Purpose: To investigate the effects of exogenous basic fibroblast growth factor-2(FGF-2) on the biological activity of endothelial progenitor cells(EPCs) exposed to high glucose conditions. Materials and Methods: 1) B...Purpose: To investigate the effects of exogenous basic fibroblast growth factor-2(FGF-2) on the biological activity of endothelial progenitor cells(EPCs) exposed to high glucose conditions. Materials and Methods: 1) Bone marrow EPCs from C57BL/6 mice were isolated and cultured in vitro. EPC purity was identified by flow cytometry and immunofluorescence staining. 2) Apoptosis was detected by TUNEL assay. Migration and tube formation ability was detected by Transwell chamber and Matrigel assays, respectively. The expression and activation of β-catenin was detected by Western blot. 3) Doppler flowmetry was used to detect the effect of FGF2 on blood flow recovery in ischemic hind limbs of mice. Results: 1) FGF-2 treatment reversed high glucose induced growth inhibition of EPCs. FGF-2 treatment also increased migration and tube formation ability of EPCs even in high glucose conditions. 2) Western blot analysis demonstrated that the percentage of activated β-catenin/total β-catenin in the high glucose group were significantly lower than that in the control group, while FGF-2 treatment reversed high glucose induced β-catenin inhibition. 3) In vivo experiments demonstrated that the blood flow recovery in ischemic hind limbs of mice was significantly improved after FGF-2 treatment. Conclusion: Exogenous FGF-2 could play a role in the functional repair of damaged EPC exposed to high glucose conditions, via the activation of the Wnt/β-catenin signaling pathway.展开更多
Prior to the generation of hematopoietic stem cells(HSCs)from the hemogenic endothelial cells(HECs)mainly in the dorsal aorta in midgestational mouse embryos,multiple hematopoietic progenitors including erythro-myeloi...Prior to the generation of hematopoietic stem cells(HSCs)from the hemogenic endothelial cells(HECs)mainly in the dorsal aorta in midgestational mouse embryos,multiple hematopoietic progenitors including erythro-myeloid progenitors and lymphoid progenitors are generated from yolk sac HECs.These HSCindependent hematopoietic progenitors have recently been identified as major contributors to functional blood cell production until birth.However,little is known about yolk sac HECs.Here,combining integrative analyses of multiple single-cell RNA-sequencing datasets and functional assays,we reveal that Neurl3-EGFP,in addition to marking the continuum throughout the ontogeny of HSCs from HECs,can also serve as a single enrichment marker for yolk sac HECs.Moreover,while yolk sac HECs have much weaker arterial characteristics than either arterial endothelial cells in the yolk sac or HECs within the embryo proper,the lymphoid potential of yolk sac HECs is largely confined to the arterial-biased subpopulation featured by the Unc5b expression.Interestingly,the B lymphoid potential of hematopoietic progenitors,but not for myeloid potentials,is exclusively detected in Neurl3-negative subpopulations in midgestational embryos.Taken together,these findings enhance our understanding of blood birth from yolk sac HECs and provide theoretical basis and candidate reporters for monitoring step-wise hematopoietic differentiation.展开更多
During embryogenesis,hematopoietic stem progenitor cells(HSPCs)are believed to be derived from hemogenic endothelial cells(HECs).Moreover,arterial feature is proposed to be a prerequisite for HECs to generate HSPCs wi...During embryogenesis,hematopoietic stem progenitor cells(HSPCs)are believed to be derived from hemogenic endothelial cells(HECs).Moreover,arterial feature is proposed to be a prerequisite for HECs to generate HSPCs with lymphoid potential.Although the molecular basis of hematopoietic stem cell-competent HECs has been delicately elucidated within the embryo proper,the functional and molecular characteristics of HECs in the extraembryonic yolk sac(YS)remain largely unresolved.In this study,we initially identified six molecularly different endothelial populations in the midgestational YS through integrated analysis of several single-cell RNA sequencing(scRNA-seq)datasets and validated the arterial vasculature distribution of Gja5+ECs using a Gja5-EGFP reporter mouse model.Further,we explored the hemogenic potential of different EC populations based on their Gja5-EGFP and CD44 expression levels.The hemogenic potential was ubiquitously detected in spatiotemporally different vascular beds on embryonic days(E)8.5–E9.5 and gradually concentrated in CD44-positive ECs from E10.0.Unexpectedly,B-lymphoid potential was detected in the YS ECs as early as E8.5 regardless of their arterial features.Furthermore,the capacity for generating hematopoietic progenitors with in vivo lymphoid potential was found in nonarterial as well as arterial YS ECs on E10.0–E10.5.Importantly,the distinct identities of E10.0–E10.5 HECs between YS and intraembryonic caudal region were revealed by further scRNA-seq analysis.Cumulatively,these findings extend our knowledge regarding the hemogenic potential of ECs from anatomically and molecularly different vascular beds,providing a theoretical basis for better understanding the sources of HSPCs during mammalian development.展开更多
基金This study was supported by grants from the General Programs of the National Natural Science Foundation of China(No.8157070468)
文摘Purpose: To investigate the effects of exogenous basic fibroblast growth factor-2(FGF-2) on the biological activity of endothelial progenitor cells(EPCs) exposed to high glucose conditions. Materials and Methods: 1) Bone marrow EPCs from C57BL/6 mice were isolated and cultured in vitro. EPC purity was identified by flow cytometry and immunofluorescence staining. 2) Apoptosis was detected by TUNEL assay. Migration and tube formation ability was detected by Transwell chamber and Matrigel assays, respectively. The expression and activation of β-catenin was detected by Western blot. 3) Doppler flowmetry was used to detect the effect of FGF2 on blood flow recovery in ischemic hind limbs of mice. Results: 1) FGF-2 treatment reversed high glucose induced growth inhibition of EPCs. FGF-2 treatment also increased migration and tube formation ability of EPCs even in high glucose conditions. 2) Western blot analysis demonstrated that the percentage of activated β-catenin/total β-catenin in the high glucose group were significantly lower than that in the control group, while FGF-2 treatment reversed high glucose induced β-catenin inhibition. 3) In vivo experiments demonstrated that the blood flow recovery in ischemic hind limbs of mice was significantly improved after FGF-2 treatment. Conclusion: Exogenous FGF-2 could play a role in the functional repair of damaged EPC exposed to high glucose conditions, via the activation of the Wnt/β-catenin signaling pathway.
基金supported by the National Key R&D Program of China(2022YFA1103501,2020YFA0112400,2021YFA1100102)the National Natural Science Foundation of China(82000111,81890991,31930054,82200121,82122004,82270118)the Program for Guangdong Introducing Innovative and Entrepreneurial Teams(2017ZT07S347).
文摘Prior to the generation of hematopoietic stem cells(HSCs)from the hemogenic endothelial cells(HECs)mainly in the dorsal aorta in midgestational mouse embryos,multiple hematopoietic progenitors including erythro-myeloid progenitors and lymphoid progenitors are generated from yolk sac HECs.These HSCindependent hematopoietic progenitors have recently been identified as major contributors to functional blood cell production until birth.However,little is known about yolk sac HECs.Here,combining integrative analyses of multiple single-cell RNA-sequencing datasets and functional assays,we reveal that Neurl3-EGFP,in addition to marking the continuum throughout the ontogeny of HSCs from HECs,can also serve as a single enrichment marker for yolk sac HECs.Moreover,while yolk sac HECs have much weaker arterial characteristics than either arterial endothelial cells in the yolk sac or HECs within the embryo proper,the lymphoid potential of yolk sac HECs is largely confined to the arterial-biased subpopulation featured by the Unc5b expression.Interestingly,the B lymphoid potential of hematopoietic progenitors,but not for myeloid potentials,is exclusively detected in Neurl3-negative subpopulations in midgestational embryos.Taken together,these findings enhance our understanding of blood birth from yolk sac HECs and provide theoretical basis and candidate reporters for monitoring step-wise hematopoietic differentiation.
基金supported by the National Key Research and Development Program of China(2020YFA0112402,2017YFA0103401,and 2016YFA0100601)the National Natural Science Foundation of China(81890991,31930054,31871173,82000111,and 81900115)+1 种基金the Program for Guangdong Introducing Innovative and Entrepreneurial Teams(2017ZT07S347)the Key Research and Development Program of Guangdong Province(2019B020234002)。
文摘During embryogenesis,hematopoietic stem progenitor cells(HSPCs)are believed to be derived from hemogenic endothelial cells(HECs).Moreover,arterial feature is proposed to be a prerequisite for HECs to generate HSPCs with lymphoid potential.Although the molecular basis of hematopoietic stem cell-competent HECs has been delicately elucidated within the embryo proper,the functional and molecular characteristics of HECs in the extraembryonic yolk sac(YS)remain largely unresolved.In this study,we initially identified six molecularly different endothelial populations in the midgestational YS through integrated analysis of several single-cell RNA sequencing(scRNA-seq)datasets and validated the arterial vasculature distribution of Gja5+ECs using a Gja5-EGFP reporter mouse model.Further,we explored the hemogenic potential of different EC populations based on their Gja5-EGFP and CD44 expression levels.The hemogenic potential was ubiquitously detected in spatiotemporally different vascular beds on embryonic days(E)8.5–E9.5 and gradually concentrated in CD44-positive ECs from E10.0.Unexpectedly,B-lymphoid potential was detected in the YS ECs as early as E8.5 regardless of their arterial features.Furthermore,the capacity for generating hematopoietic progenitors with in vivo lymphoid potential was found in nonarterial as well as arterial YS ECs on E10.0–E10.5.Importantly,the distinct identities of E10.0–E10.5 HECs between YS and intraembryonic caudal region were revealed by further scRNA-seq analysis.Cumulatively,these findings extend our knowledge regarding the hemogenic potential of ECs from anatomically and molecularly different vascular beds,providing a theoretical basis for better understanding the sources of HSPCs during mammalian development.
