title: α-1-antitrypsin gene delivery reduces inflammation, increases T-regulatory cell population size and prevents islet allograft rejection.
abstract: Antiinflammatory clinical-grade, plasma-derived human α-1 antitrypsin (hAAT) protects islets from allorejection as well as from autoimmune destruction. hAAT also interferes with disease progression in experimental autoimmune encephalomyelitis (EAE) and in collagen-induced arthritis (CIA) mouse models. hAAT increases IL-1 receptor antagonist expression in human mononuclear cells and T-regulatory (Treg) cell population size in animal models. Clinical-grade hAAT contains plasma impurities, multiple hAAT isoforms and various states of inactive hAAT. We thus wished to establish islet-protective activities and effect on Treg cells of plasmid-derived circulating hAAT in whole animals. Islet function was assessed in mice that received allogeneic islet transplants after mice were given hydrodynamic tail-vein injection with pEF-hAAT, a previously described Epstein-Barr virus (EBV) plasmid construct containing the EBV nuclear antigen 1 (EBNA1) and the family of repeat EBNA1 binding site components (designated "EF") alongside the hAAT gene. Sera collected from hAAT-expressing mice were added to lipopolysaccharide (LPS)-stimulated macrophages to assess macrophage responsiveness. Also, maturation of peritoneal cells from hAAT-expressing mice was evaluated. hAAT-expressing mice accepted islet allografts (n = 11), whereas phosphate-buffered saline-injected animals (n = 11), as well as mice treated with truncated-hAAT-plasmid (n = 6) and untreated animals (n = 20) rapidly rejected islet allografts. In hAAT-expressing animals, local Treg cells were abundant at graft sites, and the IL-1 receptor antagonist was elevated in grafts and circulation. Sera from hAAT-expressing mice, but not control mice, inhibited macrophage responses. Finally, peritoneal cells from hAAT-expressing mice exhibited a semimature phenotype. We conclude that plasmid-derived circulating hAAT protects islet allografts from acute rejection, and human plasma impurities are unrelated to islet protection. Future studies may use this in vivo approach to examine the structure-function characteristics of the protective activities of AAT by manipulation of the hAAT plasmid.
authors.affiliation: Department of Clinical Biochemistry, Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, Israel.
authors.forename: Galit
authors.initials: G
authors.lastname: Shahaf
authors.forename: Hadas
authors.initials: H
authors.lastname: Moser
authors.forename: Eyal
authors.initials: E
authors.lastname: Ozeri
authors.forename: Mark
authors.initials: M
authors.lastname: Mizrahi
authors.forename: Avishag
authors.initials: A
authors.lastname: Abecassis
authors.forename: Eli C
authors.initials: EC
authors.lastname: Lewis
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods; D006084:Graft Rejection / Q000235:genetics / Q000378:metabolism* / Q000517:prevention & control; D006801:Humans; D007106:Immune Sera / Q000276:immunology / Q000494:pharmacology; D007249:Inflammation / Q000235:genetics / Q000378:metabolism* / Q000517:prevention & control; D016381:Islets of Langerhans Transplantation / Q000379:methods*; D008070:Lipopolysaccharides / Q000494:pharmacology; D018655:Lymphocyte Count; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D010957:Plasmids / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050378:T-Lymphocytes, Regulatory / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D014184:Transplantation, Homologous; D000515:alpha 1-Antitrypsin / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
chemical_list: D007106:Immune Sera; D008070:Lipopolysaccharides; D000515:alpha 1-Antitrypsin