title: MicroRNA-126a Directs Lymphangiogenesis Through Interacting With Chemokine and Flt4 Signaling in Zebrafish.
abstract: OBJECTIVE
  MicroRNA-126 (miR-126) is an endothelium-enriched miRNA and functions in vascular integrity and angiogenesis. The application of miRNA as potential biomarker and therapy target has been widely investigated in various pathological processes. However, its role in lymphatic diseases had not been widely explored. We aimed to reveal the role of miR-126 in lymphangiogenesis and the regulatory signaling pathways for potential targets of therapy.
  
  
  RESULTS
  Loss-of-function studies using morpholino oligonucleotides and CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) system showed that silencing of miR-126a severely affected the formation of parachordal lymphangioblasts and thoracic duct in zebrafish embryos, although their development in miR-126b knockdown embryos was normal. Expression analyses by in situ hybridization and immunofluorescence indicated that miR-126a was expressed in lymphatic vessels, as well as in blood vessels. Time-lapse confocal imaging assay further revealed that knockdown of miR-126a blocked both lymphangiogenic sprouts budding from the posterior cardinal vein and lymphangioblasts extension along horizontal myoseptum. Bioinformatics analysis and in vivo report assay identified that miR-126a upregulated Cxcl12a by targeting its 5' untranslated region. Moreover, loss- and gain-of-function studies revealed that Cxcl12a signaling acted downstream of miR-126a during parachordal lymphangioblast extension, whereby Flt4 signaling acts as a cooperator of miR-126a, allowing it to modulate lymphangiogenic sprout formation.
  
  
  CONCLUSIONS
  These findings demonstrate that miR-126a directs lymphatic endothelial cell sprouting and extension by interacting with Cxcl12a-mediated chemokine signaling and Vegfc-Flt4 signal axis. Our results suggest that these key regulators of lymphangiogenesis may be involved in lymphatic pathogenesis of cardiovascular diseases.
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.).
authors: Jian
authors: J
authors: Chen
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.).
authors: Rong-Fang
authors: RF
authors: Zhu
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.).
authors: Fang-Fang
authors: FF
authors: Li
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.).
authors: Yu-Lai
authors: YL
authors: Liang
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.).
authors: Chen
authors: C
authors: Wang
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.).
authors: Yong-Wen
authors: YW
authors: Qin
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.).
authors: Shuang
authors: S
authors: Huang
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.).
authors: Xian-Xian
authors: XX
authors: Zhao
authors: From the Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences and Shanghai Jiao-Tong University School of Medicine, China (J.C., R.-F.Z., F.-F.L., Y.-L.L., C.W., Q.J.); and Department of Cardiology, Changhai Hospital, Shanghai, China (Y.-W.Q., S.H., X.-X.Z., Q.J.). qjing@sibs.ac.cn 13601713431@163.com.
authors: Qing
authors: Q
authors: Jing
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D064113:CRISPR-Cas Systems; D002465:Cell Movement; D049109:Cell Proliferation; D054377:Chemokine CXCL12 / Q000235:genetics / Q000378:metabolism*; D019295:Computational Biology; D018507:Gene Expression Regulation, Developmental; D020868:Gene Silencing; D005838:Genotype; D042583:Lymphangiogenesis; D008220:Lymphography; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D060172:Morpholinos / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D015398:Signal Transduction; D013897:Thoracic Duct / Q000196:embryology / Q000378:metabolism*; D013997:Time Factors; D059008:Time-Lapse Imaging; D040321:Vascular Endothelial Growth Factor Receptor-3 / Q000235:genetics / Q000378:metabolism*; D015027:Zebrafish / Q000196:embryology / Q000235:genetics / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D054377:Chemokine CXCL12; D035683:MicroRNAs; C549766:Mirn126 microRNA, zebrafish; D060172:Morpholinos; D029961:Zebrafish Proteins; C579453:cxcl12a protein, zebrafish; D040321:Vascular Endothelial Growth Factor Receptor-3
keywords: endothelial cells; lymphangiogenesis; microRNAs; thoracic duct; zebrafish