Virginie Millet

Bio: Virginie Millet is an academic researcher from French Institute of Health and Medical Research. The author has contributed to research in topics: Pantetheinase & Colitis. The author has an hindex of 12, co-authored 18 publications receiving 919 citations. Previous affiliations of Virginie Millet include Centre national de la recherche scientifique & Aix-Marseille University.

Short-term BMP-2 expression is sufficient for in vivo osteochondral differentiation of mesenchymal stem cells.

Abstract: Currently available murine models to evaluate mesenchymal stem cell (MSC) differentiation are based on cell injection at ectopic sites such as muscle or skin. Due to the importance of environmental factors on the differentiation capacities of stem cells in vivo, we investigated whether the peculiar synovial/cartilaginous environment may influence the lineage specificity of bone morphogenetic protein (BMP)-2-engineered MSCs. To this aim, we used the C3H10T1/2-derived C9 MSCs that express BMP-2 under control of the doxycycline (Dox)-repressible promoter, Tet-Off, and showed in vitro, using the micropellet culture system that C9 MSCs kept their potential to differentiate toward chondrocytes. Implantation of C9 cells, either into the tibialis anterior muscles or into the joints of CB17-severe combined immunodeficient bg mice led to the formation of cartilage and bone filled with bone marrow as soon as day 10. However, no differentiation was observed after injection of naive MSCs or C9 cells that were repressed to secrete BMP-2 by Dox addition. The BMP-2-induced differentiation of adult MSCs is thus independent of soluble factors present in the local environment of the synovial/cartilaginous tissues. Importantly, we demonstrated that a short-term expression of the BMP-2 growth factor is necessary and sufficient to irreversibly induce bone formation, suggesting that a stable genetic modification of MSCs is not required for stem cell-based bone/cartilage engineering.

Vanin-1-/- mice exhibit a glutathione-mediated tissue resistance to oxidative stress.

Abstract: Oxidative stress is associated with the development and persistence of numerous physiopathological disorders. To date, an active area of investigation is focused on the cellular antioxidant network and the capacity of redox changes to trigger various biological events that include cell proliferation, differentiation, apoptosis, and inflammation. Glutathione (GSH) is considered to be the major thiol-disulfide redox buffer of the cell and tissues, and regulation of its metabolism has become a major therapeutic target in tissue repair. Elimination of free radicals (i.e., reactive oxygen species [ROS]) and their toxic products occurs through the oxidation of GSH to glutathione disulfide (GSSG), subsequently regenerated by the glutathione reductase (GSH-Red). GSH homeostasis is also controlled by the gamma-glutamylcysteine synthetase (γGCS), the rate-limiting enzyme for GSH synthesis (17). The abundance of the γGCS mRNA is itself stress regulated via antioxidant response elements (AREs) found in the 5′-flanking region of the γGCS gene (21, 32, 47), and these AREs also regulate the coordinate induction of numerous oxidative stress response genes (36, 45). Gamma irradiation disrupts water molecules, producing hydroxyl radicals and thus leading to oxidative damage and apoptosis in dividing cells. For example, the rapid renewal of the small intestine epithelium renders it highly sensitive to ionizing radiation, which provokes disruption of the mucosal integrity and progenitor crypt cell loss (42). The response of the intestinal epithelium to γ-irradiation is an established model to study the dynamics of epithelial cell regeneration after injury. Monitoring of γ-irradiation-induced thymic depletion is another classical strategy to assess the radiosensitivity of an animal. On whole-body irradiation, the thymus undergoes a drastic involution involving thymocyte death and stromal cell disorganization; it is fully reconstituted by bone marrow precursor cell injection (2). This “depletion-regeneration” model is convenient to study postirradiation tissue repair (43). We have chosen this approach to investigate the early events in interactions between thymocytes and thymic stromal cells and have identified Vanin-1, a cell surface molecule expressed by a subset of stromal cells, as being involved in postirradiation thymus reconstitution (3). Vanin-1 is the prototypic member of a novel family of ectoenzymes including at least two proteins in mice (i.e., Vanin-1 and Vanin-3), three in humans (i.e., VNN1, VNN2, and VNN3), and Drosophila homologues (10, 11, 29). In mice, Vanin-1 and Vanin-3 expression is associated mostly with epithelial and myeloid cells, respectively (10, 11, 29). All Vanin molecules are pantetheinases (EC 3.5.1.-) capable of specifically hydrolyzing pantetheine into pantothenic acid (vitamin B5) and cysteamine, a sulfhydryl compound used for antioxidant properties (9, 28, 34). The most important consequence observed in Vanin-1−/− mice is the lack of cysteamine in tissues where Vanin-1 expression is predominant, allowing us to probe the relative importance of this metabolite in the postirradiation tissue response (34). We report here that Vanin-1−/− mice are more resistant to paraquat poisoning and exposure to lethal doses of γ-irradiation. Following irradiation, these mice display facilitated thymic reconstitution and a reduced apoptotic response in the small intestine, both of which are associated with a milder tissue inflammation. This protection is related to changes in the detoxifying potential of Vanin-1−/− tissues, characterized by elevated GSH stores. Importantly, intraperitoneal administration of cystamine (the disulfide form of cysteamine) abrogates the resistant phenotype of the mutant mice, suggesting that Vanin-1 regulates at least in part through cysteamine, the GSH-associated metabolism and modulates the adaptive tissue response to stress. On the other hand, Vanin gene expression is regulated during tissue reconstitution following γ-irradiation. The Vanin-1 gene promoter contains ARE-like elements involved in the enhanced expression of the gene on stress stimulation. We propose that Vanin/pantetheinase inhibitors could have useful applications in the therapy of damage due to radiation or other pro-oxidant inducers.

Vanin-1 licenses inflammatory mediator production by gut epithelial cells and controls colitis by antagonizing peroxisome proliferator-activated receptor γ activity

Abstract: Colitis involves immune cell–mediated tissue injuries, but the contribution of epithelial cells remains largely unclear. Vanin-1 is an epithelial ectoenzyme with a pantetheinase activity that provides cysteamine/cystamine to tissue. Using the 2,4,6-trinitrobenzene sulfonic acid (TNBS)-colitis model we show here that Vanin-1 deficiency protects from colitis. This protection is reversible by administration of cystamine or bisphenol A diglycidyl ether, a peroxisome proliferator-activated receptor (PPAR)γ antagonist. We further demonstrate that Vanin-1, by antagonizing PPARγ, licenses the production of inflammatory mediators by intestinal epithelial cells. We propose that Vanin-1 is an epithelial sensor of stress that exerts a dominant control over innate immune responses in tissue. Thus, the Vanin-1/pantetheinase activity might be a new target for therapeutic intervention in inflammatory bowel disease.

Tetracycline-Inducible Interleukin-10 Gene Transfer Mediated by an Adeno-Associated Virus: Application to Experimental Arthritis

Abstract: The adeno-associated viruses (AAV) offer new perspectives for cytokine gene transfer in rheumatoid arthritis (RA) because they are nonpathogenic and allow long-term transgene expression in vivo. Moreover, the use of a tetracycline-inducible promoter allows regulation of therapeutic gene expression. This study assessed the potential long-term gene regulation of a recombinant AAV vector expressing viral interleukin-10 (vIL-10) in human rheumatoid synovium and the therapeutic efficiency in a mouse model of RA. We constructed a recombinant AAV vector in which the transcription of vIL-10 cDNA is controlled by the TetON system. Transduction of human primary RA synovial cells with AAV-tetON-vIL10 conferred in vitro controlled vIL-10 expression. After intramuscular injection, both incidence and severity of collagen-induced arthritis were significantly reduced at macroscopic, radiological, and histological levels in the group of DBA1 mice treated with AAV-TetON-vIL10 vector plus doxycycline after immunization and ...

Tetracycline transcriptional silencer tightly controls transgene expression after in vivo intramuscular electrotransfer: application to interleukin 10 therapy in experimental arthritis.

Abstract: The doxycycline (Dox)-inducible reverse tetracycline transactivator (rtTA) is often used to control gene expression. However, the Tet-on system displays a high background activity. To overcome this unregulated expression we used the tetracycline-dependent transcriptional silencer (tTS), which binds the tetO inducible promoter in the absence of Dox. Controlled gene expression was analyzed in vivo by delivering combinations of Dox-regulated luciferase reporter construct, rtTA, and tTS expression plasmids into mouse muscle, using electrotransfer. Elevated luciferase expression levels were observed in the absence of doxycycline, and a 10-fold induction was obtained after drug administration. In contrast, when tTS was added, background expression was dramatically lowered by three to four orders of magnitude, and induction was maintained. The tTS system was then used to control expression of a therapeutic gene in experimental arthritis. DBA/1 mice were coinjected with plasmids encoding the antiinflammatory interleukin-10 cytokine under the control of the tetO promoter, the rtTA, and the tTS. Electrotransfer resulted in a dose-dependent increase in IL-10 expression, maintained over a 3-month period, and significant inhibitory effects on collagen-induced arthritis. We conclude that the use of tTS significantly improves the utility of the rtTA system for somatic gene transfer by reducing background activity.

TGF-β and BMP Signaling in Osteoblast Differentiation and Bone Formation

Abstract: Transforming growth factor-beta (TGF-β)/bone morphogenic protein (BMP) signaling is involved in a vast majority of cellular processes and is fundamentally important throughout life. TGF-β/BMPs have widely recognized roles in bone formation during mammalian development and exhibit versatile regulatory functions in the body. Signaling transduction by TGF-β/BMPs is specifically through both canonical Smad-dependent pathways (TGF-β/BMP ligands, receptors and Smads) and non-canonical Smad-independent signaling pathway (e.g. p38 mitogen-activated protein kinase pathway, MAPK). Following TGF-β/BMP induction, both the Smad and p38 MAPK pathways converge at the Runx2 gene to control mesenchymal precursor cell differentiation. The coordinated activity of Runx2 and TGF-β/BMP-activated Smads is critical for formation of the skeleton. Recent advances in molecular and genetic studies using gene targeting in mice enable a better understanding of TGF-β/BMP signaling in bone and in the signaling networks underlying osteoblast differentiation and bone formation. This review summarizes the recent advances in our understanding of TGF-β/BMP signaling in bone from studies of genetic mouse models and human diseases caused by the disruption of TGF-β/BMP signaling. This review also highlights the different modes of cross-talk between TGF-β/BMP signaling and the signaling pathways of MAPK, Wnt, Hedgehog, Notch, and FGF in osteoblast differentiation and bone formation.

Matrix metalloproteinases: role in arthritis.

Abstract: The irreversible destruction of the cartilage, tendon, and bone that comprise synovial joints is the hallmark of both rheumatoid arthritis (RA) and osteoarthritis (OA). While cartilage is made up of proteoglycans and type II collagen, tendon and bone are composed primarily of type I collagen. RA is an autoimmune disease afflicting numerous joints throughout the body; in contrast, OA develops in a small number of joints, usually resulting from chronic overuse or injury. In both diseases, inflammatory cytokines such as interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) stimulate the production of matrix metalloproteinases (MMPs), enzymes that can degrade all components of the extracellular matrix. The collagenases, MMP-1 and MMP-13, have predominant roles in RA and OA because they are rate limiting in the process of collagen degradation. MMP-1 is produced primarily by the synovial cells that line the joints, and MMP-13 is a product of the chondrocytes that reside in the cartilage. In addition to collagen, MMP-13 also degrades the proteoglycan molecule, aggrecan, giving it a dual role in matrix destruction. Expression of other MMPs such as MMP-2, MMP-3 and MMP-9, is also elevated in arthritis and these enzymes degrade non-collagen matrix components of the joints. Significant effort has been expended in attempts to design effective inhibitors of MMP activity and/or synthesis with the goal of curbing connective tissues destruction within the joints. To date, however, no effective clinical inhibitors exist. Increasing our knowledge of the crystal structures of these enzymes and of the signal transduction pathways and molecular mechanisms that control MMP gene expression may provide new opportunities for the development of therapeutics to prevent the joint destruction seen in arthritis.

Positive and negative selection of the T cell repertoire: what thymocytes see (and don't see).

Abstract: The fate of developing T cells is specified by the interaction of their antigen receptors with self-peptide-MHC complexes that are displayed by thymic antigen-presenting cells (APCs). Various subsets of thymic APCs are strategically positioned in particular thymic microenvironments and they coordinate the selection of a functional and self-tolerant T cell repertoire. In this Review, we discuss the different strategies that these APCs use to sample and process self antigens and to thereby generate partly unique, 'idiosyncratic' peptide-MHC ligandomes. We discuss how the particular composition of the peptide-MHC ligandomes that are presented by specific APC subsets not only shapes the T cell repertoire in the thymus but may also indelibly imprint the behaviour of mature T cells in the periphery.

Peroxisome Proliferator-Activated Receptor Alpha Target Genes

Abstract: The peroxisome proliferator-activated receptor alpha (PPARα) is a ligand-activated transcription factor involved in the regulation of a variety of processes, ranging from inflammation and immunity to nutrient metabolism and energy homeostasis. PPARα serves as a molecular target for hypolipidemic fibrates drugs which bind the receptor with high affinity. Furthermore, PPARα binds and is activated by numerous fatty acids and fatty acid-derived compounds. PPARα governs biological processes by altering the expression of a large number of target genes. Accordingly, the specific role of PPARα is directly related to the biological function of its target genes. Here, we present an overview of the involvement of PPARα in lipid metabolism and other pathways through a detailed analysis of the different known or putative PPARα target genes. The emphasis is on gene regulation by PPARα in liver although many of the results likely apply to other organs and tissues as well.