• 【对沙眼衣原体膜成分的体液免疫应答和体外受精患者卵泡液中人60 kDa热休克蛋白的表达。】 复制标题 收藏 收藏
    DOI:10.1093/humrep/12.5.925 复制DOI
    作者列表:Neuer A,Lam KN,Tiller FW,Kiesel L,Witkin SS
    BACKGROUND & AIMS: Recent evidence suggests that Chlamydia trachomatis can persist in the female upper genital tract in an unculturable state. Since unsuspected C. trachomatis infection has been associated with adverse in-vitro fertilization (IVF) outcome we sought to detect further evidence of C. trachomatis in the genital tracts of women undergoing IVF. The prevalence and distribution of antibodies to the major structural proteins of C. trachomatis in paired follicular fluid and sera of women undergoing IVF were examined. Sera and follicular fluid samples from 149 women were assayed for immunoglobulin (Ig)G and IgA antibodies to two C. trachomatis antigens, the major outer membrane protein (MOMP) and a recombinant lipopolysaccharide (rLPS) fragment. Additionally, the expression of human 60 kDa heat shock protein (hsp 60) in follicular fluid was determined. All cervical and follicular fluid samples were negative for C. trachomatis by polymerase chain reaction, ligase chain reaction and DNA probe. Sera from 60% of the subjects were positive for antichlamydial rLPS IgG; 36% were positive for anti-MOMP IgG. Similarly, rLPS-directed and MOMP-directed IgA were detected in sera of 34 and 14% of the subjects respectively. IgG antibodies to MOMP and rLPS were detected in 42 and 41% of the follicular fluid examined respectively. Anti-MOMP IgA was identified in 8.7% of the follicular fluid while 27.5% were positive for anti-rLPS IgA. Human hsp 60 expression was documented in 11.6% of the follicular fluid tested. IgA antibodies to both MOMP (P = 0.03) and rLPS (P = 0.02) in follicular fluid were associated with a failure to become pregnant after embryo transfer. IgG antibodies in sera and follicular fluid and IgA antibodies in sera were unrelated to IVF outcome. Similarly only anti-MOMP IgA (P = 0.02) and anti-rLPS IgA (P = 0.04) in follicular fluid were correlated with human hsp 60 expression in follicular fluid. The unique association between IgA antibodies to two chlamydial antigens in follicular fluid and both hsp 60 expression and IVF failure provides further support for the possibility that a persistent upper genital tract chlamydial infection contributes to IVF failure in some women.

    背景与目标: 最近的证据表明,沙眼衣原体可以在女性上生殖道中以无法培养的状态持续存在。由于未曾怀疑的沙眼衣原体感染与体外受精(IVF)不良结果有关,因此我们寻求在接受IVF的女性生殖道中发现沙眼衣原体的进一步证据。检查了在进行IVF的女性的配对卵泡液和血清中沙眼衣原体主要结构蛋白抗体的流行情况和分布。分析了来自149名妇女的血清和卵泡液样品中针对两种沙眼衣原体抗原,主要外膜蛋白(MOMP)和重组脂多糖(rLPS)片段的免疫球蛋白(Ig)G和IgA抗体。另外,测定了人60kDa热休克蛋白(hsp 60)在卵泡液中的表达。通过聚合酶链反应,连接酶链反应和DNA探针,所有子宫颈和卵泡液样品均为沙眼衣原体阴性。 60%的受试者血清抗衣原体rLPS IgG阳性; 36%的抗MOMP IgG阳性。同样,分别在34%和14%的受试者血清中检测到了rLPS导向和MOMP导向的IgA。在分别检查的卵泡液中有42%和41%检测到针对MOMP和rLPS的IgG抗体。在8.7%的卵泡液中发现了抗MOMP IgA,而抗rLPS IgA的阳性率为27.5%。在测试的卵泡液中有11.6%记录了人类hsp 60的表达。卵泡液中针对MOMP(P = 0.03)和rLPS(P = 0.02)的IgA抗体与胚胎移植后无法怀孕有关。血清和卵泡液中的IgG抗体以及血清中的IgA抗体与IVF结果无关。同样,卵泡液中只有抗MOMP IgA(P = 0.02)和抗rLPS IgA(P = 0.04)与人hsp 60在卵泡液中的表达相关。卵泡液中针对两种衣原体抗原的IgA抗体与hsp 60表达和IVF衰竭之间的独特联系为某些女性持续性上生殖道衣原体感染导致IVF衰竭的可能性提供了进一步的支持。

  • 【GABA在胎儿,出生后和成人视网膜中的表达:一项免疫组织化学研究。】 复制标题 收藏 收藏
    DOI:10.1017/s0952523800012104 复制DOI
    作者列表:Nag TC,Wadhwa S
    BACKGROUND & AIMS: The expression of GABA in the human fetal (12-25 weeks of gestation), postnatal (five-month-old), and adult (35-year-old) retinas was investigated by immunohistochemistry. GABA expression was seen as early as 12 weeks in the undifferentiated cells of the inner neuroblast zone; a few optic nerve fiber layer axons were clearly labeled, suggesting that some of the stained cell bodies were prospective ganglion cells, others could be displaced amacrine cells. From 16-17 to 24-25 weeks, intense labeling was found in the amacrine, displaced amacrine, and some ganglion cells. During this time period, horizontal cells (identified by calbindin immunohistochemistry), undergoing migration (periphery) and differentiation (center), expressed GABA prominently. In the postnatal retina, some horizontal cells were moderately labeled, but very weakly in a few cells, in the adult. The Müller cells developed immunoreactivity first weakly at 12 weeks and then moderately from 16-17 weeks onward. The staining was also evident in the postnatal and adult retinas, showing labeled processes of these glial cells. Virtually no axons in the adult optic nerve and nerve fiber layer were stained; the staining was restricted to a few, large ganglion cells and displaced amacrine cellsSome amacrines were also labeled. The possibility that GABA might play a role in horizontal cell differentiation and maturation is highlighted. Other evidences suggest that GABA might play a role in metabolism during retinal development.

    背景与目标: 通过免疫组织化学研究了GABA在人胎儿(妊娠12-25周),产后(5个月大)和成年(35岁)视网膜中的表达。早在12周内神经母细胞区未分化的细胞中就可以看到GABA的表达。一些视神经纤维层轴突被清楚地标记,表明某些染色的细胞体是预期的神经节细胞,其他可能是置换的无长突细胞。从16-17周到24-25周,在无长蛋白,移位的无长蛋白和一些神经节细胞中发现了强烈的标记。在这段时间内,水平细胞(通过钙结合蛋白免疫组织化学鉴定),经历迁移(外围)和分化(中心),主要表达GABA。在成年后的视网膜中,成人中一些水平细胞被中等程度标记,但少数细胞中非常弱。 Müller细胞首先在12周时出现弱免疫反应,然后在16-17周后逐渐出现免疫反应。染色在产后和成年视网膜中也很明显,显示出这些神经胶质细胞的标记过程。成人视神经和神经纤维层几乎没有染色。染色仅限于少数大型神经节细胞和置换的无长突细胞。一些无长突也被标记。强调了GABA可能在水平细胞分化和成熟中起作用的可能性。其他证据表明,GABA可能在视网膜发育过程中的代谢中发挥作用。

  • 【链霉菌A3(2)中的一种隐秘的I型聚酮合酶(cpk)基因簇。】 复制标题 收藏 收藏
    DOI:10.1007/s00203-006-0176-7 复制DOI
    作者列表:Pawlik K,Kotowska M,Chater KF,Kuczek K,Takano E
    BACKGROUND & AIMS: :The chromosome of Streptomyces coelicolor A3(2), a model organism for the genus Streptomyces, contains a cryptic type I polyketide synthase (PKS) gene cluster which was revealed when the genome was sequenced. The ca. 54-kb cluster contains three large genes, cpkA, cpkB and cpkC, encoding the PKS subunits. In silico analysis showed that the synthase consists of a loading module, five extension modules and a unique reductase as a terminal domain instead of a typical thioesterase. All acyltransferase domains are specific for a malonyl extender, and have a B-type ketoreductase. Tailoring and regulatory genes were also identified within the gene cluster. Surprisingly, some genes show high similarity to primary metabolite genes not commonly identified in any antibiotic biosynthesis cluster. Using western blot analysis with a PKS subunit (CpkC) antibody, CpkC was shown to be expressed in S. coelicolor at transition phase. Disruption of cpkC gave no obvious phenotype.
    背景与目标: :Streptomyces coelicolor A3(2)(Streptomyces属的一种模式生物)的染色体包含一个隐秘的I型聚酮化合物合酶(PKS)基因簇,该基因簇在对基因组进行测序时就可以显示出来。的ca。 54kb的簇包含三个大基因,编码PKS亚基的cpkA,cpkB和cpkC。在计算机分析中,该合成酶由一个加载模块,五个扩展模块和一个独特的还原酶(而不是典型的硫酯酶)作为末端结构域组成。所有酰基转移酶结构域都对丙二酰增量剂具有特异性,并具有B型酮还原酶。还可以在基因簇中鉴定出剪裁和调节基因。出乎意料的是,一些基因显示出与任何抗生素生物合成簇中未普遍鉴定的初级代谢产物基因高度相似。使用具有PKS亚基(CpkC)抗体的Western印迹分析,显示CpkC在过渡阶段在天蓝色链霉菌中表达。 cpkC的破坏没有明显的表型。
  • 【哪些血栓形成基因突变是复发性流产的危险因素?】 复制标题 收藏 收藏
    DOI:10.1111/j.1600-0897.2006.00419.x 复制DOI
    作者列表:Goodman CS,Coulam CB,Jeyendran RS,Acosta VA,Roussev R
    BACKGROUND & AIMS: PROBLEM:Thrombophilia has been associated with poor obstetrical outcomes. To determine the association of specific inherited thrombophilias and recurrent pregnancy loss, 10 thrombophilic genes were investigated. METHOD OF STUDY:A total of 550 women with a history of recurrent pregnancy loss had buccal swabs taken for DNA analyses of the following gene mutations: factor V G1691A, factor V H1299R (R2), factor V Y1702C, factor II prothrombin G20210A, factor XIII V34L, beta-fibrinogen -455G>A, PAI-1 4G/5G, HPA1 a/b(L33P), methylenetetrahydrofolate reductase (MTHFR) C677T, MTHFR A1298C. The frequencies of these mutations were compared with controls published in the literature. RESULTS:When examined individually, PAI-1 4G/5G (P = 0.009), factor XIII V34L (P < 0.0001), and homozygous MTHFR C667T (P < 0.0001) correlated significantly with recurrent pregnancy loss compared with controls. The frequency of the factor V Y1702C mutation was extremely low in patients and controls; thus, this gene was removed from further calculations. The remaining six mutated genes, when analyzed cumulatively, also corresponded with recurrent pregnancy loss (P < 0.0001). CONCLUSION:A panel of thrombogenic gene mutations consisting of factor V G1691A, factor V H1299R (R2), factor II prothrombin G20210A, factor XIII V34L, beta-fibrinogen -455G>A, PAI-1 4G/5G, HPA1 a/b(L33P), MTHFR C677T, and MTHFR A1298C can identify individuals at risk for recurrent pregnancy loss.
    背景与目标: 问题:血友病与产科预后不良有关。为了确定特定遗传性血栓形成症与复发性流产的关联,研究了10个血栓形成性基因。
    研究方法:总共550名有反复性流产史的妇女接受了口腔拭子以进行以下基因突变的DNA分析:因子V G1691A,因子V H1299R(R2),因子V Y1702C,因子II凝血酶原G20210A,因子XIII V34L,β-纤维蛋白原-455G> A,PAI-1 4G / 5G,HPA1a / b(L33P),亚甲基四氢叶酸还原酶(MTHFR)C677T,MTHFR A1298C。将这些突变的频率与文献中发表的对照进行了比较。
    结果:当单独检查时,与对照组相比,PAI-1 4G / 5G(P = 0.009),XIII因子V34L(P <0.0001)和纯合MTHFR C667T(P <0.0001)与复发性流产显着相关。在患者和对照组中,V Y1702C因子突变的频率非常低。因此,该基因已从进一步的计算中删除。当进行累积分析时,剩下的六个突变基因也与复发性流产相对应(P <0.0001)。
    结论:一组血栓形成基因突变,由因子V G1691A,因子V H1299R(R2),因子II凝血酶原G20210A,因子XIII V34L,β-纤维蛋白原-455G> A,PAI-1 4G / 5G,HPA1 a / b( L33P),MTHFR C677T和MTHFR A1298C可以识别有复发性流产风险的个体。
  • 【初始大鼠发育阶段对肝肿瘤促进过程中生化标志物表达的影响。】 复制标题 收藏 收藏
    DOI:10.1159/000217665 复制DOI
    作者列表:Decloître F,Lafarge-Frayssinet C,Barroso M,Lechner MC,Ouldelhkim M,Frayssinet C
    BACKGROUND & AIMS: :The phenotypic response of rat liver to a carcinogenic protocol involving initiation/selection and promotion with and without phenobarbital (PB) feeding was studied in pubertal and adult male rats. Considering the early presence of preneoplastic nodular areas, it appeared that pubertal rats, initiated at 6-7 weeks, presented a higher susceptibility to the protocol than adult rats initiated at 9-10 weeks. Altered liver phenotype was characterized by: (1) gamma-glutamyl-transpeptidase (GGT) and glutathione S-transferase (GST) activities; (2) the expression of two forms of cytochrome P-450; de novo PB-inducible P-450 II B 1,2 and P-450 II C 7 normally expressed in 45-day-old rats and PB-inducible, and (3) the expression of albumin and alpha-fetoprotein cDNAs. In the absence of PB, the susceptibility of pubertal rat liver to hepatocarcinogenesis was related to a special metabolic phenotype enriched in GGT and GST activities by comparison with the quasi-normal expression of both P-450s. Adult rat liver presented a less altered pattern closer to that of noninitiated rat liver. During PB promotion, the loss of PB inducibility of P-450 II C 7 in pubertal rat liver suggested that the hormonal status of the animals could interact with initiation to modulate specific gene expression. The late phase of PB promotion revealed the loss of highly differentiated functions (P-450s, albumin), whereas enzymatic markers associated with preneoplastic foci showed a persistent high expression.
    背景与目标: :在青春期和成年雄性大鼠中研究了大鼠肝脏对涉及开始/选择和促进(有或没有苯巴比妥(PB)喂养)致癌方案的表型反应。考虑到肿瘤前结节区域的早期存在,看来与在9-10周龄开始的成年大鼠相比,在6-7周龄开始的青春期大鼠对方案的敏感性更高。肝表型改变的特征是:(1)γ-谷氨酰转肽酶(GGT)和谷胱甘肽S-转移酶(GST)活性; (2)两种形式的细胞色素P-450的表达;从头开始PB诱导的P-450 II B 1,2和P-450 II C 7在45天大的大鼠中正常表达,并且在PB诱导中正常表达,以及(3)白蛋白和甲胎蛋白cDNA的表达。在没有PB的情况下,与两个P-450的准正常表达相比,青春期大鼠肝脏对肝癌发生的敏感性与富含GGT和GST活性的特殊代谢表型有关。成年大鼠肝脏的变化较小,与未启动大鼠肝脏的变化更接近。在PB促进过程中,青春期大鼠肝脏中P-450 II C 7的PB诱导性丧失,这表明动物的荷尔蒙状态可能与启动相互作用,从而调节特定的基因表达。 PB促进的晚期阶段揭示了高度分化的功能(P-450s,白蛋白)的丧失,而与肿瘤前病灶相关的酶标记物显示了持续的高表达。
  • 【肝细胞癌肝或外周血端粒酶表达的动态变化及其诊断意义。】 复制标题 收藏 收藏
    DOI:10.3748/wjg.v12.i31.4966 复制DOI
    作者列表:Yao DF,Wu W,Yao M,Qiu LW,Wu XH,Su XQ,Zou L,Yao DB,Meng XY
    BACKGROUND & AIMS: AIM:To investigate the dynamic alteration of telomerase expression during development of hepatocellular carcinoma (HCC) and its diagnostic implications in liver tissues or peripheral blood mononuclear cells for HCC. METHODS:Dynamic expressions of liver telomerase during malignant transformation of hepatocytes were observed in Sprague-Dawly (SD) rats fed with 0.05% of 2-fluoenyacetamide (2-FAA). Total RNA and telomerase were extracted from rat or human liver tissues. The telomerase activities in livers and in circulating blood were detected by a telomeric repeat amplification protocol-enzyme-linked immunosorbent assay (TRAP-ELISA), and its diagnostic value was investigated in patients with benign or malignant liver diseases. RESULTS:The hepatoma model displayed the dynamic expression of hepatic telomerase during HCC development. The telomerase activities were consistent with liver total RNA levels (r = 0.83, P<0.01) at the stages of degeneration, precancerosis, and cancerization of hepatocytes. In HCC patients, the telomerase levels in HCC tissues were significantly higher than in their adjacent non-cancerous tissues, but liver total RNA levels were lower in the former than in the latter. Although the circulating telomerase of HCC patients was abnormally expressed among patients with chronic liver diseases, the telomerase activity was a non-specific marker for HCC diagnosis, because the incidence was 15.7% in normal control, 25% in chronic hepatitis, 45.9% in liver cirrhosis, and 85.2% in HCC, respectively when absorbance value of telomerase activity was more than 0.2. If the value was over 0.6, the incidence was 60% in HCC group and 0% in any of the others (P<0.01) except in two cases with liver cirrhosis. However, the combination of circulating telomerase with serum alpha-fetoprotein level could increase the positive rate and the accuracy (92.6%, 125 of 135) of HCC diagnosis. CONCLUSION:The overexpression of telomerase is associated with HCC development, and its abnormality in liver tissues or in peripheral blood could be a useful marker for diagnosis and prognosis of HCC.
    背景与目标: 目的:研究肝细胞癌(HCC)发生过程中端粒酶表达的动态变化及其在肝组织或外周血单核细胞中的诊断意义。
    方法:在喂食0.05%2-氟乙酰胺(2-FAA)的Sprague-Dawly(SD)大鼠中观察肝细胞恶性转化过程中肝端粒酶的动态表达。从大鼠或人肝脏组织中提取总RNA和端粒酶。通过端粒重复扩增方案-酶联免疫吸附测定(TRAP-ELISA)检测肝脏和循环血液中的端粒酶活性,并对其在良性或恶性肝病患者中的诊断价值进行研究。
    结果:肝癌模型在肝癌发生过程中表现出肝端粒酶的动态表达。在变性,癌前期和肝细胞癌化阶段,端粒酶活性与肝脏总RNA水平一致(r = 0.83,P <0.01)。在HCC患者中,HCC组织中的端粒酶水平显着高于其相邻的非癌性组织,但前者的肝脏总RNA水平低于后者。尽管在慢性肝病患者中HCC患者的循环端粒酶异常表达,但端粒酶活性不是HCC诊断的非特异性标志物,因为正常对照组的发生率为15.7%,慢性肝炎为25%,肝为45.9%当端粒酶活性的吸光度值大于0.2时,肝硬化和HCC中的85.2%。如果该值超过0.6,则除两名肝硬化患者外,HCC组的发生率为60%,其他任何一组的发生率为0%(P <0.01)。但是,循环端粒酶与血清甲胎蛋白水平的结合可以提高HCC诊断的阳性率和准确性(92.6%,第125页)。
    结论:端粒酶高表达与肝癌的发生有关,其在肝组织或外周血中的异常可能是诊断和预后的有用标志。
  • 【拉坦前列素对人小梁网细胞中基质金属蛋白酶及其组织抑制剂表达的影响。】 复制标题 收藏 收藏
    DOI:10.1167/iovs.06-0036 复制DOI
    作者列表:Oh DJ,Martin JL,Williams AJ,Russell P,Birk DE,Rhee DJ
    BACKGROUND & AIMS: PURPOSE:To determine the effect of latanoprost on the expression of human matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in the trabecular meshwork (TM). METHODS:Total RNA was isolated, and qualitative RT-PCR was performed to detect the mRNA of MMPs and TIMPs in human TM tissue and explant cultures of TM endothelial cells. Cultures of TM cells were treated with vehicle control or latanoprost acid for 24 hours. Real-time RT-PCR of cell cultures from five different donors was performed to determine relative changes in expression. GAPDH served as an endogenous control. RESULTS:The mRNA of MMP-1, -2, -3, -11, -12, -14, -15, -16, -17, -19, and -24 and of TIMP-1 to -4 was present in TM tissue and cultures of TM cells. MMP-9 was not found. In control TM endothelial cells, the relative expression of MMP mRNA were MMP-2 and -14 > MMP-16, -19, and -24 > MMP-15 > MMP-11 and -17 > MMP-1 and -3 > MMP-12. The relative expressions of TIMP mRNA were TIMP-1 > TIMP-2 and -3 > TIMP-4. Latanoprost increased MMP-1 (in four of five cultures), MMP-3 (in four of five cultures), MMP-17 (in three of five cultures), MMP-24 (in all five cultures), TIMP-2, -3, and -4 expression (in three of five cultures); MMP-11 and -15 were downregulated. CONCLUSIONS:Contrary to the expected result, latanoprost seems to have a significant effect on TM cells. The transcription of the genes for MMP-1, -3, -17, and -24 is increased by latanoprost treatment. TIMP-2, -3, and -4 are also upregulated. The upregulation of these TIMPs may compensate for the increase of those MMPs. The absence of MMP-9 and concurrent upregulation of a greater number of TIMPs may explain the limited effect of latanoprost on TM outflow.
    背景与目标: 目的:确定拉坦前列素对小梁网(TM)中人基质金属蛋白酶(MMPs)和金属蛋白酶组织抑制剂(TIMPs)表达的影响。
    方法:分离总RNA,进行定性RT-PCR检测人TM组织和TM内皮细胞外植体培养物中MMP和TIMP的mRNA。用媒介物对照或拉坦前列素酸处理TM细胞的培养物24小时。对来自五个不同供体的细胞培养物进行了实时RT-PCR,以确定表达的相对变化。 GAPDH用作内源性对照。
    结果:MMP-1,-2,-3,-11,-12,-14,-15,-16,-17,-19和-24和TIMP-1至-4的mRNA均存在于TM组织和TM细胞培养物。找不到MMP-9。在对照TM内皮细胞中,MMP mRNA的相对表达为MMP-2和-14> MMP-16,-19和-24> MMP-15> MMP-11和-17> MMP-1和-3> MMP -12。 TIMP mRNA的相对表达为TIMP-1> TIMP-2和-3> TIMP-4。拉坦前列素可提高MMP-1(在五种文化中的四种),MMP-3(在五种文化中的四种),MMP-17(在五种文化中的三种),MMP-24(在五种文化中),TIMP-2,- 3和-4表达(在五种文化中的三种); MMP-11和-15下调。
    结论:与预期结果相反,拉坦前列素似乎对TM细胞有显着影响。拉坦前列素处理可增加MMP-1,-3,-17和-24基因的转录。 TIMP-2,-3和-4也被上调。这些TIMP的上调可以补偿那些MMP的增加。 MMP-9的缺乏和大量TIMP的同时上调可能解释了拉坦前列素对TM外流的作用有限。
  • 【大鼠角膜缘和中央角膜上皮中基因表达(SAGE)的系列分析。】 复制标题 收藏 收藏
    DOI:10.1167/iovs.06-0216 复制DOI
    作者列表:Adachi W,Ulanovsky H,Li Y,Norman B,Davis J,Piatigorsky J
    BACKGROUND & AIMS: PURPOSE:To identify genes preferentially expressed in the stem-cell-rich limbal epithelium of the rat cornea. METHODS:The limbal and central corneal epithelial cells of 6-week-old rats were isolated by microdissection. Serial analysis of gene expression (SAGE) libraries were constructed and analyzed, and in situ hybridization, reverse transcription-polymerase chain reaction (RT-PCR) and cDNA cloning were conducted by conventional procedures. RESULTS:The rat limbal and central corneal epithelial SAGE libraries consisted of 41,894 and 40,691 tags, respectively. After annotation, this was reduced to 759 transcripts specific for the limbal library and 844 transcripts specific for the central corneal library; 2292 transcripts overlapped. Transcripts encoding proteins with metabolic functions comprised the major functional category in both libraries. In situ hybridization and/or RT-PCR results of 12 of the most abundant, highly enriched transcripts in the limbal epithelium were in general agreement with the SAGE data and showed that these proteins are also expressed in the conjunctival epithelium. Interesting limbal-enriched transcripts encode WDNM1-like protein (similar to WDNM1/Expi, a putative secreted proteinase and inhibitor of metastasis), mesothelin (a cancer marker), marapsin (a trypsin-like serine protease that may control cell growth and migration), K4 and K15 (both cytokeratins), and membrane-spanning four-domain subfamily A member 8B. WDNM1-like protein was cloned and confirmed as a member of the four-disulfide core family. CONCLUSIONS:The SAGE results extend the database of genes expressed in the rodent cornea and suggest an association between several genes preferentially expressed in the limbal epithelium with cellular proliferation and migration.
    背景与目标: 目的:鉴定在大鼠角膜的富含干细胞的角膜缘上皮细胞中优先表达的基因。
    方法:采用显微解剖技术分离6周龄大鼠角膜缘和角膜上皮细胞。构建并分析基因表达(SAGE)库的序列分析,并通过常规方法进行原位杂交,逆转录-聚合酶链反应(RT-PCR)和cDNA克隆。
    结果:大鼠角膜缘和角膜上皮SAGE文库分别由41,894和40,691个标签组成。注释后,该数目减少为角膜缘文库特异的759个转录物和中央角膜文库特异的844个转录物。 2292个成绩单重叠。编码具有代谢功能的蛋白质的转录本在两个文库中均属于主要功能类别。角膜缘上皮细胞中12种最丰富,高度富集的转录本的原位杂交和/或RT-PCR结果与SAGE数据基本一致,表明这些蛋白也在结膜上皮细胞中表达。有趣的角膜缘丰富的转录本编码WDNM1样蛋白(类似于WDNM1 / Expi,一种推测的分泌蛋白酶和转移抑制剂),间皮素(一种癌症标志物),marapsin(一种胰蛋白酶样丝氨酸蛋白酶,可以控制细胞的生长和迁移)。 ,K4和K15(均为细胞角蛋白)和跨膜四结构域亚家族A成员8B。克隆了类似WDNM1的蛋白质,并确认其为四-二硫键核心家族的成员。
    结论:SAGE结果扩展了在啮齿动物角膜中表达的基因数据库,并暗示了在角膜缘上皮中优先表达的几个基因与细胞增殖和迁移之间的关联。
  • 【与人神经胶质瘤有关的人核糖体蛋白L14.22的全长新基因。】 复制标题 收藏 收藏
    DOI: 复制DOI
    作者列表:Qi ZY,Hui GZ,Li Y,Zhou ZX,Gu SH,Xie Y
    BACKGROUND & AIMS: BACKGROUND:This study was undertaken to obtain differentially expressed genes related to human glioma by cDNA microarray and the characterization of a novel full-length gene. METHODS:Total RNA was extracted form human glioma and normal brain tissue, and mRNA was used as a probe. The results of hybridization procedure were scanned with the computer system. The gene named 507E08 cone was subsequently analyzed by northern blot, bioinformatic approach, and protein expression. RESULTS:Fifteen differentially expressed genes were obtained from human glioma by hybridization and scanning for four times. Northern blot analysis confirmed that the 507E08 clone was low expressed in human brain tissue and over expressed in human glioma tissues. The analysis of BLASTn and BLASTx showed that the 507E08 clone was a novel full-length gene, which codes 203 amino acid of protein and is called human ribosomal protein 14.22 gene. The nucleotide sequence had been submitted to the GenBank with the accession number of AF329277. After expression in E. coli., protein yielded a major band of apparent molecular mass 22 kDa on an SDS-PAGE gel. CONCLUSIONS:cDNA microarray technology can be successfully used to identify differentially expressed genes. The novel full-length gene of human ribosomal protein 13.22 may be correlated with the development of human glioma.
    背景与目标: 背景:本研究旨在通过cDNA微阵列技术获得与人神经胶质瘤相关的差异表达基因,并鉴定一个新的全长基因。
    方法:从人脑胶质瘤和正常脑组织中提取总RNA,并以mRNA为探针。杂交程序的结果用计算机系统扫描。随后通过Northern印迹,生物信息学方法和蛋白质表达来分析名为507E08视锥细胞的基因。
    结果:通过杂交和扫描4次,从人脑胶质瘤中获得了15个差异表达基因。 Northern印迹分析证实507E08克隆在人脑组织中低表达而在人脑胶质瘤组织中高表达。对BLASTn和BLASTx的分析表明,507E08克隆是一个新颖的全长基因,其编码蛋白质的203个氨基酸,被称为人核糖体蛋白质14.22基因。该核苷酸序列已提交给GenBank,登录号为AF329277。在大肠杆菌中表达后,蛋白质在SDS-PAGE凝胶上产生一条表观分子量为22 kDa的主带。
    结论:cDNA微阵列技术可成功用于鉴定差异表达的基因。人核糖体蛋白13.22的新的全长基因可能与人脑胶质瘤的发展有关。
  • 【HIV-1 RNA的贩运是由异质核糖核蛋白A2的表达介导的,并且对病毒的组装也有影响。】 复制标题 收藏 收藏
    DOI:10.1111/j.1600-0854.2006.00461.x 复制DOI
    作者列表:Lévesque K,Halvorsen M,Abrahamyan L,Chatel-Chaix L,Poupon V,Gordon H,DesGroseillers L,Gatignol A,Mouland AJ
    BACKGROUND & AIMS: :Few details are known about how the human immunodeficiency virus type 1 (HIV-1) genomic RNA is trafficked in the cytoplasm. Part of this process is controlled by the activity of heterogeneous nuclear ribonucleoprotein A2 (hnRNP A2). The role of hnRNP A2 during the expression of a bona fide provirus in HeLa cells is investigated in this study. Using immunofluorescence and fluorescence in situ hybridization techniques, we show that knockdown of hnRNP A2 expression in HIV-1-expressing cells results in the rapid accumulation of HIV-1 genomic RNA in a distinct, cytoplasmic space that corresponds to the microtubule-organizing center (MTOC). The RNA exits in the nucleus and accumulates at the MTOC region as a result of hnRNP A2 knockdown even during the expression of a provirus harboring mutations in the hnRNP A2-response element (A2RE), the expression of which results in nuclear retention of genomic RNA. We also demonstrate that hnRNP A2 expression is required for downstream trafficking of genomic RNA from the MTOC in the cytoplasm. Genomic RNA localization at the MTOC that was both the result of hnRNP A2 knockdown and the overexpression of Rab7-interacting lysosomal protein had little effect on pr55Gag synthesis but negatively influenced virus production and infectivity. These data indicate that altered HIV-1 genomic RNA localization modulates viral assembly and that the MTOC serves as a central site to which HIV-1 genomic RNA converges following its exit from the nucleus, with the host protein, hnRNP A2, playing a central role in taking it to and from this site in the cell.
    背景与目标: 关于人类免疫缺陷病毒1型(HIV-1)基因组RNA如何在细胞质中运输的信息鲜为人知。该过程的一部分由异质核糖核蛋白A2(hnRNP A2)的活性控制。这项研究调查了hnRNP A2在HeLa细胞中表达真正的原病毒的过程中的作用。使用免疫荧光和荧光原位杂交技术,我们显示敲低表达HIV-1的细胞中hnRNP A2表达的表达导致HIV-1基因组RNA在与微管组织中心相对应的独特细胞质空间中的快速积累( MTOC)。即使hnRNP A2反应元件(A2RE)中携带有突变的原病毒表达,RNA仍会通过hnRNP A2敲除而留在细胞核中并在MTOC区域积聚,其表达会导致基因组RNA的核保留。我们还证明hnRNP A2表达是从细胞质MTOC下游运输基因组RNA所必需的。基因组RNA在MTOC处的定位既是hnRNP A2敲除的结果,又是与Rab7相互作用的溶酶体蛋白的过表达,对pr55Gag的合成影响很小,但对病毒的产生和感染性产生负面影响。这些数据表明,改变的HIV-1基因组RNA定位可调节病毒装配,MTOC充当HIV-1基因组RNA从细胞核退出后向其汇聚的中心位点,宿主蛋白hnRNP A2发挥着核心作用。将其带入和移出该单元格中的此站点。
  • 【罕见的母体mRNA编码调控蛋白,这些蛋白控制着海胆胚胎中谱系特异性基因的表达。】 复制标题 收藏 收藏
    DOI:10.1073/pnas.87.20.7953 复制DOI
    作者列表:Cutting AE,Höög C,Calzone FJ,Britten RJ,Davidson EH
    BACKGROUND & AIMS: :The prevalence of mRNAs coding for the sea urchin embryo regulatory factors P3A1 and P3A2 was measured by single-strand probe excess solution hybridization. P3A1 and P3A2 are not homologous proteins, though they both bind specifically to a particular cis-regulatory sequence. Interaction at this target site is known to be required for lineage-specific expression of an aboral ectoderm-specific gene and probably for several other genes as well. Genome blot hybridizations show that both factors are encoded by single-copy genes. Maternal mRNAs for both factors are present at less than 10(3) molecules per egg, which places them in the rare mRNA class. During development to the mesenchyme blastula stage, the amount of P3A1 mRNA (per embryo) increases severalfold while that of P3A2 remains approximately constant. Specification of the aboral ectoderm founder cells and of their initial patterns of gene expression must occur during early to mid-cleavage stage. Therefore, the regulatory proteins needed for this process must be produced by this stage. We show that the quantities of the P3A proteins that can be synthesized from the numbers of mRNA molecules present in the large blastomeres of the early embryo are sufficient to be functional, because these proteins will be accumulated in the nuclei. Thus maternal P3A1 or P3A2 proteins asre not required, nor were these detected in earlier studies. Furthermore, differential spatial (as well as temporal) distribution of both of these newly synthesized factor species could result from the unequal cleavage pattern utilized in the sea urchin egg.
    背景与目标: :通过单链探针过量溶液杂交测量编码海胆胚胎调节因子P3A1和P3A2的mRNA的发生率。尽管P3A1和P3A2都与特定的顺式调控序列特异性结合,但它们不是同源蛋白。已知该靶标位点的相互作用是特定于宗族的外胚层特异性基因的谱系表达所必需的,并且可能还需要其他几个基因。基因组印迹杂交显示这两个因子均由单拷贝基因编码。每个鸡蛋的母体mRNA均少于10(3)个分子,这使它们处于罕见的mRNA类中。在发育到间充质囊阶段期间,P3A1 mRNA(每个胚胎)的数量增加了几倍,而P3A2的数量则保持大致恒定。胎盘外胚层基础细胞及其基因表达的初始模式的规范必须在卵裂早期至中期进行。因此,必须在该阶段生产该过程所需的调节蛋白。我们表明,可以从早期胚胎的大型卵裂球中存在的mRNA分子数量合成的P3A蛋白数量足以发挥功能,因为这些蛋白将积聚在细胞核中。因此,不需要母体P3A1或P3A2蛋白,也不需要在早期研究中检测到这些蛋白。此外,这两种新合成的因子物种在空间(以及时间)上的差异分布可能是由于海胆卵中使用的不均等卵裂模式造成的。
  • 【苯巴比妥依赖和戒断大鼠脑中谷氨酸受体,c-fos mRNA表达和激活蛋白-1(AP-1)DNA结合活性的变化。】 复制标题 收藏 收藏
    DOI:10.1016/s0006-8993(97)00134-0 复制DOI
    作者列表:Tanaka S,Kiuchi Y,Numazawa S,Oguchi K,Yoshida T,Kuroiwa Y
    BACKGROUND & AIMS: We studied changes in glutamate receptors, expression of immediate early genes, and AP-1 DNA binding activity in the brains of phenobarbital (PB)-dependent and -withdrawn rats to investigate the possible involvement of activation of glutamate receptors in PB withdrawal syndrome. PB-dependent rats were prepared by feeding drug-admixed food for 5 weeks. Autoradiographic analysis showed that binding of [3H(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imin e (MK-801), an antagonist of N-methyl-D-aspartic acid (NMDA) receptors, increased significantly in the cerebral cortices of PB-dependent and 24-h-withdrawn rats. However, [3H]MK-801 binding in the hippocampus and [3H]6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and [3H]kainic acid binding in the hippocampus and cerebral cortex were essentially unchanged in both groups. PB withdrawal seizures were followed by increased expression of c-fos mRNA in the hippocampus and cerebral cortex and of c-jun mRNA in the cerebral cortex. The induction of c-fos and c-jun mRNA was suppressed by administration of MK-801. Furthermore, PB withdrawal enhanced AP-1 DNA binding activity in the brain. The present findings suggest functional enhancement of glutamatergic neurotransmission during the development of PB withdrawal syndrome.

    背景与目标: 我们研究了苯巴比妥(PB)依赖和戒断大鼠大脑中谷氨酸受体的变化,立即早期基因的表达以及AP-1 DNA结合活性,以研究PB戒断综合征中谷氨酸受体活化的可能。通过喂食药物混合的食物5周来制备PB依赖的大鼠。放射自显影分析表明,结合物[3H()-5-甲基-10,11-二氢-5H-二苯并[a,d]环庚烯-5,10-亚胺e(MK-801)是一种N-甲基-拮抗剂D-天冬氨酸(NMDA)受体,在PB依赖和24小时戒断大鼠的大脑皮层中显着增加。然而,在海马中的[3H] MK-801结合以及在海马和大脑皮层中的[3H] 6-氰基-7-硝基喹喔啉-2,3-二酮(CNQX)和[3H]海藻酸结合基本上没有变化。组。 PB抽搐发作后,海马和大脑皮层中c-fos mRNA的表达增加,大脑皮层中c-jun mRNA的表达增加。通过施用MK-801抑制了c-fos和c-jun mRNA的诱导。此外,PB撤离可增强大脑中AP-1 DNA的结合活性。目前的发现表明,在PB戒断综合征的发展过程中,谷氨酸能神经传递的功能增强。

  • 【电离辐射以肺内不同的组织学模式介导细胞粘附分子的表达。】 复制标题 收藏 收藏
    DOI: 复制DOI
    作者列表:Hallahan DE,Virudachalam S
    BACKGROUND & AIMS: Inflammatory cell infiltration of the lung is a predominant histopathological change that occurs during radiation pneumonitis. Emigration of inflammatory cells from the circulation requires the interaction between cell adhesion molecules on the vascular endothelium and molecules on the surface of leukocytes. We studied the immunohistochemical pattern of expression of cell adhesion molecules in lungs from mice treated with thoracic irradiation. After X-irradiation, the endothelial leukocyte adhesion molecule 1 (ELAM-1; E-selectin) was primarily expressed in the pulmonary endothelium of larger vessels and minimally in the microvascular endothelium. Conversely, the intercellular adhesion molecule 1 (ICAM-1; CD54) was expressed in the pulmonary capillary endothelium and minimally in the endothelium of larger vessels. Radiation-mediated E-selectin expression was first observed at 6 h, whereas ICAM-1 expression initially increased at 24 h after irradiation. ICAM-1 and E-selectin expression persisted for several days. P-selectin is constitutively expressed in Weibel-Palade bodies in the endothelium, which moved to the vascular lumen within 30 min after irradiation. P-selectin was not detected in the pulmonary endothelium at 6 h after irradiation. The radiation dose required for increased cell adhesion molecule expression within the pulmonary vascular endothelium was 2 Gy, and expression increased in a dose-dependent manner. These data demonstrate that ICAM-1 and E-selectin expression is increased in the pulmonary endothelium following thoracic irradiation. The pattern of expression of E-selectin, P-selectin, and ICAM-1 is distinct from one another.

    背景与目标: 肺炎性细胞浸润是放射性肺炎期间发生的主要组织病理学变化。炎性细胞从循环系统的迁移需要血管内皮细胞粘附分子与白细胞表面分子之间的相互作用。我们研究了用胸腔照射治疗的小鼠肺中细胞粘附分子表达的免疫组织化学模式。 X射线照射后,内皮白细胞粘附分子1(ELAM-1; E-选择素)主要在较大血管的肺内皮中表达,而在微血管内皮中则最低表达。相反,细胞间粘附分子1(ICAM-1; CD54)在肺毛细血管内皮中表达,而在较大血管的内皮中表达最少。辐射介导的E-选择素表达首先在6 h观察到,而ICAM-1表达最初在辐射后24 h增加。 ICAM-1和E-选择素表达持续数天。 P-选择蛋白在内皮的Weibel-Palade体中组成性表达,在照射后30分钟内移至血管腔。照射后6 h在肺内皮中未检测到P-选择素。肺血管内皮细胞中细胞粘附分子表达增加所需的辐射剂量为2 Gy,并且表达以剂量依赖性方式增加。这些数据表明,胸腔照射后,肺内皮中的ICAM-1和E-选择蛋白表达增加。 E-选择素,P-选择素和ICAM-1的表达模式彼此不同。

  • 【雌二醇通过上调Fas和Fas配体表达来增加人冠状动脉内皮细胞的凋亡。】 复制标题 收藏 收藏
    DOI:10.1210/jc.2006-1225 复制DOI
    作者列表:Seli E,Guzeloglu-Kayisli O,Cakmak H,Kayisli UA,Selam B,Arici A
    BACKGROUND & AIMS: CONTEXT:In animal models, estrogen inhibits atherogenesis by inhibiting many of the early steps of atherosclerotic plaque formation. However, the lack of cardioprotective effect by postmenopausal hormone replacement therapy and possible increase in cardiovascular events observed during the first year after the initiation of hormone replacement therapy may suggest that once the plaque is formed, estrogen may have additional effects that may counteract its beneficial outcomes. Indeed, the effect of estrogen on plaque stability has not been identified. OBJECTIVE:We hypothesized that 17beta-estradiol (E2) may cause increased apoptosis in human coronary artery endothelial cells (HCAECs). This effect would explain an adverse effect on plaque stability in vivo. INTERVENTION(S) AND MAIN OUTCOME MEASURE(S):The effect of E2 on apoptosis, cell proliferation, and expression of proapoptotic molecules Fas and Fas ligand (FasL) in cultured HCAECs was evaluated. RESULTS:HCAECs in culture treated with E2 showed an increase in DNA strand breaks and nuclear fragmentation indicative of apoptosis. E2 treatment also induced a significant concentration-dependent increase in Fas mRNA and protein expressions in HCAECs. Moreover, the expression of FasL mRNA and secretion of FasL protein by HCAECs were enhanced in response to E2 treatments. CONCLUSIONS:E2 increases the apoptosis in cultured HCAECs. Enhanced Fas and FasL expressions in response to E2 suggest that activation of the Fas/FasL pathway may be a mediator of the proapoptotic effects of E2 in these cells.
    背景与目标: 背景:在动物模型中,雌激素通过抑制动脉粥样硬化斑块形成的许多早期步骤来抑制动脉粥样硬化。但是,绝经后激素替代疗法缺乏心脏保护作用,并且在开始激素替代疗法后的第一年内观察到的心血管事件可能增加,这表明一旦形成斑块,雌激素可能会产生额外的作用,从而抵消其有益结果。实际上,尚未确定雌激素对斑块稳定性的作用。
    目的:我们假设17β-雌二醇(E2)可能导致人冠状动脉内皮细胞(HCAEC)凋亡增加。该作用将解释对体内斑块稳定性的不利影响。
    干预和主要观察指标:评估了E2对培养的HCAEC中细胞凋亡,细胞增殖以及促凋亡分子Fas和FasL配体(FasL)表达的影响。
    结果:用E2处理的培养物中的HCAECs显示DNA链断裂的增加和核碎裂指示凋亡。 E2处理还诱导了HCAECs Fas mRNA和蛋白表达的浓度依赖性显着增加。而且,响应于E2处理,HCAECs的FasL mRNA的表达和FasL蛋白的分泌得到增强。
    结论:E2增加了培养的HCAECs的细胞凋亡。响应E2增强的Fas和FasL表达表明Fas / FasL途径的激活可能是这些细胞中E2促凋亡作用的介质。
  • 15 bcl-2 expression in pilomatricoma. 复制标题 收藏 收藏

    【bcl-2在pilomatricoma中的表达。】 复制标题 收藏 收藏
    DOI:10.1097/00000372-199706000-00009 复制DOI
    作者列表:Farrier S,Morgan M
    BACKGROUND & AIMS: Pilomatricoma is a distinctive tumor characterized by a dual population of proliferating basophilic cells and diagnostic shadow cells, believed to arise from the hair matrix. The normal hair matrix undergoes defined cycles of growth (anagen), regression (catagen), and resting (telogen) that are regulated by programmed cell death (apoptosis). bcl-2 is a proto-oncogene that helps to suppress apoptosis in both benign and malignant tumors. In addition, both apoptosis and bel-2 are critical factors in normal hair follicle development. In order to clarify the role of bcl-1, we used immunohistochemical means to study 10 cases of histologically proven pilomatricoma for bcl-2 expression. The study design included both positive and negative controls. All of the pilomatricomas in our series were strongly decorated by bcl-2 immunostaining. Based on our findings of increased bcl-2 staining, we concluded that the faulty suppression of apoptosis contributes to the pathogenesis of pilomatricoma.

    背景与目标: 口唇瘤是一种独特的肿瘤,其特征是增殖的嗜碱性细胞和诊断性影子细胞由双重人群组成,据信它们是由毛发基质引起的。正常的头发基质会经历特定的生长(生长期),消退(退化期)和静止(休止期)周期,这些周期受程序性细胞死亡(细胞凋亡)的调节。 bcl-2是一种原癌基因,有助于抑制良性和恶性肿瘤中的细胞凋亡。此外,凋亡和bel-2都是正常毛囊发育的关键因素。为了阐明bcl-1的作用,我们使用免疫组化方法研究了10例经组织学证实的pilomatricoma的bcl-2表达。研究设计包括阳性和阴性对照。 bcl-2免疫染色强烈修饰了我们系列中的所有pilomatricomas。根据bcl-2染色增加的发现,我们得出结论,凋亡的错误抑制是导致毛细血管瘤的发病原因。

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