title: Alginate microencapsulated hepatocytes optimised for transplantation in acute liver failure.
abstract: OBJECTIVE
  Intraperitoneal transplantation of alginate-microencapsulated human hepatocytes is an attractive option for the management of acute liver failure (ALF) providing short-term support to allow native liver regeneration. The main aim of this study was to establish an optimised protocol for production of alginate-encapsulated human hepatocytes and evaluate their suitability for clinical use.
  
  
  METHODS
  Human hepatocyte microbeads (HMBs) were prepared using sterile GMP grade materials. We determined physical stability, cell viability, and hepatocyte metabolic function of HMBs using different polymerisation times and cell densities. The immune activation of peripheral blood mononuclear cells (PBMCs) after co-culture with HMBs was studied. Rats with ALF induced by galactosamine were transplanted intraperitoneally with rat hepatocyte microbeads (RMBs) produced using a similar optimised protocol. Survival rate and biochemical profiles were determined. Retrieved microbeads were evaluated for morphology and functionality.
  
  
  RESULTS
  The optimised HMBs were of uniform size (583.5±3.3 µm) and mechanically stable using 15 min polymerisation time compared to 10 min and 20 min (p<0.001). 3D confocal microscopy images demonstrated that hepatocytes with similar cell viability were evenly distributed within HMBs. Cell density of 3.5×10(6) cells/ml provided the highest viability. HMBs incubated in human ascitic fluid showed better cell viability and function than controls. There was no significant activation of PBMCs co-cultured with empty or hepatocyte microbeads, compared to PBMCs alone. Intraperitoneal transplantation of RMBs was safe and significantly improved the severity of liver damage compared to control groups (empty microbeads and medium alone; p<0.01). Retrieved RMBs were intact and free of immune cell adherence and contained viable hepatocytes with preserved function.
  
  
  CONCLUSIONS
  An optimised protocol to produce GMP grade alginate-encapsulated human hepatocytes has been established. Transplantation of microbeads provided effective metabolic function in ALF. These high quality HMBs should be suitable for use in clinical transplantation.
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Suttiruk
authors.initials: S
authors.lastname: Jitraruch
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Anil
authors.initials: A
authors.lastname: Dhawan
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Robin D
authors.initials: RD
authors.lastname: Hughes
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Celine
authors.initials: C
authors.lastname: Filippi
authors.affiliation: British Heart Foundation Centre of Excellence Cardiovascular Division, King's College London School of Medicine, London, United Kingdom.
authors.forename: Daniel
authors.initials: D
authors.lastname: Soong
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Christina
authors.initials: C
authors.lastname: Philippeos
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Sharon C
authors.initials: SC
authors.lastname: Lehec
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Nigel D
authors.initials: ND
authors.lastname: Heaton
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Maria S
authors.initials: MS
authors.lastname: Longhi
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine, London, United Kingdom.
authors.forename: Ragai R
authors.initials: RR
authors.lastname: Mitry
mesh_terms: D000464:Alginates / Q000737:chemistry*; D000818:Animals; D002470:Cell Survival; D002478:Cells, Cultured; D018920:Coculture Techniques; D004195:Disease Models, Animal; D005688:Galactosamine / Q000009:adverse effects; D020723:Glucuronic Acid / Q000737:chemistry; D022781:Hepatocytes / Q000166:cytology* / Q000637:transplantation*; D006603:Hexuronic Acids / Q000737:chemistry; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000166:cytology*; D017114:Liver Failure, Acute / Q000139:chemically induced / Q000401:mortality / Q000628:therapy*; D008115:Liver Regeneration; D008297:Male; D008863:Microspheres; D051381:Rats; D017207:Rats, Sprague-Dawley; D014183:Transplantation, Heterologous
chemical_list: D000464:Alginates; D006603:Hexuronic Acids; D005688:Galactosamine; D020723:Glucuronic Acid