二氧化硫吸入对小鼠脾细胞凋亡的诱导作用

目的:探讨二氧化硫大剂量吸入对小鼠脾细胞凋亡和脾脏组织学结构的影响.方法:以不同浓度的二氧化硫分别对小鼠连续染毒7d,用透射电镜法、DNA琼脂糖凝胶电泳法和流式细胞技术观察小鼠脾脏组织学结构和细胞凋亡改变.结果:在二氧化硫染毒组小鼠脾脏的红髓区和白髓区均有典型的脾细胞凋亡发生,在边缘区可见大量核变形淋巴细胞,此外还可见巨噬细胞出现明显的凋亡改变和网状内皮细胞受损.168mg/m3二氧化硫染毒可引起小鼠脾细胞凋亡率增加.结论:大剂量二氧化硫吸入可引起小鼠脾脏超微结构改变,在一定剂量和时间范围内可引起脾细胞凋亡加速,从而对机体的免疫功能造成一定损伤.     

用SYBR-Green Ⅰ实时荧光PCR结合融解曲线分析法诊断α-地中海贫血

目的建立自动化、高通量、准确快速检测缺失型α-地中海贫血基因型的技术。方法应用SYBR-Green1进行两个实时荧光聚合酶链反应(real-time fluorescence polymerase chain reaction with SYBR-Green1,SYBR-PCR),检测左缺(-α4.2)、右缺(-α3.7)等位基因,同时进行融解曲线(dissociation curve,DC)和Tm(melting temperature)值分析。PCR产物重组到pCR2.1,重组子梯度稀释作为模板检测灵敏度,确定两种等位基因型(-α4.2,-α3.7)的检测下限,并对110份DNA样品进行检测。结果检测-α4.2和-α3.7的PCR产物长度分别为1.65kb、1.9kb,Tm分别为(81.5±0.5)℃、(82.5±0.5)℃,检测下限分别为9×102个拷贝、4.3×102个拷贝。该检测技术的灵敏度较常规PCR结合琼脂糖凝胶电泳法高10倍。结论SYBR-Green1实时荧光PCR结合融解曲线分析及Tm分析可以灵敏、准确地检测-α4.2、-α3.7、αα(包括αTα)及--SEA4种等位基因,从而为各种缺失型α-地中海贫血做出基因诊断。该技术具有自动化程度高,不需荧光标记探针,成本低,易质控,防污染,高通量等优点,适于临床推广应用。     

NaCl味觉刺激对钠缺乏大鼠臂旁核c-fos表达的影响

目的:探讨臂旁核(PBN)在钠平衡行为调节中的作用。方法:成年雄性SD大鼠20只,分为对照组、糖精溶液口腔灌流组(Sac组)、NaCl溶液口腔灌流组(NaCl组)、注射furosemide后NaCl溶液口腔灌流组(Furo-NaCl组),观察大鼠对味刺激的摄取反应和各组PBN各亚核内c-fos表达水平差异。结果:正常大鼠对糖精溶液显示嗜好性反应,对NaCl溶液显示厌恶性反应,但钠缺乏大鼠对NaCl也表现出嗜好性摄取反应。Sac组PBN各亚核内c-fos表达水平均高于对照组;NaCl刺激增加内侧亚核(ms)内的c-fos表达,但减少外部外侧亚核(els)内的c-fos表达;Furo- NaCl组els和中央外侧亚核(cls)内c-fos表达显著高于其他3组,ms内的c-fos表达低于NaCl组,但仍高于对照组和Sac组。结论:els和cls在钠平衡的味觉嗜好调节中发挥重要作用。     

Stromal cell-derived factor-1/CXCR4 signaling modifies the capillary-like organization of human embryonic stem cell-derived endothelium in vitro.

The molecular mechanisms that regulate human blood vessel formation during early development are largely unknown. Here we used human ESCs (hESCs) as an in vitro model to explore early human vasculogenesis. We demonstrated that stromal cell-derived factor-1 (SDF-1) and CXCR4 were expressed concurrently with hESC-derived embryonic endothelial differentiation. Human ESC-derived embryonic endothelial cells underwent dose-dependent chemotaxis to SDF-1, which enhanced vascular network formation in Matrigel. Blocking of CXCR4 signaling abolished capillary-like structures induced by SDF-1. Inhibition of the SDF-1/CXCR4 signaling pathway by AMD3100, a CXCR4 antagonist, disrupted the endothelial sprouting outgrowth from human embryoid bodies, suggesting that the SDF-1/CXCR4 axis plays a critical role in regulating initial vessel formation, and may function as a morphogen during human embryonic vascular development.     

Neuroprotective potential of ceftriaxone in in vitro models of stroke

Astrocytic glutamate transporters are considered an important target for neuroprotective therapies as the function of these transporters is abnormal in stroke and other neurological disorders associated with excitotoxicity. Recently, Rothstein et al., [Rothstein JD, Patel S, Regan MR, Haenggeli C, Huang YH, Bergles DE, Jin L, Dykes Hoberg M, Vidensky S, Chung DS, Toan SV, Bruijn LI, Su ZZ, Gupta P, Fisher PB (2005) Beta-lactam antibiotics offer neuroprotection by increasing glutamate transporter expression. Nature 433:73-77] reported that beta-lactam antibiotics (including ceftriaxone, which easily crosses the blood-brain barrier) increase glutamate transporter 1 (GLT-1) expression and reduce cell death resulting from oxygen-glucose deprivation (OGD) in dissociated embryonic cortical cultures. To determine whether a similar neuroprotective mechanism operates in more mature neurons, which show a different pattern of response to ischemia than primary cultures, we exposed acute hippocampal slices obtained from rats treated with ceftriaxone for 5 days (200 mg/kg; i.p.) to OGD. Whole-cell patch clamp recording of glutamate-induced N-methyl-d-aspartate (NMDA) currents from CA1 pyramidal neurons showed a larger potentiation of these currents after application of 15 microM dl-threo-beta-benzyloxyaspartic acid (TBOA; a potent blocker of glutamate transporters) in ceftriaxone-injected animals than in untreated animals, indicating increased glutamate transporter activity. Western blot analysis did not reveal GLT-1 upregulation in the hippocampus. Delay to OGD-induced hypoxic spreading depression (HSD) recorded in slices obtained from ceftriaxone-treated rats was longer (6.3+/-0.2 vs. 5.2+/-0.2 min; P<0.001) than that in the control group, demonstrating a neuroprotective action of the antibiotic in this model. The effect of ceftriaxone was also tested in organotypic hippocampal slices obtained from P7-9 rats (>14 days in vitro). OGD or glutamate (3.5-5.0 mM) damaged CA1 pyramidal neurons as assessed by propidium iodide (PI) fluorescence. Similar damage was observed after pre-treatment with ceftriaxone (10-200 microM; 5 days) and ceftriaxone exposure did not result in GLT-1 upregulation as assayed by Western blot. Treatment of slice cultures with dibutyryl cAMP (100-250 microM; 5 days) increased GLT-1 expression but did not reduce cell damage induced by OGD or glutamate. Thus we confirm the neuroprotective effect of antibiotic exposure on OGD-induced injury, but suggest that this action is related to independent modulation of transporter activity rather than to the level of GLT-1 protein expression. In addition, our results indicate that the protective effects of beta-lactam antibiotics are highly dependent on the experimental model.     

Induction of specific immune responses by severe acute respiratory syndrome coronavirus spike DNA vaccine with or without interleukin-2 immunization using different vaccination routes in mice

DNA vaccines induce humoral and cellular immune responses in animal models and humans. To analyze the immunogenicity of the severe acute respiratory syndrome (SARS) coronavirus (CoV), SARS-CoV, spike DNA vaccine and the immunoregulatory activity of interleukin-2 (IL-2), DNA vaccine plasmids pcDNA-S and pcDNA-IL-2 were constructed and inoculated into BALB/c mice with or without pcDNA-IL-2 by using three different immunization routes (the intramuscular route, electroporation, or the oral route with live attenuated Salmonella enterica serovar Typhimurium). The cellular and humoral immune responses were assessed by enzyme-linked immunosorbent assays, lymphocyte proliferation assays, enzyme-linked immunospot assays, and fluorescence-activated cell sorter analyses. The results showed that specific humoral and cellular immunities could be induced in mice by inoculating them with SARS-CoV spike DNA vaccine alone or by coinoculation with IL-2-expressing plasmids. In addition, the immune response levels in the coinoculation groups were significantly higher than those in groups receiving the spike DNA vaccine alone. The comparison between the three vaccination routes indicated that oral vaccination evoked a vigorous T-cell response and a weak response predominantly with subclass immunoglobulin G2a (IgG2a) antibody. However, intramuscular immunization evoked a vigorous antibody response and a weak T-cell response, and vaccination by electroporation evoked a vigorous response with a predominant subclass IgG1 antibody response and a moderate T-cell response. Our findings show that the spike DNA vaccine has good immunogenicity and can induce specific humoral and cellular immunities in BALB/c mice, while IL-2 plays an immunoadjuvant role and enhances the humoral and cellular immune responses. Different vaccination routes also evoke distinct immune responses. This study provides basic information for the design of DNA vaccines against SARS-CoV.     

复合肌肉动作电位与周围神经损伤程度的相关性研究

目的:分析45例腓总神经不全损伤患者复合肌肉动作电位（CMAP）的波幅、时程,探讨其与周围神经损伤程度的关系。方法 :选择2012年1月至2012年12月就诊于我院的45例不同程度腓总神经不全损伤患者,根据患者胫前肌肌力级别将45例患者分为三组（n=15）:胫前肌肌力4-5级（P₁组）、胫前肌肌力2-3级（P₂组）、胫前肌肌力1级（P₃组）。应用肌电诱发电位仪记录45例患者的CMAP波形,测量CMAP波幅与时程。采用SPSS13.0统计软件对数据进行秩和检验,分析各组间CMAP波幅与时程的差别。结果 :（1）P₁组、P₂组和P₃组腓总神经-胫前肌CMAP平均波幅分别为7.1±0.2mv、3.3±0.3mv和0.5±0.1mv;P₂组、P₃组与P₁组相比,波幅均有显著性差异（P<0.05）;P₃组与P₂组相比,波幅亦有显著性差异（P<0.05）。（2）P₁组、P₂组和P₃组腓总神经-胫前肌CMAP平均时程分别为11.4±0.4ms、16.9±0.6ms和23.3±1.2ms;P₂组、P₃组与P₁组相比,时程均有显著性差异（P<0.05）;P₃组与P₂组相比,时程亦有显著性差异（P<0.05）。结论 :（1）CMAP波幅、时程有助于评价周围神经受损的严重程度。（2）CMAP波幅降低提示有功能的周围神经轴突数量减少,时程延长反映了周围神经的脱髓鞘损害。     

显微手术治疗高血压性脑出血的临床体会

目的探讨小骨窗开路经颞叶颞叶皮质显微外科清除血肿的手术方法,提高对高血压脑出血的治疗水平。方法回顾性分析我科自2007年3月至2010年3月基底节区高血压性脑出血患者大骨瓣开颅减压以及采用经颞叶皮质入路以显微外科手术治疗126例,总结其手术方法,比较其入路与其他手术入路的优缺点,探讨手术技巧。结果 126例患者中完全清除血肿90例、大部分清除血肿15例,再出血5例、存活105例、死亡21例。根据日常生活活动能力评分（activities of daily living,ADL）分级,手术后随访3个月,105例生存者预后达I级者6例,Ⅱ级43例,Ⅲ级18例,Ⅳ级26例,Ⅴ级13例。结论显微手术治疗基底节区脑出血,手术创伤小,时间短,止血可靠,安全有效。手术脑组织损伤小,可缩短昏迷时间,减少并发症,可提高患者生存率和生存质量。     

Hyperhomocysteinemia Associates with Small Vessel Disease More Closely Than Large Vessel Disease

Background: Hyperhomocysteinemia was believed to be an independent risk factor for stroke and associate with small vessel disease (SVD) related stroke and large vessel disease (LVD) related stroke differently. However it''s still unclear which type of stroke associated with homocysteine (HCY) more strongly because the conclusions of previous studies were contradictory. In this study we focused on the subclinical angiopathies of stroke, i.e., SVD and LVD instead of stroke subtypes and sought to compare the associations between HCY level and different angiopathies. Methods: 324 non-stroke patients were enrolled. Sex, age, HCY level and other vascular risk factors were collected. MRI and angiographies were used to determine the type of angiopathies and their severity, i.e., the scores of leukoaraiosis (LA), plaques and numbers of silent brain infarctions (SBI). LVD was defined as the presence of atherosclerotic plaques of cerebral arteries. SVD was defined as the presence of either LA or SBI. 230 patients were deemed to have LVD; 180 patients were deemed to have SVD. Spearman''s correlation test and logistic regression were used to analyze the association between HCY level and different angiopathies. Results: The correlation between HCY level and scores of plaques was weaker than that of the scores of LA and numbers of SBI. Hyperhomocysteinemia was an independent risk factor for SVD (OR = 1.315, P <0.001), whereas the association between HCY level and LVD was not that significant (OR = 1.058, P = 0.075). Conclusion: HCY level associated with SVD more strongly than LVD.     

Survey on hospital‐acquired urinary tract infection in neurological intensive care unit

This study aimed to explore the causes, incidence, and risk factors of urinary tract infection patients in neurological intensive care unit (ICU). Patients (n = 916) admitted to the neurological ICU from January 2005 to December 2010 were retrospectively surveyed for urinary tract infections. There were 246 patients in neurological ICU who were diagnosed with hospital‐acquired urinary tract infection during that period of time (26.9%). Forty‐three cases were upper urinary tract infection, and 203 cases were lower urinary tract infection. The top three strains were Escherichia coli, Enterococcus faecalis, and Klebsiella pneumoniae. Older age (UTI rate, 22.6%), female patients (21.7%), hospital stay for more than 7 days (16.7%), diabetes (11.7%), and catheterization (21.1%) were the risk factors for hospital‐acquired urinary tract infection. There is a high incidence of nosocomial urinary tract infection in the neurological intensive care unit. Active prevention program and surveillance need to be carried out in neurological ICU, especially in those with risk factors.