Revealing novel molecular pathways in exosome production and pathological effects in the context of autophagy deficiency

Abstract

Extracellular vehicles (EVs) are defined as membrane‐surrounded, nanometer‐sized vesicles released by cells into the extracellular space in a highly regulated manner. Exosomes, a subclass of EVs with an average size of 40 to 160 nm in diameter, play critical roles in cell‐cell communications and in pathogenesis of many diseases, including liver diseases. However, the molecular mechanism of exosome production in specific disease scenario is not known. Here we use a mouse model of liver injury caused by autophagy deficiency to investigate the induction mechanism and the role of exosome in the pathogenesis.We found that there was an increased serum presence of liver‐specific exosomes when hepatic autophagy function was disabled through nanoparticle tracking analysis (NTA). These EVs had the average size of 100 nm, and expressed hepatocyte‐derived TSG101 and CYP2E1. Notably we defined specific markers that had not been reported before, such as Syntenin‐2, which is a Syndecan‐binding protein. This molecule is closely related to the formation of multivesicular bodies (MVBs), which is considered as the precursor of EVs. Exosome production can be increased or decreased by Syntenin‐2 overexpression or knockdown, respectively, indicating that this molecule is crucial for the exosome production. In addition, increased expression of Syntenin‐2 and increased production of exosomes in autophagy deficient mice were reversed by co‐deletion of an anti‐oxidative gene, Nrf2. Thus, we have defined a new pathway from Nrf2 to Syntenin‐2 in controlling the production of a liver specific exosome in the context of autophagy deficiency.To under the pathological role of these exosomes, blood EVs isolated from autophagy deficient mice were incubated with macrophages derived from the bone marrows (BMDM), which are recruited to the liver and contribute to the hepatic inflammation. We found that BMDM could effectively internalize the exosomes and are then activated for the production of a number of pro‐inflammatory genes compared to BMDM co‐cultured with exosomes isolated from wild type mice. Sequence profiling of miRNAs carried by these exosomes identified miR‐3072‐3p that can stimulate BMDM to upregulate the expression of the pro‐inflammatory genes, suggesting that these cells can be activated by the exosomes via miRNA‐mediated signaling.In summary, our work has revealed novel molecular components in promoting the production of liver‐specific exosomes under autophagy deficiency, which in turn enhances the inflammatory response in the liver through a unique miRNA‐mediated pathway.