Pre-transplant Transcriptional Signature in Peripheral Blood Mononuclear Cells of Acute Renal Allograft Rejection

Acute rejection (AR) is closely associated with renal allograft dysfunction. Here, we utilised RNA sequencing (RNA-Seq) and bioinformatic methods to characterise the peripheral blood mononuclear cells (PBMCs) of patients with acute renal allograft rejection. Pretransplant blood samples were collected from 32 kidney allograft donors and 42 corresponding recipients with biopsies classified as T cell-mediated rejection (TCMR, n = 18), antibody-mediated rejection (ABMR, n = 5), and normal/non-specific changes (non-AR, n = 19). The patients with TCMR and ABMR were assigned to the AR group, and the patients with normal/non-specific changes (n = 19) were assigned to the non-AR group. We analysed RNA-Seq data for identifying differentially expressed genes (DEGs), and then gene ontology (GO) analysis, Reactome, and ingenuity pathway analysis (IPA), protein—protein interaction (PPI) network, and cell-type enrichment analysis were utilised for bioinformatics analysis. We identified DEGs in the PBMCs of the non-AR group when compared with the AR, ABMR, and TCMR groups. Pathway and GO analysis showed significant inflammatory responses, complement activation, interleukin-10 (IL-10) signalling pathways, classical antibody-mediated complement activation pathways, etc., which were significantly enriched in the DEGs. PPI analysis showed that IL-10, VEGFA, CXCL8, MMP9, and several histone-related genes were the hub genes with the highest degree scores. Moreover, IPA analysis showed that several proinflammatory pathways were upregulated, whereas antiinflammatory pathways were downregulated. The combination of NFSF14+TANK+ANKRD 33 B +HSPA1B was able to discriminate between AR and non-AR with an AUC of 92.3% (95% CI 82.8–100). Characterisation of PBMCs by RNA-Seq and bioinformatics analysis demonstrated gene signatures and biological pathways associated with AR. Our study may provide the foundation for the discovery of biomarkers and an in-depth understanding of acute renal allograft rejection.

P1678ASSOCIATION OF EPIDERMAL GROWTH FACTOR(EGF) AND EGF RECEPTOR POLYMORPHISMS WITH END STAGE RENAL DISEASE AND ACUTE RENAL ALLOGRAFT REJECTION IN KOREAN POPULATION

Background and Aims Epidermal growth factor (EGF) has been found to be associated with development and repair mechanisms of several renal diseases. In this study, we hypothesized that single nucleotide polymorphisms (SNPs) of the EGF or its receptor genes might have association with end stage renal disease (ESRD) or acute renal allograft rejection (AR) in Korean patients. Method 347 recipients of the first renal transplants for ESRD including 63 AR patients among them and 289 healthy adults were included in the study. Five EGF gene SNPs (rs11568835, rs11568943, rs2237051, rs11569017, rs3756261) and four EGFR gene SNPs (rs1140475, rs2293347, rs1050171, rs6965469) were analyzed. The genotypes of these SNPs were analyzed using AxiomTM genome-wide human assay. For statistical analysis we used SNPStats and Haploview version 4.2. Multiple logistic regression models (codominant, dominant, recessive and Log-additive) were produced to obtain odds ratio (OR), 95% confidence interval (CI), and p value. Results One SNP (rs11569017) in EGF gene showed significant association with ESRD but not with AR. Another SNP (rs11568835) in EGF gene showed significant association with susceptibility to AR but not with ESRD. One SNP (rs1050171) in EGFR gene showed significant association with susceptibility to AR but not with ESRD. Conclusion We suggest that genetic polymorphisms of EGF and EGFR gene may be associated with the risk of development of ESRD and AR in the Korean population.

The Use of Donor-Derived Cell-Free DNA for Assessment of Allograft Rejection and Injury Status

Patient monitoring after kidney transplantation (KT) for early detection of allograft rejection remains key in preventing allograft loss. Serum creatinine has poor predictive value to detect ongoing active rejection as its increase is not sensitive, nor specific for acute renal allograft rejection. Diagnosis of acute rejection requires allograft biopsy and histological assessment, which can be logistically challenging in some cases and carries inherent risk for complications related to procedure. Donor-derived cell-free DNA (dd-cfDNA), DNA of donor origin in the blood of KT recipient arising from cells undergoing injury and death, has been examined as a potential surrogate marker for allograft rejection. A rise in dd-cfDNA levels precedes changes in serum creatinine allows early detections and use as a screening tool for allograft rejection. In addition, when used in conjunction with donor-specific antibodies (DSA), it increases the pre-biopsy probability of antibody-mediated rejection (ABMR) aiding the decision-making process. Advancements in noninvasive biomarker assays such as dd-cfDNA may offer the opportunity to improve and expand the spectrum of available diagnostic tools to monitor and detect risk for rejection and positively impact outcomes for KT recipients. In this this article, we discussed the evolution of dd-cfDNA assays and recent evidence of assessment of allograft rejection and injury status of KT by the use of dd-cfDNA.

Renal epithelial cells retain primary cilia during human acute renal allograft rejection injury

Objectives Primary cilia are sensory organelles which co-ordinate several developmental/repair pathways including hedgehog signalling. Studies of human renal allografts suffering acute tubular necrosis have shown that length of primary cilia borne by epithelial cells doubles throughout the nephron and collecting duct, and then normalises as renal function returns. Conversely the loss of primary cilia has been reported in chronic allograft rejection and linked to defective hedgehog signalling. We investigated the fate of primary cilia in renal allografts suffering acute rejection. Results Here we observed that in renal allografts undergoing acute rejection, primary cilia were retained, with their length increasing 1 week after transplantation and remaining elevated. We used a mouse model of acute renal injury to demonstrate that elongated renal primary cilia in the injured renal tubule show evidence of smoothened accumulation, a biomarker for activation of hedgehog signalling. We conclude that primary cilium-mediated activation of hedgehog signalling is still possible during the acute phase of renal allograft rejection.