Vascular malformations are non-neoplastic expansions of blood vessels that arise due to errors during angiogenesis. They are a heterogeneous group of sporadic or inherited vascular disorders characterized by localized lesions of arteriovenous, capillary, or lymphatic origin. Vascular malformations that occur inside bone tissue are rare. Herein, we report loss-of-function mutations in ELMO2 (which translates extracellular signals into cellular movements) that are causative for autosomal-recessive intraosseous vascular malformation (VMOS) in five different families. Individuals with VMOS suffer from life-threatening progressive expansion of the jaw, craniofacial, and other intramembranous bones caused by malformed blood vessels that lack a mature vascular smooth muscle layer. Analysis of primary fibroblasts from an affected individual showed that absence of ELMO2 correlated with a significant downregulation of binding partner DOCK1, resulting in deficient RAC1-dependent cell migration. Unexpectedly, elmo2-knockout zebrafish appeared phenotypically normal, suggesting that there might be human-specific ELMO2 requirements in bone vasculature homeostasis or genetic compensation by related genes. Comparative phylogenetic analysis indicated that elmo2 originated upon the appearance of intramembranous bones and the jaw in ancestral vertebrates, implying that elmo2 might have been involved in the evolution of these novel traits. The present findings highlight the necessity of ELMO2 for maintaining vascular integrity, specifically in intramembranous bones.

Germ-line chimaerism is a powerful technique that has proved to be useful to produce viable gametes when transplanted blastomeres colonize the germinal ridges in recipient embryos and obtaining offspring from such transplanted cells. In fish, ionizing radiations were commonly used for embryo penalization to cancelling the cell participation of recipient embryos in development and in gamete production. The ultraviolet (UV) radiation when compared with other radiation types is cheaper, easier and no special installations are required for its use. So, the aim of this work was to establish the optimal UV radiation dose to be applied in zebrafish embryos at mid-blastula transition stage of development, in order to use them as penalized recipient embryos in futures chimaerism assays. A UV germicide lamp was used as radiation source (0.529 mW/cm(2)). Four exposure levels and three exposure times of UV radiation were tested. The survival rates obtained with the non-dechorionated embryos without lid group suggested that it could be the optimal exposure level to achieve the objective proposed. With the obtained results, we concluded that this UV radiation dose for 60 and 30 s are optimal parameters to penalize recipient wild and gold strain zebrafish embryos, respectively in chimaerism assays, but without involving their survival and apparently normal development.

Germline chimaerism (intra or interspecific) is a technique of great potential in aquaculture. It allows specimens to be obtained that produce gametes whose origins lie in the cells of the donor organism. Chimaerism is usually performed at the mid blastula transition (MBT) stage since this is the last in which embryonic cells remain completely totipotent. Zebrafish are photoperiodic in their egg-laying behaviour and show rapid embryonic development. For chimaerism to be successful, it is of interest to establish the maximum time over which embryonic development can be reversibly arrested. This paper reports the effect on survival of subjecting zebrafish embryos at different stages of development to a water temperature of 16 deg C for different lengths of time. The maximum exposure time after which these embryos were able to resume development following low-temperature-induced developmental arrest became shorter as the embryonic stage exposed became earlier. At the MBT stage, the maximum safe exposure time was 2 h; longer exposure times led to problems in development and survival.

The blastema is a regenerative tissue with remarkable pluripotency. The aim of this work done on zebrafish (Danio rerio) was to define technical procedures required for obtaining and integrating blastema cells into embryos at the mid blastula transition stage (MBT) and the effect on survival, as well as the capacity to produce pigmented chimaeras. Wild type blastema cells were injected into gold type MBT embryos (E). Wild MBT blastomere cells were also injected into gold type MBT embryos as a control (C 1 ). A second control group, C 2 , was not subjected to any manipulation. Survival was evaluated at 24, 48 and 72 h after performing the chimaerism, and the rate of adult chimaeras evaluated. The results showed significant differences in embryo survival between the E and C 1 groups in embryo survival at 24 and 48 h postchimaerism (24 h: E-83.49% vs C 1 -54.8%, p < 0.05; 48 h: E-98.83% vs C 1 -85.13%, p < 0.05). There was no significant difference, at any time, between E and C 2 . The results at 72 h for E and the controls (E-89.41%; C 1 -84.12% and C 2 -92.55%) indicate that insertion of blastema cells does not have a negative effect on embryo development. The results in adults (E: 0 chimaeras from 7 specimens; C 1 : 5 chimaeras from 17 specimens) suggest that the dedifferentiation grade of the blastema cells may not be enough to generate germ-line chimaeras, but their condition of potentially dedifferentiating cells may be an advantage when using them as donor nuclei in somatic cloning by nuclear transplant.Additional key words: biodiversity, chimaerism, embryo, zebrafish. Resumen Evaluación de las células de la blastema de aleta caudal como donantes en quimeras intraespecíficas de pez cebraEl blastema es un tejido regenerativo con una pluripotencia remarcable. El objetivo del presente trabajo realizado en pez cebra (Danio rerio) es definir los procedimientos técnicos requeridos para la obtención e integración de células de la blastema en embriones en estadio MBT y su efecto en la supervivencia, así como también la capacidad de dar lugar a quimeras pigmentadas. Se inyectaron células de blastema de la estirpe silvestre en embriones gold en estadio MBT (E) y como control se inyectaron blastómeras silvestres MBT en embriones gold MBT (C 1 ). El segundo grupo control (C 2 ), no fue sometido a manipulación. Se evaluó la tasa de supervivencia a 24, 48 y 72 h post-quimerismo y la tasa de quimeras adultas. Los resultados mostraron diferencias significativas en la supervivencia embrionaria a las 24 y 48 h entre el grupo E y el C 1 (24 h: E-83,49% vs C 1 -54,8%, p<0,05; 48 h: E-98,83% vs C 1 -85,13%, p<0,05). No se detectaron diferencias significativas entre los grupos E y C 2 en ningún momento. Los resultados obtenidos a las 72 h, tanto en E como en los controles (E-89,41%; C 1 -84,12% y C 2 -92,55%) sugieren que la inserción de células de blastema no causa efectos negativos en el desarrollo del embrión. Los resultados obtenidos en adultos (E: 0 quimeras de 7 especímenes; C 1 : 5 quimeras de 17 especí-menes) apun...

No abstract