Background: Diabetes mellitus is one of the major health problems for people all over the world today. According to international diabetes federation reports, diabetes affects 382 million people worldwide. Environment...Background: Diabetes mellitus is one of the major health problems for people all over the world today. According to international diabetes federation reports, diabetes affects 382 million people worldwide. Environmental pollutants have deleterious effects on glucose metabolism and cause insulin resistance. We aimed to investigate the effects of the environmental pollutants benzo(a)pyrene, and the therapeutic potential of resveratrol. Methods: 20 μM of benzo(a)pyrene was administered after 48 h of resveratrol (80 μM) application for 24 h in INS-1 (832/13) insulinoma cells. The cells were treated with 20 μM benzo(a)pyrene for 24 hours after 48 hours initial preconditions with 10 μM resveratrol. Oxidative stress status, insulin secretion and apoptosis were analyzed by molecular techniques. Results: Though resveratrol increased the antioxidant status which was decreased by benzo(a)pyrene, interestingly, it increased the oxidative status. Resveratrol increased benzo(a)pyrene-depleted reduced glutathione levels to the control level. The mRNA expression levels of beta-cell functions associated with genes insulin-1, insulin-2 and sirtuin-1 were upregulated by resveratrol. Resveratrol treatment elevated the insulin concentration of culture medium, and the mRNA expression of forkhead box protein-1 gene. Resveratrol upregulated benzo(a)pyrene-downregulated p53 gene expression. On the other hand, benzo(a)pyrene-downregulated mRNA expression of B-cell lymphoma-2 was induced by resveratrol treatment. Conclusion: The data showed that resveratrol could reverse the oxidative alterations, functional impairments and the carcinogenetic effects of benzo(a)pyrene in pancreas beta-cells.展开更多
Most type 2 diabetics are accompanied with deficiency of insulin secretion and hyperglycemia. It has reported that glucose-stimulated insulin secretion of β cell (GSIS)
目的探究慢病毒介导RNAi沉默SGMS2基因的单克隆细胞系构建中最佳感染复数(multiplicity of infection,MOI)及BSD基因筛选抗生素(blasticidin)浓度。方法荧光标记小鼠SGMS2干扰阴性对照慢病毒并按照MOI值0、10、30、60、120(TU number/ce...目的探究慢病毒介导RNAi沉默SGMS2基因的单克隆细胞系构建中最佳感染复数(multiplicity of infection,MOI)及BSD基因筛选抗生素(blasticidin)浓度。方法荧光标记小鼠SGMS2干扰阴性对照慢病毒并按照MOI值0、10、30、60、120(TU number/cell)分别侵染INS-1空白细胞,培养72 h后使用荧光显微镜拍照并计算细胞的荧光比率(%)及死亡率(%),以确定最佳MOI值。小鼠胰岛素瘤INS-1空白细胞中加入0、1、2、3μg/m L blasticidin,第7天时采用MTT法检测细胞的死亡率,以确定细胞抗生素敏感浓度。使用SGMS2干扰阴性对照慢病毒及SGMS2干扰慢病毒(病毒滴度:1×108TU/m L)按照最佳MOI值侵染细胞,并用blasticidin敏感浓度进行阳性细胞筛选,获得混合系细胞。当细胞的荧光率达90%时,进行单克隆稳转细胞系的构建。结果最佳MOI值为60,此时细胞的荧光率达100%,但细胞的死亡率<0.5%,细胞保持原有的形态。当blasticidin敏感浓度为2μg/m L,此时空白细胞失去原有的贴壁性,全部死亡。INS-1-SEMS2细胞第2次检测的Ct值28.21大于第1次检测的Ct值27.58,且siRNA的干扰效率为77.78%,siRNA成功表达,混合稳转细胞系构建成功。成功构建小鼠胰岛素瘤INS-1-SEMS2单克隆细胞系。结论慢病毒介导RNAi沉默基因SGMS2的单克隆细胞系构建成功。展开更多
Purpose Dinutuximab beta is a monoclonal antibody used only in glioblastoma.Streptozotocin is an agent that is particularly toxic to pancreatic beta cells.Dinutuximab beta causes cytotoxicity through natural killer ce...Purpose Dinutuximab beta is a monoclonal antibody used only in glioblastoma.Streptozotocin is an agent that is particularly toxic to pancreatic beta cells.Dinutuximab beta causes cytotoxicity through natural killer cells and neutrophils and shows effects.In this study,cytotoxicity was induced by streptozotocin without natural killer cells and neutrophils.Gaining the ability to show the effect of Dinutuximab beta without a natural killer was the first aim of this study.This will be especially important in cases where the immune system is deficient,such as cancer.The second aim of the study was to investigate the effects of Dinutuximab beta on cell viability and cell death in insulinoma under the conditions created.Methods The effect of Dinutuximab beta in the presence of natural killer cells in vivo was created by the application of Streptozotocin to Beta-cell tumors of the pancreas in vitro.The cell viability was determined with WST-1 assay.Reactive oxygen species were measured by using dichlorofluorescein diacetate as a spectrophotometer.The cells were marked with DAPI to indicate apoptotic markers(nuclear condensation and fragmentation)with the confocal microscope.GLUT2(Glucose transporter 2),IR(Insulin receptor),INS1,and INS2 expression levels were analyzed with q-RT-PCR.Results The cell cytotoxicity was induced by Streptozotocin.The cells proliferated with the administration of Dinutuximab beta alone.The result of Dinutuximab beta administered following Streptozotocin administration resulted in more cell death,increased ROS levels,GLUT2,Ins1,and Ins2 mRNA expression levels,and decreased IR mRNA expression levels.Furthermore,the cells predominantly died via apoptosis showing cytoplasmic condensation and DNA fragmentation.Conclusions The lethal effect of Dinutuximab beta without a natural killer was provided by Streptozotocin in Beta cell tumors of the pancreas.展开更多
基金supported by the Scientific Research Projects Unit of Afyon Kocatepe University(Project No:10.TIP.18/MSc Thesis in Medical Biochemistry)NATO-SPS Program(Project No:MYP-G5266).
文摘Background: Diabetes mellitus is one of the major health problems for people all over the world today. According to international diabetes federation reports, diabetes affects 382 million people worldwide. Environmental pollutants have deleterious effects on glucose metabolism and cause insulin resistance. We aimed to investigate the effects of the environmental pollutants benzo(a)pyrene, and the therapeutic potential of resveratrol. Methods: 20 μM of benzo(a)pyrene was administered after 48 h of resveratrol (80 μM) application for 24 h in INS-1 (832/13) insulinoma cells. The cells were treated with 20 μM benzo(a)pyrene for 24 hours after 48 hours initial preconditions with 10 μM resveratrol. Oxidative stress status, insulin secretion and apoptosis were analyzed by molecular techniques. Results: Though resveratrol increased the antioxidant status which was decreased by benzo(a)pyrene, interestingly, it increased the oxidative status. Resveratrol increased benzo(a)pyrene-depleted reduced glutathione levels to the control level. The mRNA expression levels of beta-cell functions associated with genes insulin-1, insulin-2 and sirtuin-1 were upregulated by resveratrol. Resveratrol treatment elevated the insulin concentration of culture medium, and the mRNA expression of forkhead box protein-1 gene. Resveratrol upregulated benzo(a)pyrene-downregulated p53 gene expression. On the other hand, benzo(a)pyrene-downregulated mRNA expression of B-cell lymphoma-2 was induced by resveratrol treatment. Conclusion: The data showed that resveratrol could reverse the oxidative alterations, functional impairments and the carcinogenetic effects of benzo(a)pyrene in pancreas beta-cells.
文摘Most type 2 diabetics are accompanied with deficiency of insulin secretion and hyperglycemia. It has reported that glucose-stimulated insulin secretion of β cell (GSIS)
文摘目的探究慢病毒介导RNAi沉默SGMS2基因的单克隆细胞系构建中最佳感染复数(multiplicity of infection,MOI)及BSD基因筛选抗生素(blasticidin)浓度。方法荧光标记小鼠SGMS2干扰阴性对照慢病毒并按照MOI值0、10、30、60、120(TU number/cell)分别侵染INS-1空白细胞,培养72 h后使用荧光显微镜拍照并计算细胞的荧光比率(%)及死亡率(%),以确定最佳MOI值。小鼠胰岛素瘤INS-1空白细胞中加入0、1、2、3μg/m L blasticidin,第7天时采用MTT法检测细胞的死亡率,以确定细胞抗生素敏感浓度。使用SGMS2干扰阴性对照慢病毒及SGMS2干扰慢病毒(病毒滴度:1×108TU/m L)按照最佳MOI值侵染细胞,并用blasticidin敏感浓度进行阳性细胞筛选,获得混合系细胞。当细胞的荧光率达90%时,进行单克隆稳转细胞系的构建。结果最佳MOI值为60,此时细胞的荧光率达100%,但细胞的死亡率<0.5%,细胞保持原有的形态。当blasticidin敏感浓度为2μg/m L,此时空白细胞失去原有的贴壁性,全部死亡。INS-1-SEMS2细胞第2次检测的Ct值28.21大于第1次检测的Ct值27.58,且siRNA的干扰效率为77.78%,siRNA成功表达,混合稳转细胞系构建成功。成功构建小鼠胰岛素瘤INS-1-SEMS2单克隆细胞系。结论慢病毒介导RNAi沉默基因SGMS2的单克隆细胞系构建成功。
基金supported by the Scientific Research Project Coordination Unit of Istanbul University.Project No:34960.
文摘Purpose Dinutuximab beta is a monoclonal antibody used only in glioblastoma.Streptozotocin is an agent that is particularly toxic to pancreatic beta cells.Dinutuximab beta causes cytotoxicity through natural killer cells and neutrophils and shows effects.In this study,cytotoxicity was induced by streptozotocin without natural killer cells and neutrophils.Gaining the ability to show the effect of Dinutuximab beta without a natural killer was the first aim of this study.This will be especially important in cases where the immune system is deficient,such as cancer.The second aim of the study was to investigate the effects of Dinutuximab beta on cell viability and cell death in insulinoma under the conditions created.Methods The effect of Dinutuximab beta in the presence of natural killer cells in vivo was created by the application of Streptozotocin to Beta-cell tumors of the pancreas in vitro.The cell viability was determined with WST-1 assay.Reactive oxygen species were measured by using dichlorofluorescein diacetate as a spectrophotometer.The cells were marked with DAPI to indicate apoptotic markers(nuclear condensation and fragmentation)with the confocal microscope.GLUT2(Glucose transporter 2),IR(Insulin receptor),INS1,and INS2 expression levels were analyzed with q-RT-PCR.Results The cell cytotoxicity was induced by Streptozotocin.The cells proliferated with the administration of Dinutuximab beta alone.The result of Dinutuximab beta administered following Streptozotocin administration resulted in more cell death,increased ROS levels,GLUT2,Ins1,and Ins2 mRNA expression levels,and decreased IR mRNA expression levels.Furthermore,the cells predominantly died via apoptosis showing cytoplasmic condensation and DNA fragmentation.Conclusions The lethal effect of Dinutuximab beta without a natural killer was provided by Streptozotocin in Beta cell tumors of the pancreas.
