TY - JOUR
T1 - A Comparative Proteomic Analysis of Extracellular Vesicles Associated With Lipotoxicity
AU - Nakao, Yasuhiko
AU - Fukushima, Masanori
AU - Mauer, Amy S.
AU - Liao, Chieh Yu
AU - Ferris, Anya
AU - Dasgupta, Debanjali
AU - Heppelmann, Carrie Jo
AU - Vanderboom, Patrick M.
AU - Saraswat, Mayank
AU - Pandey, Akhilesh
AU - Nair, K. Sreekumaran
AU - Allen, Alina M.
AU - Nakao, Kazuhiko
AU - Malhi, Harmeet
N1 - Funding Information:
This work is supported by NIH grant DK111378 (to HM) and DK115594 (to AA), the Mayo Foundation (to HM), the Clinical Core of the Mayo Clinic Center for Cell Signaling in Gastroenterology (P30DK084567), and Kanae Foundation Foreign Study Grant (to YN). We thank the Mayo Clinic Medical Genome Facility-Proteomics Core (a shared resource of the Mayo Clinic Cancer Center (NCI P30 CA15083)).
Publisher Copyright:
Copyright © 2021 Nakao, Fukushima, Mauer, Liao, Ferris, Dasgupta, Heppelmann, Vanderboom, Saraswat, Pandey, Nair, Allen, Nakao and Malhi.
PY - 2021/11/4
Y1 - 2021/11/4
N2 - Extracellular vesicles (EVs) are emerging mediators of intercellular communication in nonalcoholic steatohepatitis (NASH). Palmitate, a lipotoxic saturated fatty acid, activates hepatocellular endoplasmic reticulum stress, which has been demonstrated to be important in NASH pathogenesis, including in the release of EVs. We have previously demonstrated that the release of palmitate-stimulated EVs is dependent on the de novo synthesis of ceramide, which is trafficked by the ceramide transport protein, STARD11. The trafficking of ceramide is a critical step in the release of lipotoxic EVs, as cells deficient in STARD11 do not release palmitate-stimulated EVs. Here, we examined the hypothesis that protein cargoes are trafficked to lipotoxic EVs in a ceramide-dependent manner. We performed quantitative proteomic analysis of palmitate-stimulated EVs in control and STARD11 knockout hepatocyte cell lines. Proteomics was performed on EVs isolated by size exclusion chromatography, ultracentrifugation, and density gradient separation, and EV proteins were measured by mass spectrometry. We also performed human EV proteomics from a control and a NASH plasma sample, for comparative analyses with hepatocyte-derived lipotoxic EVs. Size exclusion chromatography yielded most unique EV proteins. Ceramide-dependent lipotoxic EVs contain damage-associated molecular patterns and adhesion molecules. Haptoglobin, vascular non-inflammatory molecule-1, and insulin-like growth factor-binding protein complex acid labile subunit were commonly detected in NASH and hepatocyte-derived ceramide-dependent EVs. Lipotoxic EV proteomics provides novel candidate proteins to investigate in NASH pathogenesis and as diagnostic biomarkers for hepatocyte-derived EVs in NASH patients.
AB - Extracellular vesicles (EVs) are emerging mediators of intercellular communication in nonalcoholic steatohepatitis (NASH). Palmitate, a lipotoxic saturated fatty acid, activates hepatocellular endoplasmic reticulum stress, which has been demonstrated to be important in NASH pathogenesis, including in the release of EVs. We have previously demonstrated that the release of palmitate-stimulated EVs is dependent on the de novo synthesis of ceramide, which is trafficked by the ceramide transport protein, STARD11. The trafficking of ceramide is a critical step in the release of lipotoxic EVs, as cells deficient in STARD11 do not release palmitate-stimulated EVs. Here, we examined the hypothesis that protein cargoes are trafficked to lipotoxic EVs in a ceramide-dependent manner. We performed quantitative proteomic analysis of palmitate-stimulated EVs in control and STARD11 knockout hepatocyte cell lines. Proteomics was performed on EVs isolated by size exclusion chromatography, ultracentrifugation, and density gradient separation, and EV proteins were measured by mass spectrometry. We also performed human EV proteomics from a control and a NASH plasma sample, for comparative analyses with hepatocyte-derived lipotoxic EVs. Size exclusion chromatography yielded most unique EV proteins. Ceramide-dependent lipotoxic EVs contain damage-associated molecular patterns and adhesion molecules. Haptoglobin, vascular non-inflammatory molecule-1, and insulin-like growth factor-binding protein complex acid labile subunit were commonly detected in NASH and hepatocyte-derived ceramide-dependent EVs. Lipotoxic EV proteomics provides novel candidate proteins to investigate in NASH pathogenesis and as diagnostic biomarkers for hepatocyte-derived EVs in NASH patients.
KW - DAMP
KW - StAR-related lipid transfer domain 11
KW - exosome
KW - hepatocyte
KW - microvesicle
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U2 - 10.3389/fcell.2021.735001
DO - 10.3389/fcell.2021.735001
M3 - Article
AN - SCOPUS:85119445637
VL - 9
JO - Frontiers in Cell and Developmental Biology
JF - Frontiers in Cell and Developmental Biology
SN - 2296-634X
M1 - 735001
ER -