Tubule dysfunction and injury and future risk of sepsis-associated acute kidney injury.

BACKGROUND: Novel biomarkers can quantify both kidney tubule function, including proximal tubule reabsorptive (urine α-1 microglobulin (uα1m)) and tubule protein synthesis capacities (urine uromodulin (uUMOD)), and tubular injury (urine neutrophil gelatinase-associated lipocalin (uNGAL)). In a blood pressure trial, we reported that lower reabsorptive and synthetic protein capacity at times of health predicted future risk of acute kidney injury (AKI), but most AKI was related to hemodynamic causes in this trial. Associations between tubular function and injury and future AKI related to other causes is unknown. MATERIALS AND METHODS: We performed a case-control study in REGARDS, a population-based cohort study, among participants who provided urine at the baseline visit. We matched each septic AKI case by age, sex, race, and time from baseline to hospital admission 1 : 1 to a participant with sepsis who did not develop AKI (controls). Using conditional logistic regression, we evaluated the associations of uα1m, uUMOD, urine ammonium, and uNGAL with septic AKI. RESULTS: Mean age was 69 ± 8 years, 44% were female, and 39% were Black participants. Median baseline eGFR among cases and controls was 73 (55, 90) and 82 (65, 92) mL/min/1.73m2, and median albuminuria was 19 (8, 87) vs. 9 (5, 22) mg/g, respectively. No independent associations were observed between the tubule function or injury markers and subsequent risk of septic AKI once models were adjusted for baseline albuminuria, estimated glomerular filtration rate, and other risk factors. CONCLUSION: Among community participants, tubule function and injury markers at times of health were not independently associated with future risk of septic AKI.

Long Non-Coding RNA Small Nucleolar RNA Host Gene 5 (SNHG5) Regulates Renal Tubular Damage in Diabetic Nephropathy via Targeting MiR-26a-5p.

The study explored the diagnostic value of SNHG5 in diabetic nephropathy (DN) and investigated the role and mechanism on DN via establishing the in vitro HK2 cell model. This study recruited 62 types 2 diabetes mellitus (T2DM) patients, 58 DN patients and 60 healthy controls (HC). The expressions of serum SNHG5 and miR-26a-5p were measured by RT-qPCR analysis. The diagnostic value of SNHG5 in DN was assessed by ROC curve. The in vitro cell model was built to estimate the effects of SNHG5 on cell viability, cell apoptosis, inflammation response and oxidative stress. Serum SNHG5 was increased in DN patients (relative expression: 2.04±0.34) and had the diagnostic value in DN. After HK2 cells were treated with high glucose, the cell viability decreased and apoptosis increased, and the production of inflammatory cytokines and ROS enhanced significantly. It was noticed that inhibition of SNHG5 could reverse the above phenomenon caused by high glucose. Besides, serum miR-26a-5p was diminished in DN patients, and luciferase reporter gene revealed that miR-26a-5p is direct target of SNHG5. These results indicated that inhibition of SNHG5 may mitigate HG-induced renal tubular damage via targeting miR-26a-5p, which providing a new insight into the mechanism of renal tubule damage in DN patients.

CPT1α maintains phenotype of tubules via mitochondrial respiration during kidney injury and repair.

Impaired energy metabolism in proximal tubular epithelial cells (PTECs) is strongly associated with various kidney diseases. Here, we characterized proximal tubular phenotype alternations during kidney injury and repair in a mouse model of folic acid nephropathy, in parallel, identified carnitine palmitoyltransferase 1α (CPT1α) as an energy stress response accompanied by renal tubular dedifferentiation. Genetic ablation of Cpt1α aggravated the tubular injury and interstitial fibrosis and hampered kidney repair indicate that CPT1α is vital for the preservation and recovery of tubular phenotype. Our data showed that the lipid accumulation and mitochondrial mass reduction induced by folic acid were persistent and became progressively more severe in PTECs without CPT1α. Interference of CPT1α reduced capacities of mitochondrial respiration and ATP production in PTECs, and further sensitized cells to folic acid-induced phenotypic changes. On the contrary, overexpression of CPT1α protected mitochondrial respiration and prevented against folic acid-induced tubular cell damage. These findings link CPT1α to intrinsic mechanisms regulating the mitochondrial respiration and phenotype of kidney tubules that may contribute to renal pathology during injury and repair.

Netrin-1 and clusterin: Innovative potential diagnostic biomarkers for early renal damage in ß-thalassemia major children.

BACKGROUND: Children with ß-thalassemia major (ß-TM) suffer from tubular dysfunction even before the onset of any renal impairment symptoms and/or clinical signs. Therefore, identifying innovative biomarkers allowing early renal damage detection has focused attention. AIM: This study aims to preliminary assess Netrin-1(NTN-1) and clusterin (CLU) in ß-TM children and explore their possible roles as surrogate noninvasive biomarkers of renal tubular dysfunction. SUBJECTS AND METHODS: In this study, 40 ß-TM children and 30 healthy children were enrolled. Routine serum and urinary biochemical variables were determined. Urinary NTN-1 and CLU levels were measured using ELISA and their mRNA expression in PBMCs were assayed using real-time PCR. Serum TNF-α, MDA levels and GST activity were measured. RESULTS: Urinary NTN-1 and CLU concentrations and mRNA relative expression levels in PBMCs were significantly increased in ß-TM children relative to controls. Oxidative stress and inflammatory markers revealed significant elevation in ß-TM children compared to controls. The change in these parameters correlated significantly with other renal parameters. ROC curves analysis showed that urinary NTN-1 and CLU levels are of promising diagnostic performance. CONCLUSION: Our results suggest that NTN-1 and CLU are qualified as new noninvasive biomarker panels for early detection of renal injury in ß-TM children. Moreover, urinary NTN-1 is recommended as a precise one during the clinical practices.

PEGylated and Acylated Elabela Analogues Show Enhanced Receptor Binding, Prolonged Stability, and Remedy of Acute Kidney Injury.

Acute kidney injury (AKI), mostly caused by renal ischemia-reperfusion (I/R) injury and nephrotoxins, is characterized by rapid deterioration in renal-functions without effective drug treatment available. Through activation of a G protein-coupled receptor APJ, a furin-cleaved fragment of Elabela (ELA[22-32], E11), an endogenous APJ ligand, protects against renal I/R injury. However, the poor plasma stability and relatively weak APJ-binding ability of E11 limit its application. To address these issues, we rationally designed and synthesized a set of E11 analogues modified by palmitic acid (Pal) or polyethylene glycol; improved plasma stability and APJ-binding capacity of these analogues were achieved. In cultured renal tubular cells, these analogues protected against hypoxia-reperfusion or cisplatin-caused injury. For renal I/R-injured mice, these analogues showed improved reno-protective effects than E11; notably, Pal-E11 showed therapeutic effects at 24 h post I/R injury. These results present ELA analogues as potential therapeutic options in managing AKI.

XJB-5-131 inhibited ferroptosis in tubular epithelial cells after ischemia-reperfusion injury.

Regulated necrosis has been reported to exert an important role in the pathogenesis of various diseases, including renal ischemia-reperfusion (I/R) injury. Damage to renal tubular epithelial cells and subsequent cell death initiate the progression of acute kidney injury (AKI) and subsequent chronic kidney disease (CKD). We found that ferroptosis appeared in tubular epithelial cells (TECs) of various human kidney diseases and the upregulation of tubular proferroptotic gene ACSL4 was correlated with renal function in patients with acute kidney tubular injury. XJB-5-131, which showed high affinity for TECs, attenuated I/R-induced renal injury and inflammation in mice by specifically inhibiting ferroptosis rather than necroptosis and pyroptosis. Single-cell RNA sequencing (scRNA-seq) indicated that ferroptosis-related genes were mainly expressed in tubular epithelial cells after I/R injury, while few necroptosis- and pyroptosis-associated genes were identified to express in this cluster of cell. Taken together, ferroptosis plays an important role in renal tubular injury and the inhibition of ferroptosis by XJB-5-131 is a promising therapeutic strategy for protection against renal tubular cell injury in kidney diseases.

Role of DNA damage in the progress of chronic tubule­interstitial injury.

Tubulointerstitial fibrosis (TIF) is a common final endpoint of chronic allograft nephropathy. Over the years, several hypotheses have been developed to explain the progression of TIF, including mechanisms such as inflammation, epithelial­mesenchymal transition, senescence, chronic hypoxia and reactive oxygen species. Furthermore, TIF is reportedly induced by the 'damage­proliferation­death' cycle. In the present study, an AA renal fibrosis model was established in vitro to investigate whether the vicious proliferation­death cycle is a pathophysiological process of TIF following chronic injury to the kidneys. Results from the present study revealed that cell death was associated with the entrance of cells into the cell cycle. Genetic knockdown of p21 was observed to increase cell cycle progression and the proliferative rate of cells, which overall promoted increased rates of cell death. In addition, the activation of the DNA damage response (DDR) signaling pathway was demonstrated to be crucial to the initiation of the vicious cycle of 'proliferation­death'. Ataxia telangiectasia mutated (ATM) is an important molecule of the DDR and the genetic knockdown of ATM induced apoptosis, increased cell proliferation and promoted cell death. The increase in apoptosis was suggested to be due to the decreased expression levels of p21 observed following the genetic knockdown of ATM. In conclusion, the present study suggested that the crosstalk between the ATM and p21 protein may serve an important role in the regulation of the 'proliferation­death' cycle in the progress of chronic tubulointerstitial injury.

Injured tubular epithelial cells activate fibroblasts to promote kidney fibrosis through miR-150-containing exosomes.

The kidney injury induced by ischemia-reperfusion (IR) usually comes with irreversible renal fibrosis, a process that develops into chronic kidney disease (CKD), but the underlying cellular mechanism has yet to be determined. To test our hypothesis that exosomes are tightly connected with kidney fibrosis following AKI, we studied the role of exosomes and the transfer of specific miRNA among other genetic components in injured tubular epithelial cells (TECs). We utilized an experimental IR mice model to simulate the fibrotic environment in injured tissue and detect the production of exosomes, and found that exosome deficiency could significantly alleviate the degree of kidney fibrosis following IR administration. MiRNA profiling of exosomes extracted from renal tissue samples with or without ischemia-reperfusion injury (IRI) revealed that miR-150 was markedly increased as a compelling profibrotic molecule, as evidenced by the fact that overexpression of miR-150 facilitated renal fibrosis. Exosomes isolated from hypoxia TECs also induced the increased production of miR-150. In cocultured fibroblasts with TECs-derived exosomes, we confirmed a direct uptake of exosomal miR-150 by fibroblasts. Finally, we verified that in vivo ischemia mice pretreated with exosomes enriched in miR-150 developed more profibrotic manifestations. Thus, our current study indicated that TECs have the ability to employ exosomes to initiate the activation and proliferation of fibroblasts via direct shuttling of miR-150-containing exosomes during reparative responses, and that exosome/miR-150 provides the groundwork for research to develop more personalized therapeutic approaches for controlling tissue fibrosis.

Weight-gain induced changes in renal perfusion assessed by contrast-enhanced ultrasound precede increases in urinary protein excretion suggestive of glomerular and tubular injury and normalize after weight-loss in dogs.

Early detection of obesity-related glomerulopathy in humans is challenging as it might not be detected by routine biomarkers of kidney function. This study's aim was to use novel kidney biomarkers and contrast-enhanced ultrasound (CEUS) to evaluate the effect of obesity development and weight-loss on kidney function, perfusion, and injury in dogs. Sixteen healthy lean adult beagles were assigned randomly but age-matched to a control group (CG) (n = 8) fed to maintain a lean body weight (BW) for 83 weeks; or to a weight-change group (WCG) (n = 8) fed the same diet to induce obesity (week 0-47), to maintain stable obese weight (week 47-56) and to lose BW (week 56-83). At 8 time points, values of systolic blood pressure (sBP); serum creatinine (sCr); blood urea nitrogen (BUN); serum cystatin C (sCysC); urine protein-to-creatinine ratio (UPC); and urinary biomarkers of glomerular and tubular injury were measured. Glomerular filtration rate (GFR) and renal perfusion using CEUS were assayed (except for week 68). For CEUS, intensity- and time-related parameters representing blood volume and velocity were derived from imaging data, respectively. At 12-22% weight-gain, cortical time-to-peak, representing blood velocity, was shorter in the WCG vs. the CG. After 37% weight-gain, sCysC, UPC, glomerular and tubular biomarkers of injury, urinary immunoglobulin G and urinary neutrophil gelatinase-associated lipocalin, respectively, were higher in the WCG. sBP, sCr, BUN and GFR were not significantly different. After 23% weight-loss, all alterations were attenuated. Early weight-gain in dogs induced renal perfusion changes measured with CEUS, without hyperfiltration, preceding increased urinary protein excretion with potential glomerular and tubular injury. The combined use of routine biomarkers of kidney function, CEUS and site-specific urinary biomarkers might be valuable in assessing kidney health of individuals at risk for obesity-related glomerulopathy in a non-invasive manner.

Neutrophil Gelatinase-Associated Lipocalin for the Early Prediction of Acute Kidney Injury in ST-Segment Elevation Myocardial Infarction Patients Treated with Primary Percutaneous Coronary Intervention.

INTRODUCTION AND OBJECTIVE: Neutrophil gelatinase-associated lipocalin (NGAL), a glycoprotein released by renal tubular cells, can be used as a marker of early tubular damage. We evaluated plasma NGAL level utilization for the identification of acute kidney injury (AKI) among ST-elevation myocardial infarction (STEMI) patients undergoing primary coronary intervention (PCI). METHODS: 131 STEMI patients treated with PCI were prospectively included. Plasma NGAL levels were drawn prior to PCI (0 h) and 24 h afterwards. AKI was defined per KDIGO criteria of serum creatinine increase. Receiver-operating characteristic (ROC) methods were used to identify optimal sensitivity and specificity for the observed NGAL range. RESULTS: Overall AKI incidence was 14%. NGAL levels were significantly higher for patients with AKI at both 0 h (164 ± 42 vs. 95 ± 30; p < 0.001) and 24 h (142 ± 41 vs. 93 ± 36; p < 0.001). Per ROC curve analysis, an optimal cutoff value of NGAL (>120 ng/mL) predicted AKI with 80% sensitivity and specificity (AUC 0.881, 95%, CI 0.801-0.961, p < 0.001). In a multivariate logistic regression model, NGAL levels were independently associated with AKI at 0 h (OR 1.044, 95% CI 1.013-1.076; p = 0.005) and 24 h (OR 1.018, 95% CI 1.001-1.036; p = 0.04). CONCLUSIONS: Elevated NGAL levels, suggesting renal tubular damage, are independently associated with AKI in STEMI patients undergoing primary PCI.

Downregulation of MiR-218 can alleviate high-glucose-induced renal proximal tubule injury by targeting GPRC5A.

The purpose of this study was to explore the functional implication of microRNA-218 (miR-218) in diabetic nephropathy (DN) through high-glucose-stimulated renal proximal tubule impairment. Biological function experiments showed that miR-218 and inflammatory factors TNF-α and IL-1ß were highly expressed in renal proximal tubule under high-glucose conditions. Inhibiting miR-218 alleviated renal tubular cell injury, which was represented by miR-218 inhibitor facilitating renal tubular cell vitality whilst reducing its apoptosis and levels of inflammation factors. In addition, we confirmed that miR-218 directly targeted GPRC5A and negatively regulated its expression. Co-transfection assay showed that overexpression of GPRC5A accentuated the mitigated action of miR-218 inhibitor on renal proximal tubule cell injury induced by high-glucose. Accordingly, these data indicated that downregulation of miR-218 can assuage high-glucose-resulted renal tubular cell damage, and its ameliorative effect was achieved by negative regulation of GPRC5A, which provides a novel direction for unearthing the pathogenesis and even further biological treatment of DN.

Cabin1 involves in renal tubular epithelial cells mitochondrial dysfunction through SIRT1/p53 pathway.

Background: Angiotensin II (AngII) induced Calcineurin binding protein 1 (Cabin1) protein expression significantly increased during Renal tubular epithelial cells (RTEC) injury. However, the detailed function of Cabin1 protein in RTEC was not characterized well. In this study, we aimed to explore the downstream target of Cabin1 in vitro model.Methods: Rat kidney epithelial cells were cultured and stimulated with AngII. Electron microscopy was performed to observe mitochondrial morphology change. Immunofluorescence staining was detected to observe the distribution of cytoskeleton and Cabin1. Mitochondrial morphology change and protein expression were detected by electrical microscopy and western blot.Results: AngII induced the disruption of cytoskeleton at 24 and 48 h. Western blot analysis showed AngII significantly induced the overexpression of Cabin1. AngII induced a great deal of small, long and irregular mitochondria in RTEC, aspect ratio which reflects the length-to-width ratio of mitochondria remarkably increased at 12 and 24 h. Knocking down Cabin1 aggravated mitochondrial morphological abnormality in AngII treated RTEC. In comparison with control, Cabin1, p53 and cyto C level were significantly increased in AngII treated cells, while SIRT1 level was obviously decreased. Knocked down Cabin1 plus AngII stimulated, SIRT1 was further decreased, while p53 and cyto C were significantly increased.Conclusions: Cabin1 involves in RTEC mitochondrial dysfunction through SIRT1/p53 pathway. Cabin1 may be used as a new marker for the mechanisms of RTEC injury.

Sirt6 attenuates hypoxia-induced tubular epithelial cell injury via targeting G2/M phase arrest.

Acute kidney injury (AKI) is a condition that has a high incidence and death rate. Unfortunately, the kidney may not recover completely after AKI, which then develops to chronic kidney disease (CKD). Therefore, it is necessary to identify potential curative targets to avoid its development to CKD. As an NAD+ -dependent deacetylase, sirtuin 6 (Sirt6) has been linked to different types of biological processes. In the present work, our group investigated the role of Sirt6 in tubular epithelial cells (TECs) under hypoxic stress. Sirt6 expression was examined in mouse kidney following ischemia/reperfusion (IR) injury and hypoxia-challenged TECs. Using Sirt6 plasmid and small interfering RNA, we also investigated how, in regard to inflammation and epithelial-to-mesenchymal transition, Sirt6 affects hypoxia-triggered injury. In addition, cell cycle was detected in hypoxia-challenged TECs. Sirt6 was downregulated in the kidney of mice with IR injury and hypoxia-challenged TECs. Consequently, Sirt6 depletion aggravated hypoxia-induced injury and G2/M phase arrest. Sirt6 overexpression attenuated hypoxia-triggered damage and G2/M phase arrest in TECs. Sirt6 prevented hypoxia-triggered TEC damage via suppressing G2/M phase arrest. Thus, Sirt6 is a possible candidate for alleviating the effects of kidney injury.

SGK1 Attenuates Oxidative Stress-Induced Renal Tubular Epithelial Cell Injury by Regulating Mitochondrial Function.

Mitochondrial dysfunction has been implicated in the early stages or progression of many renal diseases. Improving mitochondrial function and homeostasis has the potential to protect renal function. Serum- and glucocorticoid-induced kinase 1 (SGK1) is known to regulate various cellular processes, including cell survival. In this study, we intend to demonstrate the effect and molecular mechanisms of SGK1 in renal tubular cells upon oxidative stress injury and to determine whether regulation of mitochondrial function is implicated in this process. HK-2 cells were exposed to H2O2, and cell viability and apoptosis were dynamically detected by the CCK-8 assay and annexin-V/PI staining. The concentrations of cellular reactive oxygen species (ROS) and adenosine triphosphate (ATP) and the expression of the SGK1/GSK3ß/PGC-1α signaling pathway were analyzed by flow cytometry or western blot. In addition, shRNA targeting SGK1 and SB216763 were added into the culture medium before H2O2 exposure to downregulate SGK1 and GSK3ß, respectively. Cell viability and mitochondrial functions, including mitochondrial membrane potential (Δψm), Cytochrome C release, mtDNA copy number, and mitochondrial biogenesis, were examined. Protein levels and SGK1 activation were significantly stimulated by H2O2 exposure. HK-2 cells with SGK1 inhibition were much more sensitive to H2O2-induced oxidative stress injury than control group cells, as they exhibited increased apoptotic cell death and mitochondrial dysfunction involving the deterioration of cellular ATP production, ROS accumulation, mitochondrial membrane potential reduction, and release of Cytochrome C into the cytoplasm. Studies on SGK1 knockdown also indicated that SGK1 is required for the induction of proteins associated with mitochondrial biogenesis, including PGC-1α, NRF-1, and TFAM. Moreover, the deleterious effects of SGK1 suppression on cell apoptosis and mitochondrial function, including mitochondrial biogenesis, were related to the phosphorylation of GSK3ß and partially reversed by SB216763 treatment. H2O2 leads to SGK1 overexpression in HK-2 cells, which protects human renal tubule cells from oxidative stress injury by improving mitochondrial function and inactivating GSK3ß.

Monoclonal Gammopathy of Renal Significance and its Associated Experimental Models.

Monoclonal gammopathy of renal significance (MGRS) is a state of circulating monoclonal immunoglobulin (Ig) and light chains that cause kidney injury without definite evidence of multiple myeloma (MM). Although chemotherapy is used to treat many variants of MGRS and has been recently recommended, relatively limited clinical validation studies are available. A few transgenic models of MM reveal renal deposition of monoclonal Ig and light chains. We have demonstrated that the XBP1s-transgenic mouse model from early plasma cell dyscrasia to MM reveals monoclonal IgG/kappa deposition at the subendothelial spaces of the glomeruli, mimicking proliferative glomerulonephritis with monoclonal immunoglobulin deposits. Inhibition of a key immune-modulator, gp96/grp94, genetically or pharmacologically results in a significant reduction of plasma cells within the bone marrow and reduced renal deposition of monoclonal IgG and kappa light chain. This article will review the emerging role of in vitro and animal models from plasma cell dyscrasia to MM in understanding the renal deposition of monoclonal Ig and light chains, along with its potential treatment strategies.

Inhibition of xanthine oxidoreductase protects against contrast-induced renal tubular injury by activating adenosine monophosphate-activated protein kinase.

Reactive oxygen species (ROS) play a pivotal role in the development of contrast-induced nephropathy (CIN). The inhibition of xanthine oxidoreductase is known to reduce levels of ROS. We investigated whether febuxostat could attenuate oxidative stress via the activation of adenosine monophosphate-activated protein kinase (AMPK) against CIN. In a mouse model of CIN, renal impairment and tubular injury substantially increased, whereas febuxostat attenuated renal injury. Plasma and kidney xanthine oxidoreductase levels were decreased by febuxostat. Febuxostat administration was accompanied by the upregulation of AMPK phosphorylation and the inhibition of nicotinamide-adenine dinucleotide phosphate oxidase (Nox)1 and Nox2, followed by the inhibition of hypoxia-inducible factor-1α (HIF-1α) and heme oxygenase-1 expressions and the suppression of transcription factor forkhead box O (FoxO)1 and FoxO3a phosphorylation. Cell survival was significantly reduced after iohexol administration and febuxostat ameliorated iohexol-induced cell death in proximal tubular (HK-2) cells. Furthermore, febuxostat enhanced AMPK phosphorylation and inhibited Nox1, Nox2, and HIF-1α expression in iohexol-exposed HK-2 cells. Finally, these processes decrease ROS in both in vivo and in vitro models of CIN. AMPK inhibition using small interfering RNA blunted the antioxidative effects of febuxostat in iohexol-treated HK-2 cells. Febuxostat attenuated CIN by modulating oxidative stress through AMPK-NADPH oxidase-HIF-1α signaling.

Effect of M2 Macrophages on Injury and Apoptosis of Renal Tubular Epithelial Cells Induced by Calcium Oxalate Crystals.

BACKGROUND: M2 macrophages have important roles in diseases such as tumours, cardiovascular diseases and renal diseases. This study aimed to determine the effects and protective mechanism of M2 macrophages against oxidative stress injury and apoptosis induced by calcium oxalate crystals (CaOx) in renal tubular epithelial cells (HK-2) under coculture conditions. METHODS: THP-1 cells were induced to differentiate into M2 macrophages by using phorbol-12-myristate-13-acetate, IL-4 and IL-13. Morphological features were observed by microscopy. Phenotypic markers were identified by reverse transcription-polymerase chain reaction, Western blot and enzyme-linked immunosorbent assay (ELISA). HK-2 cells were treated with 0.5 mg/mL CaOx crystals and co-cultured with M2 macrophages or apocynin. The viability of HK-2 cells was detected by CCK-8 assay. The lactate dehydrogenase (LDH) activity of HK-2 cells was analysed using a microplate reader. The apoptosis of HK-2 cells was examined by flow cytometry and Hoechst 33258 staining. Reactive oxygen species (ROS) expression and mitochondrial membrane potential in HK-2 cells were detected by a fluorescence microplate reader. Western blot analysis was conducted to detect the expression of p47phox, Bcl-2, cleaved caspase-3, cytochrome c, p38 MAPK, phospho-p38 MAPK, Akt and phospho-Akt. RESULTS: The results of morphology, reverse transcription-polymerase chain reaction, Western blot and ELISA showed that THP-1 cells were successfully polarised to M2 macrophages. The results of co-culture suggested that M2 macrophages or apocynin significantly increased the cell viability and decreased the LDH activity and apoptosis rate after HK-2 cells were challenged with CaOx crystals. The expression of the p47phox protein and the concentration of ROS were reduced, the release of mitochondrial membrane potential and the expression of the Bcl-2 protein were upregulated and the protein expression of cleaved caspase-3 and cytochrome c was downregulated. The expression of the phosphorylated form of p38 MAPK increased. Under coculture conditions with M2 macrophages, the Akt protein of HK-2 cells treated with CaOx crystals was dephosphorylated, but the phosphorylated form of Akt was not reduced by apocynin. CONCLUSIONS: M2 macrophages reduced the oxidative stress injury and apoptosis of HK-2 cells by downregulating the activation of NADPH oxidase, reducing the production of ROS, inhibiting the phosphorylation of p38 MAPK and enhancing the phosphorylation of Akt. We have revealed one of the possible mechanisms by which M2 macrophages reduce the formation of kidney stones.

Induction of Fibrosis and Autophagy in Kidney Cells by Vinyl Chloride.

Vinyl chloride (VC) is a noninfective occupational risk factor. It is found in industrial chemicals, volatile organic compounds, cigarette smoke ingredients, etc. It is a kind of toxic gas that causes many diseases. VC exposure causes an increased risk of liver fibrosis and can result in angiosarcoma of the liver. Previous studies have shown that high-doses of VC exposure in mice resulted in acute death with marked tubular necrosis of the renal cortex. In this study, we assessed the nephrotoxicity of VC in vitro and in vivo. As a result, we demonstrated that VC induced fibrosis-associated protein expression, such as connective tissue growth factor (CTGF), plasminogen activator inhibitor-1 (PAI-1) and collagen 1, and autophagy-associated protein expression, such as Beclin 1 and LC3-II, in kidney cells. The beclin1 siRNA experiments found that autophagy inhibited VC-induced fibrosis. Blood urea nitrogen (BUN) and creatinine levels were increased after VC treatment. Furthermore, VC caused glomerulosclerosis and tubular injury in mouse kidney tissues. Kidney tissue sections showed that VC induced fibrosis and autophagy in mouse kidney tissues. In summary, the results of VC-induced fibrosis suggest that autophagy plays an important role in kidney damage. VC may cause nephrotoxicity, and the results illustrate the importance of considering the toxicological hazards of VC in kidney cells.

The Mitochondria-Targeted Metabolic Tubular Injury in Diabetic Kidney Disease.

BACKGROUND/AIMS: Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease (ESRD) worldwide, and the importance of tubular injury has been highlighted in recent years. However, the underlying mechanisms and effective therapeutic targets are still unclear. In this study, we investigated mtDNA, mitochondrial dynamics, function and metabolic pathways to determine if mitochondrial damage plays a critical role in the development of tubular injury in DKD patients. METHODS: A cross-sectional study was carried out among healthy controls (HCs, n = 65), diabetes patients without kidney disease (DCs, n = 48) and DKD patients (n = 60). Serum, peripheral blood mononuclear cells (PBMCs) and kidney biopsy specimens were obtained from participants. Metabolomics was employed to investigate cellular metabolism. RESULTS: DKD patients had decreased mtDNA copy numbers and increased mtDNA damage compared to DCs. Mitochondrial fragmentation was specifically presented in tubules, but not in podocytes of DKD patients. The accumulation of damaged mtDNA and fragmented mitochondria resulted in increased reactive oxygen species (ROS) generation, activation of apoptosis and loss of mitochondrial membrane potential (ΔΨm) in tubules and PBMCs. Furthermore, glycolysis and tricarboxylic acid (TCA) cycle was perturbed, and increased dihydroxyacetone phosphate (DHAP) and decreased succinyl-CoA synthetase (SCS) respectively in these two metabolic pathways were identified as potential biomarkers for tubular injury in DKD. CONCLUSION: Our study indicates that mitochondrial damage could be the hallmark of tubular injury in DKD patients, and this would provide a novel and attractive therapeutic target to improve this disease.

The role of nuclear factor erythroid-2-related factor 2 expression in radiocontrast-induced nephropathy.

Radiocontrast-induced nephropathy (CIN) is the third most common cause of acute renal failure. The pathophysiology of CIN is related to tubular injury caused by oxidative stress, and nuclear factor erythroid-2-related factor 2 (Nrf2) is critical in coordinating intracellular antioxidative processes. We thus investigated the role of Nrf2 in CIN. CIN was established in mice and in NRK-52E cells via iohexol administration according to the protocols of previous studies. To determine the role of Nrf2 in CIN, Nrf2 expression was reduced in vivo using Nrf2 knockout (KO) mice (B6.129 × 1-Nfe2 l2tm1Ywk/J) and in vitro with siRNA treatment targeting Nrf2. Increased Nrf2 expression was observed after iohexol treatment both in vivo and in vitro. Serum creatinine at 24 h after iohexol injection was significantly higher in KO mice than in wild-type (WT) mice. Histologic examination showed that iohexol-induced tubular vacuolization and structural disruption were aggravated in Nrf2 KO mice. Significant increases in apoptosis and F4/80(+) inflammatory cell infiltration were demonstrated in KO mice compared to WT mice. In addition, the increase in reactive oxygen species after iohexol treatment was augmented by Nrf2 inhibition both in vivo and in vitro. Nrf2 may be implicated in the pathogenesis of CIN via the modulation of antioxidant, anti-apoptotic, and anti-inflammatory processes.