Proximal Tubular Cell Damage Research Articles

NRF2 Activation by AR-20007 Preserves Renal Tubular Epithelial Cells from Antimycin A-Induced Cell Death via Glutathione Metabolism Regulation.

Oxidative stress plays a crucial role in the development and progression of various kidney diseases. Nuclear factor erythroid 2-related factor 2 (NRF2) is the primary transcription factor that protects cells from oxidative stress by regulating cytoprotective genes including those involved in the antioxidant glutathione (GSH) pathway. GSH maintains cellular redox status and affects redox signaling, cell proliferation, and cell death. Antimycin A, an inhibitor of complex III of the electron transport chain, causes oxidative stress and reduces GSH levels. In this study, we induced mitochondrial damage in rat renal proximal tubular cells using antimycin A and investigated cellular viability and levels of NRF2 and GSH. Treatment with antimycin A altered the expression of antioxidant genes, including reduction in the transcription of glutathione-cysteine ligase subunits (Gclc and Gclm) and glutathione reductase (Gsr1), followed by a reduction in total GSH content with a concomitant decrease in NRF2 protein expression. AR-20007, previously described as an NRF2 activator, stabilizes and increases NRF2 protein expression in cells. By stimulating NRF2, AR-20007 increased the expression of antioxidant and detoxifying enzymes, thereby enhancing protection against oxidative stress induced by antimycin A. These data suggest that NRF2 activation effectively inhibits antimycin A-induced oxidative stress and that NRF2 may be a promising therapeutic target for preventing cell death during acute kidney injury.

SGLT2 inhibitor dapagliflozin reduces proximal tubular cell damage biomarkers in patients with acute heart failure

SGLT2 inhibitor dapagliflozin reduces proximal tubular cell damage biomarkers in patients with acute heart failure

Cadmium Induces Kidney Iron Deficiency and Chronic Kidney Injury by Interfering with the Iron Metabolism in Rats.

Cadmium (Cd) is a common environmental pollutant and occupational toxicant that seriously affects various mammalian organs, especially the kidney. Iron ion is an essential trace element in the body, and the disorder of iron metabolism is involved in the development of multiple pathological processes. An iron overload can induce a new type of cell death, defined as ferroptosis. However, whether iron metabolism is abnormal in Cd-induced nephrotoxicity and the role of ferroptosis in Cd-induced nephrotoxicity need to be further elucidated. Sprague Dawley male rats were randomly assigned into three groups: a control group, a 50 mg/L CdCl2-treated group, and a 75 mg/L CdCl2-treated group by drinking water for 1 month and 6 months, respectively. The results showed that Cd could induce renal histopathological abnormalities and dysfunction, disrupt the mitochondria's ultrastructure, and increase the ROS and MDA content. Next, Cd exposure caused GSH/GPX4 axis blockade, increased FTH1 and COX2 expression, decreased ACSL4 expression, and significantly decreased the iron content in proximal tubular cells or kidney tissues. Further study showed that the expression of iron absorption-related genes SLC11A2, CUBN, LRP2, SLC39A14, and SLC39A8 decreased in proximal tubular cells or kidneys after Cd exposure, while TFRC and iron export-related gene SLC40A1 did not change significantly. Moreover, Cd exposure increased SLC11A2 gene expression and decreased SLC40A1 gene expression in the duodenum. Finally, NAC or Fer-1 partially alleviated Cd-induced proximal tubular cell damage, while DFO and Erastin further aggravated Cd-induced cell damage. In conclusion, our results indicated that Cd could cause iron deficiency and chronic kidney injury by interfering with the iron metabolism rather than typical ferroptosis. Our findings suggest that an abnormal iron metabolism may contribute to Cd-induced nephrotoxicity, providing a novel approach to preventing kidney disease in clinical practice.

Angiotensin 1-7 Exerts Antioxidant Effects, Suppresses Mammalian Target of Rapamycin (mTOR) Signaling, and Inhibits Apoptosis in Renal Proximal Tubular Cells

Oxidative stress is one of the crucial pathogenic factors involved in the progression of renal injury. Angiotensin (ANG) 1-7, a bioactive heptapeptide of the renin-angiotensin-aldosterone system is known to exert antioxidant and nephroprotective effects. However, the cellular mechanism involved in the beneficial effect of ANG 1-7 is not clear. Here, we assessed ANG 1-7’s effect on H2O2-mediated oxidative damage in the human proximal tubular (HK2) cells and the underlying mechanisms. HK2 cells were incubated with H2O2 (500µM, 4h) pre-treated with and without ANG 1-7 (100nM, 24h), and reactive oxygen species (ROS) generation, mitochondrial dysfunction, endoplasmic reticulum (ER) stress, apoptosis and mammalian target of rapamycin (mTOR) signaling were determined. H2O2 induced an increase in oxidative and ER stress together with loss of mitochondrial membrane potential, decreased ATP levels, and induced apoptosis in HK2 cells. Moreover, H2O2 treatment resulted in the activation of mTOR complexes (mTORC1 and mTORC2) in these cells. ANG 1-7 significantly attenuated H2O2-induced ROS generation, ER stress and apoptosis, and also improved mitochondrial function. Additionally, pre-treatment of ANG 1-7 inhibited the H2O2-mediated mTOR activation. These effects of ANG 1-7 were blocked by co-treatment with the Mas receptor (MasR) inhibitor, A779. Furthermore, transfection of HK2 cells with Mas receptor siRNA also abolished the inhibitory effect of ANG 1-7 on mTOR activities. In conclusion, ANG 1-7 via MasR mitigates oxidative stress, suppresses mTOR signaling, and protects HK2 cells from ER stress, mitochondrial dysfunction, and apoptosis, suggesting ANG 1-7-MasR renoprotective effects.

Valproic acid treatment attenuates cisplatin-induced kidney injury by suppressing proximal tubular cell damage.

Cisplatin treatment is effective against several types of carcinomas. However, it frequently leads to kidney injury, which warrants effective prevention methods. Sodium valproic acid is a prophylactic drug candidate with a high potential for clinical application against cisplatin-induced kidney injury. Therefore, in this study, we aimed to elucidate the mechanism underlying the prophylactic effect of valproic acid on cisplatin-induced kidney injury in a mouse model and HK2 and PODO cells with cisplatin-induced toxicity. In the mouse model of cisplatin-induced kidney injury, various renal function parameters and tubular damage scores were worsened by cisplatin, but they were significantly improved upon combination with valproic acid. No difference was observed in cisplatin accumulation between the cisplatin-treated and valproic acid-treated groups in whole blood and the kidneys. The mRNA expression levels of proximal tubular damage markers, apoptosis markers, and inflammatory cytokines significantly increased in the cisplatin group 72 h after cisplatin administration but significantly decreased upon combination with valproic acid. In HK2 cells, a human proximal tubular cell line, the cisplatin-induced decrease in cell viability was significantly suppressed by co-treatment with valproic acid. Valproic acid may inhibit cisplatin-induced kidney injury by suppressing apoptosis, inflammatory responses, and glomerular damage throughout the kidneys by suppressing proximal tubular cell damage. However, prospective controlled trials need to evaluate these findings before their practical application.

Urinary exosomal miRNA-663a shows variable expression in diabetic kidney disease patients with or without proteinuria

Heterogeneity in the Diabetic Kidney Disease (DKD) diagnosis makes its rational therapeutics challenging. Although albuminuria characterizes DKD, reports also indicate its prevalence among non-proteinuric. Recent understanding of disease progression has thus inclined the focus on proximal tubular cell damage besides the glomeruli. A non-invasive approach exploiting exosomal miRNA derived from human kidney proximal tubular cell line was, hence, targeted. Upon miRNA profiling, three miRNAs, namely, hsa-miR-155-5p, hsa-miR-28-3p, and hsa-miR-425-5p were found to be significantly upregulated, while hsa-miR-663a was downregulated under diabetic conditions. Among these, hsa-miR-663a downregulation was more pronounced in non-proteinuric than proteinuric DKD subjects and was thus selected for the bioinformatics study. Ingenuity Pathway Analysis (IPA) narrowed on to IL-8 signaling and inflammatory response as the most enriched ‘canonical pathway’ and ‘disease pathway’ respectively, during DKD. Further, the putative gene network generated from these enriched pathways revealed experimentally induced diabetes, renal tubular injury, and decreased levels of albumin as part of mapping under ‘disease and function’. Genes target predictions and annotations by IPA reiterated miR-663a’s role in the pathogenesis of DKD following tubular injury. Overall, the observations might offer an indirect reflection of the underlying mechanism between patients who develop proteinuria and non-proteinuria.

Exosomes derived from calcium oxalate-treated macrophages promote apoptosis of HK-2 cells by promoting autophagy

ABSTRACT Calcium oxalate (CaOx) crystals are the main component of kidney stones. Macrophages have the function of eliminating these crystals, and the underlying mechanism remains unclear. Here, we attempted to determine the role of macrophage-derived exosomes exposed to CaOx crystals in regulating apoptosis of human proximal tubular cells (HK-2). Exosomes (CaOx-Exo) were isolated from CaOx-treated macrophages and then incubated with HK-2 cells. CaOx-Exo treatment reduced cell viability and promoted apoptosis of HK-2 cells. The expression of Caspase-3 and Bax was increased, and Bcl-2 expression was decreased in HK-2 cells following CaOx-Exo treatment. Moreover, CaOx-Exo treatment caused an increase of LC3-II/LC3-I ratio and Beclin-1 expression and a downregulation of p62 in HK-2 cells. GFP-LC3 puncta were increased in HK-2 cells following CaOx-Exo treatment. Additionally, CaOx-Exo-treated HK-2 cells were treated with 3-methyladenine (3-MA) to inhibit autophagy activity. 3-MA treatment weakened the impact of CaOx-Exo on cell viability and apoptosis of HK-2 cells. 3-MA treatment also reduced the LC3-II/LC3-I ratio and Beclin-1 expression and enhanced p62 expression in CaOx-Exo-treated HK-2 cells. In conclusion, these data demonstrated that exosomes derived from CaOx-treated macrophages promote apoptosis of HK-2 cells by promoting autophagy. Thus, this work suggests that macrophage-derived exosomes may play a vital role in CaOx-induced human proximal tubular cell damage.

Vitamin B12 does not increase cell viability after hydrogen peroxide induced damage in mouse kidney proximal tubular cells and brain endothelial cells

Vitamin B12 (B12) is an essential co-factor for two enzymes in mammalian metabolism and can also act as a mimetic of superoxide dismutase (SOD) converting superoxide (O2•−) to hydrogen peroxide (H2O2). High oral dose B12 decreases renal O2•− and post-ischemia/reperfusion injury in mice and protects against damage induced by hypoxia/reperfusion in mouse kidney proximal tubular cells (BU.MPT). O2•− is unstable and rapidly converted to H2O2. H2O2 mediates oxidative stress associated with O2•−. Whether B12 protects against damage induced by H2O2 is unknown. Both BU.MPT cells and mouse brain endothelial cells (bEdn.3) were applied to test the effects of B12 on H2O2-induced cytotoxicity. Both types of cells were treated with different doses of H2O2 with or without different doses of B12. Cell viability was analyzed 24 h later. H2O2 caused cell death only at a very high dose, and high pharmacological dose of B12 did not prevent this detrimental effect in either cell type. In bEnd.3 cells, transcriptional levels of heme oxygenase-1 (HO-1) increased, while nuclear factor erythroid 2-related factor 2 (Nrf2) decreased by H2O2. The levels of transcripts were not affected by the B12 treatment. We conclude that the cytotoxic effects of exogenous H2O2 in BU.MPT and bEdn.3 cells are not prevented by B12.

DNA repair factor KAT5 prevents ischemic acute kidney injury through glomerular filtration regulation

SummaryThe “preconditioning effect” in AKI is a phenomenon in which an episode of ischemia-reperfusion results in tolerance to subsequent ischemia-reperfusion injury. However, its relationship between DNA damage repair has not been elucidated. Here, we show the role of KAT5 in the preconditioning effect. Preconditioning attenuated DNA damage in proximal tubular cells with elevated KAT5 expression. Ischemia-reperfusion (IR) injuries were exacerbated, and preconditioning effect vanished in proximal tubular-cell-specific KAT5 knockout mice. Investigation of tubuloglomerular feedback (TGF) by MALDI-IMS and urinary adenosine revealed that preconditioning caused attenuated TGF at least in part via KAT5. In addition, K-Cl cotransporter 3 (KCC3) expression decreased in damaged proximal tubular cells, which may be involved in accelerated TGF following IR. Furthermore, KAT5 induced KCC3 expression by maintaining chromatin accessibility and binding to the KCC3 promoter. These results suggest a novel mechanism of the preconditioning effect mediated by the promotion of DNA repair and attenuation of TGF through KAT5.

Tannic acid attenuates vascular calcification-induced proximal tubular cells damage through paracrine signaling

Vascular calcification is common in chronic kidney disease; however, the extent to which such condition can affect the renal microvasculature and the neighboring cell types is unclear. Our induced-calcification model in renal proximal tubular (PT) cells exhibited endoplasmic reticulum (ER) stress and oxidative damage, leading to apoptosis. Here, we utilized such calcification in mouse vascular smooth muscle (MOVAS-1) cells as a vascular calcification model, because it exhibited reactive oxygen species (ROS) generation, ER and oxidative stress, inflammatory, and apoptotic gene expressions. To demonstrate whether the vascular calcification condition can dictate the function of the adjacent PT cell layer, we utilized a Transwell multilayer culture system by combining those MOVAS-1 cells in the bottom chamber and polarized PT cells in the upper chamber to show the dimensional cross-signaling effect. Interestingly, calcification of MOVAS-1 cells, in this co-culture, induced H2O2 and lactate dehydrogenase (LDH) release leading to store-operated Ca2+ entry, ROS generation, and activation of oxidative, inflammatory, and apoptotic gene expressions in PT cells through paracrine signaling. Interestingly, application of tannic acid (TA) to either calcified MOVAS-1 or uncalcified PT cells diminished such detrimental pathway activation. Furthermore, the TA-mediated protection was much higher in the PT cells when applied on the calcified MOVAS-1 cells, and the delayed the pathological effects in neighboring PT cells can well be via paracrine signaling. Together, these results provide evidence of vascular calcification-induced PT cell damage, and the protective role of TA in preventing such pathological consequences, which can potentially be used as a nephroprotective remedy.

Renal effect of severe hypoxia evaluated By NGAL measurements: An in vivo and in vitro study.

To investigate possible renal damage in healthy men exposed to extreme hypobaric hypoxia, using urinary Neutrophil Gelatinase-Associated Lipocalin (NGAL) concentration as biomarker. The value of NGAL as a biomarker of proximal tubular cell damage under hypoxic conditions was also tested in vitro experiments. NGAL was assayed in a cohort of air cadets (n = 16) exposed to hypobaric hypoxia in a hypobaric chamber during their training program. In all subjects, urine creatinine (Cr) and urinary NGAL levels were measured immediately before, 3, and 24 h after hypobaric environment exposure. Three in vitro experiments using proximal tubular cell cultures were also performed to measure NGAL gene expression, NGAL secretion in the culture medium and to evaluate apoptosis under two cycles of hypoxia and reoxygenation. In the in vivo study, geometric means of urinary NGAL/Cr ratio measured 24 h after hypobaric hypoxia in the hypobaric chamber were significantly lower than baseline values (13.4 vs 25.9 ng/mg, p = 0.01). In cell cultures, hypoxia down-regulated NGAL gene expression without significantly changing NGAL secretion in the culture medium. Hypoxia significantly increased the percentage of apoptotic/necrotic cells, especially after the second hypoxia-reoxygenation cycle. Exposure to hypobaric-hypoxic environments does not cause significant and irreversible renal tubular injury in vivo and in vitro, except than in a late stage. The hypoxic insult does not seem to be mirrored by an increase of urinary NGAL in healthy men nor of NGAL gene expression in HK-2 cell culture or secretion in the culture medium in the in vitro conditions reported in the present study.

Potential Protection Effect of ER Homeostasis of N6-(2-Hydroxyethyl)adenosine Isolated from Cordyceps cicadae in Nonsteroidal Anti-Inflammatory Drug-Stimulated Human Proximal Tubular Cells.

Nonsteroidal anti-inflammatory drugs (NSAIDs) belong to a class of universally and commonly used anti-inflammatory analgesics worldwide. A diversity of drawbacks of NSAIDs have been reported including cellular oxidative stress, which in turn triggers the accumulation of unfolded proteins, enhancing endoplasmic reticulum stress, and finally resulting in renal cell damage. Cordyceps cicadae (CC) has been used as a traditional medicine for improving renal function via its anti-inflammatory effects. N6-(2-hydroxyethyl)adenosine (HEA), a physiologically active compound, has been reported from CC mycelia (CCM) with anti-inflammatory effects. We hypothesize that HEA could protect human proximal tubular cells (HK–2) from NSAID-mediated effects on differential gene expression at the mRNA and protein levels. To verify this, we first isolated HEA from CCM using Sephadex® LH–20 column chromatography. The MTT assay revealed HEA to be nontoxic up to 100 µM toward HK–2 cells. The HK–2 cells were pretreated with HEA (10–20 µM) and then insulted with the NSAIDs diclofenac (DCF, 200 µM) and meloxicam (MXC, 400 µM) for 24 h. HEA (20 µM) effectively prevented ER stress by attenuating ROS production (p < 0.001) and gene expression of ATF–6, PERK, IRE1α, CDCFHOP, IL1β, and NFκB within 24 h. Moreover, HEA reversed the increase of GRP78 and CHOP protein expression levels induced by DCF and MXC, and restored the ER homeostasis. These results demonstrated that HEA treatments effectively protect against DCF- and MXC-induced ER stress damage in human proximal tubular cells through regulation of the GRP78/ATF6/PERK/IRE1α/CHOP pathway.

Induction of oxidative stress and DNA damage in human renal proximal tubular cells by aristolochic acid

Aristolochic acid 1 (AAI) is found primarily in the plant Aristolochia. Consumption of products containing AAI has been linked with permanent kidney damage and urothelial carcinoma. This study applied human proximal tubule epithelial cell line (HK-2) to examine the relationship among AAI-induced intracellular oxidative stress, DNA damage and MAP kinase activation. High concentrations of AAI caused a decrease in cell viability and an increase in the activity of caspase 3. AAI treatment also led to a dose-dependent increase of reactive oxygen species (ROS) in HK-2 cells, and the presence of antioxidant glutathione (GSH) effectively inhibited ROS generation. Stimulating HK-2 cultures with AAI also led to GSH depletion. Results from single cell gel electrophoresis (SCGE) assays demonstrated that AAI showed the ability to increase the levels of DNA strand breaks in HK-2 cells. Up-regulation of luciferase activity driven by the Nrf2 binding element was also observed after 200 μM AAI treatment. Exposure of HK-2 cells with AAI activated both ERK1/2 and p38 kinase phosphorylation, while only the MEK1/2 inhibitor, U0126, significantly decreased the levels of AAI-mediated ROS. In addition, both U0126 and SB202190 effectively reversed the levels of DNA damage triggered by AAI. This suggests that AAI treatment of HK-2 results in ROS formation and DNA damage. Furthermore, ROS generation occurs via the MEK/ERK1/2 signaling pathway, whereas DNA damage occurs via both the ERK1/2 and p38 pathways.

Lipotoxicity, Nutrient-Sensing Signals, and Autophagy in Diabetic Nephropathy.

Diabetic nephropathy is a leading cause of proteinuria, kidney fibrosis, and subsequent end-stage renal disease. The renal prognosis of diabetic patients with refractory proteinuria is extremely poor. Therefore, identification of novel therapeutic targets to combat this serious condition and improve renal prognosis is urgently necessary. In diabetic patients, in addition to blood glucose levels, serum levels of free fatty acids (FFAs) are chronically elevated, even during postprandial periods. Of the various types of FFAs, saturated FFAs are highly cytotoxic and their levels are elevated in the serum of patients with diabetes. Thus, an increase in saturated FFAs is currently thought to contribute to proximal tubular cell damage and podocyte injury in diabetic nephropathy. Therefore, protecting both types of kidney cells from saturated FFA-related lipotoxicity may become a novel therapeutic approach for diabetic patients with refractory proteinuria. Interestingly, accumulating evidence suggests that controlling intracellular nutrient signals and autophagy can ameliorate the FFA-related kidney damage. Here, we review the evidence indicating possible mechanisms underlying cell injury caused by saturated FFAs and cell protective roles of intracellular nutrient signals and autophagy in diabetic nephropathy.

L-ornithine activates Ca2+ signaling to exert its protective function on human proximal tubular cells

Oxidative stress and reactive oxygen species (ROS) generation can be influenced by G-protein coupled receptor (GPCR)-mediated regulation of intracellular Ca2+ ([Ca2+]i) signaling. ROS production are much higher in proximal tubular (PT) cells; in addition, the lack of antioxidants enhances the vulnerability to oxidative damage. Despite such predispositions, PT cells show resiliency, and therefore must possess some inherent mechanism to protect from oxidative damage. While the mechanism in unknown, we tested the effect of l-ornithine, since it is abundantly present in PT luminal fluid and can activate Ca2+-sensing receptor (CaSR), a GPCR, expressed in the PT luminal membrane. We used human kidney 2 (HK2) cells, a PT cell line, and performed Ca2+ imaging and electrophysiological experiments to show that l-ornithine has a concentration-dependent effect on CaSR activation. We further demonstrate that the operation of CaSR activated Ca2+ signaling in HK-2 cells mediated by the transient receptor potential canonical (TRPC) dependent receptor-operated Ca2+ entry (ROCE) using pharmacological and siRNA inhibitors. Since PT cells are vulnerable to ROS, we simulated such deleterious effects using genetically encoded peroxide-induced ROS production (HyperRed indicator) to show that the l-ornithine-induced ROCE mediated [Ca2+]i signaling protects from ROS production. Furthermore, we performed cell viability, necrosis and apoptosis assays, and mitochondrial oxidative gene expression to establish that presence of l-ornithine rescued the ROS-induced damage in HK-2 cells. Moreover, l-ornithine-activation of CaSR can reverse ROS production and apoptosis via mitogen-activated protein kinase p38 activation. Such nephroprotective role of l-ornithine can be useful as the translational option for reversing kidney diseases involving PT cell damage due to oxidative stress or crystal nephropathies.

Protective effect of urolithin a on cisplatin-induced nephrotoxicity in mice via modulation of inflammation and oxidative stress

Limitation of widely used anti-cancer agent cisplatin for a patient is nephrotoxicity. Nephrotoxicity is presentable in mice by injecting cisplatin at 25 mg/kg with 3 days endpoint. We used the same model to understand the protective role of urolithin A. Cisplatin-induced renal damages measured by histological damage in proximal tubular cells and by the increase in serum neutrophil gelatinase-associated lipocalin (NGAL), blood urea nitrogen (BUN), creatinine and urinary Kidney Injury Molecule-1 (KIM-1). Urolithin A pretreatment reduced all the above renal damage parameters in a significant way. Urolithin A attenuated cisplatin-induced pro-inflammatory cytokine/chemokine tumor necrosis factor α (TNFα), interleukin 23 (IL-23), interleukin 18 (IL-18) and macrophage inflammatory protein 2 (MIP2). Cisplatin-induced CD11b positive macrophages in kidneys reduced by urolithin A. Urolithin A also attenuated cisplatin-induced renal oxidative/nitrative stress, which was measured by lipid peroxidation(4-hydroxy-2-nonenal or 4-HNE protein adducts) and protein nitration. Urolithin A cisplatin-induced kidney injury in mice through the down regulation of inflammatory cytokines/chemokine, immune cells, and oxidative/nitrative stress thus improving cisplatin-induced proximal tubular cell death.

Melamine promotes calcium crystal formation in three-dimensional microfluidic device

Melamine, which induces proximal tubular (PT) cell damage has a greater nephrotoxic effect when combined with cyanuric and uric acids; however, it is unknown whether such effect can stimulate calcium phosphate (CaP)/calcium oxalate (CaOx) stone formation. Here, we show that melamine acts as an inducer of CaP, CaOx and CaP + CaOx (mixed) crystal formations in a time and concentration-dependent manner by stabilizing those crystals and further co-aggregating with melamine. To explore the physiological relevance of such melamine-augmented calcium crystal formation, we used 2-dimensional (2D) and 3D microfluidic (MF) device, embedded with PT cells, which also resembled the effect of melamine-stimulated CaP, CaOx and mixed crystal formation. Significantly, addition of preformed CaP and/or CaOx crystal in the presence of melamine, further potentiated those crystal formations in 3D MFs, which helped the growth and aggregation of mixed crystals. Our data show that the mechanism of such predisposition of stone formation could be largely due to co-crystallization between melamine and CaP/CaOx and pronounced effect on induction of stone-forming pathway activation in 3D MF. Taken together, melamine-induced CaP and/or CaOx crystal formation ex-vivo will help us in understanding the larger role of melamine as an environmental toxicant in producing the pathology in similar cellular microenvironments.

The Involvement of Glucose Transporter Gene Expression in Cadmium Renal Toxicity

Cadmium (Cd) is an environmental contaminant known to exert renal toxicity. Proximal tubular cell damage is characterized as Cd-induced renal toxicity. Previous studies revealed that Cd caused the gene expression disruption in proximal tubular cells. However, the transcription factors that regulate the expression of genes associated with Cd renal toxicity are poorly understood. Our previous study demonstrated MEF2A (myocyte enhancer factor 2A) transcription factor that is involved in Cd toxicity in HK-2 human proximal tubular cells, using protein/DNA binding array and RNAi method. In this study, it was investigated which downstream factor of MEF2A is involved in Cd toxicity in HK-2 cells. MEF2A is reported to regulate the gene expression of SLC2A4, which encodes GLUT4 (glucose transporter 4) protein. We found that SLC2A4 expression is regulated by MEF2A in HK-2 cells. Moreover, Cd decreased not only the mRNA level of SLC2A4 but also the protein level of GLUT4 in HK-2 cells. In addition, SLC2A4 knockdown by siRNA conferred the toxicity to HK-2 cells. On the other hand, GLUT2 protein is reported to be expressed abundantly in the mammalian kidney. However, the knockdown of the GLUT2 encoding gene, SLC2A2, did not induce the toxicity in HK-2 cells. Furthermore, MEF2A knockdown did not inhibit the gene expression of SLC2A2 in HK-2 cells. These results suggest that the pathway of Cd renal toxicity through the decrease in activity of MEF2A involves the suppression of SLC2A4 gene expression.

Ameliorative Effect of Daidzein on Cisplatin-Induced Nephrotoxicity in Mice via Modulation of Inflammation, Oxidative Stress, and Cell Death.

Oxidative stress and inflammation are part and parcel of cisplatin-induced nephrotoxicity. The purpose of this work is to study the role of soy isoflavone constituent, daidzein, in cisplatin-induced renal damage. Cisplatin-induced nephrotoxicity was evident by the histological damage in proximal tubular cells and by the increase in serum neutrophil gelatinase-associated lipocalin (NGAL), blood urea nitrogen (BUN), creatinine, and urinary kidney injury molecule-1 (KIM-1). Cisplatin-induced cell death was shown by TUNEL staining and caspase-3/7 activity. Daidzin treatment reduced all kidney injury markers (NGAL, BUN, creatinine, and KIM-1) and attenuated cell death (apoptotic markers). In cisplatin-induced kidney injury, renal oxidative/nitrative stress was manifested by the increase in lipid peroxidation and protein nitration. Cisplatin induced the reactive oxygen species-generating enzyme NOX-2 and impaired antioxidant defense enzyme activities such as glutathione peroxidase (GPX) and superoxide dismutase (SOD) activities. Cisplatin-induced oxidative/nitrative stress was attenuated by daidzein treatment. Cisplatin induced CD11b-positive macrophages in kidneys and daidzein attenuated CD11b-positive cells. Daidzein attenuated cisplatin-induced inflammatory cytokines tumor necrosis factor α (TNFα), interleukin 10 (IL-10), interleukin 18 (IL-18), and monocyte chemoattractant protein-1 (MCP-1). Daidzein attenuated cell death in vitro. Our data suggested that daidzein attenuated cisplatin-induced kidney injury through the downregulation of oxidative/nitrative stress, immune cells, inflammatory cytokines, and apoptotic cell death, thus improving kidney regeneration.

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.