6533b822fe1ef96bd127d954
RESEARCH PRODUCT
Oxysterol mixture in hypercholesterolemia-relevant proportion causes oxidative stress-dependent eryptosis.
Carla GentileAlessandro AttanzioAntonio CillaM. A. LivreaLuisa TesoriereMario Allegrasubject
ErythrocytesPhysiologyEryptosisApoptosisPharmacologylcsh:PhysiologyAntioxidantschemistry.chemical_compoundPhospholipid scramblingSettore BIO/10 - Biochimicapolycyclic compoundslcsh:QD415-436PhosphatidylserineKetocholesterolsProtein Kinase Clcsh:QP1-981OxysterolsPhosphatidylserineErythrocyteCalphostin CBiochemistryCaspaseslipids (amino acids peptides and proteins)AntioxidantReactive Oxygen SpecieHumanProgrammed cell deathOxysterolHypercholesterolemiachemistry.chemical_elementPhosphatidylserinesCalciumCalcium ChannelDinoprostonelcsh:BiochemistryOxysterolLipid oxidationHumansCalphostinHypercholesterolemia Human red blood cell Oxysterols Eryptosis Oxidative stressKetocholesterolApoptosiOxidative StreCaspaseOxidative StresschemistryCalciumCalcium ChannelsReactive Oxygen SpeciesEryptosiHuman red blood celldescription
Background/Aims: Oxysterol activity on the erythrocyte (RBC) programmed cell death (eryptosis) had not been studied yet. Effects of an oxysterol mixture in hyper-cholesterolemic-relevant proportion, and of individual compounds, were investigated on RBCs from healthy humans. Methods: Membrane phosphatidylserine (PS) externalization, calcium entry, ROS production, amino-phospholipid translocase (APLT) activity were evaluated by cytofluorimetric assays, cell volume from forward scatter. Prostaglandin PGE2 was measured by ELISA; GSH-adducts and lipoperoxides by spectrophotometry. Involvement of protein kinase C and caspase was investigated by inhibitors staurosporin, calphostin C, and Z-DEVD-FMK, respectively. Results: Oxysterols caused PS externalization and cell shrinkage, associated with PGE2 release, opening of PGE2-dependent calcium channels, ROS production, GSH depletion, membrane lipid oxidation. Addition of antioxidants prevented Ca2+ influx and eryptosis. Calcium removal prevented cell shrinkage, with small effect (-20%) on the PS exposure, whereas ROS generation was unaltered. Either in the presence or absence of calcium i) oxysterols inhibited APLT, ii) staurosporin, calphostin C, Z-DEVD-FMK blunted and iii) antioxidants fully prevented the oxysterol-induced PS externalization. Only 7-ketocholesterol and cholestan-3β,5α,6β-triol were individually active. Eryptosis was observed in RBCs isolated after ex vivo spiking of human whole blood with the oxysterol mixture. Conclusions: Oxysterols induce an oxidative stress-dependent eryptosis, involving calcium-independent mechanisms. Eryptotic activity of oxysterols may be relevant in vivo.
year | journal | country | edition | language |
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2014-01-01 | Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology |