BACKGROUND:Most pancreatic carcinomas are clinically insensitive to chemotherapeutics.The exact mechanisms of their apoptosis and multiple drug resistance are obscure at present.This study was undertaken to explore th...BACKGROUND:Most pancreatic carcinomas are clinically insensitive to chemotherapeutics.The exact mechanisms of their apoptosis and multiple drug resistance are obscure at present.This study was undertaken to explore the influence of chemotherapy on anti-proliferation,apoptosis and the cell cycle,and lay a fundamental basis for further research into the apoptotic mechanisms and prevention of multiple drug resistance in pancreatic carcinoma. METHODS:The human pancreatic carcinoma cell line BxPC-3 was cultured in vitro.The growth inhibition rate,cell cycle and apoptotic rate of cells treated with 5-fluorouracil（5-FU）,sulfasalazine alone or a combination at different concentrations were evaluated with the MTT method and flow cytometry.Phase-contrast microscopy was used to observe morphological changes in the cells treated with 5-FU,sulfasalazine or both for 24 hours. RESULTS:The growth inhibition rate of the BxPC-3 cells treated with 5-FU and sulfasalazine significantly increased in a time-and dose-dependent manner.The growth inhibition rate of the cells treated with 5-FU gradually increased,but decreased at different concentrations of sulfasalazine for a prolonged period.The apoptotic rate of the BxPC-3 cells induced by sulfasalazine（200 mg/L）, 5-FU（100 mg/L）or both for 12 hours were（2.68±0.36）%, （6.59±0.90）%,and（10.52±0.55）%,respectively,compared with the corresponding control values were（3.17±0.08）%, （1.50±0.06）%,and（4.08±0.31）%[(t=2.33（P】0.05）,9.78 and 17.56（P【0.01）].It increased to（7.63±0.68）%,（40.43± 1.79）%,and（64.69±0.82）%for 48 hours,in comparison with the control that was（29.20±2.18）%,（5.61±0.13）%,and（12.02±0.52）%[t=17.06,33.66 and 94.51（P【0.01）]. The apoptotic rate,proportion of cells in S-phase and proliferative index rose after use of 5-FU（12.5,25,50,75, and 100 mg/L）alone for 24 hours.However,the apoptotic rate at augmented concentrations of sulfasalazine for 24 hours slowly increased from（1.47±0.08）%to（3.45± 0.28）%,the proportion of cells in G0/G1-phase increased from（35.13±0.32）%to（54.32±1.45）%,the proportion of cells in S-phase decreased from（45.37±1.48）%to（16.67± 2.73）%,and the proliferative index gradually lowered.The proportion of G0/G1-phase cells treated by 5-FU（100 mg/L） and sulfasalazine（200 mg/L）increased from（43.31±1.52）% （12 hours）to（85.05±0.24）%（48 hours）compared with the corresponding controls[t=7.93（12 hours）,21.30（48 hours）,P【0.01],and the proportion of cells in S-phase decreased from（11.63±1.11）%（12 hours）to（4.47±0.68）% （48 hours）in contrast to the controls[t=37.68（12 hours）, 8.60（48 hours）,P【0.01].Most cells after the combined use of the two agents for 24 hours displayed pyknosis and oval shape by phase-contrast microscopy.The cells treated with 5-FU（100 mg/L）for 24 hours were pyknotic and oval shaped.A few of cells in the group treated with sulfasalazine（200 mg/L）were pyknotic at 24 hours. CONCLUSIONS:Sulfasalazine may enhance the inhibitory proliferation and apoptosis effect on BxPC-3 cells induced by 5-FU,which is closely related to synergistically the cell cycle arrested in G<sub>0</sub>/G<sub>1</sub>-phase.展开更多
目的探讨甲基转移酶5(methyltransferase-like 5,METTL5)在三阴乳腺癌(triple-negative breast cancer,TNBC)中的作用和潜在机制。方法采用免疫组织化学方法和Western blot检测TNBC肿瘤组织和细胞系中METTL5的表达情况。用靶向METTL5的s...目的探讨甲基转移酶5(methyltransferase-like 5,METTL5)在三阴乳腺癌(triple-negative breast cancer,TNBC)中的作用和潜在机制。方法采用免疫组织化学方法和Western blot检测TNBC肿瘤组织和细胞系中METTL5的表达情况。用靶向METTL5的shRNA(shRNA-METTL5)转染TNBC细胞后,用CCK-8、集落形成、伤口愈合以及Transwell实验分别检测细胞增殖活性、迁移与侵袭,Western blot检测Wnt/β-catenin信号关键蛋白的表达。构建异种移植瘤模型,验证敲降METTL5对TNBC细胞在体内生长以及Wnt/β-catenin信号活性的影响。结果METTL5在TNBC肿瘤组织和细胞系中表达上调(P<0.01)。敲降METTL5可抑制TNBC细胞的增殖、迁移和侵袭并降低了Wnt/β-catenin信号分子β-catenin、细胞周期蛋白(Cyclin)D1、基质金属蛋白酶(MMP)-2和MMP-7的表达(均P<0.01)。体内实验显示,敲降METTL5减缓了移植瘤生长和Wnt/β-catenin信号活性。结论敲降METTL5能抑制TNBC细胞的增殖、迁移与侵袭,其作用可能与抑制Wnt/β-catenin信号通路有关。展开更多
目的:研究溶质载体家族6成员9(solute carrier family 6 member 9,SLC6A9)表达对结直肠癌细胞增殖、迁移和5-氟尿嘧啶(5-fluorouracil,5-FU)药物敏感性的影响。方法:TCGA数据库分析、实时荧光定量PCR和Western blot分析检测SLC6A9在结...目的:研究溶质载体家族6成员9(solute carrier family 6 member 9,SLC6A9)表达对结直肠癌细胞增殖、迁移和5-氟尿嘧啶(5-fluorouracil,5-FU)药物敏感性的影响。方法:TCGA数据库分析、实时荧光定量PCR和Western blot分析检测SLC6A9在结肠癌组织、正常结肠细胞系(NCM460)和结直肠癌细胞系(SW620、HCT116、HT29、Lovo和SW480)中的表达。将SCL6A9过表达质粒及阴性对照(SLC6A9 OE、Vector)转染HT29细胞,将SCL6A9小干扰RNA及阴性对照(SLC6A9 siRNA1#、siRNA2#和Scramble)转染SW620细胞。划痕愈合实验和Transwell实验检测各组细胞的迁移、侵袭能力。Western blot和细胞免疫荧光检测EMT相关蛋白E-cadherin、Vimentin的表达水平。利用CCK-8法和构建裸鼠移植瘤模型检测SLC6A9过表达对结直肠癌细胞5-FU药物敏感性的影响。结果:与正常结肠组织和NCM460细胞相比,SLC6A9在结肠癌组织和结直肠癌细胞系中低表达(均P<0.05)。SLC6A9过表达引起E-cadherin蛋白表达增加,Vimentin蛋白水平降低,抑制结直肠癌细胞的迁移、侵袭(P<0.05)。SLC6A9低表达引起E-cadherin蛋白表达降低,Vimentin蛋白水平增加,促进结直肠癌细胞的迁移、侵袭能力(P<0.05)。SLC6A9过表达提高了5-FU的药物敏感性,并使肿瘤生长缓慢,质量减轻(P<0.05)。而SLC6A9低表达降低了5-FU的药物敏感性(P<0.05)。结论:SLC6A9过表达能够抑制结直肠癌细胞的迁移、侵袭和EMT进程,并增强5-FU对结直肠癌细胞的药物敏感性。展开更多
文摘BACKGROUND:Most pancreatic carcinomas are clinically insensitive to chemotherapeutics.The exact mechanisms of their apoptosis and multiple drug resistance are obscure at present.This study was undertaken to explore the influence of chemotherapy on anti-proliferation,apoptosis and the cell cycle,and lay a fundamental basis for further research into the apoptotic mechanisms and prevention of multiple drug resistance in pancreatic carcinoma. METHODS:The human pancreatic carcinoma cell line BxPC-3 was cultured in vitro.The growth inhibition rate,cell cycle and apoptotic rate of cells treated with 5-fluorouracil（5-FU）,sulfasalazine alone or a combination at different concentrations were evaluated with the MTT method and flow cytometry.Phase-contrast microscopy was used to observe morphological changes in the cells treated with 5-FU,sulfasalazine or both for 24 hours. RESULTS:The growth inhibition rate of the BxPC-3 cells treated with 5-FU and sulfasalazine significantly increased in a time-and dose-dependent manner.The growth inhibition rate of the cells treated with 5-FU gradually increased,but decreased at different concentrations of sulfasalazine for a prolonged period.The apoptotic rate of the BxPC-3 cells induced by sulfasalazine（200 mg/L）, 5-FU（100 mg/L）or both for 12 hours were（2.68±0.36）%, （6.59±0.90）%,and（10.52±0.55）%,respectively,compared with the corresponding control values were（3.17±0.08）%, （1.50±0.06）%,and（4.08±0.31）%[(t=2.33（P】0.05）,9.78 and 17.56（P【0.01）].It increased to（7.63±0.68）%,（40.43± 1.79）%,and（64.69±0.82）%for 48 hours,in comparison with the control that was（29.20±2.18）%,（5.61±0.13）%,and（12.02±0.52）%[t=17.06,33.66 and 94.51（P【0.01）]. The apoptotic rate,proportion of cells in S-phase and proliferative index rose after use of 5-FU（12.5,25,50,75, and 100 mg/L）alone for 24 hours.However,the apoptotic rate at augmented concentrations of sulfasalazine for 24 hours slowly increased from（1.47±0.08）%to（3.45± 0.28）%,the proportion of cells in G0/G1-phase increased from（35.13±0.32）%to（54.32±1.45）%,the proportion of cells in S-phase decreased from（45.37±1.48）%to（16.67± 2.73）%,and the proliferative index gradually lowered.The proportion of G0/G1-phase cells treated by 5-FU（100 mg/L） and sulfasalazine（200 mg/L）increased from（43.31±1.52）% （12 hours）to（85.05±0.24）%（48 hours）compared with the corresponding controls[t=7.93（12 hours）,21.30（48 hours）,P【0.01],and the proportion of cells in S-phase decreased from（11.63±1.11）%（12 hours）to（4.47±0.68）% （48 hours）in contrast to the controls[t=37.68（12 hours）, 8.60（48 hours）,P【0.01].Most cells after the combined use of the two agents for 24 hours displayed pyknosis and oval shape by phase-contrast microscopy.The cells treated with 5-FU（100 mg/L）for 24 hours were pyknotic and oval shaped.A few of cells in the group treated with sulfasalazine（200 mg/L）were pyknotic at 24 hours. CONCLUSIONS:Sulfasalazine may enhance the inhibitory proliferation and apoptosis effect on BxPC-3 cells induced by 5-FU,which is closely related to synergistically the cell cycle arrested in G<sub>0</sub>/G<sub>1</sub>-phase.
文摘目的:研究溶质载体家族6成员9(solute carrier family 6 member 9,SLC6A9)表达对结直肠癌细胞增殖、迁移和5-氟尿嘧啶(5-fluorouracil,5-FU)药物敏感性的影响。方法:TCGA数据库分析、实时荧光定量PCR和Western blot分析检测SLC6A9在结肠癌组织、正常结肠细胞系(NCM460)和结直肠癌细胞系(SW620、HCT116、HT29、Lovo和SW480)中的表达。将SCL6A9过表达质粒及阴性对照(SLC6A9 OE、Vector)转染HT29细胞,将SCL6A9小干扰RNA及阴性对照(SLC6A9 siRNA1#、siRNA2#和Scramble)转染SW620细胞。划痕愈合实验和Transwell实验检测各组细胞的迁移、侵袭能力。Western blot和细胞免疫荧光检测EMT相关蛋白E-cadherin、Vimentin的表达水平。利用CCK-8法和构建裸鼠移植瘤模型检测SLC6A9过表达对结直肠癌细胞5-FU药物敏感性的影响。结果:与正常结肠组织和NCM460细胞相比,SLC6A9在结肠癌组织和结直肠癌细胞系中低表达(均P<0.05)。SLC6A9过表达引起E-cadherin蛋白表达增加,Vimentin蛋白水平降低,抑制结直肠癌细胞的迁移、侵袭(P<0.05)。SLC6A9低表达引起E-cadherin蛋白表达降低,Vimentin蛋白水平增加,促进结直肠癌细胞的迁移、侵袭能力(P<0.05)。SLC6A9过表达提高了5-FU的药物敏感性,并使肿瘤生长缓慢,质量减轻(P<0.05)。而SLC6A9低表达降低了5-FU的药物敏感性(P<0.05)。结论:SLC6A9过表达能够抑制结直肠癌细胞的迁移、侵袭和EMT进程,并增强5-FU对结直肠癌细胞的药物敏感性。
