title: Primary porcine Kupffer cell phagocytosis of human platelets involves the CD18 receptor.
abstract: BACKGROUND
  Hepatic failure has been treated successfully with clinical extracorporeal perfusions of porcine livers. However, dog-to-pig and pig-to-baboon liver xenotransplant models have resulted in severe bleeding secondary to liver xenograft-induced thrombocytopenia. Kupffer cells (KC) are abundant phagocytic cells in the liver. KC express the CD11b/CD18 receptor, which has been implicated in chilled platelet binding and phagocytosis through interaction with platelet surface proteins and carbohydrates. We sought to identify the role of KC CD18 in liver xenograft-induced thrombocytopenia.
  
  
  METHODS
  Primary pig KC were characterized by flow cytometry, immunoblots, and quantitative polymerase chain reaction. Pig KC were used in inhibition assays with fluorescently labeled human platelets. The CD18 receptor was targeted for siRNA knockdown.
  
  
  RESULTS
  Domestic and α1,3-galactosyltransferase double knockout porcine KC cultures were approximately 92% positive for CD18 as detected by quantitative polymerase chain reaction and flow cytometry. Use of CD18 blocking antibodies resulted in reduction of human platelet binding and phagocytosis. Additionally, asialofetuin, not fetuin, inhibited platelet phagocytosis suggesting the involvement of an oligosaccharide-binding site. Furthermore, reduced CD18 expression by siRNA resulted in decreased human platelet binding.
  
  
  CONCLUSIONS
  Our data suggest that primary pig KC bind and phagocytose human platelets with involvement of CD18. Further understanding and modification of CD18 expression in pigs may result in a liver xenograft with reduced thrombocytopenic effects, which could be used as a bridge to allogeneic liver transplantation.
authors.affiliation: Department of Surgery, Indiana University Transplant Institute, Indiana University School of Medicine, Indianapolis, IN 46202, USA. Cburlak@iupui.edu
authors.forename: Ray K
authors.initials: RK
authors.lastname: Chihara
authors.forename: Leela L
authors.initials: LL
authors.lastname: Paris
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: Zheng-Yu
authors.initials: ZY
authors.lastname: Wang
authors.forename: A Joseph
authors.initials: AJ
authors.lastname: Tector
authors.forename: Christopher
authors.initials: C
authors.lastname: Burlak
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D001212:Asialoglycoproteins / Q000494:pharmacology; D001792:Blood Platelets / Q000502:physiology*; D018821:CD18 Antigens / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D002478:Cells, Cultured; D057899:Cytophagocytosis / Q000187:drug effects / Q000502:physiology*; D060748:Fetuins / Q000494:pharmacology; D006801:Humans; D007728:Kupffer Cells / Q000502:physiology*; D016031:Liver Transplantation / Q000009:adverse effects; D023421:Models, Animal; D034741:RNA, Small Interfering / Q000494:pharmacology; D013552:Swine; D013556:Swine, Miniature; D013921:Thrombocytopenia / Q000209:etiology; D014183:Transplantation, Heterologous / Q000009:adverse effects
chemical_list: D001212:Asialoglycoproteins; D018821:CD18 Antigens; D060748:Fetuins; D034741:RNA, Small Interfering; C017029:asialofetuin