The Role and Control of WNT Signalling in an HESC Model of Human Primitive Streak

In amniotes, gastrulation is marked by the creation of the primitive streak (PS) and is largely controlled by WNT, BMP, and ACTIVIN/NODAL signalling. Despite detailed characterization in model organisms, the human PS and the role these pathways play in its formation and patterning remains a mystery. In this work I focused on understanding the role and control of the WNT pathway in human PS development. Due to the ethical limitations of working with human embryos, I used an in vitro human embryonic stem cell (hESC) micropatterned “gastruloid” system. I first showed that in the human PS there is a conserved BMP → WNT→ NODAL signalling initiation hierarchy, and that WNT is necessary and sufficient for PS formation. Next, I found that structured subpopulations of endoderm and mesoderm emerge and self-organize depending on different BMP, WNT, and ACTIVIN/NODAL levels, and that by comparison to the mouse embryo I could arrange these subpopulations along an anterior-posterior axis. With the development of a new cell tracking technique, I was also able to identify and characterize robust cell migrations from the PS region of each gastruloid that depended on which fates the cells would ultimately adopt. Putting these pieces together, I was able to derive a rudimentary first fate map of the human PS, as well as a rough picture of the BMP, WNT, and ACTIVIN/NODAL signalling gradients that determine it. One interesting and unforeseen result from this fate map was the hint of a human “organizer” cell fate that emerged under joint WNT and ACTIVIN/NODAL stimulation. To characterize and functionally prove this organizer’s existence, I devised an ex ovo cross-species transplantation strategy grafting treated gastruloids into chick embryos. The assay demonstrated that the human cells induce and contribute autonomously to a secondary axis while inducing neural fate in the host, thus fulfilling the most stringent criteria for an organizer. This work adds an important milestone to the research program begun in 1924 with the first famous organizer experiment of Hilde Mangold and Hans Spemann, and the methods I developed have opened a door to new functional explorations and tests of early human development. Having learned more about the role of WNT in determining cell fates in the gastruloid model, I next endeavoured to understand how the spatial extent and duration of the WNT signal itself was controlled. With the use of various CRISPR/Cas9 knockout lines, I discovered that DKK1 and E-CADHERIN were the two dominant factors, with E-CADHERIN transducing boundary forces to focus WNT signalling to colony border at early times, and DKK1 controlling the late WNT pattern via cell non-autonomous negative feedback. With the help of time-lapse imaging of a fluorescent reporter line and mathematical modelling, I showed that these two factors mediate a wave of WNT signalling that spreads across the tissue to be patterned, and that this wave is a generic property of a bistable system and thus likely generalizable to other instances in development. While limited by the use of hESCs, taken together my findings provide a first glimpse into the role and control of WNT signalling early on in our own, human development

Prioritizing Populations for Conservation Using Phylogenetic Networks

In the face of inevitable future losses to biodiversity, ranking species by conservation priority seems more than prudent. Setting conservation priorities within species (i.e., at the population level) may be critical as species ranges become fragmented and connectivity declines. However, existing approaches to prioritization (e.g., scoring organisms by their expected genetic contribution) are based on phylogenetic trees, which may be poor representations of differentiation below the species level. In this paper we extend evolutionary isolation indices used in conservation planning from phylogenetic trees to phylogenetic networks. Such networks better represent population differentiation, and our extension allows populations to be ranked in order of their expected contribution to the set. We illustrate the approach using data from two imperiled species: the spotted owl Strix occidentalis in North America and the mountain pygmy-possum Burramys parvus in Australia. Using previously published mitochondrial and microsatellite data, we construct phylogenetic networks and score each population by its relative genetic distinctiveness. In both cases, our phylogenetic networks capture the geographic structure of each species: geographically peripheral populations harbor less-redundant genetic information, increasing their conservation rankings. We note that our approach can be used with all conservation-relevant distances (e.g., those based on whole-genome, ecological, or adaptive variation) and suggest it be added to the assortment of tools available to wildlife managers for allocating effort among threatened populations

Computing evolutionary distinctiveness indices in large scale analysis

We present optimal linear time algorithms for computing the Shapley values and 'heightened evolutionary distinctiveness' (HED) scores for the set of taxa in a phylogenetic tree. We demonstrate the efficiency of these new algorithms by applying them to a set of 10,000 reasonable 5139-species mammal trees. This is the first time these indices have been computed on such a large taxon and we contrast our finding with an ad-hoc index for mammals, fair proportion (FP), used by the Zoological Society of London's EDGE programme. Our empirical results follow expectations. In particular, the Shapley values are very strongly correlated with the FP scores, but provide a higher weight to the few monotremes that comprise the sister to all other mammals. We also find that the HED score, which measures a species' unique contribution to future subsets as function of the probability that close relatives will go extinct, is very sensitive to the estimated probabilities. When they are low, HED scores are less than FP scores, and approach the simple measure of a species' age. Deviations (like the Solendon genus of the West Indies) occur when sister species are both at high risk of extinction and their clade roots deep in the tree. Conversely, when endangered species have higher probabilities of being lost, HED scores can be greater than FP scores and species like the African elephant Loxondonta africana, the two solendons and the thumbless bat Furipterus horrens can move up the rankings. We suggest that conservation attention be applied to such species that carry genetic responsibility for imperiled close relatives. We also briefly discuss extensions of Shapley values and HED scores that are possible with the algorithms presented here

The methodology of on the spot accident investigations in the UK

A new ’On-The-Spot’ (OTS) accident research project is now underway in the UK. This project enables expert investigators to attend the scene of an accident within 15 minutes of the incident occurring, which allows the collection of accident data that would otherwise be quickly lost. This paper considers previous studies and the justification for a new research approach before describing methodology used on the spot and during subsequent follow-up research. Investigations focus on all types of vehicles (including damage, failures, features fitted and their contribution); the highway (including design, features, maintenance and condition); the human factors (including drivers, riders, passengers and pedestrians); and the injuries sustained. Five hundred crashes will be studied in depth each year. The project objectives include establishing an in-depth database that will permit analyses to better understand the causes of crashes and injuries, and assist in the development of solutions

Changing Pattern of Human Listeriosis, England and Wales, 2001–2004

Disease has reemerged, mainly in patients ≥60 years of age with bacteremia

Time for nutrition in medical education.

AIM: To synthesise a selection of UK medical students' and doctors' views surrounding nutrition in medical education and practice. METHODS: Information was gathered from surveys of medical students and doctors identified between 2015 and 2018 and an evaluation of nutrition teaching in a single UK medical school. Comparative analysis of the findings was undertaken to answer three questions: the perceived importance of nutrition in medical education and practice, adequacy of nutrition training, and confidence in current nutrition knowledge and skills. RESULTS: We pooled five heterogeneous sources of information, representing 853 participants. Most agreed on the importance of nutrition in health (>90%) and in a doctor's role in nutritional care (>95%). However, there was less desire for more nutrition education in doctors (85%) and in medical students (68%). Most felt their nutrition training was inadequate, with >70% reporting less than 2 hours. There was a preference for face-to-face rather than online training. At one medical school, nutrition was included in only one module, but this increased to eight modules following an increased nutrition focus. When medical students were asked about confidence in their nutrition knowledge and on advising patients, there was an even split between agree and disagree (p=0.869 and p=0.167, respectively), yet few were confident in the UK dietary guidelines. Only 26% of doctors were confident in their nutrition knowledge and 74% gave nutritional advice less than once a month, citing lack of knowledge (75%), time (64%) and confidence (62%) as the main barriers. There was some recognition of the importance of a collaborative approach, yet 28% of doctors preferred to get specialist advice rather than address nutrition themselves. CONCLUSION: There is a desire and a need for more nutrition within medical education, as well as a need for greater clarity of a doctor's role in nutritional care and when to refer for specialist advice. Despite potential selection bias and limitations in the sampling frame, this synthesis provides a multifaceted snapshot via a large number of insights from different levels of training through medical students to doctors from which further research can be developed

Xenobiotic metabolism: the effect of acute kidney injury on non-renal drug clearance and hepatic drug metabolism.

Acute kidney injury (AKI) is a common complication of critical illness, and evidence is emerging that suggests AKI disrupts the function of other organs. It is a recognized phenomenon that patients with chronic kidney disease (CKD) have reduced hepatic metabolism of drugs, via the cytochrome P450 (CYP) enzyme group, and drug dosing guidelines in AKI are often extrapolated from data obtained from patients with CKD. This approach, however, is flawed because several confounding factors exist in AKI. The data from animal studies investigating the effects of AKI on CYP activity are conflicting, although the results of the majority do suggest that AKI impairs hepatic CYP activity. More recently, human study data have also demonstrated decreased CYP activity associated with AKI, in particular the CYP3A subtypes. Furthermore, preliminary data suggest that patients expressing the functional allele variant CYP3A5*1 may be protected from the deleterious effects of AKI when compared with patients homozygous for the variant CYP3A5*3, which codes for a non-functional protein. In conclusion, there is a need to individualize drug prescribing, particularly for the more sick and vulnerable patients, but this needs to be explored in greater depth

Current and emerging approaches to noncompetitive AR inhibition

The Androgen Receptor (AR) has been shown to be a key determinant in the pathogenesis of castration-resistant prostate cancer (CRPC). Current standard of care therapies target the ligand-binding domain of the receptor, and afford improvements to life expectancy often only in the order of months before resistance occurs. Emerging preclinical and clinical compounds that inhibit receptor activity via differentiated mechanisms of action which are orthogonal to current anti-androgens show promise for overcoming treatment resistance. In this review we present an authoritative summary of molecules that non-competitively target the AR. Emerging small molecule strategies for targeting alternative domains of the AR represent a promising area of research that shows significant potential future therapies. The overall quality of lead candidates in the area of non-competitive AR inhibition is discussed, and identifies the key chemotypes and associated properties which are likely to be, or are currently, positioned for first in human applications