TMEM8C-mediated fusion is regionalized and regulated by NOTCH signalling during foetal myogenesis.

The location and regulation of fusion events within skeletal muscles during development remain unknown. Using the fusion marker myomaker (Mymk), named TMEM8C in chicken, as a readout of fusion, we identified a co-segregation of TMEM8C-positive cells and MYOG-positive cells in single-cell RNA-sequencing datasets of limbs from chicken embryos. We found that TMEM8C transcripts, MYOG transcripts and the fusion-competent MYOG-positive cells were preferentially regionalized in central regions of foetal muscles. We also identified a similar regionalization for the gene encoding the NOTCH ligand JAG2 along with an absence of NOTCH activity in TMEM8C+ fusion-competent myocytes. NOTCH function in myoblast fusion had not been addressed so far. We analysed the consequences of NOTCH inhibition for TMEM8C expression and myoblast fusion during foetal myogenesis in chicken embryos. NOTCH inhibition increased myoblast fusion and TMEM8C expression and released the transcriptional repressor HEYL from the TMEM8C regulatory regions. These results identify a regionalization of TMEM8C-dependent fusion and a molecular mechanism underlying the fusion-inhibiting effect of NOTCH in foetal myogenesis. The modulation of NOTCH activity in the fusion zone could regulate the flux of fusion events.

Including microbiome information in a multi-trait genomic evaluation: a case study on longitudinal growth performance in beef cattle.

BACKGROUND: Growth rate is an important component of feed conversion efficiency in cattle and varies across the different stages of the finishing period. The metabolic effect of the rumen microbiome is essential for cattle growth, and investigating the genomic and microbial factors that underlie this temporal variation can help maximize feed conversion efficiency at each growth stage. RESULTS: By analysing longitudinal body weights during the finishing period and genomic and metagenomic data from 359 beef cattle, our study demonstrates that the influence of the host genome on the functional rumen microbiome contributes to the temporal variation in average daily gain (ADG) in different months (ADG1, ADG2, ADG3, ADG4). Five hundred and thirty-three additive log-ratio transformed microbial genes (alr-MG) had non-zero genomic correlations (rg) with at least one ADG-trait (ranging from |0.21| to |0.42|). Only a few alr-MG correlated with more than one ADG-trait, which suggests that a differential host-microbiome determinism underlies ADG at different stages. These alr-MG were involved in ribosomal biosynthesis, energy processes, sulphur and aminoacid metabolism and transport, or lipopolysaccharide signalling, among others. We selected two alternative subsets of 32 alr-MG that had a non-uniform or a uniform rg sign with all the ADG-traits, regardless of the rg magnitude, and used them to develop a microbiome-driven breeding strategy based on alr-MG only, or combined with ADG-traits, which was aimed at shaping the rumen microbiome towards increased ADG at all finishing stages. Combining alr-MG information with ADG records increased prediction accuracy of genomic estimated breeding values (GEBV) by 11 to 22% relative to the direct breeding strategy (using ADG-traits only), whereas using microbiome information, only, achieved lower accuracies (from 7 to 41%). Predicted selection responses varied consistently with accuracies. Restricting alr-MG based on their rg sign (uniform subset) did not yield a gain in the predicted response compared to the non-uniform subset, which is explained by the absence of alr-MG showing non-zero rg at least with more than one of the ADG-traits. CONCLUSIONS: Our work sheds light on the role of the microbial metabolism in the growth trajectory of beef cattle at the genomic level and provides insights into the potential benefits of using microbiome information in future genomic breeding programs to accurately estimate GEBV and increase ADG at each finishing stage in beef cattle.

Skin carotenoid levels in lactating dairy cows as measured using multiple spatially resolved reflection spectroscopy.

This research communication describes a pilot study measuring skin carotenoid levels of lactating dairy cows. Carotenoids are natural antioxidants, involved in cell communication and immune function, protecting against oxidative stress. They are precursors of vitamin A, important for reproduction efficiency, growth and male fertility. Therefore, easy-to-use, inexpensive methods to measure carotenoids in cattle would provide interesting data for farmers to monitor the health and nutritional status of their herds. In this study, we used a commercially available sensor based on multiple spatially resolved reflection spectroscopy (MSRRS), intended for human use, to measure the carotenoid content in bovine skin in three research herds in France, Ireland and Scotland. Carotenoid levels were measured by applying the sensor to the teat barrel, avoiding pigmented areas of skin. Mean sensor values differed significantly between herds and between diets, with pasture-based animals showing significantly higher carotenoid levels. Our results suggest that MSRRS can be used to accurately to measure skin carotenoids in cows. However, further calibration in bovines is needed to improve of the accuracy of the MSRRS sensor in cattle.

Production, characterization, and dietary supplementation effect of rumen-protected fat on ruminal function and blood parameters of sheep.

Rumen-protected fat (RPF) was produced in the 1st experimental stage through melt-emulsification technique using buriti oil (BO) as core, at concentrations of 10% (BO10), 20% (BO20), and 30% (BO30) (w/w), and carnauba wax (CW) as encapsulant material. After obtention and characterization, protected fat microspheres were tested in a 2nd experimental stage on the sheep' diet using six castrated 2-year-old male Santa Ines with initial weight 48.9 ± 5.23 kg, fistulated in rumen and distributed in a double Latin square design with 3 treatments × 3 periods, to evaluate rumen pH, temperature, protozoal count, and blood parameters. There was no difference (P > 0.05) among RPF microspheres for microencapsulation yield. However, microencapsulation efficiency increased (P < 0.05) with BO addition ranging from 36 to 61.3% for BO10 and BO30, respectively. The inclusion of BO10 in the sheep's diet did not affect the ruminal dry matter degradability (DMD) of BO over time (P > 0.05); however, BO20 and BO30 had higher (P < 0.05) DMD values than BO10. No significant differences were observed among RPF for rumen pH and temperature (P > 0.05). There was an increase (P < 0.05) in the protozoal population in the rumen environment due to the microencapsulated BO30 inclusion. There was also increase (P < 0.05) in serum albumin, cholesterol, aspartate aminotransferase (ALT), and gamma-glutamyltransferase (GGT), and a reduction (P < 0.05) in serum triglycerides of the sheep when RPF microspheres increased in the diet. Melt-emulsification proved to be a good technique for microencapsulation of buriti oil into the carnauba wax matrix. RPF from buriti oil protected into carnauba wax is recommended for sheep diet because it increases energy density, without adverse effects on the protozoal populations and blood serum metabolites from the bypass effect in the rumen.

Trackosome: a computational toolbox to study the spatiotemporal dynamics of centrosomes, nuclear envelope and cellular membrane.

During the initial stages of mitosis, multiple mechanisms drive centrosome separation and positioning. How they are coordinated to promote centrosome migration to opposite sides of the nucleus remains unclear. Here, we present Trackosome, an open-source image analysis software for tracking centrosomes and reconstructing nuclear and cellular membranes, based on volumetric live-imaging data. The toolbox runs in MATLAB and provides a graphical user interface for easy access to the tracking and analysis algorithms. It provides detailed quantification of the spatiotemporal relationships between centrosomes, nuclear envelope and cellular membrane, and can also be used to measure the dynamic fluctuations of the nuclear envelope. These fluctuations are important because they are related to the mechanical forces exerted on the nucleus by its adjacent cytoskeletal structures. Unlike previous algorithms based on circular or elliptical approximations, Trackosome measures membrane movement in a model-free condition, making it viable for irregularly shaped nuclei. Using Trackosome, we demonstrate significant correlations between the movements of the centrosomes, and identify specific oscillation modes of the nuclear envelope. Overall, Trackosome is a powerful tool that can be used to help unravel new elements in the spatiotemporal dynamics of subcellular structures.

The European Food Regulatory Environment Index: a tool to monitor progress in implementing food environment policies.

BACKGROUND: Evidence based health policy, such as that put forward in the European Food and Nutrition Action Plan 2015-2020 and the WHO Global Action Plan on the Prevention and Control of Noncommunicable Diseases, has a role in curbing the consumption of unhealthful foods and drink. We ask how countries are performing in the adoption of these policies and how the comprehensiveness of their food environment policies explains variations in consumption of unhealthful products across Europe. METHODS: In order to assess the state of policy adoption, we developed a composite indicator-the Food Regulatory Environment Index (FREI) for which we calculated unweighted and weighted formulations according to the strength of the evidence base. We used linear regression models to explain variations in the consumption of unhealthful products as well as variations in a composite indicator of obesogenic diets. RESULTS: Overall, wealthier countries in the Region perform better. The weighting of the constituent policies does not affect the rankings. We find negative associations between unweighted and weighted formulations of the Index and household consumption of sugary and carbonate drinks as well as with the composite indicator for obesogenic diets. CONCLUSIONS: The main strength of this study is the comprehensiveness and comparability of the policy data across the relatively large number of countries covered. There is a negative association that is statistically significant, between all formulations of the FREI and the household consumption of sugary and carbonated drinks. There is also a negative association between some FREI formulations and obesogenic diets.

Defining Priorities for Action and Research on the Commercial Determinants of Health: A Conceptual Review.

In recent years, the concept of commercial determinants of health (CDoH) has attracted scholarly, public policy, and activist interest. To date, however, this new attention has failed to yield a clear and consistent definition, well-defined metrics for quantifying its impact, or coherent directions for research and intervention. By tracing the origins of this concept over 2 centuries of interactions between market forces and public health action and research, we propose an expanded framework and definition of CDoH. This conceptualization enables public health professionals and researchers to more fully realize the potential of the CDoH concept to yield insights that can be used to improve global and national health and reduce the stark health inequities within and between nations. It also widens the utility of CDoH from its main current use to study noncommunicable diseases to other health conditions such as infectious diseases, mental health conditions, injuries, and exposure to environmental threats. We suggest specific actions that public health professionals can take to transform the burgeoning interest in CDoH into meaningful improvements in health. (Am J Public Health. 2021;111(12):2202-2211. https://doi.org/10.2105/AJPH.2021.306491).

Epigenetic Regulation of Myogenesis: Focus on the Histone Variants.

Skeletal muscle development and regeneration rely on the successive activation of specific transcription factors that engage cellular fate, promote commitment, and drive differentiation. Emerging evidence demonstrates that epigenetic regulation of gene expression is crucial for the maintenance of the cell differentiation status upon division and, therefore, to preserve a specific cellular identity. This depends in part on the regulation of chromatin structure and its level of condensation. Chromatin architecture undergoes remodeling through changes in nucleosome composition, such as alterations in histone post-translational modifications or exchange in the type of histone variants. The mechanisms that link histone post-translational modifications and transcriptional regulation have been extensively evaluated in the context of cell fate and differentiation, whereas histone variants have attracted less attention in the field. In this review, we discuss the studies that have provided insights into the role of histone variants in the regulation of myogenic gene expression, myoblast differentiation, and maintenance of muscle cell identity.

A comprehensive model for the diffusion and hybridization processes of nucleic acid probes in fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) has been extensively used in the past decades for the detection and localization of microorganisms. However, a mechanistic approach of the whole FISH process is still missing, and the main limiting steps for the hybridization to occur remain unclear. In here, FISH is approached as a particular case of a diffusion-reaction kinetics, where molecular probes (MPs) move from the hybridization solution to the target RNA site within the cells. Based on literature models, the characteristic times taken by different MPs to diffuse across multiple cellular barriers, as well as the reaction time associated with the formation of the duplex molecular probe-RNA, were estimated. Structural and size differences at the membrane level of bacterial and animal cells were considered. For bacterial cells, the limiting step for diffusion is likely to be the peptidoglycan layer (characteristic time of 7.94 × 102 - 4.39 × 103 s), whereas for animal cells, the limiting step should be the diffusion of the probe through the bulk (1.8-5.0 s) followed by the diffusion through the lipid membrane (1 s). The information provided here may serve as a basis for a more rational development of FISH protocols in the future.

Leaving no one behind: health equity as a catalyst for the sustainable development goals.

2019 has been a milestone year for catalyzing changes to improve health equity in the WHO European Region through concomitant progress in the sustainable development goal (SDG) targets. The WHO European Health Equity Status Report Initiative (HESRi) has captured and analyzed the relationships between inequities in health and the conditions that are essential for all to be able to live healthy and prosperous lives. The five essential conditions map directly onto a number of SDG targets, with a much broader span than SDG3 on health. They are: (i) Universal access to good-quality, affordable health services; (ii) Basic income security and social protection; (iii) Safe and decent living conditions; (iv) Inclusive social and human capital building opportunities; and (v) Decent and non-discriminatory employment and working conditions. There is certainly room for improvement in the way ahead, particularly in the availability of fine-grained and disaggregated data, and in the quality of monitoring and analysis of policy options that this would allow. However, the work of the HESRi shows that by harnessing such data it is possible to show what actions policymakers can take in the present to ensure that no one is left behind. This equity framing allows to measure whether the progress on SDGs benefits all, including those who need them most.

TGFß and FGF promote tendon progenitor fate and act downstream of muscle contraction to regulate tendon differentiation during chick limb development.

The molecular programme underlying tendon development has not been fully identified. Interactions with components of the musculoskeletal system are important for limb tendon formation. Limb tendons initiate their development independently of muscles; however, muscles are required for further tendon differentiation. We show that both FGF/ERK MAPK and TGFß/SMAD2/3 signalling pathways are required and sufficient for SCX expression in chick undifferentiated limb cells, whereas the FGF/ERK MAPK pathway inhibits Scx expression in mouse undifferentiated limb mesodermal cells. During differentiation, muscle contraction is required to maintain SCX, TNMD and THBS2 expression in chick limbs. The activities of FGF/ERK MAPK and TGFß/SMAD2/3 signalling pathways are decreased in tendons under immobilisation conditions. Application of FGF4 or TGFß2 ligands prevents SCX downregulation in immobilised limbs. TGFß2 but not FGF4 prevent TNMD and THBS2 downregulation under immobilisation conditions. We did not identify any intracellular crosstalk between both signalling pathways in their positive effect on SCX expression. Independently of each other, both FGF and TGFß promote tendon commitment of limb mesodermal cells and act downstream of mechanical forces to regulate tendon differentiation during chick limb development.

Targeting miR-9 in gastric cancer cells using locked nucleic acid oligonucleotides.

BACKGROUND: Gastric cancer is the third leading cause of cancer-related mortality worldwide. Recently, it has been demonstrated that gastric cancer cells display a specific miRNA expression profile, with increasing evidence of the role of miRNA-9 in this disease. miRNA-9 upregulation has been shown to influence the expression of E-cadherin-encoding gene, triggering cell motility and invasiveness. RESULTS: In this study, we designed LNA anti-miRNA oligonucleotides with a complementary sequence to miRNA-9 and tested their properties to both detect and silence the target miRNA. We could identify and visualize the in vitro uptake of low-dosing LNA-based anti-miRNA oligonucleotides without any carrier or transfection agent, as early as 2 h after the addition of the oligonucleotide sequence to the culture medium. Furthermore, we were able to assess the silencing potential of miRNA-9, using different LNA anti-miRNA oligonucleotide designs, and to observe its subsequent effect on E-cadherin expression. CONCLUSIONS: The administration of anti-miRNA sequences even at low-doses, rapidly repressed the target miRNA, and influenced the expression of E-cadherin by significantly increasing its levels.

Anti-miRNA oligonucleotides: A comprehensive guide for design.

MicroRNAs (miRNAs) are small, non-coding RNA molecules that regulate gene expression post-transcriptionally. As a consequence of their function towards mRNA, miRNAs are widely associated with the pathogenesis of several human diseases, making miRNAs a target for new therapeutic strategies based on the control of their expression. Indeed, numerous works were published in the past decades showing the potential use of antisense oligonucleotides to target aberrant miRNAs (AMOs) involved in several human pathologies. New classes of chemical-modified-AMOs, including locked nucleic acid oligonucleotides, have recently proved their worth in silencing miRNAs. A correct design of a specific AMOs can help to improve their performance and potency towards the target miRNA by increasing for instance nuclease resistance and target affinity. This review outlines the technologies involved to suppress aberrant miRNAs. From the design strategies used in AMOs to its application in novel miRNA-based therapeutics and detection methodologies.

Corporate practices and health: a framework and mechanisms.

BACKGROUND: The Global Burden of Disease estimates that approximately a third of deaths worldwide are attributable to behavioural risk factors that, at their core, have the consumption of unhealthful products and exposures produced by profit driven commercial entities. We use Steven Lukes' three-dimensional view of power to guide the study of the practices deployed by commercial interests to foster the consumption of these commodities. Additionally, we propose a framework to systematically study corporations and other commercial interests as a distal, structural, societal factor that causes disease and injury. Our framework offers a systematic approach to mapping corporate activity, allowing us to anticipate and prevent actions that may have a deleterious effect on population health. CONCLUSION: Our framework may be used by, and can have utility for, public health practitioners, researchers, students, activists and other members of civil society, policy makers and public servants in charge of policy implementation. It can also be useful to corporations who are interested in identifying key actions they can take towards improving population health.

Avifauna conservation and disturbance ecology in a coastal freshwater lagoon.

This work was developed in Barrinha de Mira Coastal Lagoon, Portugal. The lagoon belongs to the Natura 2000 network. Its main objectives were to monitor aquatic bird species before, during, and after a desorption intervention on this lagoon. Monitoring of waterfowl was carried out to evaluate the monthly variation of birds in Barrinha, in order to obtain useful information for region's management and planning, as well as for species preservation. In experimental design, it was decided to perform four counts per month in each pond (weekly), to evaluate the effect of increased monitoring effort in relation to previous years. The results revealed that the study area has a significant importance as a wintering zone for water birds. Some species have stable resident populations which, seasonally, see their numbers vary with migratory birds arrival and departure. Another purpose was to verify the influence of desorption intervention on the usual number of individuals by species. The results show that during and after intervention, there was a reduction of birds in the order of 89%, being more visible in the populations of Anas platyrhynchos and Fulica atra.

Austerity and Abortion in the European Union.

Economic hardship accompanying large recessions can lead families to terminate unplanned pregnancies. To assess whether abortions have risen during the recession, we collected crude abortion data from 2000 to 2012 from Eurostat for countries that had legal abortions and complete data. Declining trends in abortion ratios between 2000 and 2009 have been reversing. Excess abortions between 2010 and 2012 totaled 10.6 abortions per 1000 pregnancies ending in abortion or birth or 6701 additional abortions (95% CI 1190-9240) with stronger effects in younger ages. Economic shocks may increase recourse to abortion. Further research should explore causal pathways and protective factors.

Specific pattern of cell cycle during limb fetal myogenesis.

Tight regulation of cell proliferation and differentiation is required to ensure proper growth during development and post-natal life. The source and nature of signals regulating cell proliferation are not well identified in vivo. We investigated the specific pattern of proliferating cells in mouse limbs, using the Fluorescent ubiquitynation-based cell-cycle indicator (Fucci) system, which allowed the visualization of the G1, G1/S transition and S/G2/M phases of the cell cycle in red, yellow or green fluorescent colors, respectively. We also used the retroviral RCAS system to express a Fucci cassette in chick embryos. We performed a comprehensive analysis of the cell cycle state of myogenic cells in fetal limb muscles, adult myoblast primary cultures and isolated muscle fiber cultures using the Fucci transgenic mice. We found that myonuclei of terminally differentiated muscle fibers displayed Fucci red fluorescence during mouse and chick fetal development, in adult isolated muscle fiber (ex vivo) and adult myoblast (in vitro) mouse cultures. This indicated that myonuclei exited from the cell cycle in the G1 phase and are maintained in a blocked G1-like state. We also found that cycling muscle progenitors and myoblasts in G1 phase were not completely covered by the Fucci system. During mouse fetal myogenesis, Pax7+ cells labeled with the Fucci system were observed mostly in S/G2/M phases. Proliferating cells in S/G2/M phases displayed a specific pattern in mouse fetal limbs, delineating individualized muscles. In addition, we observed more Pax7+ cells in S/G2/M phases at muscle tips, compared to the middle of muscles. These results highlight a specific spatial regionalization of cycling cells at the muscle borders and muscle-tendon interface during fetal development.

Stable and bicistronic expression of two genes in somite- and lateral plate-derived tissues to study chick limb development.

BACKGROUND: Components of the limb musculoskeletal system have distinct mesoderm origins. Limb skeletal muscles originate from somites, while the skeleton and attachments (tendons and connective tissues) derive from limb lateral plate. Despite distinct mesoderm origins, the development of muscle, skeleton and attachments is highly coordinated both spatially and temporally to ensure complete function of the musculoskeletal system. A system to study molecular interactions between somitic-derived tissues (muscles) and lateral-plate-derived tissues (skeletal components and attachments) during limb development is missing. RESULTS: We designed a gene delivery system in chick embryos with the ultimate aim to study the interactions between the components of the musculoskeletal system during limb development. We combined the Tol2 genomic integration system with the viral T2A system and developed new vectors that lead to stable and bicistronic expression of two proteins at comparable levels in chick cells. Combined with limb somite and lateral plate electroporation techniques, two fluorescent reporter proteins were co-expressed in stoichiometric proportion in the muscle lineage (somitic-derived) or in skeleton and their attachments (lateral-plate-derived). In addition, we designed three vectors with different promoters to target muscle cells at different steps of the differentiation process. CONCLUSION: Limb somite electroporation technique using vectors containing these different promoters allowed us to target all myogenic cells, myoblasts or differentiated muscle cells. These stable and promoter-specific vectors lead to bicistronic expression either in somitic-derived myogenic cells or lateral plate-derived cells, depending on the electroporation sites and open new avenues to study the interactions between myogenic cells and tendon or connective tissue cells during limb development.