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Activation of complement pathways in xenotransplantation: an in vitro study
Alexander J Walpen1, Paul Mohacsi, Caroline Frey
1Heart Transplantation Laboratory, Swiss Cardiovascular Center Bern, University Hospital.
Transplant Immunology
|August 16, 2002
Summary
Human antibodies against Galalpha1-3Gal (anti-Gal) trigger hyperacute rejection in pig-to-human xenotransplantation. This study shows anti-Gal antibodies activate both classical and alternative complement pathways, with the classical pathway being predominant.
Area of Science:
- Immunology
- Transplantation Biology
- Complement System
Background:
- Pig-to-human xenotransplantation is hindered by hyperacute graft rejection.
- This rejection is primarily mediated by human naturally occurring antibodies against the Galalpha1-3Gal (anti-Gal) epitope on pig endothelium.
- Antibody-mediated complement activation, typically via the classical pathway, is a key mechanism.
Purpose of the Study:
- To investigate whether the alternative complement pathway is directly activated by anti-Gal antibodies or if the classical pathway is essential.
- To compare the roles of the classical and alternative complement pathways in anti-Gal-mediated hemolysis.
- To evaluate the potential of targeting the classical pathway for xenotransplantation strategies.
Main Methods:
- Development of a novel rabbit erythrocyte (E) hemolysis assay (RECH50) to simultaneously assess both classical and alternative complement pathways.
- Utilized standard complement assays (CH50 and AP50) for comparison.
- Employed flow cytometry (FACS) to detect complement and antibody deposition on rabbit E.
- Performed serum depletion studies (anti-Gal, C1q, Factor D) and repletion experiments.
Main Results:
- Anti-Gal antibodies significantly contributed to rabbit E hemolysis in the RECH50 assay (65% reduction after depletion).
- Hemolysis was also reduced in the AP50 assay (35% reduction), indicating some alternative pathway involvement.
- The RECH50 assay showed an 8.4-fold increase in lysis compared to AP50, suggesting combined pathway activation.
- Classical pathway components were detected on rabbit E, particularly when the alternative pathway was compromised.
- Higher serum dilutions showed a predominant role for the classical pathway in anti-Gal-mediated lysis.
Conclusions:
- Anti-Gal antibodies induce xenograft lysis through both classical and alternative complement pathways, with the alternative pathway playing a minor role.
- The classical complement pathway is the primary driver of anti-Gal-mediated lysis, especially at higher serum dilutions.
- Targeting early steps of the classical pathway is a promising strategy for pig-to-primate xenotransplantation.
- Preserving the alternative pathway is beneficial as it is crucial for antibacterial defense.