目的:探讨双特异性酪氨酸磷酸化调节激酶2(dual-specificity tyrosine phosphorylation-regulated kinase2,DYRK2)在胰腺癌中的表达及其临床意义.方法:应用实时荧光定量PCR、Western blot及免疫组织化学技术检测40例胰腺癌及其癌旁正常...目的:探讨双特异性酪氨酸磷酸化调节激酶2(dual-specificity tyrosine phosphorylation-regulated kinase2,DYRK2)在胰腺癌中的表达及其临床意义.方法:应用实时荧光定量PCR、Western blot及免疫组织化学技术检测40例胰腺癌及其癌旁正常组织标本中DYRK2mRNA和蛋白的表达量,并结合免疫组织化学结果及胰腺癌患者的临床病理资料进行统计学分析.结果:实时荧光定量PCR检测显示,DYRK2m R N A在胰腺癌及其癌旁组织中均有表达,但后者的表达水平明显高于前者(P<0.01);Western blot检测显示,在88.9%的癌组织中D Y R K2蛋白的表达量低于其对应的癌旁组织;免疫组织化学染色显示,DYRK2在胰腺癌组织中的阳性比例明显低于其癌旁组织(42.5%vs87.5%,X2=17.802,P<0.01),且DYRK2的表达与胰腺癌有无淋巴结转移相关(X2=6.32,P<0.05).结论:DYRK2在胰腺癌组织中表达下调,可能与胰腺癌的发生发展和淋巴结转移等恶性生物学行为相关.展开更多
目的探讨双特异性酪氨酸磷酸化调节激酶1b(DYRK1b)在宫颈病变组织和癌细胞中的表达情况及其作用.方法选取宫颈癌患者127例为研究组,同期确诊为慢性宫颈炎患者32例为对照组,收集上述患者石蜡组织标本,免疫组化SP法检测DYRK1b蛋白表达情况...目的探讨双特异性酪氨酸磷酸化调节激酶1b(DYRK1b)在宫颈病变组织和癌细胞中的表达情况及其作用.方法选取宫颈癌患者127例为研究组,同期确诊为慢性宫颈炎患者32例为对照组,收集上述患者石蜡组织标本,免疫组化SP法检测DYRK1b蛋白表达情况.依据宫颈癌细胞系He La 229和Si Ha细胞将标本分为实验组(加DYRK1b抑制剂Az191)、对照组(不加Az191),Western blod法检测细胞中DYRK1蛋白表达情况,MTT法检测细胞增殖情况,流式细胞仪检测细胞凋亡情况.结果子宫颈鳞癌阳性表达率明显高于慢性宫颈炎、低级别鳞状上皮内病变、高级别鳞状上皮内病变,差异均具有统计学意义(P<0.01);高级别鳞状上皮内病变明显高于慢性宫颈炎、低级别鳞状上皮内病变,差异具有统计学意义(P<0.05).在He La 229和Si Ha细胞中,DYRK1b蛋白表达量随Az191剂量增加而减少,且均明显低于对照组,差异具有统计学意义(P<0.01).研究组He La 229和Si Ha细胞抑制率随Az191浓度增加而提升,均明显高于对照组,差异具有统计学意义(P<0.05).10μmol/L的Az191作用后,研究组He La 229和Si Ha细胞凋亡率明显高于对照组,差异具有统计学意义(P<0.05).结论宫颈病变过程中DYRK1b蛋白表达水平逐渐提升,宫颈癌组织和细胞中均呈高表达;下调DYRK1b蛋白表达后可抑制宫颈癌细胞增殖,促进凋亡.展开更多
Objective:Mcl-1 overexpression confers acquired resistance to Bcl-2 inhibitors in non-small cell lung cancer(NSCLC),but no direct Mcl-1 inhibitor is currently available for clinical use.Thus,novel therapeutic strategi...Objective:Mcl-1 overexpression confers acquired resistance to Bcl-2 inhibitors in non-small cell lung cancer(NSCLC),but no direct Mcl-1 inhibitor is currently available for clinical use.Thus,novel therapeutic strategies are urgently needed to target Mcl-1 and sensitize the anti-NSCLC activity of Bcl-2 inhibitors.Methods:Cell proliferation was measured using sulforhodamine B and colony formation assays,and apoptosis was detected with Annexin V-FITC staining.Gene expression was manipulated using siRNAs and plasmids.Real-time PCR and Western blot were used to measure mRNA and protein levels.Immunoprecipitation and immunofluorescence were used to analyze co-localization o f dual specificity tyrosine-phosphorylation-regulated kinase 1A(DYRK1A)and Mcl-1.Results:Suppression of DYRK1A resulted in reduced Mcl-1 expression in NSCLC cells,whereas overexpression of DYRK1A significantly increased Mcl-1 expression.Suppression of DYRK1A did not alter Mcl-1 mRNA levels,but did result in an accelerated degradation o f Mcl-1 protein in NSCLC cells.Furthermore,DYRK1A mediated proteasome-dependent degradation o f Mcl-1 in NSCLC cells,and DYRK1A co-localized with Mcl-1 in NSCLC cells and was co-expressed with Mcl-1 in tumor samples from lung cancer patients,suggesting that Mcl-1 may be a novel DYRK1A substrate.We showed that combined therapy with harmine and Bcl-2 antagonists significantly inhibited cell proliferation and induced apoptosis in NSCLC cell lines as well as primary NSCLC cells.Conclusions:Mcl-1 is a novel DYRK1A substrate,and the role of DYRK1A in promoting Mcl-1 stability makes it an attractive target for decreasing Bcl-2 inhibitor resistance.展开更多
The dual-specificity tyrosine-regulated kinase (DYRK) family is evolutionarily conserved from invertebrate to mammals. DYRKs regulate cell proliferation, apoptosis, survival, and differentiation by modifying the prote...The dual-specificity tyrosine-regulated kinase (DYRK) family is evolutionarily conserved from invertebrate to mammals. DYRKs regulate cell proliferation, apoptosis, survival, and differentiation by modifying the protein activation state, cellular localization, and turnover. In contrast to several studies in cellular models, the available evidence regarding the in vivo roles of DYRKs in mammalian development is limited. This review summarizes the in vivo studies on Dyrks which provide insight into their roles in mammalian tissue development and congenital diseases. In vivo evidence obtained using knockout and genetically modified animals helps to understand and develop novel clinical therapies and drug for human congenital diseases, such as Down syndrome and neuronal disorders (associated with DYRK1A) and skeletal ciliopathies (associated with DYRK2).展开更多
文摘目的:探讨双特异性酪氨酸磷酸化调节激酶2(dual-specificity tyrosine phosphorylation-regulated kinase2,DYRK2)在胰腺癌中的表达及其临床意义.方法:应用实时荧光定量PCR、Western blot及免疫组织化学技术检测40例胰腺癌及其癌旁正常组织标本中DYRK2mRNA和蛋白的表达量,并结合免疫组织化学结果及胰腺癌患者的临床病理资料进行统计学分析.结果:实时荧光定量PCR检测显示,DYRK2m R N A在胰腺癌及其癌旁组织中均有表达,但后者的表达水平明显高于前者(P<0.01);Western blot检测显示,在88.9%的癌组织中D Y R K2蛋白的表达量低于其对应的癌旁组织;免疫组织化学染色显示,DYRK2在胰腺癌组织中的阳性比例明显低于其癌旁组织(42.5%vs87.5%,X2=17.802,P<0.01),且DYRK2的表达与胰腺癌有无淋巴结转移相关(X2=6.32,P<0.05).结论:DYRK2在胰腺癌组织中表达下调,可能与胰腺癌的发生发展和淋巴结转移等恶性生物学行为相关.
文摘目的探讨双特异性酪氨酸磷酸化调节激酶1b(DYRK1b)在宫颈病变组织和癌细胞中的表达情况及其作用.方法选取宫颈癌患者127例为研究组,同期确诊为慢性宫颈炎患者32例为对照组,收集上述患者石蜡组织标本,免疫组化SP法检测DYRK1b蛋白表达情况.依据宫颈癌细胞系He La 229和Si Ha细胞将标本分为实验组(加DYRK1b抑制剂Az191)、对照组(不加Az191),Western blod法检测细胞中DYRK1蛋白表达情况,MTT法检测细胞增殖情况,流式细胞仪检测细胞凋亡情况.结果子宫颈鳞癌阳性表达率明显高于慢性宫颈炎、低级别鳞状上皮内病变、高级别鳞状上皮内病变,差异均具有统计学意义(P<0.01);高级别鳞状上皮内病变明显高于慢性宫颈炎、低级别鳞状上皮内病变,差异具有统计学意义(P<0.05).在He La 229和Si Ha细胞中,DYRK1b蛋白表达量随Az191剂量增加而减少,且均明显低于对照组,差异具有统计学意义(P<0.01).研究组He La 229和Si Ha细胞抑制率随Az191浓度增加而提升,均明显高于对照组,差异具有统计学意义(P<0.05).10μmol/L的Az191作用后,研究组He La 229和Si Ha细胞凋亡率明显高于对照组,差异具有统计学意义(P<0.05).结论宫颈病变过程中DYRK1b蛋白表达水平逐渐提升,宫颈癌组织和细胞中均呈高表达;下调DYRK1b蛋白表达后可抑制宫颈癌细胞增殖,促进凋亡.
基金the National Natural Science Foundation of China(Grant Nos.81702887,81272473)the Key Laboratory of Clinical Cancer Pharmacology and Toxicology Research of Zhejiang Province(Grant No.2020E10021)+6 种基金the Key Medical Disciplines of Zhejiang Province(Grant No.2018-2-3)the Hangzhou Major Science and Technology Project(Grant No.20172016A01)the Zhejiang Provincial Foundation of Natural Science(Grant No.LY19H310004)the Scientific and Technological Developing Scheme of Hangzhou City(Grant No.20191203B49)the Science Research Foundation of Zhejiang Health Bureau(Grant No.2020RC026)the High-level Talents Coming Back from Abroad Innovation,the Entrepreneurship Program in Hangzhouand the Teachers Research Fund of Zhejiang University City College(Grant No.J-19006).
文摘Objective:Mcl-1 overexpression confers acquired resistance to Bcl-2 inhibitors in non-small cell lung cancer(NSCLC),but no direct Mcl-1 inhibitor is currently available for clinical use.Thus,novel therapeutic strategies are urgently needed to target Mcl-1 and sensitize the anti-NSCLC activity of Bcl-2 inhibitors.Methods:Cell proliferation was measured using sulforhodamine B and colony formation assays,and apoptosis was detected with Annexin V-FITC staining.Gene expression was manipulated using siRNAs and plasmids.Real-time PCR and Western blot were used to measure mRNA and protein levels.Immunoprecipitation and immunofluorescence were used to analyze co-localization o f dual specificity tyrosine-phosphorylation-regulated kinase 1A(DYRK1A)and Mcl-1.Results:Suppression of DYRK1A resulted in reduced Mcl-1 expression in NSCLC cells,whereas overexpression of DYRK1A significantly increased Mcl-1 expression.Suppression of DYRK1A did not alter Mcl-1 mRNA levels,but did result in an accelerated degradation o f Mcl-1 protein in NSCLC cells.Furthermore,DYRK1A mediated proteasome-dependent degradation o f Mcl-1 in NSCLC cells,and DYRK1A co-localized with Mcl-1 in NSCLC cells and was co-expressed with Mcl-1 in tumor samples from lung cancer patients,suggesting that Mcl-1 may be a novel DYRK1A substrate.We showed that combined therapy with harmine and Bcl-2 antagonists significantly inhibited cell proliferation and induced apoptosis in NSCLC cell lines as well as primary NSCLC cells.Conclusions:Mcl-1 is a novel DYRK1A substrate,and the role of DYRK1A in promoting Mcl-1 stability makes it an attractive target for decreasing Bcl-2 inhibitor resistance.
基金supported by JSPS KAKENHI(No.21K06192 to S.Y.and 17H03584,18K19484,and 20H03519 to K.Y.)the Jikei University Research Fund(to K.Y.)+1 种基金the Takeda Science Foundation(to S.Y.)the Uehara Memorial Foundation(to S.Y.and K.Y.).
文摘The dual-specificity tyrosine-regulated kinase (DYRK) family is evolutionarily conserved from invertebrate to mammals. DYRKs regulate cell proliferation, apoptosis, survival, and differentiation by modifying the protein activation state, cellular localization, and turnover. In contrast to several studies in cellular models, the available evidence regarding the in vivo roles of DYRKs in mammalian development is limited. This review summarizes the in vivo studies on Dyrks which provide insight into their roles in mammalian tissue development and congenital diseases. In vivo evidence obtained using knockout and genetically modified animals helps to understand and develop novel clinical therapies and drug for human congenital diseases, such as Down syndrome and neuronal disorders (associated with DYRK1A) and skeletal ciliopathies (associated with DYRK2).
