Extracellular vesicles released by hepatocytes from gastric infusion model of alcoholic liver disease contain a MicroRNA barcode that can be detected in blood

Abstract

Extracellular vesicles (EVs) released during cell stress, or demise, can contain a barcode of the cell origin including specific microRNAs (miRNAs). Here we tested the hypothesis that during early alcoholic steatohepatitis (ASH) development, hepatocytes (HCs) release EVs with a miRNA signature that can be measured in circulation. A time course experiment showed that after two weeks of intragastric infusion, a time-point that results in isolated steatosis, there was no increase of blood EVs. After four weeks of infusion mice developed features of early ASH accompanied by a marked increase in the level of EVs in blood (p<0.05), as well as and in culture media of isolated HCs (p<0.001) and hepatic macrophages (p<0.001) with HCs being the predominant source of EVs. The transcriptome analysis of HC-EVs from ASH mice detected differentially expressed miRNAs, including nine significantly up-regulated and four significantly down-regulated miRNAs. Target prediction and pathway analyses of the up-regulated miRNAs identified 121 potential target genes involved in inflammatory and cancer pathways such as NF-κB, EGF, Wnt, and Bcl2. Three miRNAs, let7f, miR-29a, and miR-340, were increased in blood EVs from ASH mice (p<0.05), but not in blood EVs from three other models of chronic liver injury including bile duct ligation, non-alcoholic steatohepatitis, and obese mice, as well as EVs released from hepatocytes exposed to ethanol. The blood EV level (p<0.01) and three miRNAs (p<0.05) were significantly increased in patients with ambulatory mild ALD as compared to non-alcoholics. Conclusions Our results reveal that damaged hepatocytes from ASH mice are a key EV source with a specific miRNA cargo, which are specific for ASH-related liver injury. These findings uncover EVs as a potentially novel diagnostic for ASH.

Mitochondrial DNA–enriched microparticles promote acute-on-chronic alcoholic neutrophilia and hepatotoxicity

“…Blockage of the CXCL1 receptor CXCR1/2 ameliorated alcoholic steatohepatitis in mice (14). In addition to chemokines, extracellular vesicles (EVs) are new players in cellto-cell communication and play an important role in promoting inflammation and neutrophil infiltration in a variety of diseases (15,16), including liver diseases (17)(18)(19)(20)(21)(22)(23)(24)(25). EVs are nanometer-sized, membrane-bound extracellular milieu vesicles derived from cells (17).…”

“…The functions of EVs are mediated by delivering their contents from one cell to another. For example, stressed hepatocytes can activate macrophages via the release of EVs that contain lipids, proteins, chemokines, and nucleic acids (e.g., mitochondrial DNA [mtDNA]) (17)(18)(19)(20)(21)(22)(23)(24)(25). However, whether EVs also contribute to the elevation of circulating and hepatic neutrophils induced by acute-on-chronic alcoholic liver injury in human and mice remains unknown.…”

Emricasan, a pan-caspase inhibitor, improves survival and portal hypertension in a murine model of common bile-duct ligation

“…In addition to the effects of pan-caspase inhibition on modulation of fibrogenic pathways, it has been recently shown to modulate the generation and release of hemodynamically-active microparticles (MPs) [17]. Indeed, MPs are small membrane-bound particles released from dying or activated cells in a process involving caspase 3 and Rho-associated kinase activation [4, 5]. Studies in patients with cirrhosis have demonstrated that MPs are increased in circulation of these patients and impair vasoconstrictor responses contributing to arterial vasodilatation associated with portal hypertension [6].…”

“…MPs are effective communicators that are generated by a cell of origin or parenteral cell and can act on a number of target cells in an autocrine or paracrine pathway in the extracellular environment where they are released, as well as in an endocrine manner, acting as long-range signals. Circulating MPs containing cytokeratin-18 and expressing phosphatidylserine on the surface are released by hepatocytes in a caspase dependent manner [4, 5] and have been shown to be increased in circulation of patients with cirrhosis, and in these patients to impair vasoconstrictor responses and decrease blood pressure [6], contributing to the arterial vasodilation that may represent an important contributor to the worsening of PHT. Broad-spectrum caspase inhibition via the use of pan-caspase inhibitors is a novel therapeutic approach that has been shown to have anti-inflammatory and anti-fibrotic effects in experimental models and in patients with chronic liver disease [7–10].…”

Targeting inflammation for the treatment of alcoholic liver disease

“…In addition to intracellular localization, miRNAs are also detected in body fluids, such as urine, bile, saliva, serum and plasma, et al (Mitchell et al, 2008; Shigehara et al, 2011; Weber et al, 2010). Recent studies have suggested that circulating miRNAs are present in either the protein fraction or extracellular vesicles (Bala et al, 2012; Eguchi et al, 2017; Momen-Heravi, Saha, et al, 2015). The high stability of circulating miRNAs makes them attractive for biomarker discovery in liver diseases (Szabo & Bala, 2013).…”