Extracellular vesicles from mice with alcoholic liver disease carry a distinct protein cargo and induce macrophage activation via Hsp90

Abstract

A salient feature of alcoholic liver disease (ALD) is Kupffer cell (KC) activation and recruitment of inflammatory monocytes/macrophages (Mo/MØ). These key cellular events of ALD pathogenesis may be mediated by extracellular vesicles (EVs). EVs transfer biomaterials, including proteins and miRNAs, and have recently emerged as important effectors of intercellular communication. We hypothesized that circulating EVs from mice with ALD have a protein cargo characteristic of the disease and mediate biological effects by activating immune cells. The total number of circulating EVs was increased in mice with ALD (ALD-EV) compared to pair-fed controls (control-EV). Mass spectrometry analysis of circulating EVs revealed a distinct signature for proteins involved in inflammatory responses, cellular development and cellular movement between ALD-EVs and control-EVs. We also identified uniquely important proteins in ALD-EVs that were not present in control-EVs. When ALD-EVs were injected intravenously into alcohol-naïve mice, we found evidence of uptake of ALD-EVs in recipient livers in hepatocytes and MØs. Hepatocytes isolated from mice after transfer of ALD-EVs, but not control-EVs, showed increased MCP-1 mRNA and protein expression suggesting a biological effect of ALD-EVs. Compared to control-EV recipient mice, ALD-EV recipient mice had increased numbers of F4/80^hiCD11b^lo KCs and increased percentages of TNFα⁺IL-12/23⁺ (inflammatory/M1) KCs and infiltrating monocytes (F4/80^intCD11b^hi) while the percentage of CD206⁺CD163⁺ (anti-inflammatory/M2) KCs was decreased. In vitro, ALD-EVs increased TNFα and IL-1β production in MØ and reduced CD163 and CD206 expression. We identified Heat shock protein 90 (Hsp90) in ALD-EVs as the mediator of ALD-EV-induced MØ activation. Conclusion: Our study indicates a specific protein signature of ALD-EVs and demonstrates a functional role of circulating EVs containing Hsp90 in mediating KC/MØ activation in the liver. This article is protected by copyright. All rights reserved.

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