RESUMEN
Ellis-van Creveld (EvC) syndrome is a skeletal dysplasia, characterized by short limbs, postaxial polydactyly, and dental abnormalities. EvC syndrome is also categorized as a ciliopathy because of ciliary localization of proteins encoded by the two causative genes, EVC and EVC2 (aka LIMBIN). While recent studies demonstrated important roles for EVC/EVC2 in Hedgehog signaling, there is still little known about the pathophysiological mechanisms underlying the skeletal dysplasia features of EvC patients, and in particular why limb development is affected, but not other aspects of organogenesis that also require Hedgehog signaling. In this report, we comprehensively analyze limb skeletogenesis in Evc2 mutant mice and in cell and tissue cultures derived from these mice. Both in vivo and in vitro data demonstrate elevated Fibroblast Growth Factor (FGF) signaling in Evc2 mutant growth plates, in addition to compromised but not abrogated Hedgehog-PTHrP feedback loop. Elevation of FGF signaling, mainly due to increased Fgf18 expression upon inactivation of Evc2 in the perichondrium, critically contributes to the pathogenesis of limb dwarfism. The limb dwarfism phenotype is partially rescued by inactivation of one allele of Fgf18 in the Evc2 mutant mice. Taken together, our data uncover a novel pathogenic mechanism to understand limb dwarfism in patients with Ellis-van Creveld syndrome.
Asunto(s)
Enanismo/genética , Síndrome de Ellis-Van Creveld/genética , Factores de Crecimiento de Fibroblastos/genética , Proteínas de la Membrana/genética , Animales , Modelos Animales de Enfermedad , Enanismo/patología , Síndrome de Ellis-Van Creveld/patología , Factores de Crecimiento de Fibroblastos/biosíntesis , Placa de Crecimiento/crecimiento & desarrollo , Placa de Crecimiento/patología , Humanos , Péptidos y Proteínas de Señalización Intercelular , Proteínas de la Membrana/biosíntesis , Ratones , Proteínas Mutantes/biosíntesis , Proteínas Mutantes/genética , Polidactilia/genética , Polidactilia/patología , Transducción de Señal , Anomalías Dentarias/genética , Anomalías Dentarias/patologíaRESUMEN
Recombinant mutant human granulocyte colony stimulating factor (rmhG-CSF) was pegylated, purified and characterized. rhG-CSF was mutated in position 1,3,4,5,17, and cysteine was added in C-terminal. rmhG-CSF was pegylated by PEG-Mal 20000 and separated by ion-exchange chromatography, gel filtration chromatography. Analysis of SDS-PAGE showed thar the purity of the separated PEG-rmhG-CSF was greater than 95%. and in intro and in vivo bioactivity study showed that target modified PEG-rmhG-CSF kept full bioactivity which was better than traditional pegylation method, and longer half-life was proved in mice.