European Journal of Medical Genetics 63 (2020) 104038 Available online 18 August 2020 1769-7212/© 2020 Published by Elsevier Masson SAS. Altered level of plasma exosomes in patients with Gaucher disease Shtam Tatiana a, c , Naryzhny Stanislav a, d , Kulabukhova Darya a, b , Garaeva Luiza a, c , Senkevich Konstantin a, b, e , Landa Sergey a , Varfolomeeva Elena a, c , Salogub Galina e , Verlov Nikolai a, c , Kopylov Arthur d , Zorina Elena d , Kamyshinsky Roman c, f, g , Usenko Tatiana a, b , Schwarzman Alexander a , Zakharova Ekaterina a, h , Emelyanov Anton a, b , Pchelina Sofya a, b, c, * a Petersburg Nuclear Physics Institute Named By B.P. Konstantinov of National Research Centre Kurchatov Institute, Orlova Roscha 1, Gatchina, 188300, Russian Federation b First Pavlov State Medical University of St. Petersburg, L’va Tolstogo Str. 6-8, 197022, St. Petersburg, Russian Federation c National Research Center “Kurchatov Institute”, Akademika Kurchatova Pl. 1, 123182, Moscow, Russian Federation d Institute of Biomedical Chemistry, Russian Academy of Medical Sciences, Pogodinskaja Str. 10, 119832, Moscow, Russian Federation e Almazov National Medical Research Center, Akkuratova Street 1, 197341, St. Petersburg, Russian Federation f Shubnikov Institute of Crystallography of Federal Scientifc Research Centre ‘Crystallography and Photonics’ of Russian Academy of Sciences, Leninskiy Prospect, 59, 119333, Moscow, Russian Federation g Moscow Institute of Physics and Technology, Institutsky Lane 9, Dolgoprudny, 141700, Moscow, Russian Federation h Research Centre for Medical Genetics, Moskvorechye St. 1, 115522, Moscow, Russian Federation A R T I C L E INFO Keywords: Gaucher disease GBA mutations Lysosomes Exosomes Proteomics ABSTRACT Mutations in the glucocerebrosidase gene (GBA) cause Gaucher disease (GD), the lysosomal storage disorder (LSD), and are the most common genetic risk factor of Parkinson’s disease (PD). Lysosome functionality plays a critical role for secretion of extracellular vesicles (EVs) and their content. Here we compared EVs from the blood plasma of 8 GD patients and 8 controls in terms of amounts, size distribution, and composition of their protein cargo. EVs were isolated via sequential centrifugation and characterized by сryo-electron microscopy (cryo-EM), nanoparticle tracking analysis (NTA), and dynamic light scattering (DLS). The presence of exosomal markers HSP70 and tetrasponins were analyzed by Western blot and fow cytometry. Protein profling was performed by mass-spectrometry (shotgun analysis). Here, for the frst time we reported an increased size and altered morphology in exosomes derived from blood plasma of GD patients. An increased size of plasma exosomes from GD patients compared to controls was demonstrated by cryo-EM and DLS (р<0.0001, p < 0.001, respectively) and confrmed by mode size detected by NTA (p < 0.02). Cryo-EM demonstrated an increased number of double and multilayer vesicles in plasma EVs from GD patients. We found that the EVs were enriched with the surface exosomal markers (CD9, СD63, CD81) and an exosome-associated protein HSP70 in case of the patients with the disease. Proteomic profling of exo- somal proteins did not reveal any proteins associated with PD pathogenesis. Thus, we showed that lysosomal dysfunction in GD patients lead to a striking alteration of plasma exosomes in size and morphology. 1. Introduction Extracellular vesicles (EVs) are small membrane vesicles released from different types of cells. EVs are found in many human biological fuids (e.g. blood, urine, and cerebrospinal fuid) (Xu et al., 2018). There are several categories of EVs from which exosomes (30–150 nm) derived from the endosomal system and released out of cells upon the fusion of endosomes/multivesicular bodies with a cell membrane are character- ized by the specifc exosomal markers (CD9, CD63, CD81 and others). Exosomes are believed to be involved in cellular waste clearance as well as in intercellular communication (Johnstone, 2006). EVs extracted from peripheral body liquids could have specifc protein or RNA markers * Corresponding author. Petersburg Nuclear Physics Institute named by B.P. Konstantinov of National Research Centre Kurchatov Institute, Orlova roscha 1, Gatchina, 188300, Russian Federation. E-mail address: pchelina_sn@pnpi.nrcki.ru (P. Sofya). Contents lists available at ScienceDirect European Journal of Medical Genetics journal homepage: www.elsevier.com/locate/ejmg https://doi.org/10.1016/j.ejmg.2020.104038 Received 7 April 2020; Received in revised form 4 August 2020; Accepted 13 August 2020