Accumulating evidence suggests that manganese oxide nanoparticles (NPs) show multiple enzyme-mimicking antioxidant activities, which supports their potential in redox-targeting therapeutic strategies for diseases with impaired redox signaling. However, the systemic administration of any NP requires thorough hemocompatibility testing. In this study, we assessed the hemocompatibility of synthesized Mn3O4 NPs, identifying their ability to induce spontaneous hemolysis and eryptosis or impair osmotic fragility. Concentrations of up to 20 mg/L were found to be safe for erythrocytes. Eryptosis assays were shown to be more sensitive than hemolysis and osmotic fragility as markers of hemocompatibility for Mn3O4 NP testing. Flow cytometry- and confocal microscopy-based studies revealed that eryptosis induced by Mn3O4 NPs was accompanied by Ca2+ overload, altered redox homeostasis verified by enhanced intracellular reactive oxygen species (ROS) and reactive nitrogen species (RNS), and a decrease in the lipid order of cell membranes. Furthermore, Mn3O4 NP-induced eryptosis was calpain- and caspase-dependent.

• MeSH
• buněčná membrána * metabolismus   účinky léků   MeSH
• eryptóza * účinky léků   MeSH
• erytrocyty účinky léků   metabolismus   MeSH
• hemolýza účinky léků   MeSH
• kalpain * metabolismus   MeSH
• kaspasy * metabolismus   MeSH
• lidé MeSH
• nanočástice * chemie   MeSH
• oxidy * farmakologie   chemie   MeSH
• reaktivní formy dusíku * metabolismus   MeSH
• reaktivní formy kyslíku * metabolismus   MeSH
• sloučeniny manganu * farmakologie   chemie   MeSH
• vápník * metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Hemolysis and eryptosis contribute to anemia encountered in patients undergoing chemotherapy. Eicosapentaenoic acid (EPA) is an omega-3 dietary fatty acid that has anticancer potential by inducing apoptosis in cancer cells, but its effect on the physiology and lifespan of red blood cells (RBCs) is understudied. Human RBCs were exposed to anticancer concentrations of EPA (10-100 ?M) for 24 h at 37 °C. Acetylcholinesterase (AChE) activity and hemolysis were measured by colorimetric assays whereas annexin-V-FITC and forward scatter (FSC) were employed to identify eryptotic cells. Oxidative stress was assessed by H2DCFDA and intracellular Ca2+ was measured by Fluo4/AM. EPA significantly increased hemolysis and K+ leakage, and LDH and AST activities in the supernatants in a concentration-dependent manner. EPA also significantly increased annexin-V-FITC-positive cells and Fluo4 fluorescence and decreased FSC and AChE activity. A significant reduction in the hemolytic activity of EPA was noted in the presence extracellular isosmotic urea, 125 mM KCl, and polyethylene glycol 8000 (PEG 8000), but not sucrose. In conclusion, EPA stimulates hemolysis and eryptosis through Ca2+ buildup and AChE inhibition. Urea, blocking KCl efflux, and PEG 8000 alleviate the hemolytic activity of EPA. The anticancer potential of EPA may be optimized using Ca2+ channel blockers and chelators to minimize its toxicity to off-target tissue. Keywords: EPA, Eryptosis, Hemolysis, Calcium, Anticancer.

• MeSH
• acetylcholinesterasa metabolismus   MeSH
• cholinesterasové inhibitory * farmakologie   MeSH
• eryptóza účinky léků   MeSH
• erytrocytární membrána * účinky léků   metabolismus   MeSH
• erytrocyty účinky léků   metabolismus   MeSH
• fosfatidylseriny * metabolismus   MeSH
• hemolýza * účinky léků   MeSH
• kyselina eikosapentaenová * farmakologie   MeSH
• lidé MeSH
• vápník metabolismus   MeSH
• vápníková signalizace * účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Hemocompatibility evaluation is an important step in nanotoxicological studies. It is generally accepted that nanomaterials promote lysis of erythrocytes, blood clotting, alter phagocytosis, and upregulate pro-inflammatory cytokines. However, there are no standardized guidelines for testing nanomaterials hemocompatibility despite the fact that nanomaterials enter the bloodstream and interact with blood cells. In this review, the current knowledge on the ability of nanomaterials to induce distinct cell death modalities of erythrocytes is highlighted primarily focusing on hemolysis and eryptosis. This review aims to summarize the molecular mechanisms underlying erythrotoxicity of nanomaterials and critically compare the sensitivity and efficiency of hemolysis or eryptosis assays for nanomaterials blood compatibility testing. The list of eryptosis-inducing nanomaterials is growing, but it is still difficult to generalize how physico-chemical properties of nanoparticles affect eryptosis degree and molecular mechanisms involved. Thus, another aim of this review is to raise the awareness of eryptosis as a nanotoxicological tool to encourage the corresponding studies. It is worthwhile to consider adding eryptosis to in vitro nanomaterials hemocompatibility testing protocols and guidelines.

• MeSH
• eryptóza * účinky léků   MeSH
• erytrocyty účinky léků   MeSH
• hemolýza * účinky léků   MeSH
• lidé MeSH
• nanostruktury * toxicita   MeSH
• testování materiálů metody   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Docosahexaenoic acid (DHA) is an omega-3 polyunsaturated fatty acid with promising anticancer potential. Anaemia is a frequent adverse effect of anticancer treatment caused in part by eryptosis and haemolysis. Thus, it is important to investigate the role of DHA in red blood cell (RBC) death. RBCs were treated with anticancer concentrations (10-100 μM) of DHA under different physiological conditions, and fluorescence-assisted cell sorting was employed to measure eryptotic markers. Cell membrane scrambling was detected by annexin-V-FITC labelling, cytoplasmic Ca2+ by Fluo4/AM, cell size by forward scatter (FSC), and oxidative stress by H2DCFDA. Haemolytic markers were also assayed by photometric methods. DHA caused significant phospholipid scrambling with Ca2+ accumulation, loss of cellular volume, and oxidative stress. These changes were associated with dacrocyte formation, as revealed by electron microscopy. Moreover, DHA exhibited a dual effect on membrane integrity: it was haemolytic under isotonic conditions and anti-haemolytic in hypotonic environments. Importantly, inhibition of Rac1 GTPase activity with NSC23766 significantly reduced DHA-mediated haemolysis, as did co-administration of either sucrose or polyethylene glycol 8,000. Conversely, the presence of 125 mM KCl and urea without extracellular Ca2+ significantly exacerbated DHA toxicity. In conclusion, this is the first report that identifies key biochemical mechanisms underlying the cytotoxic effects of DHA in RBCs, promoting further development and validation of DHA in anticancer therapy.

• MeSH
• eryptóza * účinky léků   MeSH
• erytrocyty účinky léků   metabolismus   MeSH
• hemolýza * účinky léků   MeSH
• kyseliny dokosahexaenové * farmakologie   MeSH
• lidé MeSH
• oxidační stres * účinky léků   MeSH
• signální transdukce účinky léků   MeSH
• vápník * metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Costunolide, a natural sesquiterpene lactone, has multiple pharmacological activities such as neuroprotection or induction of apoptosis and eryptosis. However, the effects of costunolide on pro-survival factors and enzymes in human erythrocytes, e.g. glutathione and glucose-6-phosphate dehydrogenase (G6PDH) respectively, have not been studied yet. Our aim was to determine the mechanisms underlying costunolide-induced eryptosis and to reverse this process. Phosphatidylserine exposure was estimated from annexin-V-binding, cell volume from forward scatter in flow cytometry, and intracellular glutathione [GSH]i from high performance liquid chromatography. The oxidized status of intracellular glutathione and enzyme activities were measured by spectrophotometry. Treatment of erythrocytes with costunolide dose-dependently enhanced the percentage of annexin-V-binding cells, decreased the cell volume, depleted [GSH]i and completely inhibited G6PDH activity. The effects of costunolide on annexin-V-binding and cell volume were significantly reversed by pre-treatment of erythrocytes with the specific PKC-α inhibitor chelerythrine. The latter, however, had no effect on costunolide-induced GSH depletion. Costunolide induces eryptosis, depletes [GSH]i and inactivates G6PDH activity. Furthermore, our study reveals an inhibitory effect of chelerythrine on costunolide-induced eryptosis, indicating a relationship between costunolide and PKC-α. In addition, chelerythrine acts independently of the GSH depletion. Understanding the mechanisms of G6PDH inhibition accompanied by GSH depletion should be useful for development of anti-malarial therapeutic strategies or for synthetic lethality-based approaches to escalate oxidative stress in cancer cells for their sensitization to chemotherapy and radiotherapy.

• MeSH
• apoptóza účinky léků   MeSH
• benzofenantridiny farmakologie   MeSH
• eryptóza účinky léků   genetika   MeSH
• erytrocyty účinky léků   patologie   MeSH
• glukosa-6-fosfátdehydrogenasa antagonisté a inhibitory   genetika   MeSH
• glutathion genetika   MeSH
• inhibitory enzymů farmakologie   MeSH
• lidé MeSH
• oxidační stres účinky léků   MeSH
• proteinkinasa C-alfa antagonisté a inhibitory   genetika   MeSH
• reaktivní formy kyslíku MeSH
• seskviterpeny farmakologie   MeSH
• vápník metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

In mature erythrocytes, glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase (6PGDH) yield NADPH, a crucial cofactor of the enzyme glutathione reductase (GR) converting glutathione disulfide (GSSG) into its reduced state (GSH). GSH is essential for detoxification processes in and survival of erythrocytes. We explored whether the anti-inflammatory compounds Bay 11-7082, parthenolide and dimethyl fumarate (DMF) were able to completely deplete a common target (GSH), and to impair the function of upstream enzymes of GSH recycling and replenishment. Treatment of erythrocytes with Bay 11-7082, parthenolide or DMF led to concentration-dependent eryptosis resulting from complete depletion of GSH. GSH depletion was due to strong inhibition of G6PDH activity. Bay 11-7082 and DMF, but not parthenolide, were able to inhibit the GR activity. This approach "Inhibitors, Detection of their common target that is completely depleted or inactivated when pharmacologically relevant concentrations of each single inhibitor are applied, Subsequent functional analysis of upstream enzymes for this target" (IDS), can be applied to a broad range of inhibitors and cell types according to the selected target. The specific G6PDH inhibitory effect of these compounds may be exploited for the treatment of human diseases with high NADPH and GSH consumption rates, including malaria, trypanosomiasis, cancer or obesity.

• MeSH
• dimethyl fumarát farmakologie   MeSH
• eryptóza účinky léků   MeSH
• erytrocyty enzymologie   MeSH
• glukosa-6-fosfátdehydrogenasa * antagonisté a inhibitory   metabolismus   MeSH
• lidé MeSH
• nitrily farmakologie   MeSH
• seskviterpeny farmakologie   MeSH
• sulfony farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH