The preparation of polypeptide materials in continuous flow reactors shows great potential with improved reproducibility and scalability.However,conventional polypeptide synthesis from the polymerization of N-carboxya...The preparation of polypeptide materials in continuous flow reactors shows great potential with improved reproducibility and scalability.However,conventional polypeptide synthesis from the polymerization of N-carboxyanhydride(NCA)is conducted at relatively slow rates,requiring long tubing or ending up with low-molecular-weight polymers.Inspired by recent advances in accelerated NCA polymerization,we report the crown-ether-catalyzed,rapid synthesis of polypeptide materials in cosolvents in flow reactors.The incorporation of low-polarity dichloromethane and the use of catalysts enabled fast conversion of monomers in 30 min,yielding well-defined polypeptides(up to 30 k Da)through a 20-cm tubing reactor.Additionally,random or block copolypeptides were efficiently prepared by incorporating a second NCA monomer.We believe that this work highlights the accelerated polymerization design in flow polymerization processes,offering the continuous production of polypeptide materials.展开更多
BACKGROUND As a member of the chaperonin-containing tailless complex polypeptide 1(TCP1)complex,which plays a pivotal role in ensuring the accurate folding of numerous proteins,chaperonin-containing TCP1 subunit 6A(CC...BACKGROUND As a member of the chaperonin-containing tailless complex polypeptide 1(TCP1)complex,which plays a pivotal role in ensuring the accurate folding of numerous proteins,chaperonin-containing TCP1 subunit 6A(CCT6A)participates in various physiological and pathological processes.However,its effects on cell death and cancer therapy and the underlying mechanisms need further exploration in colorectal cancer(CRC)cells.AIM To explore the effects of CCT6A on cell death and cancer therapy and the underlying mechanisms in CRC.METHODS Cell proliferation was evaluated using the MTS assay,EdU staining,and colony growth assays.The expression of CCT6A was monitored by immunoblotting and quantitative PCR.CCT6A was knocked out by CRISPR-Cas9,and overexpressed by transfecting plasmids.Autophagy was examined by immunoblotting and the mCherry-GFP-LC3 assay.To monitor apoptosis and necroptosis,immunoblotting,co-immunoprecipitation,and flow cytometry were employed.RESULTS Cisplatin(DDP)exerted cytotoxic effects on CRC cells while simultaneously downregulating the expression of CCT6A.Depletion of CCT6A amplified the cytotoxic effects of DDP,whereas overexpression of CCT6A attenuated these adverse effects.CCT6A suppressed autophagy,apoptosis,and necroptosis under both basal and DDP-treated conditions.Autophagy inhibitors significantly enhanced the cytotoxic effects of DDP,whereas a necroptosis inhibitor partially reversed the cell viability loss induced by DDP.Furthermore,inhibiting autophagy enhanced both apoptosis and necroptosis induced by DDP.CONCLUSION CCT6A negatively modulates autophagy,apoptosis,and necroptosis,and CCT6A confers resistance to DDP therapy in CRC,suggesting its potential as a therapeutic target.展开更多
Ischemic stroke is the leading cause of death in China,accounting for approximately one-third of all stroke-associated deaths worldwide.Currently,thrombolysis is employed for ischemic strokes.However,due to the limite...Ischemic stroke is the leading cause of death in China,accounting for approximately one-third of all stroke-associated deaths worldwide.Currently,thrombolysis is employed for ischemic strokes.However,due to the limited therapeutic window of thrombolytic agents,most patients do not receive the drug at the right time.Moreover,these agents are associated with risks of hemorrhage and reperfusion damage.Herein,Angiopep-2(ANG)-black phosphorus(BP)-resveratrol(RES),a drug-loaded system,was used to deliver drugs across the blood–brain barrier(BBB).ANG-BP-RES has a uniform size,stable structure,good photothermal effect,and strong drug release ability under near-infrared(NIR)irradiation and acidic conditions.Furthermore,ANG-BP-RES can efficiently target the brain and improve BBB permeability,exerting a significant therapeutic effect against ischemic brain injury,especially after NIR irradiation.ANG-BP-RES is also biocompatible and shows minimal toxicity toward cells and tissues.This study offers novel insights into the therapeutic management of ischemic brain injury.展开更多
载脂蛋白B m RNA编辑酶催化多肽3F(APOBEC3F)属固有免疫系统中的重要成员,对人免疫缺陷病毒(HIV)、乙型肝炎病毒(HBV)等多种病毒的复制具有广泛的抑制作用。为构建APOBEC3F基因的真核表达载体,并检测其在体外培养的MARC145细胞中的表达...载脂蛋白B m RNA编辑酶催化多肽3F(APOBEC3F)属固有免疫系统中的重要成员,对人免疫缺陷病毒(HIV)、乙型肝炎病毒(HBV)等多种病毒的复制具有广泛的抑制作用。为构建APOBEC3F基因的真核表达载体,并检测其在体外培养的MARC145细胞中的表达情况,通过RT-PCR从猪脾脏组织扩增APOBEC3F基因,定向克隆到表达增强型绿色荧光蛋白(EGFP)的真核表达载体p EGFP-CMV中,构建p EGFP-APOBEC3F。PCR扩增及测序显示,APOBEC3F插入载体位置、方向及序列均正确。p EGFP-APOBEC3F经脂质体介导法转染MARC145细胞,转染后24 h APOBEC3F m RNA的水平升至最高。细胞免疫化学法检测发现APOBEC3F蛋白主要在MARC145细胞质中表达。研究结果为体外研究猪APOBEC3F基因在抗猪蓝耳病中的作用奠定了基础。展开更多
目的探讨载脂蛋白B mRNA编辑酶催化多肽样蛋白3G(apolipoprotein B mRNA editing enzyme catalytic polypeptide like3G,APOBEC3G)(也称为CEM15)体外抗乙型肝炎病毒(HBV)的作用及其机制。方法脂质体转染pcDNA3.1 Human APOBEC3G-Myc-6X...目的探讨载脂蛋白B mRNA编辑酶催化多肽样蛋白3G(apolipoprotein B mRNA editing enzyme catalytic polypeptide like3G,APOBEC3G)(也称为CEM15)体外抗乙型肝炎病毒(HBV)的作用及其机制。方法脂质体转染pcDNA3.1 Human APOBEC3G-Myc-6Xhis、pcDAN3.1/His-C进入HepG2.2.15细胞,转染后,RT-PCR证实转染基因的表达,Western Blot证实蛋白的表达。通过ELISA方法检测细胞上清液中乙型肝炎表面抗原(HBsAg)及乙型肝炎e抗原(HBeAg),RT-PCR分析APOBEC3G对HBV mRNA转录的影响。结果APOBEC3G基因与蛋白在HepG2.2.15细胞都有表达,与空质粒转染组相比,pcDNA3.1 Human APOBEC3G-Myc-6Xhis转染组HBsAg含量下降70.38%,HBeAg含量下降62.88%,未转质粒细胞为空白对照组。结论APOBEC3G在体外可以抑制HBV复制,可以作为一种新型的抗病毒制剂治疗乙肝病毒感染。展开更多
基金financially supported by the National Natural Science Foundation of China(No.22101194)Natural Science Foundation of Jiangsu Province(No.BK20210733)+3 种基金Suzhou Municipal Science and Technology Bureau(No.ZXL2021447)Collaborative Innovation Center of Suzhou Nano Science&Technologythe 111 ProjectJoint International Research Laboratory of Carbon-Based Functional Materials and Devices。
文摘The preparation of polypeptide materials in continuous flow reactors shows great potential with improved reproducibility and scalability.However,conventional polypeptide synthesis from the polymerization of N-carboxyanhydride(NCA)is conducted at relatively slow rates,requiring long tubing or ending up with low-molecular-weight polymers.Inspired by recent advances in accelerated NCA polymerization,we report the crown-ether-catalyzed,rapid synthesis of polypeptide materials in cosolvents in flow reactors.The incorporation of low-polarity dichloromethane and the use of catalysts enabled fast conversion of monomers in 30 min,yielding well-defined polypeptides(up to 30 k Da)through a 20-cm tubing reactor.Additionally,random or block copolypeptides were efficiently prepared by incorporating a second NCA monomer.We believe that this work highlights the accelerated polymerization design in flow polymerization processes,offering the continuous production of polypeptide materials.
基金Supported by Shandong Provincial Natural Science Foundation,No.ZR2023MH329Project of Shandong Province Higher Educational Youth Innovation Science and Technology Program,No.2023KJ263and Natural Science Foundation of Gansu Province,China,No.22JR5RA953.
文摘BACKGROUND As a member of the chaperonin-containing tailless complex polypeptide 1(TCP1)complex,which plays a pivotal role in ensuring the accurate folding of numerous proteins,chaperonin-containing TCP1 subunit 6A(CCT6A)participates in various physiological and pathological processes.However,its effects on cell death and cancer therapy and the underlying mechanisms need further exploration in colorectal cancer(CRC)cells.AIM To explore the effects of CCT6A on cell death and cancer therapy and the underlying mechanisms in CRC.METHODS Cell proliferation was evaluated using the MTS assay,EdU staining,and colony growth assays.The expression of CCT6A was monitored by immunoblotting and quantitative PCR.CCT6A was knocked out by CRISPR-Cas9,and overexpressed by transfecting plasmids.Autophagy was examined by immunoblotting and the mCherry-GFP-LC3 assay.To monitor apoptosis and necroptosis,immunoblotting,co-immunoprecipitation,and flow cytometry were employed.RESULTS Cisplatin(DDP)exerted cytotoxic effects on CRC cells while simultaneously downregulating the expression of CCT6A.Depletion of CCT6A amplified the cytotoxic effects of DDP,whereas overexpression of CCT6A attenuated these adverse effects.CCT6A suppressed autophagy,apoptosis,and necroptosis under both basal and DDP-treated conditions.Autophagy inhibitors significantly enhanced the cytotoxic effects of DDP,whereas a necroptosis inhibitor partially reversed the cell viability loss induced by DDP.Furthermore,inhibiting autophagy enhanced both apoptosis and necroptosis induced by DDP.CONCLUSION CCT6A negatively modulates autophagy,apoptosis,and necroptosis,and CCT6A confers resistance to DDP therapy in CRC,suggesting its potential as a therapeutic target.
基金funded by the National Natural Science Foundation of China (No. 81960334)the Guiding Plan of Xinjiang Production Construction Corps (No. 2022ZD007)+4 种基金the Science and Technology Innovation Leading Talents Program of Guangdong Province (No. 2019TX05C343)the Basic and Applied Basic Research Foundation of Guangdong Province-Regional Joint Fund-Key Projects (No. 2019B1515120043)the Project supported by the State Key Laboratory of Luminescence and Applications (No. SKLA-2020-03)the support from Instrumental Analysis Center of Shenzhen University (Xili Campus)Instrumental Analysis Center of Shihezi University.
文摘Ischemic stroke is the leading cause of death in China,accounting for approximately one-third of all stroke-associated deaths worldwide.Currently,thrombolysis is employed for ischemic strokes.However,due to the limited therapeutic window of thrombolytic agents,most patients do not receive the drug at the right time.Moreover,these agents are associated with risks of hemorrhage and reperfusion damage.Herein,Angiopep-2(ANG)-black phosphorus(BP)-resveratrol(RES),a drug-loaded system,was used to deliver drugs across the blood–brain barrier(BBB).ANG-BP-RES has a uniform size,stable structure,good photothermal effect,and strong drug release ability under near-infrared(NIR)irradiation and acidic conditions.Furthermore,ANG-BP-RES can efficiently target the brain and improve BBB permeability,exerting a significant therapeutic effect against ischemic brain injury,especially after NIR irradiation.ANG-BP-RES is also biocompatible and shows minimal toxicity toward cells and tissues.This study offers novel insights into the therapeutic management of ischemic brain injury.
文摘载脂蛋白B m RNA编辑酶催化多肽3F(APOBEC3F)属固有免疫系统中的重要成员,对人免疫缺陷病毒(HIV)、乙型肝炎病毒(HBV)等多种病毒的复制具有广泛的抑制作用。为构建APOBEC3F基因的真核表达载体,并检测其在体外培养的MARC145细胞中的表达情况,通过RT-PCR从猪脾脏组织扩增APOBEC3F基因,定向克隆到表达增强型绿色荧光蛋白(EGFP)的真核表达载体p EGFP-CMV中,构建p EGFP-APOBEC3F。PCR扩增及测序显示,APOBEC3F插入载体位置、方向及序列均正确。p EGFP-APOBEC3F经脂质体介导法转染MARC145细胞,转染后24 h APOBEC3F m RNA的水平升至最高。细胞免疫化学法检测发现APOBEC3F蛋白主要在MARC145细胞质中表达。研究结果为体外研究猪APOBEC3F基因在抗猪蓝耳病中的作用奠定了基础。
文摘目的探讨载脂蛋白B mRNA编辑酶催化多肽样蛋白3G(apolipoprotein B mRNA editing enzyme catalytic polypeptide like3G,APOBEC3G)(也称为CEM15)体外抗乙型肝炎病毒(HBV)的作用及其机制。方法脂质体转染pcDNA3.1 Human APOBEC3G-Myc-6Xhis、pcDAN3.1/His-C进入HepG2.2.15细胞,转染后,RT-PCR证实转染基因的表达,Western Blot证实蛋白的表达。通过ELISA方法检测细胞上清液中乙型肝炎表面抗原(HBsAg)及乙型肝炎e抗原(HBeAg),RT-PCR分析APOBEC3G对HBV mRNA转录的影响。结果APOBEC3G基因与蛋白在HepG2.2.15细胞都有表达,与空质粒转染组相比,pcDNA3.1 Human APOBEC3G-Myc-6Xhis转染组HBsAg含量下降70.38%,HBeAg含量下降62.88%,未转质粒细胞为空白对照组。结论APOBEC3G在体外可以抑制HBV复制,可以作为一种新型的抗病毒制剂治疗乙肝病毒感染。
