Ferulic Acid, Pterostilbene, and Tyrosol Protect the Heart from ER-Stress-Induced Injury by Activating SIRT1-Dependent Deacetylation of eIF2α.

Disturbances in Endoplasmic Reticulum (ER) homeostasis induce ER stress, which has been involved in the development and progression of various heart diseases, including arrhythmias, cardiac hypertrophy, ischemic heart diseases, dilated cardiomyopathy, and heart failure. A mild-to-moderate ER stress is considered beneficial and adaptative for heart functioning by engaging the pro-survival unfolded protein response (UPR) to restore normal ER function. By contrast, a severe or prolonged ER stress is detrimental by promoting cardiomyocyte apoptosis through hyperactivation of the UPR pathways. Previously, we have demonstrated that the NAD+-dependent deacetylase SIRT1 is cardioprotective in response to severe ER stress by regulating the PERK pathway of the UPR, suggesting that activation of SIRT1 could protect against ER-stress-induced cardiac damage. The purpose of this study was to identify natural molecules able to alleviate ER stress and inhibit cardiomyocyte cell death through SIRT1 activation. Several phenolic compounds, abundant in vegetables, fruits, cereals, wine, and tea, were reported to stimulate the deacetylase activity of SIRT1. Here, we evaluated the cardioprotective effect of ten of these phenolic compounds against severe ER stress using cardiomyoblast cells and mice. Among the molecules tested, we showed that ferulic acid, pterostilbene, and tyrosol significantly protect cardiomyocytes and mice heart from cardiac alterations induced by severe ER stress. By studying the mechanisms involved, we showed that the activation of the PERK/eIF2α/ATF4/CHOP pathway of the UPR was reduced by ferulic acid, pterostilbene, and tyrosol under ER stress conditions, leading to a reduction in cardiomyocyte apoptosis. The protection afforded by these phenolic compounds was not directly related to their antioxidant activity but rather to their ability to increase SIRT1-mediated deacetylation of eIF2α. Taken together, our results suggest that ferulic acid, pterostilbene, and tyrosol are promising molecules to activate SIRT1 to protect the heart from the adverse effects of ER stress.

Draft Genome Sequence of the Environmental Fungus Scedosporium dehoogii.

Today, the genus Scedosporium comprises at least ten species with four of them, Scedosporium apiospermum, Scedosporium boydii, Scedosporium aurantiacum and Scedosporium minutisporum capable of colonizing the lungs of patients with cystic fibrosis. Scedosporium dehoogii, which is also common in the soil, has never been reported as causing human pulmonary infections. Here we report the first genome sequence for S. dehoogii, an invaluable resource to understand the genetic bases of pathogenesis in the genus Scedosporium.

Implication of the deacetylase sirtuin-1 on synovial angiogenesis and persistence of experimental arthritis.

OBJECTIVES: To decipher the phenotype of endothelial cells (ECs) derived from circulating progenitors issued from patients with rheumatoid arthritis (RA). METHODS: RA and control ECs were compared according to their proliferative capacities, apoptotic profile, response to tumour necrosis factor (TNF)-α stimulation and angiogenic properties. Microarray experiments were performed to identify gene candidates relevant to pathological angiogenesis. Identified candidates were detected by RT-PCR and western blot analysis in ECs and by immunohistochemistry in the synovium. Their functional relevance was then evaluated in vitro after gene invalidation by small interfering RNA and adenoviral gene overexpression, and in vivo in the mouse model of methyl-bovine serum albumin-(mBSA)-induced arthritis. RESULTS: RA ECs displayed higher proliferation rate, greater sensitisation to TNF-α and enhanced in vitro and in vivo angiogenic capacities. Microarray analyses identified the NAD-dependent protein deacetylase sirtuin-1 (SIRT1) as a relevant gene candidate. Decreased SIRT1 expression was detected in RA ECs and synovial vessels. Deficient endothelial SIRT1 expression promoted a proliferative, proapoptotic and activated state of ECs through the acetylation of p53 and p65, and lead the development of proangiogenic capacities through the upregulation of the matricellular protein cysteine-rich angiogenic protein-61. Conditional deletion of SIRT1 in ECs delayed the resolution of experimental methyl-bovine serum albumin-(mBSA)-induced arthritis. Conversely, SIRT1 activation reversed the pathological phenotype of RA ECs and alleviates signs of experimental mBSA-induced arthritis. CONCLUSIONS: These results support a role of SIRT1 in RA and may have therapeutic implications, since targeting angiogenesis, and especially SIRT1, might be used as a complementary therapeutic approach in RA.

Threat to Asian wild apple trees posed by gene flow from domesticated apple trees and their "pestified" pathogens.

Secondary contact between crops and their wild relatives poses a threat to wild species, not only through gene flow between plants, but also through the dispersal of crop pathogens and genetic exchanges involving these pathogens, particularly those that have become more virulent by indirect selection on resistant crops, a phenomenon known as "pestification." Joint analyses of wild and domesticated hosts and their pathogens are essential to address this issue, but such analyses remain rare. We used population genetics approaches, demographic inference and pathogenicity tests on host-pathogen pairs of wild or domesticated apple trees from Central Asia and their main fungal pathogen, Venturia inaequalis, which itself has differentiated agricultural and wild-type populations. We confirmed the occurrence of gene flow from cultivated (Malus domestica) to wild (Malus sieversii) apple trees in Asian forests, potentially threatening the persistence of Asian wild apple trees. Pathogenicity tests demonstrated the pestification of V. inaequalis, the agricultural-type population being more virulent on both wild and domesticated trees. Single nucleotide polymorphism (SNP) markers and the demographic modelling of pathogen populations revealed hybridization following secondary contact between agricultural and wild-type fungal populations, and dispersal of the agricultural-type pathogen population in wild forests, increasing the threat of disease in the wild apple species. We detected an SNP potentially involved in pathogen pestification, generating an early stop codon in a gene encoding a small secreted protein in the agricultural-type fungal population. Our findings, based on joint analyses of paired host and pathogen data sets, highlight the threat posed by cultivating a crop near its centre of origin, in terms of pestified pathogen invasions in wild plant populations and introgression in the wild-type pathogen population.

Tebuconazole induces ROS-dependent cardiac cell toxicity by activating DNA damage and mitochondrial apoptotic pathway.

Tebuconazole (TEB) is a common triazole fungicide that is widely used throughout the world in agriculture applications. We previously reported that TEB induces cardiac toxicity in rats. The aim of this study was to investigate the underlying mechanism of the toxicity induced by TEB in cardiac cells. TEB induced dose-dependent cell death in H9c2 cardiomyoblasts and in adult rat ventricular myocytes (ARVM). The comet assay and western blot analysis showed a concentration-dependent increase in DNA damage and in p53 and p21 protein levels 24 h after TEB treatment. Our findings also showed that TEB triggered the mitochondrial pathway of apoptosis as evidenced by a loss of mitochondrial transmembrane potential (ΔΨm), an increase in Bax/Bcl-2 ratio, an activation of caspase-9 and caspase-3, a cleavage of poly (ADP-ribose) polymerase (PARP) and an increase in the proportion of cells in the sub-G1 phase. In addition, TEB promoted ROS production in cardiac cells and consequently increased the amounts of MDA, the end product of lipid peroxidation. Treatment of cardiomyocytes with the ROS scavenger N-acetylcysteine reduced TEB-induced DNA damage and activation of the mitochondrial pathway of apoptosis. These results indicate that the genotoxic and cytotoxic effects of TEB are mediated through a ROS-dependent pathway in cardiac cells.

Inducible Cardiac-Specific Deletion of Sirt1 in Male Mice Reveals Progressive Cardiac Dysfunction and Sensitization of the Heart to Pressure Overload.

Heart failure is associated with profound alterations of energy metabolism thought to play a major role in the progression of this syndrome. SIRT1 is a metabolic sensor of cellular energy and exerts essential functions on energy metabolism, oxidative stress response, apoptosis, or aging. Importantly, SIRT1 deacetylates the peroxisome proliferator-activated receptor gamma co-activator 1α (PGC-1α), the master regulator of energy metabolism involved in mitochondrial biogenesis and fatty acid utilization. However, the exact role of SIRT1 in controlling cardiac energy metabolism is still incompletely understood and conflicting results have been obtained. We generated a cardio-specific inducible model of Sirt1 gene deletion in mice (Sirt1ciKO) to decipher the role of SIRT1 in control conditions and following cardiac stress induced by pressure overload. SIRT1 deficiency induced a progressive cardiac dysfunction, without overt alteration in mitochondrial content or properties. Sixteen weeks after Sirt1 deletion an increase in mitochondrial reactive oxygen species (ROS) production and a higher rate of oxidative damage were observed, suggesting disruption of the ROS production/detoxification balance. Following pressure overload, cardiac dysfunction and alteration in mitochondrial properties were exacerbated in Sirt1ciKO mice. Overall the results demonstrate that SIRT1 plays a cardioprotective role on cardiac energy metabolism and thereby on cardiac function.

Cobalamin and folate protect mitochondrial and contractile functions in a murine model of cardiac pressure overload.

PGC-1α, a key regulator of energy metabolism, seems to be a relevant therapeutic target to rectify the energy deficit observed in heart failure (HF). Since our previous work has shown positive effects of cobalamin (Cb) on PGC-1α cascade, we investigate the protective role of Cb in pressure overload-induced myocardial dysfunction. Mice were fed with normal diet (ND) or with Cb and folate supplemented diet (SD) 3weeks before and 4weeks after transverse aortic constriction (TAC). At the end, left ventricle hypertrophy and drop of ejection fraction were significantly lower in SD mice than in ND mice. Alterations in mitochondrial oxidative capacity, fatty acid oxidation and mitochondrial biogenesis transcription cascade were markedly improved by SD. In SD-TAC mice, lower expression level of the acetyltransferase GCN5 and upregulation of the methyltransferase PRMT1 were associated with a lower protein acetylation and a higher protein methylation levels. This was accompanied by a sustained expression of genes involved in mitochondrial biogenesis transcription cascade (Tfam, Nrf2, Cox1 and Cox4) after TAC in SD mice, suggesting a preserved activation of PGC-1α; this could be at least partly due to corrected acetylation/methylation status of this co-activator. The beneficial effect of the treatment would not be due to an effect of Cb and folate on oxidative stress or on homocysteinemia, which were unchanged by SD. These results showed that Cb and folate could protect the failing heart by preserving energy status through maintenance of mitochondrial biogenesis. It reinforces the concept of a metabolic therapy of HF.

SIRT1 protects cardiac cells against apoptosis induced by zearalenone or its metabolites α- and ß-zearalenol through an autophagy-dependent pathway.

Zearalenone (ZEN) is a non-steroidal estrogenic mycotoxin produced by several species of Fusarium in cereals and agricultural products. The major ZEN metabolites are α-zearalenol (α-ZOL) and ß-zearalenol (ß-ZOL). In the present study, we investigated the underlying mechanism of the toxicity induced by ZEN, α-ZOL and ß-ZOL in cardiac cells (H9c2). We show that treatment with ZEN or its metabolites induces the activation of the mitochondrial pathway of apoptosis as characterized by an increase in ROS generation, a loss of mitochondrial transmembrane potential (ΔΨm) and an activation of caspases. Besides, we demonstrate that these mycotoxins promote the activation of autophagy before the onset of apoptosis. Indeed, we observed that a short-time (6h) treatment with ZEN, α-ZOL or ß-ZOL, increased the level of Beclin-1 and LC3-II and induced the accumulation of the CytoID® autophagy detection probe. Moreover, the inhibition of autophagy by Chloroquine significantly increased cell death induced by ZEN, α-ZOL or ß-ZOL, suggesting that the activation of autophagy serves as a cardioprotective mechanism against these mycotoxins. In addition, we found that the inhibition (EX527) or the knockdown of SIRT1 (siRNA) significantly increased apoptosis induced by ZEN or its derivatives, whereas SIRT1 activation with RSV greatly prevents the cytotoxic effects of these mycotoxins. By contrast, when autophagy was inhibited by CQ, the activation of SIRT1 by RSV had no protection against the cardiotoxicity of ZEN or its metabolites, suggesting that SIRT1 protects cardiac cells by an autophagy-dependent pathway.

Mitochondria: a central target for sex differences in pathologies.

It is increasingly acknowledged that a sex and gender specificity affects the occurrence, development, and consequence of a plethora of pathologies. Mitochondria are considered as the powerhouse of the cell because they produce the majority of energy-rich phosphate bonds in the form of adenosine tri-phosphate (ATP) but they also participate in many other functions like steroid hormone synthesis, reactive oxygen species (ROS) production, ionic regulation, and cell death. Adequate cellular energy supply and survival depend on mitochondrial life cycle, a process involving mitochondrial biogenesis, dynamics, and quality control via mitophagy. It appears that mitochondria are the place of marked sexual dimorphism involving mainly oxidative capacities, calcium handling, and resistance to oxidative stress. In turn, sex hormones regulate mitochondrial function and biogenesis. Mutations in genes encoding mitochondrial proteins are the origin of serious mitochondrial genetic diseases. Mitochondrial dysfunction is also an important parameter for a large panel of pathologies including neuromuscular disorders, encephalopathies, cardiovascular diseases (CVDs), metabolic disorders, neuropathies, renal dysfunction etc. Many of these pathologies present sex/gender specificity. Here we review the sexual dimorphism of mitochondria from different tissues and how this dimorphism takes part in the sex specificity of important pathologies mainly CVDs and neurological disorders.

Citrinin induces apoptosis in human HCT116 colon cancer cells through endoplasmic reticulum stress.

The mycotoxin citrinin (CTN) is a natural contaminant of various human foods that may produce serious adverse health problems. Several studies demonstrated that citrinin exerts cytotoxic and genotoxic effects in both in vivo and in vitro systems. However, the precise mechanisms of action (MOA), particularly in intestinal cells remain unclear. The aim of the present study was to examine the precise MOA of citrinin in vitro. Data demonstrated that CTN significantly decreased the number of viable human intestinal HCT116 cells and induced apoptotic events including (1) decrease in ΔÑ°m indicative of mitochondrial membrane permeabilization, (2) activation of caspase 3, (3) elevated production of reactive oxygen species (ROS) and (4) relative persistence of plasma membrane integrity. Further, the genetic deficiency of the pro-apoptotic protein Bax protected cells against CTN-induced apoptosis, indicating that Bax is required for CTN-mediated toxicity. It was also found that CTN triggered endoplasmic reticulum (ER) stress and activated different arms of the unfolded protein response (UPR) as demonstrated by increase in expression of GRP78 (glucose-regulated protein-78), GRP94 (glucose-regulated protein-94), GADD34 (growth arrest and DNA damage-inducible protein-34), the protein disulfide isomerase associated 6 (PDIA6), CHOP (C/EBP-homologous protein) and the splicing of XBP1 (X-Box Binding Protein 1). Pretreatment of cells with the chemical chaperone 4-phenylbutyrate (PBA), known to alleviate ER stress, prevented significantly the apoptotic process triggered by CTN. Taken together, these results suggest that CTN exerts its cytotoxic effects in HCT116 cells by inducing apoptosis, at least in part, through induction of ER stress.

Emergence of new virulent populations of apple scab from nonagricultural disease reservoirs.

Plant pathogens adapt readily to new crop varieties in agrosystems, and it is crucial to understand the factors underlying the epidemic spread of new virulent strains if we are to develop more efficient strategies to control them. In this study we used multilocus microsatellite typing, molecular epidemiology tools and a large collection of isolates from cultivated, wild and ornamental apples to investigate the origin of new virulent populations of Venturia inaequalis, an ascomycete fungus causing apple scab on varieties carrying the Rvi6 resistance gene. We demonstrated a common origin at the European scale of populations infecting apples (Malus × domestica) carrying the Rvi6 resistance and Malus floribunda, the progenitor of the Rvi6 resistance. Demographic modeling indicated that the Rvi6-virulent lineage separated several thousands of years ago from populations infecting non-Rvi6 hosts, without detectable gene flow between the two lineages. These findings show that 'breakdowns' of plant resistance genes can be caused by the selection and migration of virulent genotypes from standing genetic variation maintained in environmental disease reservoirs, here ornamental crabapples. This work stresses the need to take better account of pathogen diversity in resistance screenings of breeding lines and in resistance deployment strategies, in order to enhance sustainable disease management.

When virulence originates from nonagricultural hosts: evolutionary and epidemiological consequences of introgressions following secondary contacts in Venturia inaequalis.

In pathogens, introgressions through secondary contacts between divergent populations from agricultural and nonagricultural disease reservoirs are expected to have crucial evolutionary and epidemiological implications. Despite the importance of this question for disease management, experimental demonstrations of these implications remain scarce. Recently, we identified a virulent population of the apple scab pathogen Venturia inaequalis that migrated from nonagricultural hosts to European domestic apple orchards. Here, we investigated the occurrence of gene flow between agricultural and nonagricultural populations sampled in two orchards, and thereafter its consequences on the pathogenicity of hybrids. Population genetic structure and demographic inferences based on the genotypes of 104 strains revealed a high amount of gene flow between the two populations in one orchard. In this site, mating between populations was made possible by the presence of a common host. Our results revealed an invasion of the virulent trait in the agricultural population; a main direction of introgression in hybrids from the agricultural to nonagricultural genetic backgrounds; and a population of hybrids with transgressive traits. We demonstrate a secondary contact with gene flow between divergent populations of pathogens. Our findings highlight evolutionary and epidemiological changes in pathogens and have concrete implications for sustainable disease management.

Crocin and quercetin prevent PAT-induced apoptosis in mammalian cells: Involvement of ROS-mediated ER stress pathway.

Patulin (PAT) is a secondary metabolite produced by several species of the genera of Penicillium, Aspergillus, and Byssochlamys that can be found in rotting fruits, especially in apples and apple-based products. Exposure to this mycotoxin has been reported to induce intestinal and kidney injuries. The mechanism underlying such toxicity has been linked to the induction of apoptosis which occurred with reactive oxygen species production and endoplasmic reticulum (ER) stress induction. This study aimed to evaluate the effect of the two common dietary compounds Quercetin (QUER), a natural flavonoid, and Crocin (CRO), a natural carotenoid, on PAT-induced toxicity in human colon carcinoma (HCT116) and embryonic kidney cells (HEK293). We showed that antioxidant properties of QUER and CRO help to prevent ER stress activation and lipid peroxidation as evidenced by the reduction in GRP78 and GADD34 expressions and the decrease in malondialdehyde production. Furthermore, we demonstrated their ability to re-establish the loss of the mitochondrial membrane potential to inhibit caspase 3 activation and DNA fragmentation. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1851-1858, 2016.

Combined effects of alternariols mixture on human colon carcinoma cells.

Mycotoxins are naturally occurring contaminants encountered at high levels in a wide variety of agricultural products intended for human and animal consumptions. Various Alternaria mycotoxins may occur simultaneously in small grain cereals. Considering the concomitant production of alternariol (AOH) and alternariol monomethyl ether (AME), it is expected that humans and animals are exposed to the mixture rather than to individual compounds. Therefore, we studied the interactive effects of binary mixture of alternariols (AOH and AME) on the human intestinal cell line, HCT116 cells. Exposure of HCT116 cells to low cytotoxic alternariols doses, resulted in a moderate cytotoxicity manifested by a loss in the cell viability mediated by an activation of the mitochondrial apoptotic process, associated with the opening of mitochondrial permeability transition pore (PTP) and the loss of the mitochondrial transmembrane potential (ΔΨm). However, when combined, they exert a significant increase in their toxic potential. Altogether, our study showed that AOH and AME combination is obviously additive.

Hsp90 inhibition by PU-H71 induces apoptosis through endoplasmic reticulum stress and mitochondrial pathway in cancer cells and overcomes the resistance conferred by Bcl-2.

Heat shock protein 90 (Hsp90) has recently emerged as an attractive therapeutic target in cancer treatment because of its role in stabilizing the active form of a wide range of client oncoproteins. This study investigated the mechanism of apoptosis induced by the purine-scaffold Hsp90 inhibitor PU-H71 in different human cancer cell lines and examined the role of Bcl-2 and Bax in this process. We demonstrated that Hsp90 inhibition by PU-H71 generated endoplasmic reticulum (ER) stress and activated the Unfolded Protein Response (UPR) as evidenced by XBP1 mRNA splicing and up-regulation of Grp94, Grp78, ATF4 and CHOP. In response to PU-H71-induced ER stress, apoptosis was triggered in melanoma, cervix, colon, liver and lung cancer cells, but not in normal human fibroblasts. Apoptosis was executed through the mitochondrial pathway as shown by down-regulation of Bcl-2, up-regulation and activation of Bax, permeabilization of mitochondrial membranes, release of cytochrome c and activation of caspases. We also found that, in contrast to the ER stressor thapsigargin, PU-H71 induced apoptosis in cells overexpressing Bcl-2 and thus overcame the resistance conferred by this anti-apoptotic protein. In addition, although Bax deficiency rendered cells resistant to PU-H71, combined treatment with the anticancer drugs cisplatin or melphalan greatly sensitized these cells to PU-H71. Taken together, these data suggest that inhibition of Hsp90 by PU-H71 is a promising strategy for cancer treatment, particularly in the case of tumors resistant to conventional chemotherapy.

Coalescence 2.0: a multiple branching of recent theoretical developments and their applications.

Population genetics theory has laid the foundations for genomic analyses including the recent burst in genome scans for selection and statistical inference of past demographic events in many prokaryote, animal and plant species. Identifying SNPs under natural selection and underpinning species adaptation relies on disentangling the respective contribution of random processes (mutation, drift, migration) from that of selection on nucleotide variability. Most theory and statistical tests have been developed using the Kingman coalescent theory based on the Wright-Fisher population model. However, these theoretical models rely on biological and life history assumptions which may be violated in many prokaryote, fungal, animal or plant species. Recent theoretical developments of the so-called multiple merger coalescent models are reviewed here (Λ-coalescent, beta-coalescent, Bolthausen-Sznitman, Ξ-coalescent). We explain how these new models take into account various pervasive ecological and biological characteristics, life history traits or life cycles which were not accounted in previous theories such as (i) the skew in offspring production typical of marine species, (ii) fast adapting microparasites (virus, bacteria and fungi) exhibiting large variation in population sizes during epidemics, (iii) the peculiar life cycles of fungi and bacteria alternating sexual and asexual cycles and (iv) the high rates of extinction-recolonization in spatially structured populations. We finally discuss the relevance of multiple merger models for the detection of SNPs under selection in these species, for population genomics of very large sample size and advocate to potentially examine the conclusion of previous population genetics studies.

The genetic structure of a Venturia inaequalis population in a heterogeneous host population composed of different Malus species.

BACKGROUND: Adaptation, which induces differentiation between populations in relation to environmental conditions, can initiate divergence. The balance between gene flow and selection determines the maintenance of such a structure in sympatry. Studying these two antagonistic forces in plant pathogens is made possible because of the high ability of pathogens to disperse and of the strong selective pressures exerted by their hosts. In this article, we analysed the genetic structure of the population of the apple scab fungus, Venturia inaequalis, in a heterogeneous environment composed of various Malus species. Inferences were drawn from microsatellite and AFLP data obtained from 114 strains sampled in a single orchard on nine different Malus species to determine the forces that shape the genetic structure of the pathogen. RESULTS: Using clustering methods, we first identified two specialist subpopulations: (i) a virulent subpopulation sampled on Malus trees carrying the Rvi6 resistance gene; and (ii) a subpopulation infecting only Malus trees that did not carry this resistance gene. A genome scan of loci on these two subpopulations did not detect any locus under selection. Additionally, we did not detect any other particular substructure linked to different hosts. However, an isolation-by-distance (IBD) pattern at the orchard scale revealed free gene flow within each subpopulation. CONCLUSIONS: Our work shows a rare example of a very strong effect of a resistance gene on pathogen populations. Despite the high diversity of Malus hosts, the presence of Rvi6 seems sufficient to explain the observed genetic structure. Moreover, detection of an IBD pattern at the orchard scale revealed a very low average dispersal distance that is particularly significant for epidemiologists and landscape managers for the design of scab control strategies.

Is local selection so widespread in river organisms? Fractal geometry of river networks leads to high bias in outlier detection.

Identifying local adaptation is crucial in conservation biology to define ecotypes and establish management guidelines. Local adaptation is often inferred from the detection of loci showing a high differentiation between populations, the so-called FST outliers. Methods of detection of loci under selection are reputed to be robust in most spatial population models. However, using simulations we showed that FST outlier tests provided a high rate of false-positives (up to 60%) in fractal environments such as river networks. Surprisingly, the number of sampled demes was correlated with parameters of population genetic structure, such as the variance of FST s, and hence strongly influenced the rate of outliers. This unappreciated property of river networks therefore needs to be accounted for in genetic studies on adaptation and conservation of river organisms.

Effects of Dichlorvos on cardiac cells: Toxicity and molecular mechanism of action.

In this study we aimed to understand the underlying mechanism of Dichlorvos-induced toxicity in cardiac cells. For this end, cells were treated by 170 µM of Dichlorvos (DDVP) (corresponding to the IC50) and molecular events were monitored by flow cytometry and western blotting. We have first demonstrated that cell exposure to DDVP for 24 h induced cell death by necroptosis. In fact, cell treatment with DDVP upregulated RIP1 expression and we have shown that chemical inhibition of RIP1 kinase activity by necrostatin-1 (Nec-1) greatly prevented from the induced cell death. Besides, we have demonstrated that, while there was no observed cell death following short exposure to DDVP (6 h), autophagy was enhanced, as proven by the increase in the level of both Beclin-1 and LC3-II and the accumulation of the CytoID® autophagy detection probe. Besides, when autophagy was inhibited by chloroquine (CQ) the percentage of necroptosis was significantly increased, suggesting that autophagy acts to protect cardiac cells against the toxicity induced by this pesticide. Concurrently, we have shown that the inhibition of the deacetylase sirtuin 1 (SIRT1) by EX527 or its knockdown by siRNA significantly increased DDVP-induced necroptosis, whereas when SIRT1 was activated by resveratrol (RSV) a significant decrease in DDVP-induced cell death was observed. In addition, we revealed that when the autophagy was inhibited by CQ, we can't reveal the protective effect of RSV anymore. Altogether, these results suggest that activation of SIRT1 protects cardiac cells from the toxicity of DDVP through an autophagy-dependent pathway.

EPAC1 inhibition protects the heart from doxorubicin-induced toxicity.

Anthracyclines, such as doxorubicin (Dox), are widely used chemotherapeutic agents for the treatment of solid tumors and hematologic malignancies. However, they frequently induce cardiotoxicity leading to dilated cardiomyopathy and heart failure. This study sought to investigate the role of the exchange protein directly activated by cAMP (EPAC) in Dox-induced cardiotoxicity and the potential cardioprotective effects of EPAC inhibition. We show that Dox induces DNA damage and cardiomyocyte cell death with apoptotic features. Dox also led to an increase in both cAMP concentration and EPAC1 activity. The pharmacological inhibition of EPAC1 (with CE3F4) but not EPAC2 alleviated the whole Dox-induced pattern of alterations. When administered in vivo, Dox-treated WT mice developed a dilated cardiomyopathy which was totally prevented in EPAC1 knock-out (KO) mice. Moreover, EPAC1 inhibition potentiated Dox-induced cell death in several human cancer cell lines. Thus, EPAC1 inhibition appears as a potential therapeutic strategy to limit Dox-induced cardiomyopathy without interfering with its antitumoral activity.