Novel Insights into the Bovine Polled Phenotype and Horn Ontogenesis in Bovidae

Despite massive research efforts, the molecular etiology of bovine polledness and the developmental pathways involved in horn ontogenesis are still poorly understood. In a recent article, we provided evidence for the existence of at least two different alleles at the Polled locus and identified candidate mutations for each of them. None of these mutations was located in known coding or regulatory regions, thus adding to the complexity of understanding the molecular basis of polledness. We confirm previous results here and exhaustively identify the causative mutation for the Celtic allele (PC) and four candidate mutations for the Friesian allele (PF). We describe a previously unreported eyelash-and-eyelid phenotype associated with regular polledness, and present unique histological and gene expression data on bovine horn bud differentiation in fetuses affected by three different horn defect syndromes, as well as in wild-type controls. We propose the ectopic expression of a lincRNA in PC/p horn buds as a probable cause of horn bud agenesis. In addition, we provide evidence for an involvement of OLIG2, FOXL2 and RXFP2 in horn bud differentiation, and draw a first link between bovine, ovine and caprine Polled loci. Our results represent a first and important step in understanding the genetic pathways and key process involved in horn bud differentiation in Bovidae

LIPH Expression in Skin and Hair Follicles of Normal Coat and Rex Rabbits

Natural mutations in the LIPH gene were shown to be responsible for hair growth defects in humans and for the rex short hair phenotype in rabbits. In this species, we identified a single nucleotide deletion in LIPH (1362delA) introducing a stop codon in the C-terminal region of the protein. We investigated the expression of LIPH between normal coat and rex rabbits during critical fetal stages of hair follicle genesis, in adults and during hair follicle cycles. Transcripts were three times less expressed in both fetal and adult stages of the rex rabbits than in normal rabbits. In addition, the hair growth cycle phases affected the regulation of the transcription level in the normal and mutant phenotypes differently. LIPH mRNA and protein levels were higher in the outer root sheath (ORS) than in the inner root sheath (IRS), with a very weak signal in the IRS of rex rabbits. In vitro transfection shows that the mutant protein has a reduced lipase activity compared to the wild type form. Our results contribute to the characterization of the LIPH mode of action and confirm the crucial role of LIPH in hair production

A Deletion in Exon 9 of the LIPH Gene Is Responsible for the Rex Hair Coat Phenotype in Rabbits (Oryctolagus cuniculus)

The fur of common rabbits is constituted of 3 types of hair differing in length and diameter while that of rex animals is essentially made up of amazingly soft down-hair. Rex short hair coat phenotypes in rabbits were shown to be controlled by three distinct loci. We focused on the “r1” mutation which segregates at a simple autosomal-recessive locus in our rabbit strains. A positional candidate gene approach was used to identify the rex gene and the corresponding mutation. The gene was primo-localized within a 40 cM region on rabbit chromosome 14 by genome scanning families of 187 rabbits in an experimental mating scheme. Then, fine mapping refined the region to 0.5 cM (Z = 78) by genotyping an additional 359 offspring for 94 microsatellites present or newly generated within the first defined interval. Comparative mapping pointed out a candidate gene in this 700 kb region, namely LIPH (Lipase Member H). In humans, several mutations in this major gene cause alopecia, hair loss phenotypes. The rabbit gene structure was established and a deletion of a single nucleotide was found in LIPH exon 9 of rex rabbits (1362delA). This mutation results in a frameshift and introduces a premature stop codon potentially shortening the protein by 19 amino acids. The association between this deletion and the rex phenotype was complete, as determined by its presence in our rabbit families and among a panel of 60 rex and its absence in all 60 non-rex rabbits. This strongly suggests that this deletion, in a homozygous state, is responsible for the rex phenotype in rabbits

A Deletion in Exon 9 of the LIPH Gene Is Responsible for the Rex Hair Coat Phenotype in Rabbits (Oryctolagus cuniculus)

The fur of common rabbits is constituted of 3 types of hair differing in length and diameter while that of rex animals is essentially made up of amazingly soft down-hair. Rex short hair coat phenotypes in rabbits were shown to be controlled by three distinct loci. We focused on the “r1” mutation which segregates at a simple autosomal-recessive locus in our rabbit strains. A positional candidate gene approach was used to identify the rex gene and the corresponding mutation. The gene was primo-localized within a 40 cM region on rabbit chromosome 14 by genome scanning families of 187 rabbits in an experimental mating scheme. Then, fine mapping refined the region to 0.5 cM (Z = 78) by genotyping an additional 359 offspring for 94 microsatellites present or newly generated within the first defined interval. Comparative mapping pointed out a candidate gene in this 700 kb region, namely LIPH (Lipase Member H). In humans, several mutations in this major gene cause alopecia, hair loss phenotypes. The rabbit gene structure was established and a deletion of a single nucleotide was found in LIPH exon 9 of rex rabbits (1362delA). This mutation results in a frameshift and introduces a premature stop codon potentially shortening the protein by 19 amino acids. The association between this deletion and the rex phenotype was complete, as determined by its presence in our rabbit families and among a panel of 60 rex and its absence in all 60 non-rex rabbits. This strongly suggests that this deletion, in a homozygous state, is responsible for the rex phenotype in rabbits

Evaluation d'un vaccin recombinant dérivé d'adénovirus canin chez le mouton (application à la fièvre catarrhale ovine (FCO))

La Fièvre Catarrhale Ovine est due à un virus affectant les ruminants. Les vaccins actuellement disponibles (atténués ou inactivés) ne permettent pas, ou peu, de protection croisée entre les 24 sérotypes identifiés. Notre travail de thèse a consisté à développer chez le mouton des vaccins recombinants dérivés d adénovirus canin de type 2 capables d induire une protection contre un maximum de sérotypes.Bluetongue is a viral disease that affects Ruminants. Present vaccines (which are attenuated or inactivated vaccines) do not achieve complete cross protection against the 24 serotypes of the Bluetongue Virus (BTV). During this thesis different recombinant vaccines derived from the canine type 2 adenovirus have been developed to afford a broad protection in sheep against BTV.CHATENAY M.-PARIS 11-BU Pharma. (920192101) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

Potentiel thérapeutique des cellules souches mésenchymateuses humaines dans les lésions cutanées radioinduites

Les connaissances récentes acquises sur les cellules souches adultes permettent d'envisager des applications de thérapie cellulaire pour la cicatrisation cutanée. Les cellules mésenchymateuses (CSM), issues de la moelle osseuse, sont des cellules candidates pour une telle application car elles sont multipotentes. Après un traitement anticancéreux par radiothérapie, les séquelles cutanées sont fréquentes et posent des problèmes thérapeutiques non résolus. Nous avons donc testé chez la Souris NOD/SCID la capacité de CSM d'origine humaine à participer à la cicatrisation d'une peau irradiée. Dans ce modèle, une lésion cutanée sur une patte arrière est obtenue par une exposition à 30 Gy de rayons γ. Le pic de lésions est situé autour de la troisième semaine et la cicatrisation est complète à treize semaines. Les CSM humaines sont injectées par voie intraveineuse vingt quatre heures après irradiation. L'analyse du score clinique montre une diminution significative de la sévérité des lésions chez les souris greffées. Six semaines après l'exposition, la présence de cellules dérivées des CSM humaines est détectée dans les zones cicatricielles par PCR quantitative et en histologie. Ces premiers résultats montrent que les CSM injectées par voie intraveineuse sont capables de migrer vers une lésion cutanée et de participer directement à sa réparation. Ils ouvrent des perspectives de thérapie cellulaire par les CSM pour la cicatrisation cutanée

Promotion of cancer cell invasiveness and metastasis emergence caused by olfactory receptor stimulation.

Olfactory receptors (ORs) are expressed in the olfactory epithelium, where they detect odorants, but also in other tissues with additional functions. Some ORs are even overexpressed in tumor cells. In this study, we identified ORs expressed in enterochromaffin tumor cells by RT-PCR, showing that single cells can co-express several ORs. Some of the receptors identified were already reported in other tumors, but they are orphan (without known ligand), as it is the case for most of the hundreds of human ORs. Thus, genes coding for human ORs with known ligands were transfected into these cells, expressing functional heterologous ORs. The in vitro stimulation of these cells by the corresponding OR odorant agonists promoted cell invasion of collagen gels. Using LNCaP prostate cancer cells, the stimulation of the PSGR (Prostate Specific G protein-coupled Receptor), an endogenously overexpressed OR, by β-ionone, its odorant agonist, resulted in the same phenotypic change. We also showed the involvement of a PI3 kinase γ dependent signaling pathway in this promotion of tumor cell invasiveness triggered by OR stimulation. Finally, after subcutaneous inoculation of LNCaP cells into NSG immunodeficient mice, the in vivo stimulation of these cells by the PSGR agonist β-ionone significantly enhanced metastasis emergence and spreading

Overexpression of miR-30b in the developing mouse mammary gland causes a lactation defect and delays involution

Background: MicroRNA (miRNA) are negative regulators of gene expression, capable of exerting pronounced influences upon the translation and stability of mRNA. They are potential regulators of normal mammary gland development and of the maintenance of mammary epithelial progenitor cells. This study was undertaken to determine the role of miR-30b on the establishment of a functional mouse mammary gland. miR-30b is a member of the miR-30 family, composed of 6 miRNA that are highly conserved in vertebrates. It has been suggested to play a role in the differentiation of several cell types.[br/] Methodology/Principal Findings: The expression of miR-30b was found to be regulated during mammary gland development. Transgenic mice overexpressing miR-30b in mammary epithelial cells were used to investigate its role. During lactation, mammary histological analysis of the transgenic mice showed a reduction in the size of alveolar lumen, a defect of the lipid droplets and a growth defect of pups fed by transgenic females. Moreover some mammary epithelial differentiated structures persisted during involution, suggesting a delay in the process. The genes whose expression was affected by the overexpression of miR-30b were characterized by microarray analysis.[br/] Conclusion/Significance: Our data suggests that miR-30b is important for the biology of the mammary gland and demonstrates that the deregulation of only one miRNA could affect lactation and involution

Examples of metastases found in the spine or the lungs of immunodeficient mice inoculated with LNCaP cells.

<p>Metastases are indicated by arrows. In the spine, metastases were observed using microcomputed tomography (Maximum Intensity Projection) and confirmed post-mortem by HES staining and immunohistology using anti-PSA (prostate specific antigen) and anti-PSGR antibodies (only one metastasis is presented). In the lungs, metastases were characterized by HES staining and immunohistology using anti-PSA and anti-PSGR antibodies.</p

Targeted search of ORs expressed by subclones of BON cells, using nested RT-PCR with primers specific to previously identified ORs.

<p>(X) indicates identified ORs.</p