bluuebunny/crossref_metadata_2025 huggingface dataset:
Sex differences in diet quality and health measures in US Soldiers entering initial military training
Authors: Lutz Laura J , Gaffney‐Stomberg Erin , Scisco Jenna L , Pasiakos Stefan M , McGraw Susan , Cable Sonya J , Young Andrew J , McClung James P
Published: June 2021

The Dietary Guidelines for Americans (DGA) aim to provide nutrition recommendations that promote health and the Healthy Eating Index (HEI) provides a measure of adherence to the DGA. The HEI may be a useful tool for assessing Soldiers’ diets and identifying nutrition education topics of benefit. This study assessed diet quality and health biomarkers of female (n= 71) and male (n= 117) Soldiers (20 ± 3 yr [mean ± SD]). A quantitative food frequency questionnaire (FFQ) and fasting blood were collected at the beginning of basic combat training. FFQ data were then converted to a HEI score, which ranges from 0–100. The average HEI score was 58.8 ± 10.6. Females had a higher HEI score than males (62.9 ± 11.7 v. 56.4 ± 9.2, P < 0.05) indicating females had better adherence to the DGA. Male HEI scores negatively correlated with their total cholesterol (−0.228; P < 0.05). However, there were no significant correlations between HEI scores and weight, BMI, percent body fat, HDL, LDL, triglycerides, or glucose for either sex. Females had higher HEI scores than males for intake of calories from solid fats, alcoholic beverages and added sugars, percent of energy from saturated fat, whole and total fruit servings, and total vegetable servings (P < 0.05, all such). These data indicate sex differences in diet quality and may provide insight into which topics should be the focus of nutrition education for Soldiers.

Research supported by MRMC.
Candy consumption in childhood is not predictive of cardiovascular risk factors in young adults‐ The Bogalusa Heart Study
Authors: Nicklas Theresa A. , O'Neil Carol E. , Liu Yan
Published: June 2021

Limited research has examined the relationship between candy consumption in childhood on body mass index (BMI) and other cardiovascular risk factors (CVRF) in young adulthood. The goal of this study was to examine whether candy consumption in childhood was predictive of BMI and CVRF in young adulthood. A longitudinal sample of children 10 years (y) (n=355; 61% females; 71% whites, 29% African‐Americans) who participated in cross‐sectional surveys from 1973 to 1984 and also participated in one of two post‐high school surveys as young adults (19–38 y; Mean ± SD = 23.6±2.6 y) in Bogalusa, LA were studied. Dietary data were collected using 24‐hour dietary recalls or a food frequency questionnaire. Candy consumers were defined as those consuming any amounts of candy. Candy consumption was calculated as grams consumed/day and was used as a continuous and categorical variable in linear mixed models, adjusting for appropriate covariates. Dependent variables were BMI and CVRF obtained in young adulthood. 92% of children 10 y reported consuming candy (mean=46±45g); the percentage decreased to 67% (mean=20±30g) when they were young adults. There were no associations found between candy consumption in childhood with BMI or CVRF in young adulthood suggesting that candy consumption was not predictive of health risks later in life. Supported by USDA & National Confectioners Association.
Make Physical Activity (PA) a STAPLE in Children's Lives: Development of a Mnemonic to Assist Parents of Preschoolers in Planning PA Opportunities for their Children
Authors: Golem Devon , Martin‐Biggers Jennifer , Byrd‐Bredbenner Carol
Published: June 2021

Higher PA levels during early childhood are linked with healthy weight, motor skill proficiency, and healthy PA habits later in life. Many preschoolers do not meet PA recommendations, often because parents are not aware of or do not know how to operationalize them. A comprehensive literature review (n=58 articles) conducted using IOM's LEAD framework for obesity prevention documented PA levels, recommendations, and correlates for preschoolers. A mnemonic applying literature findings for PA benefits, guidelines and tips for parents was created. The mnemonic, based on Social Cognitive Theory, includes Space (for activity), Two‐hours (goal/day) with half this time for planned Activity and half for Play (unstructured activity), Leadership (let child choose activities), and Encourage (child to be active and modeling PA). Key informant interviews (n=10 parents, n=8 experts) validated the figure and documented its perceived utility to change parent PA behavioral intentions. Overall, informants felt the mnemonic was useful and enlightening. Mnemonics, like the one developed in this study that present a synthesis of research findings in a “matter‐of‐fact” manner, could help parents explore and resolve ambivalence or resistance toward behavior change, strengthen motivation for change, and reduce preschoolers’ obesity risk and associated health outcomes.

Grant Funding Source: United States Department of Agriculture, National Institute of Food and Agriculture, Grant Number 2011–68001‐30170
Repeated measures of eating competence over a 12‐month nutrition education program revealed rebound following early decline: Implications for interventions planned to enhance eating competence
Authors: Psota Tricia , Lohse Barbara , Kris‐Etherton Penny
Published: June 2021

Eating competence (EC), an intra‐individual approach to eating and food behaviors that entrains positive biopsychosocial outcomes, is measured with the Satter EC Inventory (ecSI). Early studies show a decline in EC after short (< 4 mo) interventions, possibly resulting from increased awareness/discomfort with eating behavior. A 26‐lesson, Dietary Guidelines‐based education intervention with lower‐fat and moderate‐fat versions was assessed at baseline (BL), 4 mo, and 12 mo in overweight/obese premenopausal women (n=101) with a set of biomedical measures that included the ecSI. EC and biomedical findings did not differ between intervention groups or those completing 3 (n=40; 39.7 ± 6.2 y; BMI: 30.7 ± 4.2 kg/m2), or < 3 assessments (n=61; 38.4 ± 6.5 y; BMI: 30.9 ± 4.3 kg/m2). Replicating earlier studies, EC women had or tended toward lower BMI (P=.095), waist circumference (P=.04), uncontrolled (P=.07) and emotional eating (P=.09). ecSI decreased from BL to 4 mo (26.2 ± 1.0 to 24.9 ± 1.3) and rebounded at 12 mo (31.4 ± 1.0; P<.001, compared to BL and 4 mo). This pattern persisted for those in the lower‐fat group and when controlling for weight change. ecSI remained the same at 4 mo in the moderate‐fat group; however scores significantly increased by 12 mo (P<.001, compared to BL and 4 mo). Findings support increased intervention dosage and serial measures to accurately portray intervention effect on EC status. Funded by USDA.
Naturally Occurring Peptides in Human Milk: Identification and Evidence for Antibacterial Action
Authors: Dallas David Charles , Guerrero Andres , Khaldi Nora , Barile Daniela , German J. Bruce , Lebrilla Carlito
Published: June 2021

For over 200 million years, milk has co‐evolved with mammalian infants to be nourishing and immunoprotective. Scientists are now realizing that milk is not simply a mixture of basic nutritional building blocks, but rather a dynamic, active and personal source of nourishment to the human infant. Fragments of milk proteins released during infant digestion have an array of biological functions within the gut and systemically including opioid‐like activity, immune system modulation and antimicrobial action. These peptides are typically inactive within the sequence of the parent protein and only become active when released by proteolysis.

This study aimed to identify the extent of milk protein degradation within the breast and identify released peptides with potential bioactivity. Pooled, mature term breast milk was analyzed for presence of protein fragments by nano‐LC Chip Quadrupole Time‐of‐Flight Tandem Mass Spectrometry (Chip‐Q‐TOF). Of the 508 unique milk peptides identified, most (62%) were derived from ß‐casein. Sixty‐two peptides had significant sequence overlap with known bioactive peptides with antimicrobial or immunomodulatory functions. These 500 unique peptides represent a five‐fold increase in the number of naturally occurring peptides identified in the entire history of milk research.

Grant Funding Source: USDA
Energy intake modulates intramuscular mTORC1 signaling and ubiquitin‐mediated proteolysis during energy deficit independent of dietary protein variations
Authors: Carbone John Walter , Margolis Lee M , McClung James P , Cao Jay J , Young Andrew J , Pasiakos Stefan M
Published: June 2021

Effects of variations in habitual and acute dietary protein consumption on intracellular regulators of human skeletal muscle mass during energy deficit (ED) were investigated. Using a randomized‐block design, 32 men and 7 women consumed diets providing protein at 0.8, 1.6, or 2.4 g·kg−1·d−1 for 31 days. A 10‐day weight maintenance (WM) period preceded 21 days of ED, during which energy intake was reduced to 60% of daily requirements. Phosphorylation of mTORC1 associated signaling proteins, protein ubiquitylation (Ub), and NFκB expression were assessed in vastus lateralis muscle samples obtained after an overnight fast (FAST) and again 120 min after consuming a mixed meal (480 kcal, 20 g protein; FED) on days 10 and 31. Independent of dietary protein level and feeding state, Ub was 25% higher (P< 0.05) for ED versus WM. Energy state and feeding altered p70S6K1 phosphorylation, as FED‐ED levels were 26% lower than FED‐WM (energy‐by‐feeding, P< 0.05). Overall, Akt and rpS6 phosphorylation were higher (P< 0.05), and NFκB and Ub were lower (P< 0.05), for FED versus FAST. These data suggest that ED may upregulate intramuscular ubiquitin‐mediated proteolysis, and attenuate p70S6K1 sensitivity to feeding, although energy consumption, independent of habitual protein intake, may confer skeletal muscle protection through concomitant stimulation of mTORC1 signaling and inhibition of Ub during ED.

Supported by USAMRMC, EMU CHHS SSSA, and USDA ARS
Divergent intramuscular mTORC1 signaling responses to habitual consumption of milk‐and soy‐based energy restricted diets
Authors: Murphy Nancy E , Lin Gregory G , Wulff Charles R , Gaffney‐Stomberg Erin , Young Andrew J , McClung James P , Pasiakos Stefan M
Published: June 2021

The effects of milk‐or soy‐based, high‐protein, energy restricted (ER) diets on intracellular regulators of muscle mass are not well described. Eighty male Sprague‐Dawley rats were randomly divided into groups and fed energy adequate (EA) and ER (60% of average ad libitum EA feed intake) diets containing two protein sources (milk protein concentrate: MPC; or isoflavone‐free soy isolate: SOY) and levels (control protein: CP, 10%; high protein: HP, 35%) for 16 weeks. Basal protein expression and phosphorylation of mammalian target of rapamycin complex 1 (mTORC1) associated proteins were assessed using multiplex. Phosphorylation of mTOR was lower (P < 0.05) for ER compared to EA. Independent of protein level, p70S6K1 expression was lower (P < 0.05) for ER MPC compared to EA MPC. However, mTOR and rpS6 protein expression were higher (P < 0.05) for ER SOY than ER MPC, and TSC2, mTOR, and rpS6 were higher (P < 0.05) for ER SOY compared to EA SOY. Phosphorylation of rpS6 was also higher (P < 0.05) for EA SOY compared to EA MPC, and p70S6K1 phosphorylation was higher (P < 0.05) for SOY versus MPC, regardless of energy status and protein level. These data demonstrate divergent mTORC1 responses to soy‐and milk‐based energy restricted diets but suggest that habitual consumption of soy‐based diets may enhance protein synthetic responses to feeding by upregulating the expression of key mTORC1 signaling proteins. Supported by Dairy Research Institute and USAMRMC
Extrinsic labeling of Fe in maize, lentil and bean meals does not fully equilibrate with intrinsic Fe
Authors: Glahn Raymond Philip , Cheng Zhiqiang , Rutzke Michael
Published: June 2021

Extrinsic isotopic labeling of food Fe has been used for over 50 years to measure Fe absorption. This method is based on the assumption that complete equilibration occurs between the extrinsic and the intrinsic Fe prior to absorption. The present study tested this assumption via use of simulated in vitro digestion of varieties of maize, white beans, black beans, red beans and lentils. Prior to digestion, foods were extrinsically labeled with 58Fe at concentrations of 1, 10, 50 and 100% of the intrinsic Fe. Comparison of Fe solubility was used as the first measure of equilibration. In almost all foods, significantly more of the extrinsic Fe (up to 2–3 times) partitioned into the supernatant. The effect varied depending upon the seed coat color (ie. beans, suggesting polyphenols play a role), and the concentration of the extrinsic Fe. There was no crop in this study that consistently demonstrated equal equilibration of the extrinsic Fe with the intrinsic Fe. Moreover, different varieties of a crop exhibited different patterns of extrinsic and intrinsic Fe solubility. These observations call into question the accuracy of Fe absorption studies where isotopic extrinsic Fe was used to evaluate Fe absorption and bioavailability. It suggests that intrinsic labeling of Fe is necessary for accurate measurement of Fe absorption. Funded by USDA‐ARS.
Associations between vitamin D status and lipid profiles in military recruits
Authors: McClung James P , Gaffney‐Stomberg Erin , Lutz Laura J , Rood Jennifer C , Cable Sonya J , Pasiakos Stefan M , Young Andrew J
Published: June 2021

Recent data suggest a relationship between vitamin D status and cardiometabolic risk factors, especially in populations with cardiovascular disease, although few studies have investigated this association in healthy, young people. This study investigated the relationship between serum vitamin D (25[OH]D) and lipid profiles in military recruits. Blood biomarkers were assessed in fasted volunteers (n = 210, males = 134, females = 76, age [mean ± SD] = 20 ± 3 yrs) within one week of initially reporting for military service. In the total study population, there was a significant positive relationship between serum 25[OH]D and total cholesterol in females (P < 0.05, r = 0.241), but not males (r = 0.067). This relationship was significant in white females (P < 0.05, r = 0.365), but not in non‐white females. There were positive associations between 25[OH]D and HDL in white females (P < 0.05, r = 0.325), but not in males or non‐white females. Similarly, 25[OH]D and LDL levels were associated in white females (P = 0.06, r = 0.256), but not in males or non‐white females. These data suggest sex and race differences in relationships between serum 25[OH]D and lipid profiles in military recruits, with the strongest associations occurring in white females. Future studies should clarify the biological significance of associations between 25[OH]D and lipid profiles in healthy, young populations. Research supported by USAMRMC.
High vitamin D and calcium intakes decrease diet‐induced obesity
Authors: Sergeev Igor N
Published: June 2021

Modulation of apoptosis is emerging as a promising anti‐obesity strategy because removal of adipocytes through this process will result in reducing body fat. Effects of vitamin D on apoptosis are mediated via multiple signaling pathways that involve common regulators and effectors converging on cellular Ca2+ (Sergeev, 2005, 2009, 2012). We have previously shown that 1,25(OH)2D3 induces Ca2+signal associated with activation of Ca2+‐dependent apoptotic proteases in mature adipocytes. In this study, a mouse diet‐induced obesity (DIO) model was used to evaluate the role of vitamin D and calcium in adiposity. DIO mice fed high D3, high Ca, and especially high D3 plus high Ca diets demonstrated a decreased body and fat weight gain, improved markers of adiposity and vitamin D status (plasma concentrations of glucose, insulin, adiponectin, 25(OH)D, 1,25(OH)2D, PTH), but an increased plasma Ca2+. High D3 and Ca intakes were associated with activation of Ca2+‐dependent apoptotic proteases, calpain and caspase‐12, in adipose tissue of DIO mice. The results imply that high vitamin D and Ca intakes activate Ca2+‐mediated apoptotic pathway in adipose tissue. Targeting this pathway with vitamin D and Ca supplementation can represent an effective and affordable approach to prevention and treatment of obesity. However, this approach needs to be evaluated from a safety point of view. Supported by USDA 2009 35200 05008 and SD H325.
Garlic intake influences gene expression in whole blood
Authors: Charron Craig S. , Dawson Harry D. , Solano‐Aguilar Gloria I. , Molokin Aleksey , Vinyard Bryan T. , Milner John A. , Novotny Janet A.
Published: June 2021

Epidemiological data suggest that garlic intake is inversely associated with the progression of heart disease and cancer. A host of preclinical studies support these associations. To investigate mechanisms of garlic action in humans, we conducted a randomized crossover feeding trial in which 17 volunteers consumed a garlic‐containing meal (100 g white bread, 15 g margarine, 5 g crushed garlic) or a garlic‐free control meal (100 g white bread, 15 g margarine) after 10 days of a controlled, garlic‐free diet. Blood was collected before and 3h after test meal consumption for gene expression analysis in whole blood. Global gene expression was assessed by Illumina, followed by real‐time RT‐PCR on selected genes. Seven genes were found to be upregulated by garlic intake: AhR, ARNT, H1F1A, JUN, NFAMI, OSM, and REL. Fold‐increases in mRNA transcripts ranged from 1.6 (HIF1A) to 3.0 (NFAM1) (P<0.05). OSM and NFAM1 expression was increased in women but not men. The biological significance of these results is likely to be multifaceted. For example, AhR is involved in xenobiotic metabolism, while OSM is a pleiotropic tumor inhibitory cytokine, and JUN and REL participate in inflammation. Supported by USDA and NCI.
Astaxanthin supplementation lowers plasma triglyceride concentrations in diet‐induced obese C57BL/6J mice
Authors: Yang Yue , Pham Tho , Wegner Casey , Kim Bohkyung , Ku Chai Siah , Park Young‐Ki , Lee Ji‐Young
Published: June 2021

Elevated plasma triglyceride (TG) and cholesterol concentrations are risk factors for cardiovascular diseases. We previously reported that astaxanthin (ASTX), a xanthophyll carotenoid, lowers plasma total cholesterol and TG concentrations in apolipoprotein E knockout mice. To investigate if ASTX exerts hypolipidemic effects in the diet‐induced obesity (DIO) mice and if so, to determine an effective supplementation level, male C57BL/6J mice were fed a high fat (HF) control diet (60% energy from fat) or a HF diet supplemented with 0.003%, 0.01% or 0.03% ASTX by weight for 12 wk. Only mice fed 0.03% ASTX showed a significant reduction in plasma TG levels, suggesting this level of supplementation is necessary for ASTX to have a TG‐lowering effect. Compared with control, plasma high density lipoprotein cholesterol (HDLc) levels were significantly higher in 0.03% ASTX‐fed mice without a difference in plasma non‐HDLc levels. The ASTX supplementation also decreased plasma aspartate aminotransferase levels, suggesting it protects against high fat‐induced tissue damages. In conclusion, 0.03% ASTX in diet decreased plasma TG and increased HDLc in DIO mice and therefore, it is potentially beneficial for lowering CVD risk.

Grant Funding Source: USDA AFRI 2012–67018‐19290
Biotinylation of the c‐myc promoter binding protein MBP‐1 decreases c‐myc expression in mammary carcinoma MCF‐7 cells
Authors: Zhou Jie , Wijeratne Subhashinee S. K. , Zempleni Janos
Published: June 2021

Biotin feeding studies in mice predisposed to developing breast cancer suggest that the tumor load increases by ~100% in biotin‐deficient mice compared with controls, and that the increase is associated with metastasis rather than new tumors. The oncogene c‐myc is implicated in cancer metastasis, and its expression is repressed by binding of MBP‐1 to the c‐myc promoter. Discovery‐based studies for identifying novel biotin‐containing proteins suggest that MBP‐1 contains covalently bound biotin, judged by mass spectrometry. We hypothesized that biotinylation of MBP‐1 increases the binding of MBP‐1 to the c‐myc promoter, thereby decreasing c‐myc expression and metastasis. MCF‐7 cells were cultured in biotin‐defined media (deficient, sufficient, supplemented) and transfected with a reporter gene construct for the c‐myc promoter. Reporter activity was ~100% greater in biotin‐deficient cells compared with sufficient and supplemented controls. Likewise, c‐myc mRNA levels were 10% higher in deficient compared with sufficient cells, and 10% high in sufficient compared with supplemented cells. Biotinylation of MBP‐1 is an enzymatic process, judged by streptavidin blots. These observations are consistent with our hypothesis and preliminary studies, suggesting that biotin depletion causes an increased expression of the oncogene c‐myc, thereby increasing cancer risk.

Grant Funding Source: ARD Hatch, NIFA, and NIH
Polyunsaturated fatty acids (PUFA) modulate association between PIK3CA‐KCNMB3 variants and insulin resistance
Authors: Zheng Ju‐Sheng , Arnett Donna K , Parnell Laurence D , Lee Yu‐Chi , Ma Yiyi , Smith Caren E , Richardson Kris , Li Duo , Borecki Ingrid B , Tucker Katherine L , Ordovás José M , Lai Chao‐Qiang
Published: June 2021

The objective was to examine associations of PIK3CA‐KCNMB3 variants with glucose‐related traits and interaction with dietary fat. We investigated genetic associations and modulation by dietary fat in the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) study (n=820) and replicated the findings in the Boston Puerto Rican Health Study (BPRHS) (n=844). For KCNMB3 missense mutation rs7645550, homeostasis model assessment of insulin resistance (HOMA‐IR) was significantly lower in TT homozygotes compared with CT carriers in both GOLDN (P=0.005) and BPRHS (P=0.042); this SNP interacted with dietary n‐3: n‐6 PUFA ratio for HOMA‐IR (P=0.006) in GOLDN. For rs1183319, G allele carriers had higher fasting glucose compared with non‐carriers in GOLDN (P=0.008), and had marginally higher glycosylated hemoglobin (HbA1c) than non‐carriers in the BPRHS (P=0.053); this SNP showed significant interaction with n‐3:n‐6 ratio on HOMA‐IR in GOLDN and on HbA1c in the BPRHS. In summary, PIK3CA‐KCNMB3 variants are associated with glucose‐related traits in populations of different ancestries, and are modified by dietary PUFA.

Grant Funding Source: USDA
In vivo regulation of gene transcription by alpha and gamma‐tocopherol in murine T lymphocytes
Authors: Zingg Jean‐Marc , Han Sung Nim , Pang Eunice , Meydani Mohsen , Meydani Simin Nikbin , Azzi Angelo
Published: June 2021

Vitamin E is present in food in 8 different analogues (α‐, β‐, γ‐, δ‐tocopherols and tocotrienols). Although alpha‐tocopherol (α‐T) has been mostly studied, the other analogs also have raised interest, especially gamma‐tocopherol (γ‐T) which is abundant in the US diet. We compared the effect of dietary supplementation with adequate or high doses of α‐T or γ‐T on induction of gene expression in activated T cells. Old C57BL/6 mice were fed diets containing adequate (30 PPM) or high (500 PPM) doses of α‐T or γ‐T for 4 weeks, T cells from the spleen isolated and stimulated with plate‐bound anti‐CD3 and soluble anti‐CD28, and gene expression changes assessed by gene array analysis. The data obtained indicate significant qualitative and quantitative differences between the two analogs in regulating gene expression induced by T cell stimulation. Unique marker genes were found that could be useful for establishing the response to vitamin E for both α‐T (e.g. induced: CD40 ligand, lymphotoxin A) and γ‐T (e.g. repressed: poliovirus receptor‐related 2) supplementation. Interestingly, with high doses of supplementations, α‐T activated a higher number of genes than γ‐T, whereas γ‐T reduced a higher number of genes than α‐T in stimulated T‐cells. Whether the observed changes in gene expression in T cells after high γ‐T supplementation reduces overall inflammation or results in changes in T cell mediated function remains to be determined. Supported by USDA Contract #58–1950‐0–014.
Skeletal muscle vitamin D receptor associated with serum 25‐hydroxyvitamin D
Authors: Pojednic Rachele M. , Ceglia Lisa , Lichtenstein Alice , Dawson‐Hughes Bess , Fielding Roger
Published: June 2021

BACKGROUND

The effects of vitamin D on aging muscle may be mediated by its receptor (VDR). VDR is expressed in human skeletal muscle and when bound to the active metabolite, 1,25‐dihydroxyvitamin‐D3, regulates expression of genes that affect muscle cell differentiation and contractility. Evidence on the relationship between serum 25‐hydroxyvitamin‐D (25OHD) and VDR in human skeletal muscle is limited.

PURPOSE

To examine the association between VDR in skeletal muscle and 25OHD in older mobility‐limited adults.

METHODS

A cross‐sectional analysis of 18 mobility limited older adults (age 65–95) was conducted. Serum 25OHD and a muscle biopsy of the vastus lateralis were sampled. VDR protein expression was assessed via Western Blot.

RESULTS

Mean (±SD) 25OHD was 28.9 (±11.6) ng/mL (range: 15.0 to 53.0 ng/mL). 25OHD was strongly associated with VDR concentration (R=0.67; p=0.001). VDR concentration differed significantly between vitamin D sufficient (>;20ng/mL) and insufficient groups (≤20ng/mL, p=.02).

CONCLUSION

VDR in skeletal muscle is strongly associated with circulating 25‐Hydroxyvitamin‐D in older mobility limited adults. Individuals with sufficient 25OHD have significantly greater concentrations of VDR than those with insufficient 25OHD. This study provides support to the hypothesis that 25OHD may affect VDR protein expression in human skeletal muscle. NHLBI Training Grant T32HL69772.
Neurotoxic effects of manganese on GABAergic Iinervation in the bivalve mollusc Crassostrea virginica
Authors: Ogunnoiki Jumoke , Jackson Kelly , Catapane Edward J. , Carroll Margaret A.
Published: June 2021

High levels of airborne manganese (Mn) cause Manganism a Parkinsons‐like disease in humans by interfering with dopamine (DA) neurotransmission. Studies are showing GABA neurons also are damaged by Mn. C. virginica contains a DA and serotonin (HT) innvervation of its gill. Previously we showed Mn disrupts DA innervation. We also showed the cerebral ganglia (CG) contains GABA and within the CG GABA inhibits HT innervation of gill. Here we studied if ganglia and peripheral tissues contain GABA receptors, and if Mn effects GABA neurons within CG of C. virginica. We used 1Eantibodies (GABAA Ra1–6 ) and 2E antibodies (IgG‐FITC) to detect GABA receptors. We found GABA receptors in CG, visceral ganglia, palps and digestive tract. We also examined effects of Mn on GABA inhibition of HT neurons in CG. Beating of lateral cilia in gill cells were measured by stroboscopic microscopy. Applying HT to CG caused a dose‐dependent increase in beating. Applying GABA prior to HT prevented the increase. Acute applications of Mn (50 and 500 FM) prior to GABA prevented GABA from inhibiting HT. The study shows GABA receptors present in ganglia and peripheral tissues in C. virginica and acute Mn treatments damages GABA neurons. C. virginica preparations are good, simple test preparation to study the GABAergic system and the mechanism underlying the neurotoxic effects of Mn. The work was supported by 0516041171of NYSDOE and 0622197 of DUE Program of NSF.
Effects of antioxidants and anti‐inflammatory agents on neurotoxic effects of manganese on dopaminergic innervation of gill of the bivalve mollusc Crassostrea virginica
Authors: LaFleur Kisha , Rotibi Mojisola , Catapane Edward J. , Carroll Margaret A.
Published: June 2021

Lateral gill cilia of Crassostrea virginica are controlled by a serotonin‐dopamine (DA) innervation. DA is an excitatory neurotransmitter within ganglia, but inhibitory at gill, causing cilio‐inhibition. The mechanism of action of manganese (Mn) toxicity is not fully understood, but may be due to oxidative damage. We found several chelators prevented neurotoxic effects of Mn in C. virginica. We examined if anti‐inflammatory agents and/or antioxidants are effective in preventing neurotoxic actions of Mn in gill of C. virginica. Indomethacin (IM), an anti‐inflammatory agent with antioxidant abilities, and ascorbic acid (AA), an antioxidant with possible anti‐inflammation abilities were tested. We examined acute and short term (3 – 5 days) treatments on DA innervation. Beating of lateral cilia in gill cells was measured by stroboscopic microscopy. Acute or short‐term treatments of IM or AA (25–100 μM) had no effects on the cilio‐inhibitory effects of DA (10−6–10−3 M). When acute or short‐term Mn treated animals (25–100 μM) were pretreated with IM or AA (25–100 μM), both drugs effectively prevented neurotoxic effects of Mn, with AA being more effective than IM. The study shows antioxidants and anti‐inflammatory agents can block the neurotoxic actions of Mn and may be possible therapeutic agents in the treatment of Manganism. This work was supported by 2R25GM06003 of NIGMS, 0516041171 of NYSDOE and 0622197 of NSF.
Bioscavengers to protect against chemical warfare nerve agents: safety of adenovirus‐driven expression of butyrylcholinesterase in macaques
Authors: Hamilton Lindsey R , Myers Todd M
Published: June 2021

To date, no pretreatment against chemical warfare nerve agents offers better protection than endogenous esterases. These bioscavengers are capable of sequestering nerve agents and protecting acetylcholinesterase from inactivation, thereby preventing the toxic and lethal effects of the nerve agents. The current leading bioscavenger (butyrylcholinesterase, BChE) is purified from human serum and is quite expensive. Additionally, the enzyme is only capable of binding nerve agents stoichiometrically; thus, large amounts of human BChE are required to protect against large doses of nerve agent. A novel approach is to utilize an adenovirus to produce higher levels of in vivo expression of an esterase, thereby providing prolonged protection without the need to purify large quantities of the bioscavenger. Thus, cynomolgus macaques were injected with an adenovirus designed to increase BChE systemically. The safety and pharmacokinetic profile of the virus infection is currently ongoing. To assess efficacy of this pretreatment, monkeys will perform the temporal response differentiation task before and after infection with the virus and will be challenged with an otherwise lethal dose of nerve agent to demonstrate neurobehavioral protection. This research is supported by the Defense Threat Reduction Agency – Joint Science and Technology Office, Medical S&T Division.
Spectral profiles of acute and healed myocardial radiofrequency ablation lesions
Authors: Swift Luther M , Jaimes Rafael , Sarvazyan Narine
Published: June 2021

A spectral approach to differentiate between acute and healed myocardial lesions was explored. The long‐term goal of these studies is improved treatment of atrial fibrillation using percutaneous RF catheters. The epicardial surface of excised rat hearts was illuminated by continuous wavelength spectrum via an optical fiber coupled to a spectrofluorometer. Emitted, scattered and reflected light was acquired by a high sensitivity CCD camera. Spectra were separated into three groups: unablated tissue, acute and healed RF lesions. Structure of the lesions was confirmed by cytopathology. Spectra between all three groups differed significantly throughout the 300–800nm range. The largest difference was observed in the 300–400nm illumination range accounting for the loss of NADH fluorescence in acute lesions, but not in healed tissue. In the 400–620nm illumination range both acute and healed epicardial lesions exhibited significantly higher intensity compared to control tissue. In the 620–800nm range difference between three groups diminished. The results point to a possibility of in situ visualization of acute and chronic lesions based on the spectral differences in reflected and emitted light. The sensitivity and specificity of lesion identification can be further increased by ratiometry of images acquired at different illumination wavelengths (Supported by the NIH HL095828 & NSF CBET1231549)
Temperature influences CO2/pH‐sensitivity in locus coeruleus neurons of the bullfrog, Lithobates catesbeianus
Authors: Santin Joseph , Hartzler Lynn
Published: June 2021

The locus coeruleus (LC) is a chemoreceptive site involved in respiratory control. In bullfrogs, neurons within the LC of the brainstem increase firing rates during physiologically relevant CO2/pH changes. Given the sensitivity of these neurons to CO2/pH changes at 22°C, passive acidification and alkalinization during warming and cooling may significantly alter LC neuron CO2/pH sensitivity. We hypothesized that acute cooling of LC neurons from 22°C to 15°C would reduce and acute warming to 29°C would enhance chemosensitivity during hypercapnic acidosis (HA; ΔpH=0.4). Whole‐cell patch clamp electrophysiology was used measure membrane potentials and firing rates from LC neurons in the bullfrog brainstem slice preparation. At 15°C, sensitivity to HA was abolished (chemosensitivity index (CI)=102±2.9% vs. 185±27.7% at 22°C; p<0.05); however, at 29°C, chemosensitivity was unchanged (CI=165±23.8% vs. 203±29.3% at 22°C; p>;0.05). Cooling from 22°C to 15°C increased firing rates and membrane potentials (0.63±0.51 Hz/−38.07±3.07mV vs.1.94±1.15 Hz/−31.60±6.26 mV; p<0.05), while warming from 22°C to 29°C reduced firing rates (1.54±0.58 Hz vs. 0.68±0.55 Hz; p<0.05) with a trend toward hyperpolarization (−37.60±4.37mV vs. −39.50±5.53mV; p=0.1). These results indicate that cooling may reduce LC sensitivity to HA, likely through a temperature‐dependant mechanism. Supported by NSF ADVANCE HRD 0810989.
Effects of airway pressure, hypercapnia, and hypoxia on pulmonary vagal afferent activity in the alligator
Authors: Marschand Rachel , Wilson Jenna , Burleson Mark L , Crossley Dane A , Hedrick Michael S
Published: June 2021

The American alligator (Alligator mississippiensis) is an aquatic vertebrate that dives and has a periodic breathing pattern. The effects of airway pressure, hypercapnia, and hypoxia on dynamic and static responses of pulmonary stretch receptors (PSR) were studied in juvenile alligators (mean mass = 246 g) at room temperature (24°C). Alligators were initially anesthetized with isoflurane, pithed, tracheotomized and attached to a small animal ventilator. The vagus nerve (cranial nerve X) was exposed, sectioned and submerged in mineral oil. Nerve filaments were teased from the nerve trunk and placed over platinum electrodes for differential nerve recording. Afferent tonic and phasic activity was recorded. The majority of receptors appeared to be slowly‐adapting PSRs (SAR) with varying thresholds and degrees of adaptation and no CO2 sensitivity. Dives were simulated in order to characterize receptor activity during and after prolonged periods of inflation and deflation. Some stretch receptors showed a change in dynamic response, exhibiting inhibition after lung inflation and potentiation after lung deflation. These airway stretch receptors may be involved in buoyancy control and/or recovery of breathing patterns and lung volume during pre– and post‐diving behavior and apneic periods. Supported by NSF CAREER IOS 0845741 (DAC).
T3: Does It Really Make the Alligator Go “Tick, Tock”?
Authors: Sirsat Tushar Saoji , Sirsat Sarah K Goy , Dzialowski Edward M
Published: June 2021

Thyroid hormones (TH) are key regulators of vertebrate metabolism and are thought to regulate the development of endothermy in mammals and birds. In order to better understand the effect of TH on development of metabolic physiology of an ectothermic species, we treated neonate American alligators with triiodothryronine (T3) or the thyroperoxidase inhibitor, methimazole, for 28 days to induce hyperthyroidism and hypothyroidism, respectively. We measured whole animal O2 consumption (VO2) by flow though respirometry followed by isolated mitochondria respiration from cardiac muscle and hind limb thigh skeletal muscle. Whole animal VO2 of T3 and methimazole treated subjects differed significantly from control: hyperthyroid animals exhibited higher VO2 while hypothyroid animals showed lower VO2, independent of body mass. These differences were present after five days of exposure and were maintained through day 28. The LEAK and OXPHOS states of respiration of isolated cardiac and skeletal mitochondria from hypothyroid animals were significantly lower than control mitochondria. There were no differences in mitochondrial respiration between hyperthyroid and control animals. We provide support that T3 acts as a metabolic regulator of the American alligator at both the whole animal and mitochondrial level and plays a role in early development of neonate metabolism. Supported by NSF IOS 1146758 (EMD).
The Effects of Imatinib on the Parameters of Contraction in Rat Myometrium Exposed to Different Hormonal Levels
Authors: Uzomah Onyeka Nkolika , Pelaez Nancy
Published: June 2021

The discovery of myometrial telocytes (MT) suggests a potential role in regulating spontaneous contractile activity (SCA). Currently, the function of MT remains unknown. Here we address the hypothesis that MT mediate the action of hormones that are known to modify myometrial SCA. SCA of myometrial rings from retired breeder rats were investigated in vitro using an isometric muscle bath in the presence/absence of imatinib, a drug reported to block MT activity. Results from rats at different phases of their cycle show that imatinib causes a significant decrease in the frequency of SCA during the metestrus phase. This was true for the proximal, mid‐sectional, and distal ends of the uterine horn. However, at the metestrus phase, the force magnitude and the integral of force over time of the SCA remained unaffected by imatinib, indicating that MT may influence SCA frequency without affecting the contractile force. Preliminary results with uterus isolated during the proestrus phase showed inconsistency in contractile magnitude and frequency. We anticipate that MT activity modifies SCA according to the estrus phase of the animal and thus mediates the effect of normal hormonal changes on myometrial contractility. Research supported by the NSF Midwest AGEP Program.
Incidence of Intrazygomatic Sutures in Rhesus Macaques
Authors: Wang Qian
Published: June 2021

Craniofacial sutures are weak points compared to rigid bone on the skull thence they must be shielded from unduly high stresses so as not to disrupt vital growth processes and skeletal functions, especially in areas such as the midface. However, under rare circumstances, extra sutures may present in the midface such as intrazygomatic sutures (incidence of 2.5–6.5% in modern human populations), which arguably would cause unfavorable impact on the ontogeny and biomechanics of the facial skeletons. In this study, the skeletal collection derived from Cayo Santiago (CS) and CS‐derived Rhesus macaque (Macaca mulatta) populations in the Caribbean Primate Research Center was screened for the intrazygomatic sutures. It was hypothesized that the incidence of the intrazygomatic suture in Rhesus macaques was comparable to that in humans. Results demonstrated that, in all examinable skulls with known sex and age information (Total 2483), there are only six specimens that carry intrazygomatic sutures (two Females and four Males; three on left side and three on right side; five with one suture and one with two sutures). Thus, the overall incidence of intrazygomatic sutures in Rhesus macaques, 0.24%, is significantly lower than that in humans (p<0.001). The negative impact of intrazygomatic sutures in the midface needs more biomechanical and morphometric analyses, and insights into this rare condition may deepen our understanding of the craniofacial form and adaptation, and help to improve therapeutic philosophies in corrective and regenerative medicine. Supported by NSF HOMINID BCS‐0725183.
Mediator Affects Pol II Recruitment and Nucleosome Displacement at Heat Shock Protein (HSP) Genes in Saccharomyces cerevisiae
Authors: Moustafa Yara , Gross David S.
Published: June 2021

Mediator, an evolutionarily conserved transcriptional coactivator, plays a critical role in governing both constitutive and induced levels of heat shock protein (HSP) gene expression in budding yeast. Yet how Mediator, and in particular each of its four structural modules – Head, Middle, Tail and Kinase – participates in HSP gene regulation and crosstalk with other transcriptional coactivators remains unclear. Using the conditional depletion technique termed Anchor Away, we have studied the role of two essential Middle Module subunits, Med14 and Med7, in the transcription and chromatin remodeling of HSP genes. Depletion of either subunit disrupts integrity of Mediator, as assessed by a failure of Med20 (Head) to be recruited to induced HSP genes. Pol II recruitment is also affected, and its loss parallels that of Med7. Strikingly, quantitative depletion of Med14 has a relatively modest effect on Pol II recruitment at HSP genes. Nucleosome disassembly triggered by heat shock is likewise differentially affected by loss of Med14 and Med7. These findings suggest that Mediator integrity is important, but not essential, for Pol II recruitment and chromatin remodeling upon heat shock. The interplay between Mediator and other transcriptional coactivators (SAGA, TFIID, RSC, Swi/Snf) in regulating stress‐induced transcription is under investigation. This work is supported by NSF grant MCB‐1025025 awarded to D.S.G.
DEK is a possible IgM mRNA processing regulator
Authors: Turcios Lilia M. , Peterson Martha L.
Published: June 2021

The IgM pre‐mRNA is alternatively processed to two mRNAs that encode secreted or membrane‐associated proteins, through competing splice and cleavage‐polyadenylation reactions. The relative use of the competing reactions changes during B cell development and it is the architecture of the gene rather than specific regulatory elements that are required for this regulation. To identify potential trans‐regulators of IgM, we analysed RNA from a mouse B cell line before and after stimulation with LPS or IL‐5 to induce maturation. One of the genes identified to change is DEK, which encodes an abundant nuclear protein involved in several cellular processes, including mRNA splicing. We confirmed that DEK expression is higher in several mouse B cell lines than plasma cell lines by RT‐qPCR, consistent with the microarray data. Western blot analysis also suggests that DEK protein levels are higher in B cells compared to plasma cells. If DEK contributes to IgM mRNA processing regulation, we expect that overexpression of DEK in a plasma cell line would decrease the pA/splice expression ratio. Conversely, decreasing DEK levels in B cells by siRNA may increase this ratio. Whether the effect of DEK is direct or indirect will be assessed by ChIP assay. We are also examining other potential regulators from the microarray data to better understand IgM mRNA processing regulation. Supported by NSF grant MCB‐0919099.
Allosteric Regulation of Glyoxasomal Malate Dehydrogenase by Citrate Involves Reciprocating Active Site Communication.
Authors: Mays Jacqunae , Bell Ellis
Published: June 2021

Mitochondrial and glyoxysomal forms of malate dehydrogenase are thought to be subject to allosteric regulation by citrate. To explore the mechanisms of citrate effects on the enzyme we have used a combination of initial rate kinetics, site directed mutagenesis, circular dichroism (in conjunction with thermal melts) and ANS Fluorescence to follow effects on secondary and tertiary structure, dynamic light scattering to follow effects on quaternary structure and limited proteolysis to explore local flexibility/accessibility changes. Citrate, which binds to only one of the two otherwise identical active sites, is a partial inhibitor under all circumstances, has no effect on quaternary structure but induces changes in local flexibility/accessibility in a loop near the subunit interface. Several mutations at the subunit interface (V195A, L269A, E256Q, H90Q) impact citrate inhibition. Studies of ANS binding have also been used to explore carboxylic acid induced changes in conformation (glyoxysomal, mitochondrial and cytoplasmic). The results suggest carboxylic acid substrates of MDH induce a reciprocating subunit mechanism that may contribute to catalysis, and citrate exerts its effects by binding to the “empty” active site in this reciprocating subunit mechanism, eliciting altered subunit interactions that contribute to regulation of the overall catalytic cycle. Supported by NSF Grant MCB 0448905 to EB
Itaconic acid, a missing link in macrophage activation, is produced by IRG1
Authors: Cheng Jiongjia , Jewel Delilah , Chiles Thomas C. , Stec Boguslaw , Roberts Mary Fedarko
Published: June 2021

Itaconic acid (ITA) is a metabolite produced by primary macrophages and cell lines derived from macrophages that is dramatically increased upon activation. ITA is synthesized by a cis‐aconitate decarboxylase (cADC). Sequence searches with a known fungal cADC show little homology to ORFs in mammalian cells. Studies with the macrophage cell line RAW246.7 show that cADC activity upon stimulation is dependent on de novo protein synthesis. MS analyses of partially purified RAW 246.7 protein extracts from stimulated cells show a very large increase for immune‐responsive gene 1 protein (IRG1). IRG1 synthesis follows the time course of ITA generation (measured by 1H NMR) and siRNA knockdown of the IRG1 reduces cADC activity upon stimulation. Finally, expression of murine IRG1 in E. coli introduces cADC activity into protein extracts of the transformed bacterium. IRG1 previously was suggested to be a 2‐methylcitrate dehydratase. However, both the partially purified protein fraction from RAW246.7 cells containing cADC activity and recombinant IRG1 can not catalyze 2‐methylcitrate dehydration. This identifies IRG1 as the first mammalian cADC. Ongoing experiments seek a role for ITA in macrophages and in the mammalian immune system. This work has been supported by the U.S. Department of Energy, Office of Science, Energy Biosciences DE‐FG02–91ER20025.
Kinetic and Spectroscopic Studies of Bicupin Oxalate Oxidase and Putative Active Site Mutants
Authors: Moomaw Ellen W , Hoffer Eric , Moussatche Patricia , Salerno John , Grant Morgan , Immelman Bridget , Uberto Richard , Ozarowski Andrew , Angerhofer Alexander
Published: June 2021

Ceriporiopsis subvermispora oxalate oxidase (CsOxOx) is a Mn‐dependent enzyme that catalyzes the oxygen‐dependent oxidation of oxalate to carbon dioxide in a reaction that is coupled with the formation of hydrogen peroxide. EPR spectra indicate that the Mn is present as Mn(II). EPR spin‐trapping experiments support the existence of an oxalate‐derived radical species formed during turnover. The dominant contribution to catalysis comes from the monoprotonated form of oxalate binding to a form of the enzyme in which an active site carboxylic acid residue must be unprotonated. The pH profile of the D241A CsOxOx mutant suggests that the protonation state of aspartic acid 241 is mechanistically significant and that catalysis takes place at the N‐terminal Mn binding site. The fact that the D241S CsOxOx eliminates Mn binding to both the N‐ and C‐ terminal Mn binding sites suggests that both sites must be intact for Mn incorporation into either site. Mutation of conserved arginine residues further support that catalysis takes place at the N‐terminal Mn binding site and that both sites must be intact for Mn incorporation into either site. This work was supported by the National Science Foundation (MCB‐1041912) to EWM.