CYP17A1基因新突变致17α-羟化酶/17,20-裂解酶部分性缺陷症

目的研究1例17α-羟化酶/17,20-裂解酶部分性联合缺陷症患者CYP17A1基因突变特点,并结合患者的临床表现与基因突变类型初步探讨P450C17酶蛋白的结构与功能的关系。方法收集1例17α-羟化酶/17,20-裂解酶部分性联合缺陷症患者的临床资料及其亲属血标本,提取基因组DNA,设计7对引物扩增CYP17A1基因的8个外显子及外显子与内含子的连接区域,琼脂糖凝胶电泳鉴定PCR产物,产物胶回收后直接做为DNA双链模板测序。DNA双链模板不一致的PCR产物经克隆后测序。测序结果在核苷酸序列数据库进行比较分析。结果患者CYP17A1基因突变检测结果为5994-5995delAT/7541C>T复合杂合子。这两种突变均未见报道。推测5994-5995delAT导致I259H,274X,突变形成的截短蛋白质缺少血红素结合区域,因此是没有功能的;而通过人类P450C17酶计算机模型分析显示7541C>T导致的A398V远离酶的活性中心,推测突变可能使酶的活性减弱,而不是完全地丧失。患者临床表现为有自发不规则月经及轻度高血压、低血钾,结合激素测定结果提示肾上腺和性腺保留部分功能。因而患者的基因型与其临床表型是一致的。结论应进行突变P450C17酶的功能学研究来进一步明确结构改变对功能的影响。     

Hyaluronan in peritumoral stroma and malignant cells associates with breast cancer spreading and predicts survival

Hyaluronan (HA) is an extracellular matrix polysaccharide that promotes cell migration through its cell surface receptors and by effecting changes in the physical environment. HA expression is frequently increased in malignant tumors, whereas its association with the invasive potential and patient outcome in breast cancer has not been reported. The localization and signal intensity of HA was analyzed in 143 paraffin-embedded tumor samples of human breast carcinoma using a biotinylated HA-specific probe. In the immediate peritumoral stroma, HA signal was moderately or strongly increased in 39% and 56% of the cases, respectively. Normal ductal epithelium showed no HA, whereas in 57% of the tumors at least some of the carcinoma cells were HA positive. The intensity of the stromal HA signal and the presence of cell-associated HA were both significantly related to poor differentiation of the tumors, axillary lymph node positivity, and short overall survival of the patients. In Cox's multivariate analysis, both the intensity of stromal HA signal alone and that combined with the HA positivity in tumor cells were independent prognostic factors for overall survival. These results suggest that HA is directly involved in the spreading of breast cancer and may offer a potential target for new therapies.     

视黄酸依赖Fas/RARα融合基因转染HL-60细胞启动的凋亡效应研究

目的：检测视黄酸依赖Fas／RARα表达载体转染HL-60细胞启动的凋亡效应。方法：成功构建视黄酸依赖Fas／RARα表达载体，用脂质体转染HL-60白血病细胞，经四甲基偶氮唑盐[3-(4，5-dimethylthiaol-2-yl)-2，5-diphenyl tetrazolium bromide,MTT]、流式细胞、DNA梯形带和透射电镜等方法观测视黄酸处理后细胞凋亡效应的发生。结果：以一定剂量的视黄酸处理HL-60后，细胞增殖受抑，呈凋亡性变。结论：视黄酸及其依赖的融合基因表达可协同诱导HL-60细胞发生凋亡效应。     

自然流产模型小鼠蜕膜细胞凋亡及Bcl-2、Bax表达的研究

目的：通过比较正常妊娠模型小鼠及自然流产模型小鼠蜕膜细胞凋亡及Bcl-2、Bax蛋白的表达，从细胞及分子水平探讨自然流产的发病机制。方法：建立正常妊娠模型CBAXBALB／c和自然流产模型CBAXDBA／2。用免疫组化SABC法测定两组模型孕13天蜕膜细胞Bcl-2和Bax蛋白的表达，并通过MIAS-2000医用彩色病理图像免疫组化测量系统对其表达进行半定量分析，其结果用平均灰度值表示；同时应用DNA缺口原位末端标记技术（TUNEL）测定两组模型孕13天蜕膜细胞凋亡情况。结果：与正常妊娠模型相比，自然流产模型蜕膜细胞Bcl-2蛋白的表达降低（P〈0．01）；Bax蛋白的表达明显升高（P〈0．01）。蜕膜细胞凋亡指数，自然流产模型明显高于正常妊娠模型（P〈0．01）。结论：早孕期蜕膜组织细胞凋亡异常是自然流产的机制之一，Bcl-2／Bax途径可能是诱导早孕期蜕膜细胞凋亡的重要因素。     

CD3+木瓜提取物对ApoE基因缺陷小鼠血脂及炎性相关因素的影响

目的观察木瓜提取物对ApoE基因缺陷小鼠血脂水平、主动脉窦斑块中巨噬细胞活化及肿瘤坏死因子(TNF-α)水平的影响。方法30只雄性ApoE基因缺陷小鼠随机分为3组:对照组、低剂量组和高剂量组,低剂量组和高剂量组分别在基础饲料上添加5%和10%的木瓜提取物喷雾干燥颗粒,饲养16周后经小鼠眼眶静脉取血后处死动物,酶法测定血脂水平、采用免疫组化分析主动脉窦斑块中TNF-α的表达水平和采用免疫荧光反映斑块CD68阳性表达水平与巨噬细胞阳性表达率。结果低剂量组和高剂量组的血清TC和LDL-C显著低于对照组(P<0.05),木瓜提取物能显著下调斑块中TNF-α的表达水平以及减少巨噬细胞的阳性率。结论木瓜提取物降低了血清脂蛋白胆固醇水平(TC和LDL-C),减少了炎性因子TNF-α表达以及减弱巨噬细胞参与程度,预示其可能具有减缓动脉斑块发生发展的抗动脉粥样硬化作用。     

含氮二膦酸盐/IL-2诱导肿瘤患者外周血γδT细胞增殖的作用

目的探讨药物（含氮二膦酸盐联合IL-2，N-BP／IL-2）诱导实体肿瘤患者外周血γδT细胞增殖作用。方法采用流式细胞仪测定103例肿瘤患者和43例健康人外周血总T细胞绝对值、御细胞绝对值和相对值，测定患者外周血经药物体外刺激后御细胞增殖指数和用药前后患者外周血总T细胞、γδT细胞绝对值和相对值。结果肿瘤患者和健康人外周血总T细胞绝对值、御细胞绝对值和御细胞相对值的中位数，分别为1306细胞/μl和1522细胞/μlP=0．003），64细胞/μl和162细胞/μl（P〈0．001），5％和11％（P〈0．001）。24例经药物体外扩增试验阳性的肿瘤患者治疗前后体内外周血总T细胞绝对值、γδT细胞绝对值和御细胞相对值的中位数，分别为1283细胞/μl和1552细胞/μl（P=0．001），54细胞/μl和107细胞/μl（P〈0．001），6％和8％（P=0．008）。全部肿瘤患者中外周血γδT细胞对药物体外诱导增殖反应阳性患者的百分率为68．9％。结论肿瘤患者外周血γδT细胞计数显著低于健康人。不同肿瘤患者外周血γδT细胞对药物刺激增殖的反应性不同。N-PB／IL-2在体内有显著增加御细胞数量的药理作用。体外增殖试验可作为N-PB／IL-2治疗患者的参考。     

Analysis of the floral transcriptome uncovers new regulators of organ determination and gene families related to flower organ differentiation in Gerbera hybrida (Asteraceae)

Development of composite inflorescences in the plant family Asteraceae has features that cannot be studied in the traditional model plants for flower development. In Gerbera hybrida, inflorescences are composed of morphologically different types of flowers tightly packed into a flower head (capitulum). Individual floral organs such as pappus bristles (sepals) are developmentally specialized, stamens are aborted in marginal flowers, petals and anthers are fused structures, and ovaries are located inferior to other floral organs. These specific features have made gerbera a rewarding target of comparative studies. Here we report the analysis of a gerbera EST database containing 16,994 cDNA sequences. Comparison of the sequences with all plant peptide sequences revealed 1656 unique sequences for gerbera not identified elsewhere within the plant kingdom. Based on the EST database, we constructed a cDNA microarray containing 9000 probes and have utilized it in identification of flower-specific genes and abundantly expressed marker genes for flower scape, pappus, stamen, and petal development. Our analysis revealed several regulatory genes with putative functions in flower-organ development. We were also able to associate a number of abundantly and specifically expressed genes with flower-organ differentiation. Gerbera is an outcrossing species, for which genetic approaches to gene discovery are not readily amenable. However, reverse genetics with the help of gene transfer has been very informative. We demonstrate here the usability of the gerbera microarray as a reliable new tool for identifying novel genes related to specific biological questions and for large-scale gene expression analysis.     

细胞内外膜跨膜电位频率响应模型及滤波器特性的研究

提出球形单细胞内外膜跨膜电位频率响应模型和仿真计算方法。通过对频率响应的仿真分析，发现内外膜跨膜电位分别表现出一阶带通和低通滤波器特性，并在内膜频率响应中心频率到上限截止频率的范围内，内膜跨膜电位大于外膜同时能保持在较高水平，这将有利于诱导细胞内电处理效应。计算结果与动物实验吻合得很好，为纳秒脉冲电场治疗肿瘤的窗口参数合理选择提供了理论依据。