title: Isolation of human trophoblastic extracellular vesicles and characterization of their cargo and antiviral activity.
abstract: Primary human trophoblasts release a repertoire of extracellular vesicles (EVs). Among them are nano-sized exosomes, which we found to suppress the replication of a wide range of diverse viruses. These exosomes contain trophoblastic microRNAs (miRNAs) that are expressed from the chromosome 19 miRNA cluster and exhibit antiviral properties. Here, we report our investigation of the cargo of placental EVs, focusing on the composition and the antiviral properties of exosomes, microvesicles, and apoptotic blebs.
  
  
  
  We isolated EVs using ultracentrifugation and defined their purity using immunoblotting, electron microscopy, and nanoparticle tracking. We used liquid chromatography-electrospray ionization-mass spectrometry, protein mass spectrometry, and miRNA TaqMan card PCR to examine the phospholipids, proteins, and miRNA cargo of trophoblastic EVs and an in vitro viral infection assay to assess the antiviral properties of EVs.
  
  
  
  We found that all three EV types contain a comparable repertoire of miRNA. Interestingly, trophoblastic exosomes harbor a protein and phospholipid profile that is distinct from that of microvesicles or apoptotic blebs. Functionally, trophoblastic exosomes exhibit the highest antiviral activity among the EVs. Consistently, plasma exosomes derived from pregnant women recapitulate the antiviral effect of trophoblastic exosomes derived from in vitro cultures of primary human trophoblasts.
  
  
  
  When compared to other trophoblastic EVs, exosomes exhibit a unique repertoire of proteins and phospholipids, but not miRNAs, and a potent viral activity. Our work suggests that human trophoblastic EVs may play a key role in maternal-placental-fetal communication.
authors: Magee-Womens Research Institute, Department of OBGYN and Reproductive Sciences, University of Pittsburgh, Pittsburgh, PA 15213, USA.
authors: Yingshi
authors: Y
authors: Ouyang
authors: Magee-Womens Research Institute, Department of OBGYN and Reproductive Sciences, University of Pittsburgh, Pittsburgh, PA 15213, USA.
authors: Avraham
authors: A
authors: Bayer
authors: Magee-Womens Research Institute, Department of OBGYN and Reproductive Sciences, University of Pittsburgh, Pittsburgh, PA 15213, USA.
authors: Tianjiao
authors: T
authors: Chu
authors: Department of Environmental and Occupational Health, Center for Free Radical and Antioxidant Health, University of Pittsburgh, Pittsburgh, PA 15219, USA.
authors: Vladimir A
authors: VA
authors: Tyurin
authors: Department of Environmental and Occupational Health, Center for Free Radical and Antioxidant Health, University of Pittsburgh, Pittsburgh, PA 15219, USA.
authors: Valerian E
authors: VE
authors: Kagan
authors: T.E. Starzl Institute and Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15213, USA.
authors: Adrian E
authors: AE
authors: Morelli
authors: Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pittsburgh, PA 15219, USA. Electronic address: coynec2@pitt.edu.
authors: Carolyn B
authors: CB
authors: Coyne
authors: Magee-Womens Research Institute, Department of OBGYN and Reproductive Sciences, University of Pittsburgh, Pittsburgh, PA 15213, USA; Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pittsburgh, PA 15219, USA. Electronic address: ysadovsky@mwri.magee.edu.
authors: Yoel
authors: Y
authors: Sadovsky
mesh_terms: D055354:Exosomes / Q000378:metabolism*; D000067128:Extracellular Vesicles / Q000378:metabolism*; D005260:Female; D006801:Humans; D013058:Mass Spectrometry; D035683:MicroRNAs / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D010920:Placenta / Q000166:cytology / Q000378:metabolism*; D011247:Pregnancy; D014327:Trophoblasts / Q000166:cytology / Q000378:metabolism*
chemical_list: D035683:MicroRNAs; D010743:Phospholipids
keywords: C19MC; Exosomes; Placenta; Plasma exosomes; Trophoblastic vesicles; Viral infection