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Tropical dry forests (TDFs) are ecosystems with long drought periods, a mean temperature of 25 °C, a mean annual precipitation that ranges from 900 to 2000 mm, and that possess a high abundance of deciduous species (trees and lianas). What remains of the original extent of TDFs in the Americas remains highly fragmented and at different levels of ecological succession. It is estimated that one of the main fingerprints left by global environmental and climate change in tropical environments is an increase in liana coverage. Lianas are non-structural elements of the forest canopy that eventually kill their host trees. In this paper we evaluate the use of a terrestrial laser scanner (TLS) in combination with hemispherical photographs (HPs) to characterize changes in forest structure as a function of ecological succession and liana abundance. We deployed a TLS and HP system in 28 plots throughout secondary forests of different ages and with different levels of liana abundance. Using a canonical correlation analysis (CCA), we addressed how the VEGNET, a terrestrial laser scanner, and HPs could predict TDF structure. Likewise, using univariate analyses of correlations, we show how the liana abundance could affect the prediction of the forest structure. Our results suggest that TLSs and HPs can predict the differences in the forest structure at different successional stages but that these differences disappear as liana abundance increases. Therefore, in well known ecosystems such as the tropical dry forest of Costa Rica, these biases of prediction could be considered as structural effects of liana presence. This research contributes to the understanding of the potential effects of lianas in secondary dry forests and highlights the role of TLSs combined with HPs in monitoring structural changes in secondary TDFs.

Objective: Photovoice as a participatory research method seeks to capture and share participants' perspectives and experiences. It has been widely used in school settings to prioritise student voice in health research. Photovoice also presents opportunities for health education. There is limited synthesis on school-based Photovoice focused on nutrition and physical activity (PA). This systematic review examined the use of school-based Photovoice in relation to nutrition and PA for 10- to 18-year-olds. The research questions explored were twofold: what Photovoice approaches and implementation processes have been used in schools with respect to nutrition and PA; and what were the student learning outcomes? Methods: Six electronic databases were searched: MedLine, PsycInfo, EMBASE, ProQuest Education Journal, Cochrane Central and ERIC. The Participant, Intervention, Comparison, Outcome and Study (PICOS) framework was used to specify inclusion/exclusion criteria. Nineteen articles were selected from the search results of 2,305 articles based on fit with the established criteria. Iterative inductive thematic analysis was used to analyse the data with a focus on (1) approaches and implementation, and (2) student learning outcomes. Results: Nineteen studies were included in the review. Themes related to approaches and implementation were the importance in introductory sessions; processes for meaning-making; student-led community engagement and health promotion; and learning from challenges and limitations. Themes related to student learning outcomes included critical awareness of nutrition and PA environmental influences; capability development; and agency and empowerment. Conclusion: School-based Photovoice benefits health research by generating data with a focus on student voice. An under-valued and under-reported benefit relates to student learning outcomes, supporting the potential for healthy behaviour change. Multisectoral education and health research should value both types of goals. Photovoice can be used as a curriculum-linked pedagogical tool to add value to the pursuit of educational goals in school.

Abstract Background Untranslated regions (UTRs) are important mediators of post-transcriptional regulation. The length of UTRs and the composition of regulatory elements within them are known to vary substantially across genes, but little is known about the reasons for this variation in humans. Here, we set out to determine whether this variation, specifically in 5’UTRs, correlates with gene dosage sensitivity. Results We investigate 5’UTR length, the number of alternative transcription start sites, the potential for alternative splicing, the number and type of upstream open reading frames (uORFs) and the propensity of 5’UTRs to form secondary structures. We explore how these elements vary by gene tolerance to loss-of-function (LoF; using the LOEUF metric), and in genes where changes in dosage are known to cause disease. We show that LOEUF correlates with 5’UTR length and complexity. Genes that are most intolerant to LoF have longer 5’UTRs, greater TSS diversity, and more upstream regulatory elements than their LoF tolerant counterparts. We show that these differences are evident in disease gene-sets, but not in recessive developmental disorder genes where LoF of a single allele is tolerated. Conclusions Our results confirm the importance of post-transcriptional regulation through 5'UTRs in tight regulation of mRNA and protein levels, particularly for genes where changes in dosage are deleterious and lead to disease. Finally, to support gene-based investigation we release a web-based browser tool, VuTR, that supports exploration of the composition of individual 5'UTRs and the impact of genetic variation within them.

Background: Better understanding of the interaction between metabolism and immune response will be key to understanding physiology and disease. Tumor Necrosis Factor-alpha (TNFα) has been studied widely. However, despite the extensive knowledge about TNFα, the cytokine appears to induce not only variable, but often contradictory, effects on inflammation and cell proliferation. Despite advancements in the metabolomics field, it is still difficult to analyze the types of multi-dose, multi-time point studies needed for elucidating the varied immunologic responses induced by TNFα. Results: We studied the dose and time course effects of TNFα on murine fibroblast cultures and further elucidated these connections using selective blockade of the TNF receptors (TNFR1 and TNFR2). To streamline analysis, we developed a method to collate the metabolic pathway output from MetaboAnalyst into a single value for the Index of pathway significance (IPS). Using this metric, we tested dose-, time-, and receptor-dependent effects of TNFα signaling on cell metabolism. Guided by these results, we then demonstrate that alanine supplementation enriched TNFR1-related responses in both cell and mouse models. Conclusions: Our results suggest that TNFα, particularly when signaling through TNFR1, may preferentially use alanine metabolism for energy. These results are limited in by cell type used and immune outputs measured. However, we anticipate that our novel method may assist other researchers in identifying metabolic targets that influence their disease or model of interest through simplifying the analysis of multi-condition experiments. Furthermore, our results endorse the consideration of follow up studies in immunometabolism to improve outcomes in TNF-mediated diseases.

Abstract The terminal differentiation of osteoblasts and subsequent formation of bone marks an important phase in palate development that leads to the separation of the oral and nasal cavities. While the morphogenetic events preceding palatal osteogenesis are well explored, major gaps remain in our understanding of the molecular mechanisms driving the formation of this bony union of the fusing palate. Through bulk, single-nucleus, and spatially resolved RNA-sequencing analyses of the developing secondary palate, we identify a shift in transcriptional programming between embryonic days 14.5 and 15.5 pinpointing the onset of osteogenesis. We define spatially restricted expression patterns of key osteogenic marker genes that are differentially expressed between these developmental timepoints. Finally, we identify genes in the palate highly expressed by palate nasal epithelial cells, also enriched within palatal osteogenic mesenchymal cells. This investigation provides a relevant framework to advance palate-specific diagnostic and therapeutic biomarker discovery.

IntroductionEarly life interactions with the sow or humans can have lifelong consequences on welfare and performance in pigs. It was hypothesised that piglets that received increased maternal contact and positive human contact would display improved responses to stressful events and improved performance.MethodsNinety-eight litters were allocated to a 2 x 2 factorial design for maternal contact (MC+)/restricted maternal contact (MC-); and positive human contact (HC+)/no additional human contact (HC-). Modified farrowing crates were used to restrict maternal contact (MC-) and litters in the HC+ treatment received five minutes of daily positive human interaction (stroking). Behavioural and physiological responses were assessed during processing at three days of age and during a behaviour test in which piglets were introduced to an empty arena, novel object, human hand and human standing in the arena at 18 days of age. Observations of behaviour in the home pen and growth and survival of piglets to weaning were also assessed.ResultsAt processing, MC- piglets had more squeals (P = 0.015), tended to have more grunts (P = 0.063) and struggle attempts (P = 0.079), and had increased plasma cortisol concentrations (P = 0.009) after processing than MC+ piglets. During the behaviour test, HC+ piglets were more active (P = 0.014) and had more vocalisations (P < 0.05) in the empty arena than HC piglets. Furthermore, HC+ piglets were more likely to approach a human standing in the arena (P = 0.006) than HC- piglets. MC- piglets were less active (P = 0.008) and were less likely to approach the novel object (P = 0.049) than MC+ piglets. MC+ piglets tended to have higher weaning weights (P = 0.055) and more MC+ piglets were successfully weaned (P = 0.022) than MC- piglets. There was no significant effect of HC on piglet performance.DiscussionWhile positive handling did not affect behavioural and physiological stress response to processing, HC+ reduced fear of novelty and humans. These findings highlight the importance of early life experiences has on stress resilience early in life.

Mouse and human genetic studies indicate key roles of the Wnt10a ligand in odontogenesis. Previous studies have identified effectors and regulators of the Wnt signaling pathway actively expressed during key stages of tooth morphogenesis. However, limitations in multiplexing and spatial resolution hindered a more comprehensive analysis of these signaling molecules. Here, profiling of transcriptomes using fluorescent multiplex in situ hybridization and single-cell RNA-sequencing (scRNA-seq) provide robust insight into the synchronized expression patterns of Wnt10a, Dkk1, and Sost simultaneously during tooth development. First, we identified Wnt10a transcripts restricted to the epithelium at the stage of tooth bud morphogenesis, contrasting that of Sost and Dkk1 localization to the dental mesenchyme. By embryonic day 15.5 (E15.5), a marked shift of Wnt10a expression from dental epithelium to mesenchyme was noted, while Sost and Dkk1 expression remained enriched in the mesenchyme. By postnatal day 0 (P0), co-localization patterns of Wnt10a, Dkk1, and Sost were observed in both terminally differentiating and secreting odontoblasts of molars and incisors. Interestingly, Wnt10a exhibited robust expression in fully differentiated ameloblasts at the developing cusp tip of both molars and incisors, an observation not previously noted in prior studies. At P7 and 14, after the mineralization of dentin and enamel, Wnt10a expression was limited to odontoblasts. Meanwhile, Wnt modulators showed reduced or absent signals in molars. In contrast, strong signals persisted in ameloblasts (for Wnt10a) and odontoblasts (for Wnt10a, Sost, and Dkk1) towards the proximal end of incisors, near the cervical loop. Our scRNA-seq analysis used CellChat to further contextualize Wnt pathway-mediated communication between cells by examining ligand-receptor interactions among different clusters. The co-localization pattern of Wnt10a, Dkk1, and Sost in both terminally differentiating and secreting odontoblasts of molars and incisors potentially signifies the crucial ligand-modulator interaction along the gradient of cytodifferentiation starting from each cusp tip towards the apical region. These data provide cell type-specific insight into the role of Wnt ligands and mediators during epithelial-mesenchymal interactions in odontogenesis.

Weaning stress imposes considerable physiological challenges on piglets, often manifesting in intestinal disturbances, such as inflammation and compromised barrier function, ultimately affecting growth and health outcomes. While conventional interventions, including antimicrobials, have effectively mitigated these sequelae, concerns surrounding antimicrobial resistance necessitate the exploration of alternatives. Fucoidan, derived from brown seaweed, offers promise due to its antioxidant and anti-inflammatory effects. Previous research has been limited to the in-feed supplementation of partially purified fucoidan extracted from brown seaweed. The focus of the present study is assessing the effect of a preweaning drench with highly purified (85%) fucoidan on piglet growth, immune response, and intestinal morphology post-weaning. Forty-eight male piglets at 17 ± 3 days of age (5.67 ± 0.16 kg) were assigned to a saline (control), fucoidan, or antimicrobial group, receiving treatment as a single 18 mL oral drench three days before weaning. Monitoring for seven days post-weaning included body weight measurements, blood sample collection for the inflammatory protein assay, and small intestine morphological analysis. The findings revealed that the preweaning fucoidan drench did not elicit adverse effects on piglets. However, neither fucoidan nor antimicrobial drenches significantly enhanced growth parameters, immune markers, or intestinal morphology compared to that of the control-treated piglets (p > 0.05). The lack of response may be attributed to the high health status of the experimental cohort and the limitation of a single dosage. Future research should consider a more challenging production setting to evaluate the viability and optimal application of fucoidan as an antimicrobial alternative in the pig industry.

Abstract Cocoa products have a remarkable chemical and sensory complexity. However, in contrast to other fermentation processes in the food industry, cocoa bean fermentation is left essentially uncontrolled and is devoid of standardization. Questions of food authenticity and food quality are hence particularly challenging for cocoa. Here we provide an illustration how network science can support food fingerprinting and food authenticity research. Using a large dataset of 140 cocoa samples comprising three cocoa fermentation/processing stages and eight countries, we obtain correlation networks between the cocoa samples by computing measures of pairwise correlation from their liquid chromatography-mass spectrometry (LC-MS) profiles. We find that the topology of correlation networks derived from untargeted LC-MS profiles is indicative of the fermentation and processing stage as well as the origin country of cocoa samples. Progressively increasing the correlation threshold firstly reveals network clusters based on processing stage and later country-based clusters. We present both, qualitative and quantitative evidence through network visualization, network statistics and concepts from machine learning. In our view, this network-based approach for classifying mass spectrometry data has broad applicability beyond cocoa.

Abstract Objective To identify an age at which initiation of whole nut into the pediatric diet could be considered safe, by evaluating the age distribution of children undergoing bronchoscopy with removal of nut or seed material from the airway. Method A retrospective chart review over a ten-year period identifying children age 0–18 that have undergone bronchoscopy with retrieval of airway foreign bodies. A statistical analysis of demographic data was carried out to identify age distribution of aspiration events. Results Sixty-four cases of foreign body aspiration were identified, of which 43 (67%) were of organic origin, specifically nuts. A Fisher’s exact test was carried out on the cumulative percentage of organic foreign body aspirations to identify the age distribution of nut aspiration events. A statistically significant decrease in organic foreign body aspirations occurred at approximately 36 months of age (p = 0.004). Conclusion Foreign body aspiration is a leading cause of accidental injury or death in children. Nut and other small organic foreign bodies account for a significant portion of accidental aspiration events, however, no guidelines exist regarding appropriate age of whole nut introduction into the diet. Our study suggests that 90% of pediatric nut aspiration events occur under the age of 36 months. We suggest supervised introduction of whole nuts between the ages of 3 and 4 years. Official guidelines regarding this should be considered by professional pediatric societies. Level of evidence 4

This meta-regression analysis was conducted to identify the relationship between fibretype cross-sectional area (CSA) and frequency (%) and meat quality traits, especially tenderness (sensory and Warner-Bratzler Shear Force, WBSF). Literature searches were conducted using specific keywords which resulted in 32 peer-reviewed manuscripts that contained averages and correlation coefficients for fibre type (frequency and CSA) and quality traits of longissimus muscle for beef and pork (7 and 25 studies respectively). Correlations were analysed in meta-regression using R-Studio and linear regression was also conducted. For the combined beef and pork analysis, only pH, WBSF, and drip loss were associated with fibre type frequency and CSA (p < 0.05 for all). Limiting the analysis to pork, the key results were frequency of type I fibres were associated with decreased drip loss, increased cook loss, decreased lightness (L*) and increased sensory tenderness whereas frequency of type IIb fibres were associated with increased drip loss (p < 0.05 for all). In addition, the CSA of type I and IIb fibres was associated with colour traits lightness and redness (p < 0.05 for all). Future research should focus on fibre type across breeds and muscles to further understand the impacts of fibre type frequency and CSA on quality.

Shoulder sores in sows cause pain, may lead to early weaning and resultant piglet distress, and premature culling. Early detection and intervention is key to avoid these substantial production and welfare imposts. In this experiment we tested infrared thermography (IRT) to identify shoulder sores before wound eruption, and effectiveness of four wound healing treatments; mānuka honey (n = 11); Derisal® (n = 11); Chloromide® (n = 13) and Repiderma® (n = 16), all of which contain no antibiotics. Three hundred and ten sows (parity 2.3 ± 0.2) were monitored daily from entry to the farrowing sheds until weaning using a thermal camera. IRT successfully detected 88% of shoulder sores as hot spots and provided a lead in time of 7 days. Sixteen percent of all sows had a hotspot detected and were randomly allocated to one of four daily treatment groups. At the end of the treatment period, sore diameter was significantly reduced for all treatments, except the Repiderma® group. Sow traits had little influence on susceptibility to shoulder sores. There may be a link between prewean mortality and shoulder sores, but this requires further investigation. The use of IRT to monitor for hotspots for early intervention is validated. Future work should concentrate on methods to prevent wound eruption after detection with IRT to improve the health and welfare of both the sow and her litter.

BackgroundAmeloblasts are epithelially derived cells responsible for enamel formation through a process known as amelogenesis. Amongst the several transcription factors that are expressed during amelogenesis, both Msx2 and Sp6 transcription factors play important role. Msx2 and Sp6 mouse mutants, exhibit similar amelogenesis defects, namely enamel hypoplasia, while humans with amelogenesis imperfecta (AI) carry mutations in the human homologues of MSX2 or SP6 genes. These across species similarities in function indicate that these two transcription factors may reside in the same developmental pathway. In this paper, we test whether they work in a coordinated manner to exert their effect during amelogenesis.MethodsTwo different dental epithelial cell lines, the mouse LS8 and the rat G5 were used for either overexpression or silencing of Msx2 or Sp6 or both. Msx2 mutant mouse embryos or pups were used for in vivo studies. In situ hybridization, semi-quantitative and quantitative real time PCR were employed to study gene expression pattern. MatInspector was used to identify several potential putative Msx2 binding sites upstream of the murine Sp6 promoter region. Chromatin Immunoprecipitation (chIP) was used to confirm the binding of Msx2 to Sp6 promoter at the putative sites.ResultsUsing the above methods we identified that (i) Msx2 and Sp6 exhibit overlapping expression in secretory ameloblasts, (ii) Sp6 expression is reduced in the Msx2 mouse mutant secretoty ameloblasts, and (iii) that Msx2, like Sp6 inhibits follistatin expression. Specifically, our loss-of function studies by silencing Msx2 and/or Sp6 in mouse dental epithelial (LS8) cells showed significant downregulation of Sp6 but upregulation of Fst expression. Transient transfection of Msx2 overexpression plasmid, up-regulated Sp6 and downregulated Fst expression. Additionally, using MatInspector, we identified several potential putative Msx2 binding sites, 3.5 kb upstream of the murine Sp6 promoter region. By chIP, we confirmed the binding of Msx2 to Sp6 promoter at these sites, thus suggesting that Sp6 is a direct target of Msx2.ConclusionCollectively, these results show that Sp6 and Msx2 work in a concerted manner to form part of a network of transcription factors that operate during later stages of tooth development controlling ameloblast life cycle and amelogenesis.

MALDI imaging is a novel technique with which to study the pathophysiologies of diseases. Advancements in the field of metabolomics and lipidomics have been instrumental in mapping the signaling pathways involved in various diseases, such as cancer and neurodegenerative diseases (Parkinson’s). MALDI imaging is flexible and can handle many sample types. Researchers primarily use either formalin-fixed paraffin-embedded (FFPE) or fresh frozen tissue samples to answer their scientific questions. FFPE samples allow for easy long-term storage, but the requirement for extensive sample processing may limit the ability to provide a clear picture of metabolite distribution in biological tissue. Frozen samples require less handling, but present logistical challenges for collection and storage. A few studies, mostly focused on cancer cell lines, have directly compared the results of MALDI imaging using these two tissue fixation approaches. Herein, we directly compared FFPE and fresh frozen sample preparation for murine skin samples, and performed detailed pathway analysis to understand how differences in processing impact MALDI results from otherwise identical tissues. Our results indicate that FFPE and fresh frozen methods differ significantly in the putative identified metabolite content and distribution. The fixation methods shared only 2037 metabolites in positive mode and only 4079 metabolites in negative ion mode. However, both fixation approaches allowed for downstream fluorescent staining, which may save time and resources for samples that are clinically precious. This work represents a direct comparison of the impacts of the two main tissue processing methods on subsequent MALDI-MSI. While our results are similar to previous work in cancer tissue, they provide novel insights for those using MALDI-MSI in skin.

Abstract Background We examined the efficacy of olanzapine/fluoxetine combination (OFC) in improving health-related quality of life (QoL) in the treatment of bipolar depression in children and adolescents. Methods Patients aged 10–17 years with bipolar I disorder, depressed episode, baseline children’s depression rating scale-revised (CDRS-R) total score ≥40, Young Mania Rating Scale (YMRS) total score ≤15, and YMRS-item 1 ≤ 2 were randomized to OFC (6/25–12/50 mg/day olanzapine/fluoxetine; n = 170) or placebo (n = 85) for up to 8 weeks of double-blind treatment. Patients and parents completed the revised KINDL questionnaire for measuring health-related QoL in children and adolescents (KINDL-R) at baseline and endpoint. The mean change in CDRS-R total and item scores were used to compare improvement in symptomatology in patients taking OFC and placebo. Tests were 2-sided using a Type I error cutoff of 0.05, and no adjustments for multiple comparisons were made. Results Baseline QoL as measured by the KINDL-R was substantially impaired relative to published norms for a healthy school-based sample. OFC-treated patients demonstrated an improvement over placebo at endpoint with respect to mean change from baseline in the patient-rated KINDL-R Self-esteem subscale score (p = 0.028), and in the parent KINDL-R ratings of emotional well-being (p = 0.020), Self-esteem (p = 0.030), and Family (p = 0.006). At endpoint, OFC-treated patients still had a lower QoL compared to the normative population. OFC showed significant improvement (p ≤ 0.05) versus placebo on the CDRS-R total score and on 7 of the 17 CDRS-R items. Conclusions Patients aged 10–17 years with an acute episode of bipolar depression and their parents reported greater improvements (parents noticed improvements in more areas than did their offspring) on some aspects of QoL when treated with OFC compared with placebo. However, after 8 weeks of treatment, KINDL-R endpoint scores remained lower than those of the, presumably healthy, control population. Clinical trial registration information A Study for Assessing Treatment of Patients Ages 10–17 with Bipolar Depression; http://www.clinicaltrials.gov ; NCT00844857

Forty crossbred (Large White × Landrace × Duroc) female pigs (16.4 kg ± 0.94 kg) were used to investigate the effect of dietary lecithin supplementation on growth performance and pork quality. Pigs were randomly allocated to a commercial diet containing either 0, 3, 15 or 75 g lecithin/kg of feed during the grower and finisher growth phase. Pork from pigs consuming the diets containing 15 g and 75 g lecithin/kg had lower hardness ( P < 0.001) and chewiness ( P < 0.01) values compared to the controls. Dietary lecithin supplementation at 75 g/kg significantly increased ( P < 0.05) the linoleic acid and reduced ( P < 0.05) the myristic acid levels of pork compared to the control and the 3 g/kg and 15 g/kg lecithin supplemented treatments. Pigs fed the 75 g/kg lecithin supplemented diet had lower plasma cholesterol ( P < 0.05) at slaughter compared to pigs fed the control diet and the 3 g/kg and 15 g/kg lecithin supplemented treatments. These data indicate that dietary lecithin supplementation has the potential to improve the quality attributes of pork from female pigs.

Amy S Chappell1, Laurence A Bradley2, Curtis Wiltse1, Michael J Detke1,3,4, Deborah N D’Souza1, Michael Spaeth51Lilly Research Laboratories, Indianapolis, IN, USA; 2University of Alabama at Birmingham, Birmingham, Alabama, USA; 3Indiana University School of Medicine, Indianapolis, IN, USA; 4Harvard Medical School, Boston, MA, USA; 5Practice for Internal Medicine/Rheumatology, Graefelfing, GermanyObjective: Assess the efficacy of duloxetine 60/120 mg (N = 162) once daily compared with placebo (N = 168) in the treatment of patients with fibromyalgia, during six months of treatment.Methods: This was a phase-III, randomized, double-blind, placebo-controlled, parallel-group study assessing the efficacy and safety of duloxetine.Results: There were no significant differences between treatment groups on the co-primary efficacy outcome measures, change in the Brief Pain Inventory (BPI) average pain severity from baseline to endpoint (P = 0.053) and the Patient’s Global Impressions of Improvement (PGI-I) at endpoint (P = 0.073). Duloxetine-treated patients improved significantly more than placebo-treated patients on the Fibromyalgia Impact Questionnaire pain score, BPI least pain score and average interference score, Clinical Global Impressions of Severity scale, area under the curve of pain relief, Multidimensional Fatigue Inventory mental fatigue dimension, Beck Depression Inventory-II total score, and 36-item Short Form Health Survey mental component summary and mental health score. Nausea was the most common treatment-emergent adverse event in the duloxetine group. Overall discontinuation rates were similar between groups.Conclusions: Although duloxetine 60/120 mg/day failed to demonstrate significant improvement over placebo on the co-primary outcome measures, in this supportive study, duloxetine demonstrated significant improvement compared with placebo on numerous secondary measures.Keywords: fibromyalgia, duloxetine, placebo, double-blind, trial, pain