Abstract: SA-PO458

Enhanced Evaluation of Human Renal Biopsies Using Multicolor Flow Cytometry and Cytokine Analysis: A Focus on Transplanted Kidneys

Session Information

Category: Transplantation

  • 1702 Transplantation: Clinical and Translational

Authors

  • Muczynski, Kimberly A., University of Washington, Seattle, Washington, United States
  • Leca, Nicolae, University of Washington, Seattle, Washington, United States
  • Anderson, Susan K., University of Washington, Seattle, Washington, United States
Background

Virtually all renal disease is due to an immune response. Examination of renal biopsies with light, immunofluorescence and electron microscopy provide limited information about immune mechanisms causing kidney injury and disease activity. In adddition, information is often insufficient to direct therapy with specific agents, which becomes a frustration with new immune modulating agents being developed.

Methods

To enhance the information available from a biopsy we developed a technique for reducing a fraction of a renal biopsy to single cells for multicolor flow cytometry and for capture and quantitation of secreted cytokines present within the biopsy. As proof of concept, after evaluation of over 400 biopsies, we use our technique to suggest new criteria for evaluating rejection and renal inflammation that are clinically useful for directing therapy.

Results

A ratio of CD8+ to CD4+ lymphocytes of greater than 1.2 within the biopsy of transplanted kidneys is associated with rejection, even before it is apparent by microscopy. Elevated numbers of CD45 leukocytes and higher levels of IL-6, IL-8 and IL-10 within the kidney indicate more severe injury. Antibody binding to renal microvascular endothelial cells can be measured and corresponds to antibody-mediated forms of allograft rejection. Eculizumab binding to endothelial cells suggests complement activation, which may be independent of bound antibody. A comparison of intrarenal leukocyte subsets and their activation states to those of peripheral blood from the same donor at the time of biopsy identify significant differences supporting the need to develop techniques which interrogate the immune system within the kidney.

Conclusion

Our use of cytometry to assess intrarenal leukocyte subsets, microvascular endothelial cell properties and secreted cytokines from a fragment of a standard renal biopsy provide useful information about the immune processes in the kidney. After evaluation of over 400 biopsies we find results reliable indicators of disease activity. This information is not available from peripheral blood. While we have focused on biopsies from transplanted kidneys (due to their availability), our techniques are equally applicable to native kidneys. Cytometry can enhance renal biopsy evaluation in a clinically significant manner.

Funding

  • Private Foundation Support