成年大鼠嗅球神经干细胞的培养和鉴定

目的建立成年大鼠嗅球神经干细胞分离培养和鉴定方法,探索新的成年神经干细胞种子来源。方法用无血清方法分离培养成年大鼠嗅球来源的神经干细胞;用克隆培养、BrdU整合的方法检验培养细胞的干细胞特性;用免疫荧光细胞化学的方法检测BrdU、神经干细胞标记物Nestin和SOX2,分化的细胞标记物Tuj1、GFAP、NG2的表达。结果从成年大鼠嗅球能够分离、培养出具有自我更新、增殖的神经球,构成神经球的细胞呈Nestin和SOX2阳性,它们分化后产生Tuj1阳性的神经元、GFAP阳性的星形胶质细胞、NG2阳性的少突胶质细胞。结论成年大鼠嗅球存在神经干细胞,能够在体外进行培养、增殖、分化,是神经干细胞的新的种子来源。     

人胚嗅鞘细胞与胚胎脊髓组织联合移植对大鼠脊髓损伤的治疗作用

目的探讨人胚嗅鞘细胞（OECS）和大鼠胚胎脊髓组织（ESC）联合移植在促进大鼠横断脊髓轴索再生方面有无协同作用。方法将体外培养纯化的OECS及新鲜获取的胎鼠脊髓组织用作治疗大鼠脊髓半切洞损伤模型的移植物，通过对动物定期进行行为学评定，结合病理学观察，并通过辣根过氧化物酶-四甲基联苯胺（HRP-TMB）逆行示踪技术，评价OECS和ESC对神经元存活、纤维再生的影响。结果OECS和ESC联合移植对损伤脊髓具有明显保护作用并能促进宿主脊髓轴突再生，ESC不能帮助再生轴索返回宿主组织；而OECS除能帮助再生轴索突破胶质屏障外更能使再生轴突髓鞘化，并具有一定的迁移能力。结论OECS和ESC联合移植在促进大鼠脊髓功能恢复中起到了互补和协同的作用。     

人骨髓间充质干细胞分化为心肌样细胞的研究

目的：研究人骨髓间充质干细胞（hMSCs）在体外分离纯化、培养扩增和向心肌样细胞诱导分化的条件。方法：体外分离培养hMSCs,纯化传代至第三、四代,加入不同浓度5-氮杂胞苷（5-Aza）进行不同时间的孵育,用MTT测定细胞的生长活性,确定最佳的浓度和孵育时间。4周后行电镜,免疫组化染色及逆转录-聚合酶链式反应（RT-PCR）检测。结果：诱导后细胞呈心肌样细胞改变,电镜下可见肌丝形成,免疫组化显示部分细胞胞浆α-横纹肌肌动蛋白（α-sarcomeric actin）、肌钙蛋白-T（troponin-T）阳性。RT-PCR检测显示心肌特异转录因子GATA-4、Nkx2.5有表达。结论：hMSCs是骨髓来源的具有多向分化潜能的干细胞,在5-Aza的诱导下可向心肌样细胞分化,可成为心肌损伤移植治疗的理想细胞材料。     

人神经干细胞移植修复大鼠脊髓损伤的实验研究

目的研究人神经干细胞（hNSCs）移植对大鼠脊髓损伤的修复作用，并初步探讨其作用原理。方法分离、培养和鉴定hNSCs。24例成年SD大鼠分为移植组12例和对照组12例，均采用NYU-Ⅱ型脊髓打击器制作脊髓损伤模型，第9天移植组于损伤脊髓中心分别注入经CM-DiI标记的hNSCs混悬液，对照组注入人DMEM／F12培养液。术后第4、8周取损伤部位脊髓，免疫组织化学染色检测移植细胞的存活和分化；术后每7天行BBB评分，评定大鼠后肢运动功能恢复情况。数据进行成组设计资料的t检验。结果成功建立hNSCs的体外培养体系；移植的hNSCs在大鼠脊髓内存活超过8周，并向脊髓损伤头尾端迁移，免疫组织化学荧光染色示移植细胞可分化为神经元、少突胶质细胞和星形胶质细胞；移植组大鼠BBB评分高于对照组（P〈0．05）。结论移植到大鼠损伤脊髓中的人神经干细胞可分化为神经元和神经胶质细胞，并可促进脊髓损伤大鼠后肢运动功能恢复。     

骨髓基质细胞移植抑制缺血心肌纤维化

目的探讨骨髓基质细胞移植改善缺血心脏功能的机制。方法制作F344大鼠心肌梗死模型，将分离培养的骨髓基质细胞移植于心梗周边区（治疗组），取1、3、5、7、9、15d后缺血心肌切片，通过免疫组织化学染色及免疫荧光染色检测波形蛋白的表达，了解梗死心肌纤维化程度，并应用血液动力学在术后各时间点检测大鼠心脏功能的变化。结果治疗组心功能较同期对照组明显改善。术后7、9、15d治疗组缺血心肌纤维化程度较同期对照组明显减轻。结论骨髓基质细胞移植于缺血心肌后明显改善缺血心脏功能。其机制之一可能是抑制缺血心肌的纤维化，延缓心室重构。     

脐静脉源MSC分离及生物学特性

目的探讨脐静脉源间充质干细胞（MSC）的分离及培养,观察其进一步分化的潜能。方法用胶原酶常规消化分离脐静脉内皮细胞,传代3周后,用免疫组化法和流式细胞术检测细胞免疫表型;观察该类细胞向脂肪和成骨细胞分化的潜能,油红O染色和von Kossa染色鉴定细胞性质。结果脐静脉内皮／内皮下细胞接种3～4周后以长梭形的成纤维样细胞为主。传代至F2代时细胞形态均一为长梭形。表型检测示该类细胞高表达CD29、CD44、CD54、CD49e、HLA-ABC和CD40分子,不表达造血细胞相关分子如CD34、CD45、HLA-DR,免疫组化分析α平滑肌肌动蛋白（α-SMA）阳性、血管性血友病因子（vWF）阴性;细胞经成脂诱导后镜下可见细胞内脂滴形成,3周后油红O染色阳性;经成骨诱导3周后细胞von Kossa染色为阳性。结论脐静脉内皮和内皮下含有MSC样细胞,在适当的条件下可向脂肪细胞和成骨细胞分化。脐带可成为实验和临床研究所用MSC的又一来源。     

体外培养人脐带血间充质干细胞向胰岛细胞分化及对糖尿病治疗的实验研究

目的研究人脐带血间充质干细胞向胰岛素分泌细胞分化的潜能及其移植后对糖尿病大鼠的治疗效果。方法体外分离培养HUCB-MSCs,在胰岛细胞培养条件下经药物定向诱导其分化;免疫组化对诱导细胞进行胰岛β细胞标记鉴定;双硫腙染色鉴定锌离子表达及检测胰岛样细胞的移植效果。结果 HUCB-MSCs经诱导后,免疫细胞化学染色显示表达人胰岛素;双硫腙染色呈棕红色;移植后2周,胰岛样细胞组血糖浓度明显降低。结论 HUCB-MSCs在体外诱导培养条件下,具有向胰岛素分泌细胞分化的潜能,这种细胞可能为Ⅰ型糖尿病提供一条新的治疗途径。     

冰冻切片免疫组化检测大鼠脊髓组织NSE、NF的表达

目的观察冰冻切片免疫组化染色检测大鼠脊髓组织神经丝蛋白（NF）、神经元特异性烯醇化酶（NSE）表达情况。方法采用改良Allen重物打击法制作大鼠脊髓损伤模型，大鼠骨髓间质干细胞（rMSCs）静脉移植后对其脊髓组织行冰冻切片，免疫组化检测NSE、NF的表达情况。结果脊髓组织NSE、NF均表达，且与常规石蜡切片免疫组化染色结果基本相同。结论冰冻切片行免疫组化染色可有效检测神经元特异标志分子NSE、NF的表达。     

头面部恶性黑色素性神经鞘瘤2例及文献复习

目的 了解恶性黑色索性神经鞘瘤的临床病理特征及其鉴别诊断及预后．方法 收集2例恶性黑色索性神经鞘瘤观察其临床病理特点并进行组织化学及免疫组化染色分析。结果 所有肿瘤均具有恶性黑色素瘤的临床与组织学特征且都伴有多少量不等的黑色素。结论 恶性黑色索性神经鞘瘤少见，恶性度高，临床诊断较困难但有其病理特征．依靠镜下观察及配以组织化学染色、免疫组化染色可确诊。复习文献该肿瘤瘤细胞同时显示神经鞘瘤和黑色素细胞的分化，提示该肿瘤来自神经蝽，进一步单克隆培养实验表明神经嵴细胞具有异质性，可由全潜能细胞，多潜能细胞组成，黑色素性神经鞘瘤可以被认为是具有两种潜能细胞的增生而形成的。     

人脐带问充质干细胞体外培养、鉴定及其诱导分化的研究

目的通过体外分离、培养及鉴定人脐带间充质干细胞（humanumbilicalcordmesenchymalstemcells,hUCMSCs）,探讨其多向分化潜能。方法取正常足月新生儿脐带,采用组织贴壁培养法分离原代hUCMSCs,观察细胞生长形态。采用流式细胞仪技术检测hUCMSCs细胞表型及细胞周期。通过成神经细胞诱导和成脂肪细胞诱导,鉴定hUCMSCs的多向诱导分化能力。结果成功分离和培养hUCMSCs原代细胞,经流式细胞仪鉴定高表达间质细胞标志CD44和CDl05,阳性率为96．73％和96．10％,整合素受体CD29阳性率为99．53％;低表达造血系标志CD34和CD45阳性率为0．80％和1．91％,人白细胞抗原HLA-DR阳性率为1．41％。细胞周期检测hUCMSCs主要处于G0／G1期。hUCMSCs成神经细胞诱导,出现神经元样细胞;免疫组化检测神经巢蛋白（Nestin）呈阳性表达。hUCMSCs成脂肪细胞诱导,出现空泡样脂肪滴,油红O染色可见脂质沉积。结论hUCMSCs具有多向分化潜能,可跨胚层诱导分化为多种组织类型的细胞。     

Role of the olfactory receptor neurons in the direct transport of inhaled uranium to the rat brain

Uranium presents numerous industrial and military uses and one of the most important risks of contamination is dust inhalation. In contrast to the other modes of contamination, the inhaled uranium has been proposed to enter the brain not only by the common route of all modes of exposure, the blood pathway, but also by a specific inhalation exposure route, the olfactory pathway. To test whether the inhaled uranium enter the brain directly from the nasal cavity, male Sprague–Dawley rats were exposed to both inhaled and intraperitoneally injected uranium using the ²³⁶U and ²³³U, respectively, as tracers. The results showed a specific frontal brain accumulation of the inhaled uranium which is not observed with the injected uranium. Furthermore, the inhaled uranium is higher than the injected uranium in the olfactory bulbs (OB) and tubercles, in the frontal cortex and in the hypothalamus. In contrast, the other cerebral areas (cortex, hippocampus, cerebellum and brain residue) did not show any preferential accumulation of inhaled or injected uranium. These results mean that inhaled uranium enters the brain via a direct transfer from the nasal turbinates to the OB in addition to the systemic pathway. The uranium transfer from the nasal turbinates to the OB is lower in animals showing a reduced level of olfactory receptor neurons (ORN) induced by an olfactory epithelium lesion prior to the uranium inhalation exposure. These results give prominence to a role of the ORN in the direct transfer of the uranium from the nasal cavity to the brain.     

Delayed emergence of antidepressant efficacy following withdrawal in olfactory bulbectomized rats

Repeated antidepressant treatment attenuates the step-down passive avoidance deficit which is induced by olfactory bulbectomy in rats. Using a shuttlebox passive avoidance procedure, the effects of antidepressants were investigated after various drug withdrawal intervals. Imipramine, amitriptyline, doxepin, bupropion and mianserin were effective at 48 and usually 72 hours after withdrawal, but no significant attenuation of the deficit was seen 4 hours after withdrawal from any antidepressant tested. At least 4 to 7 days of imipramine treatment were required for efficacy. A high dose of d-amphetamine (5 mg/kg) produced similar results while tranylcypromine and haloperidol were inactive at all withdrawal intervals tested. The olfactory bulbectomy syndrome may reflect functional derotonin deficiency, which would be ameliorated through antidepressant-induced alterations in serotonin receptor sensitivity.     

小鼠鼻腔滴注纳米二氧化铈进入嗅球的研究

目的研究鼻腔滴注的纳米二氧化铈(CeO2)颗粒可否通过嗅神经到达嗅球。方法对小鼠左鼻孔滴注40μg放射性纳米CeO2材料,首次滴注后的第1、3和7天后麻醉处死小鼠,分离双侧的嗅球和嗅上皮组织,采用放射性示踪方法对双侧嗅球和嗅上皮组织中的纳米CeO2含量进行定量比较。结果左侧嗅球和嗅上皮组织中的纳米CeO2含量呈现显著的时间累积效应,右侧嗅球和嗅上皮组织中的纳米CeO2含量增加无统计学意义(P>0.05)。结论沉积在鼻腔粘膜上的纳米CeO2颗粒可以通过嗅神经到达嗅球。     

56例以嗅觉障碍为主慢性鼻-鼻窦炎的临床分析

目的探讨以嗅觉障碍为主要症状的慢性鼻-鼻窦炎患者的临床表现及治疗。方法案例来自宁德市医院接治的56例慢性鼻-鼻窦炎患者,采用手术治疗其中32例(64侧)患者。治疗效果通过主观嗅觉识别阈以及嗅觉事件相关电位检查方法进行评价。结果主观嗅觉识别阈对病程≤12个月和>12个月患者检测结果差异无统计学意义(P>0.05)。通过手术剂治疗,所有32例(64侧)患者的检测指标提高显著(P<0.01),其中24例(48侧)病程≤12个月的患者在治疗过后成效高于8例(16侧)病程>12个月的患者(P<0.05)。结论采用手术治疗能够对慢性鼻-鼻窦炎患者起到治疗作用,值得推广。     

Drug Transport into the Mammalian Brain: The Nasal Pathway and its Specific Metabolic Barrier

It is generally accepted that the rate of entry into and distribution of drugs and other xenobiotics within the central nervous system (CNS) depends on the particular anatomy of the brain microvessels forming the blood-brain barrier (BBB), and of the choroid plexus forming the blood-cerebrospinal fluid barrier (CSF), which possess tight junctions preventing the passage of most polar substances. Drug entry to the CNS also depends on the physicochemical properties of the substances, which can be metabolised during this transport to pharmacologically inactive, non-penetrating polar products. Finally, the entry of drugs may be prevented by multiple complex specialized carriers, which are able to catalyse the active transport of numerous drugs and xenobiotics out of the CNS. Nasal delivery is currently considered as an efficient tool for systemic administration of drugs that are poorly absorbed via the oral route, and increasing evidence suggests that numerous drugs and potentially toxic xenobiotics can reach the CNS by this route. This short review summarizes recent knowledge on factors controlling the nasal pathway, focusing on drug metabolising enzymes in olfactory mucosa, olfactory bulb and brain, which should constitute a CNS metabolic barrier.     

Visualization of In Vivo Olfactory Uptake and Transfer Using Fluorescein Dextran

Nasal administration of a 3 kDa fluorescein dextran (FD3) solution to rats resulted in transcellular absorption across the olfactory epithelium and transfer to the olfactory bulb within 15 min. After entering the lamina propria, FD3 was transferred in the connective tissue surrounding the olfactory nerve bundles to the olfactory bulb of the brain. More FD3 was absorbed across the olfactory epithelium than across the respiratory epithelium and to the nasal associated lymphoid tissue. Further, the amount of FD3 crossing the olfactory epithelium was region-dependent, with higher amounts absorbed in the turbinates than in the nasal septum. Plastic embedding and sectioning followed by fluorescence microscopy, enabled simultaneous visualization of FD3 in the mucosa and olfactory bulb, as well as the opportunity to store the tissue blocks for a prolonged period of time.     

Effects of olfactory bulb removal on brain norepinephrine in golden hamsters

Changes in brain norepinephrine levels after olfactory bulb removal have been found in rats. Several investigators have suggested that the dramatic behavioral effects of bilateral olfactory bulb removal in hamsters might be due to a possible similar change rather than anosmia per se. The results of this study do not support this hypothesis since no significant differences in brain norepinephrine were found between sham operated and bilaterally bulbectomized male hamsters treated identically to those used in earlier behavioral studies on olfactory bulb removal. Also, since there were no changes in the norepinephrine levels in female hamsters which were treated identically to female rats used in earlier neurochemical studies, it is probable that there is a species difference in the effects of olfactory bulb removal on brain norepinephrine.     

嗅沟脑膜瘤的早期诊断

目的探讨嗅沟脑膜瘤的早期诊断。方法回顾性分析17例嗅沟脑膜瘤的临床记录,包括临床表现、症状持续时间、神经影像学特征、手术结果和组织学类型。结果本组病例显示患者平均手术年龄44岁,症状持续时间平均17个月,头痛和精神障碍为常见症状,视乳头水肿和嗅觉功能障碍是常见体征。肿瘤的最大直径平均4.6 cm,大多数患者有瘤周水肿。肿瘤全切16例,次全切1例。88%的患者康复良好,放射学随访期间无复发。结论诊断的延误使得手术时肿瘤较大,CT和/或MR I是嗅沟脑膜瘤早期诊断的重要方法。     

嗅球结构及其对生物节律的调控

嗅球是人体控制嗅觉的关键部位。近年来研究发现,嗅球还参与了生物节律的调控,其相对独立于视交叉上核表达节律调控基因。基于嗅球与脑内的神经联系以及其对生物节律调控的作用,深入研究一些神经系统疾病和睡眠等人体自身节律可能会成为基础和临床医学研究的新热点。本文将介绍嗅球的结构,并对其参与调控生物节律的功能进行分析总结。     

Activation and toxicity of bromobenzene in nasal tissues in mice

Autoradiography of mice injected i. v. with bromobenzene-¹⁴C (BB; 25 mol/kg body wt) revealed a high concentration of non-volatile metabolites in the olfactory mucosa and in the glands around the maxillary sinuses. As determined with solvent-extracted tissue sections, there was a high level of irreversibly bound metabolites in the Bowman's glands in the olfactory mucosa, while the level of bound metabolites was low in the glands around the maxillary sinuses. Histopathological examination of the nose region of mice given a single i.p. dose of unlabelled BB ( 4.8 mmol/kg body wt) revealed degeneration and necrosis of the glands of Bowman. Degenerative changes in the olfactory epithelium were also observed. Moreover, focal degeneration and necrosis were found in the lateral nasal glands (4.8 mmol/kg body wt). Cyst-like dilatation of acini was observed in the lateral nasal glands and in the maxillary glands located around the maxillary sinus. Incubation of BB (26 M) with a homogenate prepared from the olfactory mucosa revealed an irreversible binding, which clearly exceeded that of the liver. It is suggested that BB is activated in situ to a cytotoxic metabolite that reacts with the glands of Bowman. The specific toxicity observed in the lateral nasal glands correlated with a high concentration of non-volatile but extractable metabolites in these glands.