Fetal porcine thymus engraftment, survival and CD4 reconstitution in alphaGal-KO mice is impaired in the presence of high levels of antibodies against alphaGal.
Xenospecific T-cell tolerance can be induced among murine and human T-cells by porcine thymic grafting. However, anti-alpha 1,3-galactosyltranserase (alphaGal) (Galalpha1-3Galbeta1-4GlcNAc-R) natural antibodies (NAbs) pose a major barrier to porcine xenografts in humans. We used alphaGal knockout (KO) and muchain KO mice to explore the effect of natural anti-alphaGal and other xenoantibodies on porcine thymic engraftment and to examine the potential of thymic tissue to tolerize anti-alphaGal antibody-producing cells. Thymectomized [adult thymectomy (ATX)] non-immunized and rabbit red blood cell (RRBC) pre-transplant immunized alphaGal-KO (knockout), wild-type (WT) and mu chain KO B6 mice were treated with 3Gy total body irradiation (TBI), and T and natural killer (NK) cell depleting monoclonal antibodies (mAbs). These conditioned mice were grafted with fetal porcine thymus and liver (FP THY/LIV) tissue under the kidney capsule. Flow cytometric analysis was performed to follow CD4 reconstitution as a measure of FP THY engraftment and function. Only mice with >10% CD4+ peripheral blood lymphocytes (PBL) were considered successfully engrafted. Enzyme-linked immunosorbent assay (ELISA) was used to assess the kinetics of immunoglobulin M (IgM) and IgG anti-alphaGal antibodies. Anti-pig antibodies were monitored by flow cytometry (FCM). FP THY engrafted successfully in most of the immunoglobulin deficient mice (11 out of 12, 92%) and the outcome was similar in WT B6 controls (8 out of 12, 67%). Non-immunized alphaGal-KO mice grafted with FP THY had a similar success rate (7 out of 11) to that observed in non-immunized alphaGal-WT controls (2 out of 4). In contrast, alphaGal-KO mice immunized pre-transplant with RRBC, then grafted with FP THY/LIV, showed a significant reduction in the success of thymic grafting (2 out of 9, 22%) compared with pre-transplant immunized WT controls (4 out of 7; 57%) and non-immunized alphaGal-KO mice (7 out of 11, 64%). Anti-Gal and anti-pig antibody levels were not markedly augmented by porcine thymus grafts in mice with successful thymus grafts. FP THY engraftment is impaired in the presence of high levels of anti-alphaGal xenoantibodies. However, low levels of anti-alphaGal antibodies and other mouse anti-pig NAbs appear not to play a major role in the rejection of FP THY. Although grafting FP THY expressing the alphaGal epitope did not tolerize B cells producing anti-alphaGal antibodies in a T-cell independent manner, it prevented T-cell dependent sensitization by inducing T-cell tolerance to porcine antigens.
Rodriguez-Barbosa
Jose-Ignacio
JI
Transplantation Biology Research Center, Bone Marrow Transplantation Section, Surgical Service, Massachusetts General Hospital/Harvard Medical School, Boston, MA 02129, USA.
Zhao
Yong
Y
Houser
Stuart
S
Zhao
Guiling
G
Sykes
Megan
M
D000818:Animals; D000910:Antibodies, Heterophile / Q000097:blood; D015478:Antigens, Heterophile / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D005260:Female; D016332:Fetal Tissue Transplantation; D005700:Galactosyltransferases / Q000235:genetics* / Q000276:immunology; D006085:Graft Survival / Q000276:immunology*; D007074:Immunoglobulin G / Q000097:blood; D007075:Immunoglobulin M / Q000097:blood; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D018345:Mice, Knockout; D011247:Pregnancy; D013552:Swine; D013950:Thymus Gland / Q000276:immunology / Q000637:transplantation*; D014312:Trisaccharides / Q000276:immunology*
D000910:Antibodies, Heterophile; D015478:Antigens, Heterophile; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D014312:Trisaccharides; C076569:alpha-galactosyl epitope; D005700:Galactosyltransferases; C047006:N-acetyllactosaminide alpha-1,3-galactosyltransferase