Cardiac Xenotransplantation Technology Provides Materials for
Improved Bioprosthetic Heart Valves
C.G.A. McGregor1,3,
N. Lila2, A, Carpentier2, M. Vlasin1, J.S. Logan1, G.W. Byrne1,3
1Department of Surgery, Mayo Clinic, Rochester, Minnesota, USA2Department of Cardiac Surgery, University of Paris, Paris, France3Department of Cardiothoracic Surgery, University College London, UK
Presenter Disclosure
Christopher G. A. McGregor, MB, FRCS, MD (Hons)The following relationships exist related to this presentation:
Authors McGregor and Logan are the inventors of technology related to xenotransplantation that has been licensed by the Mayo Clinic to a
commercial entity.
All other authors have no disclosure
• All humans and old world monkeys lack the xeno antigen Gal and develop preformed anti Gal antibody.
• Human anti Gal antibody is present in large quantities (1% of IgG and 1-4% of IgM) including in bioprosthetic heart valve (BHV) recipients.
• The Gal antigen is abundant with 25x106 epitopes per endothelial cell (MHC Class I is expressed on lymphocytes at a level 50 fold lower than Gal) and up to 10 11 epitopes per mg of tissue.
• All other species have the Gal antigen (a carbohydrate) and therefore do not develop anti Gal antibody.
The Gal Story - 1
GT-KO Pig Hearts are Devoid of -Gal Antigen
GT +/+
GT -/-
Nuclear transfer
Primary fibroblast culture
8 9 exon 9NeoDT
Targeting Vector
GTKO Pig Tissues negative for Gal
Kidney
Liver
Lung
Spleen
Skeletal muscle
Ligament
Tendon
Cornea
Bladder
Pericardium
Salivary
Thyroid
Aorta
Carotid
Femoral
Cecum
Duodenum
Esophagus
Rectum
Gall bladder
Thymus
•The major xeno antigen, Gal 1,3 Gal 1,4
GlcNAc ( -Gal) is present on commercial
bioprosthetic devices.Kasimir M-T, Rieder E, Seebacher G, et al, Tissue
Engineering, 11(7/8):1274-1280, 2005.
•Immune responses to -Gal are detected
in patients after implantation of BHVs.Konakci KZ, Bohle B, Blumer R, et al, Eur J Clin Invest;
35(1):17-23; 2005.
Evidence of Immunological Effects on Bioprostheses
•Glutaraldehyde fixed xenografts exhibit
greater inflammation and higher antibody
deposition compared to fresh or fixed
syngeneic grafts.Manji R, Nijjar N, Zhu L, et al, Proceedings of the Society of
Cardiothoracic Surgeons of Great Britain and Ireland;
Abstract 3, page 49, 2005.
•Higher levels of calcification found in bioprosthetic tissue exposed to graft-specific antibodies.”Zilla P, Brink J, Human P, and Bezuidenhout D. Biomaterials 29(4): 385-406; 2007
Evidence of Immunological Effects on Bioprostheses
GT
KO
W
T
Cardiac Xenotransplantation Technology Provides Materials for
Improved Bioprosthetic Heart Valves
H & E HRP-GSIB4 Gal comp.
Cardiac Xenotransplantation Technology Provides Materials for
Improved Bioprosthetic Heart Valves
Ligament TendonPericardium
Cardiac Xenotransplantation Technology Provides Materials for
Improved Bioprosthetic Heart Valves
This study tests whether binding of human anti-Gal
antibody effects calcification of wild type and GTKO
glutaraldehyde fixed porcine pericardium using a
standard rat implantation model.
Technologies for Improved Bioprosthetic Heart Valves (BHVs)
Rodent model
Gal +
Glut fixed
pericardium
Gal-KO Glut fixed
pericardium
+ anti-Gal no anti-Gal
+ anti-Gal no anti-Gal
Recovery of Pericardium 20 Days After Implantation
Pericardium prior to explant
8mm
Effect of Anti-Gal Antibodies on the Calcification of
Glutaraldehyde Fixed Porcine Pericardium
Cal
ciu
m c
on
ten
t (μ
g /
mg)
No Ab anti-Gal
Gal-KO
20
40
60
80
100
120
140
NS
NS
No Ab anti-Gal
Wild type
p = 0.005
p = 0.012
20
40
60
80
100
120
140
No antibody
anti-Gal antibody
Effect of Anti-Gal Antibodies on the Calcification of
Glutaraldehyde Fixed Porcine Pericardium
0
20
40
60
80
100
120
No Ab anti-Gal
0
20
40
60
80
100
120
No Ab anti-Gal
Calc
ium
(u
g/ m
g)
Wild typeGTKO
No antibody
anti-Gal antibody
NS
p < 0.05
NSp < 0.05
Glut/FET/Glut
+ anti-Gal Antibody
Gal +
GT-KO
Effect of Anti-Gal Antibodies on the Calcification of
Glutaraldehyde Fixed Detergent Extracted Porcine
Pericardium
Glut/FET/Glut
+ no antibody
Lila, N., McGregor, C., Carpentier, S., Rancic, J., Byrne, G., Carpentier, A.. Gal knockout pig
pericardium: New source of material for heart valve bioprostheses. Journal of Heart and Lung
Transplantation 2010.
• Glutaraldehyde fixed (GF) porcine pericardial tissues calcify in a rat model
• Anti-Gal antibodies significantly increases calcification in Gal positive but has no effect on GalKO GF pig tissue implanted in rats and rabbits.
• This study suggests that anti-Gal antibody may contribute to degeneration of BHVs.
• State of the art anti-calcification processing mitigates but does not eliminate the calcification-enhancing effects of anti-Gal antibody
• Tissue procured from GalKO pigs are resistant to the effects of anti-Gal antibodies and may become the preferred source for new potentially calcium resistant BHVs.
• If these findings are confirmed using functional GTKO BHVs, this would allow longer durability and wider application of BHVs.
Conclusions
The Gal Antigen Remains After
Glutaraldehyde Fixation
GSIB4 staining
GT+
GTKO
Human anti-Gal labeling
IgG
IgG
2nd only
2nd only
GT
+G
TK
O
• Glutaraldehyde fixed (GF) porcine pericardial tissues calcify in a rat model
• Anti-Gal antibodies significantly increase calcification in Gal positive but have no effect on GalKO GF tissue
• State of the art anti-calcification processing mitigates but does not eliminate the calcification-enhancing effects of anti-Gal antibody
• Tissue procured from GalKO pigs are resistant to the effects of anti-Gal antibodies
Conclusions
From Konakci et al 2005 Eur. J. Clin. Invest.
A
B
Cardiac Xenotransplantation Technology Provides Materials for
Improved Bioprosthetic Heart Valves
• Current commercially available BHVs of bovine or porcine origin from the major manufacturers retain the xeno antigen Gal after processing
• Preformed anti Gal antibody binds to BHVs in vivo
• Anti Gal antibody increases (is induced) after BHV implantation in patients
• The possible impact of the anti Gal antibody-Gal antigen interaction in structural valve deterioration has not been appreciated as all previously used preclinical models contain the Gal antigen and therefore lack anti Gal antibodies
The Gal Story - 2
Evidence of Immunological Effects from Bioprostheses
“We describe for the first time the presence of the -Gal epitope in clinically used porcine bioprostheses.”
Kasimir M-T, Rieder E, Seebacher G, et al, Tissue Engineering, 11(7/8):1274-1280, 2005.
“Our data suggest that implantation of bioprostheses in cardiac surgery induces a xenograft-specific immune response. Procedures diminishing the presence of -Gal on bioprostheses, such as utilization of genetically manipulated -Gal-deficient xenograft or pretreatment with -Galactosidase, might diminuate the immune response against bioprostheses and extend durability.”
Konakci KZ, Bohle B, Blumer R, et al, Eur J Clin Invest; 35(1):17-23; 2005.
“G-fixed xenograft group had significantly more valve and adventitial inflammation with more eosinophils as well as more anti-rat IgG deposition which may lead to xenograft rejection and valve destruction.”
Manji R, Nijjar N, Zhu L, et al, Proceedings of the Society of Cardiothoracic Surgeons of Great Britain and Ireland; Abstract 3, page 49, 2005.
“An almost three-times higher level of calcification was found in bioprosthetic tissue that was exposed to serum containing graft-specific antibodies.”
Zilla P, Brink J, Human P, and Bezuidenhout D. Biomaterials 29(4): 385-406; 2007
A
B
C
Bovine Porcine
Cardiac Xenotransplantation Technology Provides Materials for
Improved Bioprosthetic Heart Valves
Abstract
CARDIAC XENOTRANSPLANTATION TECHNOLOGY PROVIDES
MATERIALS FOR IMPROVED BIOPROSTHETIC HEART VALVES
Christopher GA McGregor, Nermine Lila, Michal Vlasin, John S Logan, Guerard
W. Byrne.
Department of Surgery, Mayo Clinic, Rochester, MN USA
Cardiac Surgery, University of Paris, Paris, France
Department of Cardiothoracic Surgery, University College London, UK
See Notes Page for Abstract text
