Different miRNA Profiles in Plasma Derived Small and Large Extracellular Vesicles from Patients with Neurodegenerative Diseases

Giulia Perini,Orietta Pansarasa,Stefano Bernuzzi,Raffaele Calogero ,Luca Diamanti,Alfredo Costa,Cristina Cereda,Maria Garofalo,Mauro Ceroni,Micol Avenali,Stella Gagliardi,Susanna Zucca,Daisy Sproviero,Michela Dell’Orco,Matteo Cotta Ramusino,Martina Olivero,Maddalena Arigoni,Nora I. Perrone‐Bizzozero ,Brigida Minafra,Roberta Zangaglia,Marta Giannini

doi:10.3390/ijms22052737

Abstract

Identifying biomarkers is essential for early diagnosis of neurodegenerative diseases (NDs). Large (LEVs) and small extracellular vesicles (SEVs) are extracellular vesicles (EVs) of different sizes and biological functions transported in blood and they may be valid biomarkers for NDs. The aim of our study was to investigate common and different miRNA signatures in plasma derived LEVs and SEVs of Alzheimer’s disease (AD), Parkinson’s disease (PD), Amyotrophic Lateral Sclerosis (ALS) and Fronto-Temporal Dementia (FTD) patients. LEVs and SEVs were isolated from plasma of patients and healthy volunteers (CTR) by filtration and differential centrifugation and RNA was extracted. Small RNAs libraries were carried out by Next Generation Sequencing (NGS). MiRNAs discriminate all NDs diseases from CTRs and they can provide a signature for each NDs. Common enriched pathways for SEVs were instead linked to ubiquitin mediated proteolysis and Toll-like receptor signaling pathways and for LEVs to neurotrophin signaling and Glycosphingolipid biosynthesis pathway. LEVs and SEVs are involved in different pathways and this might give a specificity to their role in the spreading of the disease. The study of common and different miRNAs transported by LEVs and SEVs can be of great interest for biomarker discovery and for pathogenesis studies in neurodegeneration.

Highlights

Neurodegenerative disorders (NDs) are a group of diseases characterized by loss of neurons within the brain and/or spinal cord [1]
In SEVS from Parkinson’s disease (PD) patients Reactome analysis identified signaling by TGF-beta family members, SLC-mediated transmembrane transport
Reactome underlined the role of miRNAs in LEVs (Table S2): (1) from Amyotrophic Lateral Sclerosis (ALS) patients in intracellular signaling by second messengers (DAG, cAMP, cGMP, IP3, Ca2+ and phosphatidylinositols), signaling by TGF-beta family members, MyD88 cascade initiated on plasma membrane and metabolism of lipids; (2) from PD patients in intracellular signaling by second messengers, SLC-mediated transmembrane transport, some of which mediate neurotransmitter uptake in the CNS and peripheral nervous system (PNS) and metabolism of lipids

Summary

Introduction

Neurodegenerative disorders (NDs) are a group of diseases characterized by loss of neurons within the brain and/or spinal cord [1]. Some studies have examined miRNAs and RNAs in EVs isolated from cultured cell media from the Central Nervous System (CNS) cells (e.g., neurons, astrocytes, microglia, and oligodendrocytes) and few have examined miRNAs in EVs in plasma of AD, PD and ALS [25,26,27]. None of these studies considered the differences between SEVs from LEVs. We previously described in ALS that SEVs and LEVs in plasma are different in dimensions and for loading of some pathological proteins for ALS (SOD1, TDP-43, p-TDP-43, and FUS) and lipids [28,29,30]. The aim was to identify common and specific small RNAs between the two subpopulation of EVs in the same NDs disease and in the four diseases in order to identify new biomarkers

Results

Specific miRNAs Pathway Analysis in SEVs of NDs

Specific miRNAs Pathway Analysis in LEVs of NDs

Discussion

Study Subjects

LEVs and SEVs Isolation

RNA Libraries Preparations

Bioinformatic Data Analysis

Conclusions