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Presenter: Elena, Campello, Padua, Italy
Authors: Claudia Maria Radu1, Elena Campello1, Luca Spiezia1, Cristiana Bulato1, Sabrina Gavasso1, Lucrezia Furian2, Emanuele Cozzi2,3, Paolo Simioni1
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Circulating microparticles plasma levels in primates recipients of hDAF and GalT-KO pig kidney
Claudia Maria Radu1, Elena Campello1, Luca Spiezia1, Cristiana Bulato1, Sabrina Gavasso1, Lucrezia Furian2, Emanuele Cozzi2,3, Paolo Simioni1
1Cardiologic, Thoracic and Vascular Sciences, University of Padua; 2Surgical and Gastrointestinal Sciences, University of Padua; 3Consortium for Research in Organ Transplantation (CORIT); Padua, Italy
Introduction: Activation of the clotting cascade, fibrin deposition and thrombosis are key features of the rejection process that takes place in xenotransplantation. In order to prevent acute humoral xenograft rejection due to immunological barrier, human decay-accelerating factor (hDAF) and a1,3-galactosyltransferase gene knockout (GalT-KO) pigs are used, but all graft from these transgenic pigs underwent rejection exhibiting thrombotic microangiopathy with platelets-rich fibrin thrombi in the microvasculature. The exact mechanisms by which coagulation activation is induced after xenotransplantation remain unclear. Microparticles (MPs) are small vesicles shed from the surface of activated or apoptotic cells that have a procoagulant activity.
Aim of the study: In human, increased levels of circulating MP are associated with a prothrombotic state We measured and characterised circulating MPs plasma levels in monkeys before and after pig hDAF and GalT-KO kidney xenotransplantation.
Materials and Methods: Blood samples were obtained at time of euthanasia from 20 healthy monkeys, 10 after hDAF and 10 after GalT-KO kidney xenograft. MPs were analyzed by flow cytometry (FC500 Cytomics, USA), with a gate defined by 1µm beads (Megamix, Stago, France) and using FITC-Annexin V, PC5-anti-CD61, PE-anti-CD62P (P-Selectin), PE-antiCD62E (E-Selectin MP) and PE-anti-CD142 antibodies in order to identify apoptotic, platelets, endothelial and tissue factor bearing MPs, respectively.
Results: The total MPs plasma levels (mean±SD) and the subgroups of MPs in total study population are summarize in Fig 1.
Conclusion: The results demonstrateddecreased circulating MPs derived from apoptotic, activated endothelial cells and TF bearing MPs in the two groups of transplanted monkeys as compared to controls. In GalT-KO kidney xenograft animals the decreased of circulating MPs are higher than hDAF transplanted animals.
The reduction of circulating cells derived in GalT-KO as compared to hDAF kidney transplanted monkeys at euthanasia may be due to a major activation of clotting cascade causing consumption of the coagulation factors.
|
Apoptotic Mps (mean±SD) |
Platelets MPs (mean±SD) |
Endothelial MPs (mean±SD) |
TF-bearing MPs (mean±SD) |
GalT-KO (MPs/uL) |
19216± 9100* |
10053±6985** |
171±90** |
91±45* |
h-DAF (MPs/uL) |
13825±5077† |
7019±4026††† |
197±46†† |
86±33† |
Controls (MPs/uL) |
31224±8668 |
34753±10186 |
325±94 |
126±18 |
* p = ns ** p < 0.001 GalT-KO vs controls
† p = ns †† p < 0.05 ††† p < 0.001 h-DAF vs controls
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