Insights into Neural Crest Evolution

Neural crest cells are unique to vertebrates and essential to the development and evolution of the craniofacial skeleton. Using a combination of DiI cell lineage tracing, transcriptomics, and analysis of key transcription factors of the Sox Family, I examined neural crest development in the sea lamprey, Petromyzon marinus, as the most basal extant vertebrate from which it is possible to get embryos. The results have uncovered distinct cranial and trunk neural crest subpopulations along the anterior-posterior axis of the lamprey embryo, with a clear separation between the two. However, no evidence of the presence of an intermediate vagal neural crest population was uncovered. Comparing cranial neural crest genes between lamprey and chick, either by examining individual candidate genes or whole genome transcriptome analysis, reveals significant changes in the cranial neural crest gene regulatory network of lamprey compared with chick. In particular, the lamprey cranial neural crest is "missing" several gnathostome cranial crest genes. We speculate that these may underlie the evolutionary divergence of craniofacial development between jawed and jawless vertebrates. Despite the absence of vagal neural crest, DiI-labeling shows that trunk neural crest-derived cells, likely homologous to mammalian Schwann cell precursors, contribute to the lamprey enteric nervous system, potentially representing the most primitive form of neural crest cells contribution to the ENS. Finally, I characterized key members of the Sox Family (Sox B-F) due to their importance in neural crest specification in other species. In comparative studies of the SoxC genes (Sox4, Sox11, and Sox12) in both lamprey and Xenopus, I found similar expression patterns and a novel key role in early neural crest specification, suggesting a conserved role of the SoxC genes amongst vertebrates. Taken together, this work represents important progress in characterizing the early evolution of the neural crest in vertebrates and its role in the transition from jawless to jawed vertebrates.

Identification of differentially expressed genes in the heart precursor cells of the chick embryo

Genetic evidence has implicated several genes as being critical for heart development. However, the inducers of these genes as well as their targets and pathways they are involved with, remain largely unknown. Previous studies in the avian embryo showed that at HH4 Cerberus (cCer) transcripts are detected in the anterior endomesoderm including the heart precursor cells and later in the left lateral plate mesoderm. We have identified a promoter element of chick cCer able to drive EGFP expression in a population of cells that consistently exit from the anterior primitive streak region, from as early as stage HH3+, and that later will populate the heart. Using this promoter element as a tool allowed us to identify novel genes previously not known to potentially play a role in heart development. In order to identify and study genes expressed and involved in the correct development and differentiation of the vertebrate heart precursor cell (HPC) lineages, a differential screening using Affymetrix GeneChip system technologies was performed. Remarkably, this screening led to the identification of more than 700 transcripts differentially expressed in the heart forming regions (HFR). Bioinformatic tools allowed us to filter the large amount of data generated from this approach and to select a few transcripts for in vivo validation. Whole-mount in situ hybridization and sectioning of selected genes showed heart and vascular expression patterns for these transcripts during early chick development. We have developed an effective strategy to specifically identify genes that are differentially expressed in the HPC lineages. Within this set we have identified several genes that are expressed in the heart, blood and vascular lineages, which are likely to play a role in their development. These genes are potential candidates for future functional studies on early embryonic patterning.

Exogenous paraoxonase-1 during oocyte maturation improves bovine embryo development in vitro.

The aims of this study were (i) to measure the direct effects of exogenous human recombinant PON1 (rPON1) on bovine oocyte maturation at the molecular level (gene expression) and (ii) to measure the carry-over effects of PON1 on pre-implantation embryo development in vitro.

Follicular wave synchronization on in vivo embryo production in Santa Ines sheep.

This study aimed to evaluate two hormonal protocols for synchronization of follicular wave emergence on in vivo embryo production in Santa Ines sheep under tropical conditions. The greater PRCL rate in GT probably contributed to the smaller number of viable embryos. Thus, it is suggested the appliance indicated the GEm protocol for in vivo embryo production in Santa Ines sheep under tropical conditions.

Route of Oxytocin administration and nonsurgical embryo recovery in Santa Inês ewes after induction synchronous estrus.

Oxytocin has been used to promote cervical dilation with the objective to access uterus both in artificial insemination and transcervical embryo recovery in sheep and goats. The objective of this study was to test the effect of two routes of oxytocin administration on nonsurgical embryo recovery efficiency in Santa Inês ewes after induction of synchronous estrus. Results of this study showed that nonsurgical transcervical embryo recovery can be efficiently done in some ewes; a higher number of individuals is needed to conclude that transcervical embryo recovery can be efficiently done in ewes and surgery embryo collections can be avoided in near to 60% of pluriparous Santa Inês ewes; and that the route of oxytocin administration did not affect the parameters evaluated.

Effects of cAMP modulators during in vitro prematuration of bovine oocytes on gap junctional communication and embryo development potential.

2016

The optimum concentration and timing of melatonin supplementation during IVP to improve bovine embryo development.

2015

Blastocoel collapse associated with melatonin in the culture media improves bovine embryo viability after vitrification.

2016

Glutathione and abscisic acid supplementation influences somatic embryo maturation and hormone endogenous levels during somatic embryogenesis in Podocarpus lambertii Klotzsch ex Endl.

Here we propose a protocol for embryogenic cultures induction, proliferation and maturation for the Brazilian conifer Podocarpus lambertii, and investigated the effect of abscisic acid (ABA) and glutathione (GSH) supplementation on the maturation phase. ABA, zeatin (Z) and salicylic acid (SA) endogenous levels were quantified. Number of somatic embryos obtained in ABA-supplemented treatment was signifi- cant higher than in ABA-free treatment, showing the relevance of ABA supplementation during somatic embryos maturation. Histological analysis showed the stereotyped sequence of developmental stages in conifer somatic embryos, reaching the late torpedo-staged embryo. GSH supplementation in maturation culture medium improved the somatic embryos number and morphological features. GSH 0 mM and GSH 0.1 mM treatments correlated with a decreased ABA endogenous level during maturation, while GSH 0.5 mM treatment showed constantlevels. Alltreatments resulted in decreased Z endogenous levels, supporting the concept that cytokinins are important during the initial cell division but not for the later stages of embryo development. The lowest SA levels found in GSH 0.5 mM treatment were coincident with early embryonic development, and this treatment resulted in the highest development of somatic embryos. Thus, a correlation between lower SA levels and improved somatic embryo formation can be hypothesized