人胚胎肺内神经内分泌细胞电镜观察

用透射电镜对不同胎龄的人胚胎肺内神经内分泌细胞进行了发生和超微结构观察。肺内支气管上皮内未分化细胞，第八周开始向神经内分泌细胞转化，提示神经内分泌细胞对早期胚胎肺的发生、发育有特殊意义。神经内分泌细胞内致密核心小泡（ＤＣＶ）及其它与内分泌活动有关的各种细胞器发达。能见到Ｐ０细胞、Ｐ１细胞、Ｐ２细胞及Ｐ３细胞等四种类型的神经内分泌细胞。Ｐ０细胞的发生及分化程度表明它可能是一种前细胞，它可以进一步转化     

Recombinant single-chain variable fragment antibodies directed against Clostridium difficile toxin B produced by use of an optimized phage display system

Recombinant antibody cloning and phage display technologies were used to produce single-chain antibodies (scFv) against Clostridium difficile toxin B. The starting material was the mouse B cell hybridoma line 5A8, which generates a monoclonal antibody against the toxin. The integrated cloning, screening, and phage display system of Krebber et al. (J. Immunol. Methods 201:35-55, 1997) allowed us to rapidly obtain toxin B-binding scFv sequences derived from the hybridoma cell line. The best candidate scFv sequences, based on preliminary enzyme-linked immunosorbent assay (ELISA) screening data were then subcloned into the compatible expression vector. Recombinant single-chain antibodies were expressed in Escherichia coli. A 29-kDa band was observed on polyacrylamide gel electrophoresis as predicted. The expressed product was characterized by immunoblotting and detection with an anti-FLAG antibody. The toxin B-binding function of the single-chain antibody was shown by a sandwich ELISA. The antibody was highly specific for toxin B and did not cross-react with material isolated from a toxin B-negative C. difficile strain. The sensitivity of the soluble single-chain antibody is significantly higher than the original monoclonal antibody based on ELISA data and could detect a minimum of 10 ng of toxin B/well. Competitive ELISAs established that the affinity of the 5A8 parent antibody and the best representative (clone 10) of the single-chain antibodies were similar and in the range of 10(-8) M. We propose that recombinant antibody technology is a rapid and effective approach to the development of the next generation of immunodiagnostic reagents.     

Monte Carlo based treatment planning for modulated electron beam radiation therapy

A Monte Carlo based treatment planning system for modulated electron radiation therapy (MERT) is presented. This new variation of intensity modulated radiation therapy (IMRT) utilizes an electron multileaf collimator (eMLC) to deliver non-uniform intensity maps at several electron energies. In this way, conformal dose distributions are delivered to irregular targets located a few centimetres below the surface while sparing deeper-lying normal anatomy. Planning for MERT begins with Monte Carlo generation of electron beamlets. Electrons are transported with proper in-air scattering and the dose is tallied in the phantom for each beamlet. An optimized beamlet plan may be calculated using inverse-planning methods. Step-and-shoot leaf sequences are generated for the intensity maps and dose distributions recalculated using Monte Carlo simulations. Here, scatter and leakage from the leaves are properly accounted for by transporting electrons through the eMLC geometry. The weights for the segments of the plan are re-optimized with the leaf positions fixed and bremsstrahlung leakage and electron scatter doses included. This optimization gives the final optimized plan. It is shown that a significant portion of the calculation time is spent transporting particles in the leaves. However, this is necessary since optimizing segment weights based on a model in which leaf transport is ignored results in an improperly optimized plan with overdosing of target and critical structures. A method of rapidly calculating the bremsstrahlung contribution is presented and shown to be an efficient solution to this problem. A homogeneous model target and a 2D breast plan are presented. The potential use of this tool in clinical planning is discussed.     

Resolving the composite trait of hypertension into its pharmacogenetic determinants by acute pharmacological modulation of blood pressure regulatory systems

Acute pharmacogenetic analysis was carried out in an intercross F2 population derived from Prague hypertensive-hypertriglyceridemic and Lewis rats. Quantitative trait loci (QTL) mapping was performed for baseline blood pressure (BP) and for BP after blockade of the renin-angiotensin system by losartan, of the sympathetic nervous system (SNS) by pentolinium, and of the nitric oxide system by N(G)-nitro- L-arginine methyl ester. Two significant loci for baseline BP were found on chromosome (Chr) 3 (logarithm of likelihood, LOD, 3.8) and Chr 5 (LOD 3.6), and one suggestive locus on Chr 1 (LOD 2.7). The QTL on Chr 3 persisted after treatment with the three agents while the QTL on Chr 5 and Chr 1 disappeared after pentolinium administration. This suggests independence of the locus on Chr 3 from each acute BP regulatory system examined, whereas the loci on Chr 5 and Chr 1 appeared to be controlled mainly by the SNS. Although not apparent at baseline, a significant locus appeared on Chr 8 (LOD 7.0) after blockade of the SNS, and NO system blockade led to the appearance of a new QTL on Chr 1 (LOD 3.6), indicating the contribution of the inhibited systems to these loci. Pharmacogenetic dissection of the BP trait is a powerful tool to unravel the underlying physiological mechanisms of QTL affecting baseline BP and to identify specific QTL for the response to drugs. This pharmocogenetic approach enabled us to determine the main causative acute BP regulatory systems and should lead to better selection of suitable antihypertensive drugs for individual patients.     

动物实验模型中多极导管心室除颤系统电除颤对心脏功能的影响

评价新型的双极和三极导管自动心室除颤系统电除颤对左心室收缩和舒张功能的影响。动物麻醉后,在X光机指导下,分别在10只犬心脏内装置双极导管自动除颤系统（组Ⅰ）;在10只猪心脏内装置三极导管自动除颤系统（组Ⅱ）;并行电除颤试验。使用食管超声心动图在电除颤前后记录二维、M型和多谱勒超声图像。组I动物接受4次电除颤,电量为64J;组Ⅱ接受平均8次电除颤,电量为210J。结果显示：左室收缩面积分数、左室等容舒张时间和二尖瓣血流E波与A波速度比值以及时间-流速积分比值等反映左室舒缩功能的指标在两组动物除颤后均无显著改变。研究表明：两种经静脉导管自动心室除颤系统中反复低能量心内膜电除颤对左室舒缩功能无明显损伤作用;研究结果为经静脉多极导管自动心室除颤系统在临床的应用和电生理研究提供了可靠的实验数据。     

构建7q32区域鼻咽癌和正常鼻咽上皮细胞部分基因的表达图谱

目的构建７ｑ３２区域鼻咽癌细胞和组织及原代培养人正常鼻咽上皮细胞的部分基因表达图谱。方法通过差异ＲＴ－ＰＣＲ和Ｎｏｒｔｈｅｒｎ杂交的方法检测定位于７ｑ３２区域的２０个ＥＳＴ在鼻咽癌细胞和鼻咽癌组织及原代培养人正常鼻咽上皮细胞ｍＲＮＡ的表达水平。结果８个ＥＳＴ在鼻咽癌细胞ＨＮＥ１和原代培养人正常鼻咽上皮细胞中表达量较一致，７个ＥＳＴ在两种细胞中均无表达，３个ＥＳＴ（Ｗ７２６８８、Ｈ１９８３０、ＡＡ１３０６３０）在鼻咽癌细胞株中表达上调，而２个ＥＳＴ（ＡＡ０７０４３７、Ｈ９０８８２）在原代培养人鼻咽上皮细胞中表达上调。在１３例鼻咽癌活检组织中３０．７％（４／１３）的ＡＡ０７０４３７表达下调，７７％（１０／１３）的Ｗ７２６８８和７７％（１０／１３）的Ｈ１９８３０表达上升。结论构建了７ｑ３２区域鼻咽癌细胞和组织及原代培养人正常鼻咽上皮细胞部分基因表达图谱，并初步认为Ａ０７０４３７的表达下调和Ｗ７２６８８、Ｈ１９８３０的过表达与鼻咽癌的发生有关。     

Age-dependent DNA methylation changes in the ITGAL (CD11a) promoter

DNA methylation patterns change with age in a complex fashion, typically with an overall decrease in genomic deoxymethylcytosine (d(m)C) content, but with local increases in some promoters that contain GC-rich sequences known as CpG islands. While the consequences of age-dependent CpG island methylation have recently been studied in organs such as the colon, less is known about the functional significance of the progressive hypomethylation of promoters lacking CpG islands, and the significance of age-dependent changes in T cell DNA methylation is completely unexplored. We asked if age-dependent DNA hypomethylation might contribute to overexpression of the T cell ITGAL gene, which encodes CD11a, a subunit of LFA-1. CD11a mRNA increased with age as well as with experimentally induced DNA hypomethylation. This increase correlated with hypomethylation of sequences flanking the ITGAL promoter in vitro and in aging. 'Patch' methylation of the region suppressed promoter function. DNA methyltransferases 1 and 3a also decreased with aging. These results indicate that hypomethylation of regions flanking the ITGAL promoter may increase CD11a expression, and suggest that age-dependent hypomethylation of promoters lacking CpG islands, perhaps due to decreased DNA methyltransferase expression, may be one mechanism contributing to increased T cell gene expression with aging.     

琥珀酸美托洛尔HPMC骨架片释放影响因素研究

以羟丙基甲基纤维素（HPMC）为骨架材料，乙基纤维素（EC）为阻滞剂，采用湿颗粒压片法制备琥珀酸美托洛尔亲水凝胶骨架片，考察HPMC用量、HPMC黏度、EC用量、制备方法、压片压力、释放介质及转速对琥珀酸美托洛尔（MS）骨架片体外释药的影响。结果表明，MS骨架片体外释药符合Higuchi方程，药物释放机制是骨架溶蚀和药物扩散的综合效应；HPMC用量与黏度、阻滞剂用量、制备方法、压片压力对释放速率均有显著性影响；释放介质的pH值及转速对释放速率无显著性影响。