bluuebunny/crossref_metadata_2025 huggingface dataset:
Slow starch digestion redefined at limit dextrin level by mucosal maltase‐glucoamylase and sucrase‐isomaltase activities
Authors: Nichols Buford L , Quezada‐Calvillo Roberto , Ao Zihua
Published: June 2021

Mucosal maltase‐glucoamylase (MGAM) and sucrase‐isomaltase (SI) digest free glucose from food starches. Amylase (AMY) amplifies these mucosal activities by production of soluble limit dextrins (LDx). This network of enzyme activities determines rate of LDx entry into either the glycemic or the fermentive pathways. We determined the roles of the mucosal activities as a function of AMY treatment and LDx substrate. Recombinant human AMY was used to produce 8 different LDx. Steady state (SS) was reached after 30 min of AMY treatment. These substrates were incubated with human jejunal lysates (LYS) or immunopurified (IP) MGAM or SI. Glucogenesis was measured by real‐time assay. Rates were normalized to maltose. Normal maize with MGAM‐IP was used as reference for calculations of % slowly digesting LDx (% RLDx) at SS. LYS SS‐rates for other LDx were less than normal maize. SS‐rate was always fastest with MGAM‐IP but all rates varied by LDx substrate. LYS activity rate was amplified by AMY ~12 X. AMY amplified SI‐IP more than MGAM‐IP. LYS SS‐assays revealed 20% RLDx for normal maize; 30% for potato; 33% for tapioca; 36% for Ae, sugary2 and waxy maize; 40% for rice; and 60% for glycogen. LYS rate best correlated with SI‐IP rate (R2 = 93). The MGAM‐IP and SI‐IP activities of jejunal LYS digested the 8 LDx to glucose with only 60‐80% efficiency. The % RLDx entering the fermentive pathway was underestimated by previous assays with fungal enzymes. Supported by USDA/ARS.
Single‐molecule experiments provide evidence for a new step in the catalytic mechanism of DNA polymerase
Authors: Christian Thomas David , Rueda David , Romano Louis J
Published: June 2021

The catalytic mechanism of DNA polymerases has been shown to involve multiple steps that precede and follow the transfer of a nucleotide to the 3′‐hydroxyl of the growing DNA chain. Here we report a single‐molecule approach to monitor the dynamics of E. coli DNA polymerase I (Klenow fragment) on a DNA template during DNA synthesis with single base‐pair resolution. Following nucleotide incorporation, the polymerase is observed translocating past the next pre‐insertion site, a distance equivalent to approximately two bases, and then backs up one base. Rate differences are observed between incorporation of pyrimidines and purines. When a mismatched primer/template junction is used, we observe the primer moving into the polymerase exonuclease site at a position clearly distinct from the translocation site. A revised model for DNA polymerase mechanism is proposed that includes a post‐insertion fidelity checking site that monitors the formation of a complementary nascent base‐pair as part of the proofreading mechanism. This study was supported by grants from the National Institutes of Health (CA40605,GM085116) and the National Science Foundation (MCB0747285).
Effects of Nucleotides on Changes in Flexibility & Allosteric Interactions in Glutamate Dehydrogenase
Authors: Scheel Carolyn , Bell Ellis
Published: June 2021

Higher eukaryotic glutamate dehydrogenases exhibit complex homo and hetero tropic regulation involving subunit interactions and ligand induced conformational changes. Previous have shown that the amino acid substrates, glutamate or norvaline have differential effects on local and global stability of the protein. To investigate the effects of nucleotide cofactors [NAD(H) and NADP(H)] and the allosteric regulators ADP and GTP similar approaches, using Limited proteolysis, Differential Scanning Calorimetry, Dynamic Light Scattering have been used in combination with Isothermal Titration Calorimetry to study ligand induced changes in local flexibility and stability. Overall these studies indicate that both activity and allosteric regulation of this complex enzyme are linked to changes in conformational flexibility rather than discrete ligand induced changes in conformation.

This research is supported by a Grant from the National Science Foundation: MCB 0448905 to EB
Systemic Characterization of Immune Suppression from Battlefield‐like Stress
Authors: Muhie Seid , Hammamieh Rasha , Cummings Christiano , Yang David , Jett Marti
Published: June 2021

This study was designed to determine impact of battlefield‐like stress on the immune response using whole genome profiling and computational analyses. We collected whole blood samples from 15 US Army Ranger Training Battalion cadets before, and after 5 and 8 weeks training. Genome wide transcriptomes were analyzed using oligo‐nucleotide arrays representing ~24,700 human genes. More than 2300 differentially regulated genes were identified at both time points. Ontological enrichment showed suppression of transcripts important in microbial pattern recognition, inflammation, chemotaxis, and antigen preparation and presentations. Profiled genes and pathways were confirmed using both quantitative real time PCR arrays and high throughput ELISAs. Our computational analyses (for regulatory motifs and networks) identified the stress response site (STRE) as common regulatory motif of a number of profiled genes. Though, population of mono‐nuclear cells was not changed significantly, in vitro exposure of leucocytes to Staphylococcus enterotoxin B, a pyrogenic toxin, showed no response, and they seemed anergic. This may mean a highly compromised protective immune response. Suppression of most molecules involved in antigen presentation and lymphocytes' activation pathways may explain why individuals or animal models exposed to chronic stress show poor response towards proteineous antigen based vaccines.

We are grateful to DTRA for funding.
Cinnamon extract improves TNFa‐ induced overproduction of intestinal apolipoproteinB 48‐containing lipoproteins
Authors: Qin Bolin , Polansky Marilyn M. , Harry Dawson , Anderson Richard A
Published: June 2021

TNFa, stimulates the overproduction of intestinal apolipoproteins. We evaluated whether a water extract of cinnamon (Cinnulin PF®) improved the dyslipidemia induced by TNFa in Triton WR‐1339 treated hamsters, and whether Cinnulin PF® inhibits the TNFa‐induced over secretion of apoB‐48 in hamster enterocytes. In vivo, acute oral intake of Cinnulin PF® (50 mg per kg BW), inhibited the postprandial overproduction of apoB‐48 lipoproteins and serum triglycerides. Ex vivo 35S labeling studies, demonstrated that CE (10 and 20 ?g/mL) inhibited the over secretion of apoB‐48 induced by TNFa treated enterocytes. To determine the effects on the mRNA levels for the related genes of lipoprotein metabolism in enterocytes, fresh isolated primary enterocytes of hamsters were pre‐treated with TNFa(10 ng per mL for 30min), then treated with and without Cinnulin PF® (10ug/mL) for 0.5, 2h and 4h. Quantitative real‐time PCR assays demonstrated that Cinnulin PF® improved the mRNA expression of IR, IRS1, IRS2, PI3K and Akt1and decreased the mRNA expression of MTP, PTEN, IL1a and b, IL6 and TNFa in TNFa treated enterocytes. In summary, our results suggest that a water extract of cinnamon reverses TNFa‐induced overproduction of intestinal apoB‐48 via regulating the gene expression of components of the insulin signaling, lipoprotein and inflammatory pathways. (Funded by USDA/ARS and Integrity Nutraceuticals).
Effect of mixed carotenoid vs. single carotenoid supplementation on lipid peroxidation in post‐menopausal women
Authors: Yeum K‐J , Yoon G‐A , Yoshida Y , Niki E , Aldini G , Rasmussen H , Lian F , Wang X‐D , Russell R M
Published: June 2021

The effects of supplements containing individual vs. mixed carotenoids on oxidative stress were determined. Healthy, post‐menopausal women (n=37, 50‐70 yrs) were assigned to one of five groups to be supplemented with either placebo, mixed carotenoids (a combination of β‐carotene, lutein and lycopene at 4 mg/d each) or 12 mg/d of single carotenoid (β‐carotene, lutein or lycopene) in this double blind placebo‐controlled randomized study. Plasma fat‐& water‐soluble antioxidants were analyzed by HPLC‐UVD & ‐ECD, respectively; total antioxidant capacity by fluorometry; lipid peroxidation by malondialdehyde using a HPLC and by total hydroxyoctadecadienoic acids (tHODE) using a GC/MS. Although the individual carotenoid supplemented (12 mg/d) groups showed a significant increase in plasma concentration of each carotenoid supplemented (p<0.05), no significant change in any biomarker of antioxidant activity or lipid peroxidation was observed. The mixed carotenoid group showed a significant decrease in lipid peroxidation at one of two time points during treatment, as determined by linoleic acid‐adjusted tHODE (p<0.05), while the placebo group showed an increase in cholesterol‐adjusted 7‐hydroxycholesterol (p<0.05) at both time treatment points. The current study indicates oxidative stress can be modestly modulated by either a single or a mixed carotenoid supplement and whether the mixed carotenoids have an advantage remains an open question. (Supported by USDA # 58‐1950‐7‐707 & BASF Ag, Germany)
Cardiometabolic risk factors in Puerto Rican adults
Authors: VanRompay Maria I. , Castaneda‐Sceppa Carmen , McKeown Nicola M , Ordovas Jose M , Tucker Katherine L
Published: June 2021

Puerto Ricans have prevalent type 2 diabetes (T2D), but few studies have examined this population. We classified 1289 Puerto Rican adults in the Boston area, aged 45 to 75 years, by T2D, impaired fasting glucose (IFG), and normal fasting glucose (NFG). We found high prevalence of T2D (39%) and IFG (24%), which were associated with lower education and physical activity, and greater lipid‐lowering and antihypertensive medication use, relative to NFG. Overall obesity (BMI = 30) and central obesity (waist circumference (WC) > 102 cm, men; > 88 cm, women) were prevalent, and higher with T2D (67.2% and 79.3%, respectively) and IFG (57.3% and 70.4%) than NFG (43.9% and 61.2%, each P < 0.0001). Of the total sample, 59‐73% had low HDL cholesterol (= 40 mg/dl men; = 50 women), 33‐70% hypertriglyceridemia (= 200 mg/dl), and 12‐32% both low HDL and hypertriglyceridemia. Of those with T2D, 73.6% had HbA1c = 7%, and 66.3% systolic blood pressure = 130 mmHg, despite frequent use of oral hypoglycemic (55.5%), insulin (25.9%) and antihypertensive medications (76.3%). Homeostasis model assessment for insulin resistance (HOMA‐IR) was significantly associated with BMI and WC and, inversely, with HDL in those with IFG and NFG. Interventions to improve the risk profile in this group are needed.

This study was supported by NIH PO 5P01‐AG023394 and by the USDA, Agriculture Research Institute agreement number 58‐1950‐7‐707.
The association of pre‐sweetened (PS) or non‐PS (NPS) ready to eat cereal (RTEC) or other breakfast (OB) with nutrient intake, diet quality, and body weight measures of children and adolescents aged 9 to 18 years: NHANES 1999‐2002
Authors: O'Neil Carol E. , Zanovec Michael T. , Cho Susan S. , Nicklas Theresa A.
Published: June 2021

The objective of this study was to examine the association of a PS or NPS RTEC or OB with nutrient intake, diet quality, and body weight measures of children aged 9‐13 (y) (n=1790) and 14‐18y (n=1826) NHANES, 1999‐2002. Nutrient intakes, mean adequacy ratios (MAR) (average percentage of the Estimated Average Requirement from 24 hour recalls of 13 vitamin and minerals), and weight of PSRTEC, NPSRTEC, and OB consumers were compared. Means + standard error were generated and regression analysis was performed adjusting for covariates. Breakfast varied by age: for 9‐13y vs. 14‐18y, 27% and 15% ate PSRTEC; 7% and 5% ate NPSRTEC, and 46% and 47% ate OB. Energy and total/added sugars were highest in PSRTEC; for 9‐13y total/added sugars were 30.6% and 21.4% of total energy and for 14‐18y total/added sugars were 29.9% and 21.1%, respectively. Total fat was lowest in 9‐13y NPSRTEC and 14‐18y PSRTEC consumers. Vitamins B6 & B12, thiamin, riboflavin, niacin, and folate were higher in RTEC than OB consumers. Calcium, potassium, magnesium, iron, and phosphorus were highest in NPSRTEC consumers compared to the two other groups. For both 9‐13y and 14‐18y, MAR was higher in RTEC than OB consumers. Body Mass Index (BMI) in OB was higher than that in NPSRTEC for both groups. Nutrient profiles vary depending on the type of breakfast consumed. Supported by USDA and Kellogg's Corporate Citizenship Fund.
Whole grain consumption and body weight measures in children and adolescents: Results from NHANES 1999‐2004
Authors: ZANOVEC MICHAEL , O'Neil Carol E. , Cho Susan S. , Nicklas Theresa A.
Published: June 2021

The objective of this study was to examine the relationship of whole grain (WG) consumption and anthropometric measures in children and adolescents using a secondary analysis of cross‐sectional data from the National Health and Nutrition Examination Survey 1999‐2004 in children and 6‐12 y (n=3,868) and adolescents13‐18 y (n=4,931) that were categorized by WG consumption: < Mean number of servings, = Mean to < 1.5, = 1.5 to < 3, and = 3 servings of WG. Outcome measures were mean body mass index (BMI), BMI percentiles, BMI z‐scores, and waist circumferences (WC). Analysis of covariance, adjusted for gender, ethnicity, cereal fiber and total energy intake was used to estimate body measures. Significance was p = 0.05. Mean WG intake was 0.59 and 0.63 servings/day in children 6‐12 y and adolescents13‐18 y, respectively. In children 6‐12 y, WG intake was not associated with BMI or WC. In adolescents 13‐18 y, BMI but not WC was significantly lower in the highest WG consumption group. Overall consumption of WG was below current recommendations of = 3 servings per day. Only in adolescents was this level of intake associated with lower BMI z‐scores. Supported by USDA and Kellogg's Corporate Citizenship Fund.
Effect of high omega‐3 fatty acid diet on markers of breast cancer risk in postmenopausal women
Authors: Orr Lindsay Rae , Redmon J Bruce , Kurzer Mindy S , Raatz Susan K
Published: June 2021

Objective

The primary objective is to determine whether diets designed to increase plasma omega‐3 fatty acid (n3) levels (a low fat diet, with or without n3), will affect sex hormone distribution in postmenopausal women in a direction associated with reduced risk of breast cancer.

Methods

We are conducting a well‐controlled feeding study to evaluate dietary fat and fatty acid effects. The diets tested in 8 week feeding periods include a "high risk" American diet (40% fat) (HF), a low fat diet (20% fat) (LF) and a low fat diet with 3% n3 (23% fat) (LFn3). Plasma estradiol (E2), estrone (E1), estrone sulfate (E1‐S), testosterone (T), androstenedione (AS), sex hormone binding globulin (SHBG), follicle stimulating hormone (FSH), dehydroepiandrosterone (DHEA), and dehydroepiandrosterone sulfate (DHEAS) from baseline and 8 weeks of each diet were analyzed by radio‐immunoassay for 10 participants.

Results

SHBG levels were significantly increased at 8 weeks with LFn3 compared to LF (p < 0.05), and there was a trend for decreased DHEAS level at 8 weeks with LFn3 compared to LF (p < 0.15). A trend for increased E2 was observed with HF compared to both LF and LFn3 at 8 weeks (p < 0.15).

Conclusions

Preliminary data suggest that LFn3 alters estrogen metabolism in a direction associated with reducing breast cancer risk in postmenopausal women.

This study was supported by DOD W81XWH‐06‐1‐0778, DOD W81XWH0410448, NIH/NCRR MO1‐00400.

Grant Funding Source

DOD W81XWH‐06‐1‐0778, DOD W81XWH0410448, NIH/NCRR MO1‐00400
Almonds ameliorate risk factors of cardiovascular disease in type 2 diabetes
Authors: Chen Chung‐Yen Oliver , Liu Jen‐Fang , Liu Yen‐Hua , Chen Chiao‐Ming
Published: June 2021

Almond consumption is associated with ameliorations in obesity, hyperlipidemia, hypertension, and hyperglycemia. The objective of this 12‐wk crossover clinical trial was to investigate the effect of almonds on risk factors of cardiovascular disease in type 2 diabetic patients with mild hyperlipidemia. After a 2‐wk run‐in period, 24 Chinese patients were randomly assigned to either a NCEP step II diet (C) or almond diet (A) for 4 wks with a 2‐wk washout period between alternative diets. Almonds were added to the C diet to replace 20% of total daily calorie intake. Addition of approximately 60 g almonds/d increased dietary intakes of fiber, magnesium, PUFA, MUFA, and vitamin E. As compared to the baseline, the A diet led to significant decreases in body fat by 1%, HOMA‐IR by 1.1%, blood sugar by 6.7%, insulin by 7.9%, total cholesterol by 8%, and LDL‐C by 13%. Almonds also increased plasma α‐tocopherol by 20% and resistance of LDL against ex vivo oxidation by 26%. Inflammatory biomarkers, IL‐6, TNF‐α, and CRP, were ameliorated by the A diet by 14, 35, and 41%, respectively. The C diet improved some of the risk factors of cardiovascular disease but not as magnificent as the A diet. Our results suggested that incorporation of almonds into a healthy diet could decrease risk of cardiovascular disease in type II diabetes. This study was funded by the Almond Board of California and USDA Agricultural Research Service.
Momordica charantia improves body weight and glucose tolerance in mice fed high fat diet.
Authors: Pina Fatima , Brown Angela , Luebcke Emily , Perkins‐Veazie Penny , Clarke Stephen , Kuvibidila Solo , Hill Molly , Lightfoot Stanley , Smith Brenda , Lucas Edralin
Published: June 2021

Obesity and diabetes are major public health concerns in the US and worldwide. A high fat diet contributes significantly in the development of these diseases. Momordica charantia (MC), also known as bitter melon, is a widely consumed vegetable in Asia and has hypoglycemic properties. This study compared the effects of freeze‐dried MC with the PPARγ agonist‐ rosiglitazone (rosi), and the known PPARα agonist‐ fenofibrate (feno), on body weight and glucose using a mouse model of diet‐induced obesity. Eight wk old male C57BL/6 mice were randomized to 8 dietary treatment groups (n=20/group): control diet (ad lib), control diet (pair fed), high fat (HF) diet, HF + 1% MC (w/w), HF + 10% MC (w/w), HF+1% MC seeds (w/w), HF + rosi (50mg/kg diet), and HF + feno (500mg/kg diet) for 8 wks. Significant differences in body weights were observed within one week of beginning the experimental diet. The final body weights of mice receiving the 10% MC were similar to the mice receiving the control diet (ad lib). Rosiglitazone and 1% MC were not able to reduce body weight; however, fenofibrate and MC seeds mildly reduced body weight. HF fed mice exhibited the highest percent body fat. Interestingly, 10% MC prevented the increase in adiposity due to high fat diet. A high fat diet containing 10% MC, similar to rosiglitazone, normalized blood glucose after a glucose tolerance test. Fenofibrate caused an enlargement of liver which was not observed in other treatment groups. The mechanism by which MC modulates glucose and body weight warrants further investigation. (Supported by USDA, #2008‐35200‐18692)

Grant Funding Source

USDA
Differential proteome profiling analysis of male and female murine skeletal muscle following a single bout of exercise
Authors: Gan Yutian , Kulp Mohini , Smith Laura , Scordilis Stylianos P
Published: June 2021

Identifying the array of proteins that change expression following a single bout of eccentrically‐biased exercise in skeletal muscle will shed light on the molecular mechanisms underlying adaptation, as well as their gender specificity. Exercise‐naive male and female mice (C57BL/10ScSnJ 000476 +/+) ran downhill (‐15°) on a treadmill at 25 m s−1 for 15 min and 800 ug of total biceps brachii extract was electrophoresed on pH 5‐8 2‐D gels. The resulting spots that changed at least +/‐ 2‐fold and were significantly different (p<0.05) relative to unexercised controls of the appropriate gender using PDQuest 8.0 were analyzed by LC/MS‐MS and identified using BioWorks 3.3.1. This analysis showed significant proteome remodeling subsequent to downhill running. The expression change patterns were followed at 0, 24, 48, 72 and 168 hrs post‐exercise (PX) and significant changes were observed in 4 NCBI KOG groups. Both myofibrillar and cytoskeletal protein remodelings were noted, as well as decreased expression of creatine kinase. Nearly all of the glycolytic enzymes were up‐regulated. The cellular heat shock protein responses were significant and specific. Roughly the same number of protein spots changed in females and males, but the females peaked at 24 hr PX, whereas in males the peak was at 72 hr PX. Supported by NSF 0420971, the Smith College Blakeslee Fund and HHMI.
The regulation of matrix metalloproteases by NHE1 in non‐small lung cancer cells
Authors: Haak Andrew , Provost Joseph J , Wallert Mark A , Dorsam Glenn
Published: June 2021

The Na+‐H+ exchanger isoform‐1 (NHE1) has plays a major role in the coordination of cell migration and cancer metastasis. One possible mechanism is based on a family of proteases, matrix metalloproteases (MMPs), which degrade the extracellular matrix allowing cells to escape tumors and migrate. Studying three non‐small cell lung carcinoma cell lines (NCI‐H358, ‐H460, ‐1299) we have demonstrated using zymogen gels that stimulation with urokinase plasminogen activator (uPA) leads to an increase in MMP9 activity. Additionally, when NHE1 is inhibited with ethylisopropyl amiloride (EIPA), MMP9 activity decreases. Here we investigated whether the increase in MMP9 activity was due to increased activation or increased expression of MMP9. We hypothesized that NHE1 drives an intracellular alkalization that promotes MMP‐9 expression. To determine this action, stimulated and EIPA inhibited cells were analyzed for the level of RNA transcription measured by qPCR. Expression of MMP9 in H460 and H358 show a 3‐4 fold increase in transcription while H1299 displays a 100 fold increase in MMP‐9 transcription when stimulated with uPA. MMP1 transcription displays a 14 and 66 fold increase in H460 and H358 under NHE1 inhibition, but the H1299 showed no increase in MMP‐1 with EIPA treatment. This work was supported by NIH grant 1R15CA135616‐01, and NSF grant MCB‐0817784.
Protein Splicing by a Non‐Canonical Mechanism
Authors: Reitter Julie , Schufreider Ann K. , Mills Kenneth V.
Published: June 2021

Protein splicing is a post‐translational process whereby an intervening polypeptide (intein) facilitates its own excision while simultaneously ligating the flanking protein sequences (exteins) with a peptide bond. Typical protein splicing begins with an amide‐to‐ester rearrangement of the scissile peptide bond at the N‐terminus of the intein followed by a nucleophilic attack from a Cys, Thr or Ser residue adjacent to the C‐terminus of the intein. The intein that interrupts the putative phage terminase from Clostridium thermocellumpromotes efficient splicing although it cannot promote the first step of protein splicing. We have examined the influence of residues flanking the scissile peptide bonds at both ends of the intein. Wildtype and mutated fusion proteins were expressed in BL21(DE3) cells and purified. We have found that the intein can promote splicing with either a nonpolar, hydrophobic or nucleophilic residue at the N‐terminus of the intein. However, splicing was eliminated by mutation of either residue in the downstream scissile bond. A positively charged residue was required at the C‐terminus of the upstream extein. This material is based upon work supported by the National Science Foundation under grant no. 0320824 and CAREER grant no. 0447647.
Nitric oxide signaling depends on biotin in Jurkat human lymphoma cells
Authors: Rodriguez‐Melendez Rocio , Zempleni Janos
Published: June 2021

Biotin affects gene expression through various signaling pathways. Previous studies suggest that cGMP‐dependent signaling also depends on biotin, but the mechanism of cGMP regulation is unknown. Here we tested the hypothesis that effects of biotin in cGMP‐dependent cell signaling are mediated by nitric oxide (NO). Human lymphoid (Jurkat) cells were cultured in media containing deficient, physiological, and pharmacological concentrations of biotin. Both levels of intracellular biotin and NO exhibited a dose‐dependent relationship in regard to biotin concentrations in culture media. Effects of biotin on NO levels were disrupted by both the NO synthase (NOS) inhibitors and NO donors. Biotin‐dependent production of NO was linked with biotin‐dependent expression of endothelial and neuronal NOS, but not the inducible iNOS. Previous studies revealed that NO is an activator of guanylate cyclase. Consistent with these previous observations, biotin‐dependent generation of NO increased the abundance of cGMP in Jurkat cells. Finally, the biotin‐dependent generation of cGMP increased the activity of protein kinase G. Collectively this study is consistent with the hypothesis that biotin‐dependent cGMP signaling in human lymphoid cells is mediated by NO. (Supported by UNL ARD Hatch Act funds, NIH DK063945, DK077816 and ES015206, USDA CSREES 2006‐35200‐17138, and NSF EPSCoR EPS‐0701892.)
Nutrition‐related self‐efficacy and outcome expectancies of freshman women
Authors: Kedem Leia Ellen , Klamm Emily L. , Evans Ellen M. , McAuley Edward , Chapman‐Novakofski Karen
Published: June 2021

The PEER project is a peer‐led intervention to promote healthy lifestyles in freshman women. Baseline data (n=157, wave‐1) included 48 items for self‐efficacy (SE) and 30 items for outcome expectancies (OE). Within SE for fruits (F), vegetables (V), low‐fat foods (LF), and high‐calcium foods (HC), strongest correlations (R) were between not liking to eat the food/not feeling like eating the food (r=.856; r=.810; r=.879; r=.878, p<.01, respectively). For V and LF, the weakest R's were between limited availability/being very hungry (r=.211, p<.01; r=.197, p<.05). The weakest R for SE in F was between no availability/when it's work to prepare (r=.238, p<.01). For HC, the weakest R was between having no HC you like/being very hungry (r=.336, p<.01). Most items for SE for making healthier food choices were significantly correlated. OE for F&V, LF, and HC had significant positive and negative R's. For eating >5 servings of F&V and >3 servings of HC, the strongest R was between feeling I'm being good to myself/other people would think I'm healthy (r=.587, p<.01; r=.771, p<.01). For eating LF every day, the strongest R was between I would be able to keep my weight where I want it/feeling that I'm being good to myself (r=.755, p<.01). Within each food type, "food would cost too much" was not correlated often. Most subjects worried about weight gain and were concerned about their health now, indicating health consciousness. Funded by USDA.
What's in that bottle? Accuracy of infant formula reconstitution among exclusively formula‐feeding mothers in East Tennessee
Authors: Kavanagh Katie F , Springer Cary
Published: June 2021

The project purpose was to assess accuracy of reconstitution of powdered infant formula among exclusively formula‐feeding mothers. Properly reconstituted infant formula provides ~20 kcal/oz (acceptable range: 18‐22 kcal/oz). Over‐dilution of infant formula may lead to an infant who cries more and continues to exhibit hunger cues, potentially leading to early introduction of other foods and fluids and predisposing to child overweight. Mothers of healthy, term infants < 4 mo of age, using powdered infant formula, were recruited from the WIC program and from a university‐housed birth registry. Mothers recorded infant intake for 24 hours and collected a 15 mL sample of infant formula from each bottle offered prior to offering the bottle to the infant. Upon collection of samples mothers were asked several questions regarding preparation of infant formula. Samples were analyzed using the loss on drying method. Twenty‐four hours of samples were collected from 18 mothers and results indicate that, though the average kcal/oz was 18.5, 26% of mothers were consistently over‐diluting the infant formula (<18 kcal/oz), with an overall average range of 14.5‐23 kcal/oz. In addition, results of the questionnaire indicate behaviors may be both intentional and unintentional. It appears that over‐dilution is a potential issue in this formula‐feeding population. Funds: USDA ERS RIDGE Program, Southern Rural Development Center.
Characterization of porcine betaine homocysteine methyltransferase (BHMT) and betaine homocysteine methyltransferase ‐2 (BHMT2) genes
Authors: Ganu Radhika Sudhir , Garrow Timothy A , Sodhi Monika , Schook Lawrence B
Published: June 2021

Betaine homocysteine methyltransferase (BHMT) and BHMT‐2 methylate homocysteine using betaine or S‐methylmethionine, respectively. These enzyme activities are found only in the liver of adult rodents, but in the liver and kidney of adult humans and pigs. Using a pig model, we seek to understand the developmental expression pattern of these enzymes in different porcine organs. Immunohistochemical staining revealed the presence of BHMT in adult liver and kidney cortex, as reported earlier, and also in fetal lungs (aged days 30, 60, 84, 90, 105). Full length cDNA of BHMT and BHMT‐2 were cloned and sequenced, and the 5' and 3' UTRs were amplified using RLM‐RACE. BHMT has a longer 5' and 3' UTR, consisting of 77 and 1142 nucleotides, whereas BHMT‐2 consisted of 17 and 893 nucleotides respectively. The predicted primary sequences of BHMT and BHMT‐2 contain 407 and 363 amino acids, respectively, and share 78% amino acid identity. Relative to BHMT‐2, BHMT has two additional regions of amino acid sequence; a 9 amino acid sequence (86‐94) in the N‐terminal region, and 34 amino acid sequence (373‐407) at the carboxy terminus. Preliminary sequencing results identified two isoforms of liver BHMT gene resulting from alternative splicing of 5' UTR. BHMT enzymes are organ/developmental stage‐dependent and further sequence analysis might reveal new splice variants that result in different protein functions.

Grant Funding Source

USDA AG 2008‐34480‐19328 & USDA‐ARS 538 AG58‐5438‐7‐317l
Cloning and characterization of 3‐hydroxyacyl‐CoA dehydrogenase from Micrococcus luteus WIUJH20
Authors: Huang Jenq‐Kuen , Dhungana Babu R , Robinson Stephanie A , Wen Lisa
Published: June 2021

Soapstock is a by‐product of edible oil refining process with low cash value. It is rich in oleic and linoleic acids which can be converted to value‐added compounds by certain microorganisms. Nocardia cholesterolicum NRRL 5767 (N. cholesterolicum NRRL 5767) is a well‐known microbe that can transform oleic acid to 10‐hydroxyoctadecanoic acid (10‐HOOD, 85%) and 10‐oxo‐octadecanoic (10‐OXOD, 10%). The 10‐OXOD contaminant complicates downstream purification of 10‐OHOD. The conversion reactions are catalyzed by oleate dehydrogenase and secondary alcohol dehydrogenase. The long‐term goal is strain improvement of N. cholesterolicum NRRL 5767 by blocking 2‐ADH to create an industrially useful microbe for the production of 10‐OHOD from oleic acid. M. luteus was used for 2‐ADH purification due to its lack of oleate hydratase which hampered the purification of 2‐ADH from N. cholesterolicum NRRL 5767.

The 2‐ADH from M. luteus WIUJH20 has been purified to homogeneity and its N‐terminal sequence determined. Using BLAST search tool, the 2‐ADH gene has been identified as belonging to the 3‐hydroxyacyl CoA dehydrogenase superfamily. The gene has been amplified by PCR and cloned into pGEX2T. The recombinant 2‐ADH protein has been overexpressed in E. coli, purified, and its Km and Vmax determined.

Supported in part by grants from USDA CSREES 04‐35504‐14712 and 02‐015470.
Molecular, mass spectral, and physiological characterization of a novel C‐type allatostatin peptide, SYWKQCAFNAVSCFamide, in crustaceans
Authors: Ackerman Rachel J. , Wiwatpanit Teerawat , Stevens Jake S. , Dickinson Patsy S. , Stemmler Elizabeth A. , Christie Andrew E.
Published: June 2021

The allatostatins comprise three structurally distinct families of neuropeptides classified as A‐, B‐, and C‐type. Known to play a role in the regulation of juvenile hormone production by the insect corpora allata, members of the A‐ and B‐type allatostatin family are found in a wide variety of arthropods; however, members of the C‐type family have been only identified in insects. C‐type allatostatins typically possess a ‐PISCF motif. Here, we report on the characterization of the first C‐type allatostatin in crustaceans, SYWKQCAFNAVSCFamide, with bridged cysteines. Using in silico mining of extant express sequence tags (ESTs) from the lobster, Homarus americanus, transcripts encoding the prepro‐hormone sequence were used to predict the mature peptide. This peptide was subsequently detected in crustacean tissue samples from species representing five different decapod infraorders using MALDI‐FTMS, suggesting broad sequence conservation. In H. americanus, a synthetic form of the peptide has been tested and found to be bioactive. Support from: MDIBL; NSF (IBN 01140, MRI‐0116416); NIH P20 RR‐016463, NCRR; HHMI Undergrad Sci Prog.
Effect of age on proteasomal activity of T cells and macrophages
Authors: ren zhihong , Shang Fu , Meydani Simin Nikbin , Wu Dayong
Published: June 2021

T cell function is impaired with aging. Proteasome activity in T cells is important for T cell activation and its activity in macrophages is required for processing antigens in order to be presented via class I major histocompatibility complex to CD8+ T cells. Since studies have demonstrated that proteasome activity decreases with aging in several tissues, we determined whether there is an age‐related change in proteasome activity in T cells and macrophages (Mφ). T cells and Mφ were isolated from young (4‐6 mo) and old (24‐26 mo) C57BL/6 mice. There was no significant age‐related difference in proteasome activity in unstimulated T cells while the anti‐CD3/CD28‐stimulated T cells from old mice had lower proteasome activity compared to those from young mice. Proteasome activity in both young and old Mφ increased slightly 15 and 30 min after LPS stimulation, returned to basal level by 120 min, and significantly decreased by 24 hrs of stimulation. The lower proteasome activity in Mφ stimulated with LPS for 24 hrs was not due to a reduction in the amount of proteasome proteins suggesting a post‐translational regulation. There was no difference between young and old mice in proteasome activity in either unstimulated or LPS‐stimulated Mφ. These results demonstrate that there is an age‐related decrease in proteasome activity in T cells, but not in Mφ. Further studies are needed to determine the relevance of these findings to the age‐related decline in T cell function. Supported by USDA #58‐1950‐7‐707 and HNRCA Pilot grant.
Inflammation status in adipose tissue and peritoneal macrophages of young and old mice in diet‐induced obesity
Authors: Wu Dayong , ren zhihong , Pae Munkyong , Han Sung Nim , Meydani Simin Nikbin
Published: June 2021

Obesity causes a low grade inflammation. We have shown that aging is also associated with upregulated inflammation in adipose tissue. In this study, we investigated effect of diet‐induced obesity on inflammation status in young and old mice. Young (2‐mo) and old (19‐mo) C57BL/6 mice were fed low fat (10%, LF) or high fat (60%, HF) diet for 5 mo. While body weight increased in both age groups, young mice gained more than the old. Of 13 mice in each age and diet group, none of the young and 2 old mice in LF group, and 2 young and 4 old mice in HF group died before the end of study. Adipose tissue from old LF mice expressed higher levels of IL‐1β, IL‐6, TFN‐α, and cyclooxygenase‐2 mRNA compared with young LF mice. Expression of all these inflammatory markers increased in young HF mice to levels comparable to those in old LF mice while old HF mice had lower expression of these markers compared with those in old LF mice. Adipocytes from old LF mice produced more IL‐6, TNF‐α, and prostaglandin (PG)E2 than those from young LF mice. HF resulted in increase of all of these markers in young, but only PGE2 in old mice. No significant diet‐ or age‐related difference was found in these markers in stromal vascular cells. PGE2 produced by peritoneal macrophages was upregulated with aging and HF induced more production of IL‐6 and PGE2 in young but not in old mice. Thus, obesity induces an inflammatory state in both visceral fat and peritoneal macrophages in young mice. The lack of such changes or even a decrease in some of inflammatory markers in old mice may be due to deteriorated health condition and loss of weight in some old mice fed HF. Supported by USDA #58‐1950‐7‐707.
Simple indicators of dietary diversity are associated with micronutrient adequacy for women of reproductive age in resource‐poor settings
Authors: Arimond Mary , Wiesmann Doris , Becquey Elodie , Daniels Melissa , Fanou Nadia , Kennedy Gina , Martin‐Prevel Yves , Torheim Liv Elin
Published: June 2021

Women of reproductive age living in resource‐poor settings are at high risk of inadequate micronutrient intakes when diets lack diversity and are dominated by staple foods. Yet comparative information on diet quality is scarce, and quantitative data on nutrient intakes is expensive and difficult to gather. We assessed the potential of simple indicators of dietary diversity, such as could be generated from household surveys, to serve as proxy indicators of micronutrient adequacy for population‐level assessment. We used 5 existing data sets (3 from Africa; 2 from Asia) with repeat 24‐hr recalls to construct 8 candidate dietary diversity scores (DDS) and to calculate the mean probability of adequacy (MPA) for 11 micronutrients. DDS varied in food group disaggregation and in minimum consumption required for a food group to count. There were large gaps between micronutrient intakes and requirements in each site. All 8 DDS were correlated with MPA in all sites; regression analysis confirmed that associations remained when controlling for energy intake. Assessment of dichotomous indicators through receiver‐operating characteristic analysis showed moderate predictive strength for the best choice indicators. Simple DDS hold promise as proxy indicators of micronutrient adequacy. [Support: USAID, through FANTA/AED]
Mechanisms of enhanced silencing in rpd3 delete strains of Saccharomyces cerevisiae
Authors: Kleinschmidt Richard Alton , Donze David
Published: June 2021

In Saccharomyces cerevisiae, heterochromatic silencing occurs at the cryptic‐mating loci HMRa and HMLα, ribosomal DNA, and telomeres. Propagation of silent heterochromatin is sequence‐independent mediated by the Silent Information Regulator proteins (e.g. Sir3p) and other factors involved in repression. Rpd3p is part of a larger histone deacetylase complex that represses transcription when targeted by promoter‐specific transcription factors. In yeast, RPD3 deletion surprisingly enhances silencing at the cryptic mating loci and telomeres (Vannier et al. 1996), even overriding the tDNA barrier element adjacent to the HMR locus (Donze and Kamakaka, 2001, Jambunathan et al 2005). To understand the mechanism of enhanced silencing in strains lacking RPD3, we mutagenized rpd3D strains containing an ADE2 marker gene adjacent to HMR and identified suppressor mutants no longer displaying enhanced silencing. We identified seven genes which may affect heterochromatin formation in rpd3Δ backgrounds: BRE1, GDH2, BRE2, GAT3, QNS1, NPT1 and RXT3. After deleting each of these genes in the rpd3Δ background, only BRE1 or BRE2 deleted strains crossed with rpd3Δ strains showed a loss of extended silencing at the HMR and a marked telomere. BRE1 and BRE2 are indirectly and directly involved in the tri‐methylation of histone H3 lysine 4 respectively. Other studies of histone H3 lysine 4 methylation have suggested that mutations in this pathway lead to a re‐distribution of Sir proteins at silenced loci (Madhani et al. 2007). We hypothesize that the increased silencing in rpd3D mutants might also be due to redistribution of chromatin proteins. Future experiments will utilize ChIP‐Seq and DamID methodologies to determine Sir3p occupancy at HMR and genome‐wide euchromatic regions in WT vs. rpd3Δ strains. Support for this project was provided by a grant from the National Science Foundation.
The Effects of Loop Mutations on the Activity and Regulation of Malate Dehydrogenase
Authors: Hedrick Shannon , Bell Ellis
Published: June 2021

Malate Dehydrogenase catalyzes the reversible reduction of Oxaloacetate using NADH, a reaction that plays a critical role in both cytosolic and mitochondrial metabolism. The reaction is regulated by both substrate inhibition and by citrate both of which appear to involve subunit communication in the homodimeric enzyme. To investigate the role that various turn regions in the molecule play in both catalysis and regulation we have, based upon clustal analysis, designed a variety of mutations to probe the role of specific residues in three specific turn regions, two near the subunit interface [residues 91‐95 and residues 203‐209‐ which appear to interact across the subunit interface] and one near the active site [residues 124‐129]. The mutations are expressed using a pQE vector and the proteins purified using Ni‐NTA resin. Initial rate kinetic studies show that various mutants affect Vmax, Km(NADH), Km(Oxaloacetate) or the substrate inhibition observed at high oxaloacetate concentrations as well as the ability of citrate to regulate the activity. The results further implicate subunit interactions as a critical component of the activity and regulation of the enzyme.

Supported by NSF Grant MCB 0448905 to EB
Dietary L‐arginine supplementation can increase expression of vascular endothelial growth factor (VEGF) in early‐weaned pigs
Authors: Yao Kang , Yin Yulong , Li Tiejun , Huang Ruilin , Wu Guoyao
Published: June 2021

This study determined the effects of dietary arginine supplementation on intestinal VEGF expression and growth performance in early‐weaned pigs. Twenty Large White × Landrace piglets (5.3 ± 0.13 kg body weight) were removed from sows at 21 d of age. Piglets were housed individually and assigned randomly to a corn‐ and soybean‐based diet supplemented with 2.05% L‐alanine (isonitrogenous control) or 1% L‐arginine (10 pigs per group). After a 7‐d period of feeding, 6 pigs were selected randomly from each group to obtain the small intestine. VEGF protein in the small intestine was determined using immunohistochemistry. Compared with control pigs, dietary supplementation with 1% L‐arginine enhanced (P<0.05) the villus height of duodenum, jejunum and ileum by 21%, 28% and 25%, respectively, and VEGF protein levels in duodenal mucosa by 14%. Arginine supplementation increased (P<0.05) the average daily gain of piglets by 38%, while decreasing (P<0.05) feed:gain ratio by 28%. These results indicate that arginine supplementation is beneficial for enhancing VEGF expression and the morphology of the small intestine, as well as the growth performance of weaned piglets. Supported by NSFC, USDA and TAMU.
Gender Dimorphism in the Exercise‐naïve Murine Skeletal Muscle Proteome
Authors: Metskas Lauren Ann , Kulp Mohini , Scordilis Stylianos P
Published: June 2021

Skeletal muscle is a plastic tissue with known gender dimorphism, especially at the metabolic level. A proteomic comparison of male (n=5) and female (n=5) murine (C57BL/10ScSnJ000476 +/+) biceps brachii was undertaken, resolving an average of 600 protein spots of MW 15–150 kDa and pI 5–8. 24 unique full‐length proteins spanning 11 KOG groups demonstrated statistically significant (p<.05) abundance differences between genders; the majority of these proteins have metabolic functions. Identified glycolytic enzymes demonstrated decreased abundance in females, while identified oxidative phosphorylation enzyme abundance differences were specific to the proteins rather than to the function group as a whole. Myoglobin and GRP78 showed the largest abundance increases of 3.9 and 5.8‐fold, respectively, whereas dihydrolipoamide S‐succinyltransferase showed the largest decrease of 5.9‐fold in the females. All three phosphorylation states of creatine kinase were decreased in females. Expression differences were significant but many subtle, usually 2‐fold or less. We conclude that while gender dimorphism is present in non‐exercised murine skeletal muscle, the proteome comparison of male and female biceps brachii in exercise‐naïve mice does not indicate any apparent hormonal regulatory basis for the dimorphism. Supported by NSF 0420971, the Smith College Blakeslee Fund and HHMI.
Pre‐partum nutrition extremes impair post‐partum maternal glucose utilization in sheep
Authors: McKnight Jason R. , Satterfield M. Carey , Bazer Fuller W. , Spencer Thomas E. , Wu Guoyao
Published: June 2021

This study determined whether malnourishment and obesity during gestation affect post‐partum maternal glucose utilization. At 120 days prior to gestation, ewes had free access to feed (induction of obesity) or received 100% of NRC nutrient requirements (control group). Beginning on day 45 of gestation, 50% of the obese ewes were restricted to 65% of NRC nutrient requirements. Following parturition, all but 6 ewes (who remained to have free access to feed) were re‐alimented to 100% of NRC nutrient requirements. At post partum day (PPD) 1 and 150, a glucose tolerance test was administered to ewes. At PPD 1 and 150, glucose uptake by both the obese and nutrient‐restricted (NR) ewes was impaired (P<0.05) in comparison with control ewes. Concentrations of glucose in plasma were greater (P<0.05) for NR and obese ewes than control ewes at 10 min post glucose administration, as well as at 15 min for NR ewes and at 30 min for obese ewes. These results indicate that both nutritional extremes negatively reduce glucose utilization by mothers and that this problem cannot be ameliorated by realimentation over a period of 5 months post‐partum. Supported by NIH and USDA grants and TAMU Faculty of Nutrition.

Grant Funding Source: NIH and USDA
Effects of western diet and wheel access on lipid profiles in mice selectively bred for high voluntary wheel running
Authors: Meek Thomas H , Eisenmann Joey C , Keeney Brooke K , Hannon Robert M , Dlugosz Elizabeth M , Garland Theodore
Published: June 2021

Increased consumption of high‐fat foods and a sedentary lifestyle are viewed as two key factors in the ongoing trends for increased rates of human obesity and cardiovascular disease. However, in experimental studies with both humans and rodents, both diet and exercise have shown varying effects on metabolic syndrome risk factors, sometimes depending on genetic background. To examine potential interactive effects of diet, exercise, and genetic background on components of the metabolic syndrome, we studied mice bred for high voluntary wheel running and their respective controls. A standard chow (14% kcal from fat) or Western diet (WD, 42% kcal from fat plus added sucrose) was given to male mice housed with or without wheels for 60 days starting at 24 days of age. Neither genetic selection history nor wheel access had an appreciable impact on the final blood lipid profile. WD was obesogenic for both HR and C, leading to increased retroperitoneal fat and liver mass, even after accounting for wheel running and calories consumed. As expected from results of previous studies with diets high in saturated fats, WD also decreased plasma triglyceride levels. WD significantly increased fasting glucose levels, but raised total and HDL cholesterol concentrations, leading to an improved total/HDL ratio, which may offset some of the negative aspects of the diet. Supported by MSU Seed Grant to JCE and NSF IOB‐0543429 to TG