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Abstract: SA-OR44

Spatial mRNA Analysis of Human Allograft Endothelium Shows Distinct Structure-Specific Endothelial Transcripts in Chronic Antibody-Mediated Rejection

Session Information

Category: Transplantation

  • 2002 Transplantation: Clinical

Authors

  • How, Ira Doressa Anne L., Massachusetts General Hospital, Boston, Massachusetts, United States
  • Tomaszewski, Kristen, Cleveland Clinic, Cleveland, Ohio, United States
  • Rios, Andrea, Massachusetts General Hospital, Boston, Massachusetts, United States
  • Morales, Samantha M., Massachusetts General Hospital, Boston, Massachusetts, United States
  • Divakar, Prajan, NanoString Technologies Inc, Seattle, Washington, United States
  • Smith, Rex Neal, Massachusetts General Hospital, Boston, Massachusetts, United States
  • Colvin, Robert B., Massachusetts General Hospital, Boston, Massachusetts, United States
  • Rosales, Ivy A., Massachusetts General Hospital, Boston, Massachusetts, United States
Background

Bulk tissue mRNA analysis has shown the importance of endothelium-associated transcripts, but structure-specific transcript differences remain unexplored. We used the NanoString GeoMx Digital Spatial Profiler (DSP) to spatially analyze transcripts in glomerular (Ge) and peritubular capillary (PTCe) endothelium in human renal allografts with and without T cell-mediated (TCMR) and chronic antibody-mediated (CAMR) rejection.

Methods

Five micron-thick formalin-fixed paraffin-embedded (FFPE) sections from 12 CAMR, 13 TCMR, 15 no rejection (NER), and 9 native (control) biopsies were hybridized with an 84-gene Immune Pathways and 10-gene custom panel attached to photocleavable oligonucleotide tags, and incubated with fluorophore-labeled antibodies (DNA, CD10, CD45, CD31). Using GeoMx DSP, Ge and PTCe areas of interest (AOIs, n=210) were segmented by CD31 expression. Oligonucleotide tags from AOIs were quantified using the NanoString nCounter MAX instrument. Data analysis was done using GeoMx DSP software.

Results

In controls, VEGFA, KDR and EHD3 are enriched in Ge while PLVAP is enriched in PTCe. Ge in CAMR with transplant glomerulopathy (TG) shows enrichment of CD59 (p=0.00005), PECAM1 (p=0.0005), COL4A1 (p=0.007), CD74 (p=0.006), HLA-DQ (p=0.02), CCND1 (p=0.005) and CXCL9 (p=0.03), while LAG3 (p=0.00005), EHD3 (p=0.003) and VEGFA (p=0.02) are decreased when compared to TCMR (Figure 1), indicating loss of VEGFA, endothelial dedifferentiation, and GBM remodelling with ectopic COL4A1. PTCe in CAMR showed IFNG-associated PTCe injury with increased IFNGR1 (p=0.001), PRF1 (p=0.005), ICAM1 (p=0.04), CD59 (p=0.02) and decreased PLVAP (p=0.001). STAT1 (p=0.05) and ITGB2 (p=0.005) are enriched in TCMR PTCe.

Conclusion

Spatial mRNA analysis identified distinct endothelial transcript expression that are potentially relevant to pathogenesis and therapeutic targets.

Funding

  • Other NIH Support