The selective mineralocorticoid receptor antagonist eplerenone is protective in mild anti-GBM glomeru-lonephritis.

BACKGROUND: Growing evidence suggests that blockade of the aldosterone-receptor may preserve kidney function by anti-inflammatory effects independent of the blood pressure. We hypothesized that the selective aldosterone-receptor antagonist eplerenone has a profound anti-inflammatory effect in the autologous phase of anti-glomerular basement membrane (GBM) glomerulonephritis (GN). METHODS: Mice received ≈200mg/kg body wt/day eplerenone via supplemented chow diet or standard chow starting at the day of immunization with rabbit IgG. Three days later the anti-GBM antibody was injected and the experiments were stopped at day 7 and 14. RESULTS: Mice receiving eplerenone showed significantly decreased albuminuria and glomerular sclerosis at day 7 and 14 after induction of anti-GBM GN. Eplerenone treatment significantly inhibited the infiltration of CD4+, CD8+ T cells and macrophages into the kidneys. Circulating levels and glomerular deposition of autologous IgG were comparable in both groups. At day 7 the pro-inflammatory cytokines MCP-1 and IL-6 were found to be significantly decreased in regional draining lymph nodes of eplerenone-treated mice, whereas the anti-inflammatory cytokine IL-10 was significantly upregulated. In line, splenocytes from eplerenone-treated nephritic mice produced significantly increased IL-10. CONCLUSION: Aldosterone-receptor blockade by eplerenone effectively attenuated proteinuria, kidney damage and the inflammatory response in anti-GBM GN by significantly decreasing pro-inflammatory cytokines in the regional draining lymph nodes of the kidney. Our results suggest that this selective aldosterone receptor antagonist is a possible additional tool in the treatment of GN.

Potential role of regulatory T cells in reversing obesity-linked insulin resistance and diabetic nephropathy.

OBJECTIVE: To assess the potential role of FoxP3-expressing regulatory T cells (Tregs) in reversing obesity-linked insulin resistance and diabetic nephropathy in rodent models and humans. RESEARCH DESIGN AND METHODS: To characterize the role of Tregs in insulin resistance, human visceral adipose tissue was first evaluated for Treg infiltration and second, the db/db mouse model was evaluated. RESULTS: Obese patients with insulin resistance displayed significantly decreased natural Tregs but an increase in adaptive Tregs in their visceral adipose tissue as compared with lean control subjects. To further evaluate the pathogenic role of Tregs in insulin resistance, the db/db mouse model was used. Treg depletion using an anti-CD25 monoclonal antibody enhanced insulin resistance as shown by increased fasting blood glucose levels as well as an impaired insulin sensitivity. Moreover, Treg-depleted db/db mice developed increased signs of diabetic nephropathy, such as albuminuria and glomerular hyperfiltration. This was paralleled by a proinflammatory milieu in both murine visceral adipose tissue and the kidney. Conversely, adoptive transfer of CD4(+)FoxP3(+) Tregs significantly improved insulin sensitivity and diabetic nephropathy. Accordingly, there was increased mRNA expression of FoxP3 as well as less abundant proinflammatory CD8(+)CD69(+) T cells in visceral adipose tissue and kidneys of Treg-treated animals. CONCLUSIONS: Data suggest a potential therapeutic value of Tregs to improve insulin resistance and end organ damage in type 2 diabetes by limiting the proinflammatory milieu.

A murine model of phosphate nephropathy.

We established a murine model of phosphate nephropathy with secondary hyperparathyroidism. db/db mice, which develop obesity and type 2 diabetes mellitus, were uninephrectomized at the age of 6 weeks and were fed either standard chow or a phosphorus-rich diet during the next 8 weeks. Thereafter, renal cryosections showed abundant tubular casts with a strong histochemical von Kossa reaction in all db/db mice on the phosphorus-rich diet but none in the controls. X-ray diffraction and Raman spectroscopy proved that these tubular casts consist mostly of hydroxyapatite Ca5(PO4)3(OH). These intraluminal precipitations were located in distal tubuli and collecting ducts and were associated with degenerative tubular changes and peritubular infiltration of T cells and macrophages. In line, kidneys of db/db mice on the phosphorus-rich diet displayed significantly increased mRNA expression of the T(H)1 cytokines interferon γ, IL-6, and tumor necrosis factor α. In addition, mice developed signs of secondary hyperparathyroidism as shown by elevated serum phosphate, decreased serum calcium, and increased parathyroid hormone, osteopontin, and fibroblast growth factor 23 levels. db/db mice on the phosphorus-rich diet also presented with significantly lower body weight, lower homeostasis model assessment of insulin resistance index, and hypertrophic cardiomyopathy. Thus, we provide a murine model of phosphate nephropathy and secondary hyperparathyroidism, which can be used for future pharmacologic and pathophysiologic studies to analyze the effect of hyperphosphatemia on renal, metabolic, and cardiovascular phenotypes.

Atorvastatin attenuates murine anti-glomerular basement membrane glomerulonephritis.

Statins mediate many of their protective effects by lowering lipids as well as by modulating inflammation. Here, we studied their potential immunomodulatory role in renal inflammation using an autoimmune mouse model of anti-glomerular basement membrane glomerulonephritis. Oral treatment with Atorvastatin dramatically reduced albuminuria and histological changes in the kidneys as compared to vehicle-treated control animals. There was a significant decrease in the Th1 and Th17 response in the regional lymph nodes draining the kidneys. This systemic effect was accompanied by decreased infiltration of the kidneys with inflammatory CD4(+) T and Th17 cells, macrophages, and neutrophils in statin-treated mice. Regulatory T cells were not altered in their number, FoxP3 expression, or suppressive capacity, but their interleukin-10 production was significantly increased by statin treatment. Hence, Atorvastatin systemically and locally decreased the Th1 and Th17 response, thereby protecting the mice against anti-glomerular basement membrane glomerulonephritis. Whether statins can be used to treat human autoimmune renal diseases will require more direct studies.

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.

Differential effects of rapamycin in anti-GBM glomerulonephritis.

The immunosuppressive mammalian target of rapamycin inhibitor rapamycin is widely used in solid-organ transplantation, but the effect of rapamycin on kidney disease is controversial. This study evaluated the effect of rapamycin in the autologous phase of anti-glomerular basement membrane (anti-GBM) glomerulonephritis. Disease was induced by preimmunizing the animals with rabbit IgG 5 d before administration of rabbit anti-mouse GBM antiserum. When rapamycin was started on the day of immunization (group 1), mice were protected from glomerulonephritis, suggested by a dramatic decrease in albuminuria, influx of inflammatory cells, and Th1-cytokine expression in the kidneys. Activation of T cells and production of autologous mouse anti-rabbit IgG were also significantly reduced in rapamycin-treated animals. In contrast, when rapamycin was started 14 d after immunization (group 2), mice had a significant increase in albuminuria and renal infiltration of inflammatory cells compared with vehicle-treated animals, and there were no differences in T and B cell responses. A significant decrease in vascular endothelial growth factor-A and an increase in IL-6 were detected in kidneys of these rapamycin-treated mice. In conclusion, rapamycin has the potential to significantly reduce the B and T cell responses and thereby protect from glomerulonephritis when administered early in disease. Once disease is established, however, rapamycin seems to worsen glomerulonephritis by disturbing the endothelial cell/vascular endothelial growth factor system in the kidney.

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.

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.

Progression of arteriovenous bypass restenosis in mice exposed to a 50 Hz magnetic field.

The controversy over whether magnetic fields (MF) produced by electrical wiring and appliances contribute to diseases such as cancer has been debated in the literature for more than 2 decades. These extremely low frequency fields at 50 or 60 Hz are omnipresent in the industrialized world and have been linked to various forms of cancer by epidemiological studies. Little has been published investigating any possible role of MF and cardiovascular disease, and this is the first study looking specifically at the effect of exposure to high-intensity MF on the development and progression of restenosis. A mouse arteriovenous bypass model was used, and mice were exposed to MF for periods of 1, 2, or 3 weeks. Neointima formation, infiltration of mononuclear cells, and heat shock protein 60 expression were all studied at the conclusion of the exposure regimen. Animals exposed to the MF for 1 week showed significantly smaller neointima formation compared with control mice exposed to a null field, although this difference was not observed in mice exposed for 2 or 3 weeks. No difference was found between mice exposed to MF and controls in any of the other parameters investigated.