Therapeutic potential of chlorotoxin-like neurotoxin from the Chinese scorpion for human gliomas

Chlorotoxin, one of the key toxins in scorpion Leiurus quinquestriatus venom, has been shown to bind specifically to glioma cell surface as a specific chloride channel blocker. In this study, a purified, recombinant chlorotoxin-like peptide from the scorpion Buthus martensii Karsch (named rBmK CTa) was characterized by in vivo and in vitro studies. The results from cell proliferation assay with human glioma (SHG-44) cells showed that rBmK CTa inhibits the growth of glioma cells in a dose-dependent manner, with an IC₅₀ value of approximately 0.28 μM. Under the same conditions, the IC₅₀ value for normal astrocytes increased to 8 μM. This clearly indicated that rBmK CTa had specific toxicity against glioma cells but not astrocytes. Results from whole-cell patch-clamp recording showed that chloride current in SHG-44 was inhibited by rBmK CTa in a voltage-dependent manner and percent inhibitions for the blocking action of rBmK CTa (0.07 and 0.14 μM) on I_Cl was 17.64 ± 3.06% and 55.86 ± 2.83%, respectively. Histological analysis of rBmK CTa treated mice showed that brain, leg muscle and cardiac muscle were the target organs of this toxin. These results suggest that rBmK CTa may have potential therapeutic application in clinical treatment of human glioma. It represents an approach for developing a novel therapeutic agent.     

ABO血型基因分型及应用

目的 :研究ABO血型基因分型的意义。方法 :采用聚合酶链反应 序列特异性引物 (PCR SSP)基因定型方法对ABO血型基因定型并观察其基因多态性分布特征和疑难血型检定。结果 :对已知ABO基因的DNA标本进行基因定型 ,证实文中的ABO基因定型方法可靠 ;对 10 4例健康、无血源关系的汉族个体ABO血型基因定型 ,结果与血清学所定表型完全符合 ;并用ABO基因分型技术解决临床输血前血型鉴定、产前胎儿血型鉴定、亲权试验及血清学亚型的正确性验证。结论 :ABO血型基因分型技术可以正确判定ABO血型疑难样本     

急性丙型肝炎患者免疫状况的研究

用间接免疫荧光法、酶联免疫吸附试验（ＥＬＩＳＡ）及ＬＤＨ释法，对１６例急性输血后丙型肝炎（抗－ＨＣＶ、ＨＣＶ－ＲＮＡ均阳性）患者，分别进行外周血单个核细胞（ＰＢＭＣ）的Ｔ细胞亚群计数、Ｔ４／Ｔ８比值、Ｔａｃ受体的检测及血清可溶性白细胞介素２受体（ｓＩＬ－２Ｒ）ＮＫ细胞活性的测定。并与正常人组比较，经ｔ检验发现，急性丙型肝炎患者的Ｔ４亚群所占百分比、Ｔ４／Ｔ８比值及ＰＢＭＣＴａｃ受体表达均明显低于正常人组（Ｐ＜０．０５），而ＮＫ细胞活性、血清ｓＩＬ－２Ｒ明显高于正常人组（Ｐ＜０．０５）。患者的这些免疫状态改变，可能对其发病机理的研究有一定意义。     

Activation of the Notch signaling pathway disturbs the CD4+/CD8+, Th17/Treg balance in rats with experimental autoimmune uveitis

Objective and design

The present study aimed to investigate the relationship between the disturbed balance of CD4⁺/CD8⁺, Th17/Treg and the activation of the Notch signaling pathway in experimental autoimmune uveitis (EAU).

Methods

An EAU rat model was induced in Lewis rats, and pathology analysis was performed by hematoxylin and eosin (H&E) staining. CD4⁺, CD8⁺, Th17, and Treg levels in spleen, lymph nodes and eye tissues were determined by flow cytometry. Meanwhile, the expression of Notch1, DLL4, IL-10, and IL-17 was determined by quantitative polymerase chain reaction (Q-PCR) and enzyme-linked immunosorbent assay (ELISA). In addition, the inhibitory effect of N-(N-(3,5-difluorophenacetyl-l-alanyl))-S-phenylglycine t-butyl ester (DAPT) on Th17 differentiation by Notch signaling in vitro was further investigated using T lymphocytes from EAU rats on day 12 post-immunization by flow cytometry.

Results

The pathological results showed that inflammatory cell infiltration occurred in ocular tissues in EAU rats. The CD4⁺/CD8⁺ and Th17/Treg ratios in EAU rats were apparently higher than those in normal control individuals. Q-PCR and ELISA analyses indicated the expression of Notch1, DLL4, IL-10, and IL-17 in EAU rats gradually increased on day 6 after immunization, peaked on day 12, and then gradually decreased. The dynamic trends in Notch1 and DLL4 expression in EAU rats were identical to those of CD4⁺/CD8⁺ and Th17/Treg levels. DAPT can significantly inhibit the activation of Notch signaling, decrease Th17 cell differentiation, and attenuate the level of the Th17 cell lineage, contributing to the balance of the Th17/Treg ratio.

Conclusion

The activation of the Notch signaling pathway can regulate Th17 and Treg cell differentiation, disrupt the CD4⁺/CD8⁺ and Th17/Treg balance, and aggravate the severity of EAU; inactivation of the Notch signaling pathway contributes to the CD4⁺/CD8⁺ and Th17/Treg balance in EAU rats. Our findings highlighted that the dynamic change in the CD4⁺/CD8⁺ and Th17/Treg ratio was consistent with the expression trend of Notch signaling in EAU rats, suggesting that Notch signaling may be a potentially important therapeutic target in clinical practice.

     

脑源性神经营养因子与脊髓损伤

<正> Spinal cord injury(SCI)is such a serious disease that itwould bring heavy burdens of both health and economy tothe victims as well as the society.Although worldwide med-ical researchers of neuroscience have made great efforts toattempt to solve this big problem for nearly a century, there     

类金刚石薄膜的碳相成分对其血小板黏附特性的影响

对不同工艺和条件制备的 7个类金刚石薄膜 (Diamondlikecarbon ,DLC)试样 ,经X光电子能谱 (XPS)碳相成分分析后 ,分别进行了血小板黏附实验、黏附血小板的形貌观察、分类计数和形态指数计算 ,并通过灰色关联分析 ,研究了碳相成分对血小板黏附量、黏附血小板的形态指数的影响。结果显示 :来自全方位离子注入离子束增强沉积工艺的DLC ,其血小板黏附量和形态指数明显小于等离子体化学气相沉积工艺制备的样品 ;在DLC的 5种碳相成分中 ,DLC碳相与血小板黏附量和形态指数的 (负 )关联度远大于其它碳相成分 ,除此之外只有C H和C O碳相与血小板形态指数的 (正 )关联度较大。表明 :(1)DLC碳相对血小板黏附的影响远较其它碳相成分为大 ,增加DLC碳相的含量是优化DLC血液相容性的关键所在 ;(2 )C H和C O碳相对黏附血小板的变形有促进作用 ,须从工艺上抑制其产生或尽可能降低其含量 ;(3)采用全方位离子注入离子束增强沉积工艺有助于改善DLC的血液相容性。这些结论对设计与改进DLC的制备工艺具有重要的指导意义。     

CT模拟定位三维适形放射治疗食管癌的临床研究

目的探讨CT模拟定位三维适形放射治疗食管癌的疗效。方法1997年4月～2002年7月,经病理证实的食管鳞状细胞癌87例患者，其中男性56例，女性31例，年龄42～75岁，平均年龄62.5岁，随机分为2组，两组患者一般情况、性别、年龄及病变部位构成经统计学检验无统计学意义，一组为采用CT模拟定位进行三维适形放射治疗，另一组为常规模拟定位放射治疗，两组的剂量都为64～66Gy／32.33次／6.4～6.6周。结果采用Kaplan—Meier法计算生存率，并用Log—Rank检验其差异性。治疗计划各指标分析采用kolmogorowSmirnov法行正态分布检验，继而行t检验，以P〈0.05为差异有统计学意义。CT模拟定位三维适形放射治疗组和常规模拟定位放射治疗组1、2、3、4年生存率分别为70.1％、55.6％、40.4％、36.3％和65.4％、50.6％、30.6％、20.9％（p〉0.05）。但是对Ⅰ～Ⅱ期患者，CT模拟定位三维适形放射治疗组和常规模拟定位放射治疗组的生存率分别为81.2％、75.5％、67.3％、61.5％和80.1％、65.7％、54.9％、29.7％（P〈0，05），局部控制率分别为75.6％、71.7％、65.5％、59.0％和65.4％、50.6％、33.3％、25.6％（P〈0.05）。而对于Ⅲ～Ⅳ期的患者，两组之间的生存率和局部控制率差异无统计学意义。结论CT模拟定位三维适形放射治疗的疗效明显好于常规模拟定位放射治疗组。特别对Ⅰ-Ⅱ期早期食管癌患者两组之间差异有统计学意义，而对晚期患者差异无统计学意义。     

Th1型细胞因子基因对结核分枝杆菌基因疫苗诱导BALB/c小鼠产生抗CFP10抗体水平的影响

目的：研究分别表达含IL—12和IL-18基因的质粒，对结核分枝杆菌(Mycobacterium tuberculosis，MTB)H37R1株CFP1O基因疫苗诱导免疫应答的影响。方法：从正常人外周血单个核细胞(PMBCs)中提取RNA，用RT—PCR扩增IL-18 cDNA，并克隆人载体pGEM—Teasy中。测序证实后，亚克隆至真核表达载体pcDNA3．1的BamH Ⅰ和EcoR Ⅰ酶切位点。将分别表达小鼠IL—12和人IL—18基因的真核表达质粒pcmlL12和pclL18，与MTB CFP10基因疫苗联合肌注免疫BALB／c小鼠，共免疫3次，每次间隔2wk。每次免疫后2wk采血、分离血清，用ELISA检测小鼠血清抗CFP10抗体的滴度。结果：用RT—PCR成功地从人PMBC的RNA中扩增出IL—18 cDNA，测序结果正确，用BamH Ⅰ和EcoR Ⅰ酶切鉴定证实，目的基因已插入载体pcDNA3．1中，阳性克隆命名为pcIL18。pcCFP10组第1次免疫后，血清抗CFP10抗体的平均滴度为1：600，末次免疫后的滴度为1：4000。pcIL18 pcCFP10组联合免疫后，血清抗CFP10抗体的滴度高于pcCFP10组，最终达1：8000。而pcmIL12 pcCFP10组联合免疫后滴度仅为1：200。结论：pcIL18与CFP10基因疫苗联合免疫，可增强CFP10抗原的特异性体液免疫应答；pcmIL12则可使CFP10基因疫苗产生的抗体水平降低。pcIL18 pcCFP10基因联合免疫是否具有增强CFP10抗原特异性细胞免疫的作用有待进一步研究。     

妊高征患者产前及产后血小板活化状态的研究

采用流式细胞术对４２例正常晚孕妇女及５０例妊高征患者产前及产后７２小时Ｐ－选择素进行对比研究。结果：妊高征患者Ｐ－选择素含量明显大于正常晚孕妇女，Ｐ－选择素含量在轻、中及重度妊高征患者呈递增趋势，差异均有显著性，产后７２小时其Ｐ－选择素含量降至正常晚孕妇女水平。提示：Ｐ－选择素可作为妊高征早期诊断及监测病情的一个重要指标     

CD54和LFA-1的相互作用在6A8α-甘露糖苷酶表达抑制的JurkatT细胞的黏附性增强中的作用

目的研究CD54和LFA-1的相互作用在6A8 α-甘露糖苷酶表达抑制的Jurkat细胞(AS)的黏附性增强中的作用。方法用细胞凝集试验确证AS细胞间黏附的增强，用细胞与细胞间黏附分子-1-人IgG的Fc片段(ICAM-1-Fc)的黏附试验和阻断性抗CD11a抗体的阻断试验研究CD54-LFA-1的作用，用单克隆抗体MEM-148检测As细胞LFA-1亲和力的变化，用单克隆抗体NKI-L16检测AS细胞LFA-1亲合力的变化，用鬼笔环肽染色细胞骨架，用Jurkat-Raji细胞间的作用作模型研究6A8α-甘露糖苷酶表达抑制对T和B细胞间黏附的影响，用ConA结合试验检测细胞中蛋白质N-糖基化的变化。结果(1)AS细胞间的黏附性增强主要与CD54及CD11a表达的增强相关，也与LFA-1亲和力的增高相关；(2)AS细胞的细胞骨架发生重排；(3)As细胞与Raji细胞间的黏附也增强；(4)ConA与AS细胞的结合增强。结论CD54和LFA-1的相互作用在AS Jurkat T细胞的黏附性增强中起重要作用。细胞骨架重排也可能起作用。As细胞的蛋白质发生了N-糖基化修饰。