title: ASGR1 expressed by porcine enriched liver sinusoidal endothelial cells mediates human platelet phagocytosis in vitro.
abstract: BACKGROUND
  Porcine liver xenografts represent a potential solution to the organ shortage, but thrombocytopenia occurs within minutes to hours after xenotransplantation, preventing clinical application. Recently, it was discovered that porcine liver sinusoidal endothelial cells (LSEC) bind and phagocytose human platelets. We examined the role of ASGR1 in binding and removing human platelets by the pig liver endothelium.
  
  
  METHODS
  Primary porcine enriched LSEC (eLSEC) were characterized by flow cytometry, immunoblot, quantitative PCR, and immunohistochemistry using confocal microscopy. Phagocytosis inhibition assays using anti-ASGR1 and an ASGR1 substrate were performed. ASGR1 was targeted for siRNA knockdown, and ASGR1-reduced cells were tested for human platelet binding and phagocytosis.
  
  
  RESULTS
  ASGR1 is expressed by eLSEC. Human platelet binding and phagocytosis by porcine eLSEC was inhibited by asialofetuin, but not fetuin, suggesting an interaction with galactose β1-4 N-acetyl glucosamine. Anti-ASGR1 antibodies inhibited human platelet binding in a dose-dependent manner. Knockdown experiments using siRNA reduced ASGR1 expression in asynchronous primary eLSEC by 40%-80%. There was a 20% reduction in translated protein significantly correlated with a 21% decrease in human platelet binding.
  
  
  CONCLUSIONS
  ASGR1 on porcine eLSEC mediates phagocytosis of xenogeneic platelets.
authors.affiliation: Department of Surgery, Indiana University School of Medicine, IU Health Transplant Institute, Indianapolis, IN 46202, USA.
authors.forename: Leela L
authors.initials: LL
authors.lastname: Paris
authors.forename: Ray K
authors.initials: RK
authors.lastname: Chihara
authors.forename: Luz M
authors.initials: LM
authors.lastname: Reyes
authors.forename: Richard A
authors.initials: RA
authors.lastname: Sidner
authors.forename: Jose L
authors.initials: JL
authors.lastname: Estrada
authors.forename: Susan M
authors.initials: SM
authors.lastname: Downey
authors.forename: Daniel P
authors.initials: DP
authors.lastname: Milgrom
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Milgrom
authors.forename: A Joseph
authors.initials: AJ
authors.lastname: Tector
authors.forename: Christopher
authors.initials: C
authors.lastname: Burlak
mesh_terms: D000818:Animals; D037263:Asialoglycoprotein Receptor / Q000235:genetics / Q000378:metabolism*; D001792:Blood Platelets / Q000378:metabolism*; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism*; D006801:Humans; D008099:Liver / Q000166:cytology*; D010587:Phagocytosis / Q000502:physiology*; D017713:Platelet Transfusion; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D013552:Swine; D014183:Transplantation, Heterologous
chemical_list: D037263:Asialoglycoprotein Receptor; D034741:RNA, Small Interfering