The key regulators and regeneration-associated genes involved in axonal regeneration of neurons after injury have not been clarified.In high-throughput sequencing,various factors influence the final sequencing results...The key regulators and regeneration-associated genes involved in axonal regeneration of neurons after injury have not been clarified.In high-throughput sequencing,various factors influence the final sequencing results,including the number and size of cells,the depth of sequencing,and the method of cell separation.There is still a lack of research on the detailed molecular expression profile during the regeneration of dorsal root ganglion neuron axon.In this study,we performed lase r-capture microdissection coupled with RNA sequencing on dorsal root ganglion neurons at 0,3,6,and 12 hours and 1,3,and 7 days after sciatic nerve crush in rats.We identified three stages after dorsal root ganglion injury:early(3-12 hours),pre-regeneration(1 day),and regeneration(3-7 days).Gene expression patterns and related function enrichment res ults showed that one module of genes was highly related to axonal regeneration.We verified the up-regulation of activating transcription factor 3(Atf3),Kruppel like factor 6(Klf6),AT-rich inte raction domain 5A(Arid5α),CAMP responsive element modulator(Crem),and FOS like 1,AP-1 transcription factor Subunit(Fosl1) in dorsal root ganglion neurons after injury.Suppressing these transcription factors(Crem,Arid5o,Fosl1 and Klf6) reduced axonal regrowth in vitro.As the hub transcription factor,Atf3 showed higher expression and activity at the preregeneration and regeneration stages.G protein-coupled estrogen receptor 1(Gper1),inte rleukin 12a(Il12α),estrogen receptor 1(ESR1),and interleukin 6(IL6) may be upstream factors that trigger the activation of Atf3 during the repair of axon injury in the early stage.Our study presents the detailed molecular expression profile during axonal regeneration of dorsal root ganglion neurons after peripheral nerve injury.These findings may provide reference for the clinical screening of molecular targets for the treatment of peripheral nerve injury.展开更多
Background:The current understanding of diabetic kidney disease(DKD)has significant gaps regarding the underlying pathogenesis.In this study,we aimed to characterize the temporal progression of DKD using a state-of-th...Background:The current understanding of diabetic kidney disease(DKD)has significant gaps regarding the underlying pathogenesis.In this study,we aimed to characterize the temporal progression of DKD using a state-of-the-art mouse model of hypertension-accelerated disease,integrating kidney biomarker analysis,histopathology,and glomerular transcriptomic profiling.Methods:Female diabetic db/db mice received a single intravenous dose of adenoassociated virus-mediated renin overexpression(ReninAAV,week 5)and underwent uninephrectomy(UNx,week 4).db/db UNx-ReninAAV mice were terminated at weeks 1,4,8,and 12(n=7–8 per group).Female db/m mice were used as healthy controls.Study endpoints included plasma and urine biochemistry,glomerulosclerosis scoring,quantitative kidney histology,and RNA sequencing of glomeruli isolated using lasercapture microdissection.Results:db/db UNx-ReninAAV mice developed progressive albuminuria(from week 4)and glomerulosclerosis(from week 8).A pathway analysis of clustered gene regulations revealed broad glomerular transcriptome perturbations with signatures of increased extracellular matrix(ECM)turnover from week 8 and early onset of metabolic dysfunction.Markers of glomerular cell types and injury exhibited temporal regulation over the course of DKD,with early and sustained downregulation of endothelial markers,heterogeneous regulation of podocyte markers,and significant mesangial and parietal epithelial aberrations.Furthermore,the upregulation of cell injury markers confirmed progressive glomerular injury in the model.Conclusion:The db/db UNx-ReninAAV mouse model exhibits distinct temporal dynamics in glomerular cell markers,metabolic dysregulation,ECM remodeling,and injury.Together,these results highlight the utility of the db/db UNx-ReninAAV model as a relevant preclinical platform for studying progressive DKD.展开更多
基金supported by the National Natural Science Foundation of China,Nos. 31730031 and 32130060the National Major Project of Research and Development,No. 2017YFA0104700the Natural Science Foundation of Jiangsu Province,No. BK20202013 (all to XSG)。
文摘The key regulators and regeneration-associated genes involved in axonal regeneration of neurons after injury have not been clarified.In high-throughput sequencing,various factors influence the final sequencing results,including the number and size of cells,the depth of sequencing,and the method of cell separation.There is still a lack of research on the detailed molecular expression profile during the regeneration of dorsal root ganglion neuron axon.In this study,we performed lase r-capture microdissection coupled with RNA sequencing on dorsal root ganglion neurons at 0,3,6,and 12 hours and 1,3,and 7 days after sciatic nerve crush in rats.We identified three stages after dorsal root ganglion injury:early(3-12 hours),pre-regeneration(1 day),and regeneration(3-7 days).Gene expression patterns and related function enrichment res ults showed that one module of genes was highly related to axonal regeneration.We verified the up-regulation of activating transcription factor 3(Atf3),Kruppel like factor 6(Klf6),AT-rich inte raction domain 5A(Arid5α),CAMP responsive element modulator(Crem),and FOS like 1,AP-1 transcription factor Subunit(Fosl1) in dorsal root ganglion neurons after injury.Suppressing these transcription factors(Crem,Arid5o,Fosl1 and Klf6) reduced axonal regrowth in vitro.As the hub transcription factor,Atf3 showed higher expression and activity at the preregeneration and regeneration stages.G protein-coupled estrogen receptor 1(Gper1),inte rleukin 12a(Il12α),estrogen receptor 1(ESR1),and interleukin 6(IL6) may be upstream factors that trigger the activation of Atf3 during the repair of axon injury in the early stage.Our study presents the detailed molecular expression profile during axonal regeneration of dorsal root ganglion neurons after peripheral nerve injury.These findings may provide reference for the clinical screening of molecular targets for the treatment of peripheral nerve injury.
基金funding from the Innovation Fund Denmark(grant number 2040-00034B).
文摘Background:The current understanding of diabetic kidney disease(DKD)has significant gaps regarding the underlying pathogenesis.In this study,we aimed to characterize the temporal progression of DKD using a state-of-the-art mouse model of hypertension-accelerated disease,integrating kidney biomarker analysis,histopathology,and glomerular transcriptomic profiling.Methods:Female diabetic db/db mice received a single intravenous dose of adenoassociated virus-mediated renin overexpression(ReninAAV,week 5)and underwent uninephrectomy(UNx,week 4).db/db UNx-ReninAAV mice were terminated at weeks 1,4,8,and 12(n=7–8 per group).Female db/m mice were used as healthy controls.Study endpoints included plasma and urine biochemistry,glomerulosclerosis scoring,quantitative kidney histology,and RNA sequencing of glomeruli isolated using lasercapture microdissection.Results:db/db UNx-ReninAAV mice developed progressive albuminuria(from week 4)and glomerulosclerosis(from week 8).A pathway analysis of clustered gene regulations revealed broad glomerular transcriptome perturbations with signatures of increased extracellular matrix(ECM)turnover from week 8 and early onset of metabolic dysfunction.Markers of glomerular cell types and injury exhibited temporal regulation over the course of DKD,with early and sustained downregulation of endothelial markers,heterogeneous regulation of podocyte markers,and significant mesangial and parietal epithelial aberrations.Furthermore,the upregulation of cell injury markers confirmed progressive glomerular injury in the model.Conclusion:The db/db UNx-ReninAAV mouse model exhibits distinct temporal dynamics in glomerular cell markers,metabolic dysregulation,ECM remodeling,and injury.Together,these results highlight the utility of the db/db UNx-ReninAAV model as a relevant preclinical platform for studying progressive DKD.
