Home Publication date: 20 February 2020 Source: Free Radical Biology and Medicine, Volume 148 Author(s): Marie-Paule Hamon, Rachel Gergondey, Aurore L'honoré, Bertrand Friguet AbstractThe ATP-dependent Lon protease is located in the mitochondrial matrix and oxidized proteins are among its primary targets for their degradation. Impairment of mitochondrial morphology and function together with apoptosis were observed in lung fibroblasts depleted for Lon expression while accumulation of carbonylated mitochondrial proteins has been reported for yeast and HeLa Lon deficient cells. In addition, age-related mitochondrial dysfunction has been associated with an impairment of Lon expression. Using a HeLa cell line stably transfected with an inducible shRNA directed against Lon, we have previously observed that Lon depletion results in a mild phenotype characterized by an increase of both production of reactive oxygen species and level of oxidized proteins (Bayot et al., 2014, Biochimie, 100: 38-47). In this study using the same cell line, we now show that Lon knockdown leads to modifications of the expression of a number of specific proteins involved in protein quality control, stress response and energy metabolism, as evidenced using a 2D gel-based proteomic approach, and to alteration of the mitochondrial network morphology. We also show that these effects are associated with decreased proliferation and can be modulated by culture conditions in galactose versus glucose containing medium. Graphical abstract
Publication date: 20 February 2020 Source: Free Radical Biology and Medicine, Volume 148 Author(s): Jelena Korać Jačić, Ljiljana Nikolić, Dalibor M. Stanković, Miloš Opačić, Milena Dimitrijević, Danijela Savić, Sanja Grgurić Šipka, Ivan Spasojević, Jelena Bogdanović Pristov AbstractUpon release in response to stress, epinephrine (Epi) may interact with labile iron pool in human plasma with potentially important (patho)physiological consequences. We have shown that Epi and Fe3+ build stable 1:1 high-spin bidentate complex at physiological pH, and that Epi does not undergo degradation in the presence of iron. However, the interactions of Epi with the more soluble Fe2+, and the impact of iron on biological activity of Epi are still not known. Herein we showed that Epi and Fe2+ build colorless complex which is stable under anaerobic conditions. In the presence of O2, Epi promoted the oxidation of Fe2+ and the formation of Epi-Fe3+ complex. Cyclic voltammetry showed that mid-point potential of Epi-Fe2+ complex is very low (−582 mV vs. standard hydrogen electrode), which explains catalyzed oxidation of Fe2+. Next, we examined the impact of iron binding on biological performance of Epi using patch clamping in cell culture with constitutive expression of adrenergic receptors. Epi alone evoked an increase of outward currents, whereas Epi in the complex with Fe3+ did not. This implies that the binding of Epi to adrenergic receptors and their activation is prevented by the formation of complex with iron. Pro-oxidative activity of Epi-Fe2+ complex may represent a link between chronic stress and cardiovascular problems. On the other hand, labile iron could serve as a modulator of biological activity of ligands. Such interactions may be important in human pathologies that are related to iron overload or deficiency. Graphical abstract
Publication date: 20 February 2020 Source: Free Radical Biology and Medicine, Volume 148 Author(s): Miłosz Wieczór, Jacek Czub AbstractOxidative damage to DNA is widely known to contribute to aging and disease. This relationship has been extensively studied for telomeres – structures that cap chromosome ends – due to their role in cell proliferation and senescence, and exceptional susceptibility to oxidation. Indeed, the repetitive telomeric DNA sequence contains the 5′-GGG-3′ motif that has the lowest ionization potential of all trinucleotides. Accordingly, experiments consistently show that telomeric oxidative lesions are more abundant and persistent than elsewhere in the genome. This led to a hypothesis that telomeres act as sensors of prolonged oxidative stress and prevent carcinogenesis, as disruption of telomeric integrity triggers senescence or apoptosis. Here, we use atomistic alchemical Molecular Dynamics simulations to perform a combinatorial assessment of changes in DNA binding affinity of telomeric proteins induced by oxidative guanine lesions. We rank lesions by their effect on telomere integrity, as well as telomeric proteins by their sensitivity to DNA oxidation. While the binding of most proteins is abolished by DNA oxidation, HOT1 emerges as a notable exception, suggesting its potential role in sensing of oxidative damage. Through statistical analysis and free energy decomposition, we also identify common trends in structural responses of protein-DNA complexes that contribute to decreased binding affinity. Graphical abstract
Publication date: 20 February 2020 Source: Free Radical Biology and Medicine, Volume 148 Author(s): Guanghui Deng, Yunjia Li, Shuoyi Ma, Zhuowei Gao, Ting Zeng, Limei Chen, Haixin Ye, Menghan Yang, Hao Shi, Xiaofen Yao, Zhiyun Zeng, Yuyao Chen, Yuhong Song, Bing Liu, Lei Gao AbstractFerroptosis is a new regulated cells death manner defined as results of iron-dependent accumulation of lipid peroxidation. However, the specific mechanisms of regulating ferroptosis remain unclear. In our present study, we demonstrated that Caveolin-1 (Cav-1) played a central role in protecting hepatocytes against ferroptosis in autoimmunity-mediated hepatitis (AIH). The down-regulated Cav-1 in liver tissues, accompanied by ferroptotic events and RNS production, were contributed to the outcome of ConA-induced hepatic damage, which were rescued by ferrostatin-1 (an inhibitor of ferroptosis) in vivo and in vitro. Additionally, Cav-1 deficiency aggravated ConA-induced hepatocellular death and ferroptosis associated with excessive nitrogen stress response. Short hairpin RNA of Cav-1 in hepatocytes promoted ferroptosis and nitrative stress in response to erastin in vitro, which was ameliorated by Cav-1 over-expression. Meanwhile, administration of the iNOS inhibitor (1400W) or ONOO- scavenger (Fe-TMPyP), diminished reactive nitrogen species (RNS), remarkably reduced hepatocytes ferroptosis and attenuated ConA-induced liver damage. Furthermore, immune inhibition by gadolinium chloride (GdCl3), a well-known Kupffer cell depletor, elevated hepatic Cav-1 but inhibited ferroptosis and nitrative stress under ConA exposure. In conclusion, these data revealed a novel molecular mechanism of ferroptosis with the Cav-1 regulation was essential for pathogenesis of ConA-induced hepatitis. Downstream of Cav-1, RNS-mediated ferroptosis was a pivotal step that drives the execution of acute immune-mediated hepatic damage. Graphical abstract
Publication date: 20 February 2020 Source: Free Radical Biology and Medicine, Volume 148 Author(s): Pan Tang, Wen-Xiang Chen, Hong-Liang Gao, Jia-Yong Dai, Yu Gu, Zi-Ang Xie, Xiong-Feng Li, Shun-Wu Fan, Xue-Sheng Jiang, Qian Lu, Zhi-Jun Hu Graphical abstract
Publication date: February 2020 Source: Redox Biology, Volume 30 Author(s): Matthew Lovatt, Khadijah Adnan, Viridiana Kocaba, Martin Dirisamer, Gary S.L. Peh, Jodhbir S. Mehta AbstractCorneal transparency is maintained by a monolayer of corneal endothelial cells. Defects in corneal endothelial cells (CEnCs) can be rectified surgically through transplantation. Fuchs’ endothelial corneal dystrophy (FECD) is the foremost cause of endothelial dysfunction and the leading indication for transplantation. Increased sensitivity of CEnCs to oxidative stress is thought to contribute to the pathogenesis of FECD through increased apoptosis. In part, this is thought to be due to loss of NRF2 expression: a global regulator of oxidative stress. We demonstrate that expression of the redox sensor, peroxiredoxin 1 (PRDX1) is selectively lost from CEnCs in FECD patient samples. We reveal that expression of PRDX1 is necessary to control the response of CEnCs to agents that cause lipid peroxidation. Iron-dependent lipid peroxidation drives non-apoptotic cell death termed ferroptosis. We establish that the inhibitor of ferroptosis, ferrostatin-1 rescues lipid peroxidation and cell death in CEnCs. Furthermore, we provide evidence that the transcription factor NRF2 similarly regulates lipid peroxidation in CEnCs. Graphical abstract
Publication date: February 2020 Source: Redox Biology, Volume 30 Author(s): Irina A. Okkelman, Nuno Neto, Dmitri B. Papkovsky, Michael G. Monaghan, Ruslan I. Dmitriev AbstractStem cells and the niche in which they reside feature a complex microenvironment with tightly regulated homeostasis, cell-cell interactions and dynamic regulation of metabolism. A significant number of organoid models has been described over the last decade, yet few methodologies can enable single cell level resolution analysis of the stem cell niche metabolic demands, in real-time and without perturbing integrity. Here, we studied the redox metabolism of Lgr5-GFP intestinal organoids by two emerging microscopy approaches based on luminescence lifetime measurement – fluorescence-based FLIM for NAD(P)H, and phosphorescence-based PLIM for real-time oxygenation. We found that exposure of stem (Lgr5-GFP) and differentiated (no GFP) cells to high and low glucose concentrations resulted in measurable shifts in oxygenation and redox status. NAD(P)H-FLIM and O2-PLIM both indicated that at high ‘basal’ glucose conditions, Lgr5-GFP cells had lower activity of oxidative phosphorylation when compared with cells lacking Lgr5. However, when exposed to low (0.5 mM) glucose, stem cells utilized oxidative metabolism more dynamically than non-stem cells. The high heterogeneity of complex 3D architecture and energy production pathways of Lgr5-GFP organoids were also confirmed by the extracellular flux (XF) analysis. Our data reveals that combined analysis of NAD(P)H-FLIM and organoid oxygenation by PLIM represents promising approach for studying stem cell niche metabolism in a live readout.
Publication date: February 2020 Source: Redox Biology, Volume 30 Author(s): Xinyu Yang, Na An, Changming Zhong, Manke Guan, Yuchen Jiang, Xinye Li, Hanlai Zhang, Liqin Wang, Yanfei Ruan, Yonghong Gao, Nian Liu, Hongcai Shang, Yanwei Xing AbstractAtrial fibrillation (AF) occurs in up to 11% of cancer patients treated with ibrutinib. The pathophysiology of ibrutinib promoted AF is complicated, as there are multiple interactions involved; the detailed molecular mechanisms underlying this are still unclear. Here, we aimed to determine the electrophysiological and molecular mechanisms of burst-pacing-induced AF in ibrutinib-treated mice. The results indicated differentially expressed proteins in ibrutinib-treated mice, identified through proteomic analysis, were found to play a role in oxidative stress-related pathways. Finally, treatment with an inhibitor of NADPH oxidase (NOX) prevented and reversed AF development in ibrutinib-treated mice. It was showed that the related protein expression of reactive oxygen species (ROS) in the ibrutinib group was significantly increased, including NOX2, NOX4, p22-phox, XO and TGF-β protein expression. It was interesting that ibrutinib group also significantly increased the expression of ox-CaMKII, p-CaMKII (Thr-286) and p-RyR2 (Ser2814), causing enhanced abnormal sarcoplasmic reticulum (SR) Ca2+ release and mitochondrial structures, as well as atrial fibrosis and atrial hypertrophy in ibrutinib-treated mice, and apocynin reduced the expression of these proteins. Ibrutinib-treated mice were also more likely to develop AF, and AF occurred over longer periods. In conclusion, our study has established a pathophysiological role for ROS signaling in atrial cardiomyocytes, and it may be that ox-CaMKII and p-CaMKII (Thr-286) are activated by ROS to increase AF susceptibility following ibrutinib treatment. We have also identified the inhibition of NOX as a potential novel AF therapy approach.
Publication date: February 2020 Source: Redox Biology, Volume 30 Author(s): Zishan Wang, Hongtian Dong, Jinghui Wang, Yulu Huang, Xiaoshuang Zhang, Yilin Tang, Qing Li, Zhaolin Liu, Yuanyuan Ma, Jiabin Tong, Li Huang, Jian Fei, Mei Yu, Jian Wang, Fang Huang AbstractThe pathological hallmarks of Parkinson's disease (PD) are the progressive loss of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc) and the presence of overactivated glial cells and neuroinflammation. Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) c-Rel subunit is closely related in the pathological progress of PD, however the roles and mechanisms of c-Rel in PD development remain unclear. Here, in neurotoxins-induced PD models, the dynamic changes of NF-κB c-Rel and its functions were evaluated. We found that c-Rel was rapidly activated in the nigrostriatal pathway, which mainly occurred in dopaminergic neurons and microglia. c-Rel could maintain neuronal survival by initiating the anti-apoptotic gene expression in MPP+-treated SH-SY5Y cells and it could inhibit microglial overactivation by suppressing the inflammatory gene expression in LPS-challenged BV2 cells. c-Rel inhibitor IT901 aggravated the damage of MPTP on dopaminergic neurons and promoted the activation of microglia in the nigrostriatal pathway of mice. Moreover, the expression of c-Rel in blood samples of PD patients decreased dramatically. Our results indicate that the NF-κB/c-Rel subunit plays an important role in neuroprotection and neuroinflammation inhibition during PD progression. Graphical abstract
Publication date: February 2020 Source: Redox Biology, Volume 30 Author(s): Joonhyuk Choi, Bong Jong Seo, Hyeonwoo La, Sang Hoon Yoon, Yean Ju Hong, Ji-Heon Lee, Hyung-Min Chung, Kwonho Hong, Jeong Tae Do AbstractPre-implantation mouse blastocyst-derived stem cells, namely embryonic stem cells (ESCs), trophoblast stem cells (TSCs), and extraembryonic endoderm (XEN) cells, have their own characteristics and lineage specificity. So far, several studies have attempted to identify these three stem cell types based on genetic markers, morphologies, and factors involved in maintaining cell self-renewal. In this study, we focused on characterizing the three stem cell types derived from mouse blastocysts by observing cellular organelles, especially the mitochondria, and analyzing how mitochondrial dynamics relates to the energy metabolism in each cell type. Our study revealed that XEN cells have distinct mitochondrial morphology and energy metabolism compared with that in ESCs and TSCs. In addition, by analyzing the energy metabolism (oxygen consumption and extracellular acidification rates), we demonstrated that differences in the mitochondria affect the cellular metabolism in the stem cells. RNA sequencing analysis showed that although ESCs are developmentally closer to XEN cells in origin, their gene expression pattern is relatively closer to that of TSCs. Notably, mitochondria-, mitochondrial metabolism-, transport/secretory action-associated genes were differentially expressed in XEN cells compared with that in ESCs and TSCs, and this feature corresponds with the morphology of the cells. Graphical abstract
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