BACKGROUND Diabetic retinopathy(DR)is the primary cause of visual problems in patients with diabetes.The Heyingwuzi formulation(HYWZF)is effective against DR.AIM To determine the HYWZF prevention mechanisms,especially...BACKGROUND Diabetic retinopathy(DR)is the primary cause of visual problems in patients with diabetes.The Heyingwuzi formulation(HYWZF)is effective against DR.AIM To determine the HYWZF prevention mechanisms,especially those underlying mitophagy.METHODS Human retinal capillary endothelial cells(HRCECs)were treated with high glucose(hg),HYWZF serum,PX-478,or Mdivi-1 in vitro.Then,cell counting kit-8,transwell,and tube formation assays were used to evaluate HRCEC proliferation,invasion,and tube formation,respectively.Transmission electron microscopy was used to assess mitochondrial morphology,and Western blotting was used to determine the protein levels.Flow cytometry was used to assess cell apoptosis,reactive oxygen species(ROS)production,and mitochondrial membrane potential.Moreover,C57BL/6 mice were established in vivo using streptozotocin and treated with HYWZF for four weeks.Blood glucose levels and body weight were monitored continuously.Changes in retinal characteristics were evaluated using hematoxylin and eosin,tar violet,and periodic acid-Schiff staining.Protein levels in retinal tissues were determined via Western blotting,immunohistochemistry,and immunostaining.RESULTS HYWZF inhibited excessive ROS production,apoptosis,tube formation,and invasion in hg-induced HRCECs via mitochondrial autophagy in vitro.It increased the mRNA expression levels of BCL2-interacting protein 3(BNIP3),FUN14 domain-containing 1,BNIP3-like(BNIP3L,also known as NIX),PARKIN,PTEN-induced kinase 1,and hypoxia-inducible factor(HIF)-1α.Moreover,it downregulated the protein levels of vascular endothelial cell growth factor and increased the light chain 3-II/I ratio.However,PX-478 and Mdivi-1 reversed these effects.Additionally,PX-478 and Mdivi-1 rescued the effects of HYWZF by decreasing oxidative stress and apoptosis and increasing mitophagy.HYWZF intervention improved the symptoms of diabetes,tissue damage,number of acellular capillaries,and oxidative stress in vivo.Furthermore,in vivo experiments confirmed the results of in vitro experiments.CONCLUSION HYWZF alleviated DR and associated damage by promoting mitophagy via the HIF-1α/BNIP3/NIX axis.展开更多
目的:探讨二氯化钴(cobaltous chloride,Co Cl2)诱导缺氧对肌细胞萎缩的调控机制。方法:本研究采用C2C12小鼠成肌细胞系作为细胞模型,分为正常组、Co Cl2组、正常+3-Methyladenine(3MA)组、Co Cl2+3MA组。正常组不作处理,Co Cl2组加入20...目的:探讨二氯化钴(cobaltous chloride,Co Cl2)诱导缺氧对肌细胞萎缩的调控机制。方法:本研究采用C2C12小鼠成肌细胞系作为细胞模型,分为正常组、Co Cl2组、正常+3-Methyladenine(3MA)组、Co Cl2+3MA组。正常组不作处理,Co Cl2组加入200μM Co Cl2诱导缺氧,正常+3MA组加入5 m M 3MA,Co Cl2+3MA组加入200μM Co Cl2及5 m M 3MA。使用吉姆萨染色观察肌管形态,多功能酶标仪检测活性氧(reactive oxygen species,ROS)表达,电镜观察自噬体形成情况,实时定量聚合酶链反应(quantita-tive real time polymerase chain reaction,QRT-PCR)和免疫印迹技术(western blotting,WB)检测缺氧诱导因子-1α(hypoxia-inducible factor-1α,HIF-1α)、Bcl2/腺病毒E1B 19k Da相关蛋白3(Bcl2/adenovirusE1B 19k Da interacting protein 3,BNIP3)、微管相关蛋白1轻链-3(microtubule associated protein1 lightchain 3,LC3)m RNA及蛋白表达;另外,通过抑制自噬观察肌肉萎缩盒F基因(muscle atrophy F-box,MAFbx)蛋白表达的变化。结果:正常组可见长条状肌管形成,Co Cl2处理后肌管萎缩、断裂。Co Cl2组ROS含量较正常组升高(t=-4.965,P=0.008),电镜可观察到Co Cl2诱导自噬体形成,同时HIF-1α、BNIP3、LC3表达增加(P<0.05)。Co Cl2与3MA共处理可减少MAFbx蛋白的表达(F=18.246,P=0.001)。结论:Co Cl2诱导C2C12骨骼肌细胞萎缩,可能与HIF-1α/BNIP3信号通路促进自噬发生有关,抑制缺氧诱导的自噬可部分减少肌萎缩。展开更多
目的 探究胰腺癌Bcl-2/腺病毒E1B-19k相关蛋白3(Bcl-2/E1B-19k-interacting protein 3,BNIP3)的蛋白表达水平及其与术后吉西他滨化疗敏感性的关系。方法 回顾性分析2016年7月至2021年8月内蒙古医科大学附属医院58例胰腺癌根治术,并在术...目的 探究胰腺癌Bcl-2/腺病毒E1B-19k相关蛋白3(Bcl-2/E1B-19k-interacting protein 3,BNIP3)的蛋白表达水平及其与术后吉西他滨化疗敏感性的关系。方法 回顾性分析2016年7月至2021年8月内蒙古医科大学附属医院58例胰腺癌根治术,并在术后规律应用吉西他滨辅助化疗或以吉西他滨为主的联合化疗患者的临床资料,选取胰腺癌术后组织切片,对其进行BNIP3免疫组织化学检测,通过实验染色强度及范围进行评分,并分析其与胰腺癌患者预后的关系。结果 与BNIP3低表达组比较,BNIP3高表达组的总生存时间(OS)[13(95%CI 11.378~14.622)个月 vs 18(95%CI 16.592~19.408)个月]和无病生存时间(DFS)[7(95%CI 4.082~9.938)个月 vs 14(95%CI 12.936~15.064)个月]均降低(P<0.05)。BNIP3的蛋白表达水平及淋巴转移情况是OS的独立预后因素,而BNIP3的蛋白表达水平及分化程度是DFS的独立预后因素。结论 胰腺癌组织中BNIP3的蛋白表达水平可能与应用吉西他滨化疗敏感性相关,BNIP3低表达患者行术后吉西他滨化疗的OS与DFS较高表达者更长。展开更多
Osteosarcoma(OS)therapy faces many challenges,especially the poor survival rate once metastasis occurs.Therefore,it is crucial to explore new OS treatment strategies that can efficiently inhibit OS metastasis.Bioactiv...Osteosarcoma(OS)therapy faces many challenges,especially the poor survival rate once metastasis occurs.Therefore,it is crucial to explore new OS treatment strategies that can efficiently inhibit OS metastasis.Bioactive nanoparticles such as zinc oxide nanoparticles(ZnO NPs)can efficiently inhibit OS growth,however,the effect and mechanisms of them on tumor metastasis are still not clear.In this study,we firstly prepared well-dispersed ZnO NPs and proved that ZnO NPs can inhibit OS metastasis-related malignant behaviors including migration,invasion,and epithelial-mesenchymal transition(EMT).RNA-Seqs found that differentially expressed genes(DEGs)in ZnO NP-treated OS cells were enriched in wingless/integrated(Wnt)and hypoxia-inducible factor-1(HIF-1)signaling pathway.We further proved that Zn^(2+)released from ZnO NPs induced downregulation ofβ-catenin expression via HIF-1α/BNIP3/LC3B-mediated mitophagy pathway.ZnO NPs combined with ICG-001,aβ-catenin inhibitor,showed a synergistic inhibitory effect on OS lung metastasis and a longer survival time.In addition,tissue microarray(TMA)of OS patients also detected much higherβ-catenin expression which indicated the role ofβ-catenin in OS development.In summary,our current study not only proved that ZnO NPs can inhibit OS metastasis by degradingβ-catenin in HIF-1α/BNIP3/LC3B-mediated mitophagy pathway,but also provided a far-reaching potential of ZnO NPs in clinical OS treatment with metastasis.展开更多
基金Supported by the National Key Research and Development Project of China,No.2019YFC1711605National Natural Science Foundation of China,No.81904257Medical Innovation Research Project of Science and Technology Commission of Shanghai Municipality,No.21Y11923100.
文摘BACKGROUND Diabetic retinopathy(DR)is the primary cause of visual problems in patients with diabetes.The Heyingwuzi formulation(HYWZF)is effective against DR.AIM To determine the HYWZF prevention mechanisms,especially those underlying mitophagy.METHODS Human retinal capillary endothelial cells(HRCECs)were treated with high glucose(hg),HYWZF serum,PX-478,or Mdivi-1 in vitro.Then,cell counting kit-8,transwell,and tube formation assays were used to evaluate HRCEC proliferation,invasion,and tube formation,respectively.Transmission electron microscopy was used to assess mitochondrial morphology,and Western blotting was used to determine the protein levels.Flow cytometry was used to assess cell apoptosis,reactive oxygen species(ROS)production,and mitochondrial membrane potential.Moreover,C57BL/6 mice were established in vivo using streptozotocin and treated with HYWZF for four weeks.Blood glucose levels and body weight were monitored continuously.Changes in retinal characteristics were evaluated using hematoxylin and eosin,tar violet,and periodic acid-Schiff staining.Protein levels in retinal tissues were determined via Western blotting,immunohistochemistry,and immunostaining.RESULTS HYWZF inhibited excessive ROS production,apoptosis,tube formation,and invasion in hg-induced HRCECs via mitochondrial autophagy in vitro.It increased the mRNA expression levels of BCL2-interacting protein 3(BNIP3),FUN14 domain-containing 1,BNIP3-like(BNIP3L,also known as NIX),PARKIN,PTEN-induced kinase 1,and hypoxia-inducible factor(HIF)-1α.Moreover,it downregulated the protein levels of vascular endothelial cell growth factor and increased the light chain 3-II/I ratio.However,PX-478 and Mdivi-1 reversed these effects.Additionally,PX-478 and Mdivi-1 rescued the effects of HYWZF by decreasing oxidative stress and apoptosis and increasing mitophagy.HYWZF intervention improved the symptoms of diabetes,tissue damage,number of acellular capillaries,and oxidative stress in vivo.Furthermore,in vivo experiments confirmed the results of in vitro experiments.CONCLUSION HYWZF alleviated DR and associated damage by promoting mitophagy via the HIF-1α/BNIP3/NIX axis.
文摘目的:探讨二氯化钴(cobaltous chloride,Co Cl2)诱导缺氧对肌细胞萎缩的调控机制。方法:本研究采用C2C12小鼠成肌细胞系作为细胞模型,分为正常组、Co Cl2组、正常+3-Methyladenine(3MA)组、Co Cl2+3MA组。正常组不作处理,Co Cl2组加入200μM Co Cl2诱导缺氧,正常+3MA组加入5 m M 3MA,Co Cl2+3MA组加入200μM Co Cl2及5 m M 3MA。使用吉姆萨染色观察肌管形态,多功能酶标仪检测活性氧(reactive oxygen species,ROS)表达,电镜观察自噬体形成情况,实时定量聚合酶链反应(quantita-tive real time polymerase chain reaction,QRT-PCR)和免疫印迹技术(western blotting,WB)检测缺氧诱导因子-1α(hypoxia-inducible factor-1α,HIF-1α)、Bcl2/腺病毒E1B 19k Da相关蛋白3(Bcl2/adenovirusE1B 19k Da interacting protein 3,BNIP3)、微管相关蛋白1轻链-3(microtubule associated protein1 lightchain 3,LC3)m RNA及蛋白表达;另外,通过抑制自噬观察肌肉萎缩盒F基因(muscle atrophy F-box,MAFbx)蛋白表达的变化。结果:正常组可见长条状肌管形成,Co Cl2处理后肌管萎缩、断裂。Co Cl2组ROS含量较正常组升高(t=-4.965,P=0.008),电镜可观察到Co Cl2诱导自噬体形成,同时HIF-1α、BNIP3、LC3表达增加(P<0.05)。Co Cl2与3MA共处理可减少MAFbx蛋白的表达(F=18.246,P=0.001)。结论:Co Cl2诱导C2C12骨骼肌细胞萎缩,可能与HIF-1α/BNIP3信号通路促进自噬发生有关,抑制缺氧诱导的自噬可部分减少肌萎缩。
文摘目的 探究胰腺癌Bcl-2/腺病毒E1B-19k相关蛋白3(Bcl-2/E1B-19k-interacting protein 3,BNIP3)的蛋白表达水平及其与术后吉西他滨化疗敏感性的关系。方法 回顾性分析2016年7月至2021年8月内蒙古医科大学附属医院58例胰腺癌根治术,并在术后规律应用吉西他滨辅助化疗或以吉西他滨为主的联合化疗患者的临床资料,选取胰腺癌术后组织切片,对其进行BNIP3免疫组织化学检测,通过实验染色强度及范围进行评分,并分析其与胰腺癌患者预后的关系。结果 与BNIP3低表达组比较,BNIP3高表达组的总生存时间(OS)[13(95%CI 11.378~14.622)个月 vs 18(95%CI 16.592~19.408)个月]和无病生存时间(DFS)[7(95%CI 4.082~9.938)个月 vs 14(95%CI 12.936~15.064)个月]均降低(P<0.05)。BNIP3的蛋白表达水平及淋巴转移情况是OS的独立预后因素,而BNIP3的蛋白表达水平及分化程度是DFS的独立预后因素。结论 胰腺癌组织中BNIP3的蛋白表达水平可能与应用吉西他滨化疗敏感性相关,BNIP3低表达患者行术后吉西他滨化疗的OS与DFS较高表达者更长。
基金supported in part by Beijing Natural Science Foundation(7192226,7222011)Beijing Chao-Yang Hospital Golden Seeds Foundation(CYJZ202148)+3 种基金National Key Research and Development Program(2021YFC2400500)National Natural Science Foundation of China(51903013,51973021,51932002,52173275)Beijing Hospitals Authority Youth Programme(QML20210402)the Beijing Municipal Health Commission(PXM 2020_026275_000002,BMHC-2021-6,BMHC-2019-9).
文摘Osteosarcoma(OS)therapy faces many challenges,especially the poor survival rate once metastasis occurs.Therefore,it is crucial to explore new OS treatment strategies that can efficiently inhibit OS metastasis.Bioactive nanoparticles such as zinc oxide nanoparticles(ZnO NPs)can efficiently inhibit OS growth,however,the effect and mechanisms of them on tumor metastasis are still not clear.In this study,we firstly prepared well-dispersed ZnO NPs and proved that ZnO NPs can inhibit OS metastasis-related malignant behaviors including migration,invasion,and epithelial-mesenchymal transition(EMT).RNA-Seqs found that differentially expressed genes(DEGs)in ZnO NP-treated OS cells were enriched in wingless/integrated(Wnt)and hypoxia-inducible factor-1(HIF-1)signaling pathway.We further proved that Zn^(2+)released from ZnO NPs induced downregulation ofβ-catenin expression via HIF-1α/BNIP3/LC3B-mediated mitophagy pathway.ZnO NPs combined with ICG-001,aβ-catenin inhibitor,showed a synergistic inhibitory effect on OS lung metastasis and a longer survival time.In addition,tissue microarray(TMA)of OS patients also detected much higherβ-catenin expression which indicated the role ofβ-catenin in OS development.In summary,our current study not only proved that ZnO NPs can inhibit OS metastasis by degradingβ-catenin in HIF-1α/BNIP3/LC3B-mediated mitophagy pathway,but also provided a far-reaching potential of ZnO NPs in clinical OS treatment with metastasis.
