Ballistic transfer of minimalistic immunologically defined expression constructs for IL4 and CTLA4 into the corneal epithelium in mice after orthotopic corneal allograft transplantation.
BACKGROUND
Experiments were performed to determine whether corneal epithelium transfected with minimalistic immunologically defined expression constructs for the extracellular fragment of CTLA4 and for interleukin-4 (IL-4) or interleukin-10 (IL-10) is able to modulate an allospecific immune response after orthotopic corneal grafting in mice.


METHODS
Six groups of BALB/c (H-2d) mice received a C3H (H-2k) corneal graft and dexamethasone eye drops until day 11. Five groups of BALB/c mice had gold particles delivered into the corneal epithelium by Gene Gun on day 10 after transplantation. In four groups, minimalistic immunologically defined gene expression (MIDGE) vectors were delivered into the corneal epithelium by ballistic transfer. The levels of expressed IL-4 and IL-10 were determined by an enzyme-linked immunosorbent assay (ELISA) in shock-frozen homogenized corneas. The expression kinetics of Gene-Gun-transfected corneas were determined by measuring luciferase in lysed whole corneas at different time intervals.


RESULTS
Luciferase expression was detectable during the first 5 days following transfection. ELISA was used to determine IL-4 and IL-10 expression in corneal tissue 36 h after transfection. Ballistic IL-4 and CTLA4 gene transfer significantly prolonged corneal graft survival in comparison with the gold-treated control group and the IL-10-treated group.


CONCLUSIONS
The beneficial effect of IL-4 and CTLA4, but not IL-10 gene transfer into the corneal epithelium by MIDGE vectors was demonstrated for the first time in corneal transplantation.
König Merediz
S A
SA
Freie Universität Berlin, Institut für Molekularbiologie und Bioinformatik, Fachbereich Humanmedizin, Germany.
Zhang
E P
EP
Wittig
B
B
Hoffmann
F
F
D000069594:Abatacept; D000818:Animals; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000096:biosynthesis / Q000235:genetics*; D060908:CTLA-4 Antigen; D002460:Cell Line; D016039:Corneal Transplantation / Q000502:physiology*; D003587:Cytomegalovirus / Q000235:genetics; D004247:DNA / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism; D019573:Epithelium, Corneal / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D005822:Genetic Vectors; D018796:Immunoconjugates; D015847:Interleukin-4 / Q000096:biosynthesis / Q000235:genetics*; D008264:Macrophages / Q000166:cytology; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D011993:Recombinant Fusion Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection / Q000379:methods*; D014184:Transplantation, Homologous
D015703:Antigens, CD; D000943:Antigens, Differentiation; D060908:CTLA-4 Antigen; C556707:Ctla4 protein, mouse; D018796:Immunoconjugates; D011993:Recombinant Fusion Proteins; D015847:Interleukin-4; D004815:Epidermal Growth Factor; D000069594:Abatacept; D004247:DNA