肿瘤坏死因子凋亡相关配体基因的克隆和表达

目的：克隆、表达和鉴定肿瘤坏死因子凋亡相关配体(TRAIL)．方法：从培养的人外周血淋巴细胞中提取总RNA，经RT-PCR获得TRAIL基因．将该基因克隆到pGEM-Teasy载体中，测序鉴定．将TRAIL基因插入pBV220表达载体中，42℃诱导表达4～5h，进行SDS-PAGE分析．免疫印迹法鉴定TRAIL蛋白的表达．结果：DNA测序证明，获得了TRAIL基因，其序列与GenBank中报道序列完全一致．SDSPAGE分析表明，TRAIL蛋白获得高效表达，分子质量为17ku，表达量约占菌体总蛋白的300k，．免疫印迹法鉴定显示，该蛋白可与鼠抗人TRAIL mAb产生阳性反应．结论：成功克隆和表达了TRAIL基因．     

Detection of disseminated tumor cells in the lymph nodes of colorectal cancer patients using a real-time polymerase chain reaction assay

Introduction Postoperative treatment for colorectal cancer depends on tumor stage as defined by the International Union Against Cancer (UICC). Adjuvant chemotherapy is not recommended in patients without lymph node involvement (UICC stages I and II). As many as 20–30% of these patients, however, will develop recurrence. Aims and objectives We conducted this study to determine the presence of disseminated tumor cells in the lymph nodes by quantitative real-time polymerase chain reaction (QRT-PCR) for cytokeratin 20 (CK20) in an attempt to provide supplementary information compared to histopathological findings. Materials and methods Using a standard QRT-PCR assay, we examined primary tumors and 391 lymph nodes from 31 patients with completely resected colorectal cancer. Results Of the 31 primary tumors, 29 were positive for CK20 by QRT-PCR. Discussion An examination of the lymph nodes from the 29 patients with CK20-positive primary tumors revealed that 35 (92.1% sensitivity) of the 38 histopathologically positive lymph nodes and 54 (16.7%) of the 324 histopathologically negative lymph nodes were positive by molecular analysis. CK20 expression was detected in 10 (100%) of 10 patients with a histopathologically positive lymph node status (pN1). In 9 (47.4%) of 19 patients with negative histopathological results (pN0), we detected a CK20 mRNA signal in at least one lymph node. Whereas eight patients with histopathologically negative lymph nodes could be upstaged on the basis of the molecular findings, no patient would be downstaged. Conclusion Our results suggest that QRT-PCR for CK20 is a useful tool for the quantitative detection of micrometastases in the regional lymph nodes. We introduce a standardized procedure that integrates a molecular diagnostic technique in the clinical staging.     

心脏营养素-1治疗失神经骨骼肌萎缩过程中对正常肌肉组织的影响

目的 探讨应用心脏营养素-1(cardiotrophin-1,CT-1)治疗失神经骨骼肌萎缩的有效剂量和安全剂量,观察不同剂量的CT-1对正常骨骼肌、心肌和血管平滑肌的副作用.方法 取Swiss小鼠80只,随机分成8组,每组分别于胫神经切断术后连续腹腔注射重组小鼠CT-1(rmCT-1)40、60、80、100、120、140、160μg·kg-1·d-1,剩余1组腹腔注射等体积CT-1溶媒,28d后称量失神经侧腓肠肌、正常侧腓肠肌和心脏的湿重;测量胸主动脉平滑肌厚度;检测正常腓肠肌中α-actin和MHC Ⅱa以及心肌中β-MHC的表达、胸主动脉血管平滑肌细胞中α-actin的含量.结果 采用腹腔注射给药,当CT-1用量达到60μg·kg-1·d-1时,开始对失神经骨骼肌产生营养作用,并呈剂量依赖性增强,当给药剂量超过100 μg·kg-1·d-1后,出现对正常骨骼肌、心肌和血管平滑肌的影响.给药剂量达到120μg·kg-1·d-1时,开始促进心肌β-MHCmRNA的表达,增加正常心脏的湿重.当给药剂量达到140μg·kg-1·d-1时,开始促进正常骨骼肌α-actin和MHC Ⅱ amRNA的表达,以及骨骼肌湿重的增加,同时提高了主动脉血管平滑肌α-actin的含量,增加了血管平滑肌的厚度.结论 腹腔注射CT-1防治失神经骨骼肌萎缩的有效剂量为60 μg·kg-1·d-1,最高安全剂量为100 μg·kg-1·d-1.当给药剂量达到120 μg·kg-1·d-1时,开始出现心肌肥厚,当给药剂量达到140 μg·kg-1·d-1时,可导致正常骨骼肌的肥大和血管平滑肌的增厚.     

Review of long-term adverse effects associated with the use of chemically-modified animal and nonanimal source hyaluronic acid dermal fillers

Although only recently introduced, chemically-modified hyaluronic acid dermal fillers have gained widespread acceptance as “redefining” dermal fillers in the fields of dermatology and cosmetic facial surgery. Although hyaluronic acid-based dermal fillers have a low overall incidence of long term side effects, occasional adverse outcomes, ranging from chronic lymphoplasmacytic inflammatory reactions to classic foreign body-type granulomatous reactions have been documented. These long-term adverse events are reviewed.     

+10 Gz重复暴露大鼠脑差异表达基因cDNA文库的构建

目的构建＋10Gz重复暴露大鼠脑差异表达基因的消减cDNA文库。方法本实验用SD大鼠，分别提取暴露组与对照组的总RNA，并分离纯化mRNA，应用抑制性消减杂交技术分离＋10GI重复暴露大鼠脑差异表达基因eDNA片段并建立消减eDNA文库；利用PCR对随机挑选的75个白色菌落进行插入片段的验证，对其中70个克隆进行eDNA斑点杂交验证。结果所构建的eDNA文库扩增后包含约400个白色克隆和100个兰色克隆，随机挑选75个白色克隆入质粒载体后共获得70个阳性克隆。结论应用抑制性消减杂交技术成功构建了＋10Gz重复暴露大鼠脑差异表达基因消减eDNA文库，为进一步筛选和克隆脑损伤相关基因奠定了基础。     

伊曲康唑不良反应的国内文献综述

目的：了解伊曲康唑的不良反应及安全性。资料与方法：手工检索《中华皮肤科杂志》等3种中文专业期刊，从对照试验的文献中提取治疗例数和各种不良反应例数，计算发生率。结果：共有14725例涉及有关安全性的报道。伊曲康唑不良反应总的发生率约为1．57％。结论：伊曲康唑较为安全。     

Two novel gastric cancer-associated genes identified by differential display

AIM To clone novel gastric cancer-associated genes and investigate their roles in gastric cancer occurrence.METHODS A method called differential display was used which allows the identification of differentially expressed genes by using PAGE to display PCR-amplified cDNA fragments between gastric cancer cells and normal gastric mucosa cells. These fragments were cloned into plasmid vector pUC18. Homology analysis was made after sequencing these fragments.RESULTS Two novel genes were identified compared with sequences from GenBank. One was registered with the AD number AF 051783. In situ hybridization showed that these two novel genes expressed specifically in gastric cancer tissues.CONCLUSION The two novel genes obtained by differential display were confirmed to be gastric cancer-associated genes using in situ hybridization.     

急性白血病非系限性分化抗原的动态研究

为探讨仅表达非系限性分化抗原的急性白血病的细胞起源，采用单克隆抗体免疫酶标和聚合酶链反应技术分析了１２例初诊时仅表达非系限性分化抗原的急性白血病化疗后免疫表型及免疫球蛋白重链（ＩｇＨ）和Ｔ细胞受体（ＴＣＲ）γ基因重排的变化。结果表明：初诊时仅表达ＣＤ３８抗原的５例急性白血病，化疗后３例出现Ｔ细胞相关抗原表达，并伴ＴＣＲγ基因重排；５例初诊时仅表达ＨＬＡ－ＤＲ或ＣＤ９的急性白血病，化疗后４例出现Ｂ细胞相关抗原表达，均伴有ＩｇＨ基因重排；２例无任何抗原表达者，化疗后１例表达Ｂ细胞相关抗原，另１例表达骨髓细胞相关抗原。提示免疫标志的动态研究，有助于初诊时免疫学无法分类急性白血病细胞起源的确定。     

荧光定量聚合酶链反应在尖锐湿疣诊断中的应用

目的　探讨荧光定量聚合酶链反应 (FQ - PCR)对尖锐湿疣 (CA )的诊断价值。方法　 FQ- PCR检测病理确诊的 CA患者 36份标本和健康人 84份标本 ;FQ- PCR及病理诊断同期检测临床送检病例 2 73份标本 ,并做诊断性试验评价。结果　 36例病理确诊的 CA患者 HPV6和 HPV11的 DNA FQ- PCR全部阳性 ,平均拷贝数为 1.0× 10 7± 1.0× 10 2 / m l;84例健康人 FQ- PCR全部阴性 ,平均拷贝数为 3× 10± 2× 10 / ml。 FQ- PCR与病理诊断符合率为 10 0 % ,CA患者与健康人拷贝数的差异有显著性 (P<0 .0 5 )。FQ- PCR与病理诊断对比得出 :灵敏度为 10 0 % ,特异度为 92 % ,误诊率为 0 .0 8,漏诊率为 0 ,准确度为 98.9% ,阳性预测值为 98.8% ,阴性预测值为 10 0 % ,阳性似然比为 12 .5 ,阴性似然比为 0。结论　 FQ - PCR能准确定量 ,灵敏度高 ,特异性强 ,快速 ,简便 ,可作为尖锐湿疣早期诊断的指标