Upregulation of HNF-1ß during experimental acute kidney injury plays a crucial role in renal tubule regeneration.

Hepatocyte nuclear factor-1ß (HNF-1ß) is a transcription factor expressed in the kidney, liver, pancreas, and other organs. Mutations of HNF-1ß cause maturity-onset diabetes of the young type 5 (MODY5). The aims of this study were to investigate the functional roles of the HNF-1ß/suppressor of cytokine signaling-3 (SOCS-3) pathway in tubule damage after acute kidney injury (AKI) both in vivo and in vitro and to examine the effect of HNF-1ß on renal tubule formation. To clarify the significance of the HNF-1ß/SOCS-3 pathway in AKI, we used a rat ischemia/reperfusion (I/R) AKI model and cultured renal tubular cells (NRK-52E cells). Western blot analysis showed that HNF-1ß and polycystic kidney disease 2 (PKD2) expressions were increased at 3-12 h and 12-24 h after I/R, respectively. The expression level of SOCS-3 was decreased at 3-48 h. Immunohistological examination revealed that expression of HNF-1ß was increased in proximal tubules. Overexpression of HNF-1ß resulted in decreased SOCS-3 expression, activation of signal transducer and activator of transcription 3 (STAT3) and Erk, and increased [(3)H]thymidine uptake in the presence of hepatocyte growth factor. Furthermore, tubule formation in three-dimensional gels was inhibited by dominant-negative HNF-1ß. Our study shows that HNF-1ß is upregulated after AKI in proximal tubular cells and that HNF-1ß controls cellular proliferation and tubule formation by regulating SOCS-3 expression and STAT3/Erk activation. Therefore, the current study unravels the physiological and pathological significance of the HNF-1ß pathway in AKI.

Aldosterone stimulates nuclear factor-kappa B activity and transcription of intercellular adhesion molecule-1 and connective tissue growth factor in rat mesangial cells via serum- and glucocorticoid-inducible protein kinase-1.

Several clinical and experimental data support the hypothesis that aldosterone contributes to the progression of renal injury. To determine the signaling pathway of aldosterone in relation to fibrosis and inflammation in mesangial cells, we investigated the effects of aldosterone on expression and activation of serum- and glucocorticoid-inducible protein kinase-1 (SGK1), the activation of nuclear factor-kappa B (NF-κB activation, and the expressions of intercellular adhesion molecule-1 (ICAM-1) and connective tissue growth factor (CTGF). Aldosterone stimulated SGK1 expression, phosphorylation (Ser-256), and kinase activity. The increments of phosphorylation and expression of SGK1 induced by aldosterone were inhibited by mineralocorticoid receptor (MR) inhibitor (eplerenone). Aldosterone stimulated NF-κB activity measured by NF-κB responsive elements, luciferase assay, and the levels of inhibitor of kappa B (IκB) phosphorylation. This aldosterone-induced activation of NF-κB was inhibited by the transfection of dominant-negative SGK1. Furthermore, aldosterone augmented the promoter activities and protein expressions of ICAM-1 and CTGF. The effects of aldosterone on ICAM-1 and CTGF promoter activities and protein expressions were inhibited by the transfection of dominant-negative SGK1 and dominant-negative IκBα. We also found that the MR antagonist significantly ameliorated the glomerular injury and enhancements in SGK1, ICAM-1, and CTGF expressions induced by 1% sodium chloride and aldosterone in vivo. In conclusion, our findings suggest that aldosterone stimulates ICAM-1 and CTGF transcription via activation of SGK1 and NF-κB, which may be involved in the progression of aldosterone-induced mesangial fibrosis and inflammation. MR antagonists may serve as useful therapeutic targets for the treatment of glomerular inflammatory disease.

Serum levels of soluble secreted α-Klotho are decreased in the early stages of chronic kidney disease, making it a probable novel biomarker for early diagnosis.

BACKGROUND: α-Klotho was first identified as an aging gene and was later shown to be a regulator of phosphate metabolism. Fibroblast growth factor 23 (FGF23) is the key regulator of phosphate metabolism. Serum levels of soluble α-Klotho in chronic kidney disease (CKD) patients have not previously been determined, especially in relation with FGF23 and creatinine levels. This study was designed to investigate whether serum soluble α-Klotho levels are modulated by renal function, age, and FGF23 level in CKD patients. This study is the first report on the utility of measuring soluble α-Klotho levels in human CKD. METHODS: A total of 292 CKD patients were enrolled. Serum samples were collected, and FGF23 and soluble α-Klotho levels were measured using enzyme-linked immunosorbent assay kits. In addition, serum creatinine, hemoglobin, albumin, calcium, and phosphate levels were measured. RESULTS: Serum soluble α-Klotho levels were associated positively with estimated glomerular filtration rate (eGFR) (P < 0.0001) and inversely with serum creatinine level (P < 0.01). Interestingly, α-Klotho levels were significantly decreased in stage 2 CKD compared with stage 1 (P = 0.0001). Serum FGF23 levels were associated positively with serum creatinine and negatively with eGFR. FGF23 levels were significantly increased in stage 5 compared with stage 1 CKD. Soluble α-Klotho was associated inversely with log-transformed FGF23 level (P < 0.01). CONCLUSION: Our data indicate that soluble α-Klotho levels are significantly decreased in stage 2 CKD compared to stage 1, and not only in the advanced stages of the disease. Soluble α-Klotho may thus represent a new biomarker for the diagnosis of CKD, especially in the early stage.

Positron emission tomography/computed tomography: a clinical tool for evaluation of enthesitis in patients with spondyloarthritides.

OBJECTIVE: To evaluate the accuracy of PET/CT using [(18)F]fluorodeoxyglucose (FDG) in determining the presence of enthesitis in patients with SpAs. METHODS: Results of PET/CT scans of eight patients with SpA and seven patients with RA were retrospectively examined, with specific focus on five joints and three entheses. Volume fixation values are expressed as standardized uptake values (SUVs). Data from 20 patients with non-rheumatic (NR) diseases and 20 healthy subjects were also examined if non-specific, false positive findings were possible. We evaluated the clinical utility of PET/CT examinations in SpA, compared with MRI and Ga scintigraphy. RESULTS: Images of PET/CT scans of the shoulder, hip and knee joints revealed that FDG accumulated at the entheses in SpA and in the synovium in RA patients. The maximum SUVs [mean (s.d.)] were statistically higher in SpA patients compared with RA patients at the entheses of lumbar spinous process [4.83 (1.15) vs 1.42 (0.34); P < 0.05, respectively], pubic symphysis [3.93 (0.87) vs 1.35 (0.31); P < 0.05, respectively] and ischial tuberosity [4.76 (1.5) vs 1.35 (0.42); P < 0.05, respectively]. The positive frequencies of lumbar spinous processes and ischial tuberosity evaluated by PET/CT scan in the SpA group were significantly higher than that evaluated by MRI. CONCLUSION: MRI is now widely used to detect bone marrow oedema and enthesitis in patients with SpA. PET/CT scans offer an alternative method to identify enthesitis, and will likely contribute to the early diagnosis of SpA.

Cisplatin-induced macroautophagy occurs prior to apoptosis in proximal tubules in vivo.

BACKGROUND: Autophagy is an intracellular bulk degradation process induced by cell starvation. Autophagy was recently reported to be induced by various stresses such as hypoxia, ischemia/reperfusion, toxins, and denatured proteins, and to affect cell survival and death. Light chain 3-II (LC3-II) is specifically located on double membrane-bound autophagosomes that envelop disused proteins or organelles. METHOD: Transgenic mice in which green fluorescent protein (GFP) was fused to LC3 (LC3-GFP) were administered cisplatin (20 mg/kg). After euthanasia at times between 0 and 72 h, kidneys were excised for immunohistochemical analyses. Microscopic examinations of the generated NRK-52E cell lines stably transfected with LC3-GFP, and Western blot analyses of NRK-52E cells, were undertaken after cisplatin treatment with or without autophagy inhibitors and beclin 1 small interfering RNA (siRNA). RESULTS: Autophagosomes increased in the proximal tubular cells of transgenic mice from 12 h after cisplatin injection (20 mg/kg). The time course for this was faster than those for tubular necrosis and apoptosis. Autophagosomes also increased in NRK-52E cells after cisplatin treatment, with the time course for this being faster than that for apoptosis. When autophagy was suppressed by autophagy inhibitors or beclin 1 siRNA, the level of apoptosis was also suppressed. CONCLUSION: Autophagy occurs in proximal tubular cells after cisplatin treatment and is involved in cell death in renal tubular injury. Our data suggest that autophagy is a kind of cell damage index and that cells with activated autophagy will be scavenged by apoptosis.

Effects of angiotensin II type 1 receptor blocker on albumin-induced cell damage in human renal proximal tubular epithelial cells.

BACKGROUND/AIMS: Proteinuria is not merely a marker of chronic nephropathies, but may also be involved in the progression to end-stage renal failure. We investigated the effect of angiotensin II type 1 receptor blockers (ARBs) on albumin-induced cell damage in human renal proximal tubular epithelial cells (RPTEC). METHODS: The N-acetyl-beta-D-glucosaminidase (NAG) and 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels in the medium after albumin treatment with ARBs were determined by commercially available kits. The levels of p22(phox) protein in RPTEC were measured using Western blotting after albumin treatment with ARBs. Angiotensin II concentrations in cell media and cell lysates were assayed with a commercially available kit. RESULTS: Human albumin (0.1-10 mg/ml) dose-dependently increased NAG release and olmesartan or valsartan (10(-9)-10(-7) mol/l) showed a significant reduction on albumin (1 mg/ml)-induced NAG release in RPTEC. Albumin treatment (1 mg/ml) showed significant increases in p22(phox) protein levels in RPTEC and ARBs significantly decreased albumin-induced p22(phox) protein levels. Significant increases in 8-OHdG levels were observed in the albumin (1 mg/ml)-treated group and ARBs markedly reduced albumin-induced 8-OHdG levels in RPTEC. Human albumin dose-dependently increased angiotensin II concentrations in both cell media and lysates. CONCLUSION: These observations suggest renal tubular cell-protective properties of ARBs related to decreased oxidative stress during proteinuria.

Angiotensin II receptor blocker inhibits tumour necrosis factor-alpha-induced cell damage in human renal proximal tubular epithelial cells.

AIM: We investigated the effect of angiotensin II (AII) type 1 (AT1) and angiotensin II type 2 (AT2) receptor blockers on tumour necrosis factor alpha (TNF-alpha)-induced cell damage in human renal proximal tubular epithelial cells (RPTEC). METHODS: The lactate dehydrogenase (LDH) and N-acetyl-beta-glucosaminidase (NAG) release into the medium after TNF-alpha treatment in RPTEC were determined using modified commercial procedures. In addition, the levels of caspase 3/7 activity in RPTEC were measured after TNF-alpha treatment with AlphaTau1 or AT2 receptor blockers. Finally we investigated the change of p22phox protein levels after TNF-alpha with AlphaTau1 or AT2 receptor blockers in RPTEC. RESULTS: Tumour necrosis factor alpha (10(-8) mol/L) significantly increased LDH and NAG release into the medium from RPTEC. AlphaTau1 receptor blockers, olmesartan and valsartan (10(-9)-10(-6) mol/L) showed a significant reduction on TNF-alpha-induced LDH and NAG release in RPTEC. AT2 receptor blocker, PD123319 (10(-7)-10(-5) mol/L) also decreased TNF-alpha-induced LDH and NAG release in RPTEC. Blockade of both AlphaTau1 and AT2 receptor indicated additional reduction on TNF-alpha-induced LDH and NAG release. TNF-alpha (10(-8) mol/L) treatment showed small but significant increases of caspase 3/7 activity in RPTEC, and AT1 and AT2 receptor blockers (10(-8) mol/L) comparably decreased TNF-alpha-induced caspase 3/7 activity. Significant increases of p22phox protein levels were observed in TNF-alpha-treated group in RPTEC. However, only AlphaTau1 (10(-8) mol/L) but not AT2 (10(-5) mol/L) receptor blocker significantly decreased TNF-alpha-induced p22phox protein levels. CONCLUSION: The present study demonstrates that TNF-alpha induces renal tubular cell damage in RPTEC and AT1/AT2 receptor blockers showed cytoprotective effects probably via at least partly different mechanism.

Possible involvement of intracellular angiotensin II receptor in high-glucose-induced damage in renal proximal tubular cells.

BACKGROUND: Recent studies have identified high glucose as a potent stimulus for the intracellular synthesis of angiotensin II. However, the exact roles of angiotensin II and angiotensin II type 1 receptor blockers (ARB) in high-glucose-induced renal tubular function remain unclear. METHODS: N-Acetyl-beta-glucosaminidase (NAG), angiotensin II and 8-hydroxy-2'-deoxyguanosine (8-OHdG) concentrations in renal proximal tubular epithelial cells (RPTECs) with or without high glucose/ARB were determined using a modified commercial procedure. The changes of p22phox and cytoplasmic inhibitory kappa B (IkB) protein levels in RPTECs were measured using Western blotting. RESULTS: High-glucose treatment (4x10-2 mol/L) significantly increased NAG release, angiotensin II concentrations in cell lysates and 8-OHdG and p22phox protein levels compared with those in regular glucose medium (1.75x10(-2) mol/L). ARBs (candesartan, olmesartan or valsartan; 1x10(-9)-10(-7) mol/L) showed a significant reduction in high-glucose-induced NAG, 8-OHdG and p22phox protein levels in RPTECs. Significant decreases of cytoplasmic IkB protein levels were observed in the high-glucose-treated group in RPTECs. ARBs markedly reversed high-glucose-induced reduction of IkB protein levels in RPTECs. CONCLUSIONS: ARBs reduce high-glucose-induced oxidative stress in RPTECs possibly via blockade of intracellular as well as extracellular AT1 receptor signaling, which possibly protects renal tubular cell function during diabetic nephropathy.