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Oral Communications 6
13.3 - In Vitro Immunogenicity of Alginate Microencapsulated Human Hepatocytes
Presenter: Suttiruk, Jitraruch, London, United Kingdom Authors: Suttiruk Jitraruch1, Anil Dhawan1, Maria S. Longhi1, Robin D. Hughes1, Celine Filippi1, Sharon C. Lehec1, Fabiane B. Klem1, Ragai R. Mitry1
In Vitro Immunogenicity of Alginate Microencapsulated Human Hepatocytes
Suttiruk Jitraruch1, Anil Dhawan1, Maria S. Longhi1, Robin D. Hughes1, Celine Filippi1, Sharon C. Lehec1, Fabiane B. Klem1, Ragai R. Mitry1
1Institute of Liver Studies, King's College School of Medicine, London, United Kingdom
Background:
Transplantation of alginate microencapsulated human hepatocytes without the use of immunosuppession is an attractive approach for the management of acute liver failure. The biocompatibility of the alginate microbeads is an important issue for efficacy following transplantation into the peritoneum. The microbeads themselves or antigen shedding through microcapsule pores, may initiate a host immune response from the encapsulated cells and subsequently lead to an inflammatory reaction and cell death.
Aim:
To investigate the alloimmune response toward clinical grade microencapsulated human hepatocyte in vitro.
Materials & Methods:
Microbeads were produced using an encapsulator (250µm nozzle) with sterile grade, highly purified sodium alginate (PRONOVA™ SLG20: low viscosity, high guluronic acid). Empty and human hepatocyte (3.5x106cells/ml alginate; n=4) microbeads were polymerised in 1.2% calcium chloride. Peripheral blood mononuclear cells (PBMCs) were obtained from four healthy adults. PBMCs were either cultured alone (monoculture) or co-cultured with empty or hepatocyte alginate microbeads. The ratio of PBMCs to hepatocytes was 1:5. Microbead morphology was examined before and after co-culture with PBMCs. The frequency of activation of T-lymphocyte, B cells, NK cells and monocytes was analysed 24h post co-culture by flow cytometry using antibodies to CD3, CD4, CD8, CD25, CD14, CD40L, CD38, CD56, CD19, and CD54.
Results:
Empty and hepatocyte microbeads were of uniform shape and size (diameter: 500±100μm). After 24hr in co-culture with PBMCs, both empty and hepatocyte microbeads were intact and maintained their uniform shape with no PBMCs adherent to their surface. There was no significant difference in the level of activation markers expression on PBMCs co-cultured with empty or hepatocyte microbeads compared to PBMCs in monoculture. Interestingly, human hepatocyte microbeads decreased the frequency of CD14+CD25+ activated monocyte on PBMCs (mean 2.66 ± SEM 0.80) in co-cultures compared to PBMCs monocultures (45.65±10.98).
Conclusions:
This study provides evidence that clinical grade human hepatocyte microbead biocompatible with the human peripheral immune system in vitro.
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