To investigate the level of telomerase activation (TA) and telomerase catalytic subunit (hEST 2) gene mRNA in patients with non small cell lung cancer, and determine whether they are associated with tumor cell apopt...To investigate the level of telomerase activation (TA) and telomerase catalytic subunit (hEST 2) gene mRNA in patients with non small cell lung cancer, and determine whether they are associated with tumor cell apoptosis, stage, and clinical outcome Methods Primary tumor specimens from 58 patients untreated with chemotherapy and 10 cases of histologically benign and adjacent lung tissue were analyzed TA and hEST 2 were measured by means of a modified telomerase repeat amplification protocol (TRAP) assay and in situ hybridization (ISH), respectively Terminal deoxynucleotidyl transferase (TdT) mediated biotin dUTP nick end labeling (TUNEL) was used to evaluate apoptotic cells Reverse transcription polymerase chain reaction (RT PCR) was used to detect bcl 2 mRNA expression Results TA and hEST 2 were detected in 45 (77 6%) and 43 (74 1%) of 58 tumor specimens, respectively, and not detected in specimens of adjacent and benign lung tissue One case expressed hEST 2 as a weak positive Statistically significant positive association was found between the level of TA and hEST 2( r =0 85, P =0 001) TA and hEST 2 were associated with tumor stage, but not associated with tumor grade, gender and patient age Positive rate of bcl 2 mRNA was 38 (65 5%) of 58 tumor specimens The mean apoptotic index in the bcl 2 positive group (9 5±1 3) was lower than that in the bcl 2 negative one (19 8±2 1, P <0.05), suggesting that apoptotic index may be inversely associated with bcl 2 expression ( r =-0 48, P =0 041) Bcl 2 expression in the TA and hEST 2 positive group (92 1% and 89 4%) was higher than that in the negative one (50 0% and 45 0%, P =0 043 and P =0 032, respectively) The apoptotic index was lower in the TA or hEST 2 positive group (8 2±1 4, 10 7±1 1) than in the negative one (20 5±1 6, 24 2±2 1, P <0 05) A statistically significant inverse association was found between TA or hEST 2 and apoptotic index ( r =-0 45, P =0 02 and r = -0 51, P =0 001, respectively) Positive correlation was also detected between TA or hEST 2 and bcl 2 expression ( r =0 86, P =0 01 and r =0 73, P =0 024, respectively) The level of hEST 2 mRNA and apoptotic index were associated with clinical outcome in a multivariate cox regression analysis Conclusions High TA and hEST 2 were frequently detected in primary non small cell lung cancer untreated with chemotherapy, having high bcl 2 expression and a low tumor cell apoptotic rate This suggests that both TA and hEST 2 are correlated with the deregulation of apoptosis hEST 2 and apoptotic index have prognostic significance in patients with non small cell lung cancer展开更多
Objective To investigate the expression of human telomerase catalytic subunit,hTERT, in human non-small cell lung cancer (NSCLC) and its correlations to c-myc gene.Methods hTERT and c-myc mRNA expressions were detecte...Objective To investigate the expression of human telomerase catalytic subunit,hTERT, in human non-small cell lung cancer (NSCLC) and its correlations to c-myc gene.Methods hTERT and c-myc mRNA expressions were detected by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). Statistical correlation analysis was made to estimate whether there was interrelation between them.Results Positive rate of hTERT expression in 51 surgically resected lung cancer specimens was 86.3%,significantly higher than that in adjacent non-neoplastic lung tissues and benign lesions,which were 14.3% and 27.3% respectively. No statistical significance was observed between the frequency of hTERT expression and histologic types,degree of differentiation,TNM stages,tumor size or lymph nodes metastases. Correlation analysis revealed that the expression of c-myc gene was significantly related to that of hTERT (correlation coefficient,r =0.633,P <0.001).Conclusions hTERT may be a useful tumor marker in diagnosing lung cancer. Significant correlation between the expression of hTERT and c-myc mRNA indicates that the activation and up-regulation of hTERT might be conferred by over-expression of c-myc gene.展开更多
Telomerase is a ribonucleoprotein that catalyzes telomere elongation through the addition of TTAGGG repeats in humans. Activation of telomerase is often associated with immortalization of human cells and cancer. Parti...Telomerase is a ribonucleoprotein that catalyzes telomere elongation through the addition of TTAGGG repeats in humans. Activation of telomerase is often associated with immortalization of human cells and cancer. Partially purification of telomerase was obtained by chromatography of HeLa cell extract on DEAE-Sepharose FF,Spermine-Sepharose CL-6B and Sephacryl S-300. To dissect the human telomerase enzyme mechanism, the optimal catalytic conditions of telomerase were measured and the human telomerase activity was reconstituted using a purified hTR RNA. The optimum pH is from 6^5 to 8^5, the optimum temperature is from 35℃ to 40℃, the optimum Mg 2+ concentration is from 1 mmol/L to 2^5 mmol/L,the apparent binding constant (K m app) of telomerase for the primer is about 8^6 nmol/L. Partially purified human telomerase extract was treated with MNase to remove endogenous telomerase RNA(hTR),the telomerase activity was inactivation. Telomerase activity was restored by incubating MNase-treated extract with purified hTR. These results might facilitate the development of the telomerase enzyme mechanism.展开更多
基金This research was supported by grants from the Bejjing Natural Science Faundation(Na 7992005)and Postdoctoral Foundation fram the Nati an al Committee of Education.
文摘To investigate the level of telomerase activation (TA) and telomerase catalytic subunit (hEST 2) gene mRNA in patients with non small cell lung cancer, and determine whether they are associated with tumor cell apoptosis, stage, and clinical outcome Methods Primary tumor specimens from 58 patients untreated with chemotherapy and 10 cases of histologically benign and adjacent lung tissue were analyzed TA and hEST 2 were measured by means of a modified telomerase repeat amplification protocol (TRAP) assay and in situ hybridization (ISH), respectively Terminal deoxynucleotidyl transferase (TdT) mediated biotin dUTP nick end labeling (TUNEL) was used to evaluate apoptotic cells Reverse transcription polymerase chain reaction (RT PCR) was used to detect bcl 2 mRNA expression Results TA and hEST 2 were detected in 45 (77 6%) and 43 (74 1%) of 58 tumor specimens, respectively, and not detected in specimens of adjacent and benign lung tissue One case expressed hEST 2 as a weak positive Statistically significant positive association was found between the level of TA and hEST 2( r =0 85, P =0 001) TA and hEST 2 were associated with tumor stage, but not associated with tumor grade, gender and patient age Positive rate of bcl 2 mRNA was 38 (65 5%) of 58 tumor specimens The mean apoptotic index in the bcl 2 positive group (9 5±1 3) was lower than that in the bcl 2 negative one (19 8±2 1, P <0.05), suggesting that apoptotic index may be inversely associated with bcl 2 expression ( r =-0 48, P =0 041) Bcl 2 expression in the TA and hEST 2 positive group (92 1% and 89 4%) was higher than that in the negative one (50 0% and 45 0%, P =0 043 and P =0 032, respectively) The apoptotic index was lower in the TA or hEST 2 positive group (8 2±1 4, 10 7±1 1) than in the negative one (20 5±1 6, 24 2±2 1, P <0 05) A statistically significant inverse association was found between TA or hEST 2 and apoptotic index ( r =-0 45, P =0 02 and r = -0 51, P =0 001, respectively) Positive correlation was also detected between TA or hEST 2 and bcl 2 expression ( r =0 86, P =0 01 and r =0 73, P =0 024, respectively) The level of hEST 2 mRNA and apoptotic index were associated with clinical outcome in a multivariate cox regression analysis Conclusions High TA and hEST 2 were frequently detected in primary non small cell lung cancer untreated with chemotherapy, having high bcl 2 expression and a low tumor cell apoptotic rate This suggests that both TA and hEST 2 are correlated with the deregulation of apoptosis hEST 2 and apoptotic index have prognostic significance in patients with non small cell lung cancer
文摘Objective To investigate the expression of human telomerase catalytic subunit,hTERT, in human non-small cell lung cancer (NSCLC) and its correlations to c-myc gene.Methods hTERT and c-myc mRNA expressions were detected by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). Statistical correlation analysis was made to estimate whether there was interrelation between them.Results Positive rate of hTERT expression in 51 surgically resected lung cancer specimens was 86.3%,significantly higher than that in adjacent non-neoplastic lung tissues and benign lesions,which were 14.3% and 27.3% respectively. No statistical significance was observed between the frequency of hTERT expression and histologic types,degree of differentiation,TNM stages,tumor size or lymph nodes metastases. Correlation analysis revealed that the expression of c-myc gene was significantly related to that of hTERT (correlation coefficient,r =0.633,P <0.001).Conclusions hTERT may be a useful tumor marker in diagnosing lung cancer. Significant correlation between the expression of hTERT and c-myc mRNA indicates that the activation and up-regulation of hTERT might be conferred by over-expression of c-myc gene.
基金国家自然科学基金 (No .30 0 70 85 9)资助项目国家重点基础研究规划资助项目 (No .G2 0 0 0 0 5 70 0 1)~~
文摘Telomerase is a ribonucleoprotein that catalyzes telomere elongation through the addition of TTAGGG repeats in humans. Activation of telomerase is often associated with immortalization of human cells and cancer. Partially purification of telomerase was obtained by chromatography of HeLa cell extract on DEAE-Sepharose FF,Spermine-Sepharose CL-6B and Sephacryl S-300. To dissect the human telomerase enzyme mechanism, the optimal catalytic conditions of telomerase were measured and the human telomerase activity was reconstituted using a purified hTR RNA. The optimum pH is from 6^5 to 8^5, the optimum temperature is from 35℃ to 40℃, the optimum Mg 2+ concentration is from 1 mmol/L to 2^5 mmol/L,the apparent binding constant (K m app) of telomerase for the primer is about 8^6 nmol/L. Partially purified human telomerase extract was treated with MNase to remove endogenous telomerase RNA(hTR),the telomerase activity was inactivation. Telomerase activity was restored by incubating MNase-treated extract with purified hTR. These results might facilitate the development of the telomerase enzyme mechanism.
