环维黄杨星D对高血压大鼠脑缺血GAP-43与Neurocan表达的影响

目的 观察中药单体环维黄杨星D(CVB-D)对易卒中型肾血管性高血压大鼠(RHRSP)脑缺血一复流不同时间脑组织生长相关蛋白43(GAP-43)与神经粘蛋白(Neurocan)表达的影响.方法 采用环形银夹使SD大鼠双侧肾动脉狭窄,制成RHRSP,再用线栓法制成一侧大脑中动脉闭塞(MCAO)模型.用免疫组化方法观察CVB-D对MCAO大鼠脑缺血2h后复流1、7、14、30d脑组织GAP-43与Neurocan表达的影响,并与生理盐水组对照.结果 缺血2h后再灌注1d,对照组缺血周围半暗区出现GAP-43阳性细胞,7d明显增多,14d减少,30d明显减少,各时间点阳性细胞数表达差异有显著性意义(P＜0.01);治疗组GAP-43阳性细胞数表达在各时间点较对照组显著增加(P＜0.01).Neuroean阳性细胞数表达对照组在缺血再灌注ld出现,7d明显增多,14d达高峰,30d时下降,但仍高于假手术组水平(P＜0.05);治疗组neurocan阳性细胞数表达在缺血再灌注7、14、30d则较对照组显著减少(P＜0.01).结论 CVB-D上调RHRSP脑缺血区GAP-43阳性细胞数表达与下调Neurocan表达的作用,可能是其促进脑损伤区中枢神经修复的重要机制之一.     

脂肪来源干细胞移植对脑缺血大鼠运动功能的影响

目的 观察脂肪来源干细胞(ADAS)移植入脑缺血大鼠后存活、迁移、分化以及大鼠的运动功能障碍恢复情况,探讨ADAS移植治疗大鼠局灶性脑缺血的有效性和可能机制. 方法 将雄性SD大鼠饲养至250～300 g时,制作左侧大脑中动脉阻塞模型(MCAO),按照随机数字表法分为未处理组、对照组和移植组,每组6只.未处理组造模后不作特殊处理,对照组在造模后3h通过尾静脉注射杜氏改良培养基(DMEM),移植组造模后3 h通过尾静脉注射ADAS.造模后14 d处死大鼠,通过免疫荧光染色观察5-溴脱氧尿嘧啶核苷(BrdU)、神经元特异性烯醇化酶(NSE)、人微管相关蛋白2(MAP-2)和神经胶质纤维酸性蛋白(GFAP)的表达.造模后1、7及14 d时神经功能缺损评分评价大鼠运动功能改善情况. 结果 (1)移植后,标记了BrdU的ADAS大量出现在缺血灶周围;(2)MCAO后14d,缺血灶周围出现了少量BrdU/GFAP双染阳性细胞;同时出现少量BrdU/NSE和BrdU/MAP-2双染阳性细胞;(3)14d时移植组大鼠神经功能缺损评分与对照组相比明显降低,差异有统计学意义(P＜0.05). 结论 (1)成年大鼠ADAS在MCAO大鼠体内存活并少量分化为神经元样细胞和星形胶质细胞样细胞;(2)移植ADAS可使大鼠脑缺血所致的运动功能缺损得到改善.     

脑源性神经营养因子基因修饰的神经干细胞对脑卒中大鼠认知功能的机制研究

目的 探究脑源性神经营养因子(BDNF)基因修饰神经干细胞对脑卒中大鼠认知功能的机制。方法 体外分离并培养神经干细胞(NSCs),Nestin染色鉴定NSCs;BDNF修饰NSCs,并采用蛋白印迹法检测BDNF蛋白表达。将40只大鼠随机分为4组,即假手术组(sham组)、卒中组(MCAO组)、卒中大鼠给予NSCs组(NSCs组),卒中大鼠给予BDNF-NSCs组(BDNF-NSCs组),每组10只。对大鼠进行神经功能缺损评分,采用Morris水迷宫实验检测大鼠认知功能;苏木精-伊红染色法检测神经元损伤;TTC检测脑组织梗死面积;蛋白印迹法检测脑组织中胞内磷脂酰肌醇激酶、人磷酸化磷脂肌醇3激酶、苏氨酸蛋白激酶、磷酸化蛋白激酶蛋白表达。结果 免疫荧光细胞化学染色鉴定NSCs,培养的NSCs中巢蛋白和溴脱氧尿苷均呈阳性表达,提示所提取的NSCs为神经干细胞且具有增殖能力。和未转染组和阴性对照组相比,转染BDNF组BDNF蛋白表达明显增加(P<0.05)。和sham组相比,MCAO组大鼠神经功能评分、逃避潜伏期、脑梗死面积明显增加,穿越平台次数和脑组织中PI3K、p-PI3K、Akt、p...     

胎鼠神经干细胞移植对大鼠脊髓损伤后神经细胞凋亡及凋亡抑制基因Bcl-2表达的影响

目的研究胎鼠神经干细胞(NSCs)移植对大鼠脊髓损伤(SCI)后神经细胞凋亡及凋亡抑制基因Bcl-2表达的影响。方法 40只SD大鼠随机分为正常对照组(Normal组),脊髓损伤组(SCI组),神经干细胞组(NSC组),神经干细胞标记组(BrdU+NSCs组)。采用电控脊髓损伤打击装置制作模型,5-溴脱氧尿嘧啶核苷(Br-dU)法标记处于对数生长期的NSCs,SCI后即刻进行NSCs移植。免疫组化法观察BrdU标记NSCs的存活、迁移及凋亡抑制基因Bcl-2的表达,TUNEL法标记凋亡细胞(免疫组化及免疫荧光显色),改良Rivlin法观察大鼠后肢运动功能的恢复情况。结果 BrdU+NSCs组在损伤脊髓区域可检测到BrdU标记的阳性NSCs。BrdU+NSC组与NSC组各时间点凋亡阳性细胞数均比SCI组减少(P<0.01),Bcl-2免疫阳性细胞光密度值比SCI组明显增加(P<0.01),且Bcl-2表达高峰延长至伤后7d;移植后7d、14d、28d后肢运动功能评分较SCI组明显升高(P<0.01)。Br-dU+NSC组与NSC组之间比较无明显差异(P>0.05)。结论体外培养的胚胎大鼠NSCs可在脊髓损伤区域存活、迁移,并能通过上调Bcl-2的表达来抑制大鼠脊髓损伤后神经细胞的凋亡,从而促进大鼠瘫痪肢体功能的恢复。     

转录因子X盒结合蛋白1转染对神经干细胞在缺氧环境下grp78、EDEM表达及抗凋亡能力的影响

目的将X盒结合蛋白1基因以重组腺病毒为载体转染胚鼠海马神经干细胞,观察其在缺氧环境下是否可以使移植后干细胞内grp78、EDEM表达增加以及对干细胞抗凋亡能力的影响。方法取孕16d SD大鼠胚鼠的海马组织进行神经干细胞的分离、克隆、nestin免疫荧光检测,以及传代和扩增。将重组腺病毒Ad-XBP1-EGFP质粒转染胚鼠海马神经干细胞,得到基因修饰后的胚鼠海马神经干细胞。选取未转染的神经干细胞标记为对照组、未转染组;选取转染后的神经干细胞标记为转染组。未转染组和转染组用CoCl2诱导缺氧,24h后WesternBlot检测3组神经干细胞中xbp-1,grp78,EDEM,bcl-2及bax表达,流式细胞术检测NSCs和XBP1-NSCs的凋亡情况。结果在缺氧条件下,与未转染组相比,转染组的grp78表达增加,EDEM表达水平上升,bcl-2水平上升,而bax水平下降(P0.05);转染组神经干细胞凋亡程度较未转染组减轻(P0.05)。结论可以利用重组腺病毒作为载体成功将X盒结合蛋白1基因导入胚鼠海马神经干细胞中;转染后的神经干细胞在缺血缺氧条件下grp78、EDEM水平上升,抗凋亡能力较普通神经干细胞明显增加。     

骨髓基质细胞源神经干细胞对大鼠局灶性脑缺血神经细胞凋亡及相关蛋白表达的动态影响

目的探讨骨髓基质细胞源神经干细胞对大鼠局灶性脑缺血神经细胞凋亡及相关蛋白表达的影响。方法建立大鼠大脑中动脉缺血再灌注模型。32只健康Sprague-Dawley(SD)大鼠分为假手术组、缺血对照组、缺血骨髓基质细胞移植组和缺血骨髓基质细胞源神经干细胞移植组。分别在移植后7d和14d行脑灌注固定取材,应用免疫组化染色及原位细胞凋亡检测脑组织Bcl-2、Bax蛋白表达及凋亡细胞数。结果缺血移植组各时点的凋亡细胞数均少于缺血对照组(P<0.01),缺血移植14d组凋亡细胞数明显少于缺血移植7d组(P<0.01),骨髓基质细胞源神经干细胞移植组凋亡细胞明显少于骨髓基质细胞移植组(P<0.05)。缺血移植组Bcl-2表达显著高于缺血对照组(P<0.01)。缺血移植组Bax蛋白表达明显低于缺血对照组(P<0.01)。结论骨髓基质细胞源神经干细胞可能通过上调Bcl-2蛋白表达,下调Bax蛋白表达,对脑缺血再灌注损伤起保护作用。     

运动训练对大鼠脑缺血再灌注后神经修复及GAP-43和Neurocan表达的影响

目的 观察运动训练对大鼠脑缺血再灌注后不同时间神经修复及GAP-43与Neurocan表达的影响.方法 健康雄性Wistar大鼠72只,随机分成运动训练组、对照组、假手术组.采用线栓法制作一侧大脑中动脉闭塞(MCAO)模型,以神经功能缺损评分和Morris水迷宫试验进行神经功能评价,免疫组化法观察对脑缺血周围GAP-43与Neuorcan的表达.结果 与对照组比较,脑缺血再灌注后14d、21d,运动组的肢体运动及记忆功能明显恢复;缺血再灌注后7d,对照组缺血周围出现GAP-43阳性细胞,14d减少,21d、28d明显减少,运动组GAP-43表达在14d、21d、28d较对照组显著增加(P<0.05).Neurocan阳性细胞在对照组缺血再灌注7d出现,14d达高峰,21d、28d时下降;运动组Neurocan表达在缺血再灌注14d、21d、28d较对照组显著减少(P<0.05).结论 运动训练上调大鼠脑缺血区GAP-43表达与下调Neurocan表达,可能是其促进脑损伤区中枢神经修复的重要机制之一.     

移植时间对大鼠视神经损伤后神经干细胞迁移的影响

目的探讨移植时间对大鼠视神经损伤后神经干细胞(NSCs)在视网膜内迁移的影响。方法 18只SD大鼠行右侧视神经损伤后按随机数字表法随机平均分为3组,并分别于损伤后当天及伤后7d和14d采用微量注射法行右眼视网膜下腔移植2μl(105个/μl)转染绿色荧光蛋白基因的NSCs(GFP-NSCs)。4周后处死大鼠,取右眼球做冰冻切片,对迁移到视网膜内的移植细胞进行计数。各组切片分别用胸腺细胞表面糖蛋白、胶质纤维酸性蛋白、β微管蛋白、视紫红质抗体进行免疫标记,观察各组移植细胞在视网膜上的分化情况。结果损伤当天及伤后7和14d移植组视网膜上的GFP阳性细胞数分别为(163441.14±16876.81)个、(145736.57±27449.07)个和(117291.33±15849.19)个,两两相较,均差异显著(P<0.01)。三组的移植细胞均可在宿主视网膜内存活、迁移,且可分化为神经胶质细胞、神经元和视网膜神经节样细胞。结论大鼠视神经损伤后随着移植时间的推迟,迁移到宿主视网膜内的移植细胞量下降。     

环维黄杨星D对高血压大鼠脑缺血GAP-43 mRNA表达与细胞损伤的影响

目的观察中药单体环维黄杨星D(CVB-D)对易卒中型肾血管性高血压大鼠(RHRSP)脑缺血再灌注不同时间脑组织生长相关蛋白-43(GAP-43)mRNA表达与细胞超微结构损伤的影响。方法采用环形银夹使SD大鼠的双侧肾动脉狭窄,制成RHRSP,再用线栓法制成一侧大脑中动脉闭塞(MCAO)模型。用原位杂交等方法观察CVB-D对脑缺血2h后复流1d、7d、14d、30d不同时间点大鼠脑组织GAP-43mRNA表达、水含量、梗死面积百分率、行为学评分及细胞超微结构的干预作用。结果脑缺血2h复流后1d缺血区周围及海马可见GAP-43mRNA表达,7d明显增多至高峰,14d开始下降,30d时则明显减少,CVB-D治疗组在上述区域各时间点较对照组显著增加。脑缺血再灌注7d后,治疗组较对照组大鼠脑水含量及梗死面积显著降低,受损脑组织神经元和血管壁的超微结构亦明显改善。结论CVB-D对RHRSP缺血性脑细胞损伤有一定保护作用,其促进轴突的再生可能与上调脑组织GAP-43mRNA表达有关。     

神经干细胞在脑出血大鼠脑内移植后对IL-6、TNF-α表达的影响

目的 探讨神经干细胞(NSCs)移植入脑出血模型大鼠脑内后对脑组织内IL-6、TNF-α表达的影响.方法 体外培养大鼠NSCs;选取SD大鼠180只,随机取60只大鼠为健康对照组(A组),余120只采用Ⅳ型胶原酶诱导法建立脑出血模型,并随机分成两组:B组60只为单纯脑出血组;C组60只为脑出血+NSCs移植组,三组大鼠分别在NSCs移植后6 h、1 d、3 d、7 d、14 d进行肢体运动功能评分后,处死取脑组织匀浆,以EHSA方法分别检测三组大鼠脑组织内5个时间点IL-6、TNF-a的表达,并用统计学的方法分析.结果 与单纯脑出血组比较,脑出血+NSCs移植组能够显著降低IL-6、TNF-a表达(P＜0.05)和促进肢体功能恢复(P＜0.05).结论 大鼠NSCs脑内移植能有效降低脑出血模型大鼠脑内IL-6、TNF-a的表达,并能够改善运动神经功能缺损.     

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.