Potential Transcriptional Biomarkers in Human Transplant Renal Biopsies ofEarly Stage Differentiated Differences between Living and Cadaveric Donors
Running title:Transcriptomics in Human Transplant Renal Biopsies
Introduction: The outcome of human kidney transplantation is significantly different between living donor (LD) and cadaveric donor (CAD). Transcriptomic signatures and signaling pathways in transplant renal biopsies may facilitate disclosing underlying mechanisms and define potential biomarkers thatsubsequently benefitdiagnosis and treatment of allograft injury.
Methods: Renal biopsies from LD and CAD paired at 30 minutes and 3 months post-transplantation (n=5-6) were used for whole-genome profiling and pathway analysis. Additional 33 biopsies (n=6-13) unpaired at two time points were used for validating differentially expressed genes, and identifying potential biomarkers by quantitative RT-PCR (qPCR). Differentially expressed genes were also further correlated with renal function and histology at late time points up to 24 months.
Results: The overall gene profiles were clearly different in 30-minute and 3-month biopsies regardless of donor type, although the cold ischemic time was significantly longer in CAD in contrast to LD. There were 446 differentially expressed genes (1.5-fold, p<0.05) between two donor types at 30 minutes, which was reduced to 149 at 3 months, with 25 genes in common. Many of these genes (CAD vs. LD) such as SERPINA3, FGA and SLP1 (27, 8 and 6-fold) linked to acute responses were up-regulated at 30 minutes and were eased off at 3 months. However, some other genes were remained higher such as SLP1, GSTM1 and VCAN (3, 3 and 2-fold), which involved in inflammation and nephrotoxicity, with lower CCND1 linked to proliferation. More interestingly, up-regulated tissue remodeling associated genes (3 months vs. 30 minutes) COL3A1, MMP9 and VCAN (9, 6 and 6-fold), and down-regulated FGA (10-fold), were only seen in CAD, with different up-regulated genes SERPINA3 (8-fold) in LD. The raised COL1A1 and COL1A2 (8-10-fold for both donor types) and reduced FOSB and FOS were seen in both CAD (74 and 39-fold) and LD (50 and 21-fold) at 3 months. The validation usingqPCR in additional 33 renal biopsies confirmed dysregulated genes SERPINA3, SLP1, VCAN, FOS and TIMP1, either between CAD and LD, or 3 months and 30 minutes. Furthermore, there were 120 genes closely correlated with serum creatinine and Sirius red staining at 12 months, among which 10 genes,GSTM1, COQ2, CCND1, CFB, FTCD, UNC5CL, SERPINA3, RAI14, TSPAN7 and SOD2, were further selected and validated by qPCR in the 33 biopsies. Up-regulated (CAD vs. LD) SERPINA3, FTCD, TASPN7 and CBF were confirmed in at 30 min; and up-regulated (3 months vs. 30 minutes) VCAN and TIMP1 in CAD with fell SERPINA 3 and FOS, while only FTCD was raised in CAD at both time points and in both sets of biopsies.
Conclusions: Transcriptomic signatures shifted from acute responses to tissue damage and remodeling post-transplantation with divergent profiles between LD and CAD, which mainly linked to initial donor ischemic injury and following immune responses. Potential candidate genes such as SERPINA3, FTCD, TASPN7, CBF, as well as SLP1, VCAN, TIMP1 and FOS, might be biomarkers for monitoring allograft injury and therapeutic response, and even as early targetsfor genetic modification to improve the long-term allograft survival.