帕金森病大鼠黑质小胶质细胞及星形胶质细胞的变化

目的探讨帕金森病(PD)发病中黑质小胶质细胞和星形胶质细胞的变化。方法采用立体定向术将神经毒素6-羟基多巴胺(6-OHDA)注入大鼠右侧黑质和内侧前脑束内,制备大鼠PD模型。将制模成功的16只PD大鼠随机分为2周和8周模型组,另6只正常大鼠作为对照组。观察各组大鼠黑质致密带内多巴胺(DA)能神经元、OX-42(小胶质细胞的特异性标志物)及神经胶质纤维酸性蛋白(GFAP,星形胶质细胞的特异性标志物)阳性细胞的分布和形态变化。结果 2周和8周模型组损毁侧黑质致密部DA能神经元较健侧显著减少(P0.01),损毁侧OX-42阳性细胞的数量较健侧明显增加(P0.01),形态呈"阿米巴状"。损毁侧GFAP阳性细胞数量较健侧明显增加(P0.01),突起变短,染色加深。2周模型组和8周模型组DA能神经元及两种胶质细胞的变化情况相似。结论 PD大鼠模型中存在着小胶质细胞和星形胶质细胞的激活,且两种胶质细胞的活化程度在PD发病过程中的不同时间无明显差别。     

大鼠坐骨神经分支选择性损伤后脊髓背角毛细血管和星形胶质细胞的反应

目的探讨成年大鼠坐骨神经分支选择性结扎切断(SNI)后,腰段脊髓背角内毛细血管和星形胶质细胞的反应。方法成年雄性SD大鼠40只,随机分为光镜组(n=30)和电镜组(n=10)。光镜组又分为对照组、假手术组和SNI手术组。SNI手术为分别结扎切断左侧胫神经和腓总神经,保留腓肠神经,存活20d。部分动物在术后经尾静脉给予30g/L伊文思蓝,切取脊髓L4平面,荧光显微镜观察伊文思蓝的渗出;另一部分在术后进行常规固定取材,切片进行抗鼠IgG和GFAP的免疫组织化学反应。电镜组(对照组和SNI后20d组),脊髓切片进行常规抗大鼠IgG的免疫电镜处理。结果 1.SNI后20d,可见明显的伊文思蓝渗出红色光斑;2.SNI后GFAP阳性细胞变为反应型,数量明显增加,毛细血管壁及其周围的IgG颗粒也显著增加。3.电镜观察:SNI术后20d,IgG阳性颗粒增多,内皮细胞观察到下列变化:(1)内皮细胞向管腔内伸出许多微细的突起,有的含有IgG阳性颗粒。(2)内皮细胞的管腔侧胞膜上有许多"内吞"小凹,凹内可见IgG阳性颗粒,胞质内有含IgG阳性颗粒的小泡,在反管腔侧胞膜上有"胞吐"现象。(3)内皮细胞紧密连接出现间隙,间隙内有IgG颗粒。(4)胞质内出现囊泡,有的泡内显示IgG阳性颗粒。结论 SNI后脊髓背角内毛细血管内皮细胞发生明显的反应,大鼠IgG和伊文思蓝可经过血脊髓屏障渗出,星形胶质细胞被激活。     

哮喘大鼠脑内小胶质细胞激活的时程及形态变化研究

为探讨哮喘大鼠在连续激发不同阶段脑内小胶质细胞激活的时程及形态变化,本实验取健康雄性SD大鼠制备哮喘模型并诱发哮喘发作。48只SD大鼠随机分为对照组和实验组。对照组分为正常组和生理盐水组(n=8);实验组根据造模期间连续抗原激发的时间不同,分为连续激发3,7,14,21d组(n=8)。各组大鼠均进行呼吸功能检测、肺组织切片HE染色及脑组织抗OX-42免疫组织化学染色。结果显示:与对照组大鼠比较,实验组中各组大鼠呼吸功能各项指标均明显加重;肺组织病变程度逐渐增加;小胶质细胞由静息状态转变为激活状态,且数量明显增多。以上结果提示:哮喘大鼠脑内小胶质细胞被激活,在一定时间内小胶质细胞激活的程度随呼吸功能的降低及肺组织病变程度的增加而增加。     

弥漫性轴索损伤后胶质细胞的反应性变化

目的 探讨大鼠弥漫性轴索损伤（DAI）后胶质细胞的反应性变化规律及与轴索继发损伤的相互关系。 方法 SD 成年大鼠随机分为对照组及DAI损伤1d、2d、3d、7d组,每组8只。参照Marmarou方法制做 DAI 模型,并对大鼠的脑干部位进行胶质纤维酸性蛋白（GFAP）、离子钙结合适配器分子1（Iba1）、少突胶质细胞系转录因子2（Olig 2）、CC-1、NG2免疫组织化学染色及TUNEL染色,并且通过透射电子显微镜进一步观察脑干超微结构变化。 结果 DAI大鼠损伤后3d脑干Iba1标记的小胶质细胞细胞数目显著增多,一直持续至伤后7d,且激活的小胶质细胞呈肥大样形态学改变;DAI大鼠伤后1周内脑干GFAP标记的星形胶质细胞数目虽有所增多,但并没有统计学意义;CC-1标记的成熟少突胶质细胞在DAI后1d即开始明显降低,且随损伤时间的延长而进一步降低。DAI大鼠脑干TUNEL标记的凋亡细胞随损伤时间延长持续增多,与成熟少突胶质细胞的丢失成正相关。Olig2标记的少突胶质细胞系表达在DAI损伤后1周内各时间点均明显增高,于损伤后3d达峰值。NG2标记的少突胶质细胞前体细胞数量于损伤后3d和7d显著增多。透射电子显微镜结果示,DAI大鼠损伤呈现由髓鞘到轴索的特点,最早表现为髓鞘松解,轴索完整;且脱髓鞘会进一步促进轴索骨架崩解,最终导致轴索变性。 结论 成熟少突胶质细胞对DAI损伤具有选择易感性,髓鞘脱失是轴索继发损伤的重要因素。少突胶质细胞前体细胞增殖,并伴随小胶质细胞的激活。探索胶质细胞的动态变化将为进一步解释轴索继发损伤的病理生理学机制奠定基础。     

罗格列酮对大鼠脑出血急性期小胶质细胞和星形胶质细胞的影响

目的:研究罗格列酮对大鼠脑出血后小胶质细胞、星形胶质细胞的影响。方法:雄性SD大鼠随机分为对照组、大剂量干预组、小剂量干预组和假手术组。采用Ⅳ型胶原酶肝素生理盐水尾状核立体定向注射法建立大鼠脑出血模型,大剂量干预组给予罗格列酮2 mg·kg~(-1)·d~(-1)灌胃,小剂量干预组给予罗格列酮0.2 mg·kg~(-1)·d~(-1)灌胃,连续7 d。在术前和术后每日进行1次Longa神经功能缺损评分和尾部微量血糖测量,在术后第2天和第7天采用免疫组织化学检测脑小胶质细胞和星形胶质细胞。结果:大剂量干预组在术后第4天到第7天,小剂量干预组在术后第5天到第7天的Longa神经功能缺损评分低于对照组;大剂量干预组在术后第4天到第7天,小剂量干预组在术后第6天和第7天的微量血糖低于对照组;大剂量干预组术后第2天和第7天血肿周边活化的小胶质细胞较对照组增加,小剂量干预组术后第7天血肿周边活化的小胶质细胞较对照组增加;大剂量干预组和小剂量干预组术后第7天星形胶质细胞较对照组明显增加。结论:罗格列酮可以部分改善大鼠脑出血急性期神经功能,这可能与调节小胶质细胞和星形胶质细胞激活和功能有关。     

活化小胶质细胞致星形胶质细胞激活

目的探讨活化小胶质细胞培养液对星形胶质细胞的影响。方法 LPS激活原代培养小胶质细胞,采用活化的小胶质细胞条件培养液刺激星形胶质细胞,观察星形胶质细胞GFAP及IL-1β和TNFα的表达。结果 LPS刺激后,小胶质细胞OX42表达量上升,IL-1β和TNFα的表达量增高;小胶质细胞条件培养液可致星形胶质细胞激活,GFAP表达量上升,IL-1β和TNFα的表达量增加。结论活化小胶质细胞的条件培养液可致星形胶质细胞激活,激活的小胶质细胞和星形胶质细胞表达前炎症介质IL-1β和TNFα。     

大鼠束缚后脑内星形胶质细胞的反应

为了探讨束缚后大鼠脑内星形胶质细胞的反应,本实验将大鼠束缚于小的塑料桶内1、3或6 h,于解束后30 min处死,正常大鼠不被束缚作为对照。脑组织进行抗glial fibrillary acidic-protein(GFAP)免疫组织化学染色。结果显示:在正常大鼠脑内有少数散在静止状态的星形胶质细胞,表现为胞体小、突起细、GFAP淡染,缺乏机能定位分布特点;束缚后脑内星形胶质细胞表现为胞体肥大、突起变粗、GFAP深染、形成激活状态,其分布有明显的机能定位特点,表达于与应激反应调节相关的核团,主要有(1)端脑:扣带回、新皮质(尤其是第Ⅲ和V层)、隔外侧核、杏仁中央核;(2)间脑:丘脑室旁核、外侧膝状体、内侧膝状体、下丘脑的视上核、室旁核、第三脑室室周区、弓状核;(3)脑干:中脑的上丘浅层、中脑导水管周围灰质、下丘的皮质部;脑桥的臂旁外侧核、蓝斑、A5区;延髓的耳蜗核、延髓内脏带(MVZ)等处。反应性星形胶质细胞表达的时程变化是束缚1 h最高,3 h次之,6 h最少。本文结果表明,大鼠被束缚后全脑多处核团的星形胶质细胞发生不同程度的改变,随着束缚时间的延长,动物产生适应性,星形胶质细胞表达减少。     

大鼠脊髓挤压伤后小胶质细胞Toll样受体4表达的变化

目的探讨脊髓挤压伤后各时间点小胶质细胞Toll样受体4(TLR4)的表达变化及其与损伤面积和免疫球蛋白G渗出范围的关系。方法 48只成年雄性SD大鼠建立T8节段脊髓挤压伤模型,随机分为6组,每组8只;假手术对照组及挤压伤组动物在手术后0h、3h、24h、72h和7d用4%多聚甲醛灌注固定,取以挤压点为中心的2cm脊髓,冷冻切片后进行HE染色和免疫荧光染色,分别观察损伤面积的变化,TLR4表达和TLR4阳性小胶质细胞的分布,以及与血浆免疫球蛋白G(IgG)渗出范围的比较,并用IPP6.0软件计算各组的阳性数目。结果脊髓损伤后TLR4主要表达在小胶质细胞,在3～24h之间开始表达,伤后在72h达到高峰,7d时已经显著下降。阳性小胶质细胞的分布与IgG渗出范围一致。结论大鼠脊髓挤压伤模型中,表达在脊髓小胶质细胞上的TLR4可在脊髓继发性损伤中发挥重要作用,并与血脊髓屏障开放有关。     

脑缺血后小胶质细胞和星形胶质细胞“交叉对话”的研究进展

<正>生理状态下,小胶质细胞保持静息状态,依靠较细长的分支感应大脑实质微环境,是中枢神经系统（central nervous system,CNS）应对损伤的前哨[1]。星形胶质细胞数目多于小胶质细胞,约占所有胶质细胞总数的一半,被认为是CNS中功能最多的胶质细胞[2]。缺血性脑卒中主要表现为神经炎症和血脑屏障（blood-brain barrier,BBB）功能障碍。     

显示小胶质细胞和少突胶质细胞镀银法的改良

在中枢神经系统的神经胶质细胞中,小胶质细胞和少突胶质细胞无论在正常的生理活动或损伤的修复过程,均有重要的作用。目前尚有一些未定论的问题需待解决。因此,对这两种细胞的研究也就显得尤为重要。虽然显示小胶质细胞和少突胶质细胞的方法较多,但这些方法在具体应用时常有一些不尽人意之处。为了满足教学和科研的要求,我们对多种显示方法进行了摸索和对比,并选定Penfield氏改进的Del Rio-Hor-lega氏法进行了重要的研究和改进。(1)在进入碳酸银溶液之前,用DAB(3.3’-diaminobenzidine)和H_2O_2处理;(2)增加碳酸银配方中碳酸钠的浓度至饱和;(3)将原法中碳酸银作用的时间和温度延长和提高。经过改良后的方法,稳定,显出率和成功率高,是显示小胶     

Comparison of blood-nerve barrier disruption and matrix metalloprotease-9 expression in injured central and peripheral nerves in mice

To investigate the involvement of blood-born factors and extracellular proteases in axonal degeneration and regeneration in both PNS and CNS, we directly compared the differences of blood-nerve barrier (BNB) disruption and matrix metalloprotease-9 (MMP-9) induction between the sciatic nerve and optic nerve after crush injury in the same animal. In sciatic nerve, BNB disruption, fibrin(ogen) deposition and MMP-9 expression were observed only in the first week following injury. Neurofilament (NF) immunoreactivity dramatically decreased in the first 2 days, gradually recovered to the normal levels by day 28. In contrast, the immunoglobulin G deposits spanned from 4 h to 28 days in crushed optic nerves. Fibrin(ogen) deposition was only observed in the first 2 days, while MMP-9 induction did not occur until a week after injury but lasted for 3 weeks in the crushed optic nerves. The NF immunoreactivity did not change much until day 7 and almost completely disappeared on day 28. The decrease of NF immunoreactivity coincided with the induction of MMP-9 after optic nerve crush. These results show that BNB disruption and MMP-9 induction are differentially regulated in the PNS and CNS after injuries, and they may contribute to the different regeneration capacities of the two systems.     

小胶质细胞在中枢神经系统和胶质瘤中的免疫学作用及其相关研究进展

血脑屏障在正常情况下能够保证脑的内环境的高度稳定性,以利于中枢神经系统的机能活动。中枢神经系统内缺乏各种常见的抗原递呈细胞,但小胶质细胞可作为潜在的抗原递呈细胞将MHC分子结合的抗原肽与T细胞受体结合介导相应的免疫反应。而在胶质瘤组织中高度浸润的小胶质细胞可通过对肿瘤微环境的免疫抑制而促进肿瘤生长。一旦小胶质细胞被激活就能成为强大的免疫效应细胞,在中枢神经系统损伤和疾病中发挥多种生物学功能。     

Role of microglia in central nervous system infections

The nature of microglia fascinated many prominent researchers in the 19th and early 20th centuries, and in a classic treatise in 1932, Pio del Rio-Hortega formulated a number of concepts regarding the function of these resident macrophages of the brain parenchyma that remain relevant to this day. However, a renaissance of interest in microglia occurred toward the end of the 20th century, fueled by the recognition of their role in neuropathogenesis of infectious agents, such as human immunodeficiency virus type 1, and by what appears to be their participation in other neurodegenerative and neuroinflammatory disorders. During the same period, insights into the physiological and pathological properties of microglia were gained from in vivo and in vitro studies of neurotropic viruses, bacteria, fungi, parasites, and prions, which are reviewed in this article. New concepts that have emerged from these studies include the importance of cytokines and chemokines produced by activated microglia in neurodegenerative and neuroprotective processes and the elegant but astonishingly complex interactions between microglia, astrocytes, lymphocytes, and neurons that underlie these processes. It is proposed that an enhanced understanding of microglia will yield improved therapies of central nervous system infections, since such therapies are, by and large, sorely needed.     

尼莫地平对慢性退变中神经元的保护作用

目的　探讨尼莫地平对慢性退变中神经元的保护作用。方法　 4 2只生后 2d新生大鼠随机分成实验组和对照组 ,切断大鼠右侧坐骨神经作为实验侧 ,左侧为正常对照侧。实验组于术后 30min内、第 5、10d皮下注射尼莫地平 (2 0 μg/g体重 ) ,对照组注射等量溶剂 ,连续用药 2周 ,然后取L4～ 6脊髓 ,在光镜下观察并计数脊髓运动神经元。结果　术后 30min内和延迟 5、10d给予尼莫地平 ,可以保护平均 6 3 6 %神经元存活 ,与对照组比较统计学表明差异非常显著。延迟 5d和 10d给予尼莫地平 ,保护作用无显著性差异。结论　尼莫地平在大鼠坐骨神经横断后 30min内、第 5、10d均有神经元保护作用 ,在术后一定时间内其延迟保护作用恒定     

周围神经系统损伤的微环境与修复方式

背景:一直以来周围神经损伤在临床工作中十分常见,虽然显微外科技术能很好地恢复损伤神经的连续性,但是由于周围神经组织存在分化程度较高、再生能力较低的特点,使得神经修复效果仍不理想,严重影响患者的生活质量.目前周围神经损伤微环境尚无统一定论,常用的修复方式众多.目的:对周围神经损伤微环境及周围神经损伤修复方式进行综述.方法...     

An explanation of axonal regeneration in peripheral nerves and its failure in the central nervous system

Nerve fibres severed within peripheral nerves are able to regenerate and reinnervate the structures they formerly supplied. Most axons severed within the mammalian central nervous system (CNS) do not regenerate in this way. Regenerative axonal growth begins to occur in the CNS but ceases about two weeks after injury. Five earlier theories purporting to explain this difference are reviewed and found not to account satisfactorily for many experimental observations. A new hypothesis is advanced in which it is held that in order for regeneration to take place, the growing tips of the axons must be surrounded by extracellular fluid containing proteins (of specified identity) derived from the blood plasma. Such proteins are thought to be imbibed by the tips of the fibres and transported retrogradely to the neuronal cell-bodies. With this hypothesis it is possible to explain the success of axonal regneration in peripheral nerves and its failure in the CNS. It is also possible to account for the exceptional circumstances in which axons do regenerate in the CNS. Various experiments are suggested for testing the validity of the new hypothesis.     

Experience with examination of the spinal cord and peripheral nervous system (PNS) in mice: A brief overview

The representative areas for examination of the mouse peripheral nervous system are the spinal cord, containing central components of the peripheral nervous system that needs to be examined at least at cervical and lumbar level, the sciatic and the tibial nerve. Skeletal muscle samples should include the soleus muscle and the quadriceps femoris or long digital extensor, as well as the medial gastrocnemius. Examination can be extended to the thoracic spinal cord, lumbar dorsal root ganglia and spinal nerve roots, as well as the plantar nerve, and other areas of interest. Perfusion fixation is considered optimal for the nervous system; however, immersion fixation allows producing microscopic sections of excellent quality as well. Paraffin-embedded, hematoxylin and eosin-stained sections can be made from all areas, save for small nerves such as the tibial or plantar nerve, which are examined with advantage in hard plastic sections. It is possible to produce hard plastic sections also of the vertebral column, including the spinal cord, dorsal root ganglia and nerve roots. For special investigations, mice can be fixed in toto, decalcified, embedded and sectioned to reveal the areas of interest. In the mouse peripheral nerves, myelination progresses until the adult age. In aging peripheral nerves there is axonal atrophy, degeneration, nerve fiber loss, increase of collagen and sporadic demyelination, especially radiculoneuropathy. The dorsal root ganglia of untreated control animals show frequent cytoplasmic vacuolation. Axonal degeneration is distally, primary demyelination proximally accentuated. Mouse is not very sensitive to peripheral neurotoxicity: to induce toxic peripheral neuropathy mostly parenteral administration and/or newborn animals are used. Naturally occurring infection affecting the spinal cord and peripheral nerves is Theiler's encephalomyelitis virus inducing acute poliomyelitis or chronic demyelination. Any experimental results are to be assessed taking into account spontaneous, age-related, background changes.