Background:Neutrophils are traditionally viewed as first responders but have a short onset of action in response to traumatic brain injury(TBI).However,the heterogeneity,multifunctionality,and time-dependent modulatio...Background:Neutrophils are traditionally viewed as first responders but have a short onset of action in response to traumatic brain injury(TBI).However,the heterogeneity,multifunctionality,and time-dependent modulation of brain damage and outcome mediated by neutrophils after TBI remain poorly understood.Methods:Using the combined single-cell transcriptomics,metabolomics,and proteomics analysis from TBI patients and the TBI mouse model,we investigate a novel neutrophil phenotype and its associated effects on TBI outcome by neurological deficit scoring and behavioral tests.We also characterized the underlying mechanisms both invitro and invivo through molecular simulations,signaling detections,gene expression regulation assessments[including dual-luciferase reporter and chromatin immunoprecipitation(ChIP)assays],primary cultures or co-cultures of neutrophils and oligodendrocytes,intracellular iron,and lipid hydroperoxide concentration measurements,as well as forkhead box protein O1(FOXO1)conditional knockout mice.Results:We identified that high expression of the FOXO1 protein was induced in neutrophils after TBI both in TBI patients and the TBI mouse model.Infiltration of these FOXO1high neutrophils in the brain was detected not only in the acute phase but also in the chronic phase post-TBI,aggravating acute brain inflammatory damage and promoting late TBI-induced depression.In the acute stage,FOXO1 upregulated cytoplasmic Versican(VCAN)to interact with the apoptosis regulator B-cell lymphoma-2(BCL-2)-associated X protein(BAX),suppressing the mitochondrial translocation of BAX,which mediated the antiapoptotic effect companied with enhancing interleukin-6(IL-6)production of FOXO1high neutrophils.In the chronic stage,the“FOXO1-transferrin receptor(TFRC)”mechanism contributes to FOXO1high neutrophil ferroptosis,disturbing the iron homeostasis of oligodendrocytes and inducing a reduction in myelin basic protein,which contributes to the progression of late depression after TBI.Conclusions:FOXO1high neutrophils represent a novel neutrophil phenotype that emerges in response to acute and chronic TBI,which provides insight into the heterogeneity,reprogramming activity,and versatility of neutrophils in TBI.展开更多
高原低氧环境会引起肌力下降和运动能力退化,而抗阻训练是刺激骨骼肌生长的重要手段,叉头转录因子1(fork head box protein O 1,FoxO1)在调控骨骼肌蛋白质分解通路中承担重要角色。为探究Akt-FoxO1通路是否参与抗阻训练抑制低氧诱导的...高原低氧环境会引起肌力下降和运动能力退化,而抗阻训练是刺激骨骼肌生长的重要手段,叉头转录因子1(fork head box protein O 1,FoxO1)在调控骨骼肌蛋白质分解通路中承担重要角色。为探究Akt-FoxO1通路是否参与抗阻训练抑制低氧诱导的骨骼肌萎缩,本研究构建低氧诱导骨骼肌萎缩的大鼠模型,并模拟海拔4 000 m低氧环境下(12.4%O_2)进行抗阻训练,对比观察大鼠比目鱼肌和趾长伸肌湿重和横截面积,以及蛋白激酶B(protein kinase B,Akt)、叉头转录因子1、泛素蛋白连接酶1(muscle ring finger 1,MuRF1)的表达差异等。结果表明,低氧暴露导致大鼠趾长伸肌湿重显著下降,苏木精-伊红染色组织切片分析肌纤维横截面积、低氧环境下比目鱼肌横截面积明显下降,而低氧抗阻训练后趾长伸肌横截面积明显高于安静组。实时荧光定量PCR和蛋白质免疫印迹结果显示,低氧暴露后FoxO1和MuRF1基因表达明显上调,低氧下抗阻训练后发现,Akt基因表达明显上调而FoxO1、MuRF则明显下调。免疫荧光观察磷酸化FoxO1在细胞核内外表达情况,发现抗阻训练后FoxO1(S256)于细胞核外表达增强。上述结果表明,抗阻训练可以达到抑制低氧诱导骨骼肌萎缩的效果,Akt促进FoxO1磷酸化从而减缓骨骼肌蛋白质分解过程是抗阻训练能够抑制骨骼肌萎缩的分子机制之一。展开更多
To explore the role of forkhead box protein O1(FOXO1)in the progression of glioblastoma multiforme(GBM)and related drug resistance,we deciphered the roles of FOXO1 and miR-506 in proliferation,apoptosis,migration,inva...To explore the role of forkhead box protein O1(FOXO1)in the progression of glioblastoma multiforme(GBM)and related drug resistance,we deciphered the roles of FOXO1 and miR-506 in proliferation,apoptosis,migration,invasion,autophagy,and temozolomide(TMZ)sensitivity in the U251 cell line using in vitro and in vivo experiments.Cell viability was tested by a cell counting kit-8(CCK8)kit;migration and invasion were checked by the scratching assay;apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick-end labeling(TUNEL)staining and flow cytometry.The construction of plasmids and dual-luciferase reporter experiment were carried out to find the interaction site between FOXO1 and miR-506.Immunohistochemistry was done to check the protein level in tumors after the in vivo experiment.We found that the FOXO1-miR-506 axis suppresses GBM cell invasion and migration and promotes GBM chemosensitivity to TMZ,which was mediated by autophagy.FOXO1 upregulates miR-506 by binding to its promoter to enhance transcriptional activation.MiR-506 could downregulate E26 transformation-specific 1(ETS1)expression by targeting its 3'-untranslated region(UTR).Interestingly,ETS1 promoted FOXO1 translocation from the nucleus to the cytosol and further suppressed the FOXO1-miR-506 axis in GBM cells.Consistently,both miR-506 inhibition and ETS1 overexpression could rescue FOXO1 overactivation-mediated TMZ chemosensitivity in mouse models.Our study demonstrated a negative feedback loop of FOXO1/miR-506/ETS1/FOXO1 in GBM in regulating invasiveness and chemosensitivity.Thus,the above axis might be a promising therapeutic target for GBM.展开更多
基金This work was supported by the National Natural Science Foundation of China(82071779 and 81901626)the Science Fund for Creative Research Groups of Chongqing Municipal Education Commission of China,the grants from the Talent Foundation of Army Medical University(to Shuang-Shuang Dai)+1 种基金the Scientific Research Grant(ALJ22J003)the Chongqing Natural Science Foundation of China(CSTB2022NSCQ-MSX0177).
文摘Background:Neutrophils are traditionally viewed as first responders but have a short onset of action in response to traumatic brain injury(TBI).However,the heterogeneity,multifunctionality,and time-dependent modulation of brain damage and outcome mediated by neutrophils after TBI remain poorly understood.Methods:Using the combined single-cell transcriptomics,metabolomics,and proteomics analysis from TBI patients and the TBI mouse model,we investigate a novel neutrophil phenotype and its associated effects on TBI outcome by neurological deficit scoring and behavioral tests.We also characterized the underlying mechanisms both invitro and invivo through molecular simulations,signaling detections,gene expression regulation assessments[including dual-luciferase reporter and chromatin immunoprecipitation(ChIP)assays],primary cultures or co-cultures of neutrophils and oligodendrocytes,intracellular iron,and lipid hydroperoxide concentration measurements,as well as forkhead box protein O1(FOXO1)conditional knockout mice.Results:We identified that high expression of the FOXO1 protein was induced in neutrophils after TBI both in TBI patients and the TBI mouse model.Infiltration of these FOXO1high neutrophils in the brain was detected not only in the acute phase but also in the chronic phase post-TBI,aggravating acute brain inflammatory damage and promoting late TBI-induced depression.In the acute stage,FOXO1 upregulated cytoplasmic Versican(VCAN)to interact with the apoptosis regulator B-cell lymphoma-2(BCL-2)-associated X protein(BAX),suppressing the mitochondrial translocation of BAX,which mediated the antiapoptotic effect companied with enhancing interleukin-6(IL-6)production of FOXO1high neutrophils.In the chronic stage,the“FOXO1-transferrin receptor(TFRC)”mechanism contributes to FOXO1high neutrophil ferroptosis,disturbing the iron homeostasis of oligodendrocytes and inducing a reduction in myelin basic protein,which contributes to the progression of late depression after TBI.Conclusions:FOXO1high neutrophils represent a novel neutrophil phenotype that emerges in response to acute and chronic TBI,which provides insight into the heterogeneity,reprogramming activity,and versatility of neutrophils in TBI.
文摘高原低氧环境会引起肌力下降和运动能力退化,而抗阻训练是刺激骨骼肌生长的重要手段,叉头转录因子1(fork head box protein O 1,FoxO1)在调控骨骼肌蛋白质分解通路中承担重要角色。为探究Akt-FoxO1通路是否参与抗阻训练抑制低氧诱导的骨骼肌萎缩,本研究构建低氧诱导骨骼肌萎缩的大鼠模型,并模拟海拔4 000 m低氧环境下(12.4%O_2)进行抗阻训练,对比观察大鼠比目鱼肌和趾长伸肌湿重和横截面积,以及蛋白激酶B(protein kinase B,Akt)、叉头转录因子1、泛素蛋白连接酶1(muscle ring finger 1,MuRF1)的表达差异等。结果表明,低氧暴露导致大鼠趾长伸肌湿重显著下降,苏木精-伊红染色组织切片分析肌纤维横截面积、低氧环境下比目鱼肌横截面积明显下降,而低氧抗阻训练后趾长伸肌横截面积明显高于安静组。实时荧光定量PCR和蛋白质免疫印迹结果显示,低氧暴露后FoxO1和MuRF1基因表达明显上调,低氧下抗阻训练后发现,Akt基因表达明显上调而FoxO1、MuRF则明显下调。免疫荧光观察磷酸化FoxO1在细胞核内外表达情况,发现抗阻训练后FoxO1(S256)于细胞核外表达增强。上述结果表明,抗阻训练可以达到抑制低氧诱导骨骼肌萎缩的效果,Akt促进FoxO1磷酸化从而减缓骨骼肌蛋白质分解过程是抗阻训练能够抑制骨骼肌萎缩的分子机制之一。
基金supported by the National Natural Science Foundation of China(Nos.81402076,81872072,and 82073274)the Science Technology Commission of Shanghai Municipality(No.20S11900700)。
文摘To explore the role of forkhead box protein O1(FOXO1)in the progression of glioblastoma multiforme(GBM)and related drug resistance,we deciphered the roles of FOXO1 and miR-506 in proliferation,apoptosis,migration,invasion,autophagy,and temozolomide(TMZ)sensitivity in the U251 cell line using in vitro and in vivo experiments.Cell viability was tested by a cell counting kit-8(CCK8)kit;migration and invasion were checked by the scratching assay;apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick-end labeling(TUNEL)staining and flow cytometry.The construction of plasmids and dual-luciferase reporter experiment were carried out to find the interaction site between FOXO1 and miR-506.Immunohistochemistry was done to check the protein level in tumors after the in vivo experiment.We found that the FOXO1-miR-506 axis suppresses GBM cell invasion and migration and promotes GBM chemosensitivity to TMZ,which was mediated by autophagy.FOXO1 upregulates miR-506 by binding to its promoter to enhance transcriptional activation.MiR-506 could downregulate E26 transformation-specific 1(ETS1)expression by targeting its 3'-untranslated region(UTR).Interestingly,ETS1 promoted FOXO1 translocation from the nucleus to the cytosol and further suppressed the FOXO1-miR-506 axis in GBM cells.Consistently,both miR-506 inhibition and ETS1 overexpression could rescue FOXO1 overactivation-mediated TMZ chemosensitivity in mouse models.Our study demonstrated a negative feedback loop of FOXO1/miR-506/ETS1/FOXO1 in GBM in regulating invasiveness and chemosensitivity.Thus,the above axis might be a promising therapeutic target for GBM.
