Introduction: Cellular senescence, a process during which senescent cells accumulate and secrete an inflammatory senescence-associated secretory phenotype (SASP) factors shapes organ quality and function.  We hypothesized that accelerated senescence is critical in advancing chronic allograft deterioration. 

Methods: Fully MHC mismatched heterotopic heart transplantation was performed from BALB/c to C57BL/6 mice; recipients were treated with CTLA4-Ig (days 1 and 2); RT-PCR and IHC were performed to assess the accumulation of senescent cells and related SASP factors (p16INK4a, p21Cip/Waf, IL-6, TNFα, senescence-associated beta-galactosidase activity (SA-β-Gal); histology analysis and FACS monitored chronic changes and profiled alloimmune responses. To test the impact of senescent cells on chronic allograft deterioration, recipients were treated with senolytics (Dasatinib 5mg/kg and Quercetin 50mg/kg; p.o. biweekly).

Results: Senescent cells and SASP-related inflammatory cytokines accumulated in significantly elevated amounts in chronically rejected cardiac allografts (p21Cip/Waf, p<0.01; p16INK4a, and SA-β-Gal positive cells p<0.05, p16INK4a/IL-6 and p16INK4a/TNFα double-positive cells, p<0.05 compared to naïve heart).  Treatment with senolytics depleted senescent cells and reduced amounts of SASP factors; morphologically, we observed a significant improvement of vascular neointimal hyperplasia (p<0.0001) while the advancement of fibrosis stalled (p<0.05). Profiling alloimmune responses subsequent to the treatment with senolytics revealed increased frequencies of Treg (p<0.01), decreased Th1 (p<0.01) and Th17 (p<0.01) systemically; intragraft, CD4⁺ T cells (p<0.01), CD8⁺ T cells (p<0.05), neutrophils (p<0.05), and macrophages (p<0.05) were reduced; treatment with senolytics also prolonged graft survival significantly (MST=34.5 days vs. 49.0, p<0.05). Mechanistically, senolytics impeded the phosphorylation of STAT3 and NFkB, key inflammatory transcription factors, suggesting accelerated senescence as a driver of chronic allograft dysfunction.

Conclusions: Accelerated aging is playing a key role in the advancement of chronic allograft dysfunction. Depleting senescent cells improves morphological changes and prolongs graft survival. Targeting senescent cells may provide a novel treatment targeting chronic allograft deterioration.