BACKGROUND The poor prognosis and rising incidence of esophageal cancer highlight the need for improved therapeutics that are essential prior to treatment.LCL161 is an SMAC(second mitochondrial activator of caspases)m...BACKGROUND The poor prognosis and rising incidence of esophageal cancer highlight the need for improved therapeutics that are essential prior to treatment.LCL161 is an SMAC(second mitochondrial activator of caspases)mimic and inhibitor of apoptosis protein(IAP)antagonist which exhibits anti-tumor effects and improves the chemical sensitivity of many cancers.AIM To ascertain the effects and mechanisms of the SMAC analog LCL161 on esophageal cancer cells.METHODS MTT assay and TUNEL assay were used to detect cell proliferation and apoptosis,respectively.Western blot analysis was used to study the molecular mechanisms of LCL161-induced death of ECA109 cells.RESULTS LCL161 decreased ECA109 cell proliferation in dose-and time-dependent manner and induced apoptosis of ECA109 cells in a dose-dependent manner.Also,LCL161 induced a significant decrease in the expression of the XIAP and significant increase in the expression of Caspase-3.In addition,Bax increased significantly with increasing concentrations of LCL161,and the relative expression of Bax was significantly different between groups.CONCLUSION These findings support the hypothesis that LCL161 can inhibit proliferation and induce apoptosis in esophageal cancer cells by regulating the expression of IAP family members,suggesting that it has potential to be an effective treatment for esophageal squamous cell carcinoma.展开更多
Objective： To investigate the anti-tumor activity and molecular mechanism of Tonglian Decoction （通莲汤, TLD） on esophageal carcinoma Eca109 cells. Methods： Ecal09 cells were treated with TLD and its separated for...Objective： To investigate the anti-tumor activity and molecular mechanism of Tonglian Decoction （通莲汤, TLD） on esophageal carcinoma Eca109 cells. Methods： Ecal09 cells were treated with TLD and its separated formulae, including the clearing-heat and detoxification formula （Q）, activating-blood and promoting-qi formula （H） and nourishing-yin and blood formula （Z）. Cell proliferation was measured using the 3-（4,5-dimethyl-2- thiazolyl）-2,5-diphenyl-2-H-tetrazolium bromide assay, cell morphology was observed using a microscope, the cell cycle was measured using flow cytometry and the activity of the nuclear factor-kappa B （NF- K B） signal pathway was detected by Western blot. Results： The half maximal inhibitory concentrations of TLD, Q and H were 386, 771 and 729 rag/L, respectively. TLD, Q and H significantly inhibited cell proliferation, with 69.43%, 60.84% and 61.90% of treated cells in the G1 phase of the cell cycle. The percentage of cells in S phase increased significantly after treatment with TLD, Q, and H compared with the control group （P〈0.05）, and TLD showed the strongest effect. Z had no influence on the cell cycle compared with the control group （P〉0.05）. Western blot detection indicated slight differences in the inhibition of the NF- K B pathway by the different formulae. TLD formula strongly inhibited IKK β, NF- K B, interleukin-6 and tumor necrosis factor-α expression compared with the control group. Conclusions： TLD inhibited Eca109 cell proliferation by arresting cells in S phase. The possible mechanism might be related to inhibiting the NF- K B transduction cascade. The combination of the herbs found in the three separate formulae, H, Q and Z, work synergistically in TLD to produce the inhibitory effects of TLD treatment on Eca109 proliferation.展开更多
AIM: To study the role of P38 kinase in esophageal cancer cell apoptosis induced by genotoxin, cisplatin and the unfolded protein response (UPR) inducer, dithiothreitol(DTT).METHODS: Esophageal carcinoma cell line Eca...AIM: To study the role of P38 kinase in esophageal cancer cell apoptosis induced by genotoxin, cisplatin and the unfolded protein response (UPR) inducer, dithiothreitol(DTT).METHODS: Esophageal carcinoma cell line Eca109 was cultured in RPMI 1640 medium to 70% confluency and treated with either cisplatin, DTT, or cisplatin plus DTT in the presence or absence of P38 inhibitor, SB203580. The untreated cells served as the control. The esophageal carcinoma cell apoptosis was detected by agarose gel DNA ladder analysis and quantified by flow cytometry.The P38 phosphorylation was detected by immunohis-tochemistry using antibodies specific to phosphorylated P38 protein.RESULTS: (1) Both cisplatin and DTT induced apoptosis in the esophageal cancer cell line Eca109 as shown by DNA ladder formation; (2) As detected by antibodies specific for the phosphorylated P38 protein (p-P38), both cisplatin and DTT treatments activated the stress-activated enzyme, MAP kinase P38. The number of positive cells was about 50% for the treatment groups, comparing to that of 10% for untreated group. DTT treatment, but not cisplatin treatment, induces nuclear localization of p-P38;(3) As measured by flow cytometry, inhibition of P38 activity by SB203580 blocks DFT- and cisplatin-induced apoptosis.The rates for DTT, cisplatin, and DTT plus cisplatin-induced apoptosis were 16.8%, 17.1%, and 21.4%, respectively.Addition of the SB compound during the incubation reduced the apoptotic rate to about 7.6% for all the treatment groups, suggesting that P38 activation is essential for cisplatin- and DTT-induced apoptosis in Eca109 cells.CONCLUSION: (1) Both DTT and cisplatin were able to induce apoptosis in esophageal cancer cell line Eca109;(2) P38 MAP kinase is essential for DTT- and cisplatin-induced apoptosis in Eca109 cells; (3) P38 activation may be the common signaling component relaying the multiple upstream signaling events to the downstream cell death program.展开更多
The expression and properties of alkaline phosphatase (ALP) in Eca109 cells, a cell line derived fromhuman esophageal cancer were studied with specific inhibition assay and polyacrylamide gel electrophoresis.The resul...The expression and properties of alkaline phosphatase (ALP) in Eca109 cells, a cell line derived fromhuman esophageal cancer were studied with specific inhibition assay and polyacrylamide gel electrophoresis.The results showed that ALP of Eca109 cells was heat stable and was strongly inhibited by L-pheuylalanine, but slightly inhibited by urea. Preduisolone could causedramatic increase in activity of ALP, but no change in ALP isozyme and concomitant increase in lactic dehydrogenase activity were found after prednisolone treatment. The results suggested that placental alkaline phosphatase as an oncodevelopmental gene product could be expressed ectopically by Eca109 cells and prednisolone could specifically induce increase in its activity.展开更多
背景白藜芦醇不仅具有抗肿瘤作用,其还能增强多种化疗药物的化疗敏感性,但其对食管癌耐药细胞的顺铂敏感性的作用尚不清楚.目的探讨白藜芦醇逆转食管癌细胞Eca109/DDP的耐药效应并从铁死亡的角度分析其潜在的作用机制.方法采用MTT法确...背景白藜芦醇不仅具有抗肿瘤作用,其还能增强多种化疗药物的化疗敏感性,但其对食管癌耐药细胞的顺铂敏感性的作用尚不清楚.目的探讨白藜芦醇逆转食管癌细胞Eca109/DDP的耐药效应并从铁死亡的角度分析其潜在的作用机制.方法采用MTT法确定白藜芦醇和顺铂的最佳作用时间和浓度;检测细胞增殖、细胞内丙二醛(malondialdehyde,MDA)、谷胱甘肽(glutathione,GSH)、活性氧(reactive oxygen species,ROS)及铁离子的水平;Western blot检测铁死亡相关调控因子酰基辅酶A合成酶长链家族成员4(acyl coenzyme A synthetase long chain family member 4,ACSL4)、铁蛋白重链(ferritin heavy chain,FTH)、谷胱甘肽过氧化物酶4(glutathione peroxidase 4,GPX4)和肿瘤蛋白53(tumor protein 53,P53)的蛋白表达情况.结果MTT检测结果显示,相较于单用顺铂,联用白藜芦醇对Eca109/DDP细胞的生长抑制作用较为显著(P<0.05).此外,白藜芦醇和顺铂联用可显著降低Eca109/DDP细胞克隆形成数(P<0.05),增加细胞内铁离子、MDA和ROS水平(P<0.05),降低GSH水平(P<0.05).铁死亡抑制剂(ferrostatin-1,Fer-1)和铁离子螯合剂(deferoxamine,DFO)可部分逆转白藜芦醇对细胞增殖的抑制作用(P<0.05).Western blot的结果显示相较于其他组,白藜芦醇和顺铂联用组的FTH和GPX4的蛋白表达显著降低(P<0.05),而ACSL4和P53的蛋白表达显著增加(P<0.05).结论白藜芦醇可通过铁死亡抑制Eca109/DDP细胞的增殖并逆转顺铂耐药.展开更多
文摘BACKGROUND The poor prognosis and rising incidence of esophageal cancer highlight the need for improved therapeutics that are essential prior to treatment.LCL161 is an SMAC(second mitochondrial activator of caspases)mimic and inhibitor of apoptosis protein(IAP)antagonist which exhibits anti-tumor effects and improves the chemical sensitivity of many cancers.AIM To ascertain the effects and mechanisms of the SMAC analog LCL161 on esophageal cancer cells.METHODS MTT assay and TUNEL assay were used to detect cell proliferation and apoptosis,respectively.Western blot analysis was used to study the molecular mechanisms of LCL161-induced death of ECA109 cells.RESULTS LCL161 decreased ECA109 cell proliferation in dose-and time-dependent manner and induced apoptosis of ECA109 cells in a dose-dependent manner.Also,LCL161 induced a significant decrease in the expression of the XIAP and significant increase in the expression of Caspase-3.In addition,Bax increased significantly with increasing concentrations of LCL161,and the relative expression of Bax was significantly different between groups.CONCLUSION These findings support the hypothesis that LCL161 can inhibit proliferation and induce apoptosis in esophageal cancer cells by regulating the expression of IAP family members,suggesting that it has potential to be an effective treatment for esophageal squamous cell carcinoma.
基金Supported by the Natural Science Fundation of Hebei Province of China(No.H2013209053)Scientific and Technological Project of Administration of Traditional Chinese Medicine of Hebei Province,China(No.2014185)
文摘Objective： To investigate the anti-tumor activity and molecular mechanism of Tonglian Decoction （通莲汤, TLD） on esophageal carcinoma Eca109 cells. Methods： Ecal09 cells were treated with TLD and its separated formulae, including the clearing-heat and detoxification formula （Q）, activating-blood and promoting-qi formula （H） and nourishing-yin and blood formula （Z）. Cell proliferation was measured using the 3-（4,5-dimethyl-2- thiazolyl）-2,5-diphenyl-2-H-tetrazolium bromide assay, cell morphology was observed using a microscope, the cell cycle was measured using flow cytometry and the activity of the nuclear factor-kappa B （NF- K B） signal pathway was detected by Western blot. Results： The half maximal inhibitory concentrations of TLD, Q and H were 386, 771 and 729 rag/L, respectively. TLD, Q and H significantly inhibited cell proliferation, with 69.43%, 60.84% and 61.90% of treated cells in the G1 phase of the cell cycle. The percentage of cells in S phase increased significantly after treatment with TLD, Q, and H compared with the control group （P〈0.05）, and TLD showed the strongest effect. Z had no influence on the cell cycle compared with the control group （P〉0.05）. Western blot detection indicated slight differences in the inhibition of the NF- K B pathway by the different formulae. TLD formula strongly inhibited IKK β, NF- K B, interleukin-6 and tumor necrosis factor-α expression compared with the control group. Conclusions： TLD inhibited Eca109 cell proliferation by arresting cells in S phase. The possible mechanism might be related to inhibiting the NF- K B transduction cascade. The combination of the herbs found in the three separate formulae, H, Q and Z, work synergistically in TLD to produce the inhibitory effects of TLD treatment on Eca109 proliferation.
基金Supported by the Henan Medical Science and Technology Innovation Proiect. No. 200084
文摘AIM: To study the role of P38 kinase in esophageal cancer cell apoptosis induced by genotoxin, cisplatin and the unfolded protein response (UPR) inducer, dithiothreitol(DTT).METHODS: Esophageal carcinoma cell line Eca109 was cultured in RPMI 1640 medium to 70% confluency and treated with either cisplatin, DTT, or cisplatin plus DTT in the presence or absence of P38 inhibitor, SB203580. The untreated cells served as the control. The esophageal carcinoma cell apoptosis was detected by agarose gel DNA ladder analysis and quantified by flow cytometry.The P38 phosphorylation was detected by immunohis-tochemistry using antibodies specific to phosphorylated P38 protein.RESULTS: (1) Both cisplatin and DTT induced apoptosis in the esophageal cancer cell line Eca109 as shown by DNA ladder formation; (2) As detected by antibodies specific for the phosphorylated P38 protein (p-P38), both cisplatin and DTT treatments activated the stress-activated enzyme, MAP kinase P38. The number of positive cells was about 50% for the treatment groups, comparing to that of 10% for untreated group. DTT treatment, but not cisplatin treatment, induces nuclear localization of p-P38;(3) As measured by flow cytometry, inhibition of P38 activity by SB203580 blocks DFT- and cisplatin-induced apoptosis.The rates for DTT, cisplatin, and DTT plus cisplatin-induced apoptosis were 16.8%, 17.1%, and 21.4%, respectively.Addition of the SB compound during the incubation reduced the apoptotic rate to about 7.6% for all the treatment groups, suggesting that P38 activation is essential for cisplatin- and DTT-induced apoptosis in Eca109 cells.CONCLUSION: (1) Both DTT and cisplatin were able to induce apoptosis in esophageal cancer cell line Eca109;(2) P38 MAP kinase is essential for DTT- and cisplatin-induced apoptosis in Eca109 cells; (3) P38 activation may be the common signaling component relaying the multiple upstream signaling events to the downstream cell death program.
文摘The expression and properties of alkaline phosphatase (ALP) in Eca109 cells, a cell line derived fromhuman esophageal cancer were studied with specific inhibition assay and polyacrylamide gel electrophoresis.The results showed that ALP of Eca109 cells was heat stable and was strongly inhibited by L-pheuylalanine, but slightly inhibited by urea. Preduisolone could causedramatic increase in activity of ALP, but no change in ALP isozyme and concomitant increase in lactic dehydrogenase activity were found after prednisolone treatment. The results suggested that placental alkaline phosphatase as an oncodevelopmental gene product could be expressed ectopically by Eca109 cells and prednisolone could specifically induce increase in its activity.
文摘背景白藜芦醇不仅具有抗肿瘤作用,其还能增强多种化疗药物的化疗敏感性,但其对食管癌耐药细胞的顺铂敏感性的作用尚不清楚.目的探讨白藜芦醇逆转食管癌细胞Eca109/DDP的耐药效应并从铁死亡的角度分析其潜在的作用机制.方法采用MTT法确定白藜芦醇和顺铂的最佳作用时间和浓度;检测细胞增殖、细胞内丙二醛(malondialdehyde,MDA)、谷胱甘肽(glutathione,GSH)、活性氧(reactive oxygen species,ROS)及铁离子的水平;Western blot检测铁死亡相关调控因子酰基辅酶A合成酶长链家族成员4(acyl coenzyme A synthetase long chain family member 4,ACSL4)、铁蛋白重链(ferritin heavy chain,FTH)、谷胱甘肽过氧化物酶4(glutathione peroxidase 4,GPX4)和肿瘤蛋白53(tumor protein 53,P53)的蛋白表达情况.结果MTT检测结果显示,相较于单用顺铂,联用白藜芦醇对Eca109/DDP细胞的生长抑制作用较为显著(P<0.05).此外,白藜芦醇和顺铂联用可显著降低Eca109/DDP细胞克隆形成数(P<0.05),增加细胞内铁离子、MDA和ROS水平(P<0.05),降低GSH水平(P<0.05).铁死亡抑制剂(ferrostatin-1,Fer-1)和铁离子螯合剂(deferoxamine,DFO)可部分逆转白藜芦醇对细胞增殖的抑制作用(P<0.05).Western blot的结果显示相较于其他组,白藜芦醇和顺铂联用组的FTH和GPX4的蛋白表达显著降低(P<0.05),而ACSL4和P53的蛋白表达显著增加(P<0.05).结论白藜芦醇可通过铁死亡抑制Eca109/DDP细胞的增殖并逆转顺铂耐药.
