摘要
AIM: To investigate the expression of Hu R in pancreatic ductal adenocarcinoma(PDA) and to assess the effects of Hu R silencing on the expression of cyclooxygenase-2(COX-2) and heme oxygenase-1(HO-1) and the in vitro response to gemcitabine(GEM) treatment in pancreatic cell lines.METHODS: We compared the expression of Hu R,COX-2,and HO-1 in PDA and normal pancreatic tissueusing quantitative reverse transcription polymerase chain reaction(q RT-PCR) and western blot. In addition,the Hu R,COX-2 and HO-1 were analyzed in four types of cancer cell lines(Mia Paca2,Su.86.86,Capan-1,and Capan-2) with and without GEM treatment. Immunocytofluorescence analysis was used to investigate Hu R localization in cells. Cell viability and response to GEM after Hu R silencing were determined with the 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide test and the crystal violet clonogenic assay,respectively. To measure apoptosis,activation of caspases 3/7 was evaluated using immunofluorescence. RESULTS: In PDA tissue obtained from patients not treated with GEM,Hu R m RNA expression was 3.2 times lower(P < 0.05) and COX-2 and HO-1 m RNAexpression was 2.3-fold and 7.2-fold higher(P < 0.05),respectively,than normal pancreatic tissue(from organ donor). q RT-PCR analysis showed that Hu R,COX-2,and HO-1 m RNA were overexpressed in all cancer cell lines treated with the half maximal inhibitory concentration(IC50) dose of GEM compared with control cells(P < 0.05). Western blot analysis revealed that COX-2 and HO-1 levels were significantly decreased in cancer cells after Hu R silencing. Furthermore,HuR silencing in creased the response to GEM treatmentand decreased cell viability by 11.6%-53.7% compared to control cell lines. Caspases 3 and 7 were activated after Hu R silencing and GEM treatment in all pancreatic cancer cell lines. In comparison,treatment with GEM alone did not activate caspases 3 and 7 in the same cell lines. CONCLUSION: Hu R mediated post-transcriptional upregulation of COX-2 and HO-1 expression after GEM treatment in pancreatic cancer cells. Hu R silencing significantly increased the effectiveness of GEM treatment in vitro.
AIM: To investigate the expression of Hu R in pancreatic ductal adenocarcinoma(PDA) and to assess the effects of Hu R silencing on the expression of cyclooxygenase-2(COX-2) and heme oxygenase-1(HO-1) and the in vitro response to gemcitabine(GEM) treatment in pancreatic cell lines.METHODS: We compared the expression of Hu R,COX-2,and HO-1 in PDA and normal pancreatic tissueusing quantitative reverse transcription polymerase chain reaction(q RT-PCR) and western blot. In addition,the Hu R,COX-2 and HO-1 were analyzed in four types of cancer cell lines(Mia Paca2,Su.86.86,Capan-1,and Capan-2) with and without GEM treatment. Immunocytofluorescence analysis was used to investigate Hu R localization in cells. Cell viability and response to GEM after Hu R silencing were determined with the 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide test and the crystal violet clonogenic assay,respectively. To measure apoptosis,activation of caspases 3/7 was evaluated using immunofluorescence. RESULTS: In PDA tissue obtained from patients not treated with GEM,Hu R m RNA expression was 3.2 times lower(P < 0.05) and COX-2 and HO-1 m RNAexpression was 2.3-fold and 7.2-fold higher(P < 0.05),respectively,than normal pancreatic tissue(from organ donor). q RT-PCR analysis showed that Hu R,COX-2,and HO-1 m RNA were overexpressed in all cancer cell lines treated with the half maximal inhibitory concentration(IC50) dose of GEM compared with control cells(P < 0.05). Western blot analysis revealed that COX-2 and HO-1 levels were significantly decreased in cancer cells after Hu R silencing. Furthermore,HuR silencing in creased the response to GEM treatmentand decreased cell viability by 11.6%-53.7% compared to control cell lines. Caspases 3 and 7 were activated after Hu R silencing and GEM treatment in all pancreatic cancer cell lines. In comparison,treatment with GEM alone did not activate caspases 3 and 7 in the same cell lines. CONCLUSION: Hu R mediated post-transcriptional upregulation of COX-2 and HO-1 expression after GEM treatment in pancreatic cancer cells. Hu R silencing significantly increased the effectiveness of GEM treatment in vitro.
基金
Supported by The European Social Fund under the Global Grant measure
