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In the normal condition, endogenous formation of peroxynitrite ( ONOOˉ) from the interaction of nitric oxide and superoxide has been suggested to play a renoprotective role. However, the exact mechanism associated with renoprotection by this radical compound is not yet clearly defined. Although ONOOˉ usually inhibits renal tubular Na+K+ ATPase ( NKA) activity at high concentrations (micromolar to millimolar range [ μM-mM], achieved in pathophysiological conditions), the effects at lower concentrations (nanomolar range [nM], relevant in normal condition) remain unknown. To examine the direct effect of ONOOˉ on NKA activity, preparations of cellular membrane fraction from mouse renal tissue and from cultured HK2 cells (human proximal tubular epithelial cell lines) were incubated for 10 and 30 min each with different concentrations of ONOOˉ (10 nmol/L-200 μmol/L). NKA activity in these samples ( n = 5 in each case) was measured via a colorimetric assay capable of detecting inorganic phosphate. At high concentrations (1-200 μmol/L), ONOOˉ caused dose-dependent inhibition of NKA activity (−3.0 ± 0.6% and −36.4 ± 1.4%). However, NKA activity remained unchanged at 100 and 500 nmol/L ONOOˉ concentration, but interestingly, at lower concentrations (10 and 50 nmol/L), ONOOˉ caused small but significant increases in the NKA activity (3.3 ± 1.1% and 3.1 ± 0.6%). Pretreatment with a ONOOˉ scavenger, mercaptoethylguanidine ( MEG; 200 μmol/L), prevented these biphasic responses to ONOOˉ. This dose-dependent biphasic action of ONOO− on NKA activity may implicate that this radical compound helps to maintain sodium homeostasis either by enhancing tubular sodium reabsorption under normal conditions or by inhibiting it during oxidative stress conditions. [ABSTRACT FROM AUTHOR]

Renal inflammation modulates angiotensinogen (AGT) production in renal proximal tubular cells (RPTCs) via inflammatory cytokines, including interleukin-6, tumor necrosis factor α, and interferon-γ (IFN-γ). Among these, the effects of IFN- on AGT regulation in RPTCs are incompletely delineated. This study aimed to elucidate mechanisms by which IFN-γ regulates AGT expression in RPTCs. RPTCs were incubated with or without IFN-γ up to 48 h. AGT expression, STAT1 and STAT3 activities, and SOCS1 expression were evaluated. RNA interference studies against STAT1, SOCS1, and STAT3 were performed to elucidate a signaling cascade. IFN-γ decreased AGT expression at 6 h (0.6±0.05, ratio to control) and 12 h (0.47±0.03). In contrast, longer exposure for 24 and 48 h increased AGT expression (1.76±0.18, EC50 = 3.4 ng/ ml, and 1.45±0.08, respectively). IFN-γ treatment for 6 h strongly induced STAT1 phosphorylation and SOCS1 augmentation, and decreased STAT3 activity. However, STAT1 phosphorylation and SOCS1 augmentation waned at 24 h, while STAT3 activity increased. RNA interference studies revealed that activation of STAT1-SOCS1 axis decreased STAT3 activity. Thus, IFN-γ biphasically regulates AGT expression in RPTCs via STAT3 activity modulated by STAT1-SOCS1 axis, suggesting the STAT1-SOCS1 axis is important in IFN-γ-induced activation of the intrarenal renin-angiotensin system. [ABSTRACT FROM AUTHOR]

1. Using HIGA (high IgA of ddY) mice as an IgA nephropathy model and BALB/c mice as controls, we demonstrated that reactive oxygen species (ROS) and the renin–angiotensin system (RAS) were activated in kidneys of HIGA mice. However, it was difficult to establish an association between renal damage and changes in ROS and the RAS. Therefore, the present study was performed to determine whether renal injury is associated with changes in ROS and the RAS in HIGA mice. 2. Male HIGA mice were divided into four groups of 10 each: (i) untreated mice (HIGA + null); (ii) mice treated with the angiotensin AT1 receptor antagonist olmesartan (5 mg/kg per day; HIGA + OLM); (iii) mice treated with the superoxide dismutase mimetic tempol (50 mg/kg per day; HIGA + Tempol); and (iv) mice treated with RAS-independent antihypertensive drugs (30 mg/kg per day hydralazine, 0.6 mg/kg per day reserpine and 12 mg/kg per day hydrochlorothiazide; HIGA + HRH). Mice were treated for 5 weeks. 3. Systolic blood pressure decreased significantly in the HIGA + OLM and HIGA + HRH groups, but not in the HIGA + Tempol group, compared with HIGA + null mice. The expression of two ROS markers (4-hydroxy-2-nonenal and heme oxygenase-1) and angiotensin II as a marker of the RAS decreased significantly in HIGA + OLM and HIGA + Tempol mice, but not in HIGA + HRH mice, compared with HIGA + null mice. As a marker of renal damage, mesangial matrix expansion and the desmin-positive area decreased significantly in the HIGA + OLM and HIGA + Tempol groups, but not in HIGA + HRH group, compared with the HIGA + null group. 4. These data suggest that intrarenal ROS and RAS activation play a pivotal role in the development of IgA nephropathy model mice, from the early phase, independent of blood pressure. [ABSTRACT FROM AUTHOR]

1. Although IgA nephropathy is the most common form of primary glomerulopathy, the detailed mechanisms underlying its development remain uncertain. 2. In the present study, we used male high IgA strain of ddY (HIGA) mice as the IgA nephropathy model and age-matched male BALB/c mice as the control. Recent studies have demonstrated that reactive oxygen species (ROS)-dependent enhancement of the renin–angiotensin system (RAS) plays a potential role in the development and progression of renal injury. Therefore, in the present study we periodically measured the systolic blood pressure (SBP) of mice over the period 21–25 weeks of age and estimated markers for ROS, RAS and renal damage after mice had been killed at 25 weeks of age. 3. Markers for ROS (urinary 8-isoprostane excretion and renal 4-hydroxy-2-nonenal accumulation), RAS (renal angiotensinogen protein expression, urinary angiotensinogen excretion and renal angiotensin II) and renal damage (desmin-positive area and urinary protein excretion), as well as SBP, were significantly increased in HIGA mice compared with control BALB/c mice. 4. The data suggest that both ROS and the RAS are activated at an early phase in IgA nephropathy model mice. [ABSTRACT FROM AUTHOR]

Chymase released from mast cells produces pro‐fibrotic, inflammatory, and vasoconstrictor agents. Studies were performed to test the hypothesis that chronic chymase inhibition provides a renal protective effect in type 2 diabetes. Diabetic (db/db) and control mice (db/m) were chronically infused with a chymase‐specific inhibitor or vehicle for 8 weeks. Baseline urinary albumin excretion (UalbV) averaged 42 ± 3 and 442 ± 32 microg/d in control (n = 22) and diabetic mice (n = 27), respectively (p <.05). After administration of chymase inhibitor to diabetic mice, the change in UalbV was significantly lower (459 ± 57 microg/d) than in vehicle‐treated diabetic mice (645 ± 108 microg/d). UNGALV was not different at baseline between diabetic mice that would receive the chymase inhibitor (349 ± 56 ng/d, n = 6) and vehicle (373 ± 99 ng/d, n = 6) infusions, but increased significantly only in the vehicle‐treated diabetic mice (p <.05). Glomeruli of diabetic kidneys treated chronically with chymase inhibition demonstrated reduced mesangial matrix expansion compared to glomeruli from untreated diabetic mice. Plasma angiotensin II levels were not altered by chymase inhibitor treatment. In summary, chronic chymase inhibition slowed the progression of urinary albumin excretion in diabetic mice. In conclusion, renal chymase may contribute to the progression of albuminuria in type 2 diabetes renal disease. [ABSTRACT FROM AUTHOR]

Background: Megalin and the Ang II type 1 receptor (ATIR) are coexpressed in the renal proximal tubule where megalin is involved in endocytosis and degradation of proteins that escape the glomerular barrier. Because Ang II uptake by AT1R-expressing BN/MSV cells (a non-renal cell line) is partially mediated by megalin, we tested if megalin mediates Ang II uptake in proximal tubule cells. Methods: Cultured proximal tubule cells (NRK-52E cells) were exposed to fluorescent Ang II for 4 hours (Alexa488-Ang II, 100 nM). Olmesartan (or Candesartan 10 µM) and anti-megalin antisera (or albumin 1 mg/mL) were used to block the AT1R and the megalin pathways respectively. Flow cytometry was used to measure single cell-associated fluorescence. Confocal or fluorescence microscope systems were used for image capturing and analysis. Results: In flow cytometry experiments, both anti-megalin antisera and AT1R blockers significantly reduced Ang II uptake (30% and 60% reductions respectively, p <0.05 vs. positive control). In imaging experiments, Ang II accumulated mostly in a perinuclear area and this accumulation was reduced by treatment with albumin, AT1R blockers or both. Conclusion: Megalin mediates Ang II uptake into proximal tubule cells in addition to the AT1R. Because megalin is responsible for the scavenging of proteins in the tubular fluid, megalin-mediated Ang II uptake might help to regulate intrarenal Ang II levels. [ABSTRACT FROM AUTHOR]