Fluorotyping of HLA-C: differential detection of amplicons by sequence-specific priming and fluorogenic probing

Conventional PCR-SSP, which is based on an agarose gel-based read-out, has the disadvantages of time-consuming post-PCR steps and low potential for automation. The aim of our study was to sort out these drawbacks by establishing a fluorescence-based PCR-SSP system for HLA-C. The assay relies on the sequence-specific identification of amplicons with individually labeled probes that are cleaved during successful PCR by the 5'-3' exonuclease activity of the Taq-DNA Polymerase. The oligonucleotides are labeled with a unique and spectrally resolvable fluorescent reporter dye at the 5' terminus (FAM or TET) and a common quencher dye at the 3' terminus (TAMRA). In case of amplification, the reporter escapes from the quenching control caused by the physical separation of the dyes, resulting in a significant increase of the reporter fluorescence. This allows simultaneous and differential detection of the specific HLA (FAM) and internal control (TET) product. The HLA-C fluorotyping information is based on the individual reporter fluorescence released by 18 PCR primer mixes. Using this method, we analyzed 145 samples previously typed with conventional PCR-SSP and found a concordance rate of 100%. Furthermore, fluorotyping revealed quantitative results that may indicate the presence of homozygosity by high signal intensities. This provided extra protection not to miss new alleles which are not amplified by the current primer mixes. These features as well as the capability of high sample throughput and the possibility of automation makes fluorotyping an attractive tool for PCR-based HLA typing.     

口服Sumatriptan治疗偏头痛的临床评价

本文报告口服Sumatriptan 100mg对偏头痛急性发作119例次的治疗结果。治疗后4h内显效91例次(76.5％),好转16例次(13.4％),无效12例次(10.1％),总有效率为89.9％。对偏头痛伴随症状恶心、呕吐和畏光、畏声的缓解率分别为94.2％、96％和94.3％。     

Initial evaluation of123|-5-|-R91150, a selective 5-HT2A ligand for single-photon emission tomography, in healthy human subjects

The mapping of 5-HT₂ receptors in the brain using functional imaging techniques has been limited by a relative lack of selective radioligands. Iodine-123 labelled 4-amino-N-[1-[3-(4-fluorophenoxy)propyl]-4-methyl-4-piperidinyl]-5-iodo-2-methoxybenzamide (¹²³I-5-I-R91150 or¹²³I-R93274) is a new ligand for single-photon emission tomography (SPET), with high affinity and selectivity for 5-HT_2A receptors. This study reports on preliminary¹²³I-5-I-R91150 SPET, wholebody and blood distribution findings in five healthy human volunteers. Maximal brain uptake was approximately 2% of total body counts at 180 min post injection (p.i.). Dynamic SPET sequences were acquired with the brain-dedicated, single-slice multi-detector system SEM-810 over 200 min p.i. Early peak uptake (at 5 min p.i.) was seen in the cerebellum, a region free from 5HT_2A receptors. In contrast, radioligand binding in the frontal cortex increased steadily over time, up to a peak at approximately 100–120 min p.i. Frontal cortex-cerebellum activity ratios reached values of 1.4, and remained stable from approximately 100 min p.i. onwards. Multi-slice SPET sequences showed a pattern of regional variation of binding compatible with the autoradiographic data on the distribution of 5-HT_2A receptors in (cerebral cortex>striatum>cerebellum). These findings suggest that¹²³I-5-I-R91150 may be used for the imaging of 5-HT_2A receptors in the living human brain with SPET.     

组织胺诱导胸腺细胞凋亡的研究

用苔盼蓝及吖啶橙染色，ＤＮＡ片段检定与ＤＮＡ电泳等方法进行组织胺诱导胸腺细胞凋亡的研究，旨在探讨其可能机制及其在胸腺细胞负选择中的作用。结果表明，组织胺诱导使体外培养的胸腺细胞出现典型的细胞调亡。雷尼替丁可阻断上述作用，提示胸腺细胞存在Ｈ＿２受体，活化该受体可能在胸腺细胞的负选择中起重要作用。     

Altered enzyme activities of xenobiotic biotransformation in kidneys after subchronic administration of 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX) to rats

Activities of the xenobiotic metabolizing enzymes were measured in the liver, kidney, duodenum and lung microsomes and cytosol fractions of Wistar rats after subchronic administration of 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), a potent bacterial mutagen in chlorinated drinking water. MX was administered by gavage at the dose level of 30 mg/kg for 18 weeks (low dose), or at the dose level which was raised gradually from 45 mg/kg for 7 weeks via 60 mg/kg for 2 weeks to a clearly toxic dose of 75 mg/kg for 5 weeks (high dose). Microsomal and cytosolic preparations were made and the activities of 7-ethoxyresorufin-O-deethylase (EROD), pentoxyresorufin-O-dealkylase (PROD), NADPH-cytochrome-c-reductase, UDP-glucuronosyltransferase (UDPGT) and glutathione-S-transferase (GST) were measured. Kidneys were affected most. A dose-dependent decrease was observed in EROD (90% in males, 80% in females at the high dose) and in PROD (58% in females, at the high dose) in kidneys. An increase was, however, detected in kidney NADPH-cytochrome-c-reductase (66% in females at high dose), UDPGT (89% in males and 97% in females at high dose) and GST activities (56% in males and 50% in females at high dose). MX caused only a few changes in the enzyme activities of the liver. The EROD activity was decreased 25% to 37%, both in the livers of males and females, but the total content of P450s was not altered. Hepatic GST activity was elevated in females in a dose-dependent manner (31% and 44%). GST activity was elevated in duodenum in females (59%) at the high dose. There were no marked changes in the enzyme activities in the lungs. MX was a weak inhibitor of EROD activity both in the liver and kidney microsomes in vitro, decreasing the EROD activity by 53% and 43%, respectively at the concentration of 0.9 mM. The results indicate that MX decreases the activity of phase I metabolism enzymes, but induces phase II conjugation enzyme activities, particularly in kidneys in vivo. It is possible that these changes contribute to metabolism of MX in kidneys and renders them susceptible to MX in the course of repeated exposure.     

气功对冠心病人CM_5ST段/心率斜率的影响

37名冠心病患者随机分为气功治疗组(22人)和对照组(15人)。气功治疗组接受12周动功治疗,对照组接受安慰剂治疗,治疗前后均测定CM_5 ST段/心率斜率。结果发现动功治疗组CM_5 ST段/心率斜率减少,而对照组不变或增大,两组有显著性差异,ST段下移及常规分级运动试验其它指标在两组间无显著差异。表明CM_5ST段/心率斜率是一个敏感的心肌缺血指标。     

小檗碱对大鼠阴茎海绵体磷酸二酯酶5mRNA水平的影响

目的 :检测大鼠阴茎海绵体中磷酸二酯酶 5 (PDE5 )mRNA的表达 ,进而探讨小檗碱 (berberine ,Ber)的分子作用机制。　方法 :通过逆转录酶链反应 (RT PCR)技术检测大鼠阴茎海绵体中PDE5mRNA的表达。　结果 :大鼠阴茎海绵体中有PDE5A1和PDE5A2mRNA的表达 ,以PDE5A2为主要的异构酶。与内参照 β 肌动蛋白相比 ,对照组、Ber孵育 1和 3h组的PDE5A1及PDE5A2基因mRNA的相对表达量分别为 :0 .2 2± 0 .0 2 ,0 .4 1± 0 .0 1 ;0 .1 5± 0 .0 1 ,0 .34± 0 .0 2 ;0 .1 0± 0 .0 1 ,0 .1 2± 0 .0 1。与对照组相比 ,PDE5A1、PDE5A2的mRNA表达 ,在Ber孵育 1h组降低了 32 %和 1 7%,3h组降低了 5 5 %和 71 %,其中尤以应用Ber 3h后PDE5A2的mRNA减少最为明显 (n =5 ,P <0 .0 1 )。　结论 :Ber对NO cGMP信号通路的下游关键酶 (PDE5 )具有一定的调控作用 ,尤其是抑制PDE5A2的mRNA表达 ,为Ber治疗勃起功能障碍的分子机制之一。     

质子磁共振波谱在胶质瘤诊断中的应用

目的探讨质子磁共振波谱（^1Hmagnetic resonance spectroscopy，^1HMRS）在胶质瘤诊断及分级中的应用。方法回顾分析我院2003年3月～2005年9月质子磁共振波谱检查并经病理证实的36例胶质瘤的临床资料，根据胶质瘤恶性程度不同分为A、B2组，其中A组为低恶性组（包括星形细胞瘤Ⅰ级和Ⅱ级、室管膜瘤、少突胶质细胞瘤等）21例，B组为高恶性组（包括星形细胞瘤Ⅲ级和Ⅳ级、多形性胶质母细胞瘤、间变型室管膜瘤、髓母细胞瘤）15例。观察氮乙酰门冬氨酸（Nacetylaspartate，NAA）、胆碱（choline，Cho）、肌酸（creatine，Cr）、肌醇（Ins）的共振峰及比值。大脑对侧对应正常脑组织的结果作内对照组。结果2组胶质瘤均表现为异常的。HMRS，与大脑对侧对应正常脑组织相比，A、B2组表现为NAA／Cr含量显著下降，Cho／Cr含量显著增高（P均〈0．05），cr总量变化不大，且无规律性（P〉0．05），Ins含量表现为轻度下降（P〉0．05）。A组NAA／Cr和NAA／Cho均显著高于B组（t=4．235，P=0．011；t=2．832，P=0．031），A组Cho／Cr显著低于B组（t=-2．323，P=0．042）。结论。HMRs可提高对胶质瘤诊断和分级的准确性。