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Abstract: SA-OR095

Single Cell RNA-Seq of Sequential Kidney Biopsies in Antibody Mediated Rejection Reveals a Highly Diverse Inflammatory Response

Session Information

Category: Transplantation

  • 1801 Transplantation: Basic

Authors

  • Malone, Andrew F., Washington University School of Medicine, Saint Louis, Missouri, United States
  • Ramakrishnan, Sai Mukund, Washington University School of Medicine, Saint Louis, Missouri, United States
  • Wu, Haojia, Washington University School of Medicine, Saint Louis, Missouri, United States
  • Donnelly, Erinn L., Washington University School of Medicine, Saint Louis, Missouri, United States
  • Humphreys, Benjamin D., Washington University School of Medicine, Saint Louis, Missouri, United States
Background

Antibody mediated rejection (AMR) remains a major cause of allograft failure yet current treatments are suboptimal reflecting our poor understanding of this disease process. To better understand the impact of current AMR treatments we performed single cell RNA-seq on human kidney biopsies both at AMR diagnosis and at 1 month post treatment.

Methods

Single cell suspensions were prepared by enzymatic digestion. The 10X Genomics platform was used for library preparation. Libraries were sequenced to a depth of ~50k reads/cell. Gene-cell matrices were obtained using CellRanger and the downstream analyses were done using R and Seurat.

Results

11,592 cells (1,841 genes/cell) and 5,581 cells (1,253 genes/cell) from the pre- and post-treatment biopsy, respectively, were included in the final analysis of each biopsy. AMR treatment included rituximab, bortezomib, IVIG, high dose steroids and tocilizumab. Allograft function did not return to baseline and post treatment histology showed no improvement in Banff scores. We detected all major epithelial cell types. A combined analysis of all immune cells from both biopsies revealed 16 separate immune cell clusters (Fig A). These included distinct monocyte(x8), dendritic cell(x3), CD8 T cell, NK cell, B cell and plasma cell clusters. Treatment resulted in a modest reduction in B cell and plasma cell proportions of 0.56 to 0.44 and 0.6 to 0.4, respectively (Fig B; red, pre-treatment cells; blue, post-treatment cells). By contrast, the number of these cells expressing TNFRSF17 (BMCA, B cell maturation antigen) increased from 1.2% to 24.9%, before and after treatment. Furthermore, the number of TNFSF13B (BAFF, B cell activating factor) expressing cells remained high after treatment (42% to 32%).

Conclusion

Aggressive anti-B cell and anti-plasma cell therapy failed to substantially reduce intra-graft antibody secreting cell numbers. Elevated intra-graft antibody secreting cell number may be maintained by BAFF-BCMA signaling. scRNA-seq provides a quantitative description of the cellular landscape in rejection.

Funding

  • NIDDK Support