A 37 kb region upstream of brachyury comprising a notochord enhancer is essential for notochord and tail development

The node-streak border region comprising notochord progenitor cells (NPCs) at the posterior node and neuro-mesodermal progenitor cells (NMPs) in the adjacent epiblast is the prime organizing center for axial elongation in mouse embryos. The T-box transcription factor brachyury (T) is essential for both formation of the notochord and maintenance of NMPs, and thus is a key regulator of trunk and tail development. The T promoter controlling T expression in NMPs and nascent mesoderm has been characterized in detail; however, control elements for T expression in the notochord have not been identified yet. We have generated a series of deletion alleles by CRISPR/Cas9 genome editing in mESCs, and analyzed their effects in mutant mouse embryos. We identified a 37 kb region upstream of T that is essential for notochord function and tailbud outgrowth. Within that region, we discovered a T-binding enhancer required for notochord cell specification and differentiation. Our data reveal a complex regulatory landscape controlling cell type-specific expression and function of T in NMP/nascent mesoderm and node/notochord, allowing proper trunk and tail development

Hypoxia induces a transcriptional early primitive streak signature in pluripotent cells enhancing spontaneous elongation and lineage representation in gastruloids

The cellular microenvironment, together with intrinsic regulators, shapes stem cell identity and differentiation capacity. Mammalian early embryos are exposed to hypoxia in vivo and appear to benefit from hypoxic culture in vitro. Yet, how hypoxia influences stem cell transcriptional networks and lineage choices remain poorly understood. Here, we investigated the molecular effects of acute and prolonged hypoxia on embryonic and extra-embryonic stem cells as well as the functional impact on differentiation potential. We find a temporal and cell type-specific transcriptional response including an early primitive streak signature in hypoxic embryonic stem cells mediated by HIF1α. Using a 3D gastruloid differentiation model, we show that hypoxia-induced T expression enables symmetry breaking and axial elongation in the absence of exogenous WNT activation. When combined with exogenous WNT activation, hypoxia enhances lineage representation in gastruloids, as demonstrated by highly enriched signatures of gut endoderm, notochord, neuromesodermal progenitors and somites. Our findings directly link the microenvironment to stem cell function and provide a rationale supportive of applying physiological conditions in models of embryo development

2017 Research & Innovation Day Program

A one day showcase of applied research, social innovation, scholarship projects and activities.https://first.fanshawec.ca/cri_cripublications/1004/thumbnail.jp

Modeling mammalian trunk development in a dish

Mammalian post-implantation development comprises the coordination of complex lineage decisions and morphogenetic processes shaping the embryo. Despite technological advances, a comprehensive understanding of the dynamics of these processes and of the self-organization capabilities of stem cells and their descendants remains elusive. Building synthetic embryo-like structures from pluripotent embryonic stem cells in vitro promises to fill these knowledge gaps and thereby may prove transformative for developmental biology. Initial efforts to model the post-implantation embryo resulted in structures with compromised morphology (gastruloids). Recent approaches employing modified culture media, an extracellular matrix surrogate or extra-embryonic stem cells, however, succeeded in establishing embryo-like architecture. For example, embedding of gastruloids in Matrigel unlocked self-organization into trunk-like structures with bilateral somites and a neural tube-like structure, together with gut tissue and primordial germ cell-like cells. In this review, we describe the currently available models, discuss how these can be employed to acquire novel biological insights, and detail the imminent challenges for improving current models by in vitro engineering

Understanding the relationship between the inbreeding coefficient and multilocus heterozygosity: Theoretical expectations and empirical data

Geneticists have been interested in inbreeding and inbreeding depression since the time of Darwin. Two alternative approaches that can be used to measure how inbred an individual is involve the use of pedigree records to estimate inbreeding coefficients or molecular markers to measure multilocus heterozygosity. However, the relationship between inbreeding coefficient and heterozygosity has only rarely been investigated. In this paper, a framework to predict the relationship between the two variables is presented. In addition, microsatellite genotypes at 138 loci spanning all 26 autosomes of the sheep genome were used to investigate the relationship between inbreeding coefficient and multilocus heterozygosity. Multilocus heterozygosity was only weakly correlated with inbreeding coefficient, and heterozygosity was not positively correlated between markers more often than expected by chance. Inbreeding coefficient, but not multilocus heterozygosity, detected evidence of inbreeding depression for morphological traits. The relevance of these findings to the causes of heterozygosity-fitness correlations is discussed and predictions for other wild and captive populations are presented

Follow-up practices for children and adolescents with celiac disease: results of an international survey

Adequate follow-up in celiac disease is important to improve dietary compliance and treat disease-related symptoms and possible complications. However, data on the follow-up of celiac children is scarce. We aimed to assess current pediatric celiac follow-up practices across Europe. Pediatricians and pediatric gastroenterologists from 35 countries in Europe, Israel, Turkey, and Russia completed an anonymous survey which comprised a 52-item questionnaire developed by the ESPGHAN Special Interest Group on Celiac Disease. A total of 911 physicians, the majority of whom exclusively worked in pediatric care (83%) and academic institutions (60%), completed the questionnaire. Mean age and mean experience with celiac care were 48.7 years (+/- 10.6) and 15.7 years (+/- 9.9), respectively. The vast majority (>= 92%) always assessed anthropometry, dietary adherence, and tissue-transglutaminase IgA-antibodies at every visit, with the first visit being between 3 and 6 months after diagnosis. Other parameters (% always tested) were as follows: complete blood count (60%), iron status (48%), liver enzymes (42%), thyroid function (38%), and vitamin D (26%). Quality of life was never assessed by 35% of the responding physicians. Transition to adult care was mostly completed via a written transition report (37%) or no formal transition at all (27%).Conclusions: Follow-up of celiac children and adolescents in Europe may be improved, especially regarding a more rational use of (laboratory) tests, dietary and QoL assessment, and transition to adult care. Evidence-based advice from international scientific societies is needed.Transplantation and immunomodulatio

Generation of Mouse Pluripotent Stem Cell-derived Trunk-like Structures: An in vitro Model of Post-implantation Embryogenesis.

Post-implantation mammalian embryogenesis involves profound molecular, cellular, and morphogenetic changes. The study of these highly dynamic processes is complicated by the limited accessibility of in utero development. In recent years, several complementary in vitro systems comprising self-organized assemblies of mouse embryonic stem cells, such as gastruloids, have been reported. We recently demonstrated that the morphogenetic potential of gastruloids can be further unlocked by the addition of a low percentage of Matrigel as an extracellular matrix surrogate. This resulted in the formation of highly organized trunk-like structures (TLSs) with a neural tube that is frequently flanked by bilateral somites. Notably, development at the molecular and morphogenetic levels is highly reminiscent of the natural embryo. To facilitate access to this powerful model, here we provide a detailed step-by-step protocol that should allow any lab with access to standard cell culture techniques to implement the culture system. This will provide the user with a means to investigate early mid-gestational mouse embryogenesis at an unprecedented spatiotemporal resolution