Publications by authors named "Cécile Notarnicola"

19 Publications

  • Page 1 of 1

Prevention of vascular calcification by the endogenous chromogranin A-derived mediator that inhibits osteogenic transdifferentiation.

Basic Res Cardiol 2021 10 13;116(1):57. Epub 2021 Oct 13.

Institute of Molecular Cardiovascular Research, Medical Faculty, RWTH Aachen University, Pauwelsstrasse 30, 52074, Aachen, Germany.

The adrenal glands participate in cardiovascular (CV) physiology and the pathophysiology of CV diseases through their effects on sodium and water metabolism, vascular tone and cardiac function. In the present study, we identified a new adrenal compound controlling mesenchymal cell differentiation that regulates osteoblastic differentiation in the context of vascular calcification. This peptide was named the "calcification blocking factor" (CBF) due to its protective effect against vascular calcification and is released from chromogranin A via enzymatic cleavage by calpain 1 and kallikrein. CBF reduced the calcium content of cells and thoracic aortic rings under calcifying culture conditions, as well as in aortas from animals treated with vitamin D and nicotine (VDN animals). Furthermore, CBF prevented vascular smooth muscle cell (VSMC) transdifferentiation into osteoblast-like cells within the vascular wall via the sodium-dependent phosphate transporter PIT-1 and by inhibition of NF-κB activation and the subsequent BMP2/p-SMAD pathway. Pulse pressure, a marker of arterial stiffness, was significantly decreased in VDN animals treated with CBF. In line with our preclinical data, CBF concentration is significantly reduced in diseases characterized by increased calcification, as shown in patients with chronic kidney disease. In preparation for clinical translation, the active site of the native 19-AS long native CBF was identified as EGQEEEED. In conclusion, we have identified the new peptide CBF, which is secreted from the adrenal glands and might prevent vascular calcification by inhibition of osteogenic transdifferentiation. The anti-calcific effects of CBF and short active site may therefore promote the development of new tools for the prevention and/or treatment of vascular calcification.
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http://dx.doi.org/10.1007/s00395-021-00899-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8514386PMC
October 2021

Optimisation of cell and ex vivo culture conditions to study vascular calcification.

PLoS One 2020 6;15(3):e0230201. Epub 2020 Mar 6.

RD - Néphrologie 2, rue de Mûriers, Montpellier, France.

Medial vascular calcification (MVC) is a highly prevalent disease associated with a high risk of severe, potentially lethal, complications. While animal studies may not systematically be circumvented, in vitro systems have been proven useful to study disease physiopathology. In the context of MVC, the absence of a clinically relevant standardized in vitro method prevents the appropriate comparison and overall interpretation of results originating from different experiments. The aim of our study is to establish in vitro models mimicking in vivo vascular calcification and to select the best methods to unravel the mechanisms involved in MVC. Human aortic smooth muscle cells and rat aortic rings were cultured in different conditions. The influence of fetal calf serum (FCS), alkaline phosphatase, phosphate and calcium concentrations in the medium were evaluated. We identified culture conditions, including the herein reported Aorta Calcifying Medium (ACM), which allowed a reproducible and specific medial calcification of aortic explants. Studying cells and aortic explants cultured, the involvement of bone morphogenetic protein 2 (BMP2) pathway, fibrosis and apoptosis processes in in vitro MVC were demonstrated. Expression of osteoblastic markers was also observed suggesting the occurrence of transdifferentiation of smooth muscle cells to osteoblasts in our models. The use of these models will help researchers in the field of vascular calcification to achieve reproducible results and allow result comparison in a more consistent way.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0230201PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7060075PMC
June 2020

Combination of nutritional polyphenols supplementation with exercise training counteracts insulin resistance and improves endurance in high-fat diet-induced obese rats.

Sci Rep 2018 02 13;8(1):2885. Epub 2018 Feb 13.

PhyMedExp, University of Montpellier, INSERM U1046, CNRS UMR 9214. 34295, Montpellier, cedex 5, France.

Separately, polyphenols and exercise are known to prevent insulin resistance (IR) but their combined curative effects on established obesity and IR require further investigation. Therefore, we compared the metabolic effects of a combination of exercise and grape polyphenols supplementation in obese IR rats with high-fat diet (EXOPP) to the effect of high-fat diet alone (HF) or with a nutritional supplementation of grape polyphenols (PP) or with endurance exercise (EXO) during 8 wks. We observed an improvement of systemic and skeletal muscle insulin sensitivity in EXO and EXOPP rats. EXOPP rats compared to HF rats presented a lower insulinemia and HOMA-IR with higher liver and muscle glycogen contents. Interestingly, EXOPP rats had a 68% enhanced endurance capacity compared to EXO rats with also a higher activation of AMPK compared to sedentary and EXO rats with increased lipid oxidation. Together, our results suggest that grape polyphenols supplementation combined with exercise has a synergistic effect by increasing muscle lipid oxidation and sparing glycogen utilization which thus enhances endurance capacity. Our data highlight that in cases of established obesity and IR, the combination of nutritional grape polyphenols supplementation and exercise heighten and intensify their individual metabolic effects.
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http://dx.doi.org/10.1038/s41598-018-21287-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5811550PMC
February 2018

ER stress disturbs SR/ER-mitochondria Ca transfer: Implications in Duchenne muscular dystrophy.

Biochim Biophys Acta Mol Basis Dis 2017 09 15;1863(9):2229-2239. Epub 2017 Jun 15.

Inserm U1046, UMR CNRS 9214, Université Montpellier, CHRU Montpellier, Montpellier, France. Electronic address:

Besides its role in calcium (Ca) homeostasis, the sarco-endoplamic reticulum (SR/ER) controls protein folding and is tethered to mitochondria. Under pathophysiological conditions the unfolded protein response (UPR) is associated with disturbance in SR/ER-mitochondria crosstalk. Here, we investigated whether ER stress altered SR/ER-mitochondria links, Ca handling and muscle damage in WT (Wild Type) and mdx mice, the murine model of Duchenne Muscular Dystrophy (DMD). In WT mice, the SR/ER-mitochondria links were decreased in isolated FDB muscle fibers after injection of ER stress activator tunicamycin (TM). Ca imaging revealed an increase of cytosolic Ca transient and a decrease of mitochondrial Ca uptake. The force generating capacity of muscle dropped after TM. This impaired contractile function was accompanied by an increase in autophagy markers and calpain-1 activation. Conversely, ER stress inhibitors restored SR/ER-mitochondria links, mitochondrial Ca uptake and improved diaphragm contractility in mdx mice. Our findings demonstrated that ER stress-altered SR/ER-mitochondria links, disturbed Ca handling and muscle function in WT and mdx mice. Thus, ER stress may open up a prospect of new therapeutic targets in DMD.
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http://dx.doi.org/10.1016/j.bbadis.2017.06.009DOI Listing
September 2017

Retinoic acid maintains human skeletal muscle progenitor cells in an immature state.

Cell Mol Life Sci 2017 05 26;74(10):1923-1936. Epub 2016 Dec 26.

Inserm U1046-UMR CNRS 9214 «Physiologie et Médecine Expérimentale du cœur et des muscles-PHYMEDEXP», CHU A. De Villeneuve, Université de Montpellier, Bâtiment Crastes de Paulet, 371 avenue du doyen Giraud, 34295, Montpellier Cedex 5, France.

Muscle satellite cells are resistant to cytotoxic agents, and they express several genes that confer resistance to stress, thus allowing efficient dystrophic muscle regeneration after transplantation. However, once they are activated, this capacity to resist to aggressive agents is diminished resulting in massive death of transplanted cells. Although cell immaturity represents a survival advantage, the signalling pathways involved in the control of the immature state remain to be explored. Here, we show that incubation of human myoblasts with retinoic acid impairs skeletal muscle differentiation through activation of the retinoic-acid receptor family of nuclear receptor. Conversely, pharmacologic or genetic inactivation of endogenous retinoic-acid receptors improved myoblast differentiation. Retinoic acid inhibits the expression of early and late muscle differentiation markers and enhances the expression of myogenic specification genes, such as PAX7 and PAX3. These results suggest that the retinoic-acid-signalling pathway might maintain myoblasts in an undifferentiated/immature stage. To determine the relevance of these observations, we characterised the retinoic-acid-signalling pathways in freshly isolated satellite cells in mice and in siMYOD immature human myoblasts. Our analysis reveals that the immature state of muscle progenitors is correlated with high expression of several genes of the retinoic-acid-signalling pathway both in mice and in human. Taken together, our data provide evidences for an important role of the retinoic-acid-signalling pathway in the regulation of the immature state of muscle progenitors.
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http://dx.doi.org/10.1007/s00018-016-2445-1DOI Listing
May 2017

Homodimerization of RBPMS2 through a new RRM-interaction motif is necessary to control smooth muscle plasticity.

Nucleic Acids Res 2014 Sep 26;42(15):10173-84. Epub 2014 Jul 26.

INSERM U1046, Université Montpellier 1, Université Montpellier 2, 34295 Montpellier, France

In vertebrates, smooth muscle cells (SMCs) can reversibly switch between contractile and proliferative phenotypes. This involves various molecular mechanisms to reactivate developmental signaling pathways and induce cell dedifferentiation. The protein RBPMS2 regulates early development and plasticity of digestive SMCs by inhibiting the bone morphogenetic protein pathway through its interaction with NOGGIN mRNA. RBPMS2 contains only one RNA recognition motif (RRM) while this motif is often repeated in tandem or associated with other functional domains in RRM-containing proteins. Herein, we show using an extensive combination of structure/function analyses that RBPMS2 homodimerizes through a particular sequence motif (D-x-K-x-R-E-L-Y-L-L-F: residues 39-51) located in its RRM domain. We also show that this specific motif is conserved among its homologs and paralogs in vertebrates and in its insect and worm orthologs (CPO and MEC-8, respectively) suggesting a conserved molecular mechanism of action. Inhibition of the dimerization process through targeting a conserved leucine inside of this motif abolishes the capacity of RBPMS2 to interact with the translational elongation eEF2 protein, to upregulate NOGGIN mRNA in vivo and to drive SMC dedifferentiation. Our study demonstrates that RBPMS2 possesses an RRM domain harboring both RNA-binding and protein-binding properties and that the newly identified RRM-homodimerization motif is crucial for the function of RBPMS2 at the cell and tissue levels.
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http://dx.doi.org/10.1093/nar/gku692DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4150794PMC
September 2014

High expression of the RNA-binding protein RBPMS2 in gastrointestinal stromal tumors.

Exp Mol Pathol 2013 Apr 4;94(2):314-21. Epub 2013 Jan 4.

INSERM U1046, Université Montpellier 1, Université Montpellier 2, Montpellier, France.

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal neoplasms of the gastrointestinal tract and are often associated with KIT or PDGFRA gene mutations. GIST cells might arise from the interstitial cells of Cajal (ICCs) or from a mesenchymal precursor that is common to ICCs and smooth muscle cells (SMCs). Here, we analyzed the mRNA and protein expression of RNA-Binding Protein with Multiple Splicing-2 (RBPMS2), an early marker of gastrointestinal SMC precursors, in human GISTs (n=23) by in situ hybridization, quantitative RT-PCR analysis and immunohistochemistry. The mean RBPMS2 mRNA level in GISTs was 42-fold higher than in control gastrointestinal samples (p<0.001). RBPMS2 expression was not correlated with KIT and PDGFRA expression levels, but was higher in GISTs harboring KIT mutations than in tumors with wild type KIT and PDGFRA or in GISTs with PDGFRA mutations that were characterized by the lowest RBPMS2 levels. Moreover, RBPMS2 levels were 64-fold higher in GIST samples with high risk of aggressive behavior than in adult control gastrointestinal samples and 6.2-fold higher in high risk than in low risk GIST specimens. RBPMS2 protein level was high in 87% of the studied GISTs independently of their histological classification. Finally, by inhibiting the KIT signaling pathway in GIST882 cells, we show that RBPMS2 expression is independent of KIT activation. In conclusion, RBPMS2 is up-regulated in GISTs compared to normal adult gastrointestinal tissues, indicating that RBPMS2 might represent a new diagnostic marker for GISTs and a potential target for cancer therapy.
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http://dx.doi.org/10.1016/j.yexmp.2012.12.004DOI Listing
April 2013

The RNA-binding protein RBPMS2 regulates development of gastrointestinal smooth muscle.

Gastroenterology 2012 Sep 5;143(3):687-697.e9. Epub 2012 Jun 5.

INSERM U1046, Université Montpellier 1, Université Montpellier 2, Montpellier, France. Electronic address:

Background & Aims: Gastrointestinal development requires regulated differentiation of visceral smooth muscle cells (SMCs) and their contractile activities; alterations in these processes might lead to gastrointestinal neuromuscular disorders. Gastrointestinal SMC development and remodeling involves post-transcriptional modification of messenger RNA. We investigated the function of the RNA-binding protein for multiple splicing 2 (RBPMS2) during normal development of visceral smooth muscle in chicken and expression of its transcript in human pathophysiological conditions.

Methods: We used avian replication-competent retroviral misexpression approaches to analyze the function of RBPMS2 in vivo and in primary cultures of chicken SMCs. We analyzed levels of RBPMS2 transcripts in colon samples from pediatric patients with Hirschsprung's disease and patients with chronic pseudo obstruction syndrome (CIPO) with megacystis.

Results: RBPMS2 was expressed strongly during the early stage of visceral SMC development and quickly down-regulated in differentiated and mature SMCs. Misexpression of RBPMS2 in differentiated visceral SMCs induced their dedifferentiation and reduced their contractility by up-regulating expression of Noggin, which reduced activity of bone morphogenetic protein. Visceral smooth muscles from pediatric patients with CIPO expressed high levels of RBPMS2 transcripts, compared with smooth muscle from patients without this disorder.

Conclusions: Expression of RBPMS2 is present in visceral SMC precursors. Sustained expression of RBPMS2 inhibits the expression of markers of SMC differentiation by inhibiting bone morphogenetic protein activity, and stimulates SMC proliferation. RBPMS2 transcripts are up-regulated in patients with CIPO; alterations in RBPMS2 function might be involved in digestive motility disorders, particularly those characterized by the presence of muscular lesions (visceral myopathies).
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http://dx.doi.org/10.1053/j.gastro.2012.05.047DOI Listing
September 2012

Activity of the RhoU/Wrch1 GTPase is critical for cranial neural crest cell migration.

Dev Biol 2011 Feb 11;350(2):451-63. Epub 2010 Dec 11.

Universités Montpellier 2 et 1, CRBM, IFR122, Montpellier, France.

The neural crest (NC) is a stem cell-like population that arises at the border of neural and non-neural ectoderm. During development, NC undergoes an epithelio-mesenchymal transition (EMT), i.e. loss of epithelial junctions and acquisition of pro-migratory properties, invades the entire embryo and differentiates into a wide diversity of terminal tissues. We have studied the implication of Rho pathways in NC development and previously showed that RhoV is required for cranial neural crest (CNC) cell specification. We show here that the non-canonical Wnt response rhoU/wrch1 gene, closely related to rhoV, is also expressed in CNC cells but at later stages. Using both gain- and loss-of-function experiments, we demonstrate that the level of RhoU expression is critical for CNC cell migration and subsequent differentiation into craniofacial cartilages. In in vitro cultures, RhoU activates pathways that cooperate with PAK1 and Rac1 in epithelial adhesion, cell spreading and directional cell migration. These data support the conclusion that RhoU is an essential regulator of CNC cell migration.
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http://dx.doi.org/10.1016/j.ydbio.2010.12.011DOI Listing
February 2011

Aldehyde dehydrogenase activity promotes survival of human muscle precursor cells.

J Cell Mol Med 2011 Jan;15(1):119-33

INSERM, ERI 25, Muscle et Pathologies, Montpellier, France.

Aldehyde dehydrogenases (ALDH) are a family of enzymes that efficiently detoxify aldehydic products generated by reactive oxygen species and might therefore participate in cell survival. Because ALDH activity has been used to identify normal and malignant cells with stem cell properties, we asked whether human myogenic precursor cells (myoblasts) could be identified and isolated based on their levels of ALDH activity. Human muscle explant-derived cells were incubated with ALDEFLUOR, a fluorescent substrate for ALDH, and we determined by flow cytometry the level of enzyme activity. We found that ALDH activity positively correlated with the myoblast-CD56(+) fraction in those cells, but, we also observed heterogeneity of ALDH activity levels within CD56-purified myoblasts. Using lentiviral mediated expression of shRNA we demonstrated that ALDH activity was associated with expression of Aldh1a1 protein. Surprisingly, ALDH activity and Aldh1a1 expression levels were very low in mouse, rat, rabbit and non-human primate myoblasts. Using different approaches, from pharmacological inhibition of ALDH activity by diethylaminobenzaldehyde, an inhibitor of class I ALDH, to cell fractionation by flow cytometry using the ALDEFLUOR assay, we characterized human myoblasts expressing low or high levels of ALDH. We correlated high ALDH activity ex vivo to resistance to hydrogen peroxide (H(2) O(2) )-induced cytotoxic effect and in vivo to improved cell viability when human myoblasts were transplanted into host muscle of immune deficient scid mice. Therefore detection of ALDH activity, as a purification strategy, could allow non-toxic and efficient isolation of a fraction of human myoblasts resistant to cytotoxic damage.
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http://dx.doi.org/10.1111/j.1582-4934.2009.00942.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3822499PMC
January 2011

Intermuscular tendons are essential for the development of vertebrate stomach.

Development 2009 Mar 28;136(5):791-801. Epub 2009 Jan 28.

INSERM ERI 25, Muscle and Pathologies, Montpellier Cedex 05, France.

Gastrointestinal motility is ensured by the correct coordination of the enteric nervous system and the visceral smooth muscle cells (SMCs), and defective development of SMCs results in gut malformations and intestinal obstructions. In order to identify the molecular mechanisms that control the differentiation of the visceral mesenchyme into SMCs in the vertebrate stomach, we developed microarrays to analyze the gene expression profiles of undifferentiated and differentiated avian stomachs. We identify Scleraxis, a basic-helix-loop-helix transcription factor, as a new marker of stomach mesenchyme and find that expression of Scleraxis defines the presence of two tendons closely associated to the two visceral smooth muscles. Using targeted gene misexpression, we show that FGF signaling is sufficient to induce Scleraxis expression and to establish two tendon domains adjacent to the smooth muscle structures. We also demonstrate that the tendon organization is perturbed by altering Scleraxis expression or function. Moreover, using primary cells derived from stomach mesenchyme, we find that undifferentiated stomach mesenchyme can give rise to both SMCs and tendon cells. These data show that upon FGF activation, selected stomach mesenchymal cells are primed to express Scleraxis and to differentiate into tendon cells. Our findings identify a new anatomical and functional domain in the vertebrate stomach that we characterize as being two intermuscular tendons closely associated with the visceral SMC structures. We also demonstrate that the coordinated development of both tendon and smooth muscle domains is essential for the correct morphogenesis of the stomach.
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http://dx.doi.org/10.1242/dev.029942DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2802759PMC
March 2009

Biochemical properties of gastrokine-1 purified from chicken gizzard smooth muscle.

PLoS One 2008 5;3(12):e3854. Epub 2008 Dec 5.

INSERM ERI 25 Muscle et Pathologies, Université Montpellier 1, EA 4202, CHU Arnaud de Villeneuve, Montpellier, France.

Unlabelled: The potential role and function of gastrokine-1 (GNK1) in smooth muscle cells is investigated in this work by first establishing a preparative protocol to obtain this native protein from freshly dissected chicken gizzard. Some unexpected biochemical properties of gastrokine-1 were deduced by producing specific polyclonal antibody against the purified protein. We focused on the F-actin interaction with gastrokine-1 and the potential role and function in smooth muscle contractile properties.

Background: GNK1 is thought to provide mucosal protection in the superficial gastric epithelium. However, the actual role of gastrokine-1 with regards to its known decreased expression in gastric cancer is still unknown. Recently, trefoil factors (TFF) were reported to have important roles in gastric epithelial regeneration and cell turnover, and could be involved in GNK1 interactions. The aim of this study was to evaluate the role and function of GNK1 in smooth muscle cells.

Methodology/principal Findings: From fresh chicken gizzard smooth muscle, an original purification procedure was used to purify a heat soluble 20 kDa protein that was sequenced and found to correspond to the gastrokine-1 protein sequence containing one BRICHOS domain and at least two or possibly three transmembrane regions. The purified protein was used to produce polyclonal antibody and highlighted the smooth muscle cell distribution and F-actin association of GNK1 through a few different methods.

Conclusion/significance: Altogether our data illustrate a broader distribution of gastrokine-1 in smooth muscle than only in the gastrointestinal epithelium, and the specific interaction with F-actin highlights and suggests a new role and function of GNK1 within smooth muscle cells. A potential role via TFF interaction in cell-cell adhesion and assembly of actin stress fibres is discussed.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0003854PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2588339PMC
January 2009

Dynamic expression patterns of RhoV/Chp and RhoU/Wrch during chicken embryonic development.

Dev Dyn 2008 Apr;237(4):1165-71

INSERM, ERI 25, Muscle and Pathologies, Université Montpellier I, EA4202, Montpellier, France.

Rho GTPases play central roles in the control of cell adhesion and migration, cell cycle progression, growth, and differentiation. However, although most of our knowledge of Rho GTPase function comes from the study of the three classic Rho GTPases RhoA, Rac1, and Cdc42, recent studies have begun to explore the expression, regulation, and function of some of the lesser-known members of the Rho GTPase family. In the present study, we cloned the avian orthologues of RhoV (or Chp for Cdc42 homologous protein) and RhoU (or Wrch-1 for Wnt-regulated Cdc42 homolog-1) and examined their expression patterns by in situ hybridization analysis both during early chick embryogenesis and later on, during gastrointestinal tract development. Our data show that both GTPases are detected in the primitive streak, the somites, the neural crest cells, and the gastrointestinal tract with distinct territories and/or temporal expression windows. Although both proteins are 90% identical, our results indicate that cRhoV and cRhoU are distinctly expressed during chicken embryonic development.
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http://dx.doi.org/10.1002/dvdy.21507DOI Listing
April 2008

Expression and biological role of the prostaglandin D synthase/SOX9 pathway in human ovarian cancer cells.

Cancer Lett 2007 Oct 29;255(2):182-93. Epub 2007 May 29.

Department of Development and Differentiation, Institut de Génétique Humaine IGH CNRS UPR1142, 141, rue de la Cardonille, 34396 Montpellier, Cedex 5, France.

New therapeutic strategies for ovarian cancer include the identification of involved signaling pathways that could potentially serve as a source of biomarkers for early stages of the disease. In this study, we show that the embryonic male prostaglandin D synthase (Pgds)/SOX9 pathway is expressed at both the RNA and protein levels in different types of human ovarian tumors, pointing to Pgds and SOX9 as possible diagnostic markers for ovarian carcinomas. Using ovarian cancer cell lines, we found, first, that components of the Pgds/SOX9 pathway are expressed in these cells, and second, that treatment of these cells with prostaglandin D2 (PGD2) can inhibit their growth via its DP1 receptor and induce apoptosis. Finally, using siRNA and overexpression strategies, we demonstrate that SOX9 expression is induced by PDG2 and is responsible for PDG2-mediated growth inhibition. Accordingly, as stimulating the PGD2/DP1 signal transduction pathway upregulates SOX9 expression, either activators of this pathway or DP1 agonists may be useful as new therapeutic agents.
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http://dx.doi.org/10.1016/j.canlet.2007.04.007DOI Listing
October 2007

Transient expression of SOX9 protein during follicular development in the adult mouse ovary.

Gene Expr Patterns 2006 Oct 17;6(7):695-702. Epub 2006 Feb 17.

Department of Differentiation and Development, Institut de Génétique Humaine CNRS UPR1142, Montpellier, France.

SOX9 is an essential activating transcription factor that plays a critical role in Sertoli cell differentiation and subsequent testis cord formation. Cytoplasmic SOX9 is present in both sexes during early gonadal embryogenesis. While in males the protein is later translocated into the nucleus of pre-Sertoli cells, its expression is rapidly turned off in females. In mammalian male gonads, SOX9 activates the expression of anti-Müllerian hormone (AMH), a male hormone that initiates Müllerian ducts regression and that is also expressed in postnatal ovarian follicles. Here, we confirm that the SOX9 protein is not present in the immature ovary but also show that SOX9 is transiently expressed in the mature ovary depending on the follicular cycle. Indeed, SOX9 protein was found in the nuclear compartment of the inner cells of the theca interna cell layer which surrounds the pre-antral/antral follicles. In contrast, no expression was detected in the AMH expressing granulosa cells. While these findings exclude the possibility that SOX9 regulates AMH expression in the ovary, they show that SOX9 could nevertheless play a role in the developing follicle.
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http://dx.doi.org/10.1016/j.modgep.2006.01.001DOI Listing
October 2006

Prostaglandin D2 induces nuclear import of the sex-determining factor SOX9 via its cAMP-PKA phosphorylation.

EMBO J 2005 May 5;24(10):1798-809. Epub 2005 May 5.

Institut de Génétique Humaine, CNRS UPR1142, 141 rue de la Cardonille, Montpellier Cedex 5, France.

During mammalian gonadal development, nuclear import/export of the transcription factor SOX9 is a critical step of the Sry-initiated testis-determining cascade. In this study, we identify a molecular mechanism contributing to the SOX9 nuclear translocation in NT2/D1 cells, which is mediated by the prostaglandin D2 (PGD2) signalling pathway via stimulation of its adenylcyclase-coupled DP1 receptor. We find that activation of cAMP-dependent protein kinase A (PKA) induces phosphorylation of SOX9 on its two S64 and S181 PKA sites, and its nuclear localization by enhancing SOX9 binding to the nucleocytoplasmic transport protein importin beta. Moreover, in embryonic gonads, we detect a male-specific prostaglandin D synthase expression and an active PGD2 signal at the time and place of SOX9 expression. We thus propose a new step in the sex-determining cascade where PGD2 acts as an autocrine factor inducing SOX9 nuclear translocation and subsequent Sertoli cell differentiation.
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http://dx.doi.org/10.1038/sj.emboj.7600660DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1142593PMC
May 2005

Identifying mutations in autoinflammatory diseases: towards novel genetic tests and therapies?

Am J Pharmacogenomics 2004 ;4(2):109-18

Laboratoire de Génétique Moléculaire et Chromosomique, Hôpital Arnaud de Villeneuve, Montpellier, France.

Autoinflammatory diseases are defined as illnesses caused by primary dysfunction of the innate immune system. This new concept includes a broad number of disorders, but the spotlight has been focused for the past two years on periodic fevers (familial Mediterranean fever [FMF]; mevalonate kinase deficiency [MVK]; tumor necrosis factor [TNF] receptor-associated periodic syndrome [TRAPS]; cryopyrin-associated periodic syndrome [CAPS]), Crohn's disease and Blau syndrome, thanks to the recent understanding of their molecular basis. Indeed, until recently, these conditions were defined only by phenotypical features, the main ones being recurrent attacks of fever, abdominal pain, arthritis, and cutaneous signs, which sometimes overlap, obscuring diagnosis. The search for distinguishing signs such as periorbital edema in TRAPS, and the use of specific functional tests where available, are valuable. Needless to say, molecular screening of the causative genes has dramatically improved patient quality-of-life by providing early and accurate diagnosis, subsequently allowing for the appropriate treatment. Some patients, however, remain hard to manage despite the advent of new genetic tests, and/or due to the lack of effective treatment. The original clinical link between the aforementioned diseases can now be confirmed by a molecular one, following the exciting discovery that most of the altered proteins are related to the death domain fold (DDF) superfamily involved in inflammation and apoptosis. These molecules mediate the regulation of nuclear factor-kappa B (NF-kappa B) activation, cell apoptosis, and interleukin-1 beta secretion through cross-regulated and, sometimes, common signaling pathways. Knowledge of the defective step in autoinflammation has already led to the elucidation of the mechanisms of action of existing drugs and may allow the development of new therapies.
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http://dx.doi.org/10.2165/00129785-200404020-00005DOI Listing
July 2004

Familial Mediterranean fever is no longer a rare disease in Italy.

Eur J Hum Genet 2003 Jan;11(1):50-6

Department of Internal Medicine, Catholic University, Rome, Italy.

Familial Mediterranean fever (FMF) is an autosomal recessive disorder, characterised by short, recurrent attacks of fever with abdominal, chest or joint pain and erysipelas-like erythema. It is an ethnically restricted genetic disease, found commonly among Mediterranean populations, as well as Armenians, Turks, Arabs and Jews. Traditionally, Italians have been considered little affected by FMF, despite the geographical position of Italy (northern Mediterranean basin) and the migratory changes in its population. The objective was to characterise the demographic, clinical and genetic features of FMF in Italy. Patients of Italian origin were recruited from those referred to Italian-French medical centres for FUO (Fever of Unknown Origin) or 'surgical' emergencies; clinical history, genealogy and physical examination were recorded; all other possible infectious, neoplastic, auto-immune and metabolic diseases were excluded. Mutational analysis of the gene responsible for FMF (MEFV on 16p13.3) was performed, after which geno-phenotypical correlations were established. Italian FMF patients, 40 women and 31 men, aged from 3 to 75 years, have shown all the clinical manifestations indicative of FMF described in the literature, but with a lower incidence of amyloidosis. The genetic tests have been contributive in 42% of cases. The frequency of each different mutation has been similar to that found in a series of 'endemic' countries. The geno-phenotypical correlations have suggested the existence of genetic and/or environmental modifier-factors. Among Italians FMF seems to be more frequent than was believed in the past. The data presented are consistent with their geographical location and their history.
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http://dx.doi.org/10.1038/sj.ejhg.5200916DOI Listing
January 2003

Reduced MEFV messenger RNA expression in patients with familial Mediterranean fever.

Arthritis Rheum 2002 Oct;46(10):2785-93

Laboratoire de Génétique Moléculaire et Chromosomique, Hôpital Arnaud de Villeneuve, 371 Avenue du Doyen Gaston Giraud, 34295 Montpellier Cedex 5, France.

Objective: Familial Mediterranean fever (FMF) is the most common inherited periodic syndrome. The disease phenotype and the almost exclusive expression of the causative gene, MEFV, in leukocytes suggest that this gene plays an important role in the inflammatory cascade. Since most of the known mutations are conservative, we sought to determine how minor DNA defects can give rise to the dramatic phenotypic features seen in FMF.

Methods: To address whether the molecular basis of the phenotype-genotype correlation could be related to altered MEFV messenger RNA (mRNA) expression, we quantified the relative abundance of MEFV transcripts in peripheral blood leukocytes from patients with FMF, healthy carriers of a single MEFV mutation, and healthy control subjects.

Results: We found significantly lower expression of MEFV mRNA in genetically ascertained FMF patients than in healthy controls (0.7 versus 1.1; P = 0.00001). In healthy carriers, the mRNA levels were intermediate, suggesting a true dose-response relationship between the number of mutations and the abundance of MEFV transcripts. The difference between healthy controls and healthy carriers was significant (1.1 versus 0.8; P = 0.008), demonstrating that the decrease in mRNA expression is related to a molecular defect independent of FMF symptoms. MEFV mRNA expression was also found to be a function of the type of mutations. The lowest MEFV levels were found in healthy carriers and patients with M694V. Moreover, we observed an inverse correlation with the clinical severity score (r = -0.6, P = 0.04 and r = -0.6, P = 0.004 in patients with 1 and 2 M694V mutations, respectively).

Conclusion: Our results demonstrate that MEFV message levels are related to both the genotype and the phenotype, and suggest that the pathophysiology of FMF relies on a quantitative defect of MEFV mRNA expression.
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http://dx.doi.org/10.1002/art.10575DOI Listing
October 2002
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