The proteasome inhibitor bortezomib aggravates renal ischemia-reperfusion injury.

Bortezomib is a well-established treatment option for patients with multiple myeloma (MM). It is a selective and reversible inhibitor of the proteasome that is responsible for the degradation of many regulatory proteins that are involved in apoptosis, cell-cycle regulation, or transcription. Because patients with MM are prone to develop acute renal failure, we evaluated the influence of bortezomib on renal ischemia-reperfusion injury (IRI). Mice were subjected to renal IRI by having the renal pedicles clamped for 30 min followed by reperfusion for 3, 24, and 48 h. Mice were either pretreated with 0.5 mg/kg body wt bortezomib or vehicle intravenously 12 h before induction of IRI. Serum creatinine and tubular necrosis were significantly increased in bortezomib compared with vehicle-treated mice. The inflammatory response was found to be significantly decreased in bortezomib-treated mice as reflected by a decreased infiltration of CD4(+) T cells and a significantly decreased Th1 cytokine expression in the kidneys. In contrast, apoptosis was significantly increased in kidneys of bortezomib-treated mice compared with vehicle-treated controls. Increased numbers of TUNEL-positive cells/mm(2) and increased mRNA expression of proapoptotic factors were detected in kidneys of bortezomib-treated mice. Of note, p21, a cell senescence marker, was also significantly increased in kidneys of bortezomib-treated mice. In summary, we provide evidence that bortezomib worsens the outcome of renal IRI by leading to increased apoptosis of tubular cells despite decreased infiltrating T cells and proinflammatory mediators.

Hypoxia response and VEGF-A expression in human proximal tubular epithelial cells in stable and progressive renal disease.

Proteinuria, inflammation, chronic hypoxia, and rarefaction of peritubular capillaries contribute to the progression of renal disease by affecting proximal tubular epithelial cells (PTECs). To study the transcriptional response that separates patients with a stable course from those with a progressive course of disease, we isolated PTECs by laser capture microdissection from cryocut tissue sections of patients with proteinuric glomerulopathies (stable n=20, progressive n=11) with a median clinical follow-up of 26 months. Gene-expression profiling and a systems biology analysis identified activation of intracellular vascular endothelial growth factor (VEGF) signaling and hypoxia response pathways in progressive patients, which was associated with upregulation of hypoxia-inducible-factor (HIF)-1alpha and several HIF target genes, such as transferrin, transferrin-receptor, p21, and VEGF-receptor 1, but downregulation of VEGF-A. The inverse expression levels of HIF-1alpha and VEGF-A were significantly superior in predicting clinical outcome as compared with proteinuria, renal function, and degree of tubular atrophy and interstitial fibrosis at the time of biopsy. Interactome analysis showed the association of attenuated VEGF-A expression with the downregulation of genes that usually stimulate VEGF-A expression, such as epidermal growth factor (EGF), insulin-like growth factor-1 (IGF-1), and HIF-2alpha. In vitro experiments confirmed the positive regulatory effect of EGF and IGF-1 on VEGF-A transcription in human proximal tubular cells. Thus, in progressive but not in stable proteinuric kidney disease, human PTECs show an attenuated VEGF-A expression despite an activation of intracellular hypoxia response and VEGF signaling pathways, which might be due to a reduced expression of positive coregulators, such as EGF and IGF-1.

Kidney transplantation in patients suffering from hereditary complete complement C4 deficiency.

Hereditary complete C4 deficiency (C4def) is a very rare condition that predisposes to immune complex disease and end-stage renal failure. Whether such patients should undergo renal transplantation is debated. The clinical outcome of five transplantations in three C4def patients is described. The first patient lost one allograft after 6 years because of chronic allograft rejection. Back on dialysis, he suffered from meningitis caused by Neisseria menigitidis and Aspergillus. One year after a second transplantation under alemtuzumab induction, he developed fulminant Kaposi's sarcoma and died. His sister is now 6 years post-transplantation without complications. The third patient lost his first graft after 3 years because of chronic allograft nephropathy and recurrence of glomerulonephritis. He has now been living with a second graft for over 9 years. He suffered from pneumonia, a generalized varicella infection and Hemophilis parainfluenzae bronchitis. Patients with complete C4def are at increased risk for infection after kidney transplantation. Under certain precautions and with judicious use of immunosuppression, good long-term results are achievable.

Role of alpha/beta and gamma/delta T cells in renal ischemia-reperfusion injury.

T cells have been implicated in the pathogenesis of renal ischemia-reperfusion injury (IRI). To date existing data about the role of the T cell receptor (Tcr) are contradictory. We hypothesize that the Tcr plays a prominent role in the late phase of renal IRI. Therefore, renal IRI was induced in alpha/beta, gamma/delta T cell-deficient and wild-type mice by clamping renal pedicles for 30 min and reperfusing for 24, 48, 72, and 120 h. Serum creatinine increased equally in all three groups 24 h after ischemia but significantly improved in Tcr-deficient animals compared with wild-type controls after 72 h. A significant reduction in renal tubular injury and infiltration of CD4+ T-cells in both Tcr-deficient mice compared with wild-type controls was detected. Infiltration of alpha/beta T cells into the kidney was reduced in gamma/delta T cell-deficient mice until 72 h after ischemia. In contrast, gamma/delta T cell infiltration was equal in wild-type and alpha/beta T cell-deficient mice, suggesting an interaction between alpha/beta and gamma/delta T cells. Data from gamma/delta T cell-deficient mice were confirmed by in vivo depletion of gamma/delta T cells in C57BL/6 mice. Whereas alpha/beta T cell-deficient mice were still protected after 120 h, gamma/delta T cell-deficient mice showed a "delayed wild-type phenotype" with a dramatic increase in kidney-infiltrating alpha/beta, Tcr-expressing CD4+ T-cells. This report provides further evidence that alpha/beta T cells are major effector cells in renal IRI, whereas gamma/delta T cells play a role as mediator cells in the first 72 h of renal IRI.

Cigarette smoking and chronic allograft nephropathy.

BACKGROUND: Smoking has been demonstrated to decrease patient and graft survival after kidney transplantation. Data on histological changes associated with smoking in renal allografts are lacking. METHODS: Smoking habits before and after renal transplantation were evaluated by questionnaire in 279 patients. A transplant biopsy was performed more than 1 year after transplantation in 76 of them. Histological changes were classified according to Banff 97 criteria. Linear regression analysis and proportional odds models for histological changes including the factors age, gender, diabetes, body mass index, donor age, time since transplantation, history of acute rejection and smoking status were calculated. RESULTS: Overall 22% of patients continued smoking after transplantation, with the proportion decreasing from 38% of those transplanted before 1990 to 13% of those transplanted after 2000. Serum creatinine was non-significantly higher in smokers (2.3 +/- 2.7 mg/dl vs 1.8 +/- 1.4 mg/dl, P = 0.21). A renal biopsy was performed in 24% of non-smokers and 39% of smokers (P = 0.02), and smokers were biopsied on average 1.5 years earlier. Among biopsied patients current smokers tended to suffer more often from diabetes (25.0% vs 13.5%, P = 0.33), to develop transplant failure (33.3% vs 21.2%, P = 0.25) or experience a cardiovascular event (29.2% vs 15.4%, P = 0.16). The frequency of acute rejection was comparable between smokers and non-smokers (25.0% vs 34.6%, P = 0.40). Glomerular sclerosis was associated with diabetes (P = 0.03). Severity of vascular intimal fibrous thickening was associated with current smoking (P = 0.004), whereas the degree of arteriolar hyalinosis (P < 0.001) and chronic/sclerosing nephropathy (P = 0.05) were associated with time since transplantation. CONCLUSIONS: The number of patients who continue cigarette smoking after renal transplantation has decreased in recent years. The main allograft lesion associated with smoking is fibrous intimal thickening of small arteries.

p21 and mTERT are novel markers for determining different ischemic time periods in renal ischemia-reperfusion injury.

In many clinical settings, the duration of renal ischemia and therefore the outcome of acute renal failure cannot be determined adequately. Renal ischemia reperfusion injury is known to shorten telomeres and upregulate stress-induced genes, such as the cyclin-dependent kinase (CDK) inhibitor p21. So far, the expression and role of CDK inhibitors, as well as mouse telomerase reverse transcriptase (mTERT), has not been investigated in a model with variable lasting ischemic periods. Male C57Bl/6 mice were subjected to renal ischemia reperfusion injury by clamping both renal pedicles for 10, 20, 30, and 45 min, and the kidneys were allowed to be reperfused for 3, 24, and 48 h. Expression of different CDK inhibitors and mTERT was evaluated. Mice developed signs of acute renal failure linear to the duration of the ischemic period. Real-time PCR revealed that mTERT was only significantly upregulated in kidneys after short ischemic periods (20 min). In contrast, p21 was constantly upregulated in kidneys after long ischemic intervals (30 and 45 min), but not in kidneys, which were clamped for shorter periods. Mainly, tubular cells contributed to the observed increase in p21 expression. Targeting p21 via the selective p53 inhibitor pifithrin-alpha was able to prevent acute renal failure when administered immediately before ischemia. The expression of another CDK inhibitor, namely p16, was differentially regulated, depending on the time of reperfusion. Taken together, we detected mTERT and p21 as "indicator" genes for short and long ischemic intervals, respectively. These two proteins might also be possible new therapeutic targets in the treatment and prevention of acute renal failure.

Role of mast cells in experimental anti-glomerular basement membrane glomerulonephritis.

Recently, divergent reports on the role of mast cells (MC) in different glomerular diseases have brought our attention to their role in an accelerated model of anti-glomerular basement membrane (GBM) glomerulonephritis (GN). Genetically MC-deficient Kit(W)/Kit(W-v) mice, MC-reconstituted Kit(W)/Kit(W-v) mice and Kit+/+ control mice were subjected to anti-GBM GN. Kit(+/+) mice developed moderate proteinuria and glomerular damage following the induction of anti-GBM nephritis. In contrast, proteinuria and glomerular damage were dramatically increased in MC-deficient Kit(W)/Kit(W-v) mice. MC-reconstituted Kit(W)/Kit(W-v) mice showed proteinuria and glomerular damage comparable to Kit+/+ mice. A significant increase in infiltrating T cells and macrophages was detected in MC-deficient Kit(W)/Kit(W-v) mice as compared to Kit+/+ control mice and MC-reconstituted Kit(W)/Kit(W-v) mice. Accordingly, we observed an increase of TGF-beta1 mRNA in kidneys from Kit(W)/Kit(W-v) mice. Interestingly, we did not detect MC in the kidney using either Giemsa staining or RT-real-time PCR, but MC were found in the regional lymph nodes. Finally, mortality of Kit(W)/Kit(W-v) mice was significantly increased after the induction of anti-GBM GN due to uremia. Our report provides the first direct evidence that MC are protective in anti-GBM GN, possibly by modulating the influx of effector T cells and macrophages to inflammatory sites in the kidney.