Apoptosis Of Renal Epithelial Cells Research Articles

Transcription factor FoxM1 promotes cyst growth in PKD1 mutant ADPKD.

Autosomal dominant polycystic kidney disease (ADPKD) is driven by mutations in the PKD1 and PKD2 genes, and it is characterized by renal cyst formation, inflammation and fibrosis. Forkhead box protein M1 (FoxM1), a transcription factor of the Forkhead box (Fox) protein super family, has been reported to promote tumor formation, inflammation and fibrosis in many organs. However, the role and mechanism of FoxM1 in regulation of ADPKD progression is still poorly understood. Here, we show that FoxM1 is an important regulator of cyst growth in ADPKD. FoxM1 is upregulated in cyst-lining epithelial cells in Pkd1 mutant mouse kidneys and human ADPKD kidneys. FoxM1 promotes cystic renal epithelial cell proliferation by increasing the expression of Akt and Stat3 and the activation of ERK and Rb. FoxM1 also regulates cystic renal epithelial cell apoptosis through NF-κB signaling pathways. In addition, FoxM1 regulates the recruitment and retention of macrophages in Pkd1 mutant mouse kidneys, a process that is associated with FoxM1-mediated upregulation of monocyte chemotactic protein 1. Targeting FoxM1 with its specific inhibitor, FDI-6, delays cyst growth in rapidly progressing and slowly progressing Pkd1 mutant mouse kidneys. This study suggests that FoxM1 is a central and upstream regulator of ADPKD pathogenesis and provides a rationale for targeting FoxM1 as a therapeutic strategy for ADPKD treatment.

RORA alleviates LPS-induced apoptosis of renal epithelial cells by promoting PGC-1α transcription

ObjectiveTo explore the effect of RORA on LPS-induced renal epithelial cell apoptosis and the underlying mechanism.MethodsLPS-treated HK-2 cells were established as a cellular model of acute kidney injury. The expression of RORA or/and PGC-1α in LPS-induced HK-2 cells was altered by transfection. qRT-PCR and Western blotting were used to detect the expression changes of RORA and PGC-1α. ELISA was performed to detect the expression of IL-1β and IL-6 and the activity of caspase-3. Western blotting was applied for visualization of cleaved caspase-3. CCK-8 and flow cytometry were used to assess cell proliferation and apoptosis. Dual-luciferase reporter and ChIP-qPCR were utilized to verify the binding of RORA to PGC-1α promoter.ResultsLPS treatment decreased the expression of RORA and PGC-1α and increased that of cleaved caspase-3 in HK-2 cells. Also, LPS treatment inhibited HK-2 cell proliferation and promoted HK-2 cell apoptosis and secretion of IL-1β and IL-6. Overexpression of RORA or PGC-1α eliminated the adverse effects of LPS treatment in HK-2 cells. RORA drove the transcription of PGC-1α by binding PGC-1α promoter. Knockdown of PGC-1α offset the reduction in HK-2 cell injury caused by overexpression of RORA.ConclusionRORA reduces LPS-induced apoptosis of renal epithelial cells by promoting PGC-1α transcription.

Super-resolution microscopy reveals that Na+/K+-ATPase signaling protects against glucose-induced apoptosis by deactivating Bad

Activation of the apoptotic pathway is a major cause of progressive loss of function in chronic diseases such as neurodegenerative and diabetic kidney diseases. There is an unmet need for an anti-apoptotic drug that acts in the early stage of the apoptotic process. The multifunctional protein Na+,K+-ATPase has, in addition to its role as a transporter, a signaling function that is activated by its ligand, the cardiotonic steroid ouabain. Several lines of evidence suggest that sub-saturating concentrations of ouabain protect against apoptosis of renal epithelial cells, a common complication and major cause of death in diabetic patients. Here, we induced apoptosis in primary rat renal epithelial cells by exposing them to an elevated glucose concentration (20 mM) and visualized the early steps in the apoptotic process using super-resolution microscopy. Treatment with 10 nM ouabain interfered with the onset of the apoptotic process by inhibiting the activation of the BH3-only protein Bad and its translocation to mitochondria. This occurred before the pro-apoptotic protein Bax had been recruited to mitochondria. Two ouabain regulated and Akt activating Ca2+/calmodulin-dependent kinases were found to play an essential role in the ouabain anti-apoptotic effect. Our results set the stage for further exploration of ouabain as an anti-apoptotic drug in diabetic kidney disease as well as in other chronic diseases associated with excessive apoptosis.

LncRNA OIP5-AS1 reduces renal epithelial cell apoptosis in cisplatin-induced AKI by regulating the miR-144-5p/PKM2 axis

BackgroundThe abnormal expression of long non-coding RNA (lncRNA) Opa-interacting protein 5 antisense RNA 1 (OIP5-AS1) has been observed in many human cancers and the underlying mechanisms have been well studied. However, the function of OIP5-AS1 in acute kidney injury (AKI) remains unclear. MethodsTo explore the role of OIP5-AS1 in the progression of AKI, the cisplatin-induced AKI mouse and cell model were established. To confirm the potential protective effect of OIP5-AS1 during cisplatin-induced AKI, rescue experiments were performed. Targetscan was used to predict the potential targets of miR-144-5p. To further determine whether the effect of miR-144-5p during cisplatin-induced AKI was mediated by PMK2, the recuse experiments using PMK2 overexpressing vector was applied. ResultsOIP5-AS1 was significantly downregulated both in cisplatin-induced AKI mice and human renal tubular cell line HK-2 cells. Moreover, overexpression of OIP5-AS1 efficiently promoted cell growth and reduced cisplatin-induced apoptosis of HK-2 cells. Furthermore, OIP5-AS1 was identified as a sponge of miR-144-5p, and upregulation of miR-144-5p could significantly reverse overexpression of OIP5-AS1-induced protective effect on the damage of cisplatin to HK-2 cells. In addition, pyruvate kinase M2 (PKM2) was found to be a direct target of miR-144-5p, and overexpression of PKM2 efficiently reversed the effect of miR-144-5p mimics on the damage in cisplatin-stimulated HK-2 cells. ConclusionsOIP5-AS1 reduced the apoptosis of cisplatin-stimulated renal epithelial cells by targeting the miR-144-5p/PKM2 axis, which extended the regulatory network of lncRNAs in cisplatin-induced AKI and also provided a novel therapeutic target for AKI treatment.

Long Noncoding RNA PRNCR1 Reduces Renal Epithelial Cell Apoptosis in Cisplatin-Induced AKI by Regulating miR-182-5p/EZH1

Background/Aims: This study was designed to examine the role of long noncoding RNA PRNCR1 in cisplatin-induced acute kidney injury (AKI) in vitro and in vivo. Methods: The expression levels of PRNCR1 and miR-182-5p in cisplatin-induced AKI mice were examined. HK-2 cells were treated with cisplatin to induce cell damage. Then, the effects of PRNCR1 and miR-182-5p on cisplatin-stimulated HK-2 cell viability and apoptosis were detected by the CCK-8 and annexin V-FITC/PI method. Target genes of PRNCR1 and miR-182-5p were analyzed by bioinformatics analysis and luciferase. Results: The expression level of PRNCR1 was significantly reduced in cisplatin-induced AKI mice. In addition, overexpression of PRNCR1 attenuated the damage of cisplatin to HK-2. The expression level of miR-182-5p was significantly raised in cisplatin-induced AKI mice. MiR-182-5p was negatively regulated by PRNCR1 and leaded to an upregulation of EZH1 expression. Overexpression of PRNCR1 attenuated cisplatin-induced apoptosis by downregulating the miR-182-5p/EZH1 axis. Conclusion: LncPRNCR1 reduced the apoptosis of renal epithelial cells induced by cisplatin by modulating miR-182-5p/EZH1.

The Role of Cell-Free Plasma DNA in Patients with Cardiorenal Syndrome Type 1

Background: Recent research highlighted the potential role of circulating cell-free DNA (cfDNA), resulted by apoptosis or cell necrosis, as a prognostic marker in the setting of different clinical conditions. Cardiorenal syndrome type 1 (CRS type 1) is characterized by a rapid worsening of cardiac function leading to acute kidney injury (AKI). Apoptosis of renal epithelial cells is proposed as a mechanism involved in CRS type 1. In this study, we investigated cfDNA levels in patients with acute heart failure (AHF) and CRS type 1 and the possible correlation between cfDNA levels and inflammatory and apoptotic parameters. Methods: We enrolled 17 AHF patients and 15 CRS type 1 who exhibited AKI at the time of admission (caused by AHF) or developed AKI during the first 48 h from admission. cfDNA was extracted from plasma and quantified by real-time polymerase chain reaction. Plasma levels of NGAL, tumor necrosis factor-α, interleukin (IL)-6, IL-18, and caspase-3 were measured. Results: We observed significantly higher levels of cfDNA in patients with CRS type 1 than patients with AHF. Caspase-3, IL-6, IL-18, and NGAL levels resulted significantly increased in patients with CRS type 1. Moreover, a positive correlation between cfDNA levels and caspase-3 levels was found, as well as between cfDNA levels and IL-6 and renal parameters. Conclusion: Our study explores the premise of cfDNA as a marker for apoptosis and inflammation in CRS type 1 patients. cfDNA could potentially serve as an index for noninvasive monitoring of tissue damage and apoptosis in patients with AKI induced by AHF.

AMP-activated protein kinase contributes to cisplatin-induced renal epithelial cell apoptosis and acute kidney injury.

Cisplatin, a commonly used anticancer drug, has been shown to induce acute kidney injury, which limits its clinical use in cancer treatment. Emerging evidence has suggested that AMP-activated protein kinase (AMPK), which functions as a cellular energy sensor, is activated by various cellular stresses that deplete cellular ATP. However, the potential role of AMPK in cisplatin-induced apoptosis of renal tubular epithelial cells has not been studied. In this study, we demonstrated that cisplatin activates AMPK (Thr172 phosphorylation) in cultured renal tubular epithelial cells in a time-dependent manner, which was associated with p53 phosphorylation. Compound C, a selective AMPK inhibitor, suppressed cisplatin-induced AMPK activation, p53 phosphorylation, Bax induction, and caspase 3 activation. Furthermore, silencing AMPK expression by siRNA attenuated cisplatin-induced p53 phosphorylation, Bax induction, and caspase 3 activation. In a mouse model of cisplatin-induced kidney injury, compound C inhibited p53 phosphorylation, Bax expression, caspase 3 activation, and apoptosis, protecting the kidney from injury and dysfunction. Taken together, these results suggest that the AMPK-p53-Bax signaling pathway plays a crucial role in cisplatin-induced tubular epithelial cell apoptosis.

Macelignan protects against renal ischemia-reperfusion injury via inhibition of inflammation and apoptosis of renal epithelial cells

Ischemia-reperfusion injury(IRI) refers to tissuedamagethat occurs when blood supply returns to tissue after a period of ischemia,anoxia or hypoxia. It occurs frequently during shock, organ transplantation and heart failure. It can cause impairment or even renal failure. Macelignan is a lignin isolated from the seeds of Myristica fragrans. It has been reported to inhibit neuroinflammation and oxidative toxicity. The preventive or therapeutic effects of macelignan on renal IRI has not been reported. The present study investigated the effects of macelignan on renal IRI in rats, and the underlying mechanism(s). Healthy adult male Sprague Dawley rats (n = 50) aged 7 - 9 weeks (mean weight = 220 ± 20 g) were used in this study. The rats were randomly assigned to five groups of 10 rats each: sham treated group, IRI group and 40 mg macelignan/kg body weight (bwt) group, 80 mg macelignan/kg bwt group, and 160 mg macelignan/kg bwt group. Ischemia-reperfusion injurywas induced in the rats using standard procedure. The results showed that serum levels of creatinine, blood urea nitrogen (BUN), interleukin-6 (IL-6), tumor necrosis factor α (TNF-α) and gamma interferon (IFN-γ) were significantly higher in IRI group than in sham treated group, but were significantly and dose-dependently reduced after treatment with macelignan (p < 0.05). The activities of catalase and superoxide dismutase (SOD), and reduced glutathione (GSH) level were significantly reduced in IRI group, when compared with sham treated group, but were significantly and dose-dependently increased after treatment with macelignan (p < 0.05). However, the level of malondialdehyde (MDA) was significantly higher in IRI group than in sham treated group, but treatment with macelignan reduced it significantly and dose-dependently (p < 0.05). Macelignan also significantly and dose-dependently inhibited IRI-induced apoptosis in epithelial cells of renal tubules (p < 0.05). The results of Western blotting showed that IRI significantly upregulated the expressions of bax and caspase-3, and down-regulated the expression of bcl-2 in epithelial cells of renal tubules (p < 0.05). However, treatment with macelignan significantly and dose-dependently down-regulated the expressions of bax and caspase-3 in these cells, but significantly and dose-dependently upregulated the expression of bcl-2. These results show that macelignan confers protection on renal IRI via mechanisms involving inhibition of inflammation and apoptosis, and stimulation of natural antioxidant defense system.

Intracellular Reactive Oxygen Species Mediate the Therapeutic Effect of Induced Pluripotent Stem Cells for Acute Kidney Injury.

Aims Treatment for acute kidney injury (AKI) is challenging. Induced pluripotent stem cells (iPSCs) have great therapeutic potential. This study sought to determine whether iPSCs attenuate AKI and the role of reactive oxygen species (ROS). Results We intravenously injected isogenic iPSCs into mice 2 h after renal ischemia-reperfusion injury (IRI). The cells were selectively trafficked to ischemia/reperfusion-injured kidney where they decreased kidney ROS and inflammatory cytokines and improved kidney function and morphology. Pretreating the cells with ROS inhibitors before administration decreased iPSC engraftment and abolished the protective effect of iPSCs. In contrast, pretreating iPSCs with hydrogen peroxide increased iPSC engraftment and therapeutic effect. Although the intravenously administered iPSCs trafficked to the IRI kidney, the cells did not differentiate into proximal or distal tubular epithelial cells. In vitro, the capabilities of the iPSC-released substances to promote proliferation and decrease apoptosis of renal epithelial cells were increased by ROS pretreatment of iPSCs. Moreover, pretreatment of the iPSCs with ROS inhibitor had the opposite effect. Similarly, moderate concentrations of ROS increased while ROS inhibitors decreased iPSC mobility, adhesion to the extracellular matrix, and mitochondrial metabolism. Innovation and Conclusion. iPSCs decreased renal ischemia/reperfusion injury mainly through iPSC-released substances. The therapeutic effect, mitochondrial metabolism, mobility, and kidney trafficking of iPSCs were ROS dependent.

Imoxin attenuates high fructose-induced oxidative stress and apoptosis in renal epithelial cells via downregulation of protein kinase R pathway.

Double-stranded RNA (dsRNA)-activated protein kinase R (PKR), a ubiquitously expressed serine/threonine kinase, is a key inducer of inflammation, insulin resistance, and glucose homeostasis in obesity. Recent studies have demonstrated that PKR can respond to metabolic stress in mice as well as in humans. However, the underlying molecular mechanism is not fully understood. The aim of this study was to examine the effect of high fructose (HF) in cultured renal tubular epithelial cells (NRK-52E) derived from rat kidney and to investigate whether inhibition of PKR could prevent any deleterious effects of HF in these cells. PKR expression was determined by immunofluorescence staining and Western blotting. Oxidative damage and apoptosis were measured by flow cytometry. HF-treated renal cells developed a significant increase in PKR expression. A significant increase in reactive oxygen species generation and apoptosis was also observed in HF-treated cultured renal epithelial cells. All these effects of HF were attenuated by a selective PKR inhibitor, imoxin (C16). In conclusion, our study demonstrates PKR induces oxidative stress and apoptosis, is a significant contributor involved in vascular complications and is a possible mediator of HF-induced hypertension. Inhibition of PKR pathway can be used as a therapeutic strategy for the treatment of cardiovascular and metabolic disorders.

Effect of miR-29a on apoptosis of renal tubular epithelial cells and its mechanism

Objective To investigate the expression of miR-29a in serum of acute kidney injury patients, and whether miR-29a can affect the apoptosis of renal tubular cells by regulating the expressions of phosphatase and tensin homologue deleted on chromosome ten (PTEN). Methods Blood samples were collected from 113 patients with acute kidney injury (AKI) and 110 healthy controls. The expressions of miR-29a in serum were detected by real-fluorescence quantitative polymerase chain reaction (RT-q-PCR). HK-2 cells were cultured in vitro, miR-29a NC or miR-29a mimic was transfected into HK-2 cells, and apoptosis of HK-2 cells was induced by transforming growth factor-β1 (TGF-β1). Flow cytometry was used to investigate the apoptosis of HK-2 cells. Bioinformatics was used to predict potential target genes of miR-29a. Luciferase reporter gene test was used to verify the complementary pairing relationship between miR-29a and PTEN. The expression of PTEN was detected by Western blot. After transfection of plasmids overexpressing PTEN, the apoptosis of HK-2 cells was detected by flow cytometry. Results Compared to the healthy control group, the serum expression of miR-29a was decreased in AKI patients (P<0.05). Flow cytometry results showed that transfection of miR-29a mimic was significantly decreased HK-2 cells apoptosis compared to miR-29 NC group (P<0.05). The Luciferase reporter assay results showed that miR-29a and PTEN had complementary relationship. After transfected with overexpression of PTEN, the HK-2 cells apoptosis rate was significantly increased (P<0.05). Conclusions In the serum of AKI patients, the expression of miR-29a was decreased, overexpression of miR-29a inhibited the apoptosis of renal tubular epithelial cells by inhibiting the expression of PTEN protein. Key words: MicroRNAs/ME; Kidney tubules/CY; Epithelial cells/PA; PTEN phosphohydrolase; Apoptosis

Cell adhesion molecule-1 shedding induces apoptosis of renal epithelial cells and exacerbates human nephropathies.

Chronic kidney disease (CKD) is an important problem throughout the world, associated with the increase of blood urea nitrogen (BUN) and serum creatinine (sCre) and with renal tubular injuries. It is crucial to elucidate the molecular mechanisms of renal injuries to identify the new therapeutics and early diagnostic methods. We focused on cell adhesion molecule-1 (CADM1) protein. CADM1, its isoform SP4, is expressed in the epithelial cells of various tissues, including renal distal tubules, localized on the lateral cell membrane, mediates cell-cell adhesion via trans-homophilic binding, and interacts with various proteins. We previously reported that its expression was downregulated by post-proteolytic cleavage (α- and β-shedding) in pulmonary diseases. To investigate whether CADM1 α-shedding occurs in human nephropathies, we performed Western blotting and immunohistochemical analysis of specimens with arterionephrosclerosis (AS) and diabetic nephropathy (DN) from autopsied kidneys. CADM1 α-shedding was induced in AS and DN kidneys and derived from the decrease in full-length CADM1 (FL-CADM1) and increase of the COOH-terminal fragment (α-CTF). In particular, the reduced FL-CADM1 level was correlated with tubular and tubulointerstitial injuries and the increases in BUN and sCre levels. Apoptosis of renal tubular epithelial cells (TECs) was promoted in both nephropathies, and it was significantly correlated with the decrease in the FL-CADM1. Furthermore, FL-CADM1 knockdown by small interfering RNA downregulated anti-apoptotic Bcl-2 protein and promoted apoptosis of cultured renal TECs. The present study suggests that the reduction of FL-CADM1 leads to renal TEC apoptosis and could exacerbate renal tubular and tubulointerstitial injuries, which contribute to the development of CKD.

Caspase-11 promotes cisplatin-induced renal tubular apoptosis through a caspase-3-dependent pathway.

Renal tubular injury is the hallmark of cisplatin-induced nephrotoxicity. Caspase-11, a member of the caspase family, plays an important role in inflammation and cell death. However, its role in cisplatin-induced renal tubular injury remains unclear. In cisplatin-treated mice, caspase-11 expression was significantly elevated and the expression of caspase-11 was mainly located in renal tubule. Inhibition of caspase-11 by small-interference RNA or its inhibitor wedelolactone attenuated cisplatin-induced renal dysfunction and tubular injury. In cultured primary renal tubular epithelial cells, cisplatin significantly promoted the expression and activation of caspase-11. Inhibition of caspase-11 by small-interference RNA reduced cisplatin-induced cell apoptosis. Overexpression of caspase-11 promoted cell apoptosis by activating the caspase-3-related cell apoptosis. Furthermore, coimmunoprecipitation results showed there was a direct interaction between caspase-11 and caspase-3, and the interaction was enhanced by cisplatin. The fluorescence confocal microscopy results showed that caspase-11 and caspase-3 were colocalized in the cytoplasm of renal tubular epithelial cells. These results demonstrate that caspase-11 plays an important role in cisplatin-induced renal tubular injury. Caspase-11 promotes renal epithelial cell apoptosis by activating the caspase-3-dependent apoptotic pathway. Caspase-11 might be a potential target for therapeutic treatment against cisplatin-induced nephrotoxicity.

Star Fruit Intoxication

Now a day Acute Kidney Injury and Chronic Kidney Disease are the two common problems worldwide with increased mortality and morbidity as well as health care cost. Nephrotoxicity and neurotoxicity by the Star fruit, sometimes fatal, is far more frequent than reported. Star fruit belongs to the Oxalidaceae family, species Averrhoea carambola, is a popular fruit among Orientals. The aim of the study is to create awareness among the population regarding star fruit intoxication, to reduce the development of Acute Kidney Injury in general population, to reduce the mortality and burden of morbidity from chronic renal failure, to reduce the mortality of the patients of End stage renal disease taking regular haemodialysis and to reduce the mortality and burden of morbidity of the people suffering from Diabetes mellitus and Hypertension. Star fruit juice produces acute renal injury not only through the obstructive effect of calcium oxalate crystals, but also inducing apoptosis of renal epithelial cells, which may be caused by the levels of oxalate in the fruit. There have been reports of hiccup, confusion and occasional fatal outcomes in uremic patients after ingestion of star fruit. An excitatory neurotoxin from star fruit has been implicated although the exact nature of this toxic substance has not been identified. High quantities of oxalate in empty stomach and dehydrated state may pose an additional risk for development of renal injury. To avoid acute oxalate nephropathy, pure sour carambola juice or mild diluted post pickled juice should not be consumed in large amounts, especially in an empty stomach and dehydrated state. It is recommended that uremic patients should totally abstain from star fruit due the rare but potentially fatal complications.Anwer Khan Modern Medical College Journal Vol. 6, No. 2: July 2015, P 30-33

Puerarin attenuates renal fibrosis by reducing oxidative stress induced-epithelial cell apoptosis via MAPK signal pathways in vivo and in vitro

Puerarin (PR) is an isoflavonoid isolated from the root of the plant Pueraria lobata and has been widely used in traditional Chinese herbal medicine for the treatment of various diseases. Oxidative stress and epithelial cell apoptosis play important roles in the renal fibrotic process. The present study aimed to determine whether or not PR inhibits renal fibrosis by reducing oxidative stress induced-epithelial cell apoptosis. In vivo, unilateral ureteral obstruction (UUO) induced renal fibrosis, and epithelial cell apoptosis. A total of 24 mice were randomly assigned to four experimental groups: sham, UUO alone, UUO +50 mg/kg PR, and UUO +100 mg/kg PR. In vitro, 200 μM hydrogen peroxide (H2O2) induced epithelial cell apoptosis. The experiments were dived into four groups: control, H2O2 alone, H2O2+50 μM PR, and H2O2+100 μM PR. Tubular injury was measured in the renal cortex of the mice through periodic acid-Schiff (PAS) staining, and the extracellular matrix (ECM) was measured through Sirius red (SR), immunohistochemistry (IHC) staining, and Western blot. Renal epithelial cell apoptosis was measured through terminal deoxynucleotidyl transferase-mediated dUTP Nick-End labeling (TUNEL), flow cytometry (FCM), and Hoechst assays. The protein expression of NOX4, caspase3, ERK, P38, and JNK was assessed through Western blot. PAS staining showed that PR decreased renal tubular injury in UUO mice. SR and IHC staining demonstrated that PR decreased the accumulation of ECM. PR treatment significantly inhibited epithelial cell apoptosis according to the results of TUNEL, FCM, Hoechst, and Western blot. Furthermore, NOX4 increased in UUO mice and decreased with PR treatment. H2O2-derived oxidative stress activated epithelial apoptosis and mitogen-activated protein kinases (MAPK), and PR treatment significantly reversed it. These results suggest that PR treatment ameliorates renal fibrosis by inhibiting oxidative stress induced-epithelial cell apoptosis through MAPK signaling.

Upregulation of AMWAP: a novel mechanism for HDAC inhibitors to protect against cisplatin nephrotoxicity

Histone deacetylases have been reported to protect against renal tubular damage in several animal models of acute renal injury, including cisplatin nephrotoxicity. However, the mechanism involved is not well defined. In this study, Ranganathan et al. identify activated microglia/macrophage WAP domain protein as the novel mediator of histone deacetylase inhibitor-mediated renal protection in a murine model of cisplatin nephrotoxicity. Activated microglia/macrophage WAP-mediated renal protection is associated with suppression of inflammation and renal epithelial cell apoptosis.

Macrophage migration inhibitory factor promotes cyst growth in polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) is characterized by renal cyst formation, inflammation, and fibrosis. Macrophages infiltrate cystic kidneys, but the role of these and other inflammatory factors in disease progression are poorly understood. Here, we identified macrophage migration inhibitory factor (MIF) as an important regulator of cyst growth in ADPKD. MIF was upregulated in cyst-lining epithelial cells in polycystin-1-deficient murine kidneys and accumulated in cyst fluid of human ADPKD kidneys. MIF promoted cystic epithelial cell proliferation by activating ERK, mTOR, and Rb/E2F pathways and by increasing glucose uptake and ATP production, which inhibited AMP-activated protein kinase signaling. MIF also regulated cystic renal epithelial cell apoptosis through p53-dependent signaling. In polycystin-1-deficient mice, MIF was required for recruitment and retention of renal macrophages, which promoted cyst expansion, and Mif deletion or pharmacologic inhibition delayed cyst growth in multiple murine ADPKD models. MIF-dependent macrophage recruitment was associated with upregulation of monocyte chemotactic protein 1 (MCP-1) and inflammatory cytokine TNF-α. TNF-α induced MIF expression, and MIF subsequently exacerbated TNF-α expression in renal epithelial cells, suggesting a positive feedback loop between TNF-α and MIF during cyst development. Our study indicates MIF is a central and upstream regulator of ADPKD pathogenesis and provides a rationale for further exploration of MIF as a therapeutic target for ADPKD.

Aristolochic acid-induced apoptosis and G2 cell cycle arrest depends on ROS generation and MAP kinases activation.

Ingestion of aristolochic acids (AAs) contained in herbal remedies results in a renal disease and, frequently, urothelial malignancy. The genotoxicity of AA in renal cells, including mutagenic DNA adducts formation, is well documented. However, the mechanisms of AA-induced tubular atrophy and renal fibrosis are largely unknown. To better elucidate some aspects of this process, we studied cell cycle distribution and cell survival of renal epithelial cells treated with AAI at low and high doses. A low dose of AA induces cell cycle arrest in G2/M phase via activation of DNA damage checkpoint pathway ATM-Chk2-p53-p21. DNA damage signaling pathway is activated more likely via increased production of reactive oxygen species (ROS) caused by AA treatment then via DNA damage induced directly by AA. Higher AA concentration induced cell death partly via apoptosis. Since mitogen-activated protein kinases play an important role in cell survival, death and cell cycle progression, we assayed their function in AA-treated renal tubular epithelial cells. ERK1/2 and p38 but not JNK were activated in cells treated with AA. In addition, pharmacological inhibition of ERK1/2 and p38 as well as suppression of ROS generation with N-acetyl-L-cysteine resulted in the partial relief of cells from G2/M checkpoint and a decline of apoptosis level. Cell cycle arrest may be a mechanism for DNA repair, cell survival and reprogramming of epithelial cells to the fibroblast type. An apoptosis of renal epithelial cells at higher AA dose might be necessary to provide space for newly reprogrammed fibrotic cells.

Effects of hypoxia/reoxygenation on apoptosis and gap junction in isolated renal tubular epithelial cells of rats

Objective To evaluate the effects of hypoxia/reoxygenation (H/R) on apoptosis and gap junction in isolated renal tubular epithelial cells of rats.Methods The renal tubular epithelial cells were seeded in 6-well plates at a density of 1 × 105 cells/ml and randomly divided into 2 groups (n =18 wells each) using a random number table:control group (group C) and H/R group.In group C,the cells were cultured in a regular incubator with 21％ oxygen at 37 ℃.In group H/R,the cells were incubated in an anaerobic chamber for 24 h and then returned to a regular incubator and incubated for 4 h.Hoechst33258 staining method and flow cytometry were used to detect the cell apoptosis.Gap junction function was determined by specific parachute assay.Gap function protein connexin 43(Cx43) expression was measured by Western blot.Results Compared with group C,the apoptosis rate was significantly increased and gap junction function and Cx43 expression were decreased in group H/R (P ＜ 0.05).Conclusion H/R promotes apoptosis in isolated renal tubular epithelial cells through destroying intercellular gap junction in rats. Key words: Cell hypoxia ; Oxygen ; Kidney tubules ; Epithelial cells ; Apoptosis ; Connexin 43

Role of Annexin A5 in Cisplatin-induced Toxicity in Renal Cells: MOLECULAR MECHANISM OF APOPTOSIS

Annexin A5 belongs to a large family of calcium-binding and phospholipid-binding proteins and may act as an endogenous regulator of various pathophysiological processes. There is increasing evidence that annexin A5 is related to cytotoxicity, but the precise function of this protein has yet to be elucidated. In this study, we aimed to verify the function of annexin A5 in the apoptosis of renal epithelial cells. Real-time PCR and Western blot analysis, together with immunofluorescence analysis, showed that the expression of annexin A5 significantly increased in the presence of cisplatin in both human and rat renal epithelial cells. With regard to the mechanism of cisplatin-induced apoptosis, apoptosis-inducing factor (AIF) release into the cytosol was observed, and the underlying mechanism was identified as voltage-dependent anion channel (VDAC) oligomerization. Mitochondrial membrane potential (Δψm) was found to be greatly disrupted in cisplatin-treated cells. Moreover, cisplatin strongly induced translocation of annexin A5 into mitochondria. To understand the functional significance of annexin A5 in renal cell death, we used a siRNA-mediated approach to knock down annexin A5. Annexin A5 depletion by siRNA led to decreased annexin A5 translocation into mitochondria and significantly reduced VDAC oligomerization and AIF release. Annexin A5 siRNA also increased cell viability compared with the control. Moreover, expression of annexin A5 was induced by other nephrotoxicants such as CdCl2 and bacitracin. Taken together, our data suggest that annexin A5 may play a crucial role in cisplatin-induced toxicity by mediating the mitochondrial apoptotic pathway via the induction and oligomerization of VDAC.