Phosphorylated S6 kinase and S6 ribosomal protein are diagnostic markers of antibody-mediated rejection in heart allografts

Background: Anti-MHC Class I alloantibodies have been implicated in the processes of acute and chronic rejection. These antibodies (Ab) bind to endothelial cells (EC) and transduce signals leading to the activation of cell survival and proliferation pathways, including Src, FAK and mTOR, as well as downstream targets ERK, S6 kinase (S6K) and S6 ribosomal protein (S6RP). We tested the hypothesis that phosphorylation of S6K, S6RP and ERK in capillary endothelium may serve as an adjunct diagnostic tool for antibody-mediated rejection (AMR) in heart allografts.

Methods: Diagnosis of AMR was based on histology or immunoperoxidase staining of paraffin-embedded tissue, consistent with 2013 ISHLT criteria. Diagnosis of acute cellular rejection (ACR) was based on ISHLT criteria. Endomyocardial biopsies from 67 heart transplant recipients diagnosed with acute rejection [33 with pAMR, 18 with ACR (15 with Grade 1R, 3 with Grade ≥2R), 16 with pAMR and ACR (13 with 1R and 3 with ≥2R)] and 40 age- and gender-matched recipients without rejection were tested for the presence of phosphorylated forms of ERK, S6RP and S6K by immunohistochemistry.

Results: Immunostaining of endomyocardial biopsies with evidence of pAMR showed a significant increase in expression of p-S6K and p-S6RP in capillary EC compared with controls. A weaker association was observed between pAMR and p-ERK.

Conclusions: Biopsies diagnosed with pAMR often showed phosphorylation of S6K and S6RP, indicating that staining for p-S6K and p-S6RP is useful for the diagnosis of AMR. Our findings support a role for antibody-mediated HLA signaling in the process of graft injury.