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 - 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
SN - 0168-8278
VL - 71
SP - 1193
EP - 1205
JO - Journal of hepatology
JF - Journal of hepatology
IS - 6
ER -