Expression and functional analysis of glycosyl-phosphatidyl inositol-linked CD46 in transgenic mice.
BACKGROUND
Complement activation plays a pivotal role in hyperacute xenograft rejection. In humans, activation of complement is regulated by a number of cell surface regulatory proteins. Membrane cofactor protein (CD46) is one such regulator that protects cells by acting as a cofactor for the factor I-mediated cleavage of C3b and C4b. Transgenic animals expressing human CD46 may provide organs that are resistant to complement attack. However, attempts to generate mice expressing human CD46 using cDNA-based constructs have been largely unsuccessful.


METHODS
Transgenic mice expressing a glycosylphosphatidyl inositol (GPI)-linked form of CD46 were generated by microinjection of a hybrid CD46/CD55 cDNA under the control of the human intercellular adhesion molecule-2 promoter. Expression of CD46-GPI on the vascular endothelium was determined by immunohistochemistry. The ability of CD46-GPI to protect mouse tissues from human complement attack was determined using an ex vivo isolated perfused heart model.


RESULTS
Three founder animals expressing CD46-GPI were identified. Histological analysis showed strong and uniform expression of CD46-GPI on the vascular endothelium of all organs examined. Ex vivo perfusion of transgenic mouse hearts with human plasma showed a reduction in C3c deposition and a slightly prolonged function compared with controls.


CONCLUSIONS
High-level expression of CD46-GPI was achieved in transgenic mice by using a modified cDNA-based construct. The CD46-GPI was functional, providing some protection from complement-mediated damage in the ex vivo model, and may be useful in xenotransplantation if expressed in combination with CD55 and CD59.
Shinkel
T A
TA
Immunology Research Centre, St. Vincent's Hospital, Fitzroy, Victoria, Australia.
Cowan
P J
PJ
Barlow
H
H
Aminian
A
A
Romanella
M
M
Lublin
D M
DM
Pearse
M J
MJ
d'Apice
A J
AJ
D000818:Animals; D015703:Antigens, CD / Q000235:genetics* / Q000502:physiology*; D015815:Cell Adhesion Molecules / Q000235:genetics; D004396:Coloring Agents; D003169:Complement Inactivator Proteins / Q000235:genetics / Q000502:physiology; D004730:Endothelium, Vascular / Q000166:cytology; D016650:Fluorescein-5-isothiocyanate; D015870:Gene Expression; D017261:Glycosylphosphatidylinositols / Q000235:genetics* / Q000276:immunology / Q000502:physiology*; D006801:Humans; D051916:Membrane Cofactor Protein; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000502:physiology*; D051379:Mice; D008822:Mice, Transgenic; D011401:Promoter Regions, Genetic
D015703:Antigens, CD; C497857:CD46 protein, human; D015815:Cell Adhesion Molecules; D004396:Coloring Agents; D003169:Complement Inactivator Proteins; D017261:Glycosylphosphatidylinositols; C065289:ICAM2 protein, human; C497859:Mcp protein, mouse; D051916:Membrane Cofactor Protein; D008562:Membrane Glycoproteins; D016650:Fluorescein-5-isothiocyanate