Formation of protein adducts with Hydroperoxy-PE electrophilic cleavage products during ferroptosis.

Ferroptosis is an iron dependent form of cell death, that is triggered by the discoordination of iron, lipids, and thiols. Its unique signature that distinguishes it from other forms of cell death is the formation and accumulation of lipid hydroperoxides, particularly oxidized forms of polyunsaturated phosphatidylethanolamines (PEs), which drives cell death. These readily undergo iron-catalyzed secondary free radical reactions leading to truncated products which retain the signature PE headgroup and which can readily react with nucleophilic moieties in proteins via their truncated electrophilic acyl chains. Using a redox lipidomics approach, we have identified oxidatively-truncated PE species (trPEox) in enzymatic and non-enzymatic model systems. Further, using a model peptide we demonstrate adduct formation with Cys as the preferred nucleophilic residue and PE(26:2) +2 oxygens, as one of the most reactive truncated PE-electrophiles produced. In cells stimulated to undergo ferroptosis we identified PE-truncated species with sn-2 truncations ranging from 5 to 9 carbons. Taking advantage of the free PE headgroup, we have developed a new technology using the lantibiotic duramycin, to enrich and identify the PE-lipoxidated proteins. Our results indicate that several dozens of proteins for each cell type, are PE-lipoxidated in HT-22, MLE, and H9c2 cells and M2 macrophages after they were induced to undergo ferroptosis. Pretreatment of cells with the strong nucleophile, 2-mercaptoethanol, prevented the formation of PE-lipoxidated proteins and blocked ferroptotic death. Finally, our docking simulations showed that the truncated PE species bound at least as good to several of the lantibiotic-identified proteins, as compared to the non-truncated parent molecule, stearoyl-arachidonoyl PE (SAPE), indicating that these oxidatively-truncated species favor/promote the formation of PEox-protein adducts. The identification of PEox-protein adducts during ferroptosis suggests that they are participants in the ferroptotic process preventable by 2-mercaptoethanol and may contribute to a point of no return in the ferroptotic death process.

[Changes of chromatin structure state in thymocites at the early stage of apoptosis induced by hydrogen peroxide and radiation]. / Zminy strukturnoho stanu khromatynu v tymotsytakh na rann'omu etapi apoptozu za induktsiï peroksydom vodniu i radiatsiieiu.

The paper deals with changes in the structural state of chromatin in isolated thymocites at the early stage of apoptosis induced by hydrogen peroxide and radiation. Content of necrosis and apoptosis cells in the suspension of the isolated rat thymocites, during 3-hour incubation after X-ray irradiation in a dose of 4.5 Gy or with the presence of 0.1 microM of H2O2 by the method of double lifetime staining by fluorescent dye Hehst 33342 and propydium iodide has been estimated. Apoptogenic effect of the studied effects has been found out, the dynamics of condensation and internucleosomic chromatin fragmentation has been established. It has been shown that 100 microM alpha-tocopherol inhibited completely DNA fragmentation in the cells incubated with H2O2 and only partially in irradiated cells. Introduction of postmitochondrial supernatant, isolated from the incubated control or irradiated cells, into the cell-free system which included the ATP-regenerating system and nuclei of control thymocites did not affect the level of DNA fragmentation, while the increase of the level of fragmented DNA in nuclei was observed in the presence of the supernatant obtained by centrifugation of the cells treated by H2O2. Differences of mechanisms of thymocite apoptosis initiation, as affected by hydrogen peroxide and ionizing radiation, is discussed.

[Bile secretion and the nuclear transcriptional activity of rat liver cells under the action of bile acids]. / Sekretsiia zhovchi ta transkryptsiina aktyvnist' iader klityn pechinky shchuriv pry diï zhovchnykh kyslot.

The intensity of rat's bile secretion under the intraportal infusion of cholic and taurocholic acids has been studied. The results demonstrate that bile acids decrease bile secretion rate under the infusion of these acids in the physiological saline, and increase bile secretion rate under the infusion in saline, with hydrocarbonic ions. Previous treatment by the transcriptional inhibitor actynomycin D affected the development of the hypercholeretic action of the taurocholic acid. The transcriptional activity of the isolated nuclei of liver cells is increased under taurocholic acid action, and decreased under the cholic acid action. The results show that choleretic effects of the bile acids is realised, at least, partially, on the level of the regulatory molecular mechanisms of the cell, the transcription among them.