AIM: To clarify the associations between G-protein beta polypeptide 3 (GNB3) C825T polymorphism and risk of the irritable bowel syndrome (IBS) by a meta-analysis.
Glutamate-induced excitotoxicity plays a critical role in the neurological impairment caused by middle cerebral artery occlusion.Achyranthes bidentata polypeptides have been shown to protect against neurological funct...Glutamate-induced excitotoxicity plays a critical role in the neurological impairment caused by middle cerebral artery occlusion.Achyranthes bidentata polypeptides have been shown to protect against neurological functional damage caused by middle cerebral artery occlusion,but the underlying neuroprotective mechanisms and the relationship to glutamate-induced excitotoxicity remain unclear.Therefore,in the current study,we investigated the protective effects of Achyranthes bidentata polypeptides against glutamate-induced excitotoxicity in cultured hippocampal neurons.Hippocampal neurons were treated with Mg^2+-free extracellular solution containing glutamate(300μM)for 3 hours as a model of glutamate-mediated excitotoxicity(glutamate group).In the normal group,hippocampal neurons were incubated in Mg^2+-free extracellular solution.In the Achyranthes bidentata polypeptide group,hippocampal neurons were incubated in Mg^2+-free extracellular solution containing glutamate(300μM)and Achyranthes bidentata polypeptide at different concentrations.At 24 hours after exposure to the agents,3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and Hoechst 33258 staining were used to assess neuronal viability and nuclear m'orphology,respectively.Caspase-3 expression and activity were evaluated using western blot assay and colorimetric enzymatic assay,respectively.At various time points after glutamate treatment,reactive oxygen species in cells were detected by H2 DCF-DA,and mitochondrial membrane potential was detected by rhodamine 123 staining.To examine the effect of Achyranthes bidentata polypeptides on glutamate receptors,electrophysiological recording was used to measure the glutamate-induced inward current in cultured hippocampal neurons.Achyranthes bidentata polypeptide decreased the percentage of apoptotic cells and reduced the changes in caspase-3 expression and activity induced by glutamate.In addition,Achyranthes bidentata polypeptide attenuated the amplitude of the glutamate-induced current.Furthermore,the glutamate-induced increase in intracellular reactive oxygen species and reduction in mitochondrial membrane potential were attenuated by Achyranthes bidentata polypeptide treatment.These findings collectively suggest that Achyranthes bidentata polypeptides exert a neuroprotective effect in cultured hippocampal neurons by suppressing the overactivation of glutamate receptors and inhibiting the caspase-3-dependent mitochondrial apoptotic pathway.All animal studies were approved by the Animal Care and Use Committee,Nantong University,China(approval No.20120216-001)on February 16,2012.展开更多
Background:Apolipoprotein B mRNA editing catalytic polypeptide(APOBEC),an endogenous mutator,induces DNA damage and activates the ataxia telangiectasia and Rad3-related(ATR)-checkpoint kinase 1(Chk1)pathway.Although c...Background:Apolipoprotein B mRNA editing catalytic polypeptide(APOBEC),an endogenous mutator,induces DNA damage and activates the ataxia telangiectasia and Rad3-related(ATR)-checkpoint kinase 1(Chk1)pathway.Although cisplatin-based therapy is the mainstay for muscle-invasive bladder cancer(MIBC),it has a poor survival rate.Therefore,this study aimed to evaluate the efficacy of an ATR inhibitor combined with cisplatin in the treatment of APOBEC catalytic subunit 3B(APOBEC3B)expressing MIBC.Methods:Immunohistochemical staining was performed to analyze an association between APOBEC3B and ATR in patients with MIBC.The APOBEC3B expression in MIBC cell lines was assessed using real-time polymerase chain reaction and western blot analysis.Western blot analysis was performed to confirm differences in phosphorylated Chk1(pChk1)expression according to the APOBEC3B expression.Cell viability and apoptosis analyses were performed to examine the anti-tumor activity of ATR inhibitors combined with cisplatin.Results:There was a significant association between APOBEC3B and ATR expression in the tumor tissues obtained from patients with MIBC.Cells with higher APOBEC3B expression showed higher pChk1 expression than cells expressing low APOBEC3B levels.Combination treatment of ATR inhibitor and cisplatin inhibited cell growth in MIBC cells with a higher APOBEC3B expression.Compared to cisplatin single treatment,combination treatment induced more apoptotic cell death in the cells with higher APOBEC3B expression.Conclusion:Our study shows that APOBEC3B’s higher expression status can enhance the sensitivity of MIBC to cisplatin upon ATR inhibition.This result provides new insight into appropriate patient selection for the effective application of ATR inhibitors in MIBC.展开更多
旨在建立检测猪圆环病毒3型(Porcine circovirus type 3,PCV3)血清抗体的间接ELISA方法以及摸清广西地区PCV3的流行情况。试验以PCV3的核衣壳蛋白多肽作为包被抗原,通过对包被抗原的最佳工作浓度、血清稀释度、封闭剂、酶标抗体工作浓...旨在建立检测猪圆环病毒3型(Porcine circovirus type 3,PCV3)血清抗体的间接ELISA方法以及摸清广西地区PCV3的流行情况。试验以PCV3的核衣壳蛋白多肽作为包被抗原,通过对包被抗原的最佳工作浓度、血清稀释度、封闭剂、酶标抗体工作浓度及作用时间、底物反应时间,阴性和阳性判断标准,敏感性、特异性及重复性等进行研究,建立了检测PCV3间接ELISA方法。随后,应用建立的方法检测2021—2023年广西地区采集的912份猪血清中PCV3抗体存在的情况。抗原最佳包被量为1.25μg/孔、血清稀释度为1∶800、封闭剂为5%的脱脂奶粉、酶标抗体工作浓度为1∶10000、二抗作用时间为60 min、底物反应时间为15 min。阴性和阳性判断标准OD_(450)≥0.55,为阳性;OD_(450)≤0.48,为阴性;当0.48<OD_(450)<0.55时,为疑似阳性,需对疑似样品再次检测。该方法与猪圆环病毒2型(PCV2)、猪瘟(CSFV)、猪繁殖与呼吸综合征(PRRSV)和伪狂犬病(PRV)等病毒抗体无交叉反应。批内与批间重复性试验的变异系数分别低于5%与8%,说明基于PCV3核衣壳蛋白多肽建立的间接ELISA方法具有良好的敏感性、特异性及重复性,检测方法稳定可靠。利用所建立的方法进行血清学调查时发现,广西地区PCV3抗体阳性率达到36.62%,说明广西地区猪群中PCV3的感染率较高,应引起重视。展开更多
Termination of translation in eukaryotes requires two polypeptide chain-release factors, eRF1 and eRF3. eRF1 recognizes stop signals, whereas eRF3 is a ribosome-dependent and eRFl-dependent GTPase. Polypeptide release...Termination of translation in eukaryotes requires two polypeptide chain-release factors, eRF1 and eRF3. eRF1 recognizes stop signals, whereas eRF3 is a ribosome-dependent and eRFl-dependent GTPase. Polypeptide release factor eRF3 consists of N-terminal variable region and C-terminal conserved part. C-terminal part of eRF3 is responsible for termination of the translation, In the present study, the C-terminal of Euplotes octocarinatus eRF3 （eRF3C） and truncate eRF3C lacking 76 amino acids in C-terminal （eRF3Ct） were expressed in Escherichia coll. The recombinant GST-eRF3C and GST-eRF3Ct polypeptides were purifled by affinity chromatography using glutathione Sepharose 4B column. After enzymatic cleavage of GST tail, the eRF3C and eRF3Ct protein were obtained. Pull-down analysis showed that the recombinant GST-eRF3C and GST-eRF3Ct polypeptides interacted with E. octocarinatus polypeptide chain release factor eRF1a. This result suggested that the C-terminal of eRF3 having 76 amino acids were not required for the binding of eRFla in Euplotes octocarinatus.展开更多
文摘AIM: To clarify the associations between G-protein beta polypeptide 3 (GNB3) C825T polymorphism and risk of the irritable bowel syndrome (IBS) by a meta-analysis.
基金supported by the National Natural Science Foundation of China,No.81073079(to HMS)the Natural Science Foundation of the Jiangsu Higher Education Institute of China,No.18KJA180009(to HMS)the Science Foundation of Nantong City of China,No.MS12018043(to HMS)
文摘Glutamate-induced excitotoxicity plays a critical role in the neurological impairment caused by middle cerebral artery occlusion.Achyranthes bidentata polypeptides have been shown to protect against neurological functional damage caused by middle cerebral artery occlusion,but the underlying neuroprotective mechanisms and the relationship to glutamate-induced excitotoxicity remain unclear.Therefore,in the current study,we investigated the protective effects of Achyranthes bidentata polypeptides against glutamate-induced excitotoxicity in cultured hippocampal neurons.Hippocampal neurons were treated with Mg^2+-free extracellular solution containing glutamate(300μM)for 3 hours as a model of glutamate-mediated excitotoxicity(glutamate group).In the normal group,hippocampal neurons were incubated in Mg^2+-free extracellular solution.In the Achyranthes bidentata polypeptide group,hippocampal neurons were incubated in Mg^2+-free extracellular solution containing glutamate(300μM)and Achyranthes bidentata polypeptide at different concentrations.At 24 hours after exposure to the agents,3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and Hoechst 33258 staining were used to assess neuronal viability and nuclear m'orphology,respectively.Caspase-3 expression and activity were evaluated using western blot assay and colorimetric enzymatic assay,respectively.At various time points after glutamate treatment,reactive oxygen species in cells were detected by H2 DCF-DA,and mitochondrial membrane potential was detected by rhodamine 123 staining.To examine the effect of Achyranthes bidentata polypeptides on glutamate receptors,electrophysiological recording was used to measure the glutamate-induced inward current in cultured hippocampal neurons.Achyranthes bidentata polypeptide decreased the percentage of apoptotic cells and reduced the changes in caspase-3 expression and activity induced by glutamate.In addition,Achyranthes bidentata polypeptide attenuated the amplitude of the glutamate-induced current.Furthermore,the glutamate-induced increase in intracellular reactive oxygen species and reduction in mitochondrial membrane potential were attenuated by Achyranthes bidentata polypeptide treatment.These findings collectively suggest that Achyranthes bidentata polypeptides exert a neuroprotective effect in cultured hippocampal neurons by suppressing the overactivation of glutamate receptors and inhibiting the caspase-3-dependent mitochondrial apoptotic pathway.All animal studies were approved by the Animal Care and Use Committee,Nantong University,China(approval No.20120216-001)on February 16,2012.
基金supported by St.Vincent’s Hospital,the Research Institute of Medical Science(Grant Number:SVHR-2021-03).
文摘Background:Apolipoprotein B mRNA editing catalytic polypeptide(APOBEC),an endogenous mutator,induces DNA damage and activates the ataxia telangiectasia and Rad3-related(ATR)-checkpoint kinase 1(Chk1)pathway.Although cisplatin-based therapy is the mainstay for muscle-invasive bladder cancer(MIBC),it has a poor survival rate.Therefore,this study aimed to evaluate the efficacy of an ATR inhibitor combined with cisplatin in the treatment of APOBEC catalytic subunit 3B(APOBEC3B)expressing MIBC.Methods:Immunohistochemical staining was performed to analyze an association between APOBEC3B and ATR in patients with MIBC.The APOBEC3B expression in MIBC cell lines was assessed using real-time polymerase chain reaction and western blot analysis.Western blot analysis was performed to confirm differences in phosphorylated Chk1(pChk1)expression according to the APOBEC3B expression.Cell viability and apoptosis analyses were performed to examine the anti-tumor activity of ATR inhibitors combined with cisplatin.Results:There was a significant association between APOBEC3B and ATR expression in the tumor tissues obtained from patients with MIBC.Cells with higher APOBEC3B expression showed higher pChk1 expression than cells expressing low APOBEC3B levels.Combination treatment of ATR inhibitor and cisplatin inhibited cell growth in MIBC cells with a higher APOBEC3B expression.Compared to cisplatin single treatment,combination treatment induced more apoptotic cell death in the cells with higher APOBEC3B expression.Conclusion:Our study shows that APOBEC3B’s higher expression status can enhance the sensitivity of MIBC to cisplatin upon ATR inhibition.This result provides new insight into appropriate patient selection for the effective application of ATR inhibitors in MIBC.
基金This work was supported by grants from National Natural Science Foundation of China(No.30670282,30470239,3030038)Natural Science Foundation of Shanxi Province(No.20041079,20051065).
文摘Termination of translation in eukaryotes requires two polypeptide chain-release factors, eRF1 and eRF3. eRF1 recognizes stop signals, whereas eRF3 is a ribosome-dependent and eRFl-dependent GTPase. Polypeptide release factor eRF3 consists of N-terminal variable region and C-terminal conserved part. C-terminal part of eRF3 is responsible for termination of the translation, In the present study, the C-terminal of Euplotes octocarinatus eRF3 （eRF3C） and truncate eRF3C lacking 76 amino acids in C-terminal （eRF3Ct） were expressed in Escherichia coll. The recombinant GST-eRF3C and GST-eRF3Ct polypeptides were purifled by affinity chromatography using glutathione Sepharose 4B column. After enzymatic cleavage of GST tail, the eRF3C and eRF3Ct protein were obtained. Pull-down analysis showed that the recombinant GST-eRF3C and GST-eRF3Ct polypeptides interacted with E. octocarinatus polypeptide chain release factor eRF1a. This result suggested that the C-terminal of eRF3 having 76 amino acids were not required for the binding of eRFla in Euplotes octocarinatus.
