Extracellular vesicles (EVs) have emerged as important mediators of intercellular communication in a diverse range of biological processes. For future therapeutic applications and for EV biology research in general, understanding the in vivo fate of EVs is of utmost importance. Here we studied biodistribution of EVs in mice after systemic delivery. EVs were isolated from 3 different mouse cell sources, including dendritic cells (DCs) derived from bone marrow, and labelled with a near‐infrared lipophilic dye. Xenotransplantation of EVs was further carried out for cross‐species comparison. The reliability of the labelling technique was confirmed by sucrose gradient fractionation, organ perfusion and further supported by immunohistochemical staining using CD63‐EGFP probed vesicles. While vesicles accumulated mainly in liver, spleen, gastrointestinal tract and lungs, differences related to EV cell origin were detected. EVs accumulated in the tumour tissue of tumour‐bearing mice and, after introduction of the rabies virus glycoprotein‐targeting moiety, they were found more readily in acetylcholine‐receptor‐rich organs. In addition, the route of administration and the dose of injected EVs influenced the biodistribution pattern. This is the first extensive biodistribution investigation of EVs comparing the impact of several different variables, the results of which have implications for the design and feasibility of therapeutic studies using EVs.

Alpha-synuclein aggregation plays a central role in Parkinson's disease pathology. Direct transmission of alpha-synuclein from pathologically affected to healthy unaffected neurons may be important in the anatomical spread of the disease through the nervous system. We have demonstrated that exosomes released from alpha-synuclein over-expressing SH-SY5Y cells contained alpha-synuclein and these exosomes were capable of efficiently transferring alpha-synuclein protein to normal SH-SY5Y cells. Moreover, the incubation of cells with ammonium chloride or bafilomycin A1 to produce the lysosomal dysfunction recently reported in Parkinson's disease led to an increase in the release of alpha-synuclein in exosomes and a concomitant increase in alpha-synuclein transmission to recipient cells. This study clearly demonstrates the importance of exosomes in both the release of alpha synuclein and its transmission between cells and suggests that factors associated with PD pathology accelerate this process. These mechanisms may play an important role in PD pathology and provide a suitable target for therapeutic intervention.

The identification of α-synuclein gene mutations and multiplications as a cause of familial Parkinson disease (PD) and the presence of this protein as a major constituent of Lewy bodies (LBs) in the PD brain have focused attention on the intracellular metabolism of this protein as a central component of the pathogenetic pathway(s) of PD. Intracellular protein degradation is achieved through a combination of the ubiquitin-dependent proteasomal system (UPS) and autophagic pathways. Abnormalities of UPS function have been reported in the PD brain,¹ and inhibitors of the UPS have been reported to induce a rodent model of PD in some^2,3 but not all studies.^4,5 However, the role of the UPS in α-synuclein metabolism remains unclear. Attention has recently focused on the potential contribution of autophagy to the pathogenesis of neurodegenerative diseases.^6-8 Autophagy is composed of 3 separate pathways: macroautophagy, microautophagy, and chaperone-mediated autophagy. Macroautophagy involves the formation of double-membraned autophagosomes, which fuse with lysosomes to deliver cytoplasmic contents, including misfolded or aggregated proteins for digestion and recycling of amino acids. Microautophagy involves lysosomal pinocytosis of cytoplasmic contents and is involved in the turnover of long half-life cytosolic proteins. Chaperone-mediated autophagy is dependent on the heat shock cognate 70 (hsc70) protein and its binding to lysosomal-associated membrane protein 2A (LAMP2A), a lysosomal surface receptor. A highly specific subset of cytosolic proteins with a KFREQ motif are recognized by the hsc70 chaperone and internalized for degradation by LAMP2A lysosomal membrane receptors.⁹

Alpha-synuclein (α-Syn) aggregates are the main component of Lewy bodies, which are the characteristic pathological feature in Parkinson's disease (PD) brain. Evidence that α-Syn aggregation can be propagated between neurones has led to the suggestion that this mechanism is responsible for the stepwise progression of PD pathology. Decreasing α-Syn expression is predicted to attenuate this process and is thus an attractive approach to delay or halt PD progression. We have used α-Syn small interfering RNA (siRNA) to reduce total and aggregated α-Syn levels in mouse brains. To achieve widespread delivery of siRNAs to the brain we have peripherally injected modified exosomes expressing Ravies virus glycoprotein loaded with siRNA. Normal mice were analyzed 3 or 7 days after injection. To evaluate whether this approach can decrease α-Syn aggregates, we repeated the treatment using transgenic mice expressing the human phosphorylation-mimic S129D α-Syn, which exhibits aggregation. In normal mice we detected significantly reduced α-Syn messenger RNA (mRNA) and protein levels throughout the brain 3 and 7 days after treatment with RVG-exosomes loaded with siRNA to α-Syn. In S129D α-Syn transgenic mice we found a decreased α-Syn mRNA and protein levels throughout the brain 7 days after injection. This resulted in significant reductions in intraneuronal protein aggregates, including in dopaminergic neurones of the substantia nigra. This study highlights the therapeutic potential of RVG-exosome delivery of siRNA to delay and reverse brain α-Syn pathological conditions. © 2014 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.