Caspase-3和hTERT在Barrett’s食管中的表达及其相互关系

目的研究半胱天冬蛋白酶-3（Caspase-3）和端粒酶逆转录酶（hTERT）在Barrett’s食管（Barrett’s esophagus,BE）及食管炎伴贲门腺或胃底腺化生组织中的表达,探讨其在BE发病中的作用。方法应用免疫组织化学法（LDP）检测22例BE和38例食管炎伴贲门腺或胃底腺化生组织中Caspase-3和hTERT的表达,比较两者表达的相关性。结果Caspase-3在BE中的阳性表达率为95.45%,明显高于在食管炎伴贲门腺或胃底腺化生组织中的表达（68.42%）（P=0.000）;hTERT在BE中的阳性表达率为27.27%,明显低于在食管炎伴贲门腺或胃底腺化生组织中的表达（65.79%）（P=0.000）。BE中Caspase-3与hTERT的表达呈强负相关（r=-1,P=0.000）。结论BE中Caspase-3高表达,hTERT低表达。Caspase-3和hTERT在BE发生过程中起重要作用,可能成为预测BE发生的重要参考指标。     

反义 hTERT 基因对 K562 细胞凋亡及周期的影响简

目的：探讨反义hTERT基因抑制K562细胞端粒酶活性对其细胞周期及增殖的影响。方法：用脂质体介导反义hTERT基因和空载体质粒转染K562细胞株,绘制生长曲线,MTT法检测细胞活性,血清饥饿法同步化细胞周期,流式细胞仪检测细胞周期及细胞凋亡率变化。结果：反义hTERT基因转染后,细胞生长明显受到抑制。细胞同步化后,流式细胞仪检测显示：KA组细胞出现早期凋亡峰,细胞阻滞于S期。但正常培养48h,各组细胞无明显凋亡,周期无显著差异。结论：反义hTERT基因可明显抑制K562的增殖,细胞同步化后具有促凋亡作用。     

Expression level of hTERT is regulated by somatic mutation and common single nucleotide polymorphism at promoter region in glioblastoma

We investigated the role of somatic mutations and a common single nucleotide polymorphism (SNP) in the hTERT promoter region on hTERT expression and clinical outcomes. The hTERT promoter region was sequenced from 48 glioblastomas. hTERT expression was analyzed by quantitative real time-PCR. The association between hTERT promoter genetic changes and other genomic events and clinical variables common in gliomas were examined. C228T and C250T somatic mutations were found in 60.4% of glioblastomas, and a common SNP (T349C) was found in 66.6%. Somatic mutations and the SNP likely have opposing effects on hTERT expression. hTERT expression was significantly higher in the C228T or C250T mutated tumors. Tumors with the T349C genotype showed lower hTERT expression when C228T or C250T mutations were present. However, no significant survival differences were observed among the groups with or without hTERT promoter mutations and SNP. There was a significant association between genetic changes in the hTERT promoter and patient age as well as MGMT promoter methylation and EGFR amplification. hTERT expression is modulated by somatic mutations in the hTERT promoter as well as a common polymorphism. However, hTERT related genomic changes have limited value as an independent prognostic factor for clinical outcomes in glioblastomas.     

hTERT启动子调控下PE38KDEL表达载体的构建及其在MiaPaCa2人胰腺癌细胞中的表达

目的探讨端粒酶逆转录酶（hTERT）启动子调控铜绿假单胞菌外毒素衍生物（PE38KDEL）表达载体的构建及其在MiaPaCa2人胰腺癌细胞中的表达。方法通过PCR法扩增hTERT启动子,全基因合成PE38KDEL,将其分别亚克隆到pIRES2-EGFP及phTERTp-IRES2-EGFP质粒的多克隆位点中;均经酶切及测序鉴定。通过脂质体法将两质粒分别转染MiaPaCa2及WI-38细胞,观察荧光表达情况,通过RT-PCR检测PE38KDEL的表达。结果经酶切及测序鉴定证实质粒构建成功,pPE38KDEL-IRKS2-EGFP转染后MiaPaCa2和WI-38细胞中均有PE38KDEL基因表达;但phTERTp—PE38KDEL-IRES2-EGFP转染后仅MiaPaCa2中有PE38KDEL基因表达。结论hTERT调控下PE38KDEL表达载体构建成功,并可在MiaPaCa2人胰腺癌细胞中靶向性表达,为探讨胰腺癌的靶向基因治疗提供了实验依据。