title: The non-targeted effects of radiation are perpetuated by exosomes.
abstract: Exosomes contain cargo material from endosomes, cytosol, plasma membrane and microRNA molecules, they are released by a number of non-cancer and cancer cells into both the extracellular microenvironment and body fluids such as blood plasma. Recently we demonstrated radiation-induced non-targeted effects [NTE: genomic instability (GI) and bystander effects (BE)] are partially mediated by exosomes, particularly the RNA content. However the mechanistic role of exosomes in NTE is yet to be fully understood. The present study used MCF7 cells to characterise the longevity of exosome-induced activity in the progeny of irradiated and unirradiated bystander cells. Exosomes extracted from conditioned media of irradiated and bystander progeny were added to unirradiated cells. Analysis was carried out at 1 and 20/24 population doublings following medium/exosome transfer for DNA/chromosomal damage. Results confirmed exosomes play a significant role in mediating NTE of ionising radiation (IR). This effect was remarkably persistent, observed >20 doublings post-irradiation in the progeny of bystander cells. Additionally, cell progeny undergoing a BE were themselves capable of inducing BE in other cells via exosomes they released. Furthermore we investigated the role of exosome cargo. Culture media from cells exposed to 2 Gy X-rays was subjected to ultracentrifugation and four inoculants prepared, (a) supernatants with exosomes removed, and pellets with (b) exosome proteins denatured, (c) RNA degraded, and (d) a combination of protein-RNA inactivation. These were added to separate populations of unirradiated cells. The BE was partially inhibited when either exosome protein or exosome RNA were inactivated separately, whilst combined RNA-protein inhibition significantly reduced or eliminated the BE. These results demonstrate that exosomes are associated with long-lived signalling of the NTE of IR. Both RNA and protein molecules of exosomes work in a synergistic manner to initiate NTE, spread these effects to naïve cells, and perpetuate GI in the affected cells.
authors.affiliation: Genomic Instability Group, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP, United Kingdom.
authors.forename: Ammar
authors.initials: A
authors.lastname: Al-Mayah
authors.affiliation: Genomic Instability Group, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP, United Kingdom.
authors.forename: Scott
authors.initials: S
authors.lastname: Bright
authors.affiliation: Genomic Instability Group, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP, United Kingdom.
authors.forename: Kim
authors.initials: K
authors.lastname: Chapman
authors.affiliation: Insect Virus Research Group, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP, United Kingdom.
authors.forename: Sarah
authors.initials: S
authors.lastname: Irons
authors.affiliation: Izon Science Ltd., The Oxford Science Park, Magdalen Centre, Robert Robinson Avenue, Oxford OX4 4GA, United Kingdom.
authors.forename: Ping
authors.initials: P
authors.lastname: Luo
authors.affiliation: Chromatin and non-coding RNA, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP, United Kingdom.
authors.forename: David
authors.initials: D
authors.lastname: Carter
authors.affiliation: The New Mexico Consortium, Los Alamos, NM 87544, USA.
authors.forename: Edwin
authors.initials: E
authors.lastname: Goodwin
authors.affiliation: Genomic Instability Group, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP, United Kingdom. Electronic address: mkadhim@brookes.ac.uk.
authors.forename: Munira
authors.initials: M
authors.lastname: Kadhim
mesh_terms: D024201:Bystander Effect / Q000528:radiation effects*; D045744:Cell Line, Tumor; D055354:Exosomes / Q000378:metabolism*; D005260:Female; D042822:Genomic Instability / Q000528:radiation effects*; D006801:Humans; D012313:RNA / Q000378:metabolism*; D015398:Signal Transduction / Q000528:radiation effects*; D014965:X-Rays
chemical_list: D012313:RNA
keywords: Bystander effects; Exosome cargo; Exosomes; Genomic instability; Ionising radiation