bluuebunny/crossref_metadata_2025 huggingface dataset:
Analysis of Food Advertising in Children's TV Programming
Authors: Cosenza Emma Lucia , Morales‐Juárez Analí , Kroker‐Lobos Maria Fernanda , Vandevijvere Stefanie , Ramirez‐Zea Manuel
Published: November 2023

Background

Guatemala, as most middle‐income countries, experiences a rise of overweight and obesity in school‐aged children. Food advertising on TV has been shown to affect children's food choices and intake.

Objective

The aim of this study was to assess, for the first time, the extent and nature (e.g. persuasive techniques and claims) of food advertisements, targeted at children on the most watched TV channels featuring children's shows.

Methods

The methodology of the International Network for Food and Obesity Non‐Communicable Diseases Research, Monitoring and Action Support (INFORMAS) was used. We recorded 864h (from 6:00 am to 12:00 am) of TV programming during 4 weekend days and 4 weekdays on the 6 most viewed TV channels featuring children's shows (1 local and 5 cable channels), during the period July–September 2016, excluding school holidays. Foods advertised were classified in the following groups: sugar‐sweetened beverages, snacks and sweets, sugar‐sweetened cereals, fast food restaurants, and dairy products with sugar.

Results

Of all advertisements, 19% (2,709 of 14589) were food related commercials, of which 54% (n= 1,459) were for energy‐dense foods. Forty one percent of energy‐dense food ads used both promotional characters (i.e., cartoons, famous personalities, and elements related to kids) and premium offers (i.e., collectibles, gifts, discounts, apps). Premium offers were most of the times accompanied by promotional characters (99% of cases). Concerning claims, 22% of energy‐dense food ads used both brand benefit claims (i.e., emotive claims) and health claims (i.e., health related ingredient claims), 72% presented only brand benefit claims, and 6% depicted exclusively health claims.

Conclusion

This study revealed that Guatemalan TV channels featuring children's shows have a high frequency of energy‐dense food ads and of persuasive marketing techniques in them. There is a need of monitoring and regulating food advertisements in TV channels featuring children's shows to reduce exposure.

Support or Funding Information

International Development Research Center (IDRC)
Are Diets with a Lower Carbon Footprint More Nutritious? Evidence in the United States from the National Health and Nutrition Examination Survey
Authors: Rose Diego , Willits‐Smith Amelia , Heller Martin , Meyer Robert
Published: November 2023

Although the US is the second highest‐ranked country in the world in greenhouse gas emissions (GHGE), and food production is a significant contributor to this, little is known about the impact of individual food choices in the US on GHGE or how this might relate to nutrient intakes. To address this gap, we developed a novel approach to study the environmental impacts of individual food choices in the US. Individual 24‐hour recall data on foods consumed by adults, 18–65 years of age (N= 13,204), from the National Health and Nutrition Examination Survey (NHANES) for 2005–2010 were converted to commodity form using resources from the US Environmental Protection Agency. The GHGE of these commodities were aggregated from a comprehensive review of life cycle assessment studies, which estimate environmental impacts throughout the “life” of a product, in this case foods. After matching the food choices of individuals to these GHGE, we ranked their 1‐day diets into quintiles based on GHGE per 1000 kilocalories, and compared the lowest and highest GHGE quintile groups. We focused on 12 key nutrients of concern that were identified in the Dietary Guidelines for Americans as either being under‐consumed by the US population or over‐consumed by it. T‐tests for differences in means were conducted using Stata 13.0 and made use of sample weights and survey design parameters published with the datasets. We estimate that the production of foods consumed by U.S. adults contributed 5.49 carbon dioxide‐equivalents per person per day (95% CI = 5.38 to 5.60), a value which is within the range of GHGE impacts found in European countries. The lowest GHGE diets (bottom quintile) scored better on five of the key nutrients, with significantly (p < 0.001) more vitamin E, folate, and dietary fiber per 1000 kilocalories and significantly (p < 0.001) less sodium and saturated fat than the highest GHGE diets. However, there was a higher content of three of the nutrients ‐‐ calcium, vitamin D, and potassium ‐‐ in the highest GHGE diets. We found similar results when limiting the sample to only those recalls that respondents identified as representing a “usual” amount. These results suggest that food patterns which are less environmentally impactful are more nutritious on several key dimensions, although not on all of them. The links between food choice, nutritional outcomes and environmental impacts are nuanced, which calls for more research and understanding of these interactions.

Support or Funding Information

This work was supported by the Wellcome Trust (grant # 106854/Z/15/Z).
Regular Consumption of a Functional Symbiotic Yogurt Increases Calcium Absorption in Young Adult Women
Authors: Cornes Rafael , Peña Amparo , Sintes Celia , Albin Santiago , O'Brien Kimberly O. , Donangelo Carmen Marino
Published: November 2023

We investigated the effect of daily consumption of a yogurt containing prebiotic fiber and probiotics (functional yogurt) compared to a common yogurt (control), on calcium absorption in adult women (21–33 years; n=30) using a cross‐over design. Each yogurt was consumed during 3 weeks separated by a 3 week wash‐out period. Calcium absorption was measured at the end of each yogurt‐consumption period, using an oral isotope tracer (42Ca) administered along with a standard breakfast meal. Recovery of 42‐‐‐‐‐‐‐Ca in urine was measured in complete collections at 4h, 24h, and 36h post‐dosing using magnetic sector thermal ionization mass spectrometry. Bone mass, PTH and vitamin D status were evaluated at baseline. Bone mineral content (BMC) and density (BMD) at total body (TB), lumbar spine (LS), femoral neck (FN) and trochanter (TR) were measured by dual energy X‐ray absorptiometry. Serum 25(OH)D and intact PTH were determined by chemiluminescent assays. Temporal changes in urinary 42‐‐‐‐‐‐‐Ca recovery were analyzed by repeated‐measures ANOVA. Comparison between treatments at each time point was evaluated by paired t test. Relationships between variables were examined by correlation analysis. Average Z scores of bone measurements were adequate, 0.59 (TBBMC), 0.49 (TBBMD), 0.05 (LSBMD), 0.99 (FNBMD), −0.54 (TRBMD) although TRBMD and LSBMD Z scores were lower than −1 in 30% and 7% of the women, respectively. Serum 25(OH)D was 37.9 ± 9.9 ng/mL indicating vitamin D sufficiency. As expected, urinary 42‐‐‐‐‐‐‐Ca recovery increased over time for each type of yogurt (P<0.01). There were no significant differences by yogurt type at 4h or 24h but at 36h, 42‐‐‐‐‐‐‐Ca recovery in urine was 37% higher after consumption of the functional yogurt compared to the control yogurt (P=0.05). Urinary recovery of 42‐‐‐‐‐‐‐Ca was not related to vitamin D, PTH or bone mass measurements. These results suggest that the functional yogurt tested increased calcium absorption possibly by passive processes in the distal small intestine or in the colon.

Support or Funding Information

Cooperativa Nacional de Productores de Leche (CONAPROLE), Uruguay
Risk Factors of Poor Complementary Feeding Practices in Pakistani Children Aged 6–23 months: A Multi‐Level Analysis of the Demographic and Health Survey 2012–2013
Authors: Na Muzi , Aguayo Victor M , Arimond Mary , Stewart Christine P.
Published: November 2023

Appropriate feeding practices are crucial for survival, growth, and development in childhood. This study aims to identify risk factors associated with poor complementary feeding practices in Pakistani children. We analyzed Pakistan's Demographic and Health Survey 2012–2013 using multi‐level models to fit the multistage cluster sample of 2827 children aged 6–23 months from 489 communities. Timely introduction of solid, semi‐solid or soft foods was achieved in 67% infants aged 6–8 months. Among children aged 6–23 months, the proportion of children meeting minimum meal frequency (MMF), dietary diversity (MDD) and acceptable diet (MAD) criteria were 63%, 22% and 15%, respectively. The mean (95%CI) dietary diversity score was 1.8 (1.7, 1.9), 2.6 (2.5, 2.7) and 3.0 (2.8, 3.1) food groups in children aged 6–11, 12–17 and 18–23 months, respectively. Consumption of legumes and nuts, flesh foods, and vitamin A‐rich fruits and vegetables was low in all children (6–19%), even among children who met the minimum dietary diversity criterion (15–55%). Younger child age, especially between 6–11 months, delayed maternal postnatal checkup and household poverty were significant individual‐level risk factors that consistently increased the risk of not meeting the four criteria examined. Insufficient antenatal clinic visits (1–3 times) was independently associated with not introducing complementary foods on time and not meeting MMF. Additional significant risk factors associated with not meeting MDD and MAD included small perceived birth weight and younger maternal age. Community‐level risk factors included living in the Punjab region and having poor access to maternal and child health care services. The overall quality of complementary diets in Pakistani children was very poor. This situation calls for stronger policy and program action to promote the consumption of key nutrient‐dense foods, especially among 6–11 month old infants, while prioritizing interventions for the most vulnerable children and regions.

Support or Funding Information

UNICEF Regional Office for South Asia (ROSA)
Milk cell gene expression of mothers with low breast milk production
Authors: Geddes Donna T. , Twigger Alecia‐Jane , Savigni Donna L. , Kent Jacqueline C. , Kakulas Foteini
Published: November 2023

Introduction

Perceived or actual low milk supply is the leading cause of early breastfeeding cessation worldwide. Monitoring 24 hour milk production profiles can help diagnose low milk production which may be improved with pumping regimes and support. Despite this, some cases of low milk production persist. Human milk cells are increasingly being utilised to provide non‐invasive access to mammary epithelial cells allowing investigations into lactocyte characteristics and metabolism.

Problem Statement

The molecular mechanisms behind impaired lactation in the female mammary gland are not well understood.

Procedures/Data/Observations

This study examines human milk cell gene expression from 12 participants with low milk production profiles (<600 mL in 24 hrs) and 14 participants with normal production profiles (>600 mL in 24 hrs). Genes (n=28) were examined using quantitative reverse transcription polymerase chain reaction (qRT‐PCR) and were representative of progenitor and myoepithelial cells and lactocytes, as well as genes identified as intrinsic to milk production. Students t‐test was used to compare gene expression between the two groups. Alternatively, Welch's t‐test was used in the case of unequal variances as determined by Bartlett's test.

Results

Initial analysis found cells isolated from women with low milk production showed significantly lower expression of the genes estrogen related receptor beta (ESRRB, p=0.027) and neurotrophin receptors sortilin (SORT, p=0.010) and tyrosine receptor kinase 2 splice variant 1 (TRKB1, p=0.007) and higher expression of a progenitor marker (REX1, p=0.025) compared with cells isolated from women with normal production.

Conclusions

Preliminary findings suggest variations in cell signalling and function, examined through gene expression that might contribute to low milk production. Further investigations will potentially determine significant roles of key genes enabling successful human lactation.

Support or Funding Information

This research was funded by an unrestricted research grant from Medela AG.
α‐Tocopherol–a systematic review of intake and status globally
Authors: Eggersdorfer Manfred
Published: November 2023

Objective

Review the published literature reporting vitamin E intake levels and serum concentration in order to obtain a global overview of vitamin E status. Vitamin E is essential for human health and achieving an optimal status is associated with beneficial health outcomes. Dietary recommendations are established in many countries around the world and refer to the important role of vitamin E in preserving the integrity of the cell membrane. The outcome and comparison of intake versus actual status should provide new insights for recommendations.

Method

A search in the Pubmed/Medline database focused on population based studies published between January 1st 2000 and July 30th 2012 and resulted in 176 articles referring to 132 single studies with reported intake and/or serum concentration.

Results

Applying a Recommended Daily Allowance (RDA) of 15 mg/d to all populations with a minimum age of 14 years, 82% and 61% of mean and median data points were below the RDA and EAR, respectively. Regarding serum concentrations, globally 13% of the included data points were below the functional deficiency threshold concentration of 12 μmol/L (F.a.N. Board 2000), mostly newborns and children. Several prospective observational studies suggest that a serum α‐tocopherol concentration of 30 μmol/L and above to have beneficial effects on human health. Of the reported study populations and subpopulations, only 21% reached this threshold globally.

Conclusions

The systematic review suggests that the vitamin E status is inadequate in a substantial part of the studies populations. This review could be a useful stepping‐stone for researchers to combine existing data, fill in data gaps and to understand more about the complex field of vitamin E and its impact on human health.
Neutrophil microparticles deliver active myeloperoxidase to injured mucosa to inhibit epithelial wound healing
Authors: Sumagin Ronen , Finkielsztein Ariel , Slater Thomas , Mascarenhas Lorraine Lorraine , Mehl Lindsey , Butin‐Israeli Veronika
Published: November 2023

Neutrophil (PMN) infiltration of the intestinal mucosa often leads to severe epithelial injury, however how this process occurs is unclear. The current work describes a novel mechanism, whereby membrane derived microparticles released by tissue infiltrating PMNs (PMN‐MPs) serve as shuttles to protect and deliver active mediators to locally modulate cellular function during inflammation. Specifically, myeloperoxidase (MPO), which is abundantly expressed in PMN azurophilic granules and is used for microbial killing, was found to be mobilized to the PMN surface and subsequently released in association with PMN‐MPs upon PMN activation and binding to intestinal epithelial cells (IECs). The enzymatic activity of PMN‐MP‐associated MPO was enhanced compared to soluble protein, leading to potent inhibition of wound closure following PMN‐MP binding to IECs. Importantly, localized microinjection of PMN‐MPs into wounded colonic mucosa was sufficient to impair epithelial wound healing in vivo. PMN‐MPs/MPO‐dependent inhibition of IEC wound healing was due to impaired IEC migration and proliferation, resulting from impeded actin dynamics, cell spreading, and cell cycle arrest. Thus, our findings provide new insight into mechanisms governing PMN‐induced tissue injury, and implicate PMN‐MPs and MPO as important regulators of cellular function.

Support or Funding Information

This work was supported by grants from the National Institutes of Health (NIH) DK101675 as well as American Cancer Society IRG‐9303718.
Desmoglein‐2 Intracellular Cleavage and Intestinal Epithelial Barrier Compromise
Authors: Yulis Mark , Quiros Miguel , Hilgarth Roland , Nusrat Asma
Published: November 2023

The intestinal epithelial barrier is comprised of a monolayer of simple columnar intestinal epithelial cells (IECs) that serve to protect underlying tissues from antigens and pathogens. Under conditions of intestinal inflammation this barrier breaks down, allowing these antigens and pathogens to gain access to the underlying mucosa. The aim this project is to further our understanding of the mechanisms responsible for barrier breakdown during intestinal inflammation. Epithelial barrier properties are achieved by a series of intercellular junctions that include an Apical Junctional Complex and desmosomes. The intercellular contacts of desmosomes are established by transmembrane cadherin proteins, the desmogleins and desmocollins. Pro‐inflammatory cytokines (e.g. interferon gamma (IFN‐γ) and tumor necrosis factor alpha (TNF‐α)) induce cadherin cleavage and the resulting cadherin protein fragments influence epithelial homeostasis. Using model IECs plated on transwell permeable supports, we have observed that IFN‐γ and TNF‐α or TNF‐α related apoptosis inducing ligand (TRAIL) induce desmoglein‐2 (Dsg2) intracellular cleavage resulting in generation of an intracellular fragment (ICF). This Dsg2 intracellular cleavage is, in part, mediated by caspase activation and occurs prior to Dsg2 extracellular cleavage. Previously published data suggests that Dsg2 ICF generation is associated with redistribution of another desmosomal protein, plakoglobin (Pg). We show here that exogenous expression of Myc‐tagged Dsg2 ICF using adenoviral expression constructs results in downregulation of key Pg signaling targets (c‐Myc and Bcl‐XL), increases apoptosis, and decreassd cell proliferation. Taken together, our findings suggest that IFN‐γ and TNF‐α induced Dsg2 ICF generation activates Pg signaling thereby promoting increased epithelial apoptosis and reduced proliferation.

Support or Funding Information

Intestinal Epithelial Tight Junction Structure‐Function: R01 DK059888‐15
Intestinal P‐glycoprotein exports endogenous cannabinoids at the epithelial surface to prevent inflammation and maintain homeostasis
Authors: Szabady Rose L , Mrsny Randall J , McCormick Beth A
Published: November 2023

Neutrophil influx into the intestine plays a critical protective role during infection with pathogens, but is also associated with the severe intestinal damage seen in idiopathic inflammatory bowel disease (IBD). Transmigration of neutrophils across the epithelium and into the lumen is particularly associated with tissue destruction, increased barrier permeability, and inflammatory pathology. This transepithelial migration is driven by the chemoattractant activity of hepoxilin A3 (HXA3), an eicosanoid secreted from the epithelial surface by the MRP2 efflux pump. MRP2 is upregulated in response to inflammatory stimuli including bacterial infection or epithelial damage, and we have confirmed that MRP2 secretion of HXA3 drives neutrophil transmigration in a variety of inflammatory models. Concurrent with MRP2 upregulation, a second ABC transporter, P‐glycoprotein (P‐gp), is down‐regulated at the epithelial surface during inflammation. We therefore investigated whether P‐gp was responsible for transporting immunomodulatory lipids that act to counterbalance the MRP2/HXA3 pro‐inflammatory axis.

Mass spectrometry analysis of epithelial supernatants revealed that endocannabinoids were specifically secreted into the supernatant only in the presence of P‐gp. Further analysis revealed that specific P‐gp secreted endocannabinoids of the N‐acyl ethanolamine class were capable of suppressing neutrophil migration in vitro, and that this suppression was functional in vivo. Having established that under homeostatic conditions, P‐gp constitutively secretes anti‐inflammatory endocannabinoids, we investigated how perturbation of this system affects the development of intestinal inflammation.

P‐gp deficient mice are known to develop spontaneous inflammation, consistent with our finding that P‐gp secretes anti‐inflammatory endocannabinoids. CB2 is the peripheral cannabinoid receptor found on immune cells, and we predicted that the neutrophils of CB2‐deficient mice would be unable to respond to P‐gp secreted endocannabinoids and would thus be primed for inflammation. Induction of colitis in CB2‐deficient mice resulted in increased pathology and neutrophil influx into the intestinal lumen. Bone marrow chimera experiments confirmed that these effects resulted from a lack of CB2 in the hematopoietic compartment. These results define a key role for epithelial cells in balancing secretion of anti‐inflammatory and pro‐inflammatory lipids via surface efflux pumps in order to maintain homeostasis in the healthy intestine, yet remain poised to drive neutrophil transmigration in response to pathogens.

Support or Funding Information

These studies were supported by NIH R01 DK056754 to B.A. McCormick and R.J. Mrsny and NIH F32 DK098973 to R.L. Szabady.
Increased Intestinal Permeability Secondary To Junctional Adhesion Molecule‐A Deficiency Results In Impaired Macrophage‐Dependent Neutrophil Recruitment In A Zymosan Peritonitis Model
Authors: Luissint Anny Claude , Williams Holly C , Kim Wooki , Hilgarth Roland S , O'Leary Monique , Flemming Sven , Reed Michelle , Denning Timothy L , Nusrat Asma , Parkos Charles A
Published: November 2023

JAM‐A is a transmembrane protein expressed on endothelial and epithelial cells as well as on the surface of leukocytes. Studies with JAM‐A knockout mice (JAM‐A−/−), have provided evidence that JAM‐A may facilitate tissue recruitment of neutrophils (PMN); however mechanisms regulating JAM‐A‐dependent migration of PMN in‐vivo is not well understood. Here, we used a zymosan‐induced peritonitis model to study the recruitment of PMN in total knockout mice (JAM‐A −/−) and compared results to those obtained in mice with tissue selective deletion of JAM‐A in myelomonocytic cells (LysMcre; JAM‐Afl/fl) and the intestinal epithelium (Villin‐Cre; JAM‐Afl/fl). Consistent with a previous report, infiltration of PMN in the peritoneum in response to zymosan was decreased in JAM‐A−/− versus JAM+/+ mice. Surprisingly, in LysMcre; JAM‐A fl/fl mice, the number of PMN recruited into the peritoneum was not significantly affected by the specific loss of JAM‐A in myeloid cells compared to control littermates. Given that zymosan is a potent stimulus for peritoneal macrophages, we investigated the effect of the loss of JAM‐A on macrophage immune responses. While peritoneal macrophages from JAM‐A+/+ and JAM‐A−/− mice were similarly capable of phagocytizing zymosan, JAM‐A −/− macrophages showed significantly decreased production of the PMN chemokine CXCL1/KC that was dependent on the transcription factor NFkB. Interestingly, zymosan‐stimulated production of immune mediators from macrophages was not significantly affected by the specific loss of JAM‐A in myelomonocytic cells. These findings suggested that extrinsic factors contribute to the defective PMN migration observed in zymosan exposed JAM‐A−/− mice. We thus hypothesized that blunted peritoneal macrophage responses in JAM‐A−/− mice may be secondary to the known increase in intestinal permeability and bacterial translocation in these mice. Comparison of peritoneal macrophage subsets in total knockout and tissue selective JAM‐A deficient mice (Villin‐Cre; JAM‐Afl/fl) revealed increased numbers of poor CXCL1 producing macrophages (F4/80low) compared to high producers (F4/80high). Our results suggest that a leaky intestinal barrier and enhanced microbial translocation as observed in JAM‐A deficiency indirectly blunts neutrophil recruitment through effects on macrophage release of inflammatory mediators. Such observations highlight an important functional link between the intestinal barrier and regulation of innate immune responses.

Support or Funding Information

RO1 DK072564‐21
Microbiota‐Derived Tryptophan Metabolites Provide a Novel Pathway for Regulation of Mucosal Barrier Function
Authors: Alexeev Erica E. , Lanis Jordi M. , Kao Daniel J. , Battista Kayla D. , Kelly Caleb J. , Campbell Eric L. , Kominsky Douglas J. , Colgan Sean P.
Published: November 2023

The majority of microbial species within the human body contribute an important role to human health. Mucosal surfaces act as a primary barrier against microbial toxins and invaders, while commensal bacteria work in a dynamic and intimate interaction with the epithelium to influence host cellular and immune responses. Indole, a tryptophan metabolite produced by anaerobic bacteria, is known for its intercellular signal activity and serves as an aryl hydrocarbon receptor (AHR) ligand. AHR is a ligand‐dependent transcription factor activated by a variety of synthetic and biological molecules. The physiological function of AHR includes roles in immune regulation and mucosal barrier function. Further, IL‐10 is an anti‐inflammatory cytokine that inhibits production of numerous pro‐inflammatory mediators in various cell types. This cytokine functions through binding to the IL‐10 receptor alpha subunit (IL‐10R1). An unbiased metabolomic profile of serum from healthy and colitic mice identified significant decreases in several microbial‐derived indole derivatives (e.g. indoleacetate, indolepropionate, 5‐OH‐indoleacetate). Given that colitis is strongly associated with the induction of the epithelial IL‐10R1, we hypothesized that indole‐containing tryptophan metabolites regulate epithelial IL‐10R1 expression. Human intestinal epithelial cells were exposed to indole derivatives and examined for expression of IL‐10R1. These studies revealed a prominent induction of IL‐10R1 mRNA and protein expression following treatment with indole derivatives. Such results were explained, at least in part, by the activation of AHR. Indeed, IL‐10R1 expression was abolished when indole derivatives were administered to AHR nuclear translocator (ARNT) knockdown cells. Likewise, the AHR inhibitor, CH‐223191, significantly reduced IL‐10R1 expression in cells exposed to indole derivatives. Furthermore, ongoing work has shown a potential role for indole metabolites in alleviating disease severity in a murine model of colitis. Together, these results provide strong evidence for a novel role of tryptophan‐indole derivatives in anti‐inflammatory pathways mediated by IL‐10 signaling in intestinal epithelia. The discovery of indole metabolites as AHR ligands provides new insight to our understanding of the host‐microbial axis during inflammatory processes. These studies also identify possible avenues for incorporating the indole‐AHR pathway as novel therapies for mucosal disease.
Initial Autophagic Protection Switches to Disruption of Autophagic Flux by Lysosomal Instability during Cadmium Stress Accrual in Renal NRK‐52E Cells
Authors: Thevenod Frank , Lee Wing‐Kee , Santoyo‐Sánchez Mitzi P. , Probst Stephanie , Kerek Evan , Prenner Elmar J
Published: November 2023

The renal proximal tubule (PT) is the major target of cadmium (Cd2+) toxicity where Cd2+ causes stress and apoptosis. Autophagy is induced by cell stress, e.g. endoplasmic reticulum (ER) stress, and may contribute to cell survival or death. The role of autophagy in Cd2+‐induced nephrotoxicity remains unsettled due to contradictory results and lack of evidence for autophagic machinery damage by Cd2+.

Cd2+‐induced autophagy in the rat kidney PT cell line NRK‐52E and its role in cell death was investigated. Autophagy markers LC3‐II and p62 indicate rapid induction of autophagic flux by Cd2+ (5–10μM) after 1h, accompanied by ER stress (increased p‐PERK, p‐eIF2α, CHOP). Cd2+ exposure exceeding 3h results in p62/LC3‐II accumulation, but diminished effect of lysosomal inhibitors (bafilomycin A1, pepstatin A+E64d) on p62/LC3‐II levels, indicating decreased autophagic flux and cargo degradation. At 24h exposure, Cd2+ (5–25μM) activates intrinsic apoptotic pathways (Bax/Bcl‐2, PARP1), which are not evident earlier (≤ 6h) despite decreased cell viability by MTT assay. Autophagy inducer rapamycin (100nM) does not overcome autophagy inhibition or Cd2+‐induced cell viability loss. The autophagosome‐lysosome fusion inhibitor liensinine (5μM) increases CHOP and Bax/Bcl‐2 dependent apoptosis by low Cd2+ stress, but not by high Cd2+. Lysosomal instability by Cd2+ (5μM; 6h) is indicated by increased cellular sphingomyelin and membrane fluidity and decreased cathepsins and LAMP1.

The data suggest dual and temporal impact of Cd2+ on autophagy: Low Cd2+ stress rapidly activates autophagy counteracting damage but Cd2+ stress accrual disrupts autophagic flux and lysosomal stability, possibly resulting in lysosomal cell death.

Support or Funding Information

Center for Biomedical Training and Research (ZBAF) of the University of Witten/Herdecke (F.T.), Deutsche Forschungsgemeinschaft (TH345/11‐1 to F.T.), Max Kade Foundation (W.‐K.L.), CONACYT (Consejo Nacional de Ciencia y Tecnología, Mexico) (235537 to M.P.S.‐S.), and NSERC Discovery grant (E.J.P.)
RNA decay and quality control by the eukaryotic RNA exosome
Authors: Lima Christopher D.
Published: November 2023

RNA processing or decay by the multi‐subunit eukaryotic RNA exosome is catalyzed by two associated ribonucleases, Rrp44 (aka Dis3) and Rrp6. The activities of these enzymes are modulated and coordinated by a non‐catalytic nine‐subunit core in addition to other co‐factors that link the RNA exosome to processing complexes that deliver and prepare RNA substrates for processing and/or degradation. Results will be presented from ongoing biochemical and structural studies that are aimed at understanding how RNA exosome co‐factors coordinate the activities of the RNA exosome.

Support or Funding Information

Supported in part by a grant from the National Institutes of Health (GM118080)
Redox control of the metabolome and the aging process
Authors: Gladyshev Vadim
Published: November 2023

Biological diversity among mammals is remarkable and is reflected in their transcriptomes and metabolomes. By profiling metabolites in brain, heart, kidney, and liver tissues across mammalian species, we uncovered metabolite patterns characteristic of organs, lineages, and species longevity. Our data suggest different rates of metabolite divergence across organs and reveal patterns representing organ‐specific functions and lineage‐specific physiologies. We identified metabolites that correlated with species lifespan, some of which were previously implicated in longevity control. We similarly examined metabolite changes in several long‐lived mouse models. One process relevant to lifespan control is the methionine pathway. Reduced levels of this amino acid can extend lifespan in rodents, but whether this regimen represents a general strategy for regulating aging has been controversial. We found that methionine restriction can extend lifespan in several model organisms, and that this effect is dependent on the status of other amino acids. Methionine can also be easily oxidized, requiring a designated repair process. Overall, this study describes adjustments of the mammalian metabolome according to lifespan, phylogeny, and organ and lineage specialization, and highlights the role of redox regulation.

Support or Funding Information

National Institutes of Health
Role of hepatocyte nuclear factor 4 alpha (HNF4α) in hepatocyte regeneration
Authors: Huck Ian , Apte Udayan
Published: November 2023

Liver has remarkable capacity to regenerate upon surgical resection or chemical injury, which is critical for liver homeostasis. Recent studies have shown that HNF4α, the master regulator of hepatic differentiation, inhibits hepatocyte proliferation. However, the role of HNF4α in liver regeneration is not known. We hypothesized that hepatocytes modulate HNF4α expression and function to navigate between differentiated and proliferative states during liver regeneration. First we examined HNF4α expression and activity in liver regeneration after partial hepatectomy (PH) using three‐month‐old male C57BL/6 mice over a two‐week time course. While HNF4α mRNA expression did not change, western blot analysis showed a rapid decline in nuclear HNF4α protein levels within 1 hour post‐PH, which continued until 6 hours. Nuclear HNF4α protein levels began increasing 12 hours post‐PH but remained significantly lower than normal till 3 days post‐PH and recovered to pre‐PH levels at 5 days post‐PH. Cytoplasmic levels mimicked that nuclear expression pattern. Expression of several positive target genes of HNF4α involved in hepatocyte differentiation declined and expression of negative target genes of HNF4α involved in cell proliferation increased consistent with HNF4α protein changes. Next, we studied the role of HNF4α in liver regeneration after PH using conditional hepatocyte‐specific HNF4α knockout mice (HNF4α‐KO). Despite increased proliferative marker expression in HNF4α‐KO livers at all time points following PH, we observed 100% mortality by day 11 in the HNF4α‐KO group. qPCR analysis of HNF4α target genes 7 days post‐PH showed sustained loss of HNF4α transcriptional activity lasting until the termination phase of regeneration, indicating that hepatocytes were unable to compensate for the loss of HNF4α. This correlated with decreased hepatocyte function in HNF4α‐KO livers post‐PH as measured by serum bilirubin levels. Finally, AAV8‐mediated reexpression of HNF4α into HNF4α‐KO livers post‐PH restored HNF4α protein levels and target gene expression. HNF4α re‐expression extended survival of HNF4α‐KO mice. These data indicate that decrease in HNF4α expression and activity is required for initiation of cell cycle after PH. Furthermore, resumption of HNF4α activity is necessary for the redifferentiation of hepatocytes during termination phase of regeneration. These studies, for the first time, demonstrate that HNF4α reprogramming is a critical step in liver regeneration and could have potential therapeutic benefits.

Support or Funding Information

R01DK98414
Beta‐catenin dependent Wnt signaling promotes hepatocyte‐to‐cholangiocyte transdifferentiation
Authors: Kosar Karis , Nejak‐Bowen Kari
Published: November 2023

The Wnt/β‐catenin pathway has been identified as a critical regulator of liver development and physiology, and is involved in wide variety of liver pathologies, including atypical ductular proliferation during hepatobiliary injury. Previously, we published mice expressing a mutated non‐degradable form of β‐catenin (S45D) in liver have a significant number of hepatocytes expressing cholangiocyte markers after long‐term exposure to 0.1% 3,5‐ diethoxycarbonyl‐1,4‐dihydro‐collidine (DDC) compared to wild‐type (WT) littermates. There was also marked improvement in intrahepatic cholestasis and an upregulation of Wnt7A, ‐7B, and ‐10A in the portal triad of DDC‐fed mice. Due to these findings we hypothesize that β‐catenin dependent signaling induced hepatocyte‐to‐cholangiocyte transdifferentiation, which may alleviate some of the complications caused by cholangiopathies. To study this we exposed Mdr2 knockout (KO) mice, a murine model for chronic periductular inflammation and cholestatic liver disease, to GC‐1 diet, an analog of the T3 thyroid hormone that activates β‐catenin. We determined Mdr2 KO mice on GC‐1 diet had decreased bilirubin, liver to body weight ratios, fibrosis, and total β‐catenin compared to Mdr2 KO mice on normal diet. The ratio of non‐phosphorylated, active β‐catenin to total β‐catenin was significantly increased in Mdr2 KO mice on GC‐1 diet compared to both the Mdr2 KO and WT mice on normal diet. Surprisingly, however, there was no increase in either ductular proliferation or Sox‐9 expressing transdifferentiating hepatocytes in Mdr2 KO on GC‐1 diet, indicating the activation of an alternative pathway downstream of β‐catenin that we did not initially anticipate. Thus, we demonstrate that the protective effect of b‐catenin activation in a mouse model of cholestasis, albeit through an as‐yet unknown mechanism that will be elucidated in future studies.

Support or Funding Information

National Institute of Biomedical Imaging and Bioengineering of the National Institute of Health under Award Number T32EB0010216

UPP Academic Foundation Award 1R01DK103775
Differential Activation of Itch vs Pain C‐fibers by Mast Cell Stimulation in Mouse Skin
Authors: Jurcakova Danica , Ru Fei , Dong Xinzhong , Undem Bradley
Published: November 2023

Allergic skin reactions are often itchy, but seldom painful. We addressed the hypothesis that mast cell activation preferentially stimulates itch nerves in the mouse skin. Nerves that evoke itch in mouse skin can be identified by their expression of MrgprA3 and selective activation by chloroquine (CQ) (Cell. 2009,139:1353–65). We have developed a unique innervated isolated dorsal skin preparation in which action potential discharge in single afferent nerves terminating in the skin can be evaluated with an extracellular electrode positioned in the dorsal root ganglia. The dissected section skin includes the main arterial supply through which chemical stimuli are delivered directly to the receptive field. CQ strongly stimulated (12 ± 1Hz) only C‐fibers (0.8 ± 0.01 m/s). Among 228 C‐fibers from 138 mice, 26% were CQ and histamine sensitive; so‐called “itch C‐fibers”. Capsaicin stimulated 73% of the CQ sensitive C‐fibers. The response to CQ, but not histamine was absent in Mrgpr cluster knock out animals. The majority of capsaicin‐sensitive C‐fibers are not stimulated by CQ, but are activated by ATP or 5‐HT; a population likely comprising “pain C‐fibers”. Mice were actively sensitized to ovalbumin (OVA). OVA added to the sensitized skin activates mast cells leading to mast cell mediator release strong activation of 12 of 12 CQ‐sensitive C‐fibers (6 ± 2 Hz). Conversely OVA‐activation of mast cells did not or only very modestly activated the non‐itch, pain population of C‐fibers (1.6 ± 2 Hz, n=30). The itch fibers do not respond to 5‐HT (0 of 24 responded), so it is unlikely that 5‐HT contributes to the activation of itch nerves. CQ‐sensitive C‐fibers respond to histamine 100 μM with only 4 ± 1 Hz discharge, so it is likely that mediators in addition to histamine are involved. We have previously noted that mast cells can release MrgprC11 agonists (J Immunol. 2008, 180:2251–5). BAM8‐22, a MrgprC11 agonist, stimulated 13 of 13 CQ‐sensitive C‐fibers (12 ± 2 Hz), but not CQ‐insensitive fibers (only 3 of 29 stimulated) thereby mimicking the effect of mast cells activation. We hypothesize that mast cell mediators that stimulate Mrgpr C11 may contribute to the response.

Support or Funding Information

Supprted by NIH
A circadian zip code determines rhythmic leukocyte trafficking to tissues
Authors: He Wenyan , Kraus Kerstin , Druzd David , de Juan Alba , Ince Louise , Chen Chien‐Sin , Scheiermann Christoph
Published: November 2023

Circadian rhythms are important in aligning physiological processes to the rhythmic environment most organisms are exposed to. Recent evidence indicates the time‐of‐day to strongly influence the immune response. However, the dynamics of leukocyte migration over the course of the day are unknown. Here, we investigated an oscillatory migration behavior of key leukocyte subpopulations in murine blood, i.e. neutrophils, B cells, CD4 T cells, CD8 T cells, natural killer (NK) cells, NKT cells, eosinophils, inflammatory as well non‐inflammatory monocytes. We observed oscillations in the surface expression of adhesion molecules and chemokine receptors on leukocyte subtypes and endothelial cells of various vascular beds. To test the relevance of these oscillations, we performed function‐blocking experiments using antibodies directed against adhesion molecules, followed by homing assays. Our data indicate VCAM‐1 to be pivotal in the rhythmic recruitment of most leukocytes subsets, whereas ICAM‐1 appears to be involved most prominently in mediating rhythmic homing of T and B lymphocytes. In contrast, individual targeting of ICAM‐2, E‐selectin or P‐selectin yielded no effects on rhythmic recruitment. Furthermore, we explored the role of chemokine receptor (Gαi‐protein)‐mediated signaling in this process by pulsed pre‐incubation of leukocytes with pertussis toxin. These data additionally indicate chemokines as important regulators of rhythmic leukocyte recruitment. Together, our data demonstrate the existence of a concerted, rhythmic leukocyte‐ and tissue‐specific molecular signature in the expression of pro‐migratory factors, which allows for the recruitment of select leukocyte subsets at distinct times.

Support or Funding Information

Funded by the German Research Foundation (Emmy Noether SCHE 1645/2‐1 and SFB914 projects B09 and Z03) and the European Research Council (CIRCODE, 635872)
Sympathetic nerves regulate rhythmic leukocyte recruitment to arteries and veins
Authors: de Juan Alba , Druzd David , Zuchtriegel Gabriele , Kraus Kerstin , Reichel Christoph , Scheiermann Christoph
Published: November 2023

Recent evidence points to a critical role of the time‐of‐day in the regulation of leukocyte recruitment to tissues. Using in vivo quantitative imaging analyses across the day in inflammatory scenarios, we observed circadian (~24h) leukocyte recruitment in arteries and veins of the macro‐ and microvasculature. Interestingly however, while in arteries the number of adherent cells peaked in the morning, in veins it peaked at night. These differences were functionally associated with a vessel‐type‐specific oscillatory pattern in the expression of the adhesion molecule ICAM‐1 but not of other promigratory molecules or clock genes. ICAM‐1‐deficient mice lacked rhythmic leukocyte adhesion in both arteries and veins, which was also observed in studies using functional ICAM‐1‐blocking antibodies. Since the sympathetic nervous system (SNS) is an important orchestrator of rhythmicity in peripheral tissues, we disrupted the SNS systemically by 6‐OHDA treatment. This resulted in a lack of oscillations in both arteries and veins. To assess the functional role of direct sympathetic innervation in this process, we surgically denervated sympathetic input to vessels locally by unilaterally cutting the superior cervical ganglion. While the contralateral, nerve‐intact sides still exhibited rhythmicity in both leukocyte adhesion and ICAM‐1 expression, local denervation resulted in a loss of oscillations. Similar results were obtained by administration of beta adrenergic receptor antagonists. Interestingly, when using Ng2Cre x Bmal1flox/flox mice, in which the core clock gene Bmal1 and thus rhythmicity was selectively deleted in arteriolar Ng2+‐pericytes, an ablation of the oscillations was observed in both arteries and veins. Thus, using pharmacological, surgical and genetic approaches, our data point to an important role of arteries in regulating rhythmic leukocyte recruitment to veins.

Support or Funding Information

Funded by the German Research Foundation (Emmy Noether SCHE 1645/2‐1 and SFB914 projects B09 and Z03) and the European Research Council (CIRCODE, 635872)
Impact of Learning Approaches on Academic Performance of Preclinical Medical Students: Superficial Vs Deep
Authors: Kayali Zeina , Abdulwali Fareeha K , Peeran Ahmed A , Obad Adam S , Shareef Mohammad A
Published: November 2023

Introduction

Medical students often adopt different learning strategies and motives that guide them during studying. Preclinical students often face difficulties coping with new learning environments hence impacting their learning styles. The literature shows that studies examining the influence of surface and deep learning approaches on the academic performance of preclinical students are limited. Hence, this study aims to measure the impact of superficial and deep learning approaches on their academic performance.

Methods

The revised two‐factor version of the Study Process Questionnaire (R‐SPQ‐2F) was distributed among first, second and third year medical students at Alfaisal University. Each learning approach, surface or deep, is composed of motives and strategies. Exploratory factor analysis was conducted to validate the tool and Cronbach's alpha to assess its reliability. GPA was grouped into four groups: excellent (>3.5), good (3.0–3.5), average (2.5 – 2.9) and poor (<2.5). Regression analysis explored the prediction of academic performance by different learning styles. One‐way Analysis of Variance (ANOVA) and independent samples t‐test were used to examine the differences between learning styles, different study resources and average study time.

Results

A total of 159 students were enrolled in the study and the tool was tested to be valid and reliable. Deep learning approach predicted higher academic performance among the students (β 0.26, p=0.001). Excellent students showed higher ratings of deep learning strategies than good (p=0.025) and average students (p=0.001). Students who study for more than 8 hours daily also ranked higher in deep learning strategies than those who study for 4–6 hours (0.024) and 2–4 hours (p=0.01). Additionally, students who read books demonstrated higher self‐rating in deep motives and strategies (p=0.008 & p=0.005, respectively). Those who watch videos and utilize internet to expand medical knowledge tend to have higher ratings in deep motives with p values of 0.038 and 0.019, respectively.

Conclusion

This study demonstrates adequate validity and reliability of R‐SPQ‐2F model among the study subjects. Students with deep approach style tend to study for a longer time, use different resources and achieve a higher GPA compared to students with surface approach. This may help students with surface approach shift their learning motivations to deep motives in order to achieve a transition to deep learning approach.
Early Echocardiography experience assists in improving and retaining knowledge of Cardiac Anatomy and Physiology for medical students
Authors: Elmoselhi Adel , Eladl Mohamed , Seif Mohamed
Published: November 2023

Background

Echocardiography (Echo) has become an increasingly valuable tool in clinical patient care. Acquiring the needed skills for its use is now critical for all practitioners. Recently the medical education around the globe has been undergoing a major transformation to prioritize competency and clinical integration with basic medical sciences. Our aim is to explore the feasibility of early echocardiography experience as an effective tool to improve and sustain knowledge of cardiac anatomy and physiology to second‐year medical students.

Methods

The study was conducted on 66 medical students in a problem‐based learning (PBL) curriculum. Students were introduced to cardiac anatomy and physiology as well as the basic principles of ultrasonography before the Echo sessions. A pretest was conducted at the start of the Echo theoretical session, consisting of 10 questions related to anatomical structures and phases of the cardiac cycle. The Echo practical sessions included hands‐on experience on standardized patients with a checklist of structure and function that are commonly demonstrated in left parasternal and apical four chamber windows. An early post‐test with the same questions was repeated immediately after the Echo clinical sessions to test knowledge gained during the sessions. Another late post‐test using the same questions was conducted after five months to test the retention of the student's knowledge. Furthermore, surveys were conducted to gather students' feedback.

Results

The early post‐test outcome revealed a significant improvement in identifying the cardiac structures and function in comparison to the pre‐test. The mean of the student's scores increased from 6.58 (SD = 1.75) to 9.59 (SD = 1.04) with P value ≤ 0.0001. This significant improvement has been maintained in the late post‐test after five months with a mean of 9.2 (SD = 1.74) with insignificant correlations between post‐test and late post‐test (P Value = 0.12). Furthermore, 95% of students admitted that preceding basic knowledge is essential for identification of Echo images and 86% stated that integrating this experience is helpful in consolidating different phases of the cardiac cycle.

Conclusion

Introducing Echo spatial orientation and hands‐on experience in the early years of a medical curriculum seems to improve and enhance the retention of student's knowledge of cardiac anatomy and physiology. The Echo experience was well received and endorsed by the students and needs to be considered in new medical curricula development.

Support or Funding Information

N/A