Balazs Antus

Inhibition of matrix metalloproteinases during chronic allograft nephropathy in rats.

Background. Chronic allograft nephropathy (CAN) belongs to the major causes of long-term kidney allograft failure. One of the histologic hallmarks of CAN is interstitial fibrosis, influenced by matrix metalloproteinases (MMPs) that are controlling extracellular matrix (ECM) degradation. Whether MMPs affect the development and progression of CAN is not clear so far. To analyze the role of MMPs in CAN, we investigated the effects of an early and a late application of BAY 12–9566, an inhibitor of MMP-2, -3, and -9 on the development and progression of CAN in a rat kidney-transplantation model. Methods. Fisher kidneys were orthotopically transplanted into Lewis recipients that were treated with BAY 12–9566 (15 mg/kg per day) or vehicle either for the first 10 days after transplantation (early treatment) or from week 12 to week 20 after transplantation (late treatment). Proteinuria was analyzed every 4 weeks up to week 20 after transplantation when kidney grafts were removed for further analysis. Results. Early MMP-inhibition resulted in a significantly reduced 24-hour protein excretion that was paralleled by a lower grade of CAN after 20 weeks. However, late MMP inhibition starting at week 12 after transplantation resulted in significantly higher proteinuria and a higher grade of CAN as compared with controls. Furthermore, transforming growth factor-&bgr; and platelet-derived growth factor-B chain mRNA levels were significantly increased in these animals. Conclusions. Inhibition of MMPs early after transplantation reduced the development and progression of CAN but promoted CAN if initiated at later stages. Thus, MMPs are involved in the development and progression of CAN.

Apoptosis and treatment of chronic allograft nephropathy with everolimus

Background. Chronic allograft nephropathy (CAN) is responsible for most cases of late kidney allograft loss. However, no effective treatment is available so far. Everolimus (RAD) (40-O [2-hydroxyethyl] rapamycin) is a new immunosuppressive agent with antiproliferative and apoptosis-enhancing effects. We asked whether everolimus can ameliorate CAN even at advanced stages, whether everolimus treatment affects the level of growth factor mRNA, and whether everolimus treatment affects the number of apoptotic cells in the graft. Methods. We transplanted kidneys from Fisher rats into Lewis rats and treated recipients with everolimus over different time periods. Grafts were analyzed 20 or 28 weeks after transplantation. Results. Everolimus delayed the progression of CAN when started at an early stage. Surprisingly, everolimus even ameliorated CAN when initiated at an advanced stage. Interestingly, apoptosis was more prevalent in treated animals, particularly in those with delayed treatment as compared with controls. Conclusions. Everolimus ameliorates CAN as a result of antiproliferative or apoptosis-enhancing effects.

Opposite effects of testosterone and estrogens on chronic allograft nephropathy.

Abstract In the present study we investigated whether donor gender of the effects of sex hormones play the greater role in the development of chronic allograft nephropathy. Kidneys of male and female Fisher rats were orthotopically transplanted into castrated male Lewis recipients. Animals were treated with testosterone, estradiol, or vehicle and the kidneys were harvested 20 weeks after transplantation for histological, immunohistological, and molecular analysis. Testosterone treatment resulted in increased proteinuria and profound glomerulo‐sclerosis, irrespective of donor gender. In addition, mRNA levels of transforming growth factor‐β1 (TGF‐β1) and platelet‐derived growth factor‐A and B (PDGF‐A and B) chains were enhanced in these allografts. Estradiol reduced glomerulosclerosis and mononuclear cell infiltration in allografts of both genders that paralleled a decreased mRNA expression of TGF‐β1, PDGF‐A and B. No donor gender‐related differences were noted in vehicle‐treated animals. Our findings demonstrate that sex hormones rather than donor gender have a significant impact on chronic allograft nephropathy.

Chronic Renal Failure Leads to Reduced Flow-Dependent Dilation in Isolated Rat Skeletal Muscle Arterioles due to Lack of NO Mediation

Background: Chronic renal failure (CRF) is frequently accompanied by systemic vascular alterations which further increase the morbidity and mortality of these patients. However, the nature and the underlying mechanisms of vascular dysfunction are not completely understood. We hypothesized that – in addition to other factors – CRF alters local vasomotor mechanisms that are intrinsic to the vascular wall. Methods: Changes in the diameter of isolated, pressurized (at 80 mm Hg) gracilis skeletal muscle arterioles (diameter approximately 150 µm) of female Wistar rats were investigated by videomicroscopy. Arteriolar responses to an increase in flow and vasoactive agents in partially nephrectomized (NX) and sham-operated (control) rats were compared. Results: In NX rats, serum creatinine and urine protein excretion were increased. Compared to controls, increases in intraluminal flow (from 0 to 40 µl/min) resulted in significantly reduced dilation in arterioles of NX rats (maximum: 32 ± 4 vs. 15 ± 4 µm, p < 0.05). Inhibition of nitric oxide (NO) synthesis with L-NAME reduced the dilation of control arterioles but did not affect responses of NX arterioles. Also, dilations in response to histamine were significantly reduced in arterioles from NX rats as compared to control rats. L-NAME significantly decreased histamine-induced dilations of control arterioles, but it did not affect responses of NX arterioles. Dilations in response to the NO donor sodium nitroprusside were also significantly decreased in NX arterioles as compared to responses of control vessels, whereas responses to adenosine and norepinephrine were not significantly different in the two groups. Conclusions: We conclude that in rat skeletal muscle arterioles, CRF induced by renal mass reduction alters the mechanosensitive and agonist-induced responses of peripheral arterioles, in part by interfering with NO-signaling mechanisms. These alterations could contribute to increased peripheral vascular resistance and further aggravate the cardiovascular complications in CRF.

Extracellular Signal-Regulated Kinase and the Small GTP-Binding Protein p21Rac1 Are Involved in the Regulation of Gene Transcription by Angiotensin II

To study the role of extracellular-signal-regulated kinase (ERK) cascade and the small GTP-ase proteins in the activation of the c-fos promoter by angiotensin II (AII), transient transfection experiments were performed in CHO cells stably expressing the rat AT1A receptor. In this system AII activated ERK in 1 min and also increased the transcriptional activity of the c-fos promoter-luciferase reporter gene construct. The activation of the promoter proved to be dependent on the Ras-Raf-ERK cascade as cotransfection of expression vectors known to specifically inhibit this cascade blocked the effect of AII. Dominant-negative p21Rac1 mutant partially blocked the activation of the c-fos promoter by AII. However, activation of the c-fos promoter was independent of protein kinase C (PKC) as bisindolylmaleimide I, a specific PKC inhibitor did not block the effect of AII. These results suggest that AII activates the transcription of the c-fos through the Ras-Raf-ERK cascade. Furthermore, p21Rac1 is involved in the modulation of the c-fos promoter by AII.