title: Gestational diabetes mellitus impairs fetal endothelial cell functions through a mechanism involving microRNA-101 and histone methyltransferase enhancer of zester homolog-2.
abstract: OBJECTIVE
  Gestational diabetes mellitus (GDM) produces fetal hyperglycemia with increased lifelong risks for the exposed offspring of cardiovascular and other diseases. Epigenetic mechanisms induce long-term gene expression changes in response to in utero environmental perturbations. Moreover, microRNAs (miRs) control the function of endothelial cells (ECs) under physiological and pathological conditions and can target the epigenetic machinery. We investigated the functional and expressional effect of GDM on human fetal ECs of the umbilical cord vein (HUVECs). We focused on miR-101 and 1 of its targets, enhancer of zester homolog-2 (EZH2), which trimethylates the lysine 27 of histone 3, thus repressing gene transcription. EZH2 exists as isoforms α and β.
  
  
  RESULTS
  HUVECs were prepared from GDM or healthy pregnancies and tested in apoptosis, migration, and Matrigel assays. GDM-HUVECs demonstrated decreased functional capacities, increased miR-101 expression, and reduced EZH2- β and trimethylation of histone H3 on lysine 27 levels. MiR-101 inhibition increased EZH2 expression and improved GDM-HUVEC function. Healthy HUVECs were exposed to high or normal d-glucose concentration for 48 hours and then tested for miR-101 and EZH2 expression. Similar to GDM, high glucose increased miR-101 expression. Chromatin immunoprecipitation using an antibody for EZH2 followed by polymerase chain reaction analyses for miR-101 gene promoter regions showed that both GDM and high glucose concentration reduced EZH2 binding to the miR-101 locus in HUVECs. Moreover, EZH2-β overexpression inhibited miR-101 promoter activity in HUVECs.
  
  
  CONCLUSIONS
  GDM impairs HUVEC function via miR-101 upregulation. EZH2 is both a transcriptional inhibitor and a target gene of miR-101 in HUVECs, and it contributes to some of the miR-101-induced defects of GDM-HUVECs.
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Ilaria
authors.initials: I
authors.lastname: Floris
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Betty
authors.initials: B
authors.lastname: Descamps
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Antonella
authors.initials: A
authors.lastname: Vardeu
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Tijana
authors.initials: T
authors.lastname: Mitić
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Posadino
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Saran
authors.initials: S
authors.lastname: Shantikumar
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Graciela
authors.initials: G
authors.lastname: Sala-Newby
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Gianpiero
authors.initials: G
authors.lastname: Capobianco
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Mangialardi
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Lynsey
authors.initials: L
authors.lastname: Howard
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Salvatore
authors.initials: S
authors.lastname: Dessole
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Raul
authors.initials: R
authors.lastname: Urrutia
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.).
authors.forename: Gianfranco
authors.initials: G
authors.lastname: Pintus
authors.affiliation: From the Bristol Heart Institute, School of Clinical Sciences, University of Bristol, Bristol, United Kingdom (I.F., B.D., A.V., T.M., S.S., G.S.-N., G.M., L.H., C.E.); Laboratory of Vascular Biology, Department of Biomedical Science (I.F., A.V., M.P., G.P.) and Department of Obstetrics and Gynecology Unit (G.C., S.D.), University of Sassari, Sassari, Italy; Laboratory of Epigenetics and Chromatin Dynamics, Mayo Clinic, Rochester, MN (R.U.); and National Heart and Lung Institute, Imperial College London, London, United Kingdom (C.E.). Costanza.Emanueli@bristol.ac.uk.
authors.forename: Costanza
authors.initials: C
authors.lastname: Emanueli
mesh_terms: D017209:Apoptosis; D001665:Binding Sites; D016022:Case-Control Studies; D002465:Cell Movement; D002470:Cell Survival; D002478:Cells, Cultured; D016640:Diabetes, Gestational / Q000201:enzymology* / Q000235:genetics / Q000473:pathology / Q000503:physiopathology; D000071221:Enhancer of Zeste Homolog 2 Protein; D005260:Female; D005865:Gestational Age; D005947:Glucose / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells / Q000201:enzymology* / Q000473:pathology; D006801:Humans; D008745:Methylation; D035683:MicroRNAs / Q000378:metabolism*; D018919:Neovascularization, Physiologic; D010641:Phenotype; D063151:Polycomb Repressive Complex 2 / Q000235:genetics / Q000378:metabolism*; D011247:Pregnancy; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D015398:Signal Transduction; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection; D015854:Up-Regulation
chemical_list: D006657:Histones; C534072:MIRN101 microRNA, human; D035683:MicroRNAs; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D063151:Polycomb Repressive Complex 2; D005947:Glucose
keywords: EZH2 protein human; diabetes, gestational; endothelial cells; epigenomics; microRNAs; polycomb repressive complex 2