犬贾第虫端粒重复序列的Southern印迹杂交分析

目的确定犬贾第虫端粒DNA的重复序列，为进行端粒酶活性检测奠定基础。方法根据已发表的蓝氏贾第虫的端粒重复序列5＇-TAGGG-3’设计寡聚核苷酸探针（TAGGG）5，并以地高辛标记，将犬贾第虫基因组DNA经Bal31-EcoRI酶切后，进行Southern印迹杂交分析。结果犬贾第虫基因组DNA与探针（TAGGG）。杂交，获得了较好的杂交条带，随Bal31酶切时间的延长，杂交信号逐渐减弱。结论犬贾第虫端粒重复序列定位在染色体的末端序列与国外报道的贾第虫端粒重复序列一致，为5＇-TAGGG-3’。     

人端粒保护蛋白1的结构与功能

人类端粒DNA由富含G的短片段重复序列(TTAGGG)串联组成,包括双链和3’端突出的单链两部分,不能编码蛋白质〔1-2〕。通常认为端粒酶是调控端粒DNA长度最重要的因素,端粒酶活性的降低将导致端粒长度的下降,细胞分裂停滞。端粒酶活性通常由其催化亚单位(端粒酶逆转录酶)所决定〔3-4〕,但奇怪的是,端粒双链DNA结合蛋白TRF1与TRF2,也能调控端粒酶活性,但它们并无直接联系。最近的研究显示人端粒保护蛋白1     

改良的端粒DNA检测方法及其在无偿献血中的应用

目的 建立非同位素标记的探针杂交测定外周血白细胞端粒DNA长度的方法，并探讨其在无偿献血中的应用。方法 酚／氯仿提取基因组DNA，限制性内切酶消化，琼脂糖凝胶电泳，非同位素标记的探针Southern印迹杂交，化学发光X线片曝光显示杂交谱带，积分光密度扫描计算TRF值。结果 所测样本获得了较好的低背景杂交谱带，测得35～40岁无偿献血组TRF值平均为11．73kb，相应无献血史对照组平均为11．78kb。结论 建立了非同位素标记的探针检测端粒DNA的方法，用上述方法初步显示了固定的长期无偿献血并未对端粒的长度产生影响。     

马来丝虫非放射性寡核苷酸探针的制备及现场应用研究

目的 寻找一种高度特异，敏感的马来丝虫的检测方法，并将其用于现场检测。方法 根据马来丝虫HhaⅠ重复序列，合成马来丝虫特异性引物1对和寡核苷酸探针1条，采用地高辛加尾标记试剂盒对人工合成寡核苷酸探针进行非放射性标记，并对标记后的探针进行标记效率检测，将血液标本和蚊虫标本分别用蛋白酶K等处理液和Chelex-100处理后，结合PCR技术，对该探针的特异性和敏感展出一进行检测；并将此探针用于平远县马来丝虫病的监测。结果 经测定，标记后探针的有效浓度为1.5-2fmol/μl；结合PCR技术，该探针可检出100μl阳性血样中1条微丝蚴；50只蚊中含有1只只感染1条幼丝虫阳性蚊亦能准确检出，班氏丝虫等其它寄生虫标本的检测结果均为阴性；采自平远县的240份血液标本和1008只中华按蚊标本均为阴性。结论 该探针杂交检测方法特异性强，敏感性高，可作为一种新的马来丝虫病监测方法在现场中使用。     

马来丝虫非放射性寡核苷酸探针的制备及现场应用研究

目的　寻找一种高度特异、敏感的马来丝虫的检测方法 ,并将其用于现场检测。　方法　根据马来丝虫 Hha 重复序列 ,合成马来丝虫特异性引物 1对和寡核苷酸探针 1条 ,采用地高辛加尾标记试剂盒对人工合成寡核苷酸探针进行非放射性标记 ,并对标记后的探针进行标记效率检测 ;将血液标本和蚊虫标本分别用蛋白酶 K等处理液和 Chelex- 10 0处理后 ,结合 PCR技术 ,对该探针的特异性和敏感性进行检测 ;并将此探针用于平远县马来丝虫病的监测。　结果　经测定 ,标记后探针的有效浓度为 1.5～ 2 fmol/μl;结合 PCR技术 ,该探针可检出 10 0 μl阳性血样中 1条微丝蚴 ;5 0只蚊中含有 1只只感染 1条幼丝虫阳性蚊亦能准确检出。班氏丝虫等其它寄生虫标本的检测结果均为阴性 ;采自平远县的 2 4 0份血液标本和 10 0 8只中华按蚊标本均为阴性。　结论　该探针杂交检测方法特异性强 ,敏感性高 ,可作为一种新的马来丝虫病监测方法在现场中使用。     

端粒酶与肿瘤

端粒酶是一种特殊的逆转录酶，是一种由RNA和蛋白质组成的核糖核蛋白酶，人类的端粒酶是由端粒酶RNA成分、端粒酶催化组分和端粒酶相关蛋白三个亚单位组成。它能够利用3′－端粒作为引物，以自身RNA组分为模板来逆转录合成端粒DNA序列，并不断添加到染色体末端，从而维持端粒长度的稳定。端粒酶与肿瘤的发生发展有密切关系，本文对端粒酶活性与肿瘤的早期诊断、肿瘤细胞的病理分化程度、肿瘤细胞的耐药性、肿瘤的临床分期和肿瘤患者的生存期的关系进行综述，研究结果表明端粒活性可能与肿瘤的预后有一定的关系。     

端粒酶hTR反义寡核苷酸对K562细胞端粒酶活性的影响

目的：探讨端粒酶RNA反义寡核苷酸（antisense oligodeoxynucleotide,ASODN)对人白血病K562细胞端粒酶活性及细胞生长的影响。方法：应用与人端粒酶RNA组分模板区互补的硫代ASODN处理K562细胞，用端粒酶重复扩增分析（telomere repeat amplification protocol,TRAP)-PCR-ELISA检测法观察端粒酶活性的变化；用Annexin V分析法及流式细胞术检测凋亡细胞。结果：经ASODN作用后，细胞端粒酶活性明显受到抑制，并与ASODN的浓度和处理时间相关。作用5d后，凋亡细胞明显增加。结论：端粒酶RNA模板区ASODN可明显抑制白血病K562细胞端粒酶活性，抑制细胞生长和增殖，诱导细胞凋亡，可能成为白血病基因治疗新靶点。     

骨肉瘤特异生物诊断探针制备

目的构建与骨肉瘤细胞高亲和性与特异性结合的DNA寡核苷酸适配体库,用于骨肉瘤临床特异性诊断。方法体外合成40 nt高通量随机序列的单链DNA寡核苷酸文库(ssDNAs);骨肉瘤细胞(U-2 OS)作为目标细胞与文库ssDNAs反应;指数富集配体的系统进化(SELEX)技术用于筛选富集与靶细胞结合的寡核苷酸;聚合酶链反应(PCR)技术与亲和层析技术用于次轮反应文库的制备;应用荧光光谱跟踪筛选过程;对最后的筛选序列进行克隆、测序与保存。结果十一轮至十三轮筛选后的ssDNAs荧光光谱曲线显示了有意义重叠,因此结束筛选;对十三轮筛选的ssDNAs进行了克隆测序,依据一级结构的同源序列建立了寡核苷酸探针组群。结论应用SELEX技术最终从高通量的随机文库中获得了与U-2 OS具有高度亲和性的寡核苷酸适配体,适配体库的建立不仅仅用于骨肉瘤的特异性诊断,也将为探针-载体-抗肿瘤药物复合体的研制与实施骨肉瘤定向治疗搭建了可行性平台。     

湖北钉螺多态微卫星DNA位点筛选和特征初步分析

目的分离湖北钉螺的微卫星DNA序列,筛选多态的微卫星DNA位点并分析其特征。方法应用湖北钉螺基因组DNA的酶切片段与生物素标记的(AAT)17、(GA)25、(CCT)17、(CA)25等10个寡核苷酸探针杂交,富集、浓缩、克隆并测序,构建微卫星DNA库。挑选合适的微卫星DNA位点设计并合成引物,扩增钉螺样本经聚丙烯酰胺凝胶电泳筛选多态性。结果获得湖北钉螺微卫星DNA序列205条,GenBank注册登记号GU204044～GU204248,其中完整重复序列74条,占36.10%;非完整重复序列102条,占49.76%;复合重复序列29条,占14.15%。设计合成的20对微卫星DNA位点引物中,经鉴定显示13个位点具有多态性。结论分离建立了湖北钉螺微卫星DNA序列库,为湖北钉螺群体遗传、种群溯源等相关研究提供了分子标志。     

聚合酶链反应结合非放射性标记探针检测粪便中微小隐孢子虫

目的：利用非放射性标记探针技术检测经聚合酶链反应扩增后的人粪便中微小隐孢子虫DNA，观察该方法的特异性和敏感性。方法：用裂解法从粪便标本直接制备微小隐孢子虫模板DNA，用一对人工合成的寡核苷酸作PCR引物，扩增长度为452bp的微小隐孢子虫目的DNA片段。将扩增产物直接点在或经Southern印迹法转移到硝酸纤维素膜上，再和生物素标记的寡核苷酸探针杂交，经底物（BCIP）呈色观察。结果：阳性杂交信号只见于微小隐孢子虫DNA扩增产物，而约氏疟原虫、杜氏利什曼原虫、贾第虫、白色念珠菌、大肠杆菌和人白细胞DNA均无杂交信号。PCR结合斑点杂交或Southern印迹法检测隐孢子虫DNA 的敏感性均为011 pg。结论: 非放射性探针检测人粪便隐孢子虫DNA 的PCR 扩增产物, 具有敏感性高、特异性强、操作较简便及无放射性污染等优点。     

Research and Evaluation of the Influence of the Construction of the Gate and the Influence of the Piston Velocity on the Distribution of Gases into the Volume of the Casting

Distribution of gasses to the cast volume and volume of pores can be maintained within the acceptable limits by means of correct setting of technological parameters of casting and by selection of suitable structure and gating system arrangement. The main idea of this paper solves the issue of suitability of die casting adjustment—i.e., change of technological parameters or change of structural solution of the gating system—with regards to inner soundness of casts produced in die casting process. Parameters which were compared included height of a gate and velocity of a piston. The melt velocity in the gate was used as a correlating factor between the gate height and piston velocity. The evaluated parameter was gas entrapment in the cast at the end of the filling phase of die casting cycle and at the same time percentage of porosity in the samples taken from the main runner. On the basis of the performed experiments it was proved that the change of technological parameters, particularly of pressing velocity of the piston, directly influences distribution of gasses to the cast volume.     

Weighing the QT Intervals with the Slope or the Amplitude of the T Wave

Objective: The reproducibility of QT interval measurements is low, even for the mean QT interval based on the standard ECG. In this study we analyzed whether the reproducibility of the mean weighed QT interval was better than the simple mean QT interval. The weighing was based on the amplitude of the T wave or the slope of the steepest tangent on the terminal part of the T wave. Material and methods: 12‐lead ECGs of 130 postmyocardial infarction patients were obtained. The QT intervals were measured by the tangent‐method on two occasions by the same observer Mismatch QT intervals were defined as QT intervals that were measured at only one occasion. Sixteen ECGs were rejected. The data were split into 34 and 80 ECGs for optimization and validation of the weighing, respectively. The weighed QT dispersion was calculated as the weighed mean of the three longest minus the weighed mean of the three shortest QT intervals. Results: Weighing with the slope increased the reproducibility by 41% (P = 3 10^‐6), but weighing with the amplitude reduced it by 20% (P = 0.02). However, if measurements with errors above 75 ms were rejected, weighing with the slope or the amplitude increased the reproducibility with 26% and 20% (P = 0.02), respectively. Weighing did not change the reproducibility of the weighed QT dispersion. Conclusion: Weighing with the slope improved the reproducibility of the mean weighed QT interval. However, if measurements with errors above 75 ms were rejected, weighing with the amplitude also increased the reproducibility. Weighing did not change the reproducibility of the weighed QT dispersion. Weighing is particularly efficient at reducing the negative impact of mismatch QT intervals on the reproducibility. A.N.E. 2002;7(1):4–9     

县级以下综合医疗机构医务人员对现代结控知识的认识现状调查

目的本文旨在了解医务人员现代结控知识掌握的现状及培训效果?方法于培训前后进行问卷调查,内容包括:病例发现?结核病诊断及化疗?结果培训前疫情报告和转诊,回答正确者占75.2%?71.7%;对临床表现?查痰和诊断依据,回答正确者占83.5%?42.5%?40.8%;抗痨药物?用药方法?化疗原则?短化方案?短化疗程?治愈标准六项,回答正确者占58%?14.4%?20.8%?9.2%?17%?24.3%?培训后再次调查发现,90%以上医务人员对现代结控基本知识已掌握?结论各级医务人员现代结控知识是很贫乏的,因此,对其进行系统培训是极为必要的,此项工作省时?省力?投入少,可收到事半功倍的效果。     

大骨节病病区水,粮实验对动物的骨和软骨中化学元素组成的影响

用质子激发 X 线荧光分析方法(PIXE)测定了大骨节病病区和非病区的水、粮以及用该水粮喂养的大白鼠的肋软骨和硬骨中22种化学元素的含量。结果发现水粮中存在差异的元素反应在用该水粮喂养的大白鼠的骨、软骨中也存在差异,含量都低的元素有 P、Mn、Cu、As、Zn。都高的有铅。其中锌低在水、粮、硬骨和软骨中都完全一致呈非常显著性差异(p<0.01)。提示病区水、粮中化学元素对骨质的影响不是单一元素缺乏或过多所致,而是多种元素的复合因素。     

Assessment of the Soft-Tissue Seal at the Interface between the Base of the Fixed Denture Pontic and the Oral Mucosa

Fixed dentures (bridges) are often selected as a treatment option for a defective prosthesis. In this study, we assess the contact condition between the base of the pontic and oral mucosa, and examine the effect of prosthetic preparation and material biocompatibility. The molars were removed and replaced with experimental implants with a free-end type bridge superstructure after one week. In Experiment 1, we assessed different types of prosthetic pre-treatment: (1) the untreated control group (Con: mucosa recovering from the tooth extraction); (2) the laser irradiation group (Las: mucosa recovering after the damage caused by a CO₂ laser); and (3) the tooth extraction group (Ext: mucosa recovering immediately after the teeth extraction). In Experiment 2, five materials (titanium, zirconia, porcelain, gold-platinum alloy, and self-curing resin) were placed at the base of the bridge pontic. Four weeks after the placement of the bridge, the mucosa adjacent to the pontic base was histologically analyzed. In Experiment 1, the Con and Las groups exhibited no formation of an epithelial sealing structure on the pontic base. In the Ext group, adherent epithelium was observed. In Experiment 2, the sealing properties at the pontic interface were superior for titanium and the zirconia compared with those made of porcelain or gold-platinum alloy. In the resin group, a clear delay in epithelial healing was observed.     

高胆红素血症对肿瘤标志物测定的影响探讨

目的探讨高胆红素血症对Ca19-9、Ca24-2和CEA检测的影响.方法对320例胆管、胆囊良恶性疾病病人,15例胆囊炎病人的胆汁和血清以及10例肝硬化、10例黄疸肝炎病人进行Ca19-9、Ca24-2和CEA检测.结果在良性胆管、胆囊疾病中,Ca19-9的假阳性最高;在胆红素增高的良性疾病中,Ca19-9假阳性率达46.7%;15例胆汁和血清以及10例肝硬化和10例黄疸肝炎病人中,Ca19-9的假阳性率分别为93%、20%、80%和80%.结论高胆红素血症对Ca19-9检测影响最明显,胆囊、胆管良恶性疾病鉴别时,以Ca24-2和CEA检测为佳.     

Echocardiography in the Emergency Department: From the Perspective of the Emergency Physician and the Cardiologist

Echocardiography performed in the emergency department must adapt to this new setting for noninvasive diagnostic testing. Emergency physicians require echocardiography to provide rapid diagnosis in life-threatening emergencies. New initiatives are being proposed by emergency physicians in the delivery of this test. Cardiologists now use echocardiography in the emergency-department to make the diagnosis of heart disease earlier and with greater accuracy.