The Phenotypic Effects of Exosomes Secreted from Distinct Cellular Sources: a Comparative Study Based on miRNA Composition

Abstract

Exosomes are nano-sized vesicles composed of lipids, proteins, and nucleic acids. Their molecular landscape is diverse, and exosomes derived from different cell types have distinct biological activities. Since exosomes are now being utilized as delivery vehicles for exogenous therapeutic cargoes, their intrinsic properties and biological effects must be understood. We performed miRNA profiling and found substantial differences in the miRNA landscape of prostate cancer (PC3) and human embryonic kidney (HEK) 293 exosomes with little correlation in abundance of common miRNAs (R = 0.16). Using a systems-level bioinformatics approach, the most abundant miRNAs in PC3 exosomes but not HEK exosomes were predicted to significantly modulate integrin signaling, with integrin-β3 loss inducing macrophage M2 polarization. PC3 but not HEK exosomes downregulated integrin-β3 expression levels by 70%. There was a dose-dependent polarization of RAW 264.7 macrophages toward an M2 phenotype when treated with PC3-derived exosomes but not HEK-derived exosomes. Conversely, HEK exosomes, widely utilized as delivery vehicles, were predicted to target cadherin signaling, with experimental validation showing a significant increase in the migratory potential of MCF7 breast cancer cells treated with HEK exosomes. Even widely utilized exosomes are unlikely to be inert, and their intrinsic activity ought to be assessed before therapeutic deployment.

Boosting the Biogenesis and Secretion of Mesenchymal Stem Cell-Derived Exosomes

“…The size distribution and concentration of exosomes were characterized by nanoparticle tracking analysis (NTA) as previously described [17]. Briefly, exosomes were diluted in PBS and measured three times in three different areas for each sample, with the average used to determine exosome concentration.…”

Boosting the biogenesis and secretion of mesenchymal stem cell-derived exosomes

“…The size distribution and concentration of exosomes were characterized by nanoparticle tracking analysis (NTA) as previously described [7,8]. Briefly, exosomes were diluted in PBS and measured three times in three different areas for each sample, with the average used to determine exosome concentration.…”

“…Cellular viability was analyzed using the MTT assay as previously described [7,9,10]. MSCs were seeded into 96-well plates and cultured overnight.…”

Kinetics and Specificity of HEK293T Extracellular Vesicle Uptake using Imaging Flow Cytometry

“…EVs have also been engineered as transference vectors and loaded with therapeutic agents including gene therapies and chemical compounds as an alternative to nanoparticle therapeutics and delivery vectors [4,[10][11][12]. HEK293T cells have been widely used as EV producer cells due to their inherent rapid proliferation, high EV yield, and ease of genetic manipulation [13][14][15][16][17]. HEK293T EVs delivered gene therapies including miRNA therapeutics for breast cancer [12] and have been used to deliver chemotherapeutics and therapeutic protein constructs in a schwannoma model [18].…”