title: Direct conversion of adult skin fibroblasts to endothelial cells by defined factors.
abstract: BACKGROUND
  Cell-based therapies to augment endothelial cells (ECs) hold great therapeutic promise. Here, we report a novel approach to generate functional ECs directly from adult fibroblasts.
  
  
  RESULTS
  Eleven candidate genes that are key regulators of endothelial development were selected. Green fluorescent protein (GFP)-negative skin fibroblasts were prepared from Tie2-GFP mice and infected with lentiviruses allowing simultaneous overexpression of all 11 factors. Tie2-GFP(+) cells (0.9%), representing Tie2 gene activation, were detected by flow cytometry. Serial stepwise screening revealed 5 key factors (Foxo1, Er71, Klf2, Tal1, and Lmo2) that were required for efficient reprogramming of skin fibroblasts into Tie2-GFP(+) cells (4%). This reprogramming strategy did not involve pluripotency induction because neither Oct4 nor Nanog was expressed after 5 key factor transduction. Tie2-GFP(+) cells were isolated using fluorescence-activated cell sorting and designated as induced ECs (iECs). iECs exhibited endothelium-like cobblestone morphology and expressed EC molecular markers. iECs possessed endothelial functions such as Bandeiraea simplicifolia-1 lectin binding, acetylated low-density lipoprotein uptake, capillary formation on Matrigel, and nitric oxide production. The epigenetic profile of iECs was similar to that of authentic ECs because the promoters of VE-cadherin and Tie2 genes were demethylated. mRNA profiling showed clustering of iECs with authentic ECs and highly enriched endothelial genes in iECs. In a murine model of hind-limb ischemia, iEC implantation increased capillary density and enhanced limb perfusion, demonstrating the in vivo viability and functionality of iECs.
  
  
  CONCLUSIONS
  We demonstrated the first direct conversion of adult fibroblasts to functional ECs. These results suggest a novel therapeutic modality for cell therapy in ischemic vascular disease.
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Jung-Kyu
authors.initials: JK
authors.lastname: Han
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Sung-Hwan
authors.initials: SH
authors.lastname: Chang
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Hyun-Ju
authors.initials: HJ
authors.lastname: Cho
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Saet-Byeol
authors.initials: SB
authors.lastname: Choi
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Hyo-Suk
authors.initials: HS
authors.lastname: Ahn
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Jaewon
authors.initials: J
authors.lastname: Lee
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Heewon
authors.initials: H
authors.lastname: Jeong
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Seock-Won
authors.initials: SW
authors.lastname: Youn
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Ho-Jae
authors.initials: HJ
authors.lastname: Lee
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Yoo-Wook
authors.initials: YW
authors.lastname: Kwon
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Hyun-Jai
authors.initials: HJ
authors.lastname: Cho
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Byung-Hee
authors.initials: BH
authors.lastname: Oh
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Peter
authors.initials: P
authors.lastname: Oettgen
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.).
authors.forename: Young-Bae
authors.initials: YB
authors.lastname: Park
authors.affiliation: From National Research Laboratory for Cardiovascular Stem Cell, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., S-H.C, Hyun-Ju C., S-B.C., H-S.A., J.L., H.J., S-W.Y., H-J.L., Y-W.K., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Republic of Korea (J-K.H., Hyun-Jai C., B-H.O., Y-B.P., H-S.K.); Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA (P.O.); and Molecular Medicine & Biopharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea (H-S.K.). hyosoo@snu.ac.kr.
authors.forename: Hyo-Soo
authors.initials: HS
authors.lastname: Kim
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism; D000367:Age Factors; D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology* / Q000502:physiology; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics; D006614:Hindlimb / Q000098:blood supply; D007511:Ischemia / Q000473:pathology / Q000628:therapy*; D060588:LIM Domain Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D008819:Mice, Nude; D008822:Mice, Transgenic; D015514:Oncogene Proteins, Fusion / Q000235:genetics / Q000378:metabolism; D042787:Receptor, TIE-2 / Q000235:genetics / Q000378:metabolism; D012867:Skin / Q000166:cytology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014652:Vascular Diseases / Q000473:pathology / Q000628:therapy*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C503994:ER71 protein, mouse; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors; C467200:Foxo1 protein, mouse; D060588:LIM Domain Proteins; C513188:Lmo2 protein, mouse; D015514:Oncogene Proteins, Fusion; C103486:SIL protein, mouse; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D042787:Receptor, TIE-2; C079086:Tek protein, mouse
keywords: cell transdifferentiation; endothelial cells; fibroblasts