Hepatitis C virus (HCV) core, known to be involved in liver carcinogenesis, is processed in the endoplasmic reticulum (ER). We thus investigated the impact of three HCV core isolates on ER stress, ER calcium signalling and apoptosis. We show that HCV core constructs trigger hyperexpression of Grp78/BiP, Grp 94, calreticulin and sarco/endoplasmic reticulum calcium ATPase, inducing ER stress. By using the ER-targeted aequorin calcium probe, we found that ER calcium depletion follows ER stress in core-expressing cells. HCV core induces apoptosis through overexpression of the CHOP/ GADD153 proapoptotic factor, Bax translocation to mitochondria, mitochondrial membrane depolarization, cytochrome c release, caspase-3 and PARP cleavage. Furthermore, reversion of HCV core-induced ER calcium depletion (by transfection of SERCA2) completely abolished mitochondrial membrane depolarization, suggesting that both ER stress (through CHOP overexpression) and calcium signalling play a major role in the HCV core-mediated control of apoptosis. ER stress and apoptosis were also found in a proportion of HCV-fulllength replicon-expressing cells and in the liver of HCV core transgenic mice. In conclusion, our data demonstrate that HCV core deregulates the control of apoptosis by inducing ER stress and ER calcium depletion providing new elements to understand the mechanisms involved in HCV-related liver chronic diseases.

Normal cells reach senescence after a specific time and number of divisions, leading ultimately to cell death. Although escape from this fate may be a requisite step in neoplastic transformation, the mechanisms governing senescent cell death have not been well investigated. We show here, using normal human epidermal keratinocytes, that no apoptotic markers appear with senescence. In contrast, the expression of several proteins involved in the regulation of macroautophagy, notably Beclin-1 and Bcl-2, was found to change with senescence. The corpses occurring at the senescence growth plateau displayed a large central area delimited by the cytokeratin network that contained a huge quantity of autophagic vacuoles, the damaged nucleus, and most mitochondria. 3-methyladenine, an inhibitor of autophagosome formation, but not the caspase inhibitor zVAD, prevented senescent cell death. We conclude that senescent cells do not die by apoptosis , but as a result of high macroautophagic activity that targets the primary vital cell components.

Thermococcales are hyperthermophilic archaea found in deep-sea hydrothermal vents. They have been recently reported to produce membrane vesicles (MVs) into their culture medium. Here, we have characterized the mode of production and determined the biochemical composition of MVs from two species of Thermococcales, Thermococcus gammatolerans and Thermococcus kodakaraensis. We observed that MVs are produced by a budding process from the cell membrane reminiscent of ectosome (microparticle) formation in eukaryotes. MVs and cell membranes from the same species have a similar protein and lipid composition, confirming that MVs are produced from cell membranes. The major protein present in cell membranes and MVs of both species is the oligopeptide binding protein OppA. This protein is also abundant in MVs from cells grown in minimal medium, suggesting that OppA could be involved in processes other than peptides scavenging. We have previously shown that MVs from Thermococcales harbour DNA and protect DNA against thermodegradation. Here, we show that T. kodakaraensis cells transformed with the shuttle plasmid pLC70 release MVs harbouring this plasmid. Notably, these MVs can be used to transfer pLC70 into plasmid-free cells, suggesting that MVs could be involved in DNA transfer between cells at high temperature.

Live imaging studies of the processes of demyelination and remyelination have so far been technically limited in mammals. We have thus generated a Xenopus laevis transgenic line allowing live imaging and conditional ablation of myelinating oligodendrocytes throughout the CNS. In these transgenic pMBP-eGFP-NTR tadpoles the myelin basic protein (MBP) regulatory sequences, specific to mature oligodendrocytes, are used to drive expression of an eGFP (enhanced green fluorescent protein) reporter fused to the Escherichia coli nitroreductase (NTR) selection enzyme. This enzyme converts the innocuous prodrug metronidazole (MTZ) to a cytotoxin. Using two-photon imaging in vivo, we show that pMBP-eGFP-NTR tadpoles display a graded oligodendrocyte ablation in response to MTZ, which depends on the exposure time to MTZ. MTZ-induced cell death was restricted to oligodendrocytes, without detectable axonal damage. After cessation of MTZ treatment, remyelination proceeded spontaneously, but was strongly accelerated by retinoic acid. Altogether, these features establish the Xenopus pMBP-eGFP-NTR line as a novel in vivo model for the study of demyelination/remyelination processes and for large-scale screens of therapeutic agents promoting myelin repair.

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Water soluble poly(3-hydroxyalkanoate) containing ionic groups was designed by two successive photoactivated thiol-ene reactions. Sodium-3-mercapto-1-ethanesulfonate (SO3-) and poly(ethylene glycol) methyl ether thiol (PEG) were grafted onto poly(3-hydroxyoctanoate-co-3-hydroxyundecenoate) PHO(67)U(33) to both introduce ionic groups and hydrophilic moieties. The grafted copolymers PHO(67)SO3-(20)PEG(13) were then used as biocompatible coatings of nano-Metal Organic Frameworks surface. Scanning Electron Microscopy and Scanning Transmission Electron Microscopy coupled with Energy Dispersive X-ray characterizations have clearly demonstrated the presence of the copolymer on the MOF surface. These coated nano-MOF are stable in aqueous and physiological fluids. Cell proliferation and cytotoxicity tests performed on murine macrophages J774.A1 revealed no cytotoxic side effect. Thus, biocompatibility and stability of these novel hybrid porous MOF structures encourage their use in the development of effective therapeutic nanoparticles.

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In the previous work, the poly(4-vinyl dimethyl dipicolinate) (PVDPM) polymer grafted on poly(vinyl chloride) (PVC) and poly(ethylene terephthalate) (PET) film or industrial fibers proved its efficiency for trapping uranium and many lanthanides in a liquid environment. In this work, we propose to graft our active polymer on fibers obtained by electrospinning to increase the specific surface of our final material. To address this challenge, surface grafting of electrospun chlorinated poly(vinyl chloride) (PVC-co-CPVC) fibers using supplemental activation reducing agent atom transfer radical polymerization (SARA-ATRP) of poly(4-vinyldipicolinic acid) (PVDPA, acid form of PVDPM) were successfully prepared. The (PVC-co-CPVC)-g-PVDPA fibers show a fast increase in the degree of grafting that goes from 54% after 4 h to 369% after 72 h. Further insights demonstrate successful complexation between the new functional electrospun (PVC-co-CPVC)-g-PVDPA fibers and europium, which proves that it could be a potential candidate for scavenging lanthanides or any agent able to interact with ligands (PVDPA), which opens up innovative application perspectives in the field of polymeric materials.