Objective:To investigate the effects and mechanisms of tumor suppressor gene PTEN on the induction of anoikis of human bladder transitional carcinoma cells BIU-87.Methods:BIU-87 cells were transfected with GFP plasmid...Objective:To investigate the effects and mechanisms of tumor suppressor gene PTEN on the induction of anoikis of human bladder transitional carcinoma cells BIU-87.Methods:BIU-87 cells were transfected with GFP plasmids containing wild-type PTEN or phosphatase inactivating mutant PTEN(C124A-PTEN)in vitro.The PTEN expression and the phosphorylation levels of focal adhesion kinase(FAK)and protein kinase B(PKB/Akt)were detected by Western blotting.Flow cytometry assay and laser scanning confocal microscopy were used to analyze apoptosis in adherent and non-adherent cells.Results: Compared with the control group,PTEN expression in the cells transfected with wild-type PTEN increased to 210%–260%, while the phosphorylation level of FAK and Akt decreased 59%(P<0.01)and 89%(P<0.01),respectively.And the anoikis percentage increased from 8.32±0.57%to 37.62±2.12%.In the cells transfected with C124A-PTEN,neither the phos- phorylation of FAK and Akt nor the anoikis percentage had obviously changed,although the PTEN expression enhanced remarkably in comparison with the control.Conclusion:Through its phosphatase activity,tumor suppressor gene PTEN can suppress the phosphorylation of FAK and Akt,and induce anoikis in human bladder transitional carcinoma cells BIU-87.展开更多
Objective： To study the relationship of the telomerase activity and the p53 gene mutation in cardiac cancer. Methods： Telomerase activity and the p53 gene mutation were detected in 46 case of cardiac cancer, peri-ca...Objective： To study the relationship of the telomerase activity and the p53 gene mutation in cardiac cancer. Methods： Telomerase activity and the p53 gene mutation were detected in 46 case of cardiac cancer, peri-cancerous and 30 case of normal mucosa by TRAP-ELISA and PCR-SSCP. Results： The rate of expression of telomerase activity in cardiac cancer, peri-cancerous and normal mucosa were 82.61% （38/46）, 43.48% （20/46） and 13.33% （4/30） respectively. The rate of Exon5→,8 of p53 gene mutation were 39,13% （18/46）, 4.35% （2/46） and 0.00% respectively. There was significant difference between group cancer and without cancer （P 〈 0.01）. Mean of A^- value of telomerase is 1.89：1：0.41 in cancer group and were 1.49：1： 0.43, 0.54：1：0.45 respectively in peri-canvcerous and normal mucosa, there were significant differences in cancer group and group of without cancer （P 〈 0.05）. The rate of p53 gene mutations in group of expression of telomerase activity was 44.74% （17/38）, and 12.50% （1/8） in without expression of telomerase activity. There were significant differences between the two groups. Conclusion： The rate of expression of telomerase activity and mean of A^- value of telomerase in cardiac cancer were obviously higher than without cancer, which indicating telomerase activity was closely related with the occurrence of cardiac cancer. P53 gene mutation in cardiac cancer were higher than the tissue of without cancer, and the rate of p53 gene mutation in telomerase activity were obviously higher than the group of without cancer. This shows the p53 gene mutation can loss of function of suppressing cancer and prompt telomerase activity and cause the cardiac cancer.展开更多
AIM： Gene expression profiling provides an unique opportunity to gain insight into the development of different types of gastric cancer. Tumor sample heterogeneity is thought to decrease the sensitivity and tumor spe...AIM： Gene expression profiling provides an unique opportunity to gain insight into the development of different types of gastric cancer. Tumor sample heterogeneity is thought to decrease the sensitivity and tumor specificity of microarray analysis. Thus, microdissection and preamplification of RNA is frequently performed. However, this technique may also induce considerable changes to the expression profile. To assess the effect of gastric tumor heterogeneity on expression profiling results, we measured the variation in gene expression within the same gasbic cancer sample by performing a gene chip analysis with two RNA preparations extracted from the same tumor specimen. METHODS： Tumor samples from six intestinal T2 gastric tumors were dissected under liquid nitrogen and RNA was prepared from two separate tumor fragments. Each extraction was individually processed and hybridized to an Affymetrix U133A gene chip covering approximately 18 000 human gene transcripts. Expression profiles were analyzed using Microarray Suite 5.0 （Affymetrix） and GeneSpring 6.0 （Silicon Genetics）. RESULTS： All gastric cancers showed little variance in expression profiles between different regions of the same tumor sample. In this case, gene chips displayed mean pair wise correlation coefficients of 0.94±0.02 （mean±SD）, compared to values of 0.61±0.1 for different tumor samples. Expression of the variance between the two expression profiles as a percentage of “total change” （Affymetrix） revealed a remarkably low average value of 1.18±0.78 for comparing fragments of the same tumor sample. In contrast, comparison of fragments from different tumors revealed a percentage of 24.4±4.5. CONCLUSION： Our study indicates a low degree of expression profile variability within gastric tumor samples isolated from one patient. These data suggest that tumor tissue heterogeneity is not a dominant source of error for microarray analysis of larger tumor samples, making total RNA extraction an appropriate strategy for performing gene chip expression profiling of gastric cancer.展开更多
AIM： To evaluate bacterial cytosine deaminase （bCD） mutant D314A and 5-fluorocytosine （5-FC） for treatment of colon cancer in a mouse model. METHODS： Recombinant lentivirus vectors that contained wild-type bCD g...AIM： To evaluate bacterial cytosine deaminase （bCD） mutant D314A and 5-fluorocytosine （5-FC） for treatment of colon cancer in a mouse model. METHODS： Recombinant lentivirus vectors that contained wild-type bCD gene （bCDwt）, and bCD mutant D314A gene （bCD-D314A） with green fluorescence protein gene were constructed and used to infect hu- man colon carcinoma LoVo cells, to generate stable transfected cells, LoVo/null, LoVo/bCDwt or LoVo/bCD- D314A. These were injected subcutaneously into Balb/c nude mice to establish xenograft models. Two weeks post-LoVo cell inoculation, PBS or 5-FC （500 mg/kg） was administered by intraperitoneal （i.p.） injection once daily for 14 d. On the day after LoVo cell injection, mice were monitored daily for tumor volume and survival. RESULTS： Sequence analyses confirmed the construction of recombinant lentiviral plasmids that contained bCDwt or bCD-D314A. The lentiviral vector had high elficacy for gene delivery, and RT-PCR showed that bCDwt or bCD-D314A gene was transferred to LoVo cells. Among these treatment groups, gene delivery or 5-FC administration alone had no effect on tumor growth. However, bCDwt/5-FC or bCD-D314A/5-FC treatment inhibited tumor growth and prolonged survival of mice significantly （P 〈 0.05）. Importantly, the tumor volume in the bCD-D314A/5-FC-treated group was lower than that in the bCDwt/5-FC group （P 〈 0.05）, and bCD- D314A plus 5-FC significantly prolonged survival of mice in comparison with bCDwt plus 5-FC （P 〈 0.05）. CONCLUSION： The bCD mutant D314A enhanced significantly antitumor activity in human colon cancer xenograft models, which provides a promising approach for human colon carcinoma therapy.展开更多
基金a grant from the National Natural Science Foundation ofChina(No.30271300).
文摘Objective:To investigate the effects and mechanisms of tumor suppressor gene PTEN on the induction of anoikis of human bladder transitional carcinoma cells BIU-87.Methods:BIU-87 cells were transfected with GFP plasmids containing wild-type PTEN or phosphatase inactivating mutant PTEN(C124A-PTEN)in vitro.The PTEN expression and the phosphorylation levels of focal adhesion kinase(FAK)and protein kinase B(PKB/Akt)were detected by Western blotting.Flow cytometry assay and laser scanning confocal microscopy were used to analyze apoptosis in adherent and non-adherent cells.Results: Compared with the control group,PTEN expression in the cells transfected with wild-type PTEN increased to 210%–260%, while the phosphorylation level of FAK and Akt decreased 59%(P<0.01)and 89%(P<0.01),respectively.And the anoikis percentage increased from 8.32±0.57%to 37.62±2.12%.In the cells transfected with C124A-PTEN,neither the phos- phorylation of FAK and Akt nor the anoikis percentage had obviously changed,although the PTEN expression enhanced remarkably in comparison with the control.Conclusion:Through its phosphatase activity,tumor suppressor gene PTEN can suppress the phosphorylation of FAK and Akt,and induce anoikis in human bladder transitional carcinoma cells BIU-87.
文摘Objective： To study the relationship of the telomerase activity and the p53 gene mutation in cardiac cancer. Methods： Telomerase activity and the p53 gene mutation were detected in 46 case of cardiac cancer, peri-cancerous and 30 case of normal mucosa by TRAP-ELISA and PCR-SSCP. Results： The rate of expression of telomerase activity in cardiac cancer, peri-cancerous and normal mucosa were 82.61% （38/46）, 43.48% （20/46） and 13.33% （4/30） respectively. The rate of Exon5→,8 of p53 gene mutation were 39,13% （18/46）, 4.35% （2/46） and 0.00% respectively. There was significant difference between group cancer and without cancer （P 〈 0.01）. Mean of A^- value of telomerase is 1.89：1：0.41 in cancer group and were 1.49：1： 0.43, 0.54：1：0.45 respectively in peri-canvcerous and normal mucosa, there were significant differences in cancer group and group of without cancer （P 〈 0.05）. The rate of p53 gene mutations in group of expression of telomerase activity was 44.74% （17/38）, and 12.50% （1/8） in without expression of telomerase activity. There were significant differences between the two groups. Conclusion： The rate of expression of telomerase activity and mean of A^- value of telomerase in cardiac cancer were obviously higher than without cancer, which indicating telomerase activity was closely related with the occurrence of cardiac cancer. P53 gene mutation in cardiac cancer were higher than the tissue of without cancer, and the rate of p53 gene mutation in telomerase activity were obviously higher than the group of without cancer. This shows the p53 gene mutation can loss of function of suppressing cancer and prompt telomerase activity and cause the cardiac cancer.
基金Supported by a MeDDrive grant From the University of Dresden 2003by a grant from the Dr. Mildred Scheel Stiftung No. 70-2923
文摘AIM： Gene expression profiling provides an unique opportunity to gain insight into the development of different types of gastric cancer. Tumor sample heterogeneity is thought to decrease the sensitivity and tumor specificity of microarray analysis. Thus, microdissection and preamplification of RNA is frequently performed. However, this technique may also induce considerable changes to the expression profile. To assess the effect of gastric tumor heterogeneity on expression profiling results, we measured the variation in gene expression within the same gasbic cancer sample by performing a gene chip analysis with two RNA preparations extracted from the same tumor specimen. METHODS： Tumor samples from six intestinal T2 gastric tumors were dissected under liquid nitrogen and RNA was prepared from two separate tumor fragments. Each extraction was individually processed and hybridized to an Affymetrix U133A gene chip covering approximately 18 000 human gene transcripts. Expression profiles were analyzed using Microarray Suite 5.0 （Affymetrix） and GeneSpring 6.0 （Silicon Genetics）. RESULTS： All gastric cancers showed little variance in expression profiles between different regions of the same tumor sample. In this case, gene chips displayed mean pair wise correlation coefficients of 0.94±0.02 （mean±SD）, compared to values of 0.61±0.1 for different tumor samples. Expression of the variance between the two expression profiles as a percentage of “total change” （Affymetrix） revealed a remarkably low average value of 1.18±0.78 for comparing fragments of the same tumor sample. In contrast, comparison of fragments from different tumors revealed a percentage of 24.4±4.5. CONCLUSION： Our study indicates a low degree of expression profile variability within gastric tumor samples isolated from one patient. These data suggest that tumor tissue heterogeneity is not a dominant source of error for microarray analysis of larger tumor samples, making total RNA extraction an appropriate strategy for performing gene chip expression profiling of gastric cancer.
文摘AIM： To evaluate bacterial cytosine deaminase （bCD） mutant D314A and 5-fluorocytosine （5-FC） for treatment of colon cancer in a mouse model. METHODS： Recombinant lentivirus vectors that contained wild-type bCD gene （bCDwt）, and bCD mutant D314A gene （bCD-D314A） with green fluorescence protein gene were constructed and used to infect hu- man colon carcinoma LoVo cells, to generate stable transfected cells, LoVo/null, LoVo/bCDwt or LoVo/bCD- D314A. These were injected subcutaneously into Balb/c nude mice to establish xenograft models. Two weeks post-LoVo cell inoculation, PBS or 5-FC （500 mg/kg） was administered by intraperitoneal （i.p.） injection once daily for 14 d. On the day after LoVo cell injection, mice were monitored daily for tumor volume and survival. RESULTS： Sequence analyses confirmed the construction of recombinant lentiviral plasmids that contained bCDwt or bCD-D314A. The lentiviral vector had high elficacy for gene delivery, and RT-PCR showed that bCDwt or bCD-D314A gene was transferred to LoVo cells. Among these treatment groups, gene delivery or 5-FC administration alone had no effect on tumor growth. However, bCDwt/5-FC or bCD-D314A/5-FC treatment inhibited tumor growth and prolonged survival of mice significantly （P 〈 0.05）. Importantly, the tumor volume in the bCD-D314A/5-FC-treated group was lower than that in the bCDwt/5-FC group （P 〈 0.05）, and bCD- D314A plus 5-FC significantly prolonged survival of mice in comparison with bCDwt plus 5-FC （P 〈 0.05）. CONCLUSION： The bCD mutant D314A enhanced significantly antitumor activity in human colon cancer xenograft models, which provides a promising approach for human colon carcinoma therapy.
