为了研究注射外源性神经介素B(NMB)对小鼠睾丸中NMB及其受体和睾酮合成关键酶(St AR、CYP11A1和3β-HSD)表达及血清中睾酮浓度的影响,试验将40只雄性ICR小鼠随机分成12 h对照组(12 h Control组)、12 h试验组(12 h NMB组)、24 h对照组(24...为了研究注射外源性神经介素B(NMB)对小鼠睾丸中NMB及其受体和睾酮合成关键酶(St AR、CYP11A1和3β-HSD)表达及血清中睾酮浓度的影响,试验将40只雄性ICR小鼠随机分成12 h对照组(12 h Control组)、12 h试验组(12 h NMB组)、24 h对照组(24 h Control组)和24 h试验组(24 h NMB组),每组10只,2个对照组和2个试验组分别按体重10μL/g腹腔注射生理盐水和10 nmol/L NMB溶液;称量注射前后小鼠体重,计算饲料用量,并观察小鼠行为特征;各组小鼠分别于注射后12小时、24小时摘眼球采血并处死取睾丸,通过ELISA法检测小鼠血清中睾酮的浓度;通过实时荧光定量PCR(q PCR)和蛋白印迹法(Western-blot)检测小鼠睾丸中NMB、NMBR、St AR、CYP11A1和3β-HSD m RNA和蛋白表达量。结果表明:与12 h Control组相比,12 h NMB组小鼠行为无明显异常,体重增量、总摄食量略有减少,料重比略有增高,但差异不显著(P>0.05);NMB、NMBR、St AR、CYP11A1和3β-HSD m RNA及蛋白表达量均差异不显著(P>0.05);血清中睾酮浓度差异不显著(P>0.05)。与24 h Control组相比,24 h NMB组小鼠被毛粗乱、脾气暴躁,体重增量显著减少(P<0.05);总摄食量减少,料重比略有增高,但均差异不显著(P>0.05);NMB m RNA和蛋白表达量极显著降低(P<0.01),NMBR、St AR、CYP11A1和3β-HSD m RNA及蛋白表达量均差异不显著(P>0.05);血清中睾酮浓度差异不显著(P>0.05)。与12 h NMB组相比,24 h NMB组NMBR m RNA表达量极显著降低(P<0.01),24 h NMB组NMBR蛋白表达量显著增加(P<0.05)。说明小鼠腹腔注射NMB能够抑制小鼠摄食,但对小鼠睾丸中睾酮的合成和分泌影响不大。展开更多
Neuromedin B(NMB)is a member of mammalian bombesin-like peptide family and almost all its potent biological actions take place exclusively through its receptor NMBR,which has been identified as a G-protein couple rece...Neuromedin B(NMB)is a member of mammalian bombesin-like peptide family and almost all its potent biological actions take place exclusively through its receptor NMBR,which has been identified as a G-protein couple receptor with seven trans-membrane regions.NMB and NMBR represent an interesting signaling axis over which NMB/NMBR perform multiple biological activities,in particularly,they have been highlighted in the regulation of the immunology system.This review aims to provide a brief overview of the recent advancements in our understanding regarding the potential immunological functions of NMB and NMBR in cell proliferation,tumorigenesis,angiogenesis,neurogenic inflammation,anti-influenza A virus(IAV)infections,and briefly discusses the potential importance of these observations.All these implicated that targeting NMBR or developing analogues of NMB may represent a useful strategy for development of new anti-IAV agents.展开更多
目的:探讨神经调节素B受体(neuromedin B receptor,NMBR)在妊娠小鼠子宫平滑肌细胞中mRNA、蛋白时空表达的变化及其与分娩启动的关系和机制。方法:按妊娠阶段的不同,将小鼠分为未孕组、早孕组、中孕组、晚孕组、产时组和产后组,每组12...目的:探讨神经调节素B受体(neuromedin B receptor,NMBR)在妊娠小鼠子宫平滑肌细胞中mRNA、蛋白时空表达的变化及其与分娩启动的关系和机制。方法:按妊娠阶段的不同,将小鼠分为未孕组、早孕组、中孕组、晚孕组、产时组和产后组,每组12例。应用半定量RT-PCR和Western免疫印迹技术分别检测子宫平滑肌中NMBR,IL-6和HSP70mRNA及蛋白表达;利用NoShift转录因子分析法检测NF-κB-P65的DNA结合活性,并分析各指标与分娩的关系及其相关性。结果:产时组NMBRmRNA的相对表达量显著高于未孕、早孕、晚孕、产后组(P<0.05),但与中孕组相比差异无统计学意义(P>0.05);产时组NMBR蛋白表达量显著高于其它各组(P<0.01)。NF-κB-P65DNA结合活性产时组显著高于未孕、晚孕和产后组(P<0.05)。IL-6mRNA表达产时组显著高于未孕、晚孕和产后组(P<0.05),其蛋白表达产时组亦显著高于未孕和产后组(P<0.05)。HSP70mRNA表达量产时组显著高于未孕和产后组(P<0.05),HSP70蛋白的表达量产时组明显高于晚孕、未孕组(P<0.05)。临产前后子宫平滑肌细胞P65DNA结合活性、IL-6mRNA的表达均与NMBRmRNA的变化呈正相关(r=0.40,P<0.01;r=0.30,P<0.05);P65,HSP70mRNA的变化亦与NMBR的变化呈正相关(r=0.40,P<0.01;r=0.49,P<0.01);但HSP70与P65无相关性。结论:子宫平滑肌细胞中NMBRmRNA和蛋白表达在临产时达最高峰。NMBR可借助NF-κB P65-IL-6通路、通过调节HSP70表达参与分娩发动和维持,但其对HSP70表达的影响不依赖P65途径。展开更多
TPP-Ⅰ(tripeptidyl peptidase-Ⅰ) protein turnover was studied by observing the role of rat TPP-Ⅰ in neuromedin B(NMB) and other neuropeptides degradations in some physiological situations. The mixtures of rat TPP-Ⅰ ...TPP-Ⅰ(tripeptidyl peptidase-Ⅰ) protein turnover was studied by observing the role of rat TPP-Ⅰ in neuromedin B(NMB) and other neuropeptides degradations in some physiological situations. The mixtures of rat TPP-Ⅰ with each of NMB, other neuropeptides and Ala-Ala-Phe-MCA were made respectively under the same conditions. The reaction was observed at different timeand monitored by means of high performance liquid chromatography(HPLC) and time-of-flight mass spectrometry(TOF-MS) in vitro. NMB was broken down at the same degree as Ala-Ala-Phe-MCA by rat TPP-Ⅰ and Gly-Asn-Leu was released within 16 h, but other neuropeptides were not digested within 24 h. TPP-Ⅰ is the predominant proteolytic enzyme responsible for the intracellular degradation of neuromedin B. NMB has recently been found to be a good natural substrate for rat lysosomal TPP-Ⅰ.展开更多
神经介素B(neuromedin B,NMB)及受体(NMB receptor,NMBR)通过NF-κB信号通路参与抑制甲型流感病毒(influenza A virus,IAV)H1N1亚型(IAV/H1N1)的感染。但关于NMB和NMBR对NF-κB信号通路相关泛素蛋白酶的调控如何,尚未见报道。为探究NMB...神经介素B(neuromedin B,NMB)及受体(NMB receptor,NMBR)通过NF-κB信号通路参与抑制甲型流感病毒(influenza A virus,IAV)H1N1亚型(IAV/H1N1)的感染。但关于NMB和NMBR对NF-κB信号通路相关泛素蛋白酶的调控如何,尚未见报道。为探究NMB和NMBR调控IAV诱导的NF-κB信号通路相关的泛素蛋白酶表达的影响,本研究基于IAV/H9N2感染sh-NMBR细胞与NMB刺激的A549细胞,采用RT-PCR、qRT-PCR及Western blot(WB)分析NMB和NMBR对H9N2感染引起的NF-κB信号通路相关的E3泛素连接酶Mind bomb-2(MIB2)和Ring Finger Protein 8(RNF8)及去泛素蛋白酶Cylindromatosis(CYLD)的表达变化。结果显示:H9N2感染sh-NMBR细胞中,RNF8和CYLD表达增加,MIB2表达和P65磷酸化水平降低;NMB激活A549细胞中NMBR的表达后,诱导细胞中RNF8和CYLD的表达水平下降,MIB2表达和P65磷酸化水平增加。结果表明:NMB和NMBR通过调控IAV/H9N2感染诱导的泛素蛋白酶的表达和P65活性,从而影响IAV/H9N2感染激活的NF-κB信号通路的功能,该结果为深入研究NMB和NMBR抑制甲型流感病毒感染的作用机制提供了理论基础。展开更多
目的构建小鼠神经介素B(neuromedin B,NMB)及其受体(neuromedin B receptor,NMBR)基因慢病毒过表达质粒,并于RAW264.7细胞中表达,以期为小鼠NMB和NMBR基因过表达对RAW264.7细胞增殖及凋亡影响的深入研究奠定基础。方法经RT-PCR法扩增获...目的构建小鼠神经介素B(neuromedin B,NMB)及其受体(neuromedin B receptor,NMBR)基因慢病毒过表达质粒,并于RAW264.7细胞中表达,以期为小鼠NMB和NMBR基因过表达对RAW264.7细胞增殖及凋亡影响的深入研究奠定基础。方法经RT-PCR法扩增获得小鼠NMB和NMBR基因的编码序列(coding sequence,CDS),插入载体pCD513B-1,构建重组质粒pCD513B-1-NMB和pCD513B-1-NMBR;于HEK-293T细胞包装获得小鼠NMB和NMBR基因过表达慢病毒,通过倍比稀释法检测病毒效价。用慢病毒感染RAW264.7细胞48 h后,qPCR法检测小鼠NMB和NMBR基因mRNA的表达情况,以未感染细胞为对照。结果经双酶切鉴定证明重组质粒构建正确。小鼠NMB和NMBR基因过表达慢病毒滴度分别为6×10~6和8×10~6TU/mL。2种慢病毒转染RAW264.7细胞中小鼠NMB和NMBR基因mRNA表达量显著高于对照组(t分别为24.158和14.958,P均<0.01)。结论成功构建了小鼠NMB和NMBR基因过表达质粒,能够显著提高RAW264.7细胞NMB和NMBR基因的表达量。展开更多
文摘为了研究注射外源性神经介素B(NMB)对小鼠睾丸中NMB及其受体和睾酮合成关键酶(St AR、CYP11A1和3β-HSD)表达及血清中睾酮浓度的影响,试验将40只雄性ICR小鼠随机分成12 h对照组(12 h Control组)、12 h试验组(12 h NMB组)、24 h对照组(24 h Control组)和24 h试验组(24 h NMB组),每组10只,2个对照组和2个试验组分别按体重10μL/g腹腔注射生理盐水和10 nmol/L NMB溶液;称量注射前后小鼠体重,计算饲料用量,并观察小鼠行为特征;各组小鼠分别于注射后12小时、24小时摘眼球采血并处死取睾丸,通过ELISA法检测小鼠血清中睾酮的浓度;通过实时荧光定量PCR(q PCR)和蛋白印迹法(Western-blot)检测小鼠睾丸中NMB、NMBR、St AR、CYP11A1和3β-HSD m RNA和蛋白表达量。结果表明:与12 h Control组相比,12 h NMB组小鼠行为无明显异常,体重增量、总摄食量略有减少,料重比略有增高,但差异不显著(P>0.05);NMB、NMBR、St AR、CYP11A1和3β-HSD m RNA及蛋白表达量均差异不显著(P>0.05);血清中睾酮浓度差异不显著(P>0.05)。与24 h Control组相比,24 h NMB组小鼠被毛粗乱、脾气暴躁,体重增量显著减少(P<0.05);总摄食量减少,料重比略有增高,但均差异不显著(P>0.05);NMB m RNA和蛋白表达量极显著降低(P<0.01),NMBR、St AR、CYP11A1和3β-HSD m RNA及蛋白表达量均差异不显著(P>0.05);血清中睾酮浓度差异不显著(P>0.05)。与12 h NMB组相比,24 h NMB组NMBR m RNA表达量极显著降低(P<0.01),24 h NMB组NMBR蛋白表达量显著增加(P<0.05)。说明小鼠腹腔注射NMB能够抑制小鼠摄食,但对小鼠睾丸中睾酮的合成和分泌影响不大。
基金National Science Foundation of China(U1805231)National Program for Innovation and Entrepreneurship Training of University Students(202110389008)+2 种基金Special Fund Project for Scientific and Technological Innovation of Fujian Agriculture and Forestry University(CXZX2018015,CXZX2020060A)Key Laboratory Project of Animal Infectious Disease Control and Biotechnology in Fujian Province(2018KF03)National Science Foundation of Fujian Province(2020J01539).
文摘Neuromedin B(NMB)is a member of mammalian bombesin-like peptide family and almost all its potent biological actions take place exclusively through its receptor NMBR,which has been identified as a G-protein couple receptor with seven trans-membrane regions.NMB and NMBR represent an interesting signaling axis over which NMB/NMBR perform multiple biological activities,in particularly,they have been highlighted in the regulation of the immunology system.This review aims to provide a brief overview of the recent advancements in our understanding regarding the potential immunological functions of NMB and NMBR in cell proliferation,tumorigenesis,angiogenesis,neurogenic inflammation,anti-influenza A virus(IAV)infections,and briefly discusses the potential importance of these observations.All these implicated that targeting NMBR or developing analogues of NMB may represent a useful strategy for development of new anti-IAV agents.
文摘目的:探讨神经调节素B受体(neuromedin B receptor,NMBR)在妊娠小鼠子宫平滑肌细胞中mRNA、蛋白时空表达的变化及其与分娩启动的关系和机制。方法:按妊娠阶段的不同,将小鼠分为未孕组、早孕组、中孕组、晚孕组、产时组和产后组,每组12例。应用半定量RT-PCR和Western免疫印迹技术分别检测子宫平滑肌中NMBR,IL-6和HSP70mRNA及蛋白表达;利用NoShift转录因子分析法检测NF-κB-P65的DNA结合活性,并分析各指标与分娩的关系及其相关性。结果:产时组NMBRmRNA的相对表达量显著高于未孕、早孕、晚孕、产后组(P<0.05),但与中孕组相比差异无统计学意义(P>0.05);产时组NMBR蛋白表达量显著高于其它各组(P<0.01)。NF-κB-P65DNA结合活性产时组显著高于未孕、晚孕和产后组(P<0.05)。IL-6mRNA表达产时组显著高于未孕、晚孕和产后组(P<0.05),其蛋白表达产时组亦显著高于未孕和产后组(P<0.05)。HSP70mRNA表达量产时组显著高于未孕和产后组(P<0.05),HSP70蛋白的表达量产时组明显高于晚孕、未孕组(P<0.05)。临产前后子宫平滑肌细胞P65DNA结合活性、IL-6mRNA的表达均与NMBRmRNA的变化呈正相关(r=0.40,P<0.01;r=0.30,P<0.05);P65,HSP70mRNA的变化亦与NMBR的变化呈正相关(r=0.40,P<0.01;r=0.49,P<0.01);但HSP70与P65无相关性。结论:子宫平滑肌细胞中NMBRmRNA和蛋白表达在临产时达最高峰。NMBR可借助NF-κB P65-IL-6通路、通过调节HSP70表达参与分娩发动和维持,但其对HSP70表达的影响不依赖P65途径。
文摘TPP-Ⅰ(tripeptidyl peptidase-Ⅰ) protein turnover was studied by observing the role of rat TPP-Ⅰ in neuromedin B(NMB) and other neuropeptides degradations in some physiological situations. The mixtures of rat TPP-Ⅰ with each of NMB, other neuropeptides and Ala-Ala-Phe-MCA were made respectively under the same conditions. The reaction was observed at different timeand monitored by means of high performance liquid chromatography(HPLC) and time-of-flight mass spectrometry(TOF-MS) in vitro. NMB was broken down at the same degree as Ala-Ala-Phe-MCA by rat TPP-Ⅰ and Gly-Asn-Leu was released within 16 h, but other neuropeptides were not digested within 24 h. TPP-Ⅰ is the predominant proteolytic enzyme responsible for the intracellular degradation of neuromedin B. NMB has recently been found to be a good natural substrate for rat lysosomal TPP-Ⅰ.
文摘神经介素B(neuromedin B,NMB)及受体(NMB receptor,NMBR)通过NF-κB信号通路参与抑制甲型流感病毒(influenza A virus,IAV)H1N1亚型(IAV/H1N1)的感染。但关于NMB和NMBR对NF-κB信号通路相关泛素蛋白酶的调控如何,尚未见报道。为探究NMB和NMBR调控IAV诱导的NF-κB信号通路相关的泛素蛋白酶表达的影响,本研究基于IAV/H9N2感染sh-NMBR细胞与NMB刺激的A549细胞,采用RT-PCR、qRT-PCR及Western blot(WB)分析NMB和NMBR对H9N2感染引起的NF-κB信号通路相关的E3泛素连接酶Mind bomb-2(MIB2)和Ring Finger Protein 8(RNF8)及去泛素蛋白酶Cylindromatosis(CYLD)的表达变化。结果显示:H9N2感染sh-NMBR细胞中,RNF8和CYLD表达增加,MIB2表达和P65磷酸化水平降低;NMB激活A549细胞中NMBR的表达后,诱导细胞中RNF8和CYLD的表达水平下降,MIB2表达和P65磷酸化水平增加。结果表明:NMB和NMBR通过调控IAV/H9N2感染诱导的泛素蛋白酶的表达和P65活性,从而影响IAV/H9N2感染激活的NF-κB信号通路的功能,该结果为深入研究NMB和NMBR抑制甲型流感病毒感染的作用机制提供了理论基础。
