TY - JOUR
T1 - Integrin β1-enriched extracellular vesicles mediate monocyte adhesion and promote liver inflammation in murine NASH
AU - Guo, Qianqian
AU - Furuta, Kunimaro
AU - Lucien, Fabrice
AU - Gutierrez Sanchez, Luz Helena
AU - Hirsova, Petra
AU - Krishnan, Anuradha
AU - Kabashima, Ayano
AU - Pavelko, Kevin D.
AU - Madden, Benjamin
AU - Alhuwaish, Husam
AU - Gao, Yandong
AU - Revzin, Alexander
AU - Ibrahim, Samar H.
N1 - Funding Information:
We thank Dr. Gregory J. Gores for his thorough review of the manuscript. We thank Dr. Nathan K. LeBrasseur and his laboratory members, especially Dr. Thomas White, for their assistance in the Comprehensive Laboratory Animal Monitoring System data. We thank Dr. Bing Q. Huang for his help in the studies employing electron microscopy. We also thank Dr. Cristine Charlesworth, and the Mayo Clinic Medical Genome Facility-Proteomics Core and its supporting grant, NCI Cancer Center Support Grant 5P30 CA15083-43C1 for performing the mass spectrometry analysis, as well as the optical microscopy core NIDDK P30DK084567.
Funding Information:
This work was supported by NIH grant DK111397 (to SHI), North American Society of Pediatric Gastroenterology Hepatology and Nutrition Young Investigator Award/Nestle Nutrition Award and Gilead Science career development award (to SHI), the Mayo Clinic K2R pipeline and Children Research Center. F.L is funded by a postdoctoral fellowship from the Fonds de Recherche du Quebec-Sante (FRQS).
Funding Information:
This work was supported by NIH grant DK111397 (to SHI), North American Society of Pediatric Gastroenterology Hepatology and Nutrition Young Investigator Award /Nestle Nutrition Award and Gilead Science career development award (to SHI), the Mayo Clinic K2R pipeline and Children Research Center. F.L is funded by a postdoctoral fellowship from the Fonds de Recherche du Quebec-Sante (FRQS).
Publisher Copyright:
© 2019 European Association for the Study of the Liver
PY - 2019/12
Y1 - 2019/12
N2 - Background & Aims: Hepatic recruitment of monocyte-derived macrophages (MoMFs) contributes to the inflammatory response in non-alcoholic steatohepatitis (NASH). However, how hepatocyte lipotoxicity promotes MoMF inflammation is unclear. Here we demonstrate that lipotoxic hepatocyte-derived extracellular vesicles (LPC-EVs) are enriched with active integrin β1 (ITGβ1), which promotes monocyte adhesion and liver inflammation in murine NASH. Methods: Hepatocytes were treated with either vehicle or the toxic lipid mediator lysophosphatidylcholine (LPC); EVs were isolated from the conditioned media and subjected to proteomic analysis. C57BL/6J mice were fed a diet rich in fat, fructose, and cholesterol (FFC) to induce NASH. Mice were treated with anti-ITGβ1 neutralizing antibody (ITGβ1Ab) or control IgG isotype. Results: Ingenuity® Pathway Analysis of the LPC-EV proteome indicated that ITG signaling is an overrepresented canonical pathway. Immunogold electron microscopy and nanoscale flow cytometry confirmed that LPC-EVs were enriched with activated ITGβ1. Furthermore, we showed that LPC treatment in hepatocytes activates ITGβ1 and mediates its endocytic trafficking and sorting into EVs. LPC-EVs enhanced monocyte adhesion to liver sinusoidal cells, as observed by shear stress adhesion assay. This adhesion was attenuated in the presence of ITGβ1Ab. FFC-fed, ITGβ1Ab-treated mice displayed reduced inflammation, defined by decreased hepatic infiltration and activation of proinflammatory MoMFs, as assessed by immunohistochemistry, mRNA expression, and flow cytometry. Likewise, mass cytometry by time-of-flight on intrahepatic leukocytes showed that ITGβ1Ab reduced levels of infiltrating proinflammatory monocytes. Furthermore, ITGβ1Ab treatment significantly ameliorated liver injury and fibrosis. Conclusions: Lipotoxic EVs mediate monocyte adhesion to LSECs mainly through an ITGβ1-dependent mechanism. ITGβ1Ab ameliorates diet-induced NASH in mice by reducing MoMF-driven inflammation, suggesting that blocking ITGβ1 is a potential anti-inflammatory therapeutic strategy in human NASH. Lay summary: Herein, we report that a cell adhesion molecule termed integrin β1 (ITGβ1) plays a key role in the progression of non-alcoholic steatohepatitis (NASH). ITGβ1 is released from hepatocytes under lipotoxic stress as a cargo of extracellular vesicles, and mediates monocyte adhesion to liver sinusoidal endothelial cells, which is an essential step in hepatic inflammation. In a mouse model of NASH, blocking ITGβ1 reduces liver inflammation, injury and fibrosis. Hence, ITGβ1 inhibition may serve as a new therapeutic strategy for NASH.
AB - Background & Aims: Hepatic recruitment of monocyte-derived macrophages (MoMFs) contributes to the inflammatory response in non-alcoholic steatohepatitis (NASH). However, how hepatocyte lipotoxicity promotes MoMF inflammation is unclear. Here we demonstrate that lipotoxic hepatocyte-derived extracellular vesicles (LPC-EVs) are enriched with active integrin β1 (ITGβ1), which promotes monocyte adhesion and liver inflammation in murine NASH. Methods: Hepatocytes were treated with either vehicle or the toxic lipid mediator lysophosphatidylcholine (LPC); EVs were isolated from the conditioned media and subjected to proteomic analysis. C57BL/6J mice were fed a diet rich in fat, fructose, and cholesterol (FFC) to induce NASH. Mice were treated with anti-ITGβ1 neutralizing antibody (ITGβ1Ab) or control IgG isotype. Results: Ingenuity® Pathway Analysis of the LPC-EV proteome indicated that ITG signaling is an overrepresented canonical pathway. Immunogold electron microscopy and nanoscale flow cytometry confirmed that LPC-EVs were enriched with activated ITGβ1. Furthermore, we showed that LPC treatment in hepatocytes activates ITGβ1 and mediates its endocytic trafficking and sorting into EVs. LPC-EVs enhanced monocyte adhesion to liver sinusoidal cells, as observed by shear stress adhesion assay. This adhesion was attenuated in the presence of ITGβ1Ab. FFC-fed, ITGβ1Ab-treated mice displayed reduced inflammation, defined by decreased hepatic infiltration and activation of proinflammatory MoMFs, as assessed by immunohistochemistry, mRNA expression, and flow cytometry. Likewise, mass cytometry by time-of-flight on intrahepatic leukocytes showed that ITGβ1Ab reduced levels of infiltrating proinflammatory monocytes. Furthermore, ITGβ1Ab treatment significantly ameliorated liver injury and fibrosis. Conclusions: Lipotoxic EVs mediate monocyte adhesion to LSECs mainly through an ITGβ1-dependent mechanism. ITGβ1Ab ameliorates diet-induced NASH in mice by reducing MoMF-driven inflammation, suggesting that blocking ITGβ1 is a potential anti-inflammatory therapeutic strategy in human NASH. Lay summary: Herein, we report that a cell adhesion molecule termed integrin β1 (ITGβ1) plays a key role in the progression of non-alcoholic steatohepatitis (NASH). ITGβ1 is released from hepatocytes under lipotoxic stress as a cargo of extracellular vesicles, and mediates monocyte adhesion to liver sinusoidal endothelial cells, which is an essential step in hepatic inflammation. In a mouse model of NASH, blocking ITGβ1 reduces liver inflammation, injury and fibrosis. Hence, ITGβ1 inhibition may serve as a new therapeutic strategy for NASH.
KW - Adhesion
KW - Extracellular vesicles
KW - Fibrosis
KW - Inflammation
KW - Integrin α
KW - Integrin β
KW - Liver sinusoidal endothelial cells
KW - Mass cytometry
KW - Monocytes
KW - NASH
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U2 - 10.1016/j.jhep.2019.07.019
DO - 10.1016/j.jhep.2019.07.019
M3 - Article
C2 - 31433301
AN - SCOPUS:85073027735
VL - 71
SP - 1193
EP - 1205
JO - Journal of Hepatology
JF - Journal of Hepatology
SN - 0168-8278
IS - 6
ER -