Poster Presentation Australasian Extracellular Vesicles Conference 2018

Oxygen tension and diabetes environment regulates the expression of c19mc mirnas in placental cells and their derived exosomes (#88)

Carlos Palma 1 , Dominic Guanzon 1 , Dareen Almohammadi 1 , Katherin Scholz-Romero 1 2 , Felipe Zuñiga 2 , Emilio Diaz 3 , Harold David McIntyre 4 , Martha Lappas 5 6 , Carlos Salomon 1 2
  1. Exosome Biology Laboratory, Centre for Clinical Diagnostics, University of Queensland Centre for Clinical Research, Royal Brisbane and Women's Hospital, The University of Queensland , Brisbane, Queensland 4029, Australia
  2. Department of Clinical Biochemistry and Immunology, Faculty of Pharmacy, University of Concepción, Concepcion, Chile
  3. Faculty of Medicine, University of Concepcion, Concepcion, Chile
  4. Mater Research, Faculty of Medicine, The University of Queensland, Brisbane, QLD, Australia
  5. Obstetrics, Nutrition and Endocrinology Group, Department of Obstetrics and Gynaecology, University of Melbourne, Melbourne, Victoria, Australia
  6. Mercy Perinatal Research Centre, Mercy Hospital for Women, Melbourne, Victoria, Australia

The chromosome 19 miRNA cluster (C19MC) is the biggest miRNA gene cluster and is mainly expressed in the human placenta. In this study, we determined the relative expression of C19MC miRNAs in trophoblast cells cultured under diabetic conditions, exosomes released by these cells and exosomes isolated from plasma obtained from women diagnosed with gestational diabetes mellitus (GDM). Cells were incubated under High D-glucose (25 mM) and high insulin (1nM) under 1%, 3% or 8% O2for 48 h. Plasma samples were obtained from normal and GDM (n=12) pregnancy. Total cellular RNA was purified from cells and exosomes using miRNeasy Mini Kit (Qiagen). The relative expression of miR-512-5p, miR-517a, miR-518a-5p, miR-518b, miR-518c, miR-518e, miR-519d, miR-520c-3p, miR-525-5p and RNU6B was determined by RT-qPCR. C19MC miRNAs expression was normalised (ΔCT) to RNU6B and the data are presented as 2-ΔCT. Two-way ANOVA identified a significant effect (p<0.05) of oxygen, glucose, and insulin on all miRNAs studied except miR-512-5p. A significant effect of diabetic conditions was identified in exosomes on miR-525-5p, miR-520c-3p, miR-518c and  miR-518b. Interestingly, correlation analysis showed a positive correlation (p<0.05) between cells and exosomes under 5 mM D-glucose and 8% or 3% O2, however, 25 mM D-glucose and 1% O2abolished this association (p>0.05). Finally, the relative expression of exosomal miR-518a-5p, miR-518b, miR-518c, miR-518e, miR-520c-3p, and miR-525-5p was significantly higher in GDM compared to normal pregnancy. This study establishes that in response to metabolic challenges, trophoblast cells specifically package C19MC miRNA into the exosomes they release.