Resistance to anti-xenogeneic response by combining alpha-Gal silencing with HO-1 upregulation.
BACKGROUND
A major barrier to clinical xenotransplantation is preformed xenoreactive natural antibodies (XNA) found in higher primates which react to Galalpha(1,3)Gal (alpha-Gal) epitopes found on lower species. Accommodation of organs to xenogeneic recipients involves upregulation of cytoprotective genes and resistance to complement dependent cytotoxicity (CDC).


METHODS
To develop methods of increasing these organ-protective effects, we established an in vitro CDC model utilizing human serum as the source of XNA and porcine endothelial cells (pEC) as targets.


RESULTS
Using this system we demonstrated that downregulation of alpha-Gal epitopes by siRNA silencing of alpha1,3-galactosyltransferase (alpha-GT) led to marginal protection from CDC while alpha-Gal silencing combined with Griffonia simplicifolia isolectin B4 (GS-IB4), a lectin that specifically binds to alpha-Gal epitopes, led to complete protection. Interestingly, alpha-Gal silencing and GS-IB4 mediated effects were not associated with inhibition of XNA binding to cells, but with significant decreased E-selectin expression and cytoprotective gene HO-1 upregulation. PI3K inhibitor LY294002 could block the elevation of HO-1 protein expression and reverse the protective effect of alpha-Gal silencing and GS-IB4 against CDC.


CONCLUSIONS
These data support the use of combination approaches targeting independent accommodation mechanisms to synergistically enhance donor organ survival in a xenogeneic setting.
Zhu
Min
M
Key Laboratory of Organ Transplantation, Ministry of Education and Health, Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Zhang
Wei
W
Liu
Fang
F
Wang
Lu
L
Liu
Bin
B
Chen
Dong
D
Zhu
Xue-Hai
XH
Zhang
Wei-Jie
WJ
Ichim
Thomas E
TE
Chen
Zhi-Shui
ZS
Zhou
Ping
P
Chen
Shi
S
Chen
Gang
G
D000818:Animals; D000910:Antibodies, Heterophile / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D000920:Antibody-Dependent Cell Cytotoxicity; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D004187:Disaccharides / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology*; D004338:Drug Combinations; D042783:Endothelial Cells / Q000276:immunology / Q000378:metabolism; D005700:Galactosyltransferases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology; D020868:Gene Silencing; D051547:Heme Oxygenase-1 / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D006801:Humans; D009025:Morpholines / Q000494:pharmacology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D037121:Plant Lectins / Q000494:pharmacology; D034741:RNA, Small Interfering; D015398:Signal Transduction; D013552:Swine; D014181:Transplantation Immunology; D014183:Transplantation, Heterologous / Q000276:immunology*
D000910:Antibodies, Heterophile; D002867:Chromones; D004187:Disaccharides; D004338:Drug Combinations; C045475:Griffonia simplicifolia lectins; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; D037121:Plant Lectins; D034741:RNA, Small Interfering; C055075:galactosyl-(1-3)galactose; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C496541:HMOX1 protein, human; D051547:Heme Oxygenase-1; D005700:Galactosyltransferases; C045097:UDP-galactose-lactosylceramide alpha 1-4-galactosyltransferase