scholarly journals Calciprotein particle-induced cytotoxicity via lysosomal dysfunction and altered cholesterol distribution in renal epithelial HK-2 cells

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Rina Kunishige ◽  
Mai Mizoguchi ◽  
Asako Tsubouchi ◽  
Kenjiro Hanaoka ◽  
Yutaka Miura ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Serum Proteins ◽  
Membrane Integrity ◽  
Reactive Oxygen Species Generation ◽  
Autophagic Flux ◽  
Membrane Injury ◽  
Renal Tubular Cells ◽  
Calciprotein Particles ◽  
Renal Proximal Tubular ◽  
Renal Tubular

AbstractDietary phosphate overload induces chronic kidney disease (CKD), and calciprotein particles (CPPs), a form of nanoparticle comprising calcium phosphate and serum proteins, has been proposed to cause renal toxicity. However, the mechanism of CPP cytotoxicity in renal tubular cells is unknown. Here we show that in renal proximal tubular epithelial HK-2 cells, endocytosed CPPs accumulate in late endosomes/lysosomes (LELs) and increase their luminal pH by ~ 1.0 unit. This results in a decrease in lysosomal hydrolase activity and autophagic flux blockage without lysosomal rupture and reactive oxygen species generation. CPP treatment led to vulnerability to H2O2-induced oxidative stress and plasma membrane injury, probably because of autophagic flux blockage and decreased plasma membrane cholesterol, respectively. CPP-induced disruption of lysosomal homeostasis, autophagy flux and plasma membrane integrity might trigger a vicious cycle, leading to progressive nephron loss.

scholarly journals Exocytosis of acid sphingomyelinase by wounded cells promotes endocytosis and plasma membrane repair

2010 ◽  
Vol 189 (6) ◽  
pp. 1027-1038 ◽  
Author(s):  
Christina Tam ◽  
Vincent Idone ◽  
Cecilia Devlin ◽  
Maria Cecilia Fernandes ◽  
Andrew Flannery ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Lysosomal Enzyme ◽  
Membrane Integrity ◽  
Acid Sphingomyelinase ◽  
Membrane Repair ◽  
Membrane Injury ◽  
Cellular Survival ◽  
Plasma Membrane Repair ◽  
Membrane Resealing ◽  
Niemann Pick

Rapid plasma membrane resealing is essential for cellular survival. Earlier studies showed that plasma membrane repair requires Ca2+-dependent exocytosis of lysosomes and a rapid form of endocytosis that removes membrane lesions. However, the functional relationship between lysosomal exocytosis and the rapid endocytosis that follows membrane injury is unknown. In this study, we show that the lysosomal enzyme acid sphingomyelinase (ASM) is released extracellularly when cells are wounded in the presence of Ca2+. ASM-deficient cells, including human cells from Niemann-Pick type A (NPA) patients, undergo lysosomal exocytosis after wounding but are defective in injury-dependent endocytosis and plasma membrane repair. Exogenously added recombinant human ASM restores endocytosis and resealing in ASM-depleted cells, suggesting that conversion of plasma membrane sphingomyelin to ceramide by this lysosomal enzyme promotes lesion internalization. These findings reveal a molecular mechanism for restoration of plasma membrane integrity through exocytosis of lysosomes and identify defective plasma membrane repair as a possible component of the severe pathology observed in NPA patients.

Abstract 210: Stem Cell Conditioned Culture Media Attenuated Albumin-induced Epithelial-mesenchymal Transition in Renal Tubular Cells

Hypertension ◽  
2013 ◽  
Vol 62 (suppl_1) ◽  
Author(s):  
Junping Hu ◽  
Weiqing Han ◽  
Qing Zhu ◽  
Pin-Lan Li ◽  
Ningjun Li
Keyword(s):  
Epithelial Mesenchymal Transition ◽  
Culture Media ◽  
Mesenchymal Transition ◽  
Tubular Cells ◽  
Protein Levels ◽  
Renal Tubular Cells ◽  
Proximal Tubular ◽  
Renal Proximal Tubular Cells ◽  
Renal Proximal Tubular ◽  
Renal Tubular

Mesenchymal stem cells (MSCs) have been shown to be a promising therapy for many different diseases. Stem cell conditioned culture media (SCM) exhibit similar beneficial effects as MSCs. Albuminuria-induced epithelial-mesenchymal transition (EMT) plays an important role in progressive renal tubulointerstitial fibrosis in chronic renal disease. The present study tested the hypothesis that SCM inhibit albumin-induced EMT in cultured renal tubular cells. SCM were obtained by culturing rat adult MSCs for 3 days. Cultured renal proximal tubular cells were incubated with rat albumin (20μg/ml) and treated with SCM or control culture media. Our results showed that 48 h albumin incubation stimulated EMT in renal proximal tubular cells as shown by significant decrease in the protein levels of epithelial marker E-cadherin from 2.30 ± 0.27 to 0.87 ± 0.11 ( P < 0.05) and increase in the protein levels of mesenchymal marker fibroblast-specific protein 1 (FSP-1) (2.18±0.33 folds, P < 0.05). SCM treatment significantly inhibited these albumin-induced changes in E-cadherin and FSP-1 by 2.33±0.17 and 1.95±0.23 folds ( P < 0.05), respectively. Meanwhile, albumin increased the mRNA levels of pro-inflammatory factor monocyte chemoattractant protein-1 (MCP)-1 by nearly 30 folds compared with control. SCM almost abolished the increase of MCP-1 induced by albumin. Furthermore, Western blot results displayed that albumin rapidly decreased the cytosolic levels and increased the nuclear levels of NF-κB, indicating a translocation of NF-κB; immunofluorescence microscopy also demonstrated that albumin induced NF-κB translocation from the cytosol into nucleus. SCM blocked the translocation of NF-κB into nucleus. These results suggest that SCM attenuated albumin-induced EMT in renal tubular cells via inhibiting NF-κB activation and inflammation, which may serve as a new therapeutic approach for chronic kidney diseases. (Supported by NIH grant HL89563 and HL106042)

Sperm survival kinetics in different types of bull semen: progressive motility, plasma membrane integrity, acrosomal status and reactive oxygen species generation

2015 ◽  
Vol 27 (5) ◽  
pp. 784 ◽  
Author(s):  
Mushtaq Ahmad ◽  
Nasim Ahmad ◽  
Amjad Riaz ◽  
Muhammad Anzar
Keyword(s):  
Plasma Membrane ◽  
Membrane Permeability ◽  
Sperm Motility ◽  
Membrane Integrity ◽  
Reactive Oxygen Species Generation ◽  
Plasma Membrane Permeability ◽  
Plasma Membrane Integrity ◽  
Bull Semen ◽  
Different Types ◽  
Sperm Survival

This study was designed to compare the kinetics of sperm survival in different types of bull semen. Fresh ejaculates from four bulls were pooled, diluted in Tris-citric acid-egg yolk-glycerol extender, cooled to 4°C, frozen in LN2 and thawed at 37°C. Fresh, diluted, cooled and frozen–thawed semen were incubated at 37°C, and evaluated at 0, 2, 4, 6, 12 and 24 h after the beginning of incubation. In Experiment 1, progressive sperm motility, normal acrosomes and plasma membrane integrity and asymmetry were determined. In Experiment 2, generation of superoxide anion (O2•) along with plasma membrane permeability and generation of hydrogen peroxide (H2O2) along with plasma membrane integrity were assessed. In Experiment 1, frozen–thawed semen had shorter survival times for progressive sperm motility, and spermatozoa with intact plasma membranes and acrosomes (IPM-IACR) as compared with other types of semen (P < 0.05). Fresh spermatozoa underwent a necrotic pathway, diluted and cooled spermatozoa underwent an apoptosis-like pathway and frozen–thawed spermatozoa underwent both necrotic and apoptosis-like pathways. In Experiment 2, spermatozoa in all four types of semen exhibited O2•– generation and increased plasma membrane permeability, and became necrotic without H2O2 generation during incubation (P < 0.05). In conclusion, frozen–thawed semen had shorter sperm longevity, which has important implications relating to the timing of artificial insemination. Different types of semen followed different death pathways. During incubation, spermatozoa in all types of semen generated O2•–, which increased the permeability and compromised the integrity of the plasma membrane.

A tubule cell model for ifosfamide nephrotoxicity

2005 ◽  
Vol 83 (6) ◽  
pp. 499-508 ◽  
Author(s):  
Katarina Aleksa ◽  
Naomi Halachmi ◽  
Shinya Ito ◽  
Gideon Koren
Keyword(s):  
Cell Model ◽  
Tubule Cell ◽  
Tubular Cells ◽  
Renal Tubular Cells ◽  
Cyp Enzymes ◽  
Affect Cell Viability ◽  
Renal Proximal Tubular Cells ◽  
Renal Proximal Tubular ◽  
Renal Tubular ◽  
Pharmacological Method

Mechanisms leading to ifosfamide (IF)-induced renal damage have not been fully elucidated. Recent work suggests that localized renal tubular metabolism of IF and the production of the nephrotoxic chloroacetaldehyde may lead to nephrotoxicity. Presently no pharmacological method to reduce IF nephrotoxicity has been identified. The objectives of this study were to establish a tubule cell model for IF nephrotoxicity, to verify whether renal proximal tubular cells have the necessary cytochrome P450 (CYP) enzymes to oxidize IF, and whether they can metabolize IF to chloroacetaldehyde. CYP3A, and 2B mRNA and protein were identified in LLCPK-1 cells. The cells metabolized the R- and S-IF enantiomers to their respective 2- and 3-dechloroethylifosfamide metabolites, by-products of chloroacetal dehyde formation. Metabolite production was both time and concentration-dependent. IF did not affect cell viability. In contrast, glutathione-depleted cells showed time and dose-dependent damage. The presence of the relevant CYP enzymes in renal tubular cells along with their ability to metabolize IF to its 2- and 3-dechloroethylifosfamide metabolites suggests that nephrotoxic damage may result from the localized production of chloroacetaldehyde. Glutathione is a major defence mechanism against IF toxicity, thus pharmacological methods for replenishing intracellular glutathione may be effective in modulating IF-induced nephrotoxicity. Key words: LLCPK-1, metabolism, ifosfamide, renal, CYP3A, CYP2B.

scholarly journals Canagliflozin reduces cisplatin uptake and activates Akt to protect against cisplatin-induced nephrotoxicity

2020 ◽  
Vol 318 (4) ◽  
pp. F1041-F1052
Author(s):  
Zhixia Song ◽  
Jiefu Zhu ◽  
Qingqing Wei ◽  
Guie Dong ◽  
Zheng Dong
Keyword(s):  
Protective Effect ◽  
Blood Glucose Control ◽  
Mitochondrial Pathway ◽  
Akt Inhibitor ◽  
Renal Tubular Cells ◽  
Survival Pathways ◽  
Proximal Tubular ◽  
Renal Proximal Tubular ◽  
Renal Tubular

Cisplatin is a widely used chemotherapy drug with notorious nephrotoxicity. Na+-glucose cotransporter 2 inhibitors are a class of novel antidiabetic agents that may have other effects in the kidneys besides blood glucose control. In the present study, we demonstrated that canagliflozin significantly attenuates cisplatin-induced nephropathy in C57BL/6 mice and suppresses cisplatin induced renal proximal tubular cell apoptosis in vitro. The protective effect of canagliflozin was associated with inhibition of p53, p38 and JNK activation. Mechanistically, canagliflozin partially reduced cisplatin uptake by kidney tissues in mice and renal tubular cells in culture. In addition, canagliflozin enhanced the activation of Akt and inhibited the mitochondrial pathway of apoptosis during cisplatin treatment. The protective effect of canagliflozin was diminished by the phosphatidylinositol 3-kinase/Akt inhibitor LY294002. Notably, canagliflozin did not affect the chemotherapeutic efficacy of cisplatin in A549 and HCT116 cancer cell lines. These results suggest a new application of canagliflozin for renoprotection in cisplatin chemotherapy. Canagliflozin may protect kidneys by reducing cisplatin uptake and activating cell survival pathways.

scholarly journals Autophagy Deficiency in Renal Proximal Tubular Cells Leads to an Increase in Cellular Injury and Apoptosis under Normal Fed Conditions

2019 ◽  
Vol 21 (1) ◽  
pp. 155 ◽  
Author(s):  
Chigure Suzuki ◽  
Isei Tanida ◽  
Juan Alejandro Oliva Trejo ◽  
Soichiro Kakuta ◽  
Yasuo Uchiyama
Keyword(s):  
Renal Injury ◽  
Kidney Injury ◽  
Renal Tissue ◽  
Proximal Tubular Cells ◽  
Tubular Cells ◽  
Renal Tubular Cells ◽  
Proximal Tubular ◽  
Renal Proximal Tubular Cells ◽  
Renal Proximal Tubular ◽  
Renal Tubular

Renal proximal tubular epithelial cells are significantly damaged during acute kidney injury. Renal proximal tubular cell-specific autophagy-deficient mice show increased sensitivity against renal injury, while showing few pathological defects under normal fed conditions. Considering that autophagy protects the proximal tubular cells from acute renal injury, it is reasonable to assume that autophagy contributes to the maintenance of renal tubular cells under normal fed conditions. To clarify this possibility, we generated a knock out mouse model which lacks Atg7, a key autophagosome forming enzyme, in renal proximal tubular cells (Atg7flox/flox;KAP-Cre+). Analysis of renal tissue from two months old Atg7flox/flox;KAP-Cre+ mouse revealed an accumulation of LC3, binding protein p62/sequestosome 1 (a selective substrate for autophagy), and more interestingly, Kim-1, a biomarker for early kidney injury, in the renal proximal tubular cells under normal fed conditions. TUNEL (TdT-mediated dUTP Nick End Labeling)-positive cells were also detected in the autophagy-deficient renal tubular cells. Analysis of renal tissue from Atg7flox/flox;KAP-Cre+ mice at different age points showed that tubular cells positive for p62 and Kim-1 continually increase in number in an age-dependent manner. Ultrastructural analysis of tubular cells from Atg7flox/flox;KAP-Cre+ revealed the presence of intracellular inclusions and abnormal structures. These results indicated that autophagy-deficiency in the renal proximal epithelial tubular cells leads to an increase in injured cells in the kidney even under normal fed conditions.

scholarly journals Atrasentan increased the expression of klotho by mediating miR-199b-5p and prevented renal tubular injury in diabetic nephropathy

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Wen-Ling Kang ◽  
Gao-Si Xu
Keyword(s):  
Diabetic Nephropathy ◽  
High Glucose ◽  
Tubular Injury ◽  
Mice Model ◽  
Renal Tubular Cells ◽  
Proximal Tubular ◽  
Renal Proximal Tubular ◽  
Renal Tubular

Abstract Atrasentan is a promising therapy for treating diabetic nephropathy (DN). Here we evaluated whether atrasentan down-regulated the miR-199b-5p expression, thereby increasing klotho and preventing renal tubular injury in DN. One-hundred patients with type 2 diabetes mellitus (T2DM) and 40 healthy subjects were included. A DN mice model was established by an injection of streptozotocin (STZ). Human renal proximal tubular epithelial HK-2 cells were exposed to high glucose (20 mmol/L). Treated the mice and HK-2 cells with atrasentan and we then investigated whether and how miR-199b-5p and Klotho were involved in preventing renal tubular injury in DN. In patients, the serum miR-199b-5p level increased and the klotho concentration decreased in accordance with elevated albuminuria. Atrasentan down-regulated miR-199b-5p and up-regulated klotho of the DN mice and HK-2 cells exposed to high glucose. High glucose promoted the binding of histone H3 to the miR-199b-5p promoter and atrasentan canceled this effect. MiR-199b-5p targeted the 3′ UTR of klotho. Overexpression of miR-199b-5p canceled the effects of atrasentan on klotho expression and apoptosis of renal tubular cells in both in vivo and in vitro. The increased serum klotho, mediated by miR-199b-5p, is a possible mechanism by which atrasentan prevents renal tubular injury in DN.

scholarly journals Perfluorooctane sulfonate induces autophagy-associated apoptosis through oxidative stress and the activation of extracellular signal–regulated kinases in renal tubular cells

PLoS ONE ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. e0245442
Author(s):  
Li-Li Wen ◽  
Yen-Ting Chen ◽  
Yuan-Chii Gladys Lee ◽  
Tsui-Ling Ko ◽  
Hsiu-Chu Chou ◽  
...  
Keyword(s):  
Perfluorooctane Sulfonate ◽  
Mitogen Activated Protein Kinase ◽  
Prolonged Treatment ◽  
Autophagic Flux ◽  
Protein Accumulation ◽  
Acridine Orange Staining ◽  
Tubular Cells ◽  
Renal Tubular Cells ◽  
Extracellular Signal ◽  
Renal Tubular

Perfluorooctane sulfonate (PFOS) is among the most abundant organic pollutants and is widely distributed in the environment, wildlife, and humans. Its toxic effects and biological hazards are associated with its long elimination half-life in humans. However, how it affects renal tubular cells (RTCs) remains unclear. In this study, PFOS was observed to mediate the increase in reactive oxygen species (ROS) generation, followed by the activation of the extracellular-signal-regulated kinase 1/2 (ERK1/2) pathway, which induced autophagy in RTCs. Although PFOS treatment induced autophagy after 6 h, prolonged treatment (24 h) reduced the autophagic flux by increasing lysosomal membrane permeability (LMP), leading to increased p62 protein accumulation and subsequent apoptosis. The increase in LMP was visualized through increased green fluorescence with acridine orange staining, and this was attenuated by 3-methyladenine, an autophagy inhibitor. N-acetyl cysteine and an inhibitor of the mitogen-activated protein kinase kinases (U0126) attenuated autophagy and apoptosis. Taken together, these results indicate that ROS activation and ROS-mediated phosphorylated ERK1/2 activation are essential to activate autophagy, resulting in the apoptosis of PFOS-treated RTCs. Our findings provide insight into the mechanism of PFOS-mediated renal toxicity.

scholarly journals Empagliflozin Ameliorates Free Fatty Acid Induced-Lipotoxicity in Renal Proximal Tubular Cells via the PPARγ/CD36 Pathway in Obese Mice

2021 ◽  
Vol 22 (22) ◽  
pp. 12408
Author(s):  
Chiang-Chi Huang ◽  
Chia-An Chou ◽  
Wei-Yu Chen ◽  
Jenq-Lin Yang ◽  
Wen-Chin Lee ◽  
...  
Keyword(s):  
Body Weight Gain ◽  
Specific Inhibitor ◽  
Proximal Tubular Cells ◽  
Tubular Cells ◽  
Renal Tubular Cells ◽  
Cd36 Expression ◽  
Proximal Tubular ◽  
Renal Proximal Tubular Cells ◽  
Renal Proximal Tubular ◽  
Renal Tubular

High serum levels of free fatty acids (FFAs) could contribute to obesity-induced nephropathy. CD36, a class B scavenger receptor, is a major receptor mediating FFA uptake in renal proximal tubular cells. Empagliflozin, a new anti-diabetic agent, is a specific inhibitor of sodium-glucose co-transporter 2 channels presented on renal proximal tubular cells and inhibits glucose reabsorption. In addition, empagliflozin has shown renoprotective effects. However, the mechanism through which empagliflozin regulates CD36 expression and attenuates FFA-induced lipotoxicity remains unclear. Herein, we aimed to elucidate the crosstalk between empagliflozin and CD36 in FFA-induced renal injury. C57BL/6 mice fed a high-fat diet (HFD) and palmitic acid-treated HK-2 renal tubular cells were used for in vivo and in vitro assessments. Empagliflozin attenuated HFD-induced body weight gain, insulin resistance, and inflammation in mice. In HFD-fed mice, CD36 was upregulated in the tubular area of the kidney, whereas empagliflozin attenuated CD36 expression. Furthermore, empagliflozin downregulated the expression of peroxisome proliferator-activated receptor (PPAR)-γ. Treatment with a PPARγ inhibitor (GW9662) did not further decrease PPARγ expression, whereas a PPARγ antagonist reversed this effect; this suggested that empagliflozin may, at least partly, decrease CD36 by modulating PPARγ. In conclusion, empagliflozin can ameliorate FFA-induced renal tubular injury via the PPARγ/CD36 pathway.

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