Establishment of a relationship between blastomere geometry and YAP localisation during compaction.

Precise patterning within the three-dimensional context of tissues, organs and embryos implies that cells can sense their relative position. During preimplantation development, outside and inside cells rely on apicobasal polarity and the Hippo pathway to choose their fate. Despite recent findings suggesting that mechanosensing might be central to this process, the relationship between blastomere geometry (i.e. shape and position) and the Hippo pathway effector YAP remains unknown. We used a highly quantitative approach to analyse information on the geometry and YAP localisation of individual blastomeres of mouse and human embryos. We identified the proportion of exposed cell surface area as most closely correlating with the nuclear localisation of YAP. To test this relationship, we developed several hydrogel-based approaches to alter blastomere geometry in cultured embryos. Unbiased clustering analyses of blastomeres from such embryos revealed that this relationship emerged during compaction. Our results therefore pinpoint the time during early embryogenesis when cells acquire the ability to sense changes in geometry and provide a new framework for how cells might integrate signals from different membrane domains to assess their relative position within the embryo.

Advances in Catchment Science, Hydrochemistry, and Aquatic Ecology Enabled by High-Frequency Water Quality Measurements.

High-frequency water quality measurements in streams and rivers have expanded in scope and sophistication during the last two decades. Existing technology allows in situ automated measurements of water quality constituents, including both solutes and particulates, at unprecedented frequencies from seconds to subdaily sampling intervals. This detailed chemical information can be combined with measurements of hydrological and biogeochemical processes, bringing new insights into the sources, transport pathways, and transformation processes of solutes and particulates in complex catchments and along the aquatic continuum. Here, we summarize established and emerging high-frequency water quality technologies, outline key high-frequency hydrochemical data sets, and review scientific advances in key focus areas enabled by the rapid development of high-frequency water quality measurements in streams and rivers. Finally, we discuss future directions and challenges for using high-frequency water quality measurements to bridge scientific and management gaps by promoting a holistic understanding of freshwater systems and catchment status, health, and function.

Distribution and transport of microplastic and fine particulate organic matter in urban streams.

Plastic litter is accumulating in ecosystems worldwide. Rivers are a major source of plastic litter to oceans. However, rivers also retain and transform plastic pollution. While methods for calculating particle transport dynamics in rivers are well established, they are infrequently used to quantify the transport and retention of microplastics (i.e., particles < 5 mm) in flowing waters. Measurements of microplastic movement in rivers are needed for a greater understanding of the fate of plastic litter at watershed and global scales, and to inform pollution prevention strategies. Our objectives were to (1) quantify the abundance of microplastics within different river habitats and (2) adapt organic matter "spiraling" metrics to measure microplastic transport concurrent with fine particulate organic matter (FPOM). We quantified microplastic and FPOM abundance across urban river habitats (i.e., surface water, water column, benthos), and calculated downstream particle velocity, index of retention, turnover rate, and spiraling length for both particle types. Microplastic standing stock was assessed using a habitat-specific approach, and estimates were scaled up to encompass the study reach. Spatial distribution of particles demonstrated that microplastics and FPOM were retained together, likely by hydrodynamic forces that facilitate particle sinking or resuspension. Microplastic particles had a higher downstream particle velocity and lower index of retention relative to FPOM, suggesting that microplastics were retained to a lesser degree than FPOM in the study reaches. Microplastics also showed lower turnover rates and longer spiraling lengths relative to FPOM, attributed to the slow rates of plastic degradation. Thus, rivers are less retentive of microplastics than FPOM, although both particles are retained in similar locations. Because microplastics are resistant to degradation, individual particles can be transported longer distances prior to mineralization than FPOM, making it likely that microplastic particles will encounter larger bodies of water and interact with various aquatic biota in the process. These empirical assessments of particle transport will be valuable for understanding the fate and transformation of microplastic particles in freshwater resources and ultimately contribute to the refinement of global plastic budgets.

Improved specificity of TALE-based genome editing using an expanded RVD repertoire.

Transcription activator-like effector (TALE) proteins have gained broad appeal as a platform for targeted DNA recognition, largely owing to their simple rules for design. These rules relate the base specified by a single TALE repeat to the identity of two key residues (the repeat variable diresidue, or RVD) and enable design for new sequence targets via modular shuffling of these units. A key limitation of these rules is that their simplicity precludes options for improving designs that are insufficiently active or specific. Here we address this limitation by developing an expanded set of RVDs and applying them to improve the performance of previously described TALEs. As an extreme example, total conversion of a TALE nuclease to new RVDs substantially reduced off-target cleavage in cellular studies. By providing new RVDs and design strategies, these studies establish options for developing improved TALEs for broader application across medicine and biotechnology.

ASPP2 maintains the integrity of mechanically stressed pseudostratified epithelia during morphogenesis.

During development, pseudostratified epithelia undergo large scale morphogenetic events associated with increased mechanical stress. Using a variety of genetic and imaging approaches, we uncover that in the mouse E6.5 epiblast, where apical tension is highest, ASPP2 safeguards tissue integrity. It achieves this by preventing the most apical daughter cells from delaminating apically following division events. In this context, ASPP2 maintains the integrity and organisation of the filamentous actin cytoskeleton at apical junctions. ASPP2 is also essential during gastrulation in the primitive streak, in somites and in the head fold region, suggesting that it is required across a wide range of pseudostratified epithelia during morphogenetic events that are accompanied by intense tissue remodelling. Finally, our study also suggests that the interaction between ASPP2 and PP1 is essential to the tumour suppressor function of ASPP2, which may be particularly relevant in the context of tissues that are subject to increased mechanical stress.

Dynamic shapes of the zygote and two-cell mouse and human.

Mouse zygote morphokinetics were measured during interphase, the mitotic period, cytokinesis, and two-cell stage. Sequences of rounder-distorted-rounder shapes were revealed, as were changing patterns of cross section area. A calcium chelator and an actin-disrupting agent inhibited the area changes that occurred between pronuclear envelope breakdown and cytokinesis. During cell division, two vortices developed in each nascent cell and they rotated in opposite directions at each end of the cell, a pattern that sometimes persisted for up to 10 h. Exchange with the environment may have been promoted by these shape and area cycles and persisting circulation in the cytoplasm may have a similar function between a cell's interior and periphery. Some of these movements were sporadically also seen in human zygotes with abnormal numbers of pronuclei and the two-cell stages that developed from these compromised human zygotes.

The Importance of Food Composition Data for Estimating Micronutrient Intake: What Do We Know Now and into the Future?

Food composition tables and databases (FCT/FCDB) centralize data on the energy and nutrient content of foods of a certain country or region. They are essential for many activities related to nutrition. The main factors that can affect the quality of FCT/FCDB are the sources of the data, coverage of foods and components, food description, and component identification. Around 100 countries have published at least one FCT/FCDB, although many of them are outdated and vary considerably in terms of data quality, documentation, and accessibility. A great number of those FCT/FCDB contain very few up-to-date analytical data obtained for food composition purposes, resulting in many data being estimated or copied from publicly available FCT/FCDB from other countries. In addition, many other natural factors that can affect the composition of foods are often not reflected in FCT/FCDB, including biodiversity, maturation degree, soil, and harvest season. Therefore, the use of low-quality FCT/FCDB to convert food consumption data into energy and nutrient intakes may introduce errors resulting in under- or overestimated intake for a certain component. These wrong conclusions may lead to inappropriate or inefficient nutrition and health-related policies, especially to improve micronutrient status in populations and individuals.

Management of deep-seated malformations with photodynamic therapy: a new guiding imaging modality.

Photodynamic therapy is an emerging technology and results from the interactions between a photosensitiser, oxygen and light. The delivery of light may either be by surface illumination or by interstitial application. We describe the first clinical application of ultrasound-guided interstitial photodynamic therapy (US-iPDT). A total of 23 treatments with meta-tetra-hydroxyphenyl chlorine (mTHPC) and ultrasound-guided interstitial photodynamic therapy were performed on 21 patients with various conditions at the Head & Neck Centre, University College London Hospital. The needles could be clearly identified during insertion in all 23 treatments, and it was possible to guide parallel needle insertions using ultrasound. Although the resolution of ultrasound is not as high as that of other imaging modalities [i.e. computed tomography (CT) and magnetic resonance imaging (MRI)] it was satisfactory in identifying the centre and the peripheries of the pathological lesions. Ultrasound is very easy to perform, non-invasive, relatively inexpensive, quick, convenient, non-ionising, suited to the imaging of soft tissues and does not cause any discomfort. Ultrasound can be used to guide 'real-time' photodynamic therapy in deep-seated tumours and other malformations and can augment the information from other imaging modalities without affecting the patient's treatment outcome.

Development of the FAO/INFOODS/IZINCG Global Food Composition Database for Phytate.

Phytate is widely distributed in the plant kingdom, and its significance for human nutrition has been often described. Data on phytate is available in very few composition tables, for a limited number of foods and mainly for raw products. With the aim of publishing the first global repository of analytical data on phytate, data on moisture, phytate, zinc, iron and calcium were compiled. Other aspects, such as the analytical method used, biodiversity and processing, were considered, and phytate: mineral ratios were calculated when possible. From a comprehensive literature search, over 250 references were compiled, generating 3377 entries: 39% for raw and 61% for processed foods. Most of the entries were for cereals (35%), followed by legumes (27%) and vegetables (11%). The most common analytical methods used were indirect precipitation (26%) and anion exchange (25%), while separate determination of IPs is the most recommended. The database can be used as a tool for nutrition workers to include into food composition tables and to develop programmes related to mineral deficiencies. These data will be useful for designing diets with enhanced mineral bioavailability and for improving the estimates for nutrient requirements. The database is available at the INFOODS (www.fao.org/infoods/infoods/tables-and-databases/en) and IZiNCG webpages (www.izincg.org).

Importance and use of reliable food composition data generation by nutrition/dietetic professionals towards solving Africa's nutrition problem: constraints and the role of FAO/INFOODS/AFROFOODS and other stakeholders in future initiatives.

Despite the rich biodiversity of the African continent and the tremendous progress so far made in food production, Africa is still struggling with the problems of food insecurity, hunger and malnutrition. To combat these problems, the production and consumption of nutritious and safe foods need to be promoted. This cannot be achieved without reliable data on the quantity and quality of nutrients and other components provided through these foods. Food composition data (FCD) are compiled as food composition tables (FCT) or food composition databases (FCDB). These are subsequently used for a variety of purposes, ranging from clinical practice, research, public health/education, food industry to planning and policy, as well as nutrition monitoring and surveillance. To perform these functions effectively, the importance of reliable FCT/FCDB cannot be overemphasised. Poor quality FCT/FCDB have serious consequences on the health of the population, and provide skew evidence towards developing nutrition and health-related policies. The present paper reviews different methods to generate FCT/FCDB, their importance and use in assisting nutrition/dietetic professionals in solving Africa's nutrition problems; current status of FCT/FCDB generation, compilation and dissemination in Africa, constraint to their use by professionals and the role of FAO/INFOODS/AFROFOODS and other stakeholders towards improvement and future initiatives. The information provided will create awareness on the need for up-to-date and high-quality FCT/FCDB and facilitate the identification of data gaps and prioritisation of future efforts in FCD generation, compilation and dissemination in Africa and subsequent strategies for the alleviation of the food and nutrition problems in Africa.

Citizen science datasets reveal drivers of spatial and temporal variation for anthropogenic litter on Great Lakes beaches.

Accumulation of anthropogenic litter (AL) on marine beaches and its ecological effects have been a major focus of research. Recent studies suggest AL is also abundant in freshwater environments, but much less research has been conducted in freshwaters relative to oceans. The Adopt-a-BeachTM (AAB) program, administered by the Alliance for the Great Lakes, organizes volunteers to act as citizen scientists by collecting and maintaining data on AL abundance on Great Lakes beaches. Initial assessments of the AAB records quantified sources and abundance of AL on Lake Michigan beaches, and showed that plastic AL was >75% of AL on beaches across all five Great Lakes. However, AAB records have not yet been used to examine patterns of AL density and composition among beaches of all different substrate types (e.g., parks, rocky, sandy), across land-use categories (e.g., rural, suburban, urban), or among seasons (i.e., spring, summer, and fall). We found that most AL on beaches are consumer goods that most likely originate from beach visitors and nearby urban environments, rather than activities such as shipping, fishing, or illegal dumping. We also demonstrated that urban beaches and those with sand rather than rocks had higher AL density relative to other sites. Finally, we found that AL abundance is lowest during the summer, between the US holidays of Memorial Day (last Monday in May) and Labor Day (first Monday in September) at the urban beaches, while other beaches showed no seasonality. This research is a model for utilizing datasets collected by volunteers involved in citizen science programs, and will contribute to AL management by offering priorities for AL types and locations to maximize AL reduction.

Mitochondrially targeted ZFNs for selective degradation of pathogenic mitochondrial genomes bearing large-scale deletions or point mutations.

We designed and engineered mitochondrially targeted obligate heterodimeric zinc finger nucleases (mtZFNs) for site-specific elimination of pathogenic human mitochondrial DNA (mtDNA). We used mtZFNs to target and cleave mtDNA harbouring the m.8993T>G point mutation associated with neuropathy, ataxia, retinitis pigmentosa (NARP) and the "common deletion" (CD), a 4977-bp repeat-flanked deletion associated with adult-onset chronic progressive external ophthalmoplegia and, less frequently, Kearns-Sayre and Pearson's marrow pancreas syndromes. Expression of mtZFNs led to a reduction in mutant mtDNA haplotype load, and subsequent repopulation of wild-type mtDNA restored mitochondrial respiratory function in a CD cybrid cell model. This study constitutes proof-of-principle that, through heteroplasmy manipulation, delivery of site-specific nuclease activity to mitochondria can alleviate a severe biochemical phenotype in primary mitochondrial disease arising from deleted mtDNA species.

Treatment of Kimura disease with photodynamic therapy: a case study.

We report on the application of photodynamic therapy (PDT) in the management of Kimura disease. A 58-year-old Asian male was offered this modality to assess the possibility to control disease progression. The patient was managed with surgery and the disease recurred and caused facial disfigurement. PDT was offered after careful discussion at UCLH multidiscipline meeting. The photosensitiser "mTHPC" was introduced intravenously 96h prior to delivering the light under ultrasound guidance. Magnetic resonance images showed moderate-significant reduction of the disease volume. Fourteen months post-PDT, the disease started re-growing and the patient subsequently underwent one further round of PDT which was as successful as the first round in reducing the facial disfigurement. Photodynamic therapy was very effective in controlling disease progression in this patient who suffers from Kimura disease.

Efficient immunoglobulin gene disruption and targeted replacement in rabbit using zinc finger nucleases.

Rabbits are widely used in biomedical research, yet techniques for their precise genetic modification are lacking. We demonstrate that zinc finger nucleases (ZFNs) introduced into fertilized oocytes can inactivate a chosen gene by mutagenesis and also mediate precise homologous recombination with a DNA gene-targeting vector to achieve the first gene knockout and targeted sequence replacement in rabbits. Two ZFN pairs were designed that target the rabbit immunoglobulin M (IgM) locus within exons 1 and 2. ZFN mRNAs were microinjected into pronuclear stage fertilized oocytes. Founder animals carrying distinct mutated IgM alleles were identified and bred to produce offspring. Functional knockout of the immunoglobulin heavy chain locus was confirmed by serum IgM and IgG deficiency and lack of IgM(+) and IgG(+) B lymphocytes. We then tested whether ZFN expression would enable efficient targeted sequence replacement in rabbit oocytes. ZFN mRNA was co-injected with a linear DNA vector designed to replace exon 1 of the IgM locus with ∼1.9 kb of novel sequence. Double strand break induced targeted replacement occurred in up to 17% of embryos and in 18% of fetuses analyzed. Two major goals have been achieved. First, inactivation of the endogenous IgM locus, which is an essential step for the production of therapeutic human polyclonal antibodies in the rabbit. Second, establishing efficient targeted gene manipulation and homologous recombination in a refractory animal species. ZFN mediated genetic engineering in the rabbit and other mammals opens new avenues of experimentation in immunology and many other research fields.

TMJ arthroscopy in patients with Ehlers Danlos syndrome: case series.

OBJECTIVE: The purpose of this report was to assess the outcome of temporomandibular joint (TMJ) arthroscopy in patients with temporomandibular disorders (TMDs) associated with Ehlers Danlos syndrome (EDS). STUDY DESIGN: This retrospective case series describes 18 patients with EDS who underwent arthroscopy for temporomandibular disorders. The patients' demographics were recorded, along with preoperative TMJ symptoms, Wilkes classification, mouth opening, and the presence of systemic involvement. The incidence of early and late postoperative complications and the final outcome were noted. RESULTS: All of the patients were females, with EDS Type III, and had a mean age of 34 years. A high proportion of the patients had joints other than the TMJ affected. Five patients were classified as stage II according to the Wilkes classification, 9 patients were stage III, 3 patients were stage IV, and only 1 patient was diagnosed with stage V Wilkes before intervention. Arthroscopy, followed by arthrocentesis and balloon dilatation of the affected TMJs was performed and intra-articular morphine injections were given to all patients. The main pre- and early postoperative complaint was pain, but this resolved in most cases. Improvement of mouth opening was noted from 23.4 +/- 4.2 to 27.8 +/- 5.1 mm after arthroscopy. Patients were followed for an average of 62 months and all were asymptomatic at their last review appointment. CONCLUSION: For patients where conservative measures of treating TMD are not effective, arthroscopy is a minimally invasive surgical procedure that has been shown to result in a satisfactory outcome, with no need to resort to open joint surgery. This case series is limited by its size and further research on surgical intervention on EDS patients with temporomandibular disorders is recommended.

Knockout rats via embryo microinjection of zinc-finger nucleases.

The toolbox of rat genetics currently lacks the ability to introduce site-directed, heritable mutations into the genome to create knockout animals. By using engineered zinc-finger nucleases (ZFNs) designed to target an integrated reporter and two endogenous rat genes, Immunoglobulin M (IgM) and Rab38, we demonstrate that a single injection of DNA or messenger RNA encoding ZFNs into the one-cell rat embryo leads to a high frequency of animals carrying 25 to 100% disruption at the target locus. These mutations are faithfully and efficiently transmitted through the germline. Our data demonstrate the feasibility of targeted gene disruption in multiple rat strains within 4 months time, paving the way to a humanized monoclonal antibody platform and additional human disease models.