Distinct requirements of wls, wnt9a, wnt5b and gpc4 in regulating chondrocyte maturation and timing of endochondral ossification

Formation of the mandible requires progressive morphologic change, proliferation, differentiation and organization of chondrocytes preceding osteogenesis. The Wnt signaling pathway is involved in regulating bone development and maintenance. Chondrocytes that are fated to become bone require Wnt to polarize and orientate appropriately to initiate the endochondral ossification program. Although the canonical Wnt signaling has been well studied in the context of bone development, the effects of non-canonical Wnt signaling in regulating the timing of cartilage maturation and subsequent bone formation in shaping ventral craniofacial structure is not fully understood.. Here we examined the role of the non-canonical Wnt signaling pathway (wls, gpc4, wnt5b and wnt9a) in regulating zebrafish Meckel's cartilage maturation to the onset of osteogenic differentiation. We found that disruption of wls resulted in a significant loss of craniofacial bone, whereas lack of gpc4, wnt5b and wnt9a resulted in severely delayed endochondral ossification. This study demonstrates the importance of the non-canonical Wnt pathway in regulating coordinated ventral cartilage morphogenesis and ossification

Utero-vaginal aplasia (Mayer-Rokitansky-Küster-Hauser syndrome) associated with deletions in known DiGeorge or DiGeorge-like loci

<p>Abstract</p> <p>Background</p> <p>Mayer-Rokitansky-Küster-Hauser (MRKH) syndrome is characterized by congenital aplasia of the uterus and the upper part of the vagina in women showing normal development of secondary sexual characteristics and a normal 46, XX karyotype. The uterovaginal aplasia is either isolated (type I) or more frequently associated with other malformations (type II or Müllerian Renal Cervico-thoracic Somite (MURCS) association), some of which belong to the malformation spectrum of DiGeorge phenotype (DGS). Its etiology remains poorly understood. Thus the phenotypic manifestations of MRKH and DGS overlap suggesting a possible genetic link. This would potentially have clinical consequences.</p> <p>Methods</p> <p>We searched DiGeorge critical chromosomal regions for chromosomal anomalies in a cohort of 57 subjects with uterovaginal aplasia (55 women and 2 aborted fetuses). For this candidate locus approach, we used a multiplex ligation-dependent probe amplification (MLPA) assay based on a kit designed for investigation of the chromosomal regions known to be involved in DGS.</p> <p>The deletions detected were validated by Duplex PCR/liquid chromatography (DP/LC) and/or array-CGH analysis.</p> <p>Results</p> <p>We found deletions in four probands within the four chromosomal loci 4q34-qter, 8p23.1, 10p14 and 22q11.2 implicated in almost all cases of DGS syndrome.</p> <p>Conclusion</p> <p>Uterovaginal aplasia appears to be an additional feature of the broad spectrum of the DGS phenotype. The DiGeorge critical chromosomal regions may be candidate loci for a subset of MRKH syndrome (MURCS association) individuals. However, the genes mapping at the sites of these deletions involved in uterovaginal anomalies remain to be determined. These findings have consequences for clinical investigations, the care of patients and their relatives, and genetic counseling.</p

Identification et validation de gènes candidats pour l holoprosencéphalie

L'Holoprosencéphalie (HPE) est la malformation cérébrale congénitale la plus commune (1/250 fœtus), résultant d'un défaut de clivage du cerveau antérieur. Nous travaillons sur l'HPE isolée qui représente environ 40% des cas. Quatre gènes HPE majeurs ont été mis en évidence (SHH, SIX3, ZIC2, TGIF1). Les mutations ou délétions de ces gènes nous permettent d'expliquer seulement 70% des cas, il reste donc encore d'autres gènes HPE à identifier. Dans ce but, nous recherchons des réarrangements récurrents chez nos patients grâce à la technique CGH array. Au cours de ce travail de thèse, l'étude d'une cohorte de 111 patients, nous a permis de mettre en évidence principalement deux gènes candidats. Ces travaux présentent l'identification puis la validation de l'implication de ces gènes chez les modèles animaux.Holoprosencephaly (HPE) is the most common congenital brain malformation (1 / 250 fetuses), resulting from a defect in cleavage of the forebrain.We study isolated HPE, which represents approximately 40% of cases. Four genes have been identified as responsible for the HPE (SHH, SIX3, ZIC2, TGIF1).Mutations or deletions of these genes allow us to explain only 70% of cases; we still have to identify other HPE genes.Thus, we look for recurrent rearrangements in our patients using array CGH.In this thesis, the study of a cohort of 111 patients allowed us to highlight two main candidate genes.This work presents the identification and validation of the involvement of these genes in animal models.RENNES1-BU Sciences Philo (352382102) / SudocSudocFranceF

Holoprosencephaly: An update on cytogenetic abnormalities.: holoprosencephaly and cytogenetics

International audienceHoloprosencephaly (HPE), the most common developmental defect of the forebrain and midface, is caused by a failure of midline cleavage early in gestation. Isolated HPE, which is highly genetically heterogeneous, can be due to major chromosomal abnormalities. Initially, karyotype approach led to the identification of several recurrent chromosomal anomalies predicting different HPE loci. Subsequently, several genes were isolated from these critical HPE regions, but point mutations and deletions in these genes were found only in 25% of the genetic cases. In order to identify other HPE genes, a more accurate investigation of the genome in HPE patients was necessary. To date, high-resolution cytogenetic techniques such as subtelomeric multiplex ligation-dependent probe amplification (MLPA) and microarray-based comparative genomic hybridization (array CGH) have enhanced chromosomal aberration analysis. In this article, we have updated the cytogenetic anomalies associated with HPE in a map listing all the subtelomeric and interstitial deletions that have been characterized either by karyotype, MLPA, or array CGH. The accumulation of recurrent genomic imbalances will lead to the further delineation of minimal critical HPE loci, which is the first step to the identification of new HPE genes

Production d’alevins : 2.5 Action n°19

National audienceDans ce chapitre, seules les principales méthodologies employées durant la saison 2008 ont été décrites. Elles devront évoluer dans les années à venir en fonction de l’avancée des connaissances et des retours d’expériences

Amélioration des techniques d’élevage larvaire et repeuplement : 2.6 Action n°20

Extrait de documentL’objectif de cette action était d’optimiser les croissances et les survies des alevins, dans une optique de limitation des coûts. En 2008, compte tenu du nombre important de larves produites, il n’y a eu que peu de temps pour l’expérimentation au sens strict du terme sur cet aspect. Des tests comparatifs en conditions de production ont été menés. Ils ont porté : sur les proies (quantités et mode de distribution des artémies), les caractéristiques des bassins (auges et bassins cylindro-coniques), les densités de larves, et l’âge pour le transfert en pré grossissement. Les travaux sur le sevrage précoce ont été limités à deux expérimentations

NOTCH, a new signaling pathway implicated in holoprosencephaly.

International audienceGenetics of Holoprosencephaly (HPE), a congenital malformation of the developing human forebrain, is due to multiple genetic defects. Most genes that have been implicated in HPE belong to the sonic hedgehog signaling pathway. Here we describe a new candidate gene isolated from array comparative genomic hybridization redundant 6qter deletions, DELTA Like 1 (DLL1), which is a ligand of NOTCH. We show that DLL1 is co-expressed in the developing chick forebrain with Fgf8. By treating chick embryos with a pharmacological inhibitor, we demonstrate that DLL1 interacts with FGF signaling pathway. Moreover, a mutation analysis of DLL1 in HPE patients revealed a three-nucleotide deletion. These various findings implicate DLL1 in early patterning of the forebrain and identify NOTCH as a new signaling pathway involved in HPE

Array-CGH analysis indicates a high prevalence of genomic rearrangements in holoprosencephaly: an updated map of candidate loci.

International audienceHoloprosencephaly (HPE) is the most frequent malformation of the brain. To date, 12 different HPE loci and 8 HPE genes have been identified from recurrent chromosomal rearrangements or from the sequencing of genes from Nodal and SHH pathways. Our cohort of HPE patients presents a high genetic heterogeneity. Point mutations were found in SHH, ZIC2, SIX3, and TGIF genes in about 20% of cases (with 10% in SHH). Deletions in these same genes were found in 7.5% of the patients and 4.4% presented with other subtelomeric gain or losses. Consequently, the molecular basis of HPE remains unknown in 70% of our cohorts. To detect new HPE candidate genes, we used array-CGH to refine the previous karyotype based HPE loci map. We analyzed 111 HPE patients with high-performance Agilent oligonucleotidic arrays and found that 28 presented anomalies involving known or new potential HPE loci located on different chromosomes but with poor redundancy. This study showed an impressive rate of 19 patients among 111 with de novo chromosomal anomalies giving evidence that microrearrangements could be a major molecular mechanism in HPE. Additionally, this study opens new insights on HPE candidate genes identification giving an updated HPE candidate loci map