Cytokine-primed umbilical cord mesenchymal stem cells enhanced therapeutic effects of extracellular vesicles on osteoarthritic chondrocytes

ABSTRACT The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles (“MISEV”) guidelines for the field in 2014. We now update these “MISEV2014” guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points.

Mesenchymal stem cell (MSC) exosome was previously shown to be effective in repairing critical size osteochondral defects in an immunocompetent rat model. Here we investigate the cellular processes modulated by MSC exosomes and the mechanism of action underlying the exosome-mediated responses in cartilage repair. We observed that exosome-mediated repair of osteochondral defects was characterised by increased cellular proliferation and infiltration, enhanced matrix synthesis and a regenerative immune phenotype. Using chondrocyte cultures, we could attribute the rapid cellular proliferation and infiltration during exosome-mediated cartilage repair to exosomal CD73-mediated adenosine activation of AKT and ERK signalling. Inhibitors of AKT or ERK phosphorylation suppressed exosome-mediated increase in cell proliferation and migration but not matrix synthesis. The role of exosomal CD73 was confirmed by the attenuation of AKT and ERK signalling by AMPCP, a CD73 inhibitor and theophylline, an adenosine receptor antagonist. Exosome-treated defects also displayed a regenerative immune phenotype characterised by a higher infiltration of CD163+ regenerative M2 macrophages over CD86+ M1 macrophages, with a concomitant reduction in pro-inflammatory synovial cytokines IL-1β and TNF-α. Together, these observations demonstrated that the efficient osteochondral regeneration by MSC exosomes was effected through a coordinated mobilisation of multiple cell types and activation of several cellular processes.

A transient inflammatory signal can initiate an epigenetic switch from nontransformed to cancer cells via a positive feedback loop involving NF-kappaB, Lin28, let-7, and IL-6. We identify differentially regulated microRNAs important for this switch and putative transcription factor binding sites in their promoters. STAT3, a transcription factor activated by IL-6, directly activates miR-21 and miR-181b-1. Remarkably, transient expression of either microRNA induces the epigenetic switch. MiR-21 and miR-181b-1, respectively, inhibit PTEN and CYLD tumor suppressors, leading to increased NF-kappaB activity required to maintain the transformed state. These STAT3-mediated regulatory circuits are required for the transformed state in diverse cell lines and tumor growth in xenografts, and their transcriptional signatures are observed in colon adenocarcinomas. Thus, STAT3 is not only a downstream target of IL-6 but, with miR-21, miR-181b-1, PTEN, and CYLD, is part of the positive feedback loop that underlies the epigenetic switch that links inflammation to cancer. Copyright (c) 2010 Elsevier Inc. All rights reserved.