Polycyclic aromatic hydrocarbon components contribute to the mitochondria-antiapoptotic effect of fine particulate matter on human bronchial epithelial cells via the aryl hydrocarbon receptor

<p>Abstract</p> <p>Background</p> <p>Nowadays, effects of fine particulate matter (PM_2.5) are well-documented and related to oxidative stress and pro-inflammatory response. Nevertheless, epidemiological studies show that PM_2.5exposure is correlated with an increase of pulmonary cancers and the remodeling of the airway epithelium involving the regulation of cell death processes. Here, we investigated the components of Parisian PM_2.5involved in either the induction or the inhibition of cell death quantified by different parameters of apoptosis and delineated the mechanism underlying this effect.</p> <p>Results</p> <p>In this study, we showed that low levels of Parisian PM_2.5are not cytotoxic for three different cell lines and primary cultures of human bronchial epithelial cells. Conversely, a 4 hour-pretreatment with PM_2.5prevent mitochondria-driven apoptosis triggered by broad spectrum inducers (A23187, staurosporine and oligomycin) by reducing the mitochondrial transmembrane potential loss, the subsequent ROS production, phosphatidylserine externalization, plasma membrane permeabilization and typical morphological outcomes (cell size decrease, massive chromatin and nuclear condensation, formation of apoptotic bodies). The use of recombinant EGF and specific inhibitor led us to rule out the involvement of the classical EGFR signaling pathway as well as the proinflammatory cytokines secretion. Experiments performed with different compounds of PM_2.5suggest that endotoxins as well as carbon black do not participate to the antiapoptotic effect of PM_2.5. Instead, the water-soluble fraction, washed particles and organic compounds such as polycyclic aromatic hydrocarbons (PAH) could mimic this antiapoptotic activity. Finally, the activation or silencing of the aryl hydrocarbon receptor (AhR) showed that it is involved into the molecular mechanism of the antiapoptotic effect of PM_2.5at the mitochondrial checkpoint of apoptosis.</p> <p>Conclusions</p> <p>The PM_2.5-antiapoptotic effect in addition to the well-documented inflammatory response might explain the maintenance of a prolonged inflammation state induced after pollution exposure and might delay repair processes of injured tissues.</p

Evidence for a mitochondrial localization of the retinoblastoma protein

<p>Abstract</p> <p>Background</p> <p>The retinoblastoma protein (Rb) plays a central role in the regulation of cell cycle, differentiation and apoptosis. In cancer cells, ablation of Rb function or its pathway is a consequence of genetic inactivation, viral oncoprotein binding or deregulated hyperphosphorylation. Some recent data suggest that Rb relocation could also account for the regulation of its tumor suppressor activity, as is the case for other tumor suppressor proteins, such as p53.</p> <p>Results</p> <p>In this reported study, we present evidence that a fraction of the total amount of Rb protein can localize to the mitochondria in proliferative cells taken from both rodent and human cells. This result is also supported by the use of Rb siRNAs, which substantially reduced the amount of mitochondrial Rb, and by acellular assays, in which [³⁵S]-Methionine-labeled Rb proteins bind strongly to mitochondria isolated from rat liver. Moreover, endogenous Rb is found in an internal compartment of the mitochondria, within the inner-membrane. This is consistent with the protection of Rb from alkaline treatment, which destroys any interaction of proteins that are weakly bound to mitochondria.</p> <p>Conclusion</p> <p>Although a few data regarding an unspecific cytosolic localization of Rb protein have been reported for some tumor cells, our results are the first evidence of a mitochondrial localization of Rb. The mitochondrial localization of Rb is observed in parallel with its classic nuclear location and paves the way for the study of potential as-yet-unknown roles of Rb at this site.</p

Carbon black and titanium dioxide nanoparticles elicit distinct apoptotic pathways in bronchial epithelial cells

<p>Abstract</p> <p>Background</p> <p>Increasing environmental and occupational exposures to nanoparticles (NPs) warrant deeper insight into the toxicological mechanisms induced by these materials. The present study was designed to characterize the cell death induced by carbon black (CB) and titanium dioxide (TiO₂) NPs in bronchial epithelial cells (16HBE14o- cell line and primary cells) and to investigate the implicated molecular pathways.</p> <p>Results</p> <p>Detailed time course studies revealed that both CB (13 nm) and TiO₂(15 nm) NP exposed cells exhibit typical morphological (decreased cell size, membrane blebbing, peripheral chromatin condensation, apoptotic body formation) and biochemical (caspase activation and DNA fragmentation) features of apoptotic cell death. A decrease in mitochondrial membrane potential, activation of Bax and release of cytochrome <it>c </it>from mitochondria were only observed in case of CB NPs whereas lipid peroxidation, lysosomal membrane destabilization and cathepsin B release were observed during the apoptotic process induced by TiO₂NPs. Furthermore, ROS production was observed after exposure to CB and TiO₂but hydrogen peroxide (H₂O₂) production was only involved in apoptosis induction by CB NPs.</p> <p>Conclusions</p> <p>Both CB and TiO₂NPs induce apoptotic cell death in bronchial epithelial cells. CB NPs induce apoptosis by a ROS dependent mitochondrial pathway whereas TiO₂NPs induce cell death through lysosomal membrane destabilization and lipid peroxidation. Although the final outcome is similar (apoptosis), the molecular pathways activated by NPs differ depending upon the chemical nature of the NPs.</p

On Placental Toxicology Studies and Cerium Dioxide Nanoparticles

International audienceThe human placenta is a transient organ essential for pregnancy maintenance, fetal development and growth. It has several functions, including that of a selective barrier against pathogens and xenobiotics from maternal blood. However, some pollutants can accumulate in the placenta or pass through with possible repercussions on pregnancy outcomes. Cerium dioxide nanoparticles (CeO2 NPs), also termed nanoceria, are an emerging pollutant whose impact on pregnancy is starting to be defined. CeO2 NPs are already used in different fields for industrial and commercial applications and have even been proposed for some biomedical applications. Since 2010, nanoceria have been subject to priority monitoring by the Organization for Economic Co-operation and Development in order to assess their toxicity. This review aims to summarize the current methods and models used for toxicology studies on the placental barrier, from the basic ones to the very latest, as well as to overview the most recent knowledge of the impact of CeO2 NPs on human health, and more specifically during the sensitive window of pregnancy. Further research is needed to highlight the relationship between environmental exposure to CeO2 and placental dysfunction with its implications for pregnancy outcome

Dysfunction in the mitochondrial Fe-S assembly machinery leads to formation of the chemoresistant truncated VDAC1 isoform without HIF-1α activation

International audienceBiogenesis of iron-sulfur clusters (ISC) is essential to almost all forms of life and involves complex protein machineries. This process is initiated within the mitochondrial matrix by the ISC assembly machinery. Cohort and case report studies have linked mutations in ISC assembly machinery to severe mitochondrial diseases. The voltage-dependent anion channel (VDAC) located within the mitochondrial outer membrane regulates both cell metabolism and apoptosis. Recently, the C-terminal truncation of the VDAC1 isoform, termed VDAC1-ΔC, has been observed in chemoresistant late-stage tumor cells grown under hypoxic conditions with activation of the hypoxia-response nuclear factor HIF-1α. These cells harbored atypical enlarged mitochondria. Here, we show for the first time that depletion of several proteins of the mitochondrial ISC machinery in normoxia leads to a similar enlarged mitochon-dria phenotype associated with accumulation of VDAC1-ΔC. This truncated form of VDAC1 accumulates in the absence of HIF-1α and HIF-2α activations and confers cell resistance to drug-induced apoptosis. Furthermore, we show that when hypoxia and siRNA knock-down of the ISC machinery core components are coupled, the cell phenotype is further accentuated , with greater accumulation of VDAC1-ΔC. Interestingly, we show that hypoxia promotes the downregulation of several proteins (ISCU, NFS1, FXN) involved in the early steps of mitochondrial Fe-S cluster biogenesis. Finally, we have identified the mitochondria-associated membrane (MAM) localized Fe-S protein CISD2 as a link between ISC machinery downregulation and accumulation of anti-apoptotic VDAC1-ΔC. Our results are the first to associate dysfunction in Fe-S cluster biogenesis with cleavage of VDAC1, a form which has previously been shown to promote tumor resistance to chemotherapy, and raise new perspectives for targets in cancer therapy

Regulation of iron metabolism by sideroflexin SFXN1

International audienceThe mitochondrion is at the crossroad of critical metabolic pathways, thus being a crucial player in cell fate in response to stress or infection. The mitochondrion is also an essential organelle for iron metabolism, being the place of heme and iron-sulfur clusters (Fe-S), two co-factors required for mitochondrial respiration. Iron intracellular levels are thus tightly controlled to prevent the accumulation of free ferrous iron that, when in excess, generates oxidative stress and may induce ferroptosis, an iron-dependent regulated cell death (RCD). Ferrous iron will react with ROS and generate, by Fenton reaction, lipids peroxidation. Oxidized lipids accumulation into cellular membranes can be lethal and drives ferroptosis. Ferroptosis, a physiological cell death contributing to tissue homeostasis aging and diseases as neurodegenerative, organ injury. Furthermore a number of tumor suppressors exert part of their tumor-suppression function through the induction of ferroptosis. Sideroflexins (SFXNs) are mitochondrial transporters localized in the inner mitochondrial membrane whose functions are progressively being specified. SFXNs are conserved in eukaryote forming a family of five mitochondrial proteins in mammals. The main function reported to date for Sideroflexin 1 (SFXN1) is to transport serine but SFXN1 may also be involved in iron metabolism. The serine transporter function appears to be conserved between mammalian and yeast. Our project is to study if SFXN1 can regulate cell viability, mitochondrial functions and structure, as well as its role in iron homeostasis and if SFXN1 can modulate cell sensitivity to ferroptosis inducers.References :Kory N, Wyant GA, Prakash G, Uit de Bos J, Bottanelli F, Pacold ME, Chan SH, Lewis CA, Wang T, Keys HR, Guo YE, Sabatini DM. SFXN1 is a mitochondrial serine transporter required for one-carbon metabolism. Science. 2018 Nov 16;362(6416):eaat9528. doi: 10.1126/science.aat9528. PMID: 30442778; PMCID: PMC6300058.Acoba MG, Alpergin ESS, Renuse S, Fernández-Del-Río L, Lu YW, Khalimonchuk O, Clarke CF, Pandey A, Wolfgang MJ, Claypool SM. The mitochondrial carrier SFXN1 is critical for complex III integrity and cellular metabolism. Cell Rep. 2021 Mar 16;34(11):108869. doi: 10.1016/j.celrep.2021.108869. PMID: 33730581; PMCID: PMC8048093.Tifoun N, De Las Heras JM, Guillaume A, Bouleau S, Mignotte B, Le Floch N. Insights into the Roles of the Sideroflexins/SLC56 Family in Iron Homeostasis and Iron-Sulfur Biogenesis. Biomedicines. 2021 Jan 21;9(2):103. doi: 10.3390/biomedicines9020103. PMID: 33494450; PMCID: PMC7911444.Stockwell BR. Ferroptosis turns 10: Emerging mechanisms, physiological functions, and therapeutic applications. Cell. 2022 Jul 7;185(14):2401-2421. doi: 10.1016/j.cell.2022.06.003. PMID: 35803244; PMCID: PMC9273022

The p76 Rb and p100 Rb truncated forms of the Rb protein exert antagonistic roles on cell death regulation in human cell lines

International audienceSeveral caspase-cleaved forms of the retinoblastoma protein have been described. Here, we compared the effect of full-length Rb versus the truncated p76 Rb and p100 Rb proteins on cell death regulation in five human cell lines. Interestingly, we observed that p76 Rb triggers cell death in all tested cell lines and that p100 Rb protects two cell lines against etoposide or TNF-a-induced cell death, whereas full-length Rb has no apoptotic effect. These results show that truncated forms of Rb can have specific activities in the regulation of cell death. They also suggest that caspase cleavage of Rb should not be simply assimilated to a degradation process. Finally, we show that cell death induced by p76 Rb is Bax-dependent and is diminished by Bcl-2 overexpression or by caspase inhibition and that p100 Rb could inhibit cell death by decreasing both p53 stability and caspase activity

Impact of hypoxia on proteins of the mitochondrial ISC assembly machinery.

<p>(A) Total protein extracts from HeLa cells grown in normoxia (Nx, 21% O₂) or hypoxia (Hx, 1% O₂) conditions for the indicated times were analyzed by immunoblotting using VDACs poly antibody and anti-CAIX, -ISCU, -FXN, -NFS1, -HSC20 antibodies. β-Actin was used as loading control. (<i>right panel)</i> Bar graph represents the amount of the indicated proteins relative to ß-actin level determined by quantification of n = 3 immunoblot analysis using the Odyssey System Imager. Mean and standard deviation of 3 independent experiments are shown (* p < 0.05, n = 3). (B) HeLa cells were either untreated or treated with CoCl₂ for 2 days. Total proteins were analyzed by western blotting using anti-HIF-1α, -ISCU, -FXN antibodies. Vinculin was used as loading control. (C) HeLa cells grown in normoxia (Nx, 21% O₂) or hypoxia (Hx, 1% O₂) conditions for 3 days and mRNA levels of <i>iscu</i>, <i>fxn</i> and <i>vegf A</i> were determined by RT-qPCR and normalized to 18S ribosomal rRNA levels. The bar graph presents the ratio between the relative level of each mRNA under hypoxic and normoxic conditions with a logarithmic scale. Mean and standard deviation of 6 independent experiments are shown (* p < 0.05, n = 6). (D) Total protein extracts from HeLa cells grown under normoxic (Nx, 21% O₂) or hypoxic (Hx, 1% O₂) conditions for the indicated times were analyzed by immunoblotting using anti-CIAPIN1, -NUBP1, -NARFL antibodies. β-Actin was used as loading control.</p