Kadri Rekker1, Merli Saare2, Anne Mari Roost3, Anna-Liisa Kubo4, Natasa Zarovni4, Antonio Chiesi4, Andres Salumets2, Maire Peters5. 1. Competence Centre on Reproductive Medicine and Biology, Tiigi 61b, 50410 Tartu, Estonia; Department of Obstetrics and Gynecology, University of Tartu, L. Puusepa 8, 51014 Tartu, Estonia. Electronic address: kadri.vaidla@gmail.com. 2. Competence Centre on Reproductive Medicine and Biology, Tiigi 61b, 50410 Tartu, Estonia; Department of Obstetrics and Gynecology, University of Tartu, L. Puusepa 8, 51014 Tartu, Estonia; Institute of Bio- and Translational Medicine, University of Tartu, Ravila 19, 50411 Tartu, Estonia. 3. Competence Centre on Reproductive Medicine and Biology, Tiigi 61b, 50410 Tartu, Estonia. 4. HansaBioMed Ltd, Akadeemia tee 15, 12618 Tallinn, Estonia. 5. Competence Centre on Reproductive Medicine and Biology, Tiigi 61b, 50410 Tartu, Estonia; Department of Obstetrics and Gynecology, University of Tartu, L. Puusepa 8, 51014 Tartu, Estonia.
Abstract
OBJECTIVES: Exosomes are small membrane bound vesicles secreted by most cell types. Exosomes contain various functional proteins, mRNAs and microRNAs (miRNAs) that could be used for diagnostic and therapeutic purposes. Currently, a standard method for serum exosome isolation is differential ultracentrifugation, but a search for alternative, less time-consuming and labour extensive exosomal isolation method for use in clinical settings is ongoing. The effect of serum exosome isolation method on obtained miRNA profile is not yet clear. The aim of this study was to determine to which extent selected exosome isolation methods influence the serum exosomal miRNA profile. DESIGN AND METHODS: Exosomes were isolated from blood serum of healthy individuals by ultracentrifugation and ExoQuick Precipitation methods. The expression profile of 375 miRNAs was determined by real time PCR using Exiqon miRCURY LNA™ microRNA Human panel I assays. RESULTS: Although a strong correlation of exosomal miRNA profiles was observed between the two isolation methods, distinct clusters of miRNA levels between the used methods were identified. The detected levels of two miRNAs, miR-92a and miR-486-5p, were significantly influenced by the exosome isolation method used. CONCLUSIONS: Both exosome isolation methods are suitable for serum exosomal miRNA profiling. Differences found in miRNA patterns between the two methods indicate that the observed exosomal miRNA profile is slightly affected by the extracellular vesicle isolation method.
OBJECTIVES: Exosomes are small membrane bound vesicles secreted by most cell types. Exosomes contain various functional proteins, mRNAs and microRNAs (miRNAs) that could be used for diagnostic and therapeutic purposes. Currently, a standard method for serum exosome isolation is differential ultracentrifugation, but a search for alternative, less time-consuming and labour extensive exosomal isolation method for use in clinical settings is ongoing. The effect of serum exosome isolation method on obtained miRNA profile is not yet clear. The aim of this study was to determine to which extent selected exosome isolation methods influence the serum exosomal miRNA profile. DESIGN AND METHODS: Exosomes were isolated from blood serum of healthy individuals by ultracentrifugation and ExoQuick Precipitation methods. The expression profile of 375 miRNAs was determined by real time PCR using Exiqon miRCURY LNA™ microRNA Human panel I assays. RESULTS: Although a strong correlation of exosomal miRNA profiles was observed between the two isolation methods, distinct clusters of miRNA levels between the used methods were identified. The detected levels of two miRNAs, miR-92a and miR-486-5p, were significantly influenced by the exosome isolation method used. CONCLUSIONS: Both exosome isolation methods are suitable for serum exosomal miRNA profiling. Differences found in miRNA patterns between the two methods indicate that the observed exosomal miRNA profile is slightly affected by the extracellular vesicle isolation method.
Authors: Abdelnaby Khalyfa; Leila Kheirandish-Gozal; Ahamed A Khalyfa; Mona F Philby; María Luz Alonso-Álvarez; Meelad Mohammadi; Rakesh Bhattacharjee; Joaquin Terán-Santos; Lei Huang; Jorge Andrade; David Gozal Journal: Am J Respir Crit Care Med Date: 2016-11-01 Impact factor: 21.405
Authors: Douglas W Van Pelt; Ivan J Vechetti; Marcus M Lawrence; Kathryn L Van Pelt; Parth Patel; Benjamin F Miller; Timothy A Butterfield; Esther E Dupont-Versteegden Journal: Am J Physiol Cell Physiol Date: 2020-07-08 Impact factor: 4.249