bluuebunny/crossref_metadata_2025 huggingface dataset:
Genetic influence on xanthine oxidase activity and arginine metabolites in Hispanic children
Authors: Voruganti V. Saroja , Chittoor Geetha , Haack Karin , Laston Sandra , Vazquez‐Vidal Itzel , Mehta Nitesh R , Comuzzie Anthony G , Cole Shelley A , Butte Nancy F
Published: June 2021

Xanthine oxidase (XO) catalyzes the oxidation of xanthine and hypoxanthine in the uric acid metabolic pathway generating reactive oxygen species in the process. Inhibition of XO activity is known to reduce oxidative damage and improve endothelial function. The aim of this study was to identify the genetic basis of estimated XO activity and its relationship with endothelial function biomarkers in 752 Hispanic children of the Viva La Familia Study. Xanthine, hypoxanthine and other biomarkers were measured as part of metabolomics profiling. Xanthine oxidase activity was estimated as a product/precursor ratio. Quantitative genetic analysis was conducted using a variance components decomposition‐based method accounting for kinships. Significant heritabilities were obtained for XO activity (h2 = 0.60 ±0.08; p = 1.6 × 10−14), homocysteine (h2 = 0.82± 0.07; p = 5.2 × 10−32) and arginine metabolites; arginine (h2 = 0.47± 0.09; p = 4.6 × 10−9) and dimethylarginine (h2 = 0.24 ± 0.08; p = 0.001). Significant genetic and phenotypic correlations were found between XO activity and homocysteine (rhog = −0.21 ± 0.10; p < 0.05; rhop = −0.15 ± 0.05; p < 0.0005), arginine (rhog = 0.39 ± 0.13; p < 0.005; rhop = 0.15 ± 0.05; p < 0.0005) and dimethylarginine (rhog = −0.60 ± 0.17; p < 0.0005; rhop = −0.13 ± 0.05; p < 0.001). Genome‐wide association analysis showed significant association of XO with rs7809463 in archaelysin family metallopeptidase1 (AMZ1) and rs58632700 in a gene encoding zinc finger protein 446 (ZNF446) (p < 10−8). Follow up bivariate analysis found significant evidence of pleiotropic effects of echinoderm microtubule associated protein like 6 (EML6) on XO activity and methylarginine (p < 10−8), and suggestive evidence of AMZ1 and ZNF446 on XO activity and arginine (p < 10−7). Interestingly, L‐Arginine is known to have a protective effect against reactive oxygen species and its supplementation is believed to reduce oxidative damage. The results show that shared genetic factors influence XO activity and arginine metabolites with opposing effects in Hispanic children with important implications for diseases such as obesity, diabetes and cardiovascular disease.

Support or Funding Information

National Institutes of Health (NIH) [DK080457], and the USDA/ARS [Cooperative Agreement6250‐51000‐053]
Comparison of Resuscitation Fluids in Maintaining Mitochondrial DNA Integrity in Human Endothelial Cells
Authors: Gibson Ruby S , Barr Johnny L , Dubick Michael A , Bowman Phillip D
Published: June 2021

In our continuing evaluation of resuscitation fluids in the treatment of hemorrhage, the objective of this study was to determine if standard resuscitation fluids lead to increased damage to mitochondrial DNA (mtDNA) during periods of increasing oxidative stress as associated with shock states. Mitochondria are responsible for maintaining the cellular energy balance and are involved in the triggering of apoptosis in response to oxidative stress. The majority of mitochondrial proteins are encoded by nuclear DNA, however, mitochondria retain their own genome consisting of a circular duplex molecule of approximately 16.5 kb encoding 13 polypeptides that are exclusively involved in the intracellular ATP production by the electron transport chain (ETC). MtDNA, like the ETC, is localized at the inner side of the mitochondrial inner membrane. The ETC reduces oxygen to water in four consecutive one‐electron steps and reactive oxygen species (ROS) are produced as a by‐product of this process. These reactive molecules can be converted to H2O2 spontaneously or by superoxide dismutase resulting in free hydroxyl radicals (•OH) in the presence of iron via Fenton chemistry. ROS, in particular hydroxyl radicals, exhibit a high capacity to impair proteins, lipid membranes, DNA and RNA, which are essential components of functional mitochondria. Since, many of these resuscitation fluids lack substrates for mitochondrial production of ATP we proposed that they would enhance generation of ROS. Human umbilical vein endothelial cells (HUVEC) were treated for 3 hrs to 10 mM Hepes‐buffered Hank's solution, Ringer's solution, lactated Ringer's (LR), normal saline (NR), 5% albumin (5% Alb) and Hextend. The cells were fixed in 50% isopropanol and DNA extracted with DNAzol (MRC, Cincinnati, OH) and subjected to real‐time polymerase chain reaction (RT‐PCR) with primers designed to amplify large and small regions of mitochondrial and nuclear DNA where the extent of template amplification is inversely proportional to the lesion frequency within a given DNA sequence. Detected lesions per 10kb DNA were determined. Compared to hydrogen peroxide induced DNA damage, the tested resuscitation fluids did not produce significant alterations in the mtDNA or nuclear DNA. These data suggest that based on our previous observations of a lack on intrinsic antioxidant activity in these fluids, none would offer any advantage in influencing ROS production or mitochondrial dysfunction induced by a shock state.

Support or Funding Information

This study was funded by the US Army Medical Research and Materiel Command.
Reduced Fatty Acid Binding Capacity of Human Albumin Used in Volume Resuscitation May Potentially Increase Hemolysis in Hypovolemic Shock
Authors: Penn Alexander Hayes , Dubick Michael , Filho Ivo Torres
Published: June 2021

The choice of resuscitation fluid for hypovolemic trauma patients in the absence of fresh whole blood remains controversial. Albumin solutions have shown promise in animal studies but meta‐analysis suggests they have had little to no benefit in human trials. This contradiction is currently unresolved. Unbound (cytotoxic) free fatty acids (FFAs) are generated in the intestine during ischemia and enter the mesenteric lymph, from where they likely enter the circulation, resulting in organ damage, hemolysis, coagulopathy and further ischemia. We hypothesize that resuscitation fluids with higher available binding capacities for FFAs may help prevent these events in shock. The objectives of this in‐vitro study were to determine: 1) if commercial sources of pharmaceutical‐grade human albumin contain FFAs, 2) if these FFAs result in a decreased available fatty acid binding capacity, and 3) if binding capacity will affect the level of hemolysis that occurs in whole blood when exogenous FFA is introduced in levels similar to those in shock. To achieve these objectives, we first measured FFA concentrations in human albumin via a kit. We then developed a new assay for measuring fatty acid binding capacity using exogenous oleic acid (OA), glass fiber filtration, and the FFA kit to measure binding capacity in both the human albumin and FFA‐free bovine albumin. Lastly, we mixed fresh human whole blood 4:1 with PBS, human albumin, FFA‐free bovine serum albumin (BSA), or BSA that was first saturated to its binding capacity with OA and measured the amount of hemoglobin released into the plasma after 30 minute incubation with 0 to 5 mM OA. We found that pharmaceutical‐grade 5% human albumin contained 4.6±0.3 mM FFA (N=5 lots), ~7x the physiologic concentration, leaving an available binding capacity of 4.8±0.6 mM out of a total binding capacity of 9.4±0.8 mM. This represented a significant loss of binding capacity (p<0.0001) compared to the total. FFA‐free BSA (N=5 replicates) had significantly greater available binding capacity of 7.0±1.4 mM (p=0.02) compared to the pharmaceutical‐grade human albumin. In 4:1 mixtures of blood to PBS or albumin, hemolysis was reduced after addition of all concentrations of exogenous OA with 25% FFA‐free compared to 25% pre‐saturated BSA (p<0.04). Likewise, 0.5 and 5 mM exogenous OA caused less hemolysis when blood was pre‐mixed with 5% FFA‐free BSA than with 5% human albumin (p<0.01). These preliminary findings suggest that the failure of albumin in human trials to live up to the promise of animal studies may be due to steps in the manufacturing process that reduce the FFA binding capacity of human albumin.

Support or Funding Information

Supported by US Army Medical Research & Materiel Command.
Perinatal Exposure to Brominated Flame Retardants Reduces Social Recognition in Adult Male Mice Concomitant with Reduced Amygdalar Avp Expression
Authors: Valdez Matt , Gonzalez Gwen , Gutierrez Roberto , Valdez Joseph , Lindner John , McLaughlin Thu , Leon‐Olea Martha , Curras‐Collazo Margarita
Published: June 2021

Polybrominated diphenyl ethers (PBDEs) are flame‐retardants associated with impaired cognitive and behavioral deficits in children. PBDEs are endocrine‐disrupting chemicals that also have neurotoxicological actions. In humans, PBDEs have been associated with disrupted attention, hyperactivity and lower IQ scores. This study evaluates behaviors relevant to autism spectrum disorder (ASD) and examines correlative changes in forebrain expression of genes after perinatal exposure to the industrial PBDE mixture, DE‐71. Genes for vasopressin (Avp) and oxytocin signaling (Oxtr) serve as markers for social circuits. Pituitary adenylate cyclase activating polypeptide, PACAP (Adcyap1r1), and its receptor, PAC1R (Adcyap1r1) are critical for neurodevelopment. Female C57BL/6 dams were dosed for 10 weeks (pre‐conception: 4 weeks; gestation: 3 weeks; lactation: 3 weeks). Dosing consisted of low dose (0.1 mg/kg/d), high dose (0.4 mg/kg/d), or corn oil vehicle (control) via ingestion of infused corn flakes. Exposed progeny were subjected to behavioral testing: Sociability (PND 21), Open Field (PND 30–40), Marble Burying (PND 40–50), Social Recognition (PND 55–60), and Elevated Plus Maze (PND 60–65). High dose DE‐71 treatment produced a sex‐independent decrease in repetitive behavior (F2,54=5.96, p<0.005). In males, high dose DE‐71 treatment produced hyperactivity (p<0.05). The social recognition test revealed significant preference for a novel conspecific in control males (F1,23=7.38, p<0.05) and females (F1,22=18.05, p<0.0005) but not in any DE‐71 groups. The latter could not be attributed to reduced sociability. Plasma showed reduced (p<0.05) and increased (p<0.05) plasma arginine‐vasopressin in low and high dose males, respectively. qPCR analysis revealed an apparent decrease in Avp gene expression in lateral septum and medial amygdala of low dose DE‐71 males compared to controls. In contrast, the supraoptic nucleus displayed significantly increased Avp expression in low dose DE‐71 mice (p=0.243). These mice also showed an apparent decrease in Oxtr, Adcyap1 and Adcyap1r1 expression in amygdala and in Adcyap1r1 in cingulate cortex compared to controls. Preliminary histological analysis has revealed a significant transient increase in cell density of the cingulate cortex at PND 7 (p=0.012) in high dose males compared to controls. By PND 71, the cell density of the cingulate cortex becomes significantly decreased (p<0.001) in high dose males compared to controls (n=2–3/group). We conclude that developmental PBDEs produce ASD‐relevant neurodevelopmental and behavioral abnormalities that maybe due, in part, to changes in vasopressinergic and PACAPergic circuits.

Support or Funding Information

Support: APS (G.G.), MARC (G.G., J.V, R.G.), UCMEXUS (M.C‐C., M.L‐O.), NSF (M.V.) and Sigma Xi (G.G., M.V.).
Changes in Protein Turnover, Hormonal Status, and Body Composition during Physiologically Demanding Military Training
Authors: Pasiakos Stefan M , Berryman Claire E , Margolis Lee M , Sepowitz John J , McClung Holly L , Lieberman Harris R , McClung James P , Ferrando Arny A
Published: June 2021

Physiological stress imposed during short‐term military training is often severe and intentional; producing a catabolic state and loss of lean body mass (LBM). The extent to which this occurs and metabolic responses that may contribute to the loss of LBM are not well characterized. This study examined body composition, whole‐body protein turnover (15N‐alanine), testosterone and cortisol before (BASELINE) and after (POST) 7 days of Survival, Evasion, Resistance, and Escape (SERE) School in 68 male US Marines (mean ± SD, 25 ± 2 y, 83.7 ± 9.1 kg). Total body mass (TBM) was 7% lower (−2.7 ± 1.4 kg fat and −3.1 ± 1.7 kg LBM), proteolysis 28% higher and net protein balance 132% lower POST‐SERE (P < 0.05) compared to BASELINE. Decrements in testosterone POST‐SERE (−42%; −221 ± 136 ng/dL; P < 0.05) were inversely related with increased proteolysis (r = −0.30; P < 0.05) and protein synthesis (r = −0.28; P < 0.05). Percent changes in testosterone were associated with percent loss of TBM (r = 0.37; P < 0.05) and LBM (r = 0.31; P < 0.05). Cortisol changes during SERE were associated with changes in protein synthesis (r = 0.26; P < 0.05). Percent change in cortisol was inversely associated with percent change in fat mass (r = −0.34, P < 0.05) and tended to predict the percent change in LBM (r = 0.22; P = 0.07). The physiological stress during SERE elicits a hypogonadal state and catabolism of LBM. Decrements in testosterone were quantitatively associated with the increase in proteolysis and loss of LBM.

Support or Funding Information

Supported by USAMRMC
Assessment of home food environment among healthy weight and overweight/obese school‐age children
Authors: Nepper Martha J , Chai Weiwen
Published: June 2021

Objective

To compare the differences in the availability and visibility of home healthy and unhealthy foods and family meal frequency and quality between healthy weight and overweight/obese children.

Methods

Forty‐two overweight/obese and 40 healthy weight children (9.8±2.6 years) and their parents participated in the study. In‐home assessment for food availability and visibility was conducted and children's weight and height were measured by researchers. Self‐report survey regarding demographics and family meals were completed by the parents. Logistic regression analysis was used to examine associations of demographic characteristics, home food availability and visibility and family meal variables with children's weight status.

Results

The availability of home total unhealthy foods was significantly and inversely associated with overweight/obesity among children. The association was not significant after adjusting for single‐parent household and parent working status. Lower visibility of home total unhealthy foods (OR=0.60, 95% CI=0.39–0.92, P=0.02) and lower frequency of family meals (OR=0.10, 95% CI=0.01–0.96, P=0.02) were significantly associated with children being overweight/obese after adjustment for covariates. Parents who worked full‐time (OR=4.14, 95% CI=1.62–10.54, P=0.008), single parent household (OR=5.25, 95% CI=1.72–16.07, P=0.002), and older parents (OR=1.11, 95% CI=1.03–1.20, P=0.004) were positively associated with overweight/obesity of children.

Conclusion

Helping and educating parents to create a healthier home food environment is important for both overweight/obese and healthy weight children.

Support or Funding Information

This research was supported by the University of Nebraska‐Lincoln Research Council Faculty Seed grant and in part by Agriculture and Food Research Initiative (AFRI) Grant 2011‐67002‐30202 from the U.S. Department of Agriculture (USDA) National Institute of Food and Agriculture, Childhood Obesity Prevention: Transdisciplinary Graduate Education and Training in Nutrition and Family Sciences or Child Development or Related Fields to Prevent Childhood Obesity.
Heat Stress Causes Autophagic Stalling In Oxidative Skeletal Muscle
Authors: Brownstein Alexandra , Summers Corey , Ganesan Shanthi , Hale Benjamin , Pearce Sarah , Gabler Nicholas , Ross Jason W , Rhoads Robert P. , Baumgard Lance H. , Selsby Joshua T.
Published: June 2021

Heat stress (HS) poses a threat to human and animal health and well being, and serves to undermine agricultural economics and food security. Yet, little is known about underlying pathological mechanisms caused by HS, which impedes the development of effective mitigation strategies. To address this critical need we have been working to elucidate HS‐mediated dysfunction in skeletal muscle and have implicated the mitochondria as likely contributors to cellular pathology. Given the possibility of mitochondrial dysfunction we hypothesized that cells would respond to HS by clearing damaged mitochondria via autophagy. To test this hypothesis gilts (female pigs; 35±4 kg; n=5–6/group) were housed in thermoneutral (TN; 20 ± 1°C) or HS (35°C) conditions for one (HS1) or three days (HS3). Relative protein abundance of markers of autophagosome membrane nucleation, Phosphatidylinositide 3‐kinases class III, Beclin‐1, elongation (Autophagy‐related protein 16L1), and maturation of autophagosomes (Microtubule‐associated protein A/B‐light chain 3 II) was increased (p<0.05) following 1 and 3 days of HS. Conversely, sequestosome‐1 (p62), a selective autophagy receptor consumed by autophagy and inversely associated with autophagy flux, was not detected in TN pigs, dramatically increased following 1 day of HS, and decreased by 3.4‐fold (but was still readily detectable) following 3 days of HS compared to 1 day of HS (p<0.05). Gene expression of p62 was similar between groups. In addition, mitophagy marker, BCL2/Adenovirus E1B 19kDa interacting protein 3‐like (Bnip3L) was decreased 2.5‐fold (p<0.05) after 1 and 3 days of HS and PTEN‐induced putative kinase 1 (PINK1) was decreased 7.0‐fold following 1 day of HS (p<0.05) and numerically by 2.4‐fold following 3 days of HS. Together, these data suggest that HS drives autophagy but this process stalls resulting in the accumulation of autophagosomes, suppressed mitophagy, and the preservation of dysfunctional mitochondria

Support or Funding Information

Supported by USDA grants 2014‐67015‐21627 and 2011‐67003‐30007.
Heat Stress‐induced Insulin Sensitivity in Oxidative Skeletal Muscle
Authors: Ganesan Shanthi , Pearce Sarah , Summers Corey , Gabler Nicholas , Valentine Rudy , Baumgard Lance , Rhoads Robert , Selsby Joshua
Published: June 2021

Heat‐related complications continue to be a major health concern for humans and animals and lead to potentially life‐threatening conditions ranging from heat exhaustion, heat stroke and even death. Heat stress have been shown to alter metabolic and energetic parameters in mammals and may alter glucose metabolism and insulin sensitivity. Therefore, the purpose of this investigation was to determine the extent to which short‐term HS altered markers of insulin signaling in oxidative skeletal muscle. To address this, crossbred gilts (n=8/group) were assigned to thermoneutral (TN; 24° C), HS (37° C), or pair‐fed to HS and kept under TN conditions (PFTN) groups for 12 hours. Following treatment, animals were euthanized and the semitendinosus red (STR) was recovered. HS‐induced changes were not caused by reduced feed intake as TN and PFTN were similar for nearly all measures. Twelve hours of HS increased insulin receptor protein abundance by 54% (p<0.05) and phosphorylated insulin receptor substrate (pIRS1S307) by 56% (p<0.05) compared to TN. Phosphorylation of IRS1 at this site is a negative regulator of insulin signaling. Relative protein abundance and phosphorylation of phosphoinositide 3‐kinase (PI3K) and 3‐phosphoinositide dependent protein kinase‐1 (PDK1) were similar between groups. Phosphorylation of protein kinase B (pAKTs473) was decreased by 53% (p<0.05) in HS compared to TN, which indicates that insulin signaling was suppressed through pIRS1. Conversely, HS increased phosphorylation of protein kinase C (PKC)λ/ζ and pPKCδ protein abundance by 128% and 205%, respectively, compared to TN which are known to stimulate pIRS1S307. Sarcolemma glucose transporter‐4 (Glut4) was decreased by 47% (p<0.05) in HS group compared to TN suggesting reduced glucose uptake. These data suggest that HS induces insulin sensititvity by disturbing insulin signaling via an AKT‐mediated pathway.

Support or Funding Information

This work is supported by USDA grants 2014‐67015‐21627 and 2011‐67003‐30007
Impaired Mitochondrial Clearance Contributes to Heat Stress‐mediated Muscle Dysfunction
Authors: Ganesan Shanthi , Summers Corey , Pearce Sarah , Gabler Nicholas , Valentine Rudy , Baumgard Lance , Rhoads Robert , Selsby Joshua
Published: June 2021

Prolonged exposure to high temperatures can result in heat stress (HS), which is characterized by morbidities and mortality in humans and animals and appears to cause organ‐specific injury and dysfunction. We have previously found increased free radical injury in skeletal muscle following 24 hours of HS. The purpose of this investigation was to determine the extent to which short‐term HS caused muscle dysfunction in oxidative skeletal muscle. We hypothesized that in oxidative muscle 12 hours of HS would lead to free radical injury, apoptosis, autophagy, and decreased mitochondrial content compared to muscle from thermoneutral (TN) animals. To address this hypothesis, crossbred gilts (n=8/group) were assigned to three groups: TN (24° C), HS (37° C), or pair‐fed to heat stress (PFTN; 24° C) for 12 hours. Following treatment, animals were euthanized and the semitendinosus red (STR) was recovered. TN and PFTN were similar for nearly all measures indicating changes caused by HS were not due to reduced feed intake but due to the hyperthermic environment. The relative abundance of malondialdehyde modified proteins was increased by 36% in HS (p<0.05) compared to TN, which was matched by an inadequate antioxidant response including a 48% and 170% increase in superoxide dismutase (SOD) I and II relative protein abundance, respectively, and a 26% increase in total SOD activity but a 40% reduction in catalase activity in HS compared to TN. Further, HS caused a 103% increase in caspase 3 activation and tended to increase BCL2‐Associated X Protein by 40% (p<0.06) and apoptotic protease activating factor 1 by 66% (p<0.06) indicating increased apoptotic signaling. Despite increased initiation, autophagy appeared to be inhibited by HS as the microtubule‐associated protein A/B‐light chain 3 II/I ratio was decreased by 53% and sequestosome‐1(p62) protein abundance was increased by 40% in HS compared to TN. Markers of mitochondrial content cytochrome c, cytochrome c oxidase IV, voltage dependent anion channel, pyruvate dehydrogenase and prohibitins 1 were increased in HS compared to TN. Increased mitochondrial content is likely reflective of a combination of increased accumulation of damaged mitochondria that fail to be cleared via autophagy and mitochondrial biogenesis. We postulate that in oxidative skeletal muscle HS‐mediated mitochondrial dysfunction is allowed to persist due to a failure of autophagy to clear damaged mitochondria, which results in increased oxidative stress and apoptotic signaling. These data demonstrate widespread aberrant intracellular signaling caused by HS that implicate impaired autophagy as a key feature in HS‐mediated muscle dysfunction

Support or Funding Information

This work supported by USDA grants 2014‐67015‐21627 and 2011‐67003‐30007.
Frank‐Starling Responses during Heat Stress in Healthy Aged Humans
Authors: Gagnon Daniel , Romero Steven A , Ngo Hai , Sarma Satyam , Cornwell William K , Poh Paula YS , Stoller Douglas , Levine Benjamin D , Crandall Craig G
Published: June 2021

Healthy aging markedly affects the cardiovascular adjustments to heat stress in humans. In particular, the increase in cardiac output can be attenuated by 50% in older individuals, which has previously been attributed to an inability of the aged to maintain stroke volume when cardiac filling pressures are reduced. In young individuals, a leftward shift in the operating point of the Frank‐Starling response is an important adjustment by which stroke volume is maintained during heat stress. It remains unknown how healthy aging affects the Frank‐Starling response during heat stress. We therefore tested the hypothesis that a leftward shift in the operating point of the Frank‐Starling response is absent in older individuals during whole‐body passive heat stress. Right heart catheterization was performed in 10 aged (A: 70 ± 3 y, 166 ± 9 cm, 68 ± 8 kg, 4 males/6 females) and 12 young (Y: 26 ± 5 y, 165 ± 6 cm, 64 ± 10 kg, 6 males/6 females) healthy individuals to measure pulmonary capillary wedge pressure, cardiac output (thermodilution) and pulmonary artery blood temperature. Heart rate was measured using an electrocardiogram and used to calculate stroke volume. These measurements were obtained at normothermic baseline, as well as at an increase in pulmonary artery blood temperature of 1.5°C achieved through whole‐body passive heat stress. Cardiac output increased in both groups during heat stress (A: 4.5 ± 0.7 to 7.0 ± 0.9 L/min vs. Y: 5.0 ± 0.7 to 9.5 ± 1.1 L/min), the magnitude of which was lower in the aged (P<0.01). Heart rate was similar between groups during heat stress (A: 91 ± 11 vs. Y: 98 ± 14 beats/min, p=0.17) however, the increase from baseline was attenuated in the aged (A: +29 ± 8 vs. Y: +41 ± 9 beats/min, P<0.01). Indicative of a leftward shift in the operating point of the Frank‐Starling response, stroke volume was maintained during heat stress in both groups (A: 74 ± 13 to 78 ± 14 mL/min vs. Y: 89 ± 23 to 99 ± 21 mL/min) despite reductions in pulmonary capillary wedge pressure (A: 9.0 ± 1.9 to 6.6 ± 1.4 mmHg vs. Y: 10.6 ± 1.6 to 6.6 ± 2.0 mmHg). Overall, these results suggest that stroke volume is maintained during whole‐body passive heat stress in healthy aged humans due to an appropriate leftward shift in the Frank‐Starling response. Thus, attenuated increases in cardiac output in the aged while heat stressed are primarily due to attenuated increases in heart rate.

Support or Funding Information

Funded by the National Institutes of Health (GM‐068865), Department of Defense (W81XWH‐12‐1‐0152) and ACSM Foundation Research Endowment.
Inactivity‐induced phrenic motor facilitation is initiated though TNF receptor 2 and impaired by TNF receptor 1
Authors: Braegelmann Kendra , Fischer Stephanie , Agbeh Abiye , Baker Tracy
Published: June 2021

Prolonged reductions in synaptic inputs to phrenic motor neurons results in the release of spinal, soluble TNFα (sTNFα) to initiate a long lasting increase in phrenic burst amplitude termed inactivity‐induced phrenic motor facilitation (iPMF). TNFα can activate TNF‐R1 or TNF‐R2, which are two distinct cell surface receptors that tend to initiate discrete signaling cascades. TNF‐R1 mediates the majority of TNFα activity; therefore, we hypothesized sTNFα released within or near the phrenic motor pool in response to reduced activity binds to TNF‐R1 on phrenic motor neurons to initiate iPMF. To test this hypothesis, Harlan Sprague Dawley rats were injected with small interfering RNAs directed against TNF‐R1 or TNF‐R2 in the interpleural space (100pg/side) to selectively reduce TNF‐R1 or TNF‐R2 in phrenic motor neurons. After 3 daily injections of siRNAs, iPMF was measured in anesthetized and ventilated rats for 60min following a prolonged (30min) neural apnea. As expected, control rats receiving injections of non‐targeting siRNA expressed an increase in phrenic burst amplitude for up to 60min following neural apnea (48±15,%baseline), indicating iPMF. Contrary to our hypothesis, knockdown of TNF‐R1 in phrenic motor neurons led to an enhanced iPMF following prolonged neural apnea (80±18,%baseline), whereas knockdown of TNF‐R2 completely abolished the expression of iPMF 60min post neural apnea (−8±5,%baseline,). Collectively, these data suggest that TNF‐R2 signaling in phrenic motor neurons is necessary for iPMF and that TNF‐R1 signaling may initiate mechanisms within phrenic motor neurons that oppose iPMF. Thus, both TNFα receptors may contribute to sculpting phrenic burst amplitude in response to a reduction in respiratory‐related neural inputs.

Support or Funding Information

NIH‐HL105501 and DoD SC120226
Discharge of Mid‐Cervical Neurons Following Cervical Spinal Cord Injury
Authors: Patel Shreya , Streeter Kristi , Reier Paul , Baekey David , Fuller David
Published: June 2021

Prior anatomical and neurophysiologic data suggest that mid‐cervical interneurons may contribute to the control of phrenic motor output after cervical spinal cord injury (cSCI). To explore this possibility, we recorded spinal interneuron bursting patterns and phrenic motor output in rats with chronic cSCI. Twelve weeks following lateral C2 hemi‐section (C2Hx), bilateral phrenic nerve activity was recorded in urethane anesthetized and mechanically ventilated rats (N=5). A multi‐electrode recording array was bilaterally inserted at C4 to enable simultaneous recording of mid‐cervical neuron discharge ipsilateral and contralateral to C2Hx. In this initial evaluation, neuronal discharge patterns were classified as tonic (i.e., active across the entire respiratory cycle) or phasic (i.e., active during only the inspiratory or expiratory phases). Spike triggered averaging of phrenic nerve activity based on discharge of recorded cells was used to identify phrenic motoneurons, all other cells were classified as interneurons for this analysis. Only interneurons (versus motoneurons) were identified in recordings ipsilateral to C2Hx, and these cells primarily burst in a tonic firing pattern (tonic: 91% and phasic: 9% of ipsilateral cells). In contrast, on the contralateral side phrenic motoneurons were readily recorded (26% of contralateral cells) and a greater proportion of phasic interneurons were detected (phasic: 26% and tonic: 48% of contralateral cells). Spike triggered averaging produced no evidence of direct (monosynaptic) coupling between interneurons and either the contra‐ or ipsilateral phrenic motoneuron pool. These preliminary findings confirm altered propriospinal neuron discharge in the ipsilateral vs. contralateral cord immediately caudal to chronic C2Hx injury. The precise role of mid‐cervical propriospinal neurons in respiratory recovery after cSCI remains unclear. Since we did not detect cells that were directly antecedent to the phrenic pool, our data suggest that following cSCI, “connectivity” between respiratory‐modulated spinal interneurons and the phrenic nucleus may involve polysynaptic pathways.

Support or Funding Information

Funding: APS Undergraduate Summer Research Fellowship (SP), 1R01NS080180‐01A1 (DF), 1F32NS095620‐01 (KS), and Department of Defense W81XWH‐14‐1‐0625 (PR).
Advancing Systematic Review Methodology to Better Inform Food and Nutrition Policy
Authors: Psota Tricia L. , Spill Maureen , Spahn Joanne , Gungor Darcy , Johns Kirsten , Nadaud Perrine , Wong Yat Ping , Stoody Eve Essery , Rihane Colette
Published: June 2021

Systematic reviews critically assess research studies relevant to a particular research question. The USDA's Nutrition Evidence Library (NEL) utilizes state‐of‐the‐art methodology to conduct systematic reviews evaluating scientific evidence related to food and nutrition to answer questions that inform federal policies and programs, such as the Dietary Guidelines for Americans. The NEL has continually advanced its process to stay at the forefront of the systematic review field. However, the key steps of the NEL process remain the same, including: 1) developing systematic review questions; 2) searching, screening, and sorting the literature; 3) extracting data and assessing the risk of bias of the research; 4) describing and synthesizing the evidence; 5) developing conclusion statements and grading the evidence; and 6) identifying research recommendations. Advancements to the NEL process include: 1) integrating broader input to develop, refine, and prioritize systematic review questions; 2) incorporating key definitions and confounders into the analytic framework, a visual representation of the systematic review project; 3) developing a standard inclusion/exclusion template to promote consistency across reviews and expanding the inclusion/exclusion criteria list; 4) conducting a duplication assessment to determine if other high‐quality systematic reviews exist which answer the prioritized questions; 5) combining title and abstract screening; 6) integrating dual‐screening of the literature; 7) revising the quality assessment tool to be tailored by study design and to focus on internal validity assessment, opposed to study quality; 8) revising the NEL grading rubric used to grade the totality of the evidence; 9) integrating a step to define research recommendations as a component of the evidence review, opposed to after the process; and 10) creating technical abstracts and plain‐language summaries that summarize the completed systematic reviews. In addition to process improvements, the NEL has enriched the training of experts with whom it works to streamline the conducting of systematic reviews. Also, the NEL has expanded its website by including additional systematic review materials, which increases the transparency of the process and makes nutrition research publicly available. NEL systematic reviews will be used to illustrate these improvements and to facilitate discussions which can contribute to advancing systematic review methodology for the field of nutrition science. Furthermore, future directions for advancement, such as leveraging research recommendations better, will be explored.

Support or Funding Information

USDA
Children Are Eating Savory Snack Foods: What We Eat in America, NHANES 2011–2012
Authors: Rhodes Donna G , Clemens John C , Hymes Melanie A , Moshfegh Alanna J
Published: June 2021

The savory snack market is over $28 billion yearly and advertising often targets children; however, data are limited on the consumption of such products among children. This research examined the percentage of U.S. children who consumed chips, flavored snacks, popcorn and pretzels (referred to here as savory snack foods) on a given day during 2011–2012. The studysample included nationally representative data from individuals aged 2–19 years(n=3132), participating in What We Eat in America (WWEIA), NHANES2011–2012. Dietary intake data, obtained from an in‐person 24‐hour recall, were collected using the interviewer‐administered 5‐step USDA Automated Multiple‐Pass Method. The WWEIA Food Categories were used to estimate the percentages of children who consumed potato chips, tortilla/corn/otherchips, popcorn, and pretzels/snack mix at least once, any time during the intake day. Overall, 43.9 ± 1.6% (mean ±SE) of children consumed savory snack foods on any given day. Among race/ethnicity groups, a greater percentage of non‐Hispanic black children (50.4± 2.9%) consumed savory snack foods on a day compared to non‐Hispanic white (44.0± 2.5%), Hispanic (42.5 ± 2.2%) or non‐Hispanic Asian (40.1 ± 3.3%) children. By age group, the percentage consuming was40.5% (2–5 yr), 47.8% (6–11 yr) and 42.8% (12–19 yr). With respect to amount consumed, mean intake of savory snack foods among consumers increased with age: 27 ± 3g, 38 ± 3g and 48 ± 3g among children2–5 yr, 6–11 yr and 12–19 yr, respectively. Overall, on the day consumed, they contributed 10% of total energy and7% of total sodium; mean energy was 199 kcal and mean sodium was 230mg. The results of this research highlight the wide‐spread consumption of savory snack foods by children of all ages and race/ethnic backgrounds.

Support or Funding Information

USDA, ARS
Development and Evaluation of the Transdisciplinary Obesity Prevention Research Sciences Program
Authors: Teegarden Dorothy , Bowers Jill , Hammons Amber , Destefano Lizanne , Garcia Gabriella , Koester Brenda , Parker Loran , Childress Amy , Fiese Barbara
Published: June 2021

The Transdisciplinary Obesity Prevention Research Sciences Program (TOPRS) is an applied research methods curriculum for undergraduates on the causes and consequences of childhood obesity that has been developed by a multidisciplinary team from the University of Illinois at Urbana‐Champaign, Purdue University, and California State University‐Fresno. To address the need of the contemporary workforce the course has been developed to integrate transdisciplinary approaches 1) to knowledge about the causes, consequences and correlates of obesity from an ecological framework from cell to society, 2) hands‐on research experience, and 3) a professional development series. The program development was an iterative process of integrating rigorous evaluation results with curriculum design. The class format is a “flipped” version with students viewing 10 minute lectures and class time spent in interactive discussions and activities focusing on research design and methodologies. The lectures are presented by national experts in nutritional sciences, animal science, child development, family science, early care and education, community health, and public health. The professional development activities reinforce leadership and team building skills. The pilot evaluation revealed students’ (N = 53) increased knowledge related to obesity causes and consequences. Students increased in their research‐related abilities and reported having a better understanding of the research process. Furthermore, students increased in self‐confidence in their research skills and their intention to choose a research‐oriented career by the end of the program. Finally, the program strengthened students’ understanding of the importance of collaboration and teamwork. Additionally, the program helped students understand the importance of interdisciplinary approaches by increasing appreciation of the complexity of obesity prevention. In conclusion, the innovative curriculum development will be available for use, and also represents a model for integrative, interdisciplinary program development.

Support or Funding Information

This material is based upon work that is supported by the National Institute of Food and Agriculture, U.S. Department of Agriculture.
Nutrimetabolomics fingerprinting to identify biomarkers of whole grain wheat intake
Authors: Sang Shengmin , Zhu Yingdong , Sha Wei , Wang Pei
Published: June 2021

Increased whole grain (WG) consumption has been inversely associated with the risk for developing some diet‐related disorders, such as type 2 diabetes, obesity, cancer, and cardiovascular diseases. Many epidemiological studies, however, have failed to generate consistent results on this topic due to a lack of accurate tools to assess dietary intake of whole grains. Nutritional epidemiology research needs more reliable and quantitative methods for measuring dietary intake of specific foods. Using a targeted LC/MS based metabolomic approach, we established the metabolic fingerprints of the major phytochemicals in WG wheat in human urine including metabolites of alkylresorcinols, benzoxazinoids, lignans, and phytosterols, as well as microbial‐derived metabolites. Using a non‐targeted LC/MS based global metabolomic approach, we investigated the impact of WG wheat bread intake on the endogenous urinary metabolome. Metabolic profiling revealed a number of differences relating to the consumption of WG wheat bread vs. refined grain wheat bread, such as 3,5‐dihydroxybenzoic acid, 2‐aminophenol sulfate, ferulic acid 4‐sulfate, 2‐acetamidophenol sulfate, and cysteine. Using the targeted and non‐targeted nutrimetabolomic approaches, we are able to develop multiple metabolites as the potential biomerkers of WG wheat intake.

Support or Funding Information

USDA‐NIFA grant 2012‐38821‐20012 to S. Sang
Physical Activity Level, Modeling, and Importance to Mothers of Preschool Children
Authors: Martin‐Biggers Jennifer T , Quick Virginia , Worobey John , Byrd‐Bredbenner Carol
Published: June 2021

Physical activity (PA) is an important component of obesity prevention for children and adults. This study compared maternal and child PA levels, maternal modeling of PA, and importance mothers place on PA for herself and her child to maternal and child BMI categories. Survey participants (n=550) were 72% white and 37% had a college degree or higher. Categorizing mothers by weight status indicated that 26%, 23%, 21%, and 30%, were underweight/low normal weight (BMI<22.5), high normal weight (BMI≥22.5 to <25), overweight (BMI ≥25 to <30), or obese (BMI≥30), respectively. Mean maternal PA levels were low (15.44±9.98SD; 0–42 point scale), with obese mothers (n=165) scoring significantly lower than underweight/low normal weight mothers (14.20±9.91SD vs 17.17±10.47SD, p=0.018). Using a 5‐point 2‐item scale, mothers indicated they felt modeling PA behaviors to their children was important and modeled PA 3.08±1.22SD days a week in the past month. Compared to both categories of normal weight mothers, obese mothers placed significantly less importance on modeling PA behaviors (3.91±0.89SD vs 4.24±0.84SD and 4.32±0.69SD, p<0.001) to their children and spent significantly fewer days a week in the last month modeling PA behaviors (2.80±1.23SD vs 3.35±1.14SD and 3.21±1.25SD, p<0.001). Mothers placed less importance on PA for themselves than for their children (mean 3.49±0.97SD vs 3.83±0.87SD, on a 5‐point scale). Obese mothers placed significantly less importance on PA for themselves than mothers in all other weight categories. PA level of participants’ children was moderate (26.19±11.42SD, 0–42 point scale). No significant differences were found among child BMI categories and child PA levels. The importance mothers placed on modeling PA and importance placed on PA for their children did not differ among child BMI weight categories. Mothers of overweight children reported modeling PA behaviors significantly more days per week than mothers with middle normal weight (25th to 74th percentile) children (3.41±1.27SD vs 2.85±1.17SD). The importance mothers placed on their own PA was lower among mothers of underweight children and tended to increase as child BMI increased. Findings suggest that mothers of obese children may be more sensitive to the importance of modeling PA and its importance to health. Encouragement of all parents to promote PA and model PA behaviors to children is important for obesity prevention.

Support or Funding Information

USDA NIFA #2011‐68001‐30170
Mechanical Signaling in Epithelial‐Mesenchymal Transition
Authors: Engler Adam , Yang Jing , Ondeck Matthew , Wei Spencer
Published: June 2021

Mammary epithelial cells (MECs) are classically known to respond to differences in static extracellular matrix (ECM) stiffness and resemble a mesenchymal, more malignant state lacking any polarized acinar structure on stiffer ECM, i.e. Epithelial‐Mesenchymal Transition (EMT). While this is akin to the stiff mammary tumors that one can detect with manual palpation, breast cancer fibrosis is a dynamic process that results in matrix stiffness increasing from normal ~150 to stiff 5000 Pascals (Pa) over months to years as a result of enhanced collagen expression and lysyloxidase crosslinking. I will present data on our new understanding of a molecular mechanism used by MECs interpret stiffness changes in which the EMT marker Twist changes its nuclear localization based on binding to GTPase Activating Protein (SH3 Domain) Binding Protein 2 (G3BP2). G3BP2 suppression drives tumorigenesis in mice whereas higher expression is correlated with recurrence free survival in women (Wei et al, Nature Cell Bio 2015). I will also describe our efforts to more accurately mimic the onset of tumor‐associated fibrosis using dynamic methacrylated‐hyaluronic acid (MeHA) hydrogels, whose stiffness that can be modulated from “normal” 100 Pa to “malignant” 5000 Pa, utilizing a two step polymerization process. Mammary epithelial cells cultured on 100 Pa MeHA hydrogels form and remain as polarized acini and then undergo EMT upon matrix stiffening to 5000 Pa. However, the degree of matrix stiffening and culture time prior to stiffening play a large role in acini transformation; increasing culture time on soft matrix prior to stiffening from 6 to 10 days delayed the onset malignant transformation by as much as 35%. EMT is independent of acinus size, cell density, and is not cell intrinsic, suggesting that individual acini respond to the stiffness but their epigenetic state and structure can protect a subset of acini from stiffening. These data indicates a more complex interplay of intrinsic ECM cues and acinar structure in regulating signaling that results in EMT and the onset of a malignant phenotype.

Support or Funding Information

This research was supported in part by the National Institutes of Health (DP2006460 to AJE), Department of Defense ( W81XWH‐13‐1‐013 to AJE and JY), and National Science Foundation Graduate Research Fellowship program (to MO).
Molecular Engineering for Imaging and Reprogramming in Live Cells
Authors: Wang Peter Yingxiao , Sun Jie , Lei Lei
Published: June 2021

Signaling molecules and their activities are well coordinated in space and time to regulate cellular functions in response to mechanical and chemical microenvironment. Based on fluorescent resonance energy transfer (FRET), we have developed genetically encoded biosensors to monitor the dynamic molecular activities (such as Src and FAK activities) in live cells at subcellular compartments when cells interact with their neighbors or mechanical/chemical microenvironment. In a recent study, we show that a ubiquitous signaling protein, Src Homology2 (SH2) Domain‐containing Protein Tyrosine Phosphatase 2 (Shp2), displayed unexpected plasticity of conformational changes via intramolecular interactions within Shp2 (cis‐interaction). We found that two phosphorylated regulatory tyrosines upon stimulated phosphorylation can compete for the cis‐interaction of the same SH2 domain within Shp2 to achieve plasticity. The antagonistic combination of contextual amino acid sequence and position can create arelatively small difference between the two phosphorylated tyrosines in their overall competitiveness for intramolecular conformational regulation. Enlarging this difference by swapping the sequences at the two tyrosine positions resulted in loss of conformational plasticity and reprogrammed downstream ERK signaling dynamics. We have further engineered a machinery molecule based onShp2 which allows an integrated capability of imaging and manipulation of single cell life. We suggest that this strategy can serve as a general and basic design principle for natural and synthetic proteins, with their conformations and functions tunable to regulate downstream physiological consequences. As such, molecular engineering and biosensor development can provide powerful tools for the imaging and manipulation of cellular lives at single cell levels.

Support or Funding Information

This work is supported in part by NIHHL098472, HL109142, HL121365, NSF CBET1360341, UC San Diego.
Chemical Biology And Endomembrane Trafficking In Plants
Authors: Hicks Glenn R , Zhang Chunhua , Van de Ven Wilhelmina , Li Ruixi , Raikhel Natasha
Published: June 2021

Endomembrane trafficking is essential for coordinated growth and development in plants and response to environment. In particular, key plasma membrane proteins such as PIN auxin transporters and the BRI1 brassinosteroid receptor translocate dynamically between the endosomes and plasma membrane. We utilized a previously published large‐scale chemical library screening approach in which we identified small molecules that inhibited pollen germination or tube growth. This tip growth requires exocytosis, endocytosis and other endomembrane trafficking processes

We then identified groups of small molecules that affect trafficking of PIN auxin transporters and other plasma membrane proteins in Arabidopsis roots. One of these compounds (ES2) targets the essential EXO70A1 component of the exocyst complex in Arabidopsis which is necessary for normal development. In Arabidopsis, there are 23 EXO70 genes (in mammals and yeast they are low copy number) where, among other roles, they appear to be involved in autophagy, cell division and pathogen responses. ES2 targets the exocyst complex in plants as well as mammals including humans where defective recycling/exocytosis have been correlated with disease. To our knowledge, ES2 is the first bioactive compound known to have this specificity. To examine exocytosis and its regulation in more detail, we have examined exo70 mutants, generated EXO70A1 structural data and are examining additional small molecules that may affect recycling/exocytosis.

Support or Funding Information

Acknowledgement: This work was supported by the US Department of Energy DE‐FG02‐02ER15295 (GRH, NVR).