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Abstract: FR-OR129

Extracellular Vesicles Mediate Complement Activation and Tubular Senescence in Renal Antibody-Mediated Rejection

Session Information

Category: Transplantation

  • 1901 Transplantation: Basic

Authors

  • Castellano, Giuseppe, University of Bari, BARI, Italy
  • Franzin, Rossana, University of Bari, BARI, Italy
  • Divella, Chiara, University of Bari, Bari, Italy
  • Stasi, Alessandra, University of Bari, Bari, Italy
  • Sallustio, Fabio, University of Bari, Bari, Italy
Background

Renal Antibody-Mediated Rejection (AMR) is characterized by a strong complement activation that can lead to a premature senescence in tubular epithelial cells (TEC).
EVs (Extracellular vesicles), circulating microparticles able to mediate cell-to cell communication, are emerging as pivotal in different kidney diseases. The aim of this study was to investigate whether AMR derived-EVs could induce tubular inflammaging aging and complement activation.

Methods

Renal biopsies, serum and serum-isolated-EVs from 10 Acute and Chronic AMR patients were collected. TEC culture were incubated with EVs (5E+4EVs/cells target for 24h); then to assess cellular senescence qPCR for p21, p53, Klotho and CYP1B1 and SA-β-gal staining were performed. mRNA level of C3 and CFH were also measured. Inflammaging (p16INK4a and Klotho) markers were evaluated by IHC. Endothelial cells were grown in serum free media, incubated with AMR derived-EVs for 24h, then C4d IF was performed.

Results

Renal AMR biopsies showed significant tubular senescence as indicated by p16INK4a expression; p16INK4a was significantly upregulated in Chronic compared with Acute AMR biopsies (p<0.05). In vitro, the exposure of TEC to AMR serum induced senescence as observed by the upregulation of p21 and p53 gene levels (p<0.05). Furthermore, EVs exposed-TEC were characterized by significant increase in p21, p53 and CYP1B1 gene expression and down-regulation of Klotho (p<0.05) indicating that EVs can induce tubular senescence. In accordance, EVs induced a higher number of SA-β-gal+ TEC compared with control serum (p<0.05); the cells appeared larger and polynucleated indicating typical senescence phenotype. Finally, EVs from AMR patients induced a significant increase in C3 gene expression with concomitant downregualtion in CFH in TEC associated with C4d deposition on endothelial cells in serum free medium.

Conclusion

In AMR patients, circulating EVs induced accelerated inflammaging in TEC via dys-regulation of Complement system at cellular level. This new pathogenic process might help to identify new targets for therapeutic intervention .