Cillian Clancy1, Sonja Khan1, Claire L Glynn1, Emma Holian2, Peter Dockery3, Pierce Lalor3, James A L Brown1, Myles R Joyce1,4, Michael J Kerin1, Roisin M Dwyer1. 1. Discipline of Surgery, Lambe Institute for Translational Research, National University of Ireland Galway, Galway, Ireland. 2. School of Mathematics, Statistics and Applied Mathematics, National University of Ireland Galway, Galway, Ireland. 3. Discipline of Anatomy, National University of Ireland Galway, Galway, Ireland. 4. Department of Colorectal Surgery, Galway University Hospital, Galway, Ireland.
Abstract
BACKGROUND: Cells release extracellular membrane vesicles including microvesicles known as exosomes. Exosomes contain microRNAs (miRNAs) however the full range within colorectal cancer cell secreted exosomes is unknown. OBJECTIVE: To identify the full range of exosome encapsulated miRNAs secreted from 2 colorectal cancer cell lines and to investigate engineering of exosomes over-expressing miRNAs. METHODS: Exosomes were isolated from HCT-116 and HT-29 cell lines. RNA was extracted from exosomes and microRNA array performed. Cells were engineered to express miR-379 (HCT-116-379) or a non-targeting control (HCT-116-NTC) and functional effects were determined. Exosomes secreted by engineered cells were transferred to recipient cells and the impact examined. RESULTS: Microvesicles 40-100 nm in size secreted by cell lines were visualised and confirmed to express exosomal protein CD63. HT-29 exosomes contained 409 miRNAs, HCT-116 exosomes contained 393, and 338 were common to exosomes from both cell lines. Selected targets were validated. HCT-116-379 cells showed decreased proliferation (12-15% decrease, p < 0.001) and decreased migration (32-86% decrease, p < 0.001) compared to controls. HCT-116-379 exosomes were enriched for miR-379. Confocal microscopy visualised transfer of HCT-116-379 exosomes to recipient cells. CONCLUSIONS: Colorectal cancer cells secrete a large number of miRNAs within exosomes. miR-379 decreases cell proliferation and migration, and miR-379 enriched exosomes can be engineered.
BACKGROUND: Cells release extracellular membrane vesicles including microvesicles known as exosomes. Exosomes contain microRNAs (miRNAs) however the full range within colorectal cancer cell secreted exosomes is unknown. OBJECTIVE: To identify the full range of exosome encapsulated miRNAs secreted from 2 colorectal cancer cell lines and to investigate engineering of exosomes over-expressing miRNAs. METHODS: Exosomes were isolated from HCT-116 and HT-29 cell lines. RNA was extracted from exosomes and microRNA array performed. Cells were engineered to express miR-379 (HCT-116-379) or a non-targeting control (HCT-116-NTC) and functional effects were determined. Exosomes secreted by engineered cells were transferred to recipient cells and the impact examined. RESULTS: Microvesicles 40-100 nm in size secreted by cell lines were visualised and confirmed to express exosomal protein CD63. HT-29 exosomes contained 409 miRNAs, HCT-116 exosomes contained 393, and 338 were common to exosomes from both cell lines. Selected targets were validated. HCT-116-379 cells showed decreased proliferation (12-15% decrease, p < 0.001) and decreased migration (32-86% decrease, p < 0.001) compared to controls. HCT-116-379 exosomes were enriched for miR-379. Confocal microscopy visualised transfer of HCT-116-379 exosomes to recipient cells. CONCLUSIONS:Colorectal cancer cells secrete a large number of miRNAs within exosomes. miR-379 decreases cell proliferation and migration, and miR-379 enriched exosomes can be engineered.
Authors: K P O'Brien; S Khan; K E Gilligan; H Zafar; P Lalor; C Glynn; C O'Flatharta; H Ingoldsby; P Dockery; A De Bhulbh; J R Schweber; K St John; M Leahy; J M Murphy; W M Gallagher; T O'Brien; M J Kerin; R M Dwyer Journal: Oncogene Date: 2018-01-25 Impact factor: 9.867
Authors: Jonas Dohmen; Alexander Semaan; Makbule Kobilay; Martin Zaleski; Vittorio Branchi; Anja Schlierf; Karina Hettwer; Steffen Uhlig; Gunther Hartmann; Jörg C Kalff; Hanno Matthaei; Philipp Lingohr; Stefan Holdenrieder Journal: Diagnostics (Basel) Date: 2022-06-08