Tubular Epithelial Cell Damage Research Articles

Backcross db Gene into CD-1 Background Results in Novel Type 2 Diabetic Mouse Model with Progressive Kidney Fibrosis

Kidney fibrosis is the final common pathway of progressive kidney disease including diabetic kidney disease (DKD). With the rapid increase of type 2 diabetes, DKD in type 2 diabetic patients has become a serious public health threat around the world. To establish novel therapy to combat DKD, human disease relevant animal model is essential. However, a type 2 diabetic mouse model with presenting progressive kidney fibrosis is not established yet. We have previously reported that kidneys of streptozotocin-induced diabetic CD-1 mice exhibited severe fibrosis compared to other backgrounds of mice associated with the suppression of anti-fibrotic peptide N-acetyl-seryl-aspartyl-lysyl-proline, fibroblast growth factor receptor 1, and their downstream target anti-fibrotic microRNAs (miRs) such as miR 29 and miR let-7. As the type 2 diabetic mice model, leptin receptor deficient mice in BKS background (BKSdb/db) are often utilized; the kidney fibrosis in BKSdb/db phenotype is minimum. Here we backcrossed db gene into potential fibrotic CD-1 background and found that male CD-1db/db exhibited severe kidney fibrosis when compared to male BKSdb/db. We have already performed the backcrosses for 11 generations. Body weight of CD-1db/db was comparable but drank up to 3 times more water compared to BKSdb/db. Blood glucose levels in CD-1db/db appeared to be higher when compared to BKSdb/db. When evaluated the per body weight ratio, kidney and heart weighed no significant differences between CD-1db/db and BKSdb/db. CD-1db/db exhibited severe tubulo-interstitial fibrosis, glomerulosclerosis, and tubular epithelial cells damage. Heart in CD-1db/db displayed fibrotic change as well. In conclusion, here we provide novel fibrotic type 2 DKD mouse model. CD-1db/db could be excellent model to perform experimental therapy for DKD. Disclosure Y. Mizunuma: None. K. Nitta: None. S. Takagi: None. K. Kanasaki: Advisory Panel; Self; Boehringer Ingelheim Pharmaceuticals, Inc.. Speaker's Bureau; Self; Eli Lilly and Company, Sanofi, Mitsubishi Tanabe Pharma Corporation. Research Support; Self; Boehringer Ingelheim GmbH. Speaker's Bureau; Self; Boehringer Ingelheim GmbH, Astellas, Astellas. Research Support; Self; Ono Pharmaceutical Co., Ltd., Mitsubishi Tanabe Pharma Corporation. D. Koya: None.

Effects of intermittent hypoxia exposure on renal tubular epithelial cell injury and autophagy in rats

Objective: To investigate the effects of intermittent hypoxia (IH) exposure on renal tubular epithelial cell injury and autophagy in rats. Methods: A total of 12 Sprague Dawley (SD) rats were randomly divided into two groups: normal control (NC) group and IH group, with 6 rats in each group. An IH animal model was established to observe the effects of IH on renal tubular epithelial cell damage and autophagy in rats. Microalbumin (mAlb) was detected by immunoturbidimetry. Neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1(KIM-1)in urine were assessed by enzyme-linked immunosorbent assay (ELISA) test. The pathological damage of renal tubules was observed by hematoxylin-eosin (HE) staining. The expression of autophagy marker protein light chain 3 (LC3) and Beclin-1 were detected by immunohistochemistry and immunoblotting. Results: At the end of the test, the levels of mAlb [(34.7±6.7) mg/L vs (11.1±3.3) mg/L, P=0.011], NGAL [(17.3±3.9) ng/ml vs(4.0±1.7)ng/ml, P=0.011] and KIM-1 [ (10.8±2.7) ng/ml vs (2.6±1.0) ng/ml, P=0.016] in urine of IH group were higher than those of control group, and the differences were statistically significant (all P<0.05). Pathological injury was showed by HE staining in IH group. The increase of autophagy in IH group was more obvious than that in NC group. Besides, the expression level of autophagy markers (protein LC3 and Beclin-1) was significantly higher in IH group than that in NC group (P<0.05). Conclusions: IH can not only cause renal tubular injury, but also induce increased autophagy in renal tubular epithelial cells. Autophagy activation might participate in renal tubular epithelial cell injury induced by IH.

Neutrophil Gelatinase-Associated Lipocalin Attenuates Ischemia/Reperfusion Injury in an In Vitro Model via Autophagy Activation.

BackgroundThe aim of this study was to investigate the protective effects of neutrophil gelatinase-associated lipocalin (NGAL) on hypoxia/reoxygenation (H/R) induced acute kidney injury (AKI) in vitro.Material/MethodsWe used NRK-52E cells and H/R treatments to mimic ischemia/reperfusion injury (IRI) in vitro. Experimental groups were: the control group, the H/R group, the 3-methyladenine (3-MA)+H/R group, the NGAL (0.25, 0.5, and 1 ug/mL)+H/R group, and the NGAL (0.25, 0.5, 1 ug/mL)+3-MA+H/R group. After 24 hours of culture, cell proliferation was analyzed by CCK-8 assay. Expression of LC3-II was detected by immunoblot assay. Autophagy was detected by electron microscopy.ResultsThe expression of LC3-II was increased in the H/R group compared with normoxic condition (p<0.05) and proliferation also improved. Autophagy was significantly inhibited by 3-MA, with downregulated of LC3-II, followed by decreased cell viability (p<0.05). We further detected the effect of different doses of NGAL in H/R induced injury, and found that low doses of NGAL alone slightly increased LC3-II protein accumulation, and autophagy was further induced with higher dose of NGAL treatment. Meanwhile, cell viability assays showed induced cell survival. We found that in the NGAL+3-MA group, cell viability assays revealed reduced cell damage, followed concomitantly with depressed autophagy. The formulation of autophagosomes were correlated with LC3-II protein expression in each group.ConclusionsAutophagy plays a renoprotective role in H/R injury, as well in AKI. NGAL might be related to attenuated tubular epithelial cell damage via adjusting autophagy.

Massive Proteinuria-Induced Injury of Tubular Epithelial Cells in Nephrotic Syndrome is Not Exacerbated by Furosemide

Background/Aims: Massive proteinuria, a significant sign of nephrotic syndrome (NS), has the potential to injure tubular epithelial cells (TECs). Furosemide is widely used for the treatment of edema, a common manifestation of NS. However, whether furosemide treatment affects massive proteinuria-induced TEC injury in patients with NS is unknown. Methods: The effect of furosemide on TEC damage was investigated in vitro. In addition, a clinical study was conducted to study whether the short-term treatment of nephrotic edema with furosemide could exacerbate TEC injury. Results: The proliferation of in vitro human kidney-2 (HK-2) cells exposed to massive urinary protein (8 mg/mL) significantly decreased (P<0.05), while the levels of kidney injury molecule-1 (Kim-1) and neutrophil gelatinase associated lipocalin (NGAL) in the supernatants significantly increased (P<0.05). Importantly, furosemide treatment did not further increase the expression of Kim-1 and NGAL in HK-2 cells upregulated by massive proteinuria. For the clinical study, 26 patients with NS, all prescribed the recommended dosage of prednisone (1 mg/kg/day), were randomly assigned to two groups. One group (n=13) received furosemide (60-120 mg/day, intravenously) for 1 week; the remaining participants (control group) did not receive furosemide or any other diuretics. The results showed that the 24-h urine volume in the furosemide-treated group was slightly, but not significantly, higher than that in the control group (P>0.05). In addition, serum levels of BUN, Scr, Cys C, and urinary Kim-1 and NGAL were not significantly different between the two groups (all P>0.05). Twenty-three patients underwent a renal biopsy. Of these, 22 patients exhibited vacuolar degeneration of the TECs; 8 patients showed brush border membrane shedding of the TECs; and 12 patients showed protein casts. However, there were no significant differences between the two groups (all P>0.05). Conclusion: In summary, massive proteinuria induced the injury of TECs in patients with NS, and furosemide treatment did not aggravate this injury.

Mitochondrial oxidative damage and apoptosis induced by high glucose through Rho kinase signal pathway in renal tubular epithelial cells

Objective: To investigate the role of oxidative stress in human renal tubular epithelial cells (HK-2) induced by high glucose and the underlying signal pathway in vitro. Methods: MYPT1, pro-caspase-3, PGC-1α, and Drp1 protein expressions were measured by Western blot. MnSOD2, Drp1 and PGC-1α mRNA expressions were detected by real time PCR. Results: Results showed that high glucose significantly up-regulated the protein expressions of MYPT1, pro-caspase-3 and the mRNA expression of MnSOD2 in HK-2 cells; while Rho kinase inhibitor fasudil and ROCK1 siRNA inhibited protein expressions of pro-caspase-3 and the mRNA expression of MnSOD2 in HK-2 cells induced by high glucose. Importantly, fasudil and ROCK1 siRNA markedly inhibited the expressions of mitochondrial motor proteins Drp1 and mitochondrial gene PGC-1α in HK-2 cells induced by high glucose. Conclusions: Our findings suggest that Rho kinase signal pathway is involved in mitochondrial oxidative damage and apoptosis in high glucose-induced renal tubular epithelial cells by regulating mitochondrial motor proteins Drp1 and mitochondrial gene PGC-1α. Targeting Rho kinase signal pathway might be a potential strategy for the treatment of diabetic nephropathy.

PDE/cAMP/Epac/C/EBP-β Signaling Cascade Regulates Mitochondria Biogenesis of Tubular Epithelial Cells in Renal Fibrosis.

Cyclic adenosine 3'5'-monophosphate (cAMP) is a universal second messenger that plays an important role in intracellular signal transduction. cAMP is synthesized by adenylate cyclases from adenosine triphosphate and terminated by the phosphodiesterases (PDEs). In the present study, we investigated the role of the cAMP pathway in tubular epithelial cell mitochondrial biogenesis in the pathogenesis of renal fibrosis. We found that the cAMP levels were decreased in fibrotic kidney tissues, and replenishing cAMP could ameliorate tubular atrophy and extracellular matrix deposition. The downregulation of cAMP was mainly attributed to the increased PDE4 expression in tubular epithelial cells. The inhibition of PDE4 by PDE4 siRNA or the specific inhibitor, rolipram, attenuated unilateral ureteral obstruction-induced renal interstitial fibrosis and transforming growth factor (TGF)-β1-stimulated primary tubular epithelial cell (PTC) damage. The Epac1/Rap1 pathway contributed to the main effect of cAMP on renal fibrosis. Rolipram could restore C/EBP-β and PGC-1α expression and protect the mitochondrial function and structure of PTCs under TGF-β1 stimulation. The antifibrotic role of rolipram in renal fibrosis relies on C/EBP-β and PGC-1α expression in tubular epithelial cells. Innovation and Conclusion: The results of the present study indicate that cAMP signaling regulates the mitochondrial biogenesis of tubular epithelial cells in renal fibrosis. Restoring cAMP by the PDE4 inhibitor rolipram may ameliorate renal fibrosis by targeting C/EBP-β/PGC1-α and mitochondrial biogenesis. Antioxid. Redox Signal. 29, 637-652.

The mechanism of blood concentrations of the Shenqi pill repairing injured epithelial cells of renal tubular in vitro.

In the present study, we investigated the best blood concentration of the mechanism of Shenqi pill repairing the injured epithelial cells of renal tubular in vitro. First, the injured hypoxia/reoxygenation model of rat proximal renal tubular epithelial cell strain (NRK-52E) was established. The animals were divided randomly into control, model, low concentration (5 µg/ml), moderate concentration (10 µg/ml) and high concentration (20 µg/ml) groups. The apoptotic rate was measured with flow cytometry and Jag2/Notch2/hes1 mRNA, and the protein expression was measured for 1, 3 and 7 days. It was found that in comparison to the control group, the growth of each group was prolonged with time, the levels of apoptosis, and the Jag2/Notch2/hes1 mRNA and protein expression decreased. Furthermore, the levels of the apoptotic rate, Jag2/Notch2/hes1 mRNA and protein expression of the moderate concentration and high concentration groups were significantly lower than those of the model and low-dose groups at each time-point (P<0.05). In conclusion, the Shenqi Pill alleviates the damage of renal tubular epithelial cells by inhibiting the Jag2/Notch2/hes1 signaling pathway; suitable concentration such as 10–20 µg/ml can exert protective effect.

TRIP13-deficient tubular epithelial cells are susceptible to apoptosis following acute kidney injury

Damage to renal tubular epithelial cells by genetic, environmental, or biological insults can initiate complex signaling mechanisms that promote kidney repair and functional recovery. In this study, we demonstrated that thyroid receptor interacting protein 13 (TRIP13) is a critical modulator of tubular epithelial cell repair following ischemia‐reperfusion injury (IRI), a common type of renal stressor. In Trip13Gt/Gthypomorph mice treated with unilateral renal IRI, persistent tubular epithelial cell damage was determined in the IRI-treated kidney throughout the 168 hours of experimental period compared to the contralateral kidneys. The damaged epithelial cells were associated with increased levels of DNA damage (ɣH2AX) and apoptotic markers (p53, cleaved caspase-7, and TUNEL-positive cells). Correspondingly, TRIP13 was found to directly interact with Tetratricopeptide Repeat Domain 5 (TTC5), a p53 co‐factor, and genetic knockdown of TRIP13 in murine inner medullary collecting duct cells in the presence of hydrogen peroxide showed increased activity of p53 at Serine 15. In all, these studies suggest that insufficient TRIP13 increased the susceptibility of damaged tubular epithelial cells to progress towards apoptotic cell death.

Protective effects of Genistein on human renal tubular epithelial cells damage of microwave radiation

To observe the effects of microwave irradiation on human proximal renal tubular epithelial cells (HKC) and protec-tive effects of genistein. HKC cells were divided into control group, microwave irradiation group and genistein group(n=6) re-spectively. The genistein group cells were pre-incubated with 30μmol/L genistein in DMEM for 2 hours. After irradiation for 24 hours, the concentrations of lactate dehydrogenase(LDH) and β-N-acetyl glucosaminidase(NAG) in culture solution were measured to evaluate cell injury. Cells were curetted to measure the levels of malondialdehyde (MDA) and superoxide dismutase (SOD). Cell apoptosis and necrosis were de-tected with Hoechst 33258 stain. Compared with the control group, the NAG activity of the microwave irradiation group was signifi-cantly increased(P < 0.01), and NAG activity of genistein pre-incubated group was significantly decreased(P < 0.01). The levels of LDH in microwave irradiation group were also increased significantly (P < 0.01 vs control group). LDH levels could be decreased obviously (P < 0.01 vs microwave irradiation group) after genistein pre-incubate. Hoechst 33258 fluorescent staining revealed that the nucleus crimpled, cres-cent liked and chromatin condensed, even nucleus disintegrated. Our research showed that microwave irradiation could lead to large amount of cell apoptosis and necrosis, and genistein pre-treatment could reduce the ratio of apoptosis and necrosis than that in microwave irradiation group (P < 0.01). The concentration of MDA in microwave irradiation group was higher than that in control group (P < 0.01). At the same time, the activity of SOD was significantly reduced (P < 0.01). Pre-incubated with genistein could not decrease the MDA levels, but could increase the activities of SOD (P < 0.01 vs microwave irradiation group). microwave irradiation can induce human proximal renal tubular epithelial cells injury. The protective effects of genistein may partly correlated with decreasing oxidative stress damage and cell apoptosis in HKC cells.

Stearoyl-CoA Desaturase-1 Protects Cells against Lipotoxicity-Mediated Apoptosis in Proximal Tubular Cells.

Saturated fatty acid (SFA)-related lipotoxicity is a pathogenesis of diabetes-related renal proximal tubular epithelial cell (PTEC) damage, closely associated with a progressive decline in renal function. This study was designed to identify a free fatty acid (FFA) metabolism-related enzyme that can protect PTECs from SFA-related lipotoxicity. Among several enzymes involved in FFA metabolism, we identified stearoyl-CoA desaturase-1 (SCD1), whose expression level significantly decreased in the kidneys of high-fat diet (HFD)-induced diabetic mice, compared with non-diabetic mice. SCD1 is an enzyme that desaturates SFAs, converting them to monounsaturated fatty acids (MUFAs), leading to the formation of neutral lipid droplets. In culture, retrovirus-mediated overexpression of SCD1 or MUFA treatment significantly ameliorated SFA-induced apoptosis in PTECs by enhancing intracellular lipid droplet formation. In contrast, siRNA against SCD1 exacerbated the apoptosis. Both overexpression of SCD1 and MUFA treatment reduced SFA-induced apoptosis via reducing endoplasmic reticulum stress in cultured PTECs. Thus, HFD-induced decrease in renal SCD1 expression may play a pathogenic role in lipotoxicity-induced renal injury, and enhancing SCD1-mediated desaturation of SFA and subsequent formation of neutral lipid droplets may become a promising therapeutic target to reduce SFA-induced lipotoxicity. The present study provides a novel insight into lipotoxicity in the pathogenesis of diabetic nephropathy.

Histopathological Evaluation of Contrast-Induced Acute Kidney Injury Rodent Models.

Contrast-induced acute kidney injury (CI-AKI) can occur in 3–25% of patients receiving radiocontrast material (RCM) despite appropriate preventive measures. Often patients with an atherosclerotic vasculature have to receive large doses of RCM. Thus, animal studies to uncover the exact pathomechanism of CI-AKI are needed. Sensitive and specific histologic end-points are lacking; thus in the present review we summarize the histologic appearance of different rodent models of CI-AKI. Single injection of RCM causes overt renal damage only in rabbits. Rats and mice need an additional insult to the kidney to establish a clinically manifest CI-AKI. In this review we demonstrate that the concentrating ability of the kidney may be responsible for species differences in sensitivity to CI-AKI. The most commonly held theory about the pathomechanism of CI-AKI is tubular cell injury due to medullary hypoxia. Thus, the most common additional insult in rats and mice is some kind of ischemia. The histologic appearance is tubular epithelial cell (TEC) damage; however severe TEC damage is only seen if RCM is combined by additional ischemia. TEC vacuolization is the first sign of CI-AKI, as it is a consequence of RCM pinocytosis and lysosomal fusion; however it is not sensitive as it does not correlate with renal function and is not specific as other forms of TEC damage also cause vacuolization. In conclusion, histopathology alone is insufficient and functional parameters and molecular biomarkers are needed to closely monitor CI-AKI in rodent experiments.

Epidermal growth factor receptor signaling and the progression of diabetic nephropathy.

Epidermal growth factor receptor signaling and the progression of diabetic nephropathy.

Effect of dexmedetomidine on expression of hypoxia-inducible factor-1α during hypoxia/reoxygenation in human renal tubular epithelial cells

Objective To investigate the effect of dexmedetomidine on the expression of hypoxia-inducible factor-1α (HIF-1α) during hypoxia/reoxygenation (H/R) in human renal tubular epithelial cells.Methods Human renal tubular epithelial cells (HK-2 cells) cultured in vitro were randomly divided into 4 groups (n =24 each) using a random number table:control group (group C),dexmedetomidine group (group DEX),H/R group and H/R+ dexmedetomidine group (group H/R + DEX).In group C,the cells were incubated for 28 h in an incubator filled with normoxia at 37 ℃.In group DEX,dexmedetomidine 0.1 nmol/L (final concentration) was added to the culture medium and the cells were incubated for 2 h,and then incubated for 28 h in an incubator filled with normoxia at 37 ℃.In group H/R,the cells were incubated in an anaerobic chamber for 24 h at 37 ℃,and then incubated for 4 h in an incubator filled with normoxia at 37 ℃.In group H/R + DEX,the cells were incubated for 2 h in the culture medium containing dexmedetomidine 0.1 nmol/L (final concentration),incubated in an anaerobic chamber for 24 h at 37 ℃,and then incubated for 4 h in an incubator filled with normoxia at 37 ℃.After treatment in each group,the cell viability was measured by MTT assay,cell apoptosis was measured using flow cytometry,the expression of HIF-1α mRNA was detected using RT-PCR,the expression of HIF-1α and activated caspase-3 protein was detected by Western blot,and the cell growth was observed.The apoptosis rate was calculated.Results Compared with group C,the cell viability was significantly decreased,the apoptosis rate was increased,and the expression of HIF-1α mRNA and protein and activated caspase-3 protein was up-regulated in H/.R and H/R + DEX groups,and no significant change was found in group DEX.Compared with group H/R,the cell viability was significantly increased,the apoptosis rate was decreased,the expression of HIF-1α mRNA and protein was up-regulated,the expression of activated caspase-3 protein was down-regulated,and the cell status was significantly improved in group H/R + DEX.Conclusion The mechanism by which dexmedetomidine attenuates H/ R-induced damage to human renal tubular epithelial cells may be related to up-regulated expression of HIF-1 α and inhibited cell apoptosis. Key words: Dexmedetomidine; Hypoxia-inducible factor 1, alpha subunit; Kidney tubules; Epithelial cells; Reperfusion injury

Detrimental effects of Notch1 signaling activated by cadmium in renal proximal tubular epithelial cells.

We examined the roles of Notch1 signaling and its cross-talk with other signaling pathways, including p53 and phosphatidylinositol-3-kinase (PI3K)/Akt, in cadmium-induced cellular damage in HK-2 human renal proximal tubular epithelial cells. Following exposure to cadmium chloride (CdCl2), the level of Notch intracellular domain (NICD), the cleaved form of the Notch1 receptor, was increased and accumulated in the nuclear fraction. Knockdown of Notch1 with siRNA or treatment with the γ-secretase inhibitor, DAPT (N-[N-(3,5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester), prevented CdCl2-induced morphological change of HK-2 cells and reduction of cell viability. Knockdown of Jagged1 or Jagged2, the ligands of the Notch1 receptor, partially suppressed cadmium cytotoxicity. Inhibition of p53 activity with pifithrin-α or inhibition of PI3K with LY294002 suppressed CdCl2-induced cellular damage and elevation of Notch1-NICD. In addition, treatment with the epidermal growth factor receptor (EGFR) inhibitor, AG1478, and the insulin-like growth factor-1 receptor inhibitor, PPP, suppressed both Notch1-NICD accumulation and Akt phosphorylation in HK-2 cells exposed to CdCl2. However, knockdown of Notch1 did not affect CdCl2-induced p53 accumulation and phosphorylation but suppressed phosphorylation of EGFR, Akt, and p70 S6 kinase. Depletion of Notch1 suppressed CdCl2-induced reduction of E-cadherin expression and elevation of Snail expression. Furthermore, treatment with SB216763, an inhibitor of glycogen synthase kinase-3, suppressed the potency of LY294002 treatment to reduce Snail expression in HK-2 cells exposed to CdCl2. Knockdown of Snail with siRNA partially prevented HK-2 cells from CdCl2-induced reduction of E-cadherin expression and cellular damage. These results suggest that cadmium exposure induces the activation of Notch1 signaling in renal proximal tubular cells with cooperative activation by the p53 and PI3K/Akt signaling pathways; the resultant expression of Snail, a repressor of E-cadherin expression, might lead to cellular damage by decreasing cell–cell adhesion.

Clinical implication of urinary protein markers in diabetic nephropathy and interventional effects of Chinese herbal medicine

In clinic, some urinary protein makers can dynamically and noninvasively reflect the degree of renal tubular injury in patients with diabetic nephropathy (DN). These urinary biomarkers of tubular damage are broadly divided into two categories. One is newfound, including kidney injury molecule-1 (Kim-1), neutrophil getatinase-associated lipocalin (NGAL), liver-type fatty acid-binding protein (L-FABP) and cystatin C (CysC); the other one is classical, including beta2 microglobulin (beta2-MG), retinal binding protein (RBP) and N-acetyl-beta-D-glucosaminidase (NAG). It is reported that, the increases in urinary protein markers are not only closely related to the damage of tubular epithelial cells in DN patients, but also can be ameliorated by the treatment with Chinese herbal compound preparations or Chinese herbal medicine. Recently, although urinary proteomics are used in the protein separation and identification, the traditional associated detection of urinary protein markers is more practical in clinic. At present, it is possible that the associated detection of urinary biomarkers of glomerular and tubular damages may be a feasible measure to reveal the clinical significance of urinary protein markers in DN patients and the interventional effects of Chinese herbal medicine.

Downregulation of uncoupling protein-2 by genipin exacerbates diabetes-induced kidney proximal tubular cells apoptosis

Renal tubular epithelial cell injury is a major pathological event that contributes to the development of diabetic kidney disease (DKD). Uncoupling protein-2 (UCP2), a mitochondrial membrane protein, has been reported to participate in the regulation of reactive oxygen species (ROS) generation, which contributes to tubular cell apoptosis induced by hyperglycemia. In this study, we found that genipin, a UCP2 inhibitor, dramatically boosted oxidative stress, attenuated antioxidative capacity, and exacerbated cell apoptosis accompanied with caspase-3 activation in rat renal proximal tubular cells (NRK-52E) incubated with high glucose. The present study results suggest that manipulation of UCP2 could be important in the prevention of oxidative stress damage in renal tubular epithelial cells induced by hyperglycemia in vitro.

Nuclear factor kappa B in urine sediment: a useful indicator to detect acute kidney injury in Plasmodium falciparum malaria

BackgroundAcute kidney injury (AKI) is one of the major complications of Plasmodium falciparum malaria, especially among non-immune adults. It has recently been revealed that activation of transcription factor nuclear factor kappa B (NF-κB) induces pro-inflammatory gene expression involved in the development of progressive renal inflammatory diseases. The aim of this study was to determine whether urinary sediment NF-κB p65 can act as a biomarker for AKI in patients with P. falciparum malaria.MethodsUrinary sediments from malaria patients, including Plasmodium vivax malaria, uncomplicated P. falciparum malaria, complicated P. falciparum malaria without AKI (serum creatinine-Cr <3 mg/dl) and complicated P. falciparum malaria with AKI (Cr ≥3 mg/dl) were used to determine NF-κB p65 level by sandwich enzyme-linked immunosorbent assay (ELISA). Urinary sediments obtained from healthy controls were used as a normal baseline. Correlations between levels of urinary sediment NF-κB p65 and pertinent clinical data were analysed.ResultsUrinary sediment NF-κB p65 levels were significantly increased on the day of admission (day 0) and on day 7 post-treatment in complicated P. falciparum malaria patients with AKI, compared with those without AKI (p = 0.001, p <0.001, respectively), P. vivax patients (all p <0.001) and healthy controls (all p <0.001). NF-κB p65 levels in urinary sediment cells showed a significant positive correlation with serum Cr (Day 0: rs = 0.792; p <0.001, Day 7: rs = 0.605; p <0.001) and blood urea nitrogen (BUN) (Day 0: rs = 0.839; p <0.001, Day 7: rs = 0.596; p <0.001).ConclusionsUrinary sediment NF-κB p65 level is a useful indicator for estimating renal tubular epithelial cell damage and subsequent development of AKI among patients with P. falciparum malaria.

Obesity-Mediated Autophagy Insufficiency Exacerbates Proteinuria-induced Tubulointerstitial Lesions

Obesity is an independent risk factor for renal dysfunction in patients with CKDs, including diabetic nephropathy, but the mechanism underlying this connection remains unclear. Autophagy is an intracellular degradation system that maintains intracellular homeostasis by removing damaged proteins and organelles, and autophagy insufficiency is associated with the pathogenesis of obesity-related diseases. We therefore examined the role of autophagy in obesity-mediated exacerbation of proteinuria-induced proximal tubular epithelial cell damage in mice and in human renal biopsy specimens. In nonobese mice, overt proteinuria, induced by intraperitoneal free fatty acid-albumin overload, led to mild tubular damage and apoptosis, and activated autophagy in proximal tubules reabsorbing urinary albumin. In contrast, diet-induced obesity suppressed proteinuria-induced autophagy and exacerbated proteinuria-induced tubular cell damage. Proximal tubule-specific autophagy-deficient mice, resulting from an Atg5 gene deletion, subjected to intraperitoneal free fatty acid-albumin overload developed severe proteinuria-induced tubular damage, suggesting that proteinuria-induced autophagy is renoprotective. Mammalian target of rapamycin (mTOR), a potent suppressor of autophagy, was activated in proximal tubules of obese mice, and treatment with an mTOR inhibitor ameliorated obesity-mediated autophagy insufficiency. Furthermore, both mTOR hyperactivation and autophagy suppression were observed in tubular cells of specimens obtained from obese patients with proteinuria. Thus, in addition to enhancing the understanding of obesity-related cell vulnerability in the kidneys, these results suggest that restoring the renoprotective action of autophagy in proximal tubules may improve renal outcomes in obese patients.

Influence of ethylbenzene on oxidative damage and apoptosis in rat renal epithelial cells NRK-52e

To study the oxidative damage and apoptosis of renal tubular epithelial cells (NRK-52e cell line) induced by ethylbenzene. NRK-52e cells were exposed to 30, 60, 90, 120 μmol/L ethylbenzene for 24 hours. Cell viability were measured using MTT, the activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and catalase (CAT), the contents of malondialdehyde (MDA) and glutathione (GSH) were detected respectively. PI fluorescent staining assay was applied to detect percentage of apoptosis in ethylbenzene-treated groups. Compared with control group, cell outline became clear, cell diopter increased, cell became smaller and shrinkage, some cells broke in 60 μmol/L ethylbenzene-treated group. Plenty of cells died, suspension cells increased significantly in 90 μmol/L ethylbenzene-treated group. Compared with control group, cell viability the activities of SOD and CAT and the content of GSH were significantly decreased in 60 and 90 μmol/L ethylbenzene-treated groups (P<0.05). The MDA content were remarkably elevated in 90 μmol/L ethylbenzene-treated groups (P<0.05). Ethylbenzene can induce oxidative stress and apoptosis in NRK-52e cells (P<0.05).

Phenotyping of Nod1/2 double deficient mice and characterization of Nod1/2 in systemic inflammation and associated renal disease

SummaryIt is indispensable to thoroughly characterize each animal model in order to distinguish between primary and secondary effects of genetic changes. The present study analyzed Nod1 and Nod2 double deficient (Nod1/2 DKO) mice under physiological and inflammatory conditions. Nod1 and Nod2 are members of the Nucleotide-binding domain and Leucine-rich repeat containing Receptor (NLR) family. Several inflammatory disorders, such as Crohn's disease and asthma, are linked to genetic changes in either Nod1 or Nod2. These associations suggest that Nod1 and Nod2 play important roles in regulating the immune system.Three-month-old wildtype (Wt) and Nod1/2 DKO mice were sacrificed, body and organ weight were determined, and blood was drawn. Except for lower liver weight in Nod1/2 DKO mice, no differences were found in body/organ weight between both strains. Leukocyte count and composition was comparable. No significant changes in analyzed plasma biochemical markers were found. Additionally, intestinal and vascular permeability was determined. Nod1/2 DKO mice show increased susceptibility for intestinal permeability while vascular permeability was not affected. Next we induced septic shock and organ damage by administering LPS+PGN intraperitoneally to Wt and Nod1/2 DKO mice and sacrificed animals after 2 and 24 hours. The systemic inflammatory and metabolic response was comparable between both strains. However, renal response was different as indicated by partly preserved kidney function and tubular epithelial cell damage in Nod1/2 DKO at 24 hours. Remarkably, renal inflammatory mediators Tnfα, KC and Il-10 were significantly increased in Nod1/2 DKO compared with Wt mice at 2 hours.Systematic analysis of Nod1/2 DKO mice revealed a possible role of Nod1/2 in the development of renal disease during systemic inflammation.