎鼠,定期合笼,利用PCR技术快速鉴定转基因鼠,以刚果红染色法观察脑组织切片中的β-淀粉样蛋白沉积。严格无菌条件下取正常足月剖腹产胎儿的羊膜,体外分离培养羊膜间充质干细胞,至第3代经尾静脉移植。Morris水迷宫测试移植前后小鼠学习、记忆能力。移植当天开始称重,每隔三天一次,持续三周。移植后解剖小鼠,收集全血并分离血清,进行心、肝、肾功能的血液生化和12项肿瘤标记物检测,对该细胞治疗的安全性作综合评价。 结果：移植后小鼠学习、记忆能力明显增强。移植组动物的体重增长趋势与对照组相比无明显差异（P＞0.05）,未见肿瘤和死亡发生。血液生化和12项肿瘤标记物各指标无明显差异（P＞0.05）。 结论：人羊膜间充质干细胞尾静脉移植治疗阿尔茨海默氏症小鼠明显促进小鼠学习、记忆能力,未见致死致瘤现象发生,心、肝、肾功能未受影响。利用该种干细胞移植治疗阿尔茨海默氏病是安全可行的。     

面神经损伤大鼠脑内移植绿色荧光蛋白转基因胎鼠神经干细胞的

背景：目前面神经损伤后的修复主要集中在外周神经干,但面神经损伤后会导致部分中枢运动神经元凋亡。现阶段关于干细胞植入面神经损伤大鼠脑后对面神经核团内凋亡神经元的影响相关报道甚少。 目的：观察大鼠面神经损伤后,脑内移植绿色荧光蛋白转基因胎鼠的神经干细胞的成活和迁移情况。 设计、时间及地点：随机对照动物实验,于2008-07/12在昆明医学院神经科学研究所完成。 材料：孕14~16 d的绿色荧光蛋白转基因蛋白小鼠1只,用于制备转基因神经干细胞。清洁级SD雄性大鼠24只,随机分为3组：面神经损伤转基因细胞组12只、面神经损伤细胞培养液组6只、面神经正常转基因细胞组6只。 方法：面神经损伤转基因细胞组、面神经损伤细胞培养液组大鼠建立面神经切断模型。造模后1周,行转基因神经干细胞立体定向移植,注射点为前囟后方11.30 mm、背侧9.00 mm、正中线位置。面神经损伤转基因细胞组、面神经正常转基因细胞组各注入10 μL转基因神经干细胞悬液(含5×106 个细胞),面神经损伤细胞培养液组注入10 μL神经干细胞培养液,4周后制作脑组织冰冻切片。 主要观察指标：荧光显微镜观察移植部位绿色荧光蛋白阳性神经干细胞的存活情况,及其向损伤侧面神经核团周围迁移的情况。 结果：①移植处：面神经损伤转基因细胞组、面神经正常转基因细胞组均可见数量不等的绿色荧光蛋白阳性细胞,其中部分绿色荧光蛋白阳性细胞位于血管内;面神经损伤细胞培养液组未见绿色荧光蛋白阳性细胞。②面神经核团周围：仅面神经损伤转基因细胞组可见数量不等的绿色荧光蛋白阳性细胞迁移于损伤侧面神经核团周围,而健侧面神经核周围未见绿色荧光蛋白阳性细胞;余2组双侧面神经核周围均未见绿色荧光蛋白阳性细胞。③移植处与面神经核团周围之间：未见绿色荧光蛋白阳性细胞相连。 结论：神经干细胞移植入面神经损伤大鼠脑内后,可以向损伤侧面神经核周围迁移。     

VPA对OA处理的SH-SY5Y细胞tau蛋白磷酸化水平的影响

目的研究丙戊酸钠(VPA)对冈田酸(OA)处理的人神经母细胞瘤系(SH-SY5Y)细胞中tau蛋白磷酸化的影响。方法 MTT观察OA对SH-SY5Y细胞活力的影响,制备tau蛋白过度磷酸化的细胞模型;Western blot检测OA及VPA对Thr231位点tau蛋白磷酸化的影响。结果 MTT结果显示,当OA浓度大于40 nmol/L时,细胞活力受到明显抑制,而低于此浓度的OA对细胞活力的影响不明显;Western blotting结果显示,SH-SY5Y细胞经OA(40 nmol/L,12 h)处理后,Tau蛋白磷酸化的水平明显增加,给予VPA(10 mmol/L)作用12 h后,Tau蛋白磷酸化的水平明显下降。结论 VPA可以抑制OA处理的SH-SY5Y细胞的tau蛋白过度磷酸化。     

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.     

Sense of coherence as a predictor of subjective state of health: results of 4 years of follow-up of adults

A number of cross-sectional population studies have shown that a strong sense of coherence (SOC) is associated with various aspects of good perceived health. The association does not seem to be entirely attributable to underlying associations of SOC with other variables, such as age or level of education. OBJECTIVE: The aim of the study reported here was to determine whether SOC predicted subjective state of health. METHODS: The study was carried out as a two-way panel mail survey of 1976 individuals with 4 years interval for two collections of data. The statistical method used was multivariate cumulative logistic modeling. Age, initial subjective state of health, initial occupational training level, and initial degree of social integration were included as potential explanatory variables. RESULTS: A strong SOC predicted good health in women and men. CONCLUSIONS: SOC can be interpreted as an autonomous internal resource contributing to a favorable development of subjective state of health. SOC data should, however, be regarded as complementary to and not a substitute for information already known to be associated with increased risk of future ill health.