Immunological risk stratification and tailored minimisation of immunosuppression in renal transplant recipients.

BACKGROUND: The efficacy and safety of minimisation of immunosuppression including early steroid withdrawal in kidney transplant recipients treated with Basiliximab induction remains unclear. METHODS: This retrospective cohort study reports the outcomes from 298 consecutive renal transplants performed since 1st July 2010-June 2013 treated with Basiliximab induction and early steroid withdrawal in low immunological risk patients using a simple immunological risk stratification and 3-month protocol biopsy to optimise therapy. The cohort comprised 225 low-risk patients (first transplant or HLA antibody calculated reaction frequency (CRF ≤50% with no donor specific HLA antibodies) who underwent basiliximab induction, steroid withdrawal on day 7 and maintenance with tacrolimus and mycophenolate mofetil (MMF), and 73 high-risk patients who received tacrolimus, MMF and prednisolone for the first 3 months followed by long term maintenance immunosuppression with tacrolimus and prednisolone. High-risk patients not undergoing 3-month protocol biopsy were continued on triple immunosuppression. RESULTS: Steroid withdrawal could be safely achieved in low immunological risk recipients with IL2 receptor antibody induction. The incidence of biopsy-proven acute rejection was 15.1% in the low-risk and 13.9% in the high-risk group (including sub-clinical rejection detected at protocol biopsy). One- year graft survival was 93.3% and patient survival 98.5% in the low-risk group, and 97.3 and 100% respectively in the high-risk group. Graft function was similar in each group at 1 year (mean eGFR 61.2 ± 23.4 mL/min low-risk and 64.6 ± 19.2 mL/min high-risk). CONCLUSIONS: Immunosuppression regimen comprising basiliximab induction, tacrolimus, MMF and prednisolone with early steroid withdrawal in low risk patients and MMF withdrawal in high risk patients following a normal 3-month protocol biopsy is effective in limiting acute rejection episodes and produces excellent rates of patient survival, graft function and complications.

JunD/AP1 regulatory network analysis during macrophage activation in a rat model of crescentic glomerulonephritis.

BACKGROUND: Function and efficiency of a transcription factor (TF) are often modulated by interactions with other proteins or TFs to achieve finely tuned regulation of target genes. However, complex TF interactions are often not taken into account to identify functionally active TF-targets and characterize their regulatory network. Here, we have developed a computational framework for integrated analysis of genome-wide ChIP-seq and gene expression data to identify the functional interacting partners of a TF and characterize the TF-driven regulatory network. We have applied this methodology in a rat model of macrophage dependent crescentic glomerulonephritis (Crgn) where we have previously identified JunD as a TF gene responsible for enhanced macrophage activation associated with susceptibility to Crgn in the Wistar-Kyoto (WKY) strain. RESULTS: To evaluate the regulatory effects of JunD on its target genes, we analysed data from two rat strains (WKY and WKY.LCrgn2) that show 20-fold difference in their JunD expression in macrophages. We identified 36 TFs interacting with JunD/Jun and JunD/ATF complexes (i.e., AP1 complex), which resulted in strain-dependent gene expression regulation of 1,274 target genes in macrophages. After lipopolysaccharide (LPS) stimulation we found that 2.4 fold more JunD/ATF-target genes were up-regulated as compared with JunD/Jun-target genes. The enriched 314 genes up-regulated by AP1 complex during LPS stimulation were most significantly enriched for immune response (P = 6.9 × 10⁻4) and antigen processing and presentation functions (P = 2.4 × 10⁻5), suggesting a role for these genes in macrophage LPS-stimulated activation driven by JunD interaction with Jun/ATF. CONCLUSIONS: In summary, our integrated analyses revealed a large network of TFs interacting with JunD and their regulated targets. Our data also suggest a previously unappreciated contribution of the ATF complex to JunD-mediated mechanisms of macrophage activation in a rat model of crescentic glomerulonephritis.

Combined ChIP-Seq and transcriptome analysis identifies AP-1/JunD as a primary regulator of oxidative stress and IL-1ß synthesis in macrophages.

BACKGROUND: The oxidative burst is one of the major antimicrobial mechanisms adopted by macrophages. The WKY rat strain is uniquely susceptible to experimentally induced macrophage-dependent crescentic glomerulonephritis (Crgn). We previously identified the AP-1 transcription factor JunD as a determinant of macrophage activation in WKY bone marrow-derived macrophages (BMDMs). JunD is over-expressed in WKY BMDMs and its silencing reduces Fc receptor-mediated oxidative burst in these cells. RESULTS: Here we combined Jund RNA interference with microarray analyses alongside ChIP-sequencing (ChIP-Seq) analyses in WKY BMDMs to investigate JunD-mediated control of macrophage activation in basal and lipopolysaccharide (LPS) stimulated cells. Microarray analysis following Jund silencing showed that Jund activates and represses gene expression with marked differential expression (>3 fold) for genes linked with oxidative stress and IL-1ß expression. These results were complemented by comparing whole genome expression in WKY BMDMs with Jund congenic strain (WKY.LCrgn2) BMDMs which express lower levels of JunD. ChIP-Seq analyses demonstrated that the increased expression of JunD resulted in an increased number of binding events in WKY BMDMs compared to WKY.LCrgn2 BMDMs. Combined ChIP-Seq and microarray analysis revealed a set of primary JunD-targets through which JunD exerts its effect on oxidative stress and IL-1ß synthesis in basal and LPS-stimulated macrophages. CONCLUSIONS: These findings demonstrate how genetically determined levels of a transcription factor affect its binding sites in primary cells and identify JunD as a key regulator of oxidative stress and IL-1ß synthesis in primary macrophages, which may play a role in susceptibility to Crgn.