Human Naïve Epiblast Cells Possess Unrestricted Lineage Potential

SUMMARYClassical mouse embryology has established a paradigm of early development driven by sequential lineage bifurcations. Accordingly, mouse embryonic stem cells derived from early epiblast have lost the potency to produce extraembryonic trophectoderm. We show in contrast that human naïve epiblast cells readily make trophectoderm. Inhibition of ERK signalling, instrumental in naïve stem cell propagation, unexpectedly potentiates trophectoderm formation, an effect enhanced by Nodal inhibition. Transcriptome analyses authenticate conversion into trophectoderm with subsequent production of syncitiotrophoblast, cytotrophoblast and trophoblast stem cells. Genetic perturbations indicate that NANOG suppresses and TFAP2C enables trophectoderm induction. Consistent with post-implantation progression, trophectoderm potential is extinguished in conventional human pluripotent stem cells, which instead make amnion. Finally, human embryo epiblasts from late blastocysts efficiently generate trophectoderm and differentiated trophoblast. Thus, pluripotent cells in the human embryo retain extraembryonic lineage plasticity and regenerative potential until implantation. Harnessing this unanticipated regulative capacity may be beneficial for assisted reproduction technology.

Multiplying embryos: experimental monozygotic polyembryony in mammals and its uses

Monozygotic (MZ) polyembryony is a strategy to increase the output of a single zygote, thereby producing more offspring from a limited number of oocytes. However, MZ twins and multiples (multiplets) of mammals occur rarely in nature, while their generation has been more successful experimentally. In this work, we review some of the methodological, biological and field aspects of experimental MZ polyembryony in mammals. First attempts of mechanical bisection of 2-cell stage rodent embryos provided a proof-of-principle for the survival and independent development of both blastomeres. Subsequently, experiments in other species, particularly sheep and bovine, allowed 2 methods of embryo multiplication to become routine: the separation or biopsy of blastomeres from cleavage-stage embryos and the bisection of morulae and blastocysts. We discuss how the preferable stage of bisection and the success rate can be species-specific. The scope that profited most from experimental MZ polyembryony is the production of additional copies of elite livestock individuals, the reduction of interindividual variation in test groups and the possibility of investigating discordant phenotypic traits in the same genomic background, for instance, comparing an affected twin with its healthy co-twin. By contrast, the original motivation for experimental polyembryony – efficiently generating more offspring out of the same zygote – has not been fulfilled yet. Although embryo splitting leads to an increase in quantity, there is a loss of embryo quality, thus, there is no real gain from artificially generated embryos (yet) in the field of medically assisted reproduction. In conclusion, mammalian zygotes have the regulative capacity to be polyembryonic, but this is not obligate.

Laser Cyclophotocoagulation Enhances the Regulative Capacity of Retinal Vessels in Glaucoma

Purpose: To determine the effects of laser surgical IOP reduction by means of transscleral cyclophotocoagulation (CPC) on retinal blood flow parameters in glaucoma patients using Dynamic Vessel Analysis (DVA). Materials and Methodology: 26 patients (average age: 70 years) with a long history of primary open angle glaucoma underwent CPC. The effect on the reactive capacity of retinal vessels was assessed before and 6-8 weeks after CPC by means of the Dynamic Vessel Analyzer (DVA) using flicker light provocation. Results: In our group of POAG patients, IOP was significantly reduced about approximately 20% by CPC while systemic blood pressure and heart rate were not changed. The most obvious differences between the pre- and postoperative DVA measurements could be observed in the maximal dilation of the retinal arteries which increased from 0.75 % (+/- 0.6) to 3.17 % (+/- 0.5) with an average increase of 2.4 % (p<0.01). In addition, the ability of the arteries for constriction improved significantly (p<0.05) while the dynamic responses of the veins and the initial baseline values (MU) of the vessel diameters did not change. Conclusions: Our results of DVA measurements after an IOP-lowering laser surgical intervention (CPC) reveal a significant recovery of the regulative capacity of retinal arteries in glaucoma patients that has up to now neither been properly documented nor appreciated. Future studies with long-term follow-up must determine the clinical importance of these findings for the treatment of glaucoma patients.

Hedgehog is required for activation of engrailed during regeneration of fragmented Drosophila imaginal discs

Surgically fragmented Drosophila appendage primordia (imaginal discs) engage in wound healing and pattern regulation during short periods of in vivo culture. Prothoracic leg disc fragments possess exceptional regulative capacity, highlighted by the ability of anterior cells to convert to posterior identity and establish a novel posterior compartment. This anterior/posterior conversion violates developmental lineage restrictions essential for normal growth and patterning of the disc, and thus provides an ideal model for understanding how cells change fate during epimorphic pattern regulation. Here we present evidence that the secreted signal encoded by hedgehog directs anterior/posterior conversion by activating the posterior-specific transcription factor engrailed in regulating anterior cells. In the absence of hedgehog activity, prothoracic leg disc fragments fail to undergo anterior/posterior conversion, but can still regenerate missing anterior pattern elements. We suggest that hedgehog-independent regeneration within the anterior compartment (termed integration) is mediated by the positional cues encoded by wingless and decapentaplegic. Taken together, our results provide a novel mechanistic interpretation of imaginal disc pattern regulation and permit speculation that similar mechanisms could govern appendage regeneration in other organisms.

Regulative capacity of the archenteron during gastrulation in the sea urchin

Gastrulation in the sea urchin involves an extensive rearrangement of cells of the archenteron giving rise to secondary mesenchyme at the archenteron tip followed by the foregut, midgut and hindgut. To examine the regulative capacity of this structure, pieces of the archenteron were removed or transplanted at different stages of gastrulation. After removal of any or all parts of the archenteron, the remaining veg 1 and /or veg 2 tissue regulated to replace the missing parts. Endoderm transplanted to ectopic positions also regulated to that new position in the archenteron. This ability to replace or regulate endoderm did not decline until after full elongation of the archenteron was completed. When replacement occurred, the new gut was smaller relative to the remaining embryo but the recognizable morphology of the archenteron was re-established. Long after the archenteron reveals territorial specification through expression of specific markers, the endodermal cells remain capable of being respecified to other gut regions. Thus, for much of gastrulation, the gut is conditionally specified. We propose that this regulative ability requires extensive and continuous short-range communication between cells of the archenteron in order to reorganize the tissues and position the boundaries of this structure even after experimental alterations.