Potential Novel Proteomic Biomarkers for Acute T Cell-Mediated Rejection in Kidney Transplant Recipients

Introduction Acute rejection is hazardous to long-term graft survival in kidney transplant recipients (KTRs). However, the only method to confirm acute rejection is graft biopsy, which is invasive and costs a lot of money. According to recent progress in biotechnology, non-invasive diagnostic method has been studied to diagnose acute rejection. The aim of current study is to identify novel proteomic biomarkers for early diagnosis of acute T cell-mediated rejection (TCMR) in urinary exosomes for KTRs. Materials and Methods Biopsy-proven twenty two KTRs with stable graft function (STA) and twenty five KTRs with acute TCMR were included in this study. We performed proteomic analysis to find candidate biomarkers, which reflect early stage of allograft rejection, by using nano-ultra performance liquid chromatography-tandem mass spectrometry (nano-UPLC-MS/MS) on pooled urinary exosome in both KT groups. For validation, the protein levels of candidate biomarkers in each individual urinary exosome from KTRs were measured by Western blot assay. Results A total of 169 urinary exosome proteins, 138 in STA patients and 100 in acute TCMR patients, were identified by nano-UPLC-MS/MS method. 69 proteomes were common to both STA and acute TCMR, and were quantified by label-free LC-MS/MS. 34 proteins were increased in STA patients and 10 proteins were increased in acute TCMR patients. Among them, we selected 5 candidate proteins as candidate biomarkers for early diagnosis of acute TCMR according to significance, degree of quantity variance, and information from the ExoCarta database. We confirmed the proteomic levels of five candidate biomarkers by Western blot analysis in each patient, and consequently tetraspanin-1 (TSPAN1) and hemopexin (HPX) was significantly higher in acute TCMR patients (STA:acute TCMR ratio=1:1.8, P=0.009, and 1:3.5, P=0.046, respectively). Conclusion TSPAN1 and HPX, which could be potential diagnostic proteins for acute TCMR, were discovered in urine for KTRs. Follow-up studies are needed to demonstrate clinical effectiveness of TSAPN1 and HPX to predict acute TCMR in KTRs.