title: Immune mechanisms associated with the rejection of encapsulated neonatal porcine islet xenografts.
abstract: BACKGROUND
  The immune mechanisms associated with the rejection of microencapsulated neonatal porcine islets (NPI) are not clearly understood. Therefore, in this study we characterized the immune cells and molecules that are involved in this process by examining the microencapsulated NPI xenografts at various time points post-transplantation in B6 mice.
  
  
  METHODS
  Microencapsulated NPI were transplanted into streptozotocin-induced diabetic immune-competent B6 and immune-deficient B6 rag-/- mice and blood glucose levels were monitored twice a week. Encapsulated NPI were then recovered from B6 mice at various time points post-transplantation to characterize the islets and immune response using immunohistochemical and RT-PCR analyses. To determine which T-cell subpopulation is important for the rejection of encapsulated NPI, B6 rag-/- mice with established microencapsulated NPI xenografts were reconstituted with either CD4(+) or CD8(+) T cells and a return to the diabetic state was noted. For controls, adoptive transfer experiments involved reconstitution of B6 rag-/- mice with established microencapsulated NPI with non-fractionated lymph node cells or non-reconstituted mice.
  
  
  RESULTS
  All B6 recipients of microencapsulated NPI remained diabetic throughout the study while B6 rag-/- recipients achieved normoglycemia and maintained normoglycemia for up to 100 days post-transplantation. Encapsulated NPI recovered from B6 mice at early time points (day 7 and day 14) post-transplantation were surrounded with very few layers of immune cells that increased with time post-transplantation. The extent of cellular overgrowth on the surface of encapsulated NPI has a significant correlation with islet cell death and the presence of CD4(+) T cells, B cells and macrophages. Mouse IgG antibody and complement as well as cytokines [gamma-interferon (IFN-gamma), interleukin10 (IL10)] and chemokines (monocyte chemotactic protein-1 and macrophage inflammatory protein-1alpha and beta) were detected within the microcapsules at several time points post-transplantation suggesting that these molecules can traverse the microcapsule. Mouse anti-porcine IgG antibodies in recipient sera were found to peak at 30 days post-transplantation indicating leakage of porcine xenoantigens. In contrast, microencapsulated NPI recovered from B6 rag-/- mice had no cellular overgrowth on the surface. Complement and cytokines (IL 10 but not IFN-gamma) including chemokines were detected within the microcapsules at several days post-transplantation. We also found that B6 rag-/- mice reconstituted with non-fractionated lymph node cells or CD4(+) T cells but not CD8(+) T cells became diabetic demonstrating that CD4(+) T cells are the necessary T-cell subtype for microencapsulated NPI rejection. In contrast, non-reconstituted B6 rag-/- mice remained normoglycemic for the entire duration of the study.
  
  
  CONCLUSIONS
  Our results demonstrate that CD4(+) T cells, B cells and macrophages are the immune cells recruited to and involved in the rejection of encapsulated NPI. Immune molecules secreted by these cells as well as complement can traverse the microcapsule membrane and are responsible for destroying the NPI cells. Treatment regimens which target these molecules may modify the rejection of encapsulated NPI and lead to prolonged islet xenograft survival.
authors.affiliation: Department of Surgery, Dentistry Pharmacy Centre, Surgical-Medical Research Institute, University of Alberta, Edmonton, AB, Canada.
authors.forename: Tsunehiro
authors.initials: T
authors.lastname: Kobayashi
authors.forename: George
authors.initials: G
authors.lastname: Harb
authors.forename: Ray V
authors.initials: RV
authors.lastname: Rajotte
authors.forename: Gregory S
authors.initials: GS
authors.lastname: Korbutt
authors.forename: Aaron G
authors.initials: AG
authors.lastname: Mallett
authors.forename: Hossein
authors.initials: H
authors.lastname: Arefanian
authors.forename: Dereck
authors.initials: D
authors.lastname: Mok
authors.forename: Gina R
authors.initials: GR
authors.lastname: Rayat
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D001786:Blood Glucose / Q000378:metabolism; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D002460:Cell Line; D049109:Cell Proliferation; D016207:Cytokines / Q000235:genetics; D003920:Diabetes Mellitus / Q000276:immunology / Q000378:metabolism / Q000473:pathology / Q000601:surgery; D006084:Graft Rejection / Q000276:immunology*; D007113:Immunity, Innate / Q000276:immunology; D016381:Islets of Langerhans Transplantation; D008297:Male; D051379:Mice; D012333:RNA, Messenger / Q000235:genetics; D013552:Swine; D014183:Transplantation, Heterologous
chemical_list: D001786:Blood Glucose; D016207:Cytokines; D012333:RNA, Messenger