目的探讨乙型肝炎病毒DNA聚合酶反式调节蛋白1(HBV DNA polymerase trans activated protein 1,HBVDNAPTP1)在羧酸酯酶1(carboxylesterase 1,CES1)介导的单核细胞凋亡信号通路中的作用机制。方法利用Phyre2在线工具预测得到CES1三级结构...目的探讨乙型肝炎病毒DNA聚合酶反式调节蛋白1(HBV DNA polymerase trans activated protein 1,HBVDNAPTP1)在羧酸酯酶1(carboxylesterase 1,CES1)介导的单核细胞凋亡信号通路中的作用机制。方法利用Phyre2在线工具预测得到CES1三级结构,分别用pCMV-Myc载体构建HBVDNAPTP1基因、pCMV-HA载体构建CES1基因及其截短体219-264aa和265-303aa的真核细胞表达质粒。分别转染细胞后通过免疫荧光和免疫共沉淀法确定HBVDNAPTP1和CES1互相作用的区段。后通过HBVDNAPTP1和CES1单独转染及共转染,检测其对细胞凋亡率的影响;同时在分离人外周血单个核细胞(Peripheral blood mononuclear cell,PBMC)上转染上述质粒,以确定CES1和HBVDNAPTP1对单核细胞形态的影响。利用qRT-PCR和Western blot检测不同质粒转染后下游Janus激酶-1(Janus kinase,JAK1)和信号传导及转录激活蛋白3(Signal transducer and activator of transcription 3,STAT3)的基因和蛋白的表达情况。结果HBVDNAPTP1与CES1有靶向关系,其中HBVDNAPTP1与CES1全长靶向结合效果最好,故后续实验以CES1全长序列为对象开展。CCK-8和PBMC电镜实验结果表明,与Control组和NC组相比,ov-HBVDNAPTP1组、ov-CES1组和ov-HBVDNAPTP1+ov-CES1组THP-1细胞的凋亡率均升高(P<0.001),细胞受损;与ov-HBVDNAPTP1组和ov-CES1组相比,ov-HBVDNAPTP1+ov-CES1组细胞凋亡率进一步升高(P<0.01),凋亡细胞、坏死细胞的数量增多。qRT-PCR和Western blot结果显示,HBVDNAPTP1和CES1均可以在转录水平和翻译水平上显著上调JAK1和STAT3的表达,且二者同时作用时上调趋势更加明显。结论本研究证实HBVDNAPTP1与CES1二者可协同抑制THP-1细胞增殖并诱导细胞损伤;还可协同上调JAK1/STAT3信号通路在转录和蛋白水平的表达来调控单核细胞凋亡。该发现为深入理解HBV感染相关免疫调控机制提供了新的理论依据。展开更多
Background:Head and neck squamous cell carcinoma(HNSCC)is a prevalent form of cancer globally,with chemoresistance posing a major challenge in treatment outcomes.The efficacy of the commonly used chemotherapeutic agent...Background:Head and neck squamous cell carcinoma(HNSCC)is a prevalent form of cancer globally,with chemoresistance posing a major challenge in treatment outcomes.The efficacy of the commonly used chemotherapeutic agent,cisplatin,is diminished in patients with poor prognoses.Methods:Various bioinformatics databases were utilized to examine Carboxylesterase 1(CES1)gene expression,clinicopathologic features,patient survival analysis,and gene function.An organoid model of HNSCC was established,along with the induction of drug-resistant HNSCC in the organoid model.CES1 expression was assessed using qRT-PCR and Western Blot,and differential markers were identified through transcriptome sequencing.Knockdown and overexpression models of CES1 were created in SCC-9 and patient-derived organoid(PDO)cells using shRNA and lentivirus to investigate the tumor biology and cisplatin resistance associated with CES1.Results:Research in bioinformatics has uncovered a strong correlation between the expression level of CES1 and the prognosis of HNSCC.The data suggests a significant link between CES1 expression and tobacco smoking.RNA-sequencing revealed a notable increase in CES1 expression in HNSCC-PDOcis-R cells compared to the parental PDO cells.Subsequently,we performed in vitro studies by HNSCC-PDO and SCC-9 and found that CES1-overexpressing cells exhibited reduced sensitivity to cisplatin and stronger tumor malignant biological behavior compared with CES1-knockdown cells.Conclusion:The observed association between CES1 expression and tobacco smoking implies a potential influence of smoking on the efficacy of cisplatin-based chemotherapy in HNSCC through the regulation of CES1 expression.展开更多
In traditional Chinese medicine herbs(TCM),including Radix Salviae Miltiorrhizae(Danshen),Radix Puerariae Lobatae(Gegen),Radix Angelicae Sinensis(Danggui),and Rhizoma Chuanxiong(Chuanxiong)are widely used for the prev...In traditional Chinese medicine herbs(TCM),including Radix Salviae Miltiorrhizae(Danshen),Radix Puerariae Lobatae(Gegen),Radix Angelicae Sinensis(Danggui),and Rhizoma Chuanxiong(Chuanxiong)are widely used for the prevention and treatment of cardiovascular diseases and also often co-administered with Western drugs as a part of integrative medicine practice.Carboxylesterase 1(CES1)plays a pivotal role in the metabolisms of pro-drugs,Since(S)-2-(2-(6-dimethylamino)-benzothiazole)-4,5-dihydrothiazole-4-carboxylate(NLMe)has recently been identified by us as a selective CES1 bioluminescent sensor,we developed a rapid method using this substrate for the direct measurement of CES1 activity in rats.This bioluminescence assay was applied to determine CES1 activity in rat tissues after a two-week oral administration of each of the four herbs noted above.The results demonstrated the presence of CES1 enzyme in rat blood and all tested tissues with much higher enzyme activity in the blood,liver,kidney and heart than that in the small intestine,spleen,lung,pancreas,brain and stomach.In addition,the four herbs showed tissue-specific effects on rat CES1 expression.Based on the CES1 biodistribution and its changes after treatment in rats,the possibility that Danshen,Gegen and Danggui might alter CES1 activities in human blood and kidney should be considered.In summary,a selective and sensitive bioluminescence assay was developed to rapidly evaluate CES1 activity and the effects of orally administered TCMs in rats.展开更多
用烟草花叶病毒弱毒株系N14预先接种,可以诱导烟草对病原真菌赤星病菌的系统获得性抗性(SAR),减轻病菌引起的赤星病。选择表现诱导抗性最强植株材料进行组织培养与植株再生,通过体细胞无性系变异增强和巩固SAR性状,获得SAR组成性表达的...用烟草花叶病毒弱毒株系N14预先接种,可以诱导烟草对病原真菌赤星病菌的系统获得性抗性(SAR),减轻病菌引起的赤星病。选择表现诱导抗性最强植株材料进行组织培养与植株再生,通过体细胞无性系变异增强和巩固SAR性状,获得SAR组成性表达的突变体(constitutive expresser of SAR)ces2-1。除了抗病表型,ces2-1还组成性表达多种防卫反应基因。回交实验与遗传分析表明,ces2-1是在野生型位点上的单基因显性突变。对ces2-1与野生型进行mRNA差异显示分析,得到一个ces2-1独有、在野生型中缺少的转录本,与前人报道的烟草受过氧化氢诱导的一个基因片段同源。用cDNA末端快速扩增技术克隆了这个转录本的全长序列,根据生物信息学分析与功能的初步测定,把这个基因命名为烟草受过氧化氢诱导的抗病相关基因(hydrogen peroxide-induced1,NtHPI1)。展开更多
Mammalian carboxylesterases(CEs) are key enzymes from the serine hydrolase superfamily.In the human body, two predominant carboxylesterases(CES1 and CES2) have been identified and extensively studied over the past dec...Mammalian carboxylesterases(CEs) are key enzymes from the serine hydrolase superfamily.In the human body, two predominant carboxylesterases(CES1 and CES2) have been identified and extensively studied over the past decade. These two enzymes play crucial roles in the metabolism of a wide variety of endogenous esters, ester-containing drugs and environmental toxicants. The key roles of CES in both human health and xenobiotic metabolism arouse great interest in the discovery of potent CES modulators to regulate endobiotic metabolism or to improve the efficacy of ester drugs. This review covers the structural and catalytic features of CES, tissue distributions, biological functions, genetic polymorphisms, substrate specificities and inhibitor properties of CES1 and CES2, as well as the significance and recent progress on the discovery of CES modulators. The information presented here will help pharmacologists explore the relevance of CES to human diseases or to assign the contribution of certain CES in xenobiotic metabolism. It will also facilitate medicinal chemistry efforts to design prodrugs activated by a given CES isoform, or to develop potent and selective modulators of CES for potential biomedical applications.展开更多
The mammalian carboxylesterase 1(Ces1/CES1)family comprises several enzymes that hydrolyze many xenobiotic chemicals and endogenous lipids.To investigate the pharmacological and physiological roles of Ces1/CES1,we gen...The mammalian carboxylesterase 1(Ces1/CES1)family comprises several enzymes that hydrolyze many xenobiotic chemicals and endogenous lipids.To investigate the pharmacological and physiological roles of Ces1/CES1,we generated Ces1 cluster knockout(Ces1^(-/-))mice,and a hepatic human CES1 transgenic model in the Ces1^(-/-)background(TgCES1).Ces1^(-/-)mice displayed profoundly decreased conversion of the anticancer prodrug irinotecan to SN-38 in plasma and tissues.TgCES1 mice exhibited enhanced metabolism of irinotecan to SN-38 in liver and kidney.Ces1 and hCES1 activity increased irinotecan toxicity,likely by enhancing the formation of pharmacodynamically active SN-38.Ces1^(-/-)mice also showed markedly increased capecitabine plasma exposure,which was moderately decreased in TgCES1 mice.Ces1^(-/-)mice were overweight with increased adipose tissue,white adipose tissue inflammation(in males),a higher lipid load in brown adipose tissue,and impaired blood glucose tolerance(in males).These phenotypes were mostly reversed in TgCES1 mice.TgCES1 mice displayed increased triglyceride secretion from liver to plasma,together with higher triglyceride levels in the male liver.These results indicate that the carboxylesterase 1 family plays essential roles in drug and lipid metabolism and detoxification.Ces1^(-/-)and TgCES1 mice will provide excellent tools for further study of the in vivo functions of Ces1/CES1 enzymes.展开更多
Cholesterol is an important precursor of many endogenous molecules.Disruption of cholesterol homeostasis can cause many pathological changes,leading to liver and cardiovascular diseases.CYP1A is widely involved in cho...Cholesterol is an important precursor of many endogenous molecules.Disruption of cholesterol homeostasis can cause many pathological changes,leading to liver and cardiovascular diseases.CYP1A is widely involved in cholesterol metabolic network,but its exact function has not been fully elucidated.Here,we aim to explore how CYP1A regulates cholesterol homeostasis.Our data showed that CYP1A1/2 knockout(KO)rats presented cholesterol deposition in blood and liver.The serum levels of low-density lipoprotein cholesterol,high-density lipoprotein cholesterol and total cholesterol were significantly increased in KO rats.Further studies found that the lipogenesis pathway(LXRa-SREBP1-SCD1)of KO rats was activated,and the key protein of cholesterol ester hydrolysis(CES1)was inhibited.Importantly,lansoprazole can significantly alleviate rat hepatic lipid deposition in hypercholesterolemia models by inducing CYP1A.Our findings reveal the role of CYP1A as a potential regulator of cholesterol homeostasis and provide a new perspective for the treatment of hypercholesterolemia.展开更多
文摘目的探讨乙型肝炎病毒DNA聚合酶反式调节蛋白1(HBV DNA polymerase trans activated protein 1,HBVDNAPTP1)在羧酸酯酶1(carboxylesterase 1,CES1)介导的单核细胞凋亡信号通路中的作用机制。方法利用Phyre2在线工具预测得到CES1三级结构,分别用pCMV-Myc载体构建HBVDNAPTP1基因、pCMV-HA载体构建CES1基因及其截短体219-264aa和265-303aa的真核细胞表达质粒。分别转染细胞后通过免疫荧光和免疫共沉淀法确定HBVDNAPTP1和CES1互相作用的区段。后通过HBVDNAPTP1和CES1单独转染及共转染,检测其对细胞凋亡率的影响;同时在分离人外周血单个核细胞(Peripheral blood mononuclear cell,PBMC)上转染上述质粒,以确定CES1和HBVDNAPTP1对单核细胞形态的影响。利用qRT-PCR和Western blot检测不同质粒转染后下游Janus激酶-1(Janus kinase,JAK1)和信号传导及转录激活蛋白3(Signal transducer and activator of transcription 3,STAT3)的基因和蛋白的表达情况。结果HBVDNAPTP1与CES1有靶向关系,其中HBVDNAPTP1与CES1全长靶向结合效果最好,故后续实验以CES1全长序列为对象开展。CCK-8和PBMC电镜实验结果表明,与Control组和NC组相比,ov-HBVDNAPTP1组、ov-CES1组和ov-HBVDNAPTP1+ov-CES1组THP-1细胞的凋亡率均升高(P<0.001),细胞受损;与ov-HBVDNAPTP1组和ov-CES1组相比,ov-HBVDNAPTP1+ov-CES1组细胞凋亡率进一步升高(P<0.01),凋亡细胞、坏死细胞的数量增多。qRT-PCR和Western blot结果显示,HBVDNAPTP1和CES1均可以在转录水平和翻译水平上显著上调JAK1和STAT3的表达,且二者同时作用时上调趋势更加明显。结论本研究证实HBVDNAPTP1与CES1二者可协同抑制THP-1细胞增殖并诱导细胞损伤;还可协同上调JAK1/STAT3信号通路在转录和蛋白水平的表达来调控单核细胞凋亡。该发现为深入理解HBV感染相关免疫调控机制提供了新的理论依据。
基金supported by the National Natural Science Foundation of China(No.82160386)the Guangxi Natural Science Foundation of China(No.2024GXNSFDA010032,2023GXNSFAA026189).
文摘Background:Head and neck squamous cell carcinoma(HNSCC)is a prevalent form of cancer globally,with chemoresistance posing a major challenge in treatment outcomes.The efficacy of the commonly used chemotherapeutic agent,cisplatin,is diminished in patients with poor prognoses.Methods:Various bioinformatics databases were utilized to examine Carboxylesterase 1(CES1)gene expression,clinicopathologic features,patient survival analysis,and gene function.An organoid model of HNSCC was established,along with the induction of drug-resistant HNSCC in the organoid model.CES1 expression was assessed using qRT-PCR and Western Blot,and differential markers were identified through transcriptome sequencing.Knockdown and overexpression models of CES1 were created in SCC-9 and patient-derived organoid(PDO)cells using shRNA and lentivirus to investigate the tumor biology and cisplatin resistance associated with CES1.Results:Research in bioinformatics has uncovered a strong correlation between the expression level of CES1 and the prognosis of HNSCC.The data suggests a significant link between CES1 expression and tobacco smoking.RNA-sequencing revealed a notable increase in CES1 expression in HNSCC-PDOcis-R cells compared to the parental PDO cells.Subsequently,we performed in vitro studies by HNSCC-PDO and SCC-9 and found that CES1-overexpressing cells exhibited reduced sensitivity to cisplatin and stronger tumor malignant biological behavior compared with CES1-knockdown cells.Conclusion:The observed association between CES1 expression and tobacco smoking implies a potential influence of smoking on the efficacy of cisplatin-based chemotherapy in HNSCC through the regulation of CES1 expression.
基金supported by Health and Medical Research Fund(Reference No.:12131521)from Food and Health Bureau,the Government of the Hong Kong SAR,Hong Kong,ChinaNational Natural Science Foundation of China(Grant No.:81973286,81922070,81703604 and 81973393),ChinaGeneral Research Fund(CUHK2141142)from University Grant Council of Hong Kong SAR,China。
文摘In traditional Chinese medicine herbs(TCM),including Radix Salviae Miltiorrhizae(Danshen),Radix Puerariae Lobatae(Gegen),Radix Angelicae Sinensis(Danggui),and Rhizoma Chuanxiong(Chuanxiong)are widely used for the prevention and treatment of cardiovascular diseases and also often co-administered with Western drugs as a part of integrative medicine practice.Carboxylesterase 1(CES1)plays a pivotal role in the metabolisms of pro-drugs,Since(S)-2-(2-(6-dimethylamino)-benzothiazole)-4,5-dihydrothiazole-4-carboxylate(NLMe)has recently been identified by us as a selective CES1 bioluminescent sensor,we developed a rapid method using this substrate for the direct measurement of CES1 activity in rats.This bioluminescence assay was applied to determine CES1 activity in rat tissues after a two-week oral administration of each of the four herbs noted above.The results demonstrated the presence of CES1 enzyme in rat blood and all tested tissues with much higher enzyme activity in the blood,liver,kidney and heart than that in the small intestine,spleen,lung,pancreas,brain and stomach.In addition,the four herbs showed tissue-specific effects on rat CES1 expression.Based on the CES1 biodistribution and its changes after treatment in rats,the possibility that Danshen,Gegen and Danggui might alter CES1 activities in human blood and kidney should be considered.In summary,a selective and sensitive bioluminescence assay was developed to rapidly evaluate CES1 activity and the effects of orally administered TCMs in rats.
文摘用烟草花叶病毒弱毒株系N14预先接种,可以诱导烟草对病原真菌赤星病菌的系统获得性抗性(SAR),减轻病菌引起的赤星病。选择表现诱导抗性最强植株材料进行组织培养与植株再生,通过体细胞无性系变异增强和巩固SAR性状,获得SAR组成性表达的突变体(constitutive expresser of SAR)ces2-1。除了抗病表型,ces2-1还组成性表达多种防卫反应基因。回交实验与遗传分析表明,ces2-1是在野生型位点上的单基因显性突变。对ces2-1与野生型进行mRNA差异显示分析,得到一个ces2-1独有、在野生型中缺少的转录本,与前人报道的烟草受过氧化氢诱导的一个基因片段同源。用cDNA末端快速扩增技术克隆了这个转录本的全长序列,根据生物信息学分析与功能的初步测定,把这个基因命名为烟草受过氧化氢诱导的抗病相关基因(hydrogen peroxide-induced1,NtHPI1)。
基金supported by the National Key Research and Development Program of China (2016YFC1303900,2017YFC1700200,2017YFC1702000)the National Scientific and Technological Major Projects of China (2017ZX09101004)+2 种基金the National Natural Science Foundation of China (81703604,81773687,21602219,81573501 and 81473181)Program of Shanghai Academic/Technology Research Leader (18XD1403600)the Innovative Entrepreneurship Program of High-level Talents in Dalian (2016RQ025)
文摘Mammalian carboxylesterases(CEs) are key enzymes from the serine hydrolase superfamily.In the human body, two predominant carboxylesterases(CES1 and CES2) have been identified and extensively studied over the past decade. These two enzymes play crucial roles in the metabolism of a wide variety of endogenous esters, ester-containing drugs and environmental toxicants. The key roles of CES in both human health and xenobiotic metabolism arouse great interest in the discovery of potent CES modulators to regulate endobiotic metabolism or to improve the efficacy of ester drugs. This review covers the structural and catalytic features of CES, tissue distributions, biological functions, genetic polymorphisms, substrate specificities and inhibitor properties of CES1 and CES2, as well as the significance and recent progress on the discovery of CES modulators. The information presented here will help pharmacologists explore the relevance of CES to human diseases or to assign the contribution of certain CES in xenobiotic metabolism. It will also facilitate medicinal chemistry efforts to design prodrugs activated by a given CES isoform, or to develop potent and selective modulators of CES for potential biomedical applications.
基金funded in part by the China Scholarship Council(CSC Scholarship No.201506240145 to Changpei Gan)。
文摘The mammalian carboxylesterase 1(Ces1/CES1)family comprises several enzymes that hydrolyze many xenobiotic chemicals and endogenous lipids.To investigate the pharmacological and physiological roles of Ces1/CES1,we generated Ces1 cluster knockout(Ces1^(-/-))mice,and a hepatic human CES1 transgenic model in the Ces1^(-/-)background(TgCES1).Ces1^(-/-)mice displayed profoundly decreased conversion of the anticancer prodrug irinotecan to SN-38 in plasma and tissues.TgCES1 mice exhibited enhanced metabolism of irinotecan to SN-38 in liver and kidney.Ces1 and hCES1 activity increased irinotecan toxicity,likely by enhancing the formation of pharmacodynamically active SN-38.Ces1^(-/-)mice also showed markedly increased capecitabine plasma exposure,which was moderately decreased in TgCES1 mice.Ces1^(-/-)mice were overweight with increased adipose tissue,white adipose tissue inflammation(in males),a higher lipid load in brown adipose tissue,and impaired blood glucose tolerance(in males).These phenotypes were mostly reversed in TgCES1 mice.TgCES1 mice displayed increased triglyceride secretion from liver to plasma,together with higher triglyceride levels in the male liver.These results indicate that the carboxylesterase 1 family plays essential roles in drug and lipid metabolism and detoxification.Ces1^(-/-)and TgCES1 mice will provide excellent tools for further study of the in vivo functions of Ces1/CES1 enzymes.
基金supported in whole or part by grants from the National Natural Science Foundation of China(81773808,82274010)the Science and Technology Commission of Shanghai Municipality(18430760400,China)+4 种基金the Jointed PI Program from Shanghai Changning Maternity and Infant Health Hospital(2019CNECNUPI02,China)the Fundamental Research Funds for the Central Universities(China)ECNU Construction Fund of Innovation and Entrepreneurship Laboratory(China)supported from ECNU Multifunctional Platform for Innovation(011,China)the Instruments Sharing Platform of School of Life Sciences,East China Normal University(Shanghai,China)。
文摘Cholesterol is an important precursor of many endogenous molecules.Disruption of cholesterol homeostasis can cause many pathological changes,leading to liver and cardiovascular diseases.CYP1A is widely involved in cholesterol metabolic network,but its exact function has not been fully elucidated.Here,we aim to explore how CYP1A regulates cholesterol homeostasis.Our data showed that CYP1A1/2 knockout(KO)rats presented cholesterol deposition in blood and liver.The serum levels of low-density lipoprotein cholesterol,high-density lipoprotein cholesterol and total cholesterol were significantly increased in KO rats.Further studies found that the lipogenesis pathway(LXRa-SREBP1-SCD1)of KO rats was activated,and the key protein of cholesterol ester hydrolysis(CES1)was inhibited.Importantly,lansoprazole can significantly alleviate rat hepatic lipid deposition in hypercholesterolemia models by inducing CYP1A.Our findings reveal the role of CYP1A as a potential regulator of cholesterol homeostasis and provide a new perspective for the treatment of hypercholesterolemia.
