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
• Názvy látek
• acetylcholinesterasa MeSH
• cholinesterasové inhibitory * MeSH
• fosfatidylseriny * MeSH
• kyselina eikosapentaenová * MeSH
• vápník MeSH

Homeostasis of cellular membranes is maintained by fine-tuning their lipid composition. Yeast lipid transporter Osh6, belonging to the oxysterol-binding protein-related proteins family, was found to participate in the transport of phosphatidylserine (PS). PS synthesized in the endoplasmic reticulum is delivered to the plasma membrane, where it is exchanged for phosphatidylinositol 4-phosphate (PI4P). PI4P provides the driving force for the directed PS transport against its concentration gradient. In this study, we employed an in vitro approach to reconstitute the transport process into the minimalistic system of large unilamellar vesicles to reveal its fundamental biophysical determinants. Our study draws a comprehensive portrait of the interplay between the structure and dynamics of Osh6, the carried cargo lipid, and the physical properties of the involved membranes, with particular attention to the presence of charged lipids and to membrane fluidity. Specifically, we address the role of the cargo lipid, which, by occupying the transporter, imposes changes in its dynamics and, consequently, predisposes the cargo to disembark in the correct target membrane.

• MeSH
• biologický transport MeSH
• buněčná membrána * metabolismus   MeSH
• fluidita membrány MeSH
• fosfatidylinositolfosfáty metabolismus   MeSH
• fosfatidylseriny metabolismus   MeSH
• Saccharomyces cerevisiae - proteiny * metabolismus   genetika   MeSH
• Saccharomyces cerevisiae metabolismus   MeSH
• steroidní receptory metabolismus   MeSH
• unilamelární lipozómy metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfatidylinositolfosfáty MeSH
• fosfatidylseriny MeSH
• Oxysterol Binding Proteins MeSH
• phosphatidylinositol 4-phosphate MeSH Prohlížeč
• Saccharomyces cerevisiae - proteiny * MeSH
• steroidní receptory MeSH
• unilamelární lipozómy MeSH

Calmodulin (CaM) is a ubiquitous calcium-sensitive messenger in eukaryotic cells. It was previously shown that CaM possesses an affinity for diverse lipid moieties, including those found on CaM-binding proteins. These facts, together with our observation that CaM accumulates in membrane-rich protrusions of HeLa cells upon increased cytosolic calcium, motivated us to perform a systematic search for unmediated CaM interactions with model lipid membranes mimicking the cytosolic leaflet of plasma membranes. A range of experimental techniques and molecular dynamics simulations prove unambiguously that CaM interacts with lipid bilayers in the presence of calcium ions. The lipids phosphatidylserine (PS) and phosphatidylethanolamine (PE) hold the key to CaM-membrane interactions. Calcium induces an essential conformational rearrangement of CaM, but calcium binding to the headgroup of PS also neutralizes the membrane negative surface charge. More intriguingly, PE plays a dual role-it not only forms hydrogen bonds with CaM, but also destabilizes the lipid bilayer increasing the exposure of hydrophobic acyl chains to the interacting proteins. Our findings suggest that upon increased intracellular calcium concentration, CaM and the cytosolic leaflet of cellular membranes can be functionally connected.

• Klíčová slova
• calcium, calmodulin, lipid membrane, phosphatidylethanolamine, phosphatidylserine,
• MeSH
• buněčná membrána * metabolismus   MeSH
• cytosol * metabolismus   MeSH
• fosfatidylethanolaminy metabolismus   MeSH
• fosfatidylseriny * metabolismus   MeSH
• HeLa buňky MeSH
• kalmodulin * metabolismus   chemie   MeSH
• lidé MeSH
• lipidové dvojvrstvy * metabolismus   MeSH
• simulace molekulární dynamiky * MeSH
• vápník * metabolismus   MeSH
• vazba proteinů * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfatidylethanolaminy MeSH
• fosfatidylseriny * MeSH
• kalmodulin * MeSH
• lipidové dvojvrstvy * MeSH
• phosphatidylethanolamine MeSH Prohlížeč
• vápník * MeSH

Eryptosis is a regulated cell death (RCD) of mature erythrocytes initially described as a counterpart of apoptosis for enucleated cells. However, over the recent years, a growing number of studies have emphasized certain differences between both cell death modalities. In this review paper, we underline the hallmarks of eryptosis and apoptosis and highlight resemblances and dissimilarities between both RCDs. We summarize and critically discuss differences in the impact of caspase-3, Ca2+ signaling, ROS signaling pathways, opposing roles of casein kinase 1α, protein kinase C, Janus kinase 3, cyclin-dependent kinase 4, and AMP-activated protein kinase to highlight a certain degree of divergence between apoptosis and eryptosis. This review emphasizes the crucial importance of further studies that focus on deepening our knowledge of cell death machinery and identifying novel differences between cell death of nucleated and enucleated cells. This might provide evidence that erythrocytes can be defined as viable entities capable of programmed cell destruction. Additionally, the revealed cell type-specific patterns in cell death can facilitate the development of cell death-modulating therapeutic agents.

• Klíčová slova
• Ca2+ signaling, Casein kinase 1α, Caspase-3, Regulated cell death, p38 MAPK,
• MeSH
• apoptóza * MeSH
• buněčná smrt MeSH
• eryptóza * MeSH
• erytrocyty metabolismus   MeSH
• fosfatidylseriny metabolismus   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• signální transdukce MeSH
• vápník metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• fosfatidylseriny MeSH
• reaktivní formy kyslíku MeSH
• vápník MeSH

Lipid biosensors are molecular tools used both in vivo and in vitro applications, capable of selectively detecting specific types of lipids in biological membranes. However, despite their extensive use, there is a lack of systematic characterization of their binding properties in various membrane conditions. The purpose of this study was to investigate the impact of membrane properties, such as fluidity and membrane charge, on the sensitivity of two lipid biosensors, LactC2 and P4M, to their target lipids, phosphatidylserine (PS) or phosphatidylinositol 4-phosphate (PI4P), respectively. Dual-color fluorescence cross-correlation spectroscopy, employed in this study, provided a useful technique to investigate interactions of these recombinant fluorescent biosensors with liposomes of varying compositions. The results of the study demonstrate that the binding of the LactC2 biosensor to low levels of PS in the membrane is highly supported by the presence of anionic lipids or membrane fluidity. However, at high PS levels, the presence of anionic lipids does not further enhance binding of LactC2. In contrast, neither membrane charge, nor membrane fluidity significantly affect the binding affinity of P4M to PI4P. These findings provide valuable insights into the role of membrane properties on the binding properties of lipid biosensors.

• Klíčová slova
• LUVs, LactC2, Lipid biosensors, Membrane fluidity, Membrane properties, Negatively charged lipids, P4M, Phosphatidylinositol 4-phosphate, Phosphatidylserine,
• MeSH
• biosenzitivní techniky * MeSH
• buněčná membrána metabolismus   MeSH
• fosfatidylseriny * metabolismus   MeSH
• liposomy chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfatidylseriny * MeSH
• liposomy MeSH
• phosphatidylinositol 4-phosphate MeSH Prohlížeč

Phosphatidylserine (PS) lipids are important signaling molecules and the most common negatively charged lipids in eukaryotic membranes. The signaling can be often regulated by calcium, but its interactions with PS headgroups are not fully understood. Classical molecular dynamics (MD) simulations can potentially give detailed description of lipid-ion interactions, but the results strongly depend on the used force field. Here, we apply the electronic continuum correction (ECC) to the Amber Lipid17 parameters of 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-l-serine (POPS) lipid to improve its interactions with K+, Na+, and Ca2+ ions. The partial charges of the headgroup, glycerol backbone, and carbonyls of POPS, bearing a unit negative charge, were scaled with a factor of 0.75, derived for monovalent ions, and the Lennard-Jones σ parameters of the same segments were scaled with a factor of 0.89. The resulting ECC-POPS model gives more realistic interactions with Na+ and Ca2+ cations than the original Amber Lipid17 parameters when validated using headgroup order parameters and the "electrometer concept". In ECC-lipids simulations, populations of complexes of Ca2+ cations with more than two PS lipids are negligible, and interactions of Ca2+ cations with only carboxylate groups are twice more likely than with only phosphate groups, while interactions with carbonyls almost entirely involve other groups as well. Our results pave the way for more realistic MD simulations of biomolecular systems with anionic membranes, allowing signaling processes involving PS and Ca2+ to be elucidated.

• MeSH
• draslík metabolismus   MeSH
• elektrony MeSH
• fosfatidylseriny metabolismus   MeSH
• kationty metabolismus   MeSH
• lipidové dvojvrstvy metabolismus   MeSH
• simulace molekulární dynamiky MeSH
• sodík metabolismus   MeSH
• vápník metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 1-palmitoyl-2-oleoylglycero-3-phosphoserine MeSH Prohlížeč
• draslík MeSH
• fosfatidylseriny MeSH
• kationty MeSH
• lipidové dvojvrstvy MeSH
• sodík MeSH
• vápník MeSH

Phosphatidylserine (PS) is a negatively charged lipid type commonly found in eukaryotic membranes, where it interacts with proteins via nonspecific electrostatic interactions as well as via specific binding. Moreover, in the presence of calcium ions, PS lipids can induce membrane fusion and phase separation. Molecular details of these phenomena remain poorly understood, partly because accurate models to interpret the experimental data have not been available. Here we gather a set of previously published experimental NMR data of C-H bond order parameter magnitudes, |SCH|, for pure PS and mixed PS:PC (phosphatidylcholine) lipid bilayers and augment this data set by measuring the signs of SCH in the PS headgroup using S-DROSS solid-state NMR spectroscopy. The augmented data set is then used to assess the accuracy of the PS headgroup structures in, and the cation binding to, PS-containing membranes in the most commonly used classical molecular dynamics (MD) force fields including CHARMM36, Lipid17, MacRog, Slipids, GROMOS-CKP, Berger, and variants. We show large discrepancies between different force fields and that none of them reproduces the NMR data within experimental accuracy. However, the best MD models can detect the most essential differences between PC and PS headgroup structures. The cation binding affinity is not captured correctly by any of the PS force fields-an observation that is in line with our previous results for PC lipids. Moreover, the simulated response of the PS headgroup to bound ions can differ from experiments even qualitatively. The collected experimental data set and simulation results will pave the way for development of lipid force fields that correctly describe the biologically relevant negatively charged membranes and their interactions with ions. This work is part of the NMRlipids open collaboration project ( nmrlipids.blogspot.fi ).

• MeSH
• buněčná membrána chemie   metabolismus   MeSH
• fosfatidylseriny chemie   metabolismus   MeSH
• kationty chemie   metabolismus   MeSH
• lipidové dvojvrstvy chemie   metabolismus   MeSH
• simulace molekulární dynamiky MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fosfatidylseriny MeSH
• kationty MeSH
• lipidové dvojvrstvy MeSH

Membrane surface charge is critical for the transient, yet specific recruitment of proteins with polybasic regions to certain organelles. In eukaryotes, the plasma membrane (PM) is the most electronegative compartment of the cell, which specifies its identity. As such, membrane electrostatics is a central parameter in signaling, intracellular trafficking, and polarity. Here, we explore which are the lipids that control membrane electrostatics using plants as a model. We show that phosphatidylinositol-4-phosphate (PI4P), phosphatidic acidic (PA), and phosphatidylserine (PS) are separately required to generate the electrostatic signature of the plant PM. In addition, we reveal the existence of an electrostatic territory that is organized as a gradient along the endocytic pathway and is controlled by PS/PI4P combination. Altogether, we propose that combinatorial lipid composition of the cytosolic leaflet of organelles not only defines the electrostatic territory but also distinguishes different functional compartments within this territory by specifying their varying surface charges.

• Klíčová slova
• Arabidopsis, biosensor, endocytosis, lipid signaling, membrane biology, phosphatidic acid, phosphatidylserine, phosphoinositides, plant cell biology, vesicular trafficking,
• MeSH
• Arabidopsis růst a vývoj   metabolismus   MeSH
• buněčná membrána metabolismus   MeSH
• fosfatidylinositolfosfáty metabolismus   MeSH
• fosfatidylseriny metabolismus   MeSH
• kořeny rostlin růst a vývoj   metabolismus   MeSH
• kyseliny fosfatidové metabolismus   MeSH
• organely MeSH
• proteiny huseníčku metabolismus   MeSH
• signální transdukce MeSH
• statická elektřina * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fosfatidylinositolfosfáty MeSH
• fosfatidylseriny MeSH
• kyseliny fosfatidové MeSH
• phosphatidylinositol 4-phosphate MeSH Prohlížeč
• proteiny huseníčku MeSH

Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) is a critically important regulatory lipid of the plasma membrane (PM); however, little is known about how cells regulate PM PI(4,5)P2 levels. Here, we show that the phosphatidylinositol 4-phosphate (PI4P)/phosphatidylserine (PS) transfer activity of the endoplasmic reticulum (ER)-resident ORP5 and ORP8 is regulated by both PM PI4P and PI(4,5)P2 Dynamic control of ORP5/8 recruitment to the PM occurs through interactions with the N-terminal Pleckstrin homology domains and adjacent basic residues of ORP5/8 with both PI4P and PI(4,5)P2 Although ORP5 activity requires normal levels of these inositides, ORP8 is called on only when PI(4,5)P2 levels are increased. Regulation of the ORP5/8 attachment to the PM by both phosphoinositides provides a powerful means to determine the relative flux of PI4P toward the ER for PS transport and Sac1-mediated dephosphorylation and PIP 5-kinase-mediated conversion to PI(4,5)P2 Using this rheostat, cells can maintain PI(4,5)P2 levels by adjusting the availability of PI4P in the PM.

• MeSH
• biologický transport MeSH
• buněčná membrána metabolismus   MeSH
• endoplazmatické retikulum metabolismus   MeSH
• fosfatidylinositol-4,5-difosfát metabolismus   MeSH
• fosfatidylinositolfosfáty metabolismus   MeSH
• fosfatidylseriny metabolismus   MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem metabolismus   MeSH
• HEK293 buňky MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• proteinové domény MeSH
• steroidní receptory chemie   metabolismus   MeSH
• substrátová specifita MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, N.I.H., Intramural MeSH
• Názvy látek
• fosfatidylinositol-4,5-difosfát MeSH
• fosfatidylinositolfosfáty MeSH
• fosfatidylseriny MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
• Oxysterol Binding Proteins MeSH
• phosphatidylinositol 4-phosphate MeSH Prohlížeč
• steroidní receptory MeSH

SCOPE: The aim of this work is to identify which proapoptotic pathway is induced in human colon cancer cell lines, in contact with proanthocyanidins extracted from various berries. METHODS AND RESULTS: Proanthocyanidins (Pcys) extracted from 11 berry species are monitored for proapoptotic activities on two related human colon cancer cell lines: SW480-TRAIL-sensitive and SW620-TRAIL-resistant. Apoptosis induction is monitored by cell surface phosphatidylserine (PS) detection. Lowbush blueberry extract triggers the strongest activity. When tested on the human monocytic cell line THP-1, blueberry Pcys are less effective for PS externalisation and DNA fragmentation is absent, highlighting a specificity of apoptosis induction in gut cells. In Pcys-treated gut cell lines, caspase 8 (apoptosis extrinsic pathway) but not caspase 9 (apoptosis intrinsic pathway) is activated after 3 hours through P38 phosphorylation (90 min), emphasizing the potency of lowbush blueberry Pcys to eradicate gut TRAIL-resistant cancer cells. CONCLUSION: We highlight here that berries Pcys, especially lowbush blueberry Pcys, are of putative interest for nutritional chemoprevention of colorectal cancer in view of their apoptosis induction in a human colorectal cancer cell lines.

• MeSH
• antigeny CD95 metabolismus   MeSH
• apoptóza účinky léků   MeSH
• brusnice s jedlými plody chemie   metabolismus   MeSH
• buněčné linie MeSH
• chemorezistence MeSH
• DNA metabolismus   MeSH
• fosfatidylseriny metabolismus   MeSH
• fosforylace účinky léků   MeSH
• fragmentace DNA účinky léků   MeSH
• kaspasa 3 metabolismus   MeSH
• kaspasa 8 metabolismus   MeSH
• kaspasa 9 metabolismus   MeSH
• lidé MeSH
• mitogenem aktivované proteinkinasy p38 metabolismus   MeSH
• nádory tračníku metabolismus   patologie   MeSH
• ovoce chemie   metabolismus   MeSH
• proantokyanidiny chemie   izolace a purifikace   toxicita   MeSH
• protein TRAIL toxicita   MeSH
• rostlinné extrakty chemie   MeSH
• TRAIL receptory metabolismus   MeSH
• Vaccinium vitis-idaea chemie   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antigeny CD95 MeSH
• DNA MeSH
• FAS protein, human MeSH Prohlížeč
• fosfatidylseriny MeSH
• kaspasa 3 MeSH
• kaspasa 8 MeSH
• kaspasa 9 MeSH
• mitogenem aktivované proteinkinasy p38 MeSH
• proantokyanidiny MeSH
• protein TRAIL MeSH
• rostlinné extrakty MeSH
• TRAIL receptory MeSH