MDM2 inhibitor ameliorates cisplatin-induced nephropathy via NFκΒ signal inhibition.

Cisplatin is a platinum-containing chemotherapeutic drug, which is widely used and highly effective. While effective against tumors, its use is limited by severe side effects such as nephrotoxicity and bone marrow suppression. Murine double minute 2 (MDM2) is the E3 ubiquitin ligase of the tumor suppressor gene, p53, and inhibition of MDM2 can suppress tumor cell growth. However, independent of p53, MDM2 acts as a co-transcription factor for nuclear factor-κB (NFκB), whose signaling can be involved in cisplatin-induced tubular injury. We therefore examined the effects of MDM2 inhibitor on cisplatin cytotoxicity. In order to induce acute kidney injury and to investigate MDM2 inhibitory effects, we injected cisplatin into rats with or without the MDM2 inhibitor, DS-5272, and analyzed kidney physiology/histology and NFκB signaling. Serum creatinine was significantly lower in the DS-5272 group than in the vehicle group on day 3 (0.55 ± 0.069 vs 0.70 ± 0.072 mg/dL, P < 0.05). DS-5272 also significantly decreased kidney injury molecule-1 (KIM-1) expression, improved tubular injury, and decreased apoptotic cells. Western blotting showed that cisplatin increased NFκB phosphorylation in kidneys, which was significantly suppressed by DS-5272. In vitro, we treated HEK 293 cells with cisplatin, in the absence or presence of DS-5272, and examined cytotoxicity and NFκB transcriptional activity. DS-5272 co-treatment reduced both cisplatin-induced cell death and NFκB transcriptional activity. Collectively, these findings suggest that DS-5272 can ameliorate cisplatin nephrotoxicity via NFκB signal inhibition.

The lymphotoxin ß receptor is a potential therapeutic target in renal inflammation.

Accumulation of inflammatory cells in different renal compartments is a hallmark of progressive kidney diseases including glomerulonephritis (GN). Lymphotoxin ß receptor (LTßR) signaling is crucial for the formation of lymphoid tissue, and inhibition of LTßR signaling has ameliorated several non-renal inflammatory models. Therefore, we tested whether LTßR signaling could also have a role in renal injury. Renal biopsies from patients with GN were found to express both LTα and LTß ligands, as well as LTßR. The LTßR protein and mRNA were localized to tubular epithelial cells, parietal epithelial cells, crescents, and cells of the glomerular tuft, whereas LTß was found on lymphocytes and tubular epithelial cells. Human tubular epithelial cells, mesangial cells, and mouse parietal epithelial cells expressed both LTα and LTß mRNA upon stimulation with TNF in vitro. Several chemokine mRNAs and proteins were expressed in response to LTßR signaling. Importantly, in a murine lupus model, LTßR blockade improved renal function without the reduction of serum autoantibody titers or glomerular immune complex deposition. Thus, a preclinical mouse model and human studies strongly suggest that LTßR signaling is involved in renal injury and may be a suitable therapeutic target in renal diseases.

Activation of innate immune defense mechanisms contributes to polyomavirus BK-associated nephropathy.

Polyomavirus-associated nephropathy (PVAN) is a significant complication after kidney transplantation, often leading to premature graft loss. In order to identify antiviral responses of the renal tubular epithelium, we studied activation of the viral DNA and the double-stranded RNA (dsRNA) sensors Toll-like receptor 3 (TLR3) and retinoic acid inducible gene-I (RIG-I) in allograft biopsy samples of patients with PVAN, and in human collecting duct cells in culture after stimulation by the dsRNA mimic polyriboinosinic:polyribocytidylic acid (poly(I:C)), cytokines, or infection with BK virus. Double staining using immunofluorescence for BK virus and TLR3 showed strong signals in epithelial cells of distal cortical tubules and the collecting duct. In biopsies microdissected to isolate tubulointerstitial lesions, TLR3 but not RIG-I mRNA expression was found to be increased in PVAN. Collecting duct cells in culture expressed TLR3 intracellularly, and activation of TLR3 and RIG-I by poly(I:C) enhanced expression of cytokine, chemokine, and IFN-ß mRNA. This inflammatory response could be specifically blocked by siRNA to TLR3. Finally, infection of the collecting duct cells with BK virus enhanced the expression of cytokines and chemokines. This led to an efficient antiviral immune response with TLR3 and RIG-I upregulation without activation of IL-1ß or components of the inflammasome pathway. Thus, PVAN activation of innate immune defense mechanisms through TLR3 is involved in the antiviral and anti-inflammatory response leading to the expression of proinflammatory cytokines and chemokines.

Loss of a renal graft due to recurrence of anti-GBM disease despite rituximab therapy.

The recurrence of anti-glomerular basement membrane (anti-GBM) glomerulonephritis (GN) in renal transplants is very rare. We report on a patient that developed acute renal allograft dysfunction due to anti-GBM GN relapse 18 months after transplantation. As plasmaseperation, dose escalation of MMF, steroids and cyclophosphamids did not result in an improvement of the graft function, a therapy with the anti-CD20 antibody Rituximab was established in addition to plasmaseperation, cyclophosphamid and steroids. Although this resulted in a decrease of anti-GBM antibody titer, graft function deteriorated further and a renal replacement therapy had to be initiated.

DNA oligonucleotide microarray technology identifies fisp-12 among other potential fibrogenic genes following murine unilateral ureteral obstruction (UUO): modulation during epithelial-mesenchymal transition.

BACKGROUND: Tubulointerstitial inflammation and fibrosis are pathologic hallmarks of end-stage renal disease (ESRD). Here we have used DNA microarray technology to monitor the transcriptomic responses to murine unilateral ureteral obstruction (UUO) with a view to identifying molecular modulators of tubulointerstitial fibrosis. METHODS: Using Affymetrix Mu74Av2 microarrays, gene expression 4 and 10 days postobstruction was investigated relative to control contralateral kidneys. Candidate profibrogenic genes were further investigated in epithelial cells undergoing epithelial to mesenchymal transition (EMT) in vitro. RESULTS: mRNA levels for 1091 gene/EST sequences, of a total of 12,488 displayed on the microarray, were altered twofold or greater by days 4 and 10 postobstruction compared to contralateral control kidneys. Genes were categorised into functional groups, including modulators of cytoskeletal and extracellular matrix metabolism, cell growth, signalling, and transcription/translational events. Among the potentially profibrogenic genes, whose mRNA levels were increased after UUO, were fibroblast-inducible secreted protein (fisp-12), the murine homologue of connective tissue growth factor (CTGF), collagen XVIIIalpha1, secreted protein acidic and rich in cysteine (SPARC), and src-suppressed C-kinase substrate (SSeCKS). A sustained increase in fisp-12 mRNA level was observed during EMT induced by transforming growth factor-beta1 (TGF-beta1) and epidermal growth factor (EGF). CONCLUSION: Altered gene expression in murine UUO has been demonstrated. Increased expression of fisp-12, SPARC, and SSeCKS has been shown in response to TGF-beta1 treatment and during EMT, suggesting that these genes may offer potential therapeutic targets against tubulointerstitial fibrosis.