Expression and effects of human telomerase RNA in testicular tumor

Human telomerase RNA (hTR) plays an important role in determining repeated telomere sequence and the expression of an antisense telomerase RNA that leads to telomere shortening and cell death. Using highly sensitive in situ nucleic acid hybridisation, we investigated the expression of hTR in human testicular     

Expression of Beclin1 in non small cell lung cancer and its clinical significance

Objective:To investigate the expression of autophagy related gene;Beclin1 in human non-small cell lung cancer(NSCLC)tissues.Methods:Protein expression of Beclin1 was determined by immunofluorescence staining and Western Blot,mRNA expression was analyzed by RT-PCR.Results:Immunofluorescence staining revealed that the level of Beclin1 expression in lung cancer was significantly lower than that in adjacent noncancerous tissues and normal tissues(expression rate 8.3%,P=0.000).The Beclin1 mRNA expression in lung cancer,adjacent noncancerous tissues and normal tissues was 1.372(±0.475)1.721(±0.521)and 1.553(±0.554)when F = 15.0,P < 0.01.Beclin1 protein expression in lung cancer,adjacent noncancerous tissues and normal tissues was 3.453(±0.852)5.423(±1.351)and 6.878(±0.997)F = 11.2,P < 0.01.Conclusion:The protein and mRNA expression of Beclin1 in lung cancer was much lower than those in and around cancer tissues and normal tissues,those differences having statistical significance.However in adjacent noncancerous tissues and normal tissues,the expression showed no difference.     

Polymorphism of the DNA repair gene XPA and susceptibility to lung cancer

Objective： To study the relationship between one polymorphism in the promoter of the DNA repair gene XPA and the susceptibility to lung cancer. Methods： Genotypes were determined by the PCR-restriction fragment length polymorphism （PCR-RFLP） method in 310 histologically-confirmed lung cancer cases and 341 age and sex frequency-matched cancer-free controls. Results： The XPA A23G genotype frequencies were 27.1% （AA）, 42.9% （AG）, and 30.0% （GG） in case patients and21.1% （AA）, 5218% （AG）, and 26.1% （C-G） in control subjects. Multivariate logistic regression analysis revealed that individuals carrying at least one 23G variant allele （AG ＋ GG genotypes） had a significantly decreased risk for lung cancer （adjusted OR = 0.66; 95 % CI = 0.44- 0.98） compared with the wild-type genotype （23AA）. Stratified analysis showed that the protective effect was more evident in subjects with a family history of cancer. Conclusion： These results suggest that the XPA A23G polymorphism may have a role in lung cancer susceptibility in this study population.     

Determination of telomerase activity and its clinical significance in gastric cancer and premalignant lesions

Telomerase,acomplexstructurepresentattheendsofeukaryoticchromosomes,consistsoftandemarraysofhighlyconservedhexameric(TTAGGG)repeatsinvertebrates.FunctionsoftelomerasearetoprotectgenomicDNAfromdegradationofdeleteriousrecombinationandsignificantlyresolvetheendreplicationproblem['.'J.Ithasbeenproposedthatprogressiveshorteningoftelomeraseinnormalsomaticcellsmayberegardedasamitoticclock,eventuallysignalingthesenescenceofprogrammedcelldeathinagedcells,thus,preventingfurtheraccumulatedgeneticchan…     

Expression of nerve growth factor and hypoxia inducible factor-1α and its correlation with angiogenesis in non-small cell lung cancer

Summary： In order to investigate the expression of nerve growth factor （NGF） and hypoxia inducible factor-1α （HIF-1α） and its correlation with angiogenesis in non-small cell lung cancer （NSCLC）, paraffin-embedded tissue blocks from 20 patients with NSCLC were examined. Twenty corresponding para-cancerous lung tissue specimens were obtained to serve as a control. The expression of NGF, HIF-1α, and vascular endothelial growth factor （VEGF） in the NSCLC tissues was detected by using immunohistochemistry. The microvascular density （MVD） was determined by CD31 staining. The resuits showed that the expression levels ofNGF, HIF-1α and VEGF in the NSCLC tissues were remarkably higher than those in the para-cancerous lung tissues （P〈0.05）. There was significant difference in the MVD between the NSCLC tissues （9.19±1.43） and para-cancerous lung tissues （2.23±1.19） （P〈0.05）. There were positive correlations between NGF and VEGF, between HIF-1α and VEGF, and between NGF and HIF-1α in NSCLC tissues, with the spearman correlation coefficient being 0.588, 0.519 and 0.588, respectively. In NSCLC tissues, the MVD had a positive correlation with the three factors （P〈0.05）. Theses results suggest that NGF and HIF-1α are synergically involved in the angiogenesis of NSCLC.     

Identification of p63 expression in human lung cancer： analysis by complementary DNA and tissue microarray

p6 3,anovelp5 3 relatedgene ,mappingat 3q2 7 q2 9,hasrecentlybeenidentifiedandcharacterized[1- 3] .Ithasbeenreportedthattheexpressionofp6 3wasloworabsentinasubsetoflungcancer[4 ] .However ,otherstud iesrevealedthatp6 3expressionwasfoundmainlyinlungsquamouscellcancer[5- 7] .Theroleofp6 3inlungcar cinogenesisanditsexpressionpattern ,however ,stillde servefurtherinvestigation .　　Inthisstudy ,weinvestigatedp6 3expressionbycD NAmicroarrayanalysisandimmunohistochemistryaswellaschromosomalimbalanc…     

老年肺癌患者外周血单个核细胞hTERT基因表达

目的探讨外周血单个核细胞(PBMCs)的人端粒酶反转录酶(hTERT)基因的表达水平对于老年肺癌的诊断意义.方法采用RT-PCR定量检测20例老年肺癌患者及10例正常老年人外周血单个核细胞的hTERT的基因表达水平.结果老年肺癌患者hTERTmRNA的水平明显高于正常对照组(0.927±0.115vs0.408±0.124,P<0.001).与病理类型与临床分期无相关性.结论定量测定PBMCs的hTERTmR-NA的表达水平是诊断不适合其他检查的老年肺癌患者的有效方法.     

RNA干扰人端粒酶逆转录酶基因对膀胱尿路上皮癌BIU-87细胞株端粒酶活性抑制的研究

目的:探讨RNA干扰技术使人端粒酶逆转录酶(hTERT)基因沉默,对膀胱尿路上皮癌BIU-87细胞株端粒酶活性的抑制作用。方法:针对hTERT基因设计构建多种siRNA表达载体,并转染至膀胱肿瘤BIU-87细胞,通过实时荧光定量、单四唑(MTT)检测端粒酶活性,观察siRNA表达载体对hTERT基因表达的影响。结果:实时荧光定量和MTT检测证明,所构建的载体能导致不同程度的hTERT基因沉默,并成功地抑制BIU-87细胞的生长。结论:RNA干扰膀胱尿路上皮癌细胞(BIU-87)端粒酶hTERT基因,能影响端粒酶基因的表达,为进一步研究端粒酶活性在膀胱尿路上皮癌中的作用奠定了基础。     

乳癌组织中端粒酶活性与端粒酶RNA的表达

目的:分析乳癌组织中端粒酶(telomerase)活性和端粒酶mRNA(hTR)的表达.方法:采用PCR-TRAP-ELISA法对54例乳癌、配对癌旁及30例乳腺良性病变组织中端粒酶活性进行半定量检测,运用原位杂交方法检测hTR的表达.结果:癌组织中端粒酶活性及hTR表达率高于癌旁及良性病变组织(P＜0.01).乳癌组织中端粒酶活性的表达与患者年龄、肿块大小、分化程度无关(P＞0.05),但与组织学类型及淋巴结转移关系密切(P＜0.05).结论:端粒酶激活与乳癌的发生关系密切.     

人端粒酶催化亚单位基因正反义腺病毒表达载体的构建及鉴定

目的: 构建hTERT正义和反义腺病毒表达载体. 方法: 用EcoRⅠ从pGRN145质粒上切下约3.5 kb的人端粒酶全长cDNA片段,然后连入pDC315质粒的EcoRⅠ酶切位点上,经BamHⅠ酶切鉴定出正义和反义表达载体,并对正反义重组质粒进一步测序鉴定其方向. 结果: 经BamHⅠ酶切后,正义质粒形成1.0 6.4 kb两条带. 反义重组质粒为1.0 2.5 3.9 kb三条带,与理论计算值完全一致,测序结果进一步确认了方向的正确性. 结论: 成功构建了hTERT的正、反义腺病毒表达载体.     

人胎肺发育不同阶段端粒酶逆转录酶的表达及意义

目的 检测端粒酶逆转录酶(hTERT)在人胎肺发育不同阶段的表达特征,探讨其在胎肺上皮干细胞发育、分化过程中的作用及意义。方法 对孕妇自愿捐献的胎龄为10~34周的引产胎儿肺组织,用免疫组织化学技术检测hTERT的表达。结果 胎龄10周的肺组织即检测到hTERT的表达,主要表达于正在分支的支气管原始上皮细胞浆内。随着胎肺发育,阳性表达的细胞逐渐向远端呼吸道迁移。到发育晚期,呼吸道上皮hTERT表达较前逐渐减弱,强阳性表达的细胞仅集中在细支气管的基底膜和少数新生肺泡内,在形态上类似基底细胞和Ⅱ型肺泡细胞。结论 在人胎肺发育中hTERT的表达可能是肺上皮干细胞或祖细胞维系其未分化状态和自我更新能力的重要标志;对保证支气管上皮和肺泡上皮细胞持续正常分化和再生,维持上皮组织的完整性起着重要的作用。     

靶向人端粒酶逆转录酶进行RNAi的逆转录病毒载体的构建

目的构建靶向性干扰人端粒酶逆转录酶表达的特异性小干扰RNA的真核表达载体。方法采用PCR法分别扩增增强型绿色荧光蛋白的DNA序列、U6启动子序列和靶向性干扰人端粒酶逆转录酶的特异性小干扰RNA对应的DNA序列,随后将与之相应的DNA序列片断依次克隆入真核表达载体pLXSN中,并通过限制性内切酶和测序对该重组表达载体进行鉴定。以荧光显微镜和利用流式细胞仪分析重组病毒表达EGFP蛋白的情况。以MTT方法检测初步分析重组病毒对人Hela细胞的RNAi效果。结果限制性内切酶酶切和测序结果均表明成功构建了靶向人端粒酶逆转录酶的小干扰RNA的逆转录病毒表达载体pLXSN/EGFP-U6-siTERT。以磷酸钙共转染法制备的重组逆转录病毒滴定了病毒滴度后,重组病毒感染人Hela细胞24h时用流式细胞仪测得EGFP表达的阳性率为24.1%。重组逆转录病毒感染人Hela细胞48h时,MTT实验测得细胞死亡率为53.2%。结论成功构建了针对人端粒酶逆转录酶小干扰RNA的逆转录病毒表达载体pLXSN/EGFP-U6-siTERT,并制备了重组逆转录病毒,初步观察了效应,为进一步利用小干扰RNA技术研究肿瘤的基因治疗奠定了基础。     

反义hTRT转染对SGC-7901胃癌细胞株形态学的影响

目的 探讨人端粒酶蛋白催化亚单位（hTRT）反义基因转染对SGC－7901胃癌细胞株形态学的影响;方法 采用脂质体法将hTRT正、反义真核表达载体导入SGC－7901胃癌细胞株中,从光镜、电镜水平观察转染细胞形态学的变化。结果 与未转染细胞相比,光镜下hTRT反义基因转染的SGC－7901胞体增大,胞浆丰富,分裂相减少,核浆比增大,异型性减小;透射电镜下反义基因转染细胞细胞器丰富,胞浆内出现分泌泡样结构及胞质内微囊;扫描电镜下反义基因转染细胞表面微绒毛增多,胞体周围可见大量颗粒样物质,根据AB／PAS染色结果,这些颗粒样物质有可能是其分泌的粘液颗粒。结论 hTRT反义基因有促进SGC－7901细胞分化的作用。     

人端粒酶逆转录酶在喉鳞状细胞癌中表达与c-myc的相关性

目的研究人端粒酶逆转录酶（hTERT）mRNA在喉鳞状细胞癌中和病理分级、临床参数的关系以及和c-myc的相关性。方法用原位杂交法检测40例喉鳞状细胞癌中hTERT mRNA的表达,用免疫组织化学法检测c—myc蛋白。结果喉正常组织、不典型增生、鳞癌组织中hTERT mRNA的阳性率分别为0％（0／10）、10％（1／10）、80％（32／40）,喉鳞癌中hTERT mRNA的阳性率明显高于正常组织（P〈0．05）和不典型增生（P〈0．05）,hTERT mRNA水平与c—myc的表达呈显著相关（r=0．5422,P〈0．01）。喉鳞癌中hTERT mRNA表达水平与病理分级不相关（P〉0．05）,与年龄、性别、TNM分期和淋巴结转移情况不相关（P〉0．05）。结论喉鳞癌中hTERT mRNA的阳性率明显高于正常组织和不典型增生,有助于喉部良恶性病变的鉴别,c-myc的过度表达可能是喉鳞癌端粒酶活性升高的一个重要机制。     

宫颈脱落细胞中人端粒酶RNA组分基因表达在宫颈病变中的临床价值

目的：研究宫颈脱落细胞中人端粒酶RNA组分（hTERC）基因的表达,探讨其在宫颈上皮内瘤变（CIN）及鳞状上皮细胞癌（SCC）筛查与诊断中的价值。方法：采用宫颈液基细胞学（LCT）、高危型人乳头瘤病毒DNA（HC-Ⅱ）检测及组织病理学的检测,并以组织病理学为金标准,分级盲法使用荧光原位杂交（FISH）方法检测人端粒酶RNA（hTERC）基因的表达。结果：hTERC基因扩增在细胞病理学HSIL及以上病变组扩增率显著升高（P〈0.05）。hTERC基因的扩增率与组织病理学级别、HC-Ⅱ阳性率均显著正相关（均P〈0.01）。HC-Ⅱ阳性率与细胞学、组织病理学级别均显著正相关（均P〈0.01）。结论：在宫颈病变组织中可见hTERC基因的表达、扩增与宫颈细胞学和组织学异常以及HPV病毒感染正相关,hTERC基因的扩增与否可能作为判断有无宫颈高度病变及估计预后的指标之一。