免疫源性IL-1在神经免疫调节网络传入通路中的作用

目的 阐明免疫活性细胞表达的功能信号IL-1在神经免疫调节网络信号传导过程中的作用.方法 应用双向电泳分离联合基质辅助激光解析电离-飞行时间-质谱技术,分析实验大鼠外周血清中神经免疫功能相关差异组分的氨基酸组成.结果 2D电泳在相对分子质量(Mr)约18 000、PI约5.0位置观察到1个与IL-1物理性质相似的特异性差异表达点;MALDI质谱检测、肽质量指纹图谱分析在相关数据库检索结果中报告了18个来源的19个相关肽段序列,共247个氨基酸,但相关结果不能满足任何已知物质的氨基酸序列排序要求.结论 实验否定了免疫活性细胞释放到血液中的IL-1参与神经免疫调节网络功能的可能;是否存在网络神经元多极换元换能后间接活化中枢脑区功能的可能,还有待进一步实验研究.     

神经元凋亡与神经免疫调节网络功能下调模式

目的 探讨神经免疫调节网络功能下调模式.方法 应用大鼠全基因芯片检测技术分析免疫大鼠下丘脑外侧区和杏仁核前区神经免疫调节功能相关神经元凋亡等的相关信号表达;应用数据库对差异基因功能树进行分析.结果 实验组大鼠基因表达谱分析显示有632个差异基因,经功能树相关分析显示其中34个基因参与信号传导功能的已知功能基因,涵盖了已知凋亡信号和凋亡调节信号相关传导路.结论 在LH 和AA脑区存在以神经元凋亡方式实现神经免疫调节功能相关的网络功能下调模式.     

神经免疫调节网络上传信号的细胞来源

目的通过比较不同免疫状态的Balb/c鼠和裸鼠中枢神经免疫调节相关脑区功能的差异,探讨外周免疫系统释放的神经免疫调节网络上传信号的细胞来源。方法 Balb/c鼠和裸鼠各50只,每种动物随机分成免疫2、4、6、8d组和健康对照组。应用免疫组织化学技术检测不同免疫时间点动物的神经免疫调节相关脑区下丘脑外侧区(lateral hypothalamic area,LH)和杏仁核前区(anterior amygdaloid area,AA)功能活化信号白细胞介素-1β(IL-1β)的表达水平,分析T细胞功能缺陷对神经免疫调节功能的影响。结果与健康对照组比较,Balb/c鼠免疫2、4、6d组LH、AA脑区IL-1β表达明显增高(P<0.05),且免疫后2d其表达即增高,免疫后4d达高峰,免疫后6d开始下降(均P<0.05),免疫后8d其表达量降至基础生理水平(P>0.05)。裸鼠免疫组LH、AA脑区IL-1β表达趋势与Balb/c鼠结果相似。裸鼠免疫组LH、AA脑区IL-1β表达水平与Balb/c鼠比较无统计学差异(P>0.05)。结论 Balb/c鼠与裸鼠经抗原免疫注射后均可启动LH、AA脑区的神经免疫调节功能,提示免疫系统功能活化后,向中枢神经系统上传的神经免疫调节功能启动信号由树突状细胞和T辅助细胞共同提呈,但主要来自树突状细胞。     

17-β雌二醇对血管性痴呆大鼠认知功能及脑内GDNF表达的影响

目的 研究17-β雌二醇对血管性痴呆(VD)大鼠脑组织胶源性神经营养因子(GNDF)表达的影响.方法 结扎双侧颈总动脉制备慢性前脑缺血动物模型,应用Y迷宫,免疫组化检测腹腔注射17-β雌二醇60d后血管性痴呆大鼠认知功能以及脑组织中GDNF表达.结果 与正常对照组对比,大鼠学习尝试次数,记忆测试10次的正确次数在造模60d后差异有统计学意义(P＜0.01),腹腔注射17-β雌二醇60d后,大鼠认知功显著改善;造模60d后的大鼠,其脑内GDNF免疫阳性细胞数较正常对照组显著增高,雌二醇组大鼠脑组织内GDNF免疫阳性细胞数较模型组显著增高.结论 腹腔注射17-β雌二醇可显著改善VD大鼠的认知功能,增加VD大鼠脑内GDNF表达.     

雷公藤内酯醇对实验性自身免疫性脑脊髓炎大鼠血-脑脊液屏障的影响

目的 探讨雷公藤内酯醇(Tri)对实验性自身免疫性脑脊髓炎(EAE)大鼠血-脑脊液屏障(BCB)的免疫调节作用.方法 制备Wistar大鼠EAE模型,应用Tri治疗EAE,评估EAE组、Tri治疗组的临床症状,在不同时间点测定脑脊液(CSF)和外周血白蛋白(Alb)水平并计算二者比值(QA),通过QA值观察BCB变化.于免疫后(p.I.)6、8、10、12、14、16 d处死动物,用免疫组化技术检测脑和脊髓细胞间黏附分子-1(ICAM-1)和干扰素-γ(IFN-γ)表达.结果 免疫后14 d为EAE组发病高峰期,Tri治疗组临床症状评分(0.14±0.08)较EAE组(2.19±1.68)低(P<0.01).免疫后 8 d EAE组QA达最大值(0.30±0.05),较佐剂组(0.13±0.05)明显增高(P<0.01).免疫后6-14 d,EAE组与佐剂组、Tri治疗组比较QA值差异均有统计学意义(F=28.34,P<0.01).免疫后14 d Tri治疗组 QA达最大值(0.19±0.02),QA波峰时间较EAE组延迟且峰值明显较低.免疫后12-16 d,EAE组与佐剂组、Tri治疗组比较ICAM-1阳性表达高(F=17.11,F=29.87,均P<0.01);免疫后10-16 d,Tri治疗组较EAE组IFN-γ阳性表达低(F=447.92,P<0.01).结论 EAE中BCB损伤早于临床症状出现.Tri能有效抑制EAE进展,可能与其下调ICAM-1、IFN-γ表达,恢复BCB正常功能有关.     

急性脑血管病患者血清和脑脊液中胰岛素样生长因子-1的测定及其临床意义

目的　探讨急性脑血管病患者血清及脑脊液中胰岛素样生长因子 - 1(IGF - 1)浓度的变化及其临床意义。方法　观察 40例脑血管病患者和 2 0例正常人血清及脑脊液IGF - 1浓度。病例组 :脑梗死组2 0例 ,脑出血组 2 0例 ,所有患者均在发病后第 3天抽取静脉血 2ml,其中脑梗死组在抽血当天作腰穿抽取脑脊液 2ml。IGF - 1采用酶联免疫分析方法测定。应用SPSS统计软件包进行统计分析 ,组间比较采用t检验。结果　病例组与对照组血清中IGF - 1浓度有明显统计学差异 (P <0 .0 1) ,脑梗死组与脑出血组血清中IGF - 1浓度无统计学差异 (P >0 .0 5 )。脑梗死组与对照组脑脊液中IGF - 1浓度有明显统计学差异 (P <0 .0 1)。结论　脑梗死和脑出血患者急性期血清IGF - 1浓度明显降低 ,脑梗死组患者急性期脑脊液IGF - 1浓度明显增高。     

脑脊液中细胞源性囊泡对间充质干细胞向神经前体细胞分化的影响

目的探讨脑脊液微囊泡在间充质干细胞(MSCs)向神经干细胞分化中的作用。方法从人脑脊液中分离微囊泡,从小鼠骨髓中分离原代MSCs。将MSCs传代后在神经分化诱导培养基和脑脊液微囊泡作用下神经分化诱导。细胞形态学观察细胞突起数目和长度,western blot检测细胞神经特异性蛋白表达水平变化。结果单纯神经诱导液组单位面积内细胞突起长度为(2.2±0.4)mm,突起数量为(94±12)个;神经诱导液加脑脊液微囊泡诱导的细胞中细胞突起更长(5.7±1.2)mm,数目也更多(178.2±32)个,差异具有统计学意义(P0.01)。微囊泡组细胞中NSE、Nestin和MAP-2表达水平高于单纯神经诱导液组细胞,差异具有统计学意义(P0.01)。结论脑脊液微囊泡可有效促进MSCs向神经干细胞分化,有望为干细胞移植治疗神经功能缺损提供了一个新的思路。     

实验性自身免疫性脑脊髓炎大鼠血-脑脊液屏障功能及动态变化

目的　探讨实验性自身免疫性脑脊髓炎 ( EAE)大鼠血 -脑脊液屏障功能的动态变化及其作用 ,以求进一步揭示 EAE发病机制。方法　检测 EAE大鼠免疫后第 4、6、8、1 0、1 2、1 4、1 6、1 8、2 0天血清和脑脊液 ( CSF)中清蛋白 ( ALB)含量 ,其 CSF与血清 ALB比值 ( QA)作为评价血 -脑脊液屏障损害的指标。结果　 QA值免疫后第 8天即出现显著性升高 ,早于临床症状出现 ;随免疫时间的延长 QA值呈逐渐增高后缓慢下降趋势 ,并且 QA值与EAE大鼠病情评分呈显著正相关 ( r =0 .81 ,P =0 .0 0 1 )。结论　血 -脑脊液屏障的早期损害在 EAE发病过程中具关键作用。     

ERK信号通路在DPA诱导神经干细胞分化中的作用

目的探讨ERK信号通路在二十二碳戊烯酸(docosapentenoic acid,DPA)诱导神经干细胞(neural stem cells,NSCs)分化中的作用。方法原代培养SD大鼠来源神经干细胞,用不同浓度DPA和血清对照组进行神经干细胞诱导分化;用图像分析软件Image-pro Plus 2. 0采集突起长度和突触形成率;免疫组织化学方法检测NES在的表达率; Western blot法检测βⅢ-tubulin的表达和ERK信号途径磷酸化水平。结果与对照组相比神经突起以及突触形成在DPA作用下随着浓度增加而增加,差异有统计学意义(P 0. 05,P 0. 01);与对照组相比NSE阳性细胞表达率随着DAP浓度增加而增加(P 0. 05,P 0. 01);与对照组相比细胞骨架蛋白βⅢ-tubulin的表达增加,差异有统计学意义(P 0. 05,P 0. 01);与对照组相比ERK信号分子磷酸化水平升高,差异有统计学意义(P0. 05,P 0. 01)。结论 DPA促进NSCs分化中神经突起的表达和神经元的分化,活化的ERK信号途径通过信号转导可能是诱导神经突起表达的重要机制。     

散发性克-雅氏病患者脑脊液中抗炎性细胞因子IL-4水平的检测

目的研究散发性克-雅氏病(sCJD)患者脑脊液中抗炎性细胞因子IL-4的水平,探讨抗炎性细胞因子在CJD疾病过程中的作用。方法病例分为CJD组、其它痴呆组和对照组。采用westernblot法定性分析各组脑脊液中IL-4的表达,用酶联免疫吸附实验检测脑脊液中IL-4的浓度。结果CJD患者脑脊液中IL-4水平明显升高,与其它痴呆组和对照组相比有统计学意义(P<0.05)。结论抗炎性细胞因子IL-4在CJD患者脑脊液中含量增加,在其神经变性过程中可能发挥免疫调节作用,并且有可能成为CJD的生物学标志物。     

Immunoregulatory properties of T-cell lines derived from the systemic and intrathecal compartments: a phenotypic and functional study

To determine whether immune regulation can differ within the intrathecal and systemic compartments, we compared phenotypic markers and functional properties of in vitro anti-CD3 monoclonal antibody-stimulated, interleukin 2-expanded lymphoid cell lines simultaneously derived from peripheral blood and cerebrospinal fluid of individual donors (n = 9). We found that the proportions of total CD8+ T cells and of the putative CD8+ suppressor effector subset (CD28-) were lower in the cell lines derived from cerebrospinal fluid compared with cultures derived from peripheral blood (p less than 0.025 and p less than 0.005, respectively; paired t test), whereas the total CD4+ T-cell proportion was higher (p less than 0.025). For a donor subgroup with "normal" peripheral blood cell-mediated activated suppressor function (63 +/- 2%), mean suppressor cell function mediated by unfractionated or CD8(+)-enriched cells derived from cerebrospinal fluid was significantly lower (38 +/- 7%; p less than 0.01, paired t test). For a donor subgroup with "low" peripheral blood cell-mediated suppression (-1 +/- 10%), suppression mediated by cerebrospinal fluid cells was also "low" (9 +/- 12%). Our results support the postulate that the immune response may be differentially regulated between the central nervous system and peripheral blood compartments.     

INCREASED FIBRINOLYTIC ACTIVITY AND FIBRIN DEGRADATION PRODUCTS AFTER EXPERIMENTAL INTRACEREBRAL HAEMORRAGE

The fibrinolytic activity of the peripheral blood and cerebrospinal fluid was studied in 10 dogs in which intracerebral haematomas had been produced by injection of blood into the brain. Blood and cerebrospinal fluid samples were obtained at regular intervals before and after operation. The fibrinolytic activity of these two fluids were estimated by the fibrin plate method and by assays for fibrin degradation products. After the production of the haematoma, signs of increased fibrinolytic activity were found in blood and cerebrospinal fluid one day after the intracerebral injection in 6 animals. A correlation was found between the size of the haematoma and the level of the fibrinolytic activity of blood and cerebrospinal fluid. Determination of fibrin degradation products in blood and cerebrospinal fluid and the measurement of fibrinolytic activity by the plate method complement each other and are therefore recommended in this type of investigation.     

Common T-cell receptor V beta usage in oligoclonal T lymphocytes derived from cerebrospinal fluid and blood of patients with multiple sclerosis.

T-cell populations were investigated in the blood and cerebrospinal fluid of patients with multiple sclerosis and other neurological diseases. Individual T cells were directly cloned from the cerebrospinal fluid and blood before in vitro expansion, and their clonotypes were compared by Southern blot analysis of the rearrangement patterns of their T-cell receptor beta chain and gamma chain genes. This allowed the determination of whether two T cell clones shared the same T-cell receptor and thus arose from identical, clonally expanded (oligoclonal) progenitor T cells. As an extension of previous studies, oligoclonal T-cell clones were identified in both cerebrospinal fluid and blood populations in 5 of 9 patients with inflammatory demyelinating disease among a total of 486 blood and cerebrospinal fluid T-cell clones. In contrast, no clonally expanded T-cell populations were found among a total of 424 clones derived from either blood of 4 normal control subjects or blood and cerebrospinal fluid of 8 patients with other neurological diseases. Analysis of T-cell receptor V beta genes among 4 oligoclonal T-cell populations derived from 3 patients with multiple sclerosis demonstrated common usage of the V beta 12 gene segment. These data suggest that oligoclonal T cells share similar specificities and that clonal expansion may have resulted from specific stimulation by an antigen. Moreover, identical clones between blood and cerebrospinal fluid were observed in 3 of 9 patients with demyelinating disease, thus providing further evidence of an equilibrium between peripheral and central nervous system immune compartments.     

耐药性癫痫患者血清和脑脊液P-糖蛋白的表达水平变化及其临床意义

目的 观察耐药性癫痫患者血清和脑脊液中多药耐药基因-1、P-糖蛋白的表达水平变化及其临床意义。方法 选取2014年1月-2016年4月本院收治的耐药性癫痫患者以及同期在本院接受抗癫痫药物治疗控制良好的癫痫患者各85例,分别作为耐药性癫痫组和治疗有效组。另选取健康人群85例作为健康对照组。采用酶联免疫吸附测定法及荧光定量聚合酶链反应技术检测所有受试人员的外周血清及脑脊液中多药耐药基因-1及P-糖蛋白表达水平,比较不同类型癫痫患者以上两项指标的表达水平变化。结果 与治疗有效组、健康对照组比较,难治性癫痫组患者血清P-糖蛋白及MDR1-mRNA表达水平较其他2组均明显增高(P<0.05,P<0.01); 治疗有效组与健康对照组比较,血清P-糖蛋白及MDR1-mRNA表达水平无明显差异(P>0.05)。与治疗有效组比较,耐药性癫痫组脑脊液P-糖蛋白及MDR1-mRNA表达水平明显增高(P<0.05)。耐药性癫痫组中不同发作类型患者血清MDR1-mRNA、P-糖蛋白表达水平比较,均无明显差异(P均>0.05)。结论 外周血及脑脊液中多药耐药基因-1、P-糖蛋白的表达水平可以作为癫痫耐药的一项重要指标。     

T-cell subsets and interleukin-6 response in Rasmussen's encephalitis

To evaluate the immunopathogenesis in Rasmussen's encephalitis, peripheral lymphocyte subsets and interleukin-6 analysis were performed in three patients. Magnetic resonance spectroscopy and diffusion-weighted magnetic resonance imaging were performed to assess neuronal injury in the affected hemisphere. Before initiation of immune therapy, percentage of cytotoxic T cells was found to be increased in peripheral blood obtained from patients compared with a group of age-matched normal control subjects. During follow-up, percentage of cytotoxic T cells returned to the normal ranges only in one patient who had an early functional hemispherectomy. All three patients had significantly increased interleukin-6 concentration in cerebrospinal fluid and serum compared with the mean values of patients with acute viral encephalitis. The magnitude of interleukin-6 response in the patients correlated with the neuronal loss and atrophy on magnetic resonance spectroscopy and diffusion-weighted magnetic resonance imaging studies. The patient, who had a fulminant course and an early hemispherectomy, had higher interleukin-6 concentration in cerebrospinal fluid and serum than those of the other two. Detection of an increased percentage of cytotoxic T cells in peripheral blood supports the presence of a T cell-mediated inflammatory pathogenesis in Rasmussen's encephalitis. However, elevated interleukin-6 response might reflect the magnitude of the inflammatory process in the affected hemisphere.     

视神经脊髓炎谱系疾病患者脑脊液蛋白水平与致残状况的相关性研究

目的探讨视神经脊髓炎谱系疾病(Neuromyelitis optica disorders,NMOSDs)患者脑脊液蛋白与临床致残状况的关系及其临床意义。方法回顾性分析2010年1月~2016年2月我院神经内科确诊的108例NMOSD患者临床与生化资料,根据临床扩展致残量表评分(Expanded disability status scale,EDSS)将患者分为独立行走受限与不受限组,并比较两组患者间的临床、生化特征;根据脑脊液蛋白值将患者分为脑脊液蛋白正常组与异常组,并比较两组间EDSS评分的变化;分析脑脊液蛋白与临床致残状况的相关性。结果独立行走受限组与不受限组之间患者年龄、二便障碍、外周血白细胞(WBC)、中性粒细胞比(N%)、脑脊液蛋白、脑脊液Ig G较有统计学差异(P0.05);性别、病程、视力损害、AQP4抗体、24 h鞘内合成率、寡克隆带比较无统计学差异(P0.05),两组不同脑脊液蛋白水平与患者EDSS值比较有统计学差异(T=3.13,P=0.002);脑脊液蛋白水平与独立行走受限呈正相关(r=0.286,P0.01)。结论 NMOSD患者脑脊液蛋白值越高,患者致残发生的可能性越大,脑脊液蛋白有助于视神经脊髓炎谱系疾病患者致残状况的病情评估。     

Immune complexes in serum and cerebrospinal fluid of multiple sclerosis patients and patients with other neurological diseases

Paired serum and cerebrospinal fluid (CSF) specimens from 30 multiple sclerosis (MS) patients and 30 patients with other neurological diseases (ONDs) were analyzed for the presence of immune complexes (ICs). With each of the 4 tests used, ICs were found more frequently in sera from both MS and OND patients than in sera from healthy blood donors. IC-positivity for MS and OND patient CSF varied from 10-33 % and from 10-17 % in different tests. The number of IC-positive sera or CSF in MS patients did not differ significantly from those in OND patients. For both MS and OND patients, the positivity pattern for serum and CSF specimens in each IC test was essentially unique. Furthermore, because several CSF IC-positive and serum IC-negative paired specimens were found, intrathecal IC formation may be independent of IC formation in peripheral blood. The presence of ICs in serum or CSF did not correlate with the clinical status of or laboratory data on the MS patients, nor was a correlation found with the diagnosis of the OND patients. In total, these results suggest that the presence or absence of ICs in MS or OND patients may simply reflect changes in the immunological regulation of individual patients.     

Clinical stabilization and effective B-lymphocyte depletion in the cerebrospinal fluid and peripheral blood of a patient with fulminant relapsing-remitting multiple sclerosis

BACKGROUND: The anti-CD20 monoclonal antibody rituximab effectively depletes B lymphocytes and has been successfully used in the therapy of immune-mediated disorders of the peripheral nervous system. A limited effect of rituximab on B lymphocytes in the cerebrospinal fluid compartment of patients with primary progressive multiple sclerosis (MS) was recently reported. OBJECTIVE: To determine the effect of rituximab on clinical, magnetic resonance imaging, and immunological variables in a patient with relapsing-remitting MS. DESIGN: A patient with relapsing-remitting MS was treated with rituximab. The patient was repeatedly examined clinically and by magnetic resonance imaging. The frequency of peripheral blood and cerebrospinal fluid B lymphocytes was assessed by flow cytometry before, during, and after rituximab therapy. RESULTS: Rituximab monotherapy resulted in significant clinical improvement. Inflammatory surrogate markers on magnetic resonance imaging were also reduced. B lymphocytes were depleted in the cerebrospinal fluid and peripheral blood. CONCLUSIONS: Our data demonstrate beneficial clinical effects of rituximab in relapsing-remitting MS, mediated through modulation of humoral systemic and central nervous system intrinsic immune responses. Clinical trials should determine optimal therapeutic strategies for patients with relapsing-remitting MS.     

Oxidant and antioxidant parameters in the treatment of meningitis

The aim of this study was to assess the effects of meningitis treatment on the serum and cerebrospinal-fluid oxidant and antioxidant status in children with bacterial meningitis. Forty children with bacterial meningitis, at ages ranging from 4 months to 12 years (mean age, 4 years), were enrolled in the study. Within 8 hours after admission (before treatment) and 10 days after clinical and laboratory indications of recovery (after treatment), cerebrospinal fluid and venous blood were collected. Thirty-seven healthy children (mean age, 4 years) were enrolled as control subjects, and only venous blood was collected. Serum total oxidant status, lipid hydroperoxide, oxidative stress index, uric acid, albumin, and ceruloplasmin levels were lower in the patient group after treatment (P<0.05). Serum total antioxidant capacity levels, vitamin C, total bilirubin, and catalase concentrations were not significantly altered by treatment (P>0.05). However, cerebrospinal fluid total oxidant status, lipid hydroperoxide, and oxidative stress index levels were higher, and cerebrospinal fluid total antioxidant capacity levels were lower after treatment than before treatment (P<0.05). In conclusion, we demonstrated that serum oxidative stress was lower, and cerebrospinal fluid oxidative stress was higher, after rather than before treatment in children with bacterial meningitis.     

协同刺激分子在格林-巴利综合征患者 脑脊液、血及周围神经中的表达

目的　探讨协同刺激分子在格林 巴利综合征 (GBS)患者脑脊液、血和周围神经组织中的表达及其作用。方法　用RT PCR法及原位杂交法观察GBS患者外周血、脑脊液和周围神经组织中协同刺激分子B7 1、B7 2、CD2 8、CTLA 4的表达。结果　GBS患者外周血、CSF和周围神经组织中协同刺激分子CD2 8、B7 1、B7 2在炎性细胞上的表达明显高于对照组 ;GBS患者CSF中CD2 8、B7 1、B7 2mRNA的表达明显高于外周血。结论　在GBS发病过程中协同刺激分子的表达对T细胞的活化 ,进而导致体液免疫和细胞免疫反应起重要作用。