SRY基因诊断在临床中的应用

应用聚合酶链式反应（ＰＣＲ）扩增技术，对１０例染色体核型为４６，ＸＹ和１例为４６，ＸＸ／４６，ＸＹ的性反转、睾丸女性化、两性畸形及男性生殖器发育不良的患者进行了性别决定区（ＳＲＹ）基因检测。结果表明９例４６，ＸＹ和１例４６，ＸＸ／４６，ＸＹ患者的ＳＲＹ基因存在，１例４６，ＸＹ女性患者的ＳＲＹ基因缺失。该结果表明能够用分子生物学技术对性反转、性发育异常的病因进行分析     

An Evolutionarily Conserved Region in the Second lntron of the Human Nestin Gene Directs Gene Exmession to CNS Progenitor Cells and to Early Neural Ciest Cells

Central nervous system (CNS) progenitor cells transiently proliferate in the embryonic neural tube and give rise to neurons and glial cells. A characteristic feature of the CNS progenitor cells is expression of the intermediate filament nestin and it was previously shown that the rat nestin second intron functions as an enhancer, directing gene expression to CNS progenitor cells. In this report we characterize the nestin enhancer in further detail. Cloning and sequence analysis of the rat and human nestin second introns revealed local domains of high sequence similarity in the 3' portion of the introns. Transgenic mice were generated with the most conserved 714 bp in the 3' portion of the intron, or with the complete, 1852 bp, human second intron, coupled to the reporter gene lacZ. The two constructs gave a very similar nestin-like expression pattern, indicating that the important control elements reside in the 714 bp element. Expression was observed starting in embryonic day (E)7.5 neural plate, and at E10.5 CNS progenitor cells throughout the neural tube expressed lacZ. At E12.5, lacZ expression was more restricted and confined to proliferating regions in the neural tube. An interesting difference, compared to the rat nestin second intron, was that the human intron at E10.5 mediated lacZ expression also in early migrating neural crest cells, which is a site of endogenous nestin expression. In conclusion, these data show that a relatively short, evolutionarily conserved region is sufficient to control gene expression in CNS progenitor cells, but that the same region differs between rodents and primates in its capacity to control expression in neural crest cells.     

环氧化酶-2基因启动子甲基化及表达与胃黏膜病变的关系

目的 探讨环氧化酶2(COX-2)基因启动子区甲基化水平和蛋白表达与胃黏膜病变的关系,并对其相关的影响因素进行研究.方法 以1201例患有不同胃黏膜病变的高危人群为研究对象,采用免疫组织化学方法榆测COX-2表达,用亚硫酸氢钠-变性高效液相色谱(DHPLC)对COX-2启动子甲基化率进行定量分析,采用13C尿素呼气实验(13C-UBT)对幽门螺旋杆菌(H priori)感染状况进行测定.结果 COX-2甲基化率中位数随胃黏膜病变的加重逐渐升高,在浅表性胃炎和慢性萎缩性胃炎(SG/CAG)、肠上皮化生(IM)及不确定性异型增生和异璎增生(Ind DYS/DYS)病变中分别为10.6%、11.8%、13.8%,各病变组之间差异有统计学意义(X2=8.312,P=0.016).分层分析显示,在H pylori感染阴性病例中,COX-2甲基化率仍随病变加重明显升高,在SG/CAG、IM、Ind DYS/DYS病变中其中位数分别为8.8%、10.6%、14.1%(X2=6.629,P=0.036).进一步分析发现,COX-2甲基化率随着COX-2表达强度的增强而降低,由COX-2弱阳性表达的13.3%降至强阳性表达的7.6%(X2=10.400,P=0.015).结论 COX-2启动子甲基化水平与胃黏膜病变程度及H pylori感染状况密切相关,并与COX-2表达强度呈负相关,说明COX-2启动子区异常甲基化可能在胃黏膜病变的演变过程中起重要作用.     

精氨酸加压素（AVP）的衍生物AVP4—8不能作用于AVPV1a受体而诱发电流反应

在微量注射大量肝脏ｍＲＮＡ之后，通过电压箝方法进行功能鉴定，两栖类卵母细胞成功地表达了ＡＶＰＶ１ａ受体。但在灌流ＡＶ４－８溶液时，却不能诱导卵母细胞产生内向振荡电流反应。提示ＡＶＰ４－８不能通过ＡＶＰＶ１ａ受体而介导生理学效应。     

赖型钩体flaB2与VR1012中的CpG基序分析

目的：对问号赖型钩端螺旋体（赖型钩体）DNA疫苗[包括内鞭毛蛋白基因（flaB2)和质粒DNA表达载体（VR1012）]的CpG基序（CpG motifs)进行分析，为DNA疫苗免疫机制的阐明和提高DNA疫苗的效能奠定基础。方法：以flaB2与VR1012构建重组DNA的免疫原，对flaB2及VR1012全核苷酸序列进行计算机分析（分类、计数和定位）。结果：CpG的“C”的侧翼为两个嘌呤，“G”的侧翼为两个嘧啶，在flaB2中共3个，分别为GACGCT，GACGTC和GACGCC；在VR1012中共11个，分别为GACGTC1个，GACGCT2个，GACGCC1个，GACGTT1个，GGCGTT2个，GGCGCT2个，GGCGCC1个，AACGCT1个，其中特别重要的TGACGTCA4个和TAACGCCA有1个，位于5'端456－463；509－516；592－599；778－785和486－493；4个TGACGTCA和1个TAACGCCA均位于5'端且相对集中。结论：赖型钩体flaB2与VR1012构成的DNA疫苗含有TGACGTCA等CpG，这些基序又称免疫刺激序列，构成了DNA疫苗中的佐剂。     

TSG-6基因在3T3-L1脂肪细胞诱导分化中表达水平的变化

[目的] 探讨TSG-6基因在3T3-L1脂肪细胞诱导分化中表达水平的变化。[方法] 采用细胞培养和RT-PCR技术，检测细胞诱导分化不同时段脂肪细胞中TSG-6基因的表达水平。[结果] ①随着脂肪细胞逐渐分化成熟，TSG-6基因mRNA表达水平逐渐升高；②TSG-6基因表达水平除在细胞分化第0-2d、第3-5d和第7-10d各时段内差异无显著性(P＞0.05)外，其余各时段之间表达水平差异均有显著性(P＜0.05)。[结论] TSG-6基因与细胞分化以及脂原形成可能相关。     

泪腺肿瘤中P53蛋白表达的意义

目的了解P53基因在泪腺肿瘤的表达及其临床病理学意义。方法应用免疫组织化学ABC法检测57例泪腺上皮性肿瘤患者和10例正常人泪腺组织中P53蛋白的表达情况，并与病理分级、复发相联系。结果10例正常人泪腺组织中P53表达全部为阴性；恶性肿瘤患者的表达率为48．6％，良性肿瘤中P53达率为18％，二者相比有显著性差异（P＜0．025）；复发患者阳性率为76．9％，明显高于未复发患为31．8％（P＝0．015）。结论P53基因参与了泪腺肿瘤的发生发展并与肿瘤的分化和复发有关。     

心血管疾病基因治疗的基础研究Ⅵ．一种新的体内基因转移方法

将医用缝合线浸泡在质粒ＤＮＡ溶液中，然后缝合在大鼠股四头肌上，可以观察到外源基因在肌肉组织中高效、稳定地表达，其表达量高于磷酸钙共沉淀和ＤＮＡ沉淀块埋植法，较肌肉直接注射法表达量高１０倍左右。     

Sensitive Detection of p53 Gene Mutations in Esophageal Endoscopic Biopsy Specimens by Cell Sorting Combined with Polymerase Chain Reaction Single-strand Conformation Polymorphism Analysis

For the rapid and sensitive detection of p53 gene mutations in esophageal endoscopic biopsy specimens, we combined cell sorting with the polymerase chain reaction and single-strand conformation polymorphism (PCR-SSCP) analysis. Mutations in exons 5–8 of the p53 gene were investigated by FCR-SSCF analysis using 10³ sorted nuclei obtained from each endoscopic biopsy specimen of 16 patients with esophageal cancer. DNAs extracted from their respective surgical specimens were investigated by a conventional method of PCR-SSCP analysis. Mutations in the biopsy specimens were detected in 6 of the 12 aneuploid tumors but in none of the 4 diploid tumors. After tumor cell enrichment by cell sorting, one mutation in exon 8 became apparent, which could not be detected from the surgical specimen by a conventional method of PCR-SSCP analysis. This method should improve the sensitivity of detecting p53 gene mutations, and provides additional information concerning the DNA ploidy pattern in the tumors.