**Older Adults with Physical Frailty and Sarcopenia Show Increased Levels of Circulating Small Extracellular Vesicles with a Specific Mitochondrial Signature**

**Anna Picca 1,**†**, Ra**ff**aella Beli 2,**†**, Riccardo Calvani 1,\*, Hélio José Coelho-Júnior 3, Francesco Landi 1,3, Roberto Bernabei 1,3, Cecilia Bucci 2, Flora Guerra 2,\* and Emanuele Marzetti 1,3**


Received: 22 March 2020; Accepted: 13 April 2020; Published: 15 April 2020

**Abstract:** Mitochondrial dysfunction and systemic inflammation are major factors in the development of sarcopenia, but the molecular determinants linking the two mechanisms are only partially understood. The study of extracellular vesicle (EV) trafficking may provide insights into this relationship. Circulating small EVs (sEVs) from serum of 11 older adults with physical frailty and sarcopenia (PF&S) and 10 controls were purified and characterized. Protein levels of three tetraspanins (CD9, CD63, and CD81) and selected mitochondrial markers, including adenosine triphosphate 5A (ATP5A), mitochondrial cytochrome C oxidase subunit I (MTCOI), nicotinamide adenine dinucleotide reduced form (NADH):ubiquinone oxidoreductase subunit B8 (NDUFB8), NADH:ubiquinone oxidoreductase subunit S3 (NDUFS3), succinate dehydrogenase complex iron sulfur subunit B (SDHB), and ubiquinol-cytochrome C reductase core protein 2 (UQCRC2) were quantified by Western immunoblotting. Participants with PF&S showed higher levels of circulating sEVs relative to controls. Protein levels of CD9 and CD63 were lower in the sEV fraction of PF&S older adults, while CD81 was unvaried between groups. In addition, circulating sEVs from PF&S participants had lower amounts of ATP5A, NDUFS3, and SDHB. No signal was detected for MTCOI, NDUFB8, or UQCRC2 in either participant group. Our findings indicate that, in spite of increased sEV secretion, lower amounts of mitochondrial components are discarded through EV in older adults with PF&S. In-depth analysis of EV trafficking might open new venues for biomarker discovery and treatment development for PF&S.

**Keywords:** aging; biomarkers; mitophagy; mitochondrial dynamics; mitochondrial quality control; mitochondrial-derived vesicles (MDVs); exosomes; mitochondrial-lysosomal axis
