Abstract: TH-PO022

Calcineurin Inhibitor-Induced Endothelial Cell Injury – A Role for Complement

Session Information

Category: Transplantation

  • 1701 Transplantation: Basic and Experimental

Authors

  • Teoh, Chia Wei, The Hospital for Sick Children, Toronto, Ontario, Canada
  • Riedl, Magdalena, The Hospital for Sick Children, Toronto, Ontario, Canada
  • Bruno, Valentina, None, Toronto, Ontario, Canada
  • Robinson, Lisa, The Hospital for Sick Children, Toronto, Ontario, Canada
  • Licht, Christoph, The Hospital for Sick Children, Toronto, Ontario, Canada
Background

Calcineurin inhibitors (CNI) are associated with nephrotoxicity, endothelial cell (EC) dysfunction and thrombotic microangiopathy (TMA). Evolving evidence suggests a central role for complement dysregulation in the pathogenesis of CNI-induced TMA. However, the exact mechanism of CNI-induced complement-mediated injury remains unknown. We hypothesize that CNIs induce complement-mediated EC injury and impair complement regulation on EC surfaces.

Methods

Complement activation (C3c, C9) and regulation (CD46, CD55, CD59), and complement factor H (CFH) surface binding were assessed by flow cytometry. CFH surface cofactor activity was assessed by a surface cofactor assay. EC cytotoxicity was detected via LDH assay. Blood outgrowth EC (BOECs) from healthy donors and an aHUS patient with MCP mutation were incubated with cyclosporine (CsA) and subsequently exposed to 50% normal human serum (NHS) as source for complement. EC glycocalyx was assessed with wheat germ agglutinin staining on confocal microscopy.

Results

The sequence of CsA incubation and 50% NHS resulted in a dose and time dependent enhancement of EC complement deposition and EC death, exacerbated by serum starvation and sensitization with anti-CD59 antibody. An optimal balance of EC survival and CNI effect was obtained with CsA 10 mcg/ml for 24 hours. CsA led to upregulation of CD46, CD55 and CD59 on EC surface. CsA diminished the EC glycocalyx with subsequent decreased CFH surface binding and surface cofactor activity. In further support of our in-vitro findings, MCP-deficient BOECs exposed to CsA had significantly higher surface complement deposition compared to healthy controls.

Conclusion

Our findings suggest a role for complement-mediated EC injury induced by CsA, with CFH surface dysregulation playing a key role in CsA-induced complement activation on EC surfaces. MCP-deficient BOECs were genetically predisposed to be more susceptible to CsA-induced endothelial complement deposition. CsA-induced abolishment of EC glycocalyx may be the key mechanism leading to alternative pathway dysregulation, and warrants further studies.