scholarly journals ROS-Mediated Necroptosis Is Involved in Iron Overload-Induced Osteoblastic Cell Death

2020 ◽  
Vol 2020 ◽  
pp. 1-22 ◽  
Author(s):  
Qing Tian ◽  
Bo Qin ◽  
Yufan Gu ◽  
Lijun Zhou ◽  
Songfeng Chen ◽  
...  
Keyword(s):  
Cell Death ◽  
Iron Overload ◽  
Mitochondrial Permeability Transition ◽  
Cell Damage ◽  
Critical Role ◽  
Osteoblastic Cells ◽  
Osteoblastic Cell ◽  
Ros Generation ◽  
Mptp Opening

Excess iron has been reported to lead to osteoblastic cell damage, which is a crucial pathogenesis of iron overload-related osteoporosis. However, the cytotoxic mechanisms have not been fully documented. In the present study, we focused on whether necroptosis contributes to iron overload-induced osteoblastic cell death and related underlying mechanisms. Here, we showed that the cytotoxicity of iron overload in osteoblastic cells was mainly due to necrosis, as evidenced by the Hoechst 33258/PI staining, Annexin-V/PI staining, and transmission electronic microscopy. Furthermore, we revealed that iron overload-induced osteoblastic necrosis might be mediated via the RIPK1/RIPK3/MLKL necroptotic pathway. In addition, we also found that iron overload was able to trigger mitochondrial permeability transition pore (mPTP) opening, which is a critical downstream event in the execution of necroptosis. The key finding of our experiment was that iron overload-induced necroptotic cell death might depend on reactive oxygen species (ROS) generation, as N-acetylcysteine effectively rescued mPTP opening and necroptotic cell death. ROS induced by iron overload promote necroptosis via a positive feedback mechanism, as on the one hand N-acetylcysteine attenuates the upregulation of RIPK1 and RIPK3 and phosphorylation of RIPK1, RIPK3, and MLKL and on the other hand Nec-1, siRIPK1, or siRIPK3 reduced ROS generation. In summary, iron overload induced necroptosis of osteoblastic cells in vitro, which is mediated, at least in part, through the RIPK1/RIPK3/MLKL pathway. We also highlight the critical role of ROS in the regulation of iron overload-induced necroptosis in osteoblastic cells.

scholarly journals RIPK3 Induces Cardiomyocyte Necroptosis via Inhibition of AMPK-Parkin-Mitophagy in Cardiac Remodelling after Myocardial Infarction

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Pingjun Zhu ◽  
Kun Wan ◽  
Ming Yin ◽  
Peng Hu ◽  
Yifan Que ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Mitochondrial Permeability Transition ◽  
Critical Role ◽  
Permeability Transition ◽  
Cardiac Remodelling ◽  
Coronary Artery Ligation ◽  
Artery Ligation ◽  
Genetic Ablation ◽  
Mptp Opening

Receptor-interacting protein 3- (RIPK3-) modulated necroptosis plays a critical role in cardiac remodelling after myocardial infarction (MI). However, the precise regulatory mechanism is not fully elucidated yet. In the present study, we showed that RIPK3 expression was upregulated in myocardial tissue after MI in a mouse model by coronary artery ligation, as well as in the cardiomyocytes following hypoxic injury in vitro. The increase of RIPK3 expression was found to be accompanied by severe cardiac remodelling, cardiac dysfunction, and higher mortality. Elevated RIPK3 expression subsequently abrogated the AMPK pathway that was accompanied by inhibition of Parkin-mediated mitophagy. Loss of mitophagy increased the opening of mitochondrial permeability transition pore (mPTP), which ultimately induced the cardiomyocyte necroptosis. In contrast, genetic ablation of Ripk3 induced the AMPK/Parkin-mitophagy pathway, favouring a prosurvival state that eventually inhibited mPTP opening and induced the necroptosis of cardiomyocytes in the post-MI cardiac remodelling. In conclusion, our results revealed a key mechanism by which necroptosis could be mediated by RIPK3 via the AMPK/Parkin-mitophagy/mPTP opening axis, which provides a potential therapeutic target in the management of heart failure after MI.

Abstract 294: Mitochondrial Inner Membrane Protein Plays a Critical Role in the Ischemic Myocardial Dysfunction and Determines the Outcome of Cardiac Recovery

2017 ◽  
Vol 121 (suppl_1) ◽  
Author(s):  
Jean C Bopassa
Keyword(s):  
Mitochondrial Permeability Transition ◽  
Myocardial Dysfunction ◽  
Myocardial Infarct ◽  
Ischemia Reperfusion ◽  
Critical Role ◽  
Ros Generation ◽  
Myocardial Infarct Size ◽  
Retention Capacity ◽  
Mptp Opening ◽  
The Impact

Disorders characterized by ischemia/reperfusion (I/R), such as myocardial infarction and stroke, is the leading cause of death in the US. Indeed, reoxygenation of cardiomyocytes after an ischemic insult leads to mitochondrial Ca 2+ overload and increase in ROS generation that triggers the opening of mitochondrial permeability transition pore (mPTP), making this event crucial in the mechanism of cell death after I/R. Several proteins have been proposed contributing to formation and function of mPTP, but its exact molecular identity and mechanism still need to be elucidated. Cyclophylin D (CypD) deleted mice displayed a marked desensitization of the mPTP to Ca 2+ overload and oxidative stress establishing it as a key component of mPTP. However, CypD null mitochondria still exhibit mPTP activity. Using 2D-DIGE and mass spectrometry, we identified mitofilin, which plays an important role of maintaining mitochondrial cristae morphology, as a protein that the expression is reduced after I/R versus sham. We thus investigated the impact of mitofilin regulation in the mPTP formation. We found that: i) mitofilin -/- mice subjected to I/R exhibited an increase in myocardial infarct size, a reduced cardiac functional recovery, a reduced mitochondrial Ca 2+ retention capacity required to induce the mPTP opening and an increase in mitophagy compared to WT; ii) Knockdown of mitofilin in H9c2 myoblasts with siRNA led to: increase in apoptosis via activation of AIF-PARP1 pathway that is associated with S phase arrest of the cell cycle and nuclear fragmentation, increase in mitochondrial cristae disorganization, in ROS production and Calpain activity, as well as decrease in intracellular ATP production and mitochondrial membrane potential versus scramble siRNA; iii) mitofilin structurally bound to CypD and this interaction is abridged after the mPTP opening triggered by Ca 2+ overload after I/R. These results indicate that mitofilin plays an essential role in the mechanism of mPTP formation and cardiomyocyte death.

scholarly journals Pioglitazone Is a Mild Carrier-Dependent Uncoupler of Oxidative Phosphorylation and a Modulator of Mitochondrial Permeability Transition

2021 ◽  
Vol 14 (10) ◽  
pp. 1045
Author(s):  
Ekaterina S. Kharechkina ◽  
Anna B. Nikiforova ◽  
Konstantin N. Belosludtsev ◽  
Tatyana I. Rokitskaya ◽  
Yuri N. Antonenko ◽  
...  
Keyword(s):  
Adverse Effects ◽  
Oxidative Phosphorylation ◽  
Mitochondrial Permeability Transition ◽  
Kidney Diseases ◽  
Adenine Nucleotides ◽  
Permeability Transition ◽  
Protective Effects ◽  
Retention Capacity ◽  
Mptp Opening

Pioglitazone (PIO) is an insulin-sensitizing antidiabetic drug, which normalizes glucose and lipid metabolism but may provoke heart and liver failure and chronic kidney diseases. Both therapeutic and adverse effects of PIO can be accomplished through mitochondrial targets. Here, we explored the capability of PIO to modulate the mitochondrial membrane potential (ΔΨm) and the permeability transition pore (mPTP) opening in different models in vitro. ΔΨm was measured using tetraphenylphosphonium and the fluorescent dye rhodamine 123. The coupling of oxidative phosphorylation was estimated polarographically. The transport of ions and solutes across membranes was registered by potentiometric and spectral techniques. We found that PIO decreased ΔΨm in isolated mitochondria and intact thymocytes and the efficiency of ADP phosphorylation, particularly after the addition of Ca2+. The presence of the cytosolic fraction mitigated mitochondrial depolarization but made it sustained. Carboxyatractyloside diminished the PIO-dependent depolarization. PIO activated proton transport in deenergized mitochondria but not in artificial phospholipid vesicles. PIO had no effect on K+ and Ca2+ inward transport but drastically decreased the mitochondrial Ca2+-retention capacity and protective effects of adenine nucleotides against mPTP opening. Thus, PIO is a mild, partly ATP/ADP-translocase-dependent, uncoupler and a modulator of ATP production and mPTP sensitivity to Ca2+ and adenine nucleotides. These properties contribute to both therapeutic and adverse effects of PIO.

Cultures of human osteoblastic cells from dialysis patients: Influence of bone turnover rate on in vitro secretion of interleukin-6 and osteoblastic cell markers

2001 ◽  
Vol 37 (1) ◽  
pp. 30-37 ◽  
Author(s):  
M. Carmen Sánchez ◽  
M. Auxiliadora Bajo ◽  
Rafael Selgas ◽  
Alberto Mate ◽  
M. Jesús Sánchez-Cabezudo ◽  
...  
Keyword(s):  
Bone Turnover ◽  
Interleukin 6 ◽  
Turnover Rate ◽  
Osteoblastic Cells ◽  
Osteoblastic Cell ◽  
Dialysis Patients ◽  
Cell Markers ◽  
Human Osteoblastic Cells

Abstract 322: Metformin Inhibits Mitochondrial Permeability Transition Pore Opening in Human Cardiac Mitochondria

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
Larisa Emelyanova ◽  
Sirisha Gudlawar ◽  
Milanka Cosic ◽  
Mahek Mirza ◽  
Frahan Rizvi ◽  
...  
Keyword(s):  
Cell Death ◽  
Membrane Potential ◽  
Fluorescent Dyes ◽  
Mitochondrial Permeability Transition ◽  
Permeability Transition ◽  
Cardiac Mitochondria ◽  
Oral Antidiabetic Agent ◽  
Major Health Problem ◽  
Mptp Opening ◽  
Safranin O

Background: Type II diabetes mellitus is a major health problem contributing to increased morbidity and mortality with associated cardiovascular diseases. Metformin, an oral antidiabetic agent, has cardioprotective properties independent of their glucose lowering effect; however, mechanisms underlying cardioprotection remain poorly defined. We hypothesized that the cardioprotective effect of metformin appears to be associated with inhibition of mitochondrial permability transition pore (mPTP) that could be beneficial in diabetic hearts with attenuated endogenous cardioprotective responses. Purpose: The aim of the study was to determine the protective effect of metformin on mPTP opening in mitochondria from human myocardium.. Methods: Mitochondria freshly isolated from the left atrial appendage of nondiabetic patients undergoing cardiac surgery were loaded with calcium- (fluo-3) and membrane potential-sensitive (Safranin-O) fluorescent dyes and challenged with sequential pulses of Ca2+ (10 μM every 3 m) in the absence and presence of different concentrations of metformin (2.5, 5, 10 mM). The sensitivity of mitochondria toward mPTP opening was assessed by abrupt release of mitochondrial Ca2+ (fluo-5N fluorescence), with simultaneous dissipation of mitochondrial membrane potential (safranin O fluorescence) and mitochondrial swelling (decrease in light scattering). Results: Metformin caused a dose-dependent inhibition of Ca2+-induced mPTP opening with delayed mitochondrial Ca2+ release, depolarization and swelling. The tolerance of metformin-treated mitochondria to Ca2+-induced mPTP opening when compared to controls was increased from 457±71 to 600±74 nmol/mg protein at 2.5mM, 674±10 nmol/mg protein at 5 mM and 750±77 nmol/mg protein at 10 mM. Cyclosporin A (1 μM), a known mPTP inhibitor, has marginal incremental effect on metformin-induced inhibition of mPTP opening. Conclusion: In human cardiac mitochondria, metformin inhibited calcium-overload-mediated mPTP opening that otherwise leads to mitochondrial energetic failure and cell death. Thus, metformin may help restore attenuated cardioprotection in diabetic hearts by increasing tolerance to stress-induced mPTP opening and, thus, preventing cell death.

Abstract 321: Obesity Causes Enhanced Sensitivity Of The Mitochondrial Permeability Transition Pore

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
Judith Bernal-Ramírez ◽  
Adriana Riojas-Hernández ◽  
Flor E Morales-Marroquín ◽  
Elvía M Domínguez-Barragán ◽  
David Rodríguez-Mier ◽  
...  
Keyword(s):  
Cell Death ◽  
Cardiac Dysfunction ◽  
Mitochondrial Permeability Transition Pore ◽  
Mitochondrial Permeability Transition ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
H2o2 Production ◽  
Mitochondrial Permeability ◽  
Removal Capacity ◽  
Mptp Opening

Several mechanisms have been implicated in heart failure (HF) development due to obesity, including altered Ca2+ homeostasis and mitochondrial increased reactive oxygen species (ROS). Besides their metabolic role, mitochondria are important cell death regulators, since their disruption induces apoptosis. The mitochondrial permeability transition pore (MPTP) formation is key in this process. Ca2+ and ROS are known inducers of MPTP, and mitochondria are the main ROS generators. However, it has not been demonstrated that MPTP formation is involved in cardiac cell death due to obesity. Therefore, the aim of this work was to determine whether Ca2+ alterations and/or MPTP opening underlie cardiac dysfunction. We used obese Zucker fa/fa rats (32 weeks old), displaying concentric hypertrophy and cardiac dysfunction. We measured: i) Systolic and diastolic Ca2+ signaling in isolated myocytes, in basal conditions and upon β-adrenergic stimulation (β-AS), and ii) in vitro mitochondrial function: respiration, ROS production and MPTP opening. We found that the main alteration in Ca2+ signaling in fa/fa myocytes was a decrease in SERCA Ca2+ removal capacity, since Ca2+ transient amplitude and spark frequency were unchanged. Furthermore, in fa/fa myocytes, β-AS response was preserved. On the other hand, fa/fa mitochondria respiration, in state 3 decreased, but was unchanged in state 4, when glutamate/malate were used as substrate, resulting in an small decrease in respiratory control. In addition, fa/fa mitochondria were more sensitive to MPTP opening, induced by Ca2+ and carboxyatractiloside (CAT). Moreover, fa/fa mitochondria showed increased H2O2 production, and in exposed thiol groups in the adenine nucleotide translocase, a regulatory MPTP component. Since Ca2+ signaling is relatively normal in fa/fa cells, it does not seem to be the main contributor to the cardiac contractile dysfunction. However, given that fa/fa mitochondria showed decrease respiratory performance, were more susceptible to MPTP opening, and showed enhanced H2O2 production. We conclude that fa/fa mitochondria were more vulnerable to enhanced oxidative stress, causing MPTP opening, which could be exacerbated by SERCA slower Ca2+ removal capacity, leading to myocyte apoptosis.

scholarly journals P1238EVALUATION OF AN IN VITRO CO-CULTURE MODEL FOR TESTING EFFECTS OF CYTOPROTECTIVE ADDITIVES IN PERITONEAL DIALYSIS FLUIDS ON CARDIOVASCULAR OUTCOME

2020 ◽  
Vol 35 (Supplement_3) ◽  
Author(s):  
Juan Manuel Sacnun ◽  
Rebecca Herzog ◽  
Maria Bartosova ◽  
Claus Schmitt ◽  
Klaus Kratochwill
Keyword(s):  
Endothelial Cells ◽  
Cell Death ◽  
Peritoneal Dialysis ◽  
Cardiovascular Risk ◽  
Cell Damage ◽  
Peritoneal Membrane ◽  
Cytoskeleton Reorganization ◽  
Harmful Effects

Abstract Background and Aims The composition of all currently available peritoneal dialysis (PD) fluids triggers morphological and functional changes in the peritoneal membrane. Periodic exposure leads to vasculopathy, hypervascularization, and diabetes-like damage of vessels, eventually leading to failure of the technique. Patients undergoing dialysis generally, have a high risk of cardiovascular events. It is currently unclear if there is a mechanistic link between peritoneal membrane failure and cardiovascular risk. In vitro and in vivo studies have shown that cytoprotective additives (e.g. dipeptide alanyl-glutamine (AlaGln) or kinase inhibitor lithium chloride (LiCl)) to PDF reduce peritoneal damage. Here, we developed an experimental model for investigating effects of these cytoprotective additives in PDF in the cardiovascular context. Method For modelling the peritoneal membrane in vitro, mesothelial and endothelial cells were co-cultured in transwell plates. Mesothelial cells were grown in the upper compartment and primary human umbilical vein endothelial cells (HUVEc) or primary microvascular cells were grown in the lower compartment. PDF with or without cytoprotective compounds, was added to the upper compartment to only expose mesothelial cells directly to different dilutions of the fluid. Effects on cell damage was assessed by quantification of lactate-dehydrogenase (LDH) release and live-dead staining of cells. Proteome profiles were analysed for both cell-types separately and in combination using two-dimensional difference gel electrophoresis (2D-DiGE) and liquid chromatography coupled to mass spectrometry (LC-MS). In vitro findings were related to PD-induced arteriolar changes based on abundance profiles of micro-dissected omental arterioles of children treated with conventional PD-fluids and age-matched controls with normal renal function. Results Marked cellular injury of HUVEc after PD-fluid exposure was associated with a molecular landscape of the enriched biological process clusters ‘glucose catabolic process’, ‘cell redox homeostasis’, ‘RNA metabolic process’, ‘protein folding’, ‘regulation of cell death’, and ‘actin cytoskeleton reorganization’ that characterize PD-fluid cytotoxicity and counteracting cellular repair process respectively. PDF-induced cell damage was reduced by AlaGln and LiCl both in mesothelial and endothelial cells. Proteome analysis revealed perturbation of major cellular processes including regulation of cell death and cytoskeleton reorganization. Selected markers of angiogenesis, oxidative stress, cell junctions and transdifferentiation were counter-regulated by the additives. Co-cultured cells yielded differently regulated pathways following PDF exposure compared to separate culture. Comparison to human arterioles confirmed overlapping protein regulation between endothelial cells in vitro and in vivo, proving harmful effects of PD-fluids on endothelial cells leading to drastic changes of the cellular process landscape. Conclusion In summary, this study shows harmful effects of PD-fluids also effecting endothelial cells and elucidates potential mechanisms by which cytoprotective additives may counteract the signalling axis between local peritoneal damage and systemic vasculopathy. An in vitro co-culture system may be an attractive approach to simulate the peritoneal membrane for testing direct and indirect effects of cytoprotective additives in PDF. When cultured and stressed in close proximity cells may respond differently. Characterisation of PD-induced perturbations will allow identifying molecular mechanisms linking the peritoneal and cardiovascular context, offering therapeutic targets to reduce current limitations of PD and ultimately decreasing cardiovascular risk of dialysis patients.

scholarly journals Effect of Chitosan Nanoparticle-Loaded Thymus serpyllum on Hydrogen Peroxide-Induced Bone Marrow Stromal Cell Damage

2019 ◽  
Vol 2019 ◽  
pp. 1-12
Author(s):  
Salma Baig ◽  
Ainnul Hamidah Syahadah Azizan ◽  
Hanumantha Rao Balaji Raghavendran ◽  
Elango Natarajan ◽  
Sangeetha Naveen ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Cell Death ◽  
Cell Damage ◽  
Radical Scavenging Activity ◽  
Radical Scavenging ◽  
Potential Candidate ◽  
Protective Effects ◽  
Active Components ◽  
Thymus Serpyllum

We have determined the protective effects of Thymus serpyllum (TS) extract and nanoparticle-loaded TS on hydrogen peroxide-induced cell death of mesenchymal stromal cells (MSCs) in vitro. Gas chromatography–mass spectroscopy confirmed the spectrum of active components in the extract. Out of the three different extracts, the hexane extract showed significant free radical scavenging activity. Treatment of MSCs with H2O2 (hydrogen peroxide) significantly increased intracellular cell death; however, pretreatment with TS extract and nanoparticle-loaded TS (200 μg/ml) suppressed H2O2-induced elevation of Cyt-c and MMP13 and increased the survival rates of MSCs. H2O2-induced (0.1 mM) changes in cytokines were attenuated in the extract and nanoparticles by pretreatment and cotreatment at two time points (p<0.05). H2O2 increased cell apoptosis. In contrast, treatment with nanoparticle-loaded TS suppressed the percentage of apoptosis considerably (p<0.05). Therefore, TS may be considered as a potential candidate for enhancing the effectiveness of MSC transplantation in cell therapy.

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