bluuebunny/crossref_metadata_2025 huggingface dataset:
Rats consuming bran from black and brown sorghums have lower short chain fatty acid concentrations and fewer aberrant colonic crypts
Authors: Turner Nancy D. , Taddeo Stella S. , Lewis Jayme B. , McDonough Cassandra M. , Rooney Lloyd W.
Published: June 2021

Diets containing bran isolated from brown and black sorghum grain produce fewer (68 and 38%, respectively, P < 0.04) high multiplicity aberrant crypt foci (HMACF) in azoxymethane‐ (AOM) injected rats, relative to a cellulose diet. Because of the potential for short chain fatty acids (SCFA) to influence colon carcinogenesis, we determined SCFA concentrations in feces (reported as µM/g wet feces) from rats (n=10/diet) consuming diets containing 6% fiber from either cellulose or bran isolated from white, brown (contains tannins), or black (contains anthocyanins) sorghum. Diets were fed for 10 wk, with two AOM injections (15 mg/kg BW) given in wk 3 and 4. Total SCFA were affected (P < 0.0001) by diet; greatest for white sorghum bran (32.0 µM), intermediate for cellulose (22.6 µM), and lowest for black or brown sorghum bran (15.4 or 14.7 µM, respectively). Butyrate concentrations were affected (P < 0.0001) by diet; highest for white sorghum bran (7.8 µM), lowest for brown sorghum bran (1.07 µM), while concentrations were intermediate and similar for black sorghum bran and cellulose (2.7 and 3.6 µM, respectively). Differences among diets were the same when butyrate was expressed as a percentage of SCFA. These data suggest that there are significant changes either in the microbiota within the lumen or in the transport of SCFA into colonocytes, both of which can influence colon carcinogenesis. Funded by USDA 58‐5430‐5‐339 and NIEHS P30‐ES09106.
Food forms and beta glucans contents in sorghum, fonio and rice can influence their glycemic indexes
Authors: Traore Djibril , Ndoye A , Hamaker Bruce R. , Stoecker Barbara , Betts Nancy , Guiro A. T.
Published: June 2021

Several studies have shown that beta‐glucans have positive effects on cholesterol, glucose and insulin metabolism. In this present study we wanted to know whether beta‐glucan contents in cereals and their food forms could influence glycemic indexes (GI). For that purpose, we determined the glycemic indexes of three cereals grown in Africa alleged to display positive effects on diabetes.

Samples of fonio, sorghum, and rice as reference, were processed into their porridge and couscous forms and given to 15 Senegalese young adult males (22‐25 of age) for Glycemic indexes assessment following the World Health Organization guidelines. Prior to, beta‐glucan contents in selected cereals were determined in previous studies, and during the trial, total bioavailable sugar was assessed using the Megazyme enzymatic method developed by Mc Cleary.

Fonio, which had an overall lower glycemic index (GI =66), contained higher beta‐glucan content than others. Sorghum (GI =72) that ranked second had a higher Beta‐glucan content compared to rice (GI = 95). In addition, this study showed that cereals porridges displayed lower glycemic indexes than their couscous forms, indicative of the fact that food forms may play a critical role in glucose metabolism.

Grant Funding Source

USDA/Institute of Food Science Technology Dakar
Measurement of Menadione in Urine by HPLC
Authors: Al Rajabi Ala , Choi Sang‐Woon , Peterson James , Booth Sarah L
Published: June 2021

Mammals convert phylloquinone to MK‐4, with menadione as a possible intermediate. We developed and validated a method measuring urinary menadione. A high performance liquid chromatography (HPLC) method with a C30 column, fluorescence detection and post‐column zinc reduction was developed. The mobile phase was composed of 95 % methanol and 5 % DI H2O. Aqueous solution (2 M zinc chloride, 1 M acetic acid, and 1 M sodium acetate) was added to both methanol and H2O. MK‐2 was used as an internal standard. The standard calibration curve was linear with a correlation coefficient (R2) of 0.99 for both menadione and MK‐2. Sample preparation involved hydrolysis of menadiol conjugates and oxidizing the released menadiol to menadione. Menadione was then extracted using iso‐octane. Drying extraction solvent prior to HPLC injection resulted in high menadione losses. This was resolved by avoiding complete evaporation. We were able to detect < 0.05 pmole menadione/ injection. Menadione was detected in archived urine samples from subjects receiving 500 µg/d phylloquinone. The assay was validated by "spiking recovery" and by removing the zinc column and observing the disappearance of the menadione peak. This HPLC method presents a sensitive and reproducible way to detect menadione in urine, and can be used to elucidate the role played by menadione in MK‐4 formation. Research support: USDA ARS Cooperative Agreement 58‐1950‐7‐707.

Grant Funding Source

USDA ARS Cooperative Agreement 58‐1950‐7‐707
Differential proteome profiling analysis of male and female murine skeletal muscle following a repeat bout of exercise
Authors: Metskas Lauren‐Ann , Kulp Mohini , Scordilis Stylianos P
Published: June 2021

Skeletal muscle is a plastic tissue that adapts to exercise in many ways. Differential proteome profiling can be used to obtain a global view of proteins that change expression following a repeat bout of eccentrically‐biased exercise, as well as its gender specificity. Exercise‐naive male and female mice (C57BL/10ScSnJ 000476 +/+) ran downhill twice one week apart (‐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 statistically significantly different (p<0.05) relative to the unexercised controls using PDQuest 8.0 were analyzed by LC/MS‐MS and identified using BioWorks 3.3.1. Significant proteome remodeling occurs subsequent to the repeat bout. The expression change patterns were followed at 0, 24, 48, 72 and 168 hrs post‐repeat bout; significant changes were observed in myofibrillar and cytoskeletal proteins, creatine kinase and many of the glycolytic enzymes. The cellular stress responses were unique to individual heat shock and oxidative stress proteins. More protein spots changed in females (299) than in males (236); the female changes peaked at 24 hr post‐exercise, whereas the male changes did not show much variation over the week following the repeat bout. Supported by NSF 0420971, the Smith College Blakeslee Fund and HHMI.
Analysis of oxidative stress in CNS cells by integration of Atomic Force Microscopy (AFM), fluorescence microscopy and amperometry
Authors: D'Agostino Dominic P , Olson James E , Cuevas Javier , Dean Jay B
Published: June 2021

Three sensitive methodologies were used to measure and characterize hyperoxia‐induced oxidative stress in CNS cells (in vitro). A commercially available AFM (Bioscope SZ; Veeco) was mated with an inverted fluorescence microscope (Nikon TE2000‐E) for simultaneous acquisition of AFM and fluorescence data. The system is housed in a hyperbaric chamber (Riemers Systems Inc.), allowing for cellular measurements at normobaric and hyperbaric pressures (0‐85 psig), including hyperbaric oxygen (HBO2). AFM was used to resolve changes in membrane ultrastructure in response to oxidative stimuli (e.g. hyperoxia, H2O2). Fluorescence microscopy was used to detect superoxide with Dihydroethidium (DHE). An amperometric biosensor (ISO‐HPO‐100; WPI) was used to measure nM levels of H2O2. Results from AFM scans of hyperoxia‐treated glutaraldehyde‐fixed (1%) U87 glioblastoma cells revealed nanoscopic membrane surface blebbing. Hyperoxia (95% O2 and 3.5 ATA O2) induced a dose‐dependent increase in average membrane roughness (Ra), which correlated with elevated malondialdehyde (MDA) production. Similar results in Ra levels and MDA production were observed with exogenous H2O2 (0.2‐2mM). Amperometric biosensors detected increased cellular H2O2 production during hyperoxia, which was elevated 156 ± 8% (95% O2) and 276 ± 52% (4 ATA O2) above baseline (20% O2). In conclusion, these novel techniques (used alone or in combination) have provided a highly sensitive and complimentary means to detect reactive oxygen species (ROS) and assess oxidative stress from hyperoxia. ONR grant N000140610105 (DPD), ONR‐DURIP equipment grant N000140210643 (JBD)
Chronic hypoxia influences the role of extracellular Ca2+ influx during serotonin‐mediated contractility of pulmonary arteries from adult sheep
Authors: Goyal Ravi , Vemulakonda Srilakshmi , Mittal Ashwani , Loftin Matthew , Vranken Kurt , Nguyen Dan , Blood Arlin B , Pearce William J , Longo Lawrence D , Wilson Sean M
Published: June 2021

Chronic hypoxia (CH) influences Ca2+‐dependent pulmonary arterial (PA) reactivity, and enhanced vasoreactivity is a risk factor in pulmonary disease. Using wire‐myography, Western immunoblot, and Ca2+ imaging approaches of PA isolated from adult sheep housed under normoxic conditions or at 12,470 feet for ~ 110 days (CH), the hypothesis that CH alters Ca2+‐dependent contractility was tested. Serotonin (5‐HT; 10 μM) dependent PA contractility in CH was dependent on extracellular Ca2+, and 10 mM Ni2+ inhibited contractility and Ca2+ signaling. However, select Ca2+ entry pathways were altered by CH. CH reduced the role for L‐type Ca2+ channels (CaL) as vasorelaxation to 10 μM nifedipine was blunted and α‐subunit protein expression was reduced, although NIF reduced Ca2+ signals to control levels in CH. CH also reduced the role for non‐selective cation channels (NSCC), as determined by serial application of 50 μM SKF 96365 to NIF treated arteries. Similarly, inhibition of reverse mode‐Na+‐ Ca2+ exchange (NCX) with 10 μM KB‐R7943 reduced contractility more dramatically in PA from normoxic sheep. Given extracellular Ca2+ removal and Ni2+ reduced contractility in CH while the roles for CaL, NSCC, and NCX were restricted, SKF‐insensitive basal Ca2+ influx appears important to 5‐HT‐dependent contractility in PA from CH sheep. Support from NIH, NSF, UM, and LLUMC.
Changes in pulmonary arterial smooth muscle structure with maturation and chronic hypoxia in sheep
Authors: McCrate Brian , Webster Melissa , Osman Noah , Nguyen Dan , Goyal Ravi , Pearce William J , Longo Lawrence D , Wilson Sean M
Published: June 2021

Chronic hypoxia (CH) can cause structural changes in the lung where the smooth muscle layer thickens due to myocyte hypertrophy and hyperplasia. CH compromises pulmonary arterial (PA) contractility and Ca2+‐signaling in the sheep fetus as well as the non‐pregnant ewe. Yet, the influence of CH on sheep PA and myocyte structure is unresolved, which is important as this relates to pulmonary pathologies. We therefore tested the hypothesis that CH thickens the smooth muscle layer and causes myocyte hypertrophy by evaluating the structure of PAs isolated from late‐gestational fetuses or adults maintained under normoxic conditions or exposed to CH by housing animals at 12,470 ft for ~ 110 days. To visualize smooth muscle cells, arterial segments were stained with anti‐α‐smooth muscle actin while nuclei were stained with DAPI. 3‐D images were made on a laser scanning confocal microscope by optical sample sectioning. The thickness of the smooth muscle layer was determined, as was myocyte and nuclear length, width, circumference, area and cell density. Fetal cells were smaller than adult and CH induced cellular hypertrophy in fetal and adult myocytes, although CH did not thicken the smooth muscle layer or alter cell density. Similarly, CH caused nuclear enlargement. Overall, these CH‐dependent changes in cell and nuclear morphometry are consistent with PA pathogenesis. Support from NSF, NIH, UM, LLUMC.
A case for centralizing undergraduate summer research programs: the DeBakey Research‐Intensive Community
Authors: Gatson Sarah N , Stewart Randolph H , Laine Glen A , Quick Christopher M
Published: June 2021

We identify three problems with the standard undergraduate research program model. 1) Students fit into an existing research program rather than designing one; 2) Students suffer from anemic mentoring; 3) Students are isolated from their peers as they develop into practicing scientists. The fundamental reason for these problems is that effective mentoring is resource intensive, and single faculty cannot mentor more than a very few students at once. We address the underlying problem of this inefficiency and radically increase the number of novices that participate in authentic research by bringing together faculty with active research programs, graduate students seeking experience in research management, and multi‐disciplinary teams of novices eager to participate in original research. Teams attend regular workshops and use a common online communication portal to develop their research, and are directed by a graduate student, who is in turn advised by faculty members. This model has been used successfully since 2004 to enable over 200 novices to co‐author original research. The resulting Research‐Intensive Community model may be used as a resource in developing curricula, publications, and future graduate students. This model was employed to integrate four independent summer programs funded by NSF and NIH, resulting in decreased cost and increased research productivity.
Evolutionary implications of sexual dimorphism in the human midcarpal joint
Authors: Kivell Tracy Lynne , Guimont Isabelle
Published: June 2021

Research has revealed significant size differences between human male and female carpal bones but it is unknown if there are significant shape differences as well. Since humans have a relatively low degree of body mass dimorphism, differences in shape would not be expected but, if present, may (1) represent a phylogenetic retention from our more sexually‐dimorphic evolutionary relatives or (2) imply subtle functional differences in the male and female wrist. This study investigated allometry and shape variation in carpals in extant hominoids. We found that the human midcarpal joint displayed twice as many significantly sexually dimorphic shape variables compared to other hominoids. Human midcarpal morphology was generally great ape‐like, not solely Pan‐ or African ape‐like, and often followed a sex relationship (e.g., females larger than males) that was found only in Asian apes. Human scaphoid morphology was the unique relative to other hominoids while the human lunate showed the most sexual dimorphism, which together may imply variation in radiocarpal joint function. Although some sexual dimorphism was also found in other hominoids, the morphological patterns displayed in humans suggest the development of the new shape variation rather than phylogenetic lag. The functional implications, if any, are important for the locomotor reconstruction of Australopithecus and the origin of human bipedalism.

Grant Funding Source

NSF
NHE1 is Critical in the Urokinase Plasminogen Activator‐ Mediated Activation of MMP‐9
Authors: Rastedt Dani , Wallert Mark , Provost Joseph J
Published: June 2021

The Na+/H+ exchanger isoform 1 (NHE1) plays a key role in cytoskeletal anchoring and regulation of intracellular pH, at the leading edge of migrating cells. This distribution and activity of NHE1 in metastatic cancer cells shows a similarity to the localization of the urokinase plasminogen activator receptor (uPAR) resulting in the stimulation of growth factor pathways and degradation of the extracellular matrix. In this study, the mechanism of uPAR activation was examined for its impact in proliferation, stress fiber formation, and tumor growth. Additionally, the amino terminal fragment of uPA (uPA‐ATF) was also used to activate the uPA response. In H1299, uPA increased ERK activation but no significant change occurred in H460 and H358. In the presence of 10 nM uPA or uPA‐ATF, cellular proliferation increased by 20‐30% in H358 and H460, whereas H1299 showed no significant changes. To investigate whether uPAR activation stimulated cell migration, stress fiber formation was measured. All three cell lines showed a significant increase in stress fibers with uPA stimulation which was blocked by inhibition of NHE1. Finally, to determine whether migration and proliferation could combine to support new tumor formation, a soft agar assay was performed. H460 cells showed a significant increase in large tumor formation. Supported by NIH Grant 1R15CA135616‐01, And NSF Grant MCB‐0817784.
Design of a cell‐responsive de‐PEGylation DNA delivery system
Authors: Myrick Stephanie Lynne , Millili Peter G. , Sullivan Millicent O.
Published: June 2021

We are interested in formulating efficient non‐viral DNA delivery vehicles by incorporating multiple functional layers that can be sequentially cleaved off during the delivery process to expose new features. We have established a simplified model vehicle consisting of a polycation‐condensed DNA core attached via linker peptides to a layer of poly(ethylene) glycol (PEG). PEG has been shown to protect biomaterials from salt‐induced aggregation and immune recognition, but PEG coatings can inhibit DNA delivery at the target cell. This project is focused on validating cell‐responsive de‐PEGylation of the vehicle prior to delivery via the cleavage of linker peptides sensitive to matrix metalloproteinase‐1 (MMP‐1), a collagen‐degrading enzyme upregulated by fibroblasts as they migrate through the extracellular matrix. This migration is observed in tumor stroma and at sites of injury, making MMP‐1 secretion a useful targeting signal. Having established the conditions that stimulate MMP‐1 expression by human dermal fibroblasts via Western blotting and immunostaining, we are now investigating the efficiency of vehicle cleavage using light scattering in combination with salt aggregation assays (Figure 1) to detect PEG layer release. Further experiments in this area and cell studies are ongoing. Supported by HHMI, NSF, and UDRF.
Investigation of a conserved serine residue in a yeast casein kinase 1 protein kinase
Authors: Miller Jessica , Robinson Lucy C , Brame Cynthia J
Published: June 2021

Yck1/2 are essential and redundant plasma membrane‐associated casein kinase 1 (CK1) protein kinases in budding yeast. In addition to sequence features common to Ser/Thr protein kinases, CK1 proteins share features unique to this subfamily. We identified the peptide EQSRRDD as such a feature and investigated its role in the function of Yck2 (E258‐D263). Based on location of the conserved peptide in a 3‐D model of CK1 from S. pombe, we propose that this sequence is necessary for substrate binding. We focused on S260 within the conserved sequence, generating mutant alleles that were both integrated into the yeast chromosome to allow investigation of growth phenotypes and expressed as beta‐galactosidase fusion proteins for in vitro kinase assays. Deletion of S260 eliminates Yck2 function. S260T and S260A substitutions have little effect on Yck2 localization, in vitro activity on casein, or growth phenotypes of yeast expressing only the mutant allele. However, S260E substitution results in decreased kinase activity as well as significantly slower growth of yeast expressing only the mutant allele. We are testing the possibility that S260E impairs substrate recognition. Supported by NSF grant MCB‐0517204.
Green tea improves carbohydrate and lipid metabolism and regulates cardiac mRNA expression related to insulin, lipid and inflammatory signaling pathways
Authors: Qin Bolin , Polansky Marilyn M. , Dawson Harry , Anderson Richard A.
Published: June 2021

Green tea has multiple benefits to human health. Epidemiologic studies demonstrate that the consumption of green tea polyphenols (GTP) reduces the risk of coronary artery disease and cardiac events. To explore the underlying mechanisms of action at a molecular level, we examined the effects of GTP on the expression of the cardiac genes involved in insulin, lipid and inflammatory signaling pathways, using quantitative real‐time PCR. Rats, fed a high fructose diet for four weeks to induce insulin resistance, were divided into two groups: the control group and GTP group (200 mg/kg body weight daily dissolved in distilled water for 6 weeks). There were no significant differences in average food intake and bodyweight gain between groups. GTP reduced serum blood glucose, cholesterol, triglycerides, free fatty acids and LDL‐C levels, compared with the controls. Supplementation with GTP also increased insulin receptor, IRS1, IRS2, PI3K, Akt1, GLUT1 and GLUT4 mRNA levels in the myocardium; increased cardiac CD36, PTEN, PPARg mRNA levels; but decreased the inflammatory factors: TNFa and IL1ß mRNA levels. In summary, our data demonstrate that green tea polyphenols improve glucose and lipid metabolism and regulate the cardiac gene expression related to insulin, lipid and inflammatory pathways in fructose‐fed insulin resistant rats. (Funded by USDA/ARS and Integrity Nutraceuticals).
Boron (B) deprivation increases plasma homocysteine and decreases liver S‐adenosylmethionine (SAM) and S‐adenosylhomocysteine (SAH) in rats
Authors: Nielsen Forrest H
Published: June 2021

The diverse effects of B deprivation suggest that B affects a biomolecule involved in a variety of biochemical reactions. An experiment was conducted to determine whether dietary B affects the liver concentration of SAM, a frequently used enzyme substrate, especially for methylation reactions that yield SAH. SAH is hydrolyzed to yield homocysteine (Hcy). Weanling male rats (15 per treatment) were fed a ground corn‐casein based diet (0.15 mg B/kg) containing sucrose and fructose to increase oxidative stress. Experimental variables were dietary supplements of 0 and 3 mg B/kg and fat sources (affect the response to B) of 75 g corn oil, 75 g canola oil, or 65 g fish oil plus 10 g linoleic acid/kg. When euthanized at age 18 wk, rats fed the B‐low diet were considered B‐deprived because they had decreased B concentrations in femur, tibia, and kidney. Decreased plasma C‐reactive protein indicated that fish oil decreased oxidative stress; the decrease was more noticeable in B‐deprived rats. Boron deprivation regardless of dietary oil decreased liver SAM and SAH, and increased plasma Hcy. Fish oil decreased liver SAM, which attenuated the SAM response to B. Femur, tibia and liver iron (bound by SAM) was decreased in B‐supplemented and fish oil‐fed rats. The findings suggest that B is bioactive through affecting the formation or utilization of SAM, and this effect is enhanced by oxidative stress. (Supported by USDA)
Effects of Long‐Term Soy vs Casein Protein Intake and Obesity on Serum Insulin‐Like Growth Factor 1 (IGF‐1) and IGFBP‐3 Levels using Obese Female Zucker Rat Model.
Authors: Hakkak Reza , Shaaf Saeid , Korourian Soheila
Published: June 2021

Elevated serum IGF‐1 is an increased risk for breast cancer development. The main objective of this study was to investigate the effects of high‐isoflavone soy protein diet (hi‐isof soy) and obesity on serum IGF‐1 and binding proteins (BP) IGFBP‐3 levels. Lean (n= 54) and obese female (n= 45) rats were maintained on either casein or hi‐isof soy protein diet for 155 days. All rats were orally gavaged at age 50 days with 65 mg/kg DMBA. Rats were weighed once a week and were killed 155 days post‐DMBA treatment. Blood was drawn at different ages (43, 99 and 197 d) to measure serum IGF‐1 and IGFBP‐3 by ELISA. Obese soy‐fed rats gained significantly (P<0.001) more weight than obese casein rats. At age 43 d, no differences were found on serum IGF‐1 levels among the groups but IGFBP‐3 was increased by hi‐isof soy diet in lean group compared to casein fed lean rats. At the age 99 d, obesity increased (P<0.05) serum IGF‐1 for both casein and high‐isof soy diet compared to the lean group and this increase was maintained for the rest of the experiment. At age 99 d ,obesity increased IGFBP‐3 levels in the casein but not hi‐isof soy diet group (P<0.001). At end of the Exp. (197 d); obesity increased IGFBP‐3 (P<0.05) both in casein and soy fed rats. Our results suggest that dietary protein source can affect serum IGF‐1 and IGFBP‐3 in obese female Zucker rats (Supported by USDA and ABI to RH).
Total Fat, n‐3 Fatty Acids and Vitamin D3 in Selected Fish Species Sampled Under USDA's National Food and Nutrient Analysis Program (NFNAP)
Authors: Exler Jacob , Byrdwell W. Craig , Holden Joanne M. , Lemar Linda E. , Patterson Kristine Y. , Pehrsson Pamela R. , Phillips Katerhine M.
Published: June 2021

As part of the USDA's National Food and Nutrient Analysis Program (NFNAP), new analytical data were determined for the 20 most frequently consumed raw fish identified for FDA's voluntary nutrition labeling. Samples of 21 fish species were purchased from 12 supermarkets nationwide according to a statistical sampling plan. The samples were processed at Virginia Tech (a cooperator of USDA's Nutrient Data Laboratory), and detailed information on each sample was recorded. Sample composites and QC materials were analyzed by qualified laboratories. Because fish are a natural source of n‐3 fatty acids and vitamin D3, data for these nutrients and total fat are being reported. Results for n‐3 fatty acids show that the highest amounts were found in salmon and rainbow trout, closely related species, and lower amounts were in shrimp, scallops, and tuna. Preliminary results for vitamin D3 in wild salmon species show that values vary from 10 mcg/100g or less in foreign caught samples to between 10 and 20 mcg/100g in domestic caught samples. Nutrient profiles for these species will be released to the USDA National Nutrient Database for Standard Reference (http://www.ars.usda.gov/nutrientdata), which is used to support the USDA/NHANES food consumption surveys, food and nutrition policy, and consumer education.

Support by USDA, NIH #Y1CN5010, FDA, and the Beverage Institute for Health & Wellness (Coca‐Cola, Inc.)
Parathyroid hormone (PTH) values, Vitamin D status, Fetal bone Growth and Placental RANK‐L expression among Pregnant Adolescents
Authors: Essley Bridget Victoria , Foehr Marisa , Cooper Elizabeth , McIntyre Allison , Kent Tera , McNanley Thomas , Witter Frank , Harris Zena , Lafond Julie , O'Brien Kimberly
Published: June 2021

Pregnant adolescents may be at risk for suboptimal vitamin D status and bone loss across pregnancy. RANK/RANK‐L/OPG interactions are integral to osteoclast activity and alterations in RANK‐L expression may be associated with the critical balance between maternal bone loss and fetal bone growth during pregnancy. To assess these issues, vitamin D status (25‐hydroxyvitamin D) was monitored in pregnant adolescents (<18 years old) and their neonates in relation to longitudinal measures of fetal femur and humerus growth, and placental expression of RANK‐L. Fetal femur and humerus length was obtained by sonogram three times across gestation to assess fetal bone growth. Blood samples were collected from each adolescent mid‐gestation and at parturition when cord blood and placental tissue were also obtained. In 25 adolescents studied to date, expression of soluble RANK‐L in the placenta approached significance as a predictor of fetal bone growth (p=.066). At delivery, 71% (n=17) of teens had elevated PTH (>46 pg/mL). 41% (n=43) of teens and 63% of neonates (n=38), had suboptimal vitamin D levels (<20 ng/mL). Recruitment and analyses are on‐going and additional relationships between serum osteoprotegerin (OPG), maternal bone loss, and fetal bone accrual across pregnancy will be explored.

Grant Funding Source

USDA Grant No: 2008‐01857
Obesity is more common among women in the most rural places within rural counties
Authors: Olson Christine M , Strawderman Myla S
Published: June 2021

While it is well‐known that the prevalence of obesity is higher among women living in rural areas compared to urban areas, little is know about the geographic distribution of obesity within rural areas of the US. This study addressed whether obesity was more prevalent in childbearing women who lived in the most rural locations in 8 rural counties in Upstate New York. The zip codes of 552 healthy pregnant women who entered prenatal care were determined and classified into one of three area types: a city; those with incorporated villages; and those with no incorporated areas (the most rural areas). The number of women in each group was 63, 314, and 175, and the prevalence of obesity was 24%, 21% and 36%, respectively (p = 0.001). The risk of obesity in early pregnancy was then examined by type of geographic area, controlling for individual‐level covariates: education, income, parity and age. Women living in the most rural areas were 2.20 (95% CI: 1.46, 3.33) times more likely to be obese than women in the other zip code groups and women who were college graduates were only about half as likely to be obese as women with less education (95% CI: 0.29, 0.77). Within rural communities, obesity is more prevalent among women in the most rural places.

Supported by NIH Grant No. HD29549 & USDA Grant No. 2005‐35215‐15427
Sequence and expression analysis of hypoxia‐inducible factors in the killifish, Fundulus heteroclitus
Authors: Rees Bernard Bowman , Karchner Sibel I , Skripnikova Elena , Ferry Mark J , Townley Ian K , Hahn Mark E
Published: June 2021

In fish and other vertebrates, the hypoxia inducible factor (HIF) family of transcription factors is implicated in the regulation of genes whose expression underlies cellular, tissue, and organismal responses to low oxygen. HIF functions as a dimer composed of an oxygen‐dependent alpha subunit and a constitutively expressed beta subunit (also known as the Aryl Hydrocarbon Receptor Nuclear Translocator, ARNT). Previous studies have demonstrated the presence of HIF‐2α and ARNT2 in the estuarine fish, Fundulus heteroclitus. In the present study, we cloned and sequenced HIF‐1α and HIF‐3α from F. heteroclitus and the closely‐related F. grandis. In both species, there is polymorphism in the nucleotide and deduced amino acid sequences of HIF‐1α and HIF‐3α, suggesting that positive selection may be occurring in those genes. The expression of HIF‐1, 2, and 3α transcripts in brain, gill, gonad, heart, liver, and muscle tissues was measured by quantitative PCR. HIF‐α copy number differed among tissues, but within a given tissue HIF expression was relatively unchanged during long term exposure of fish to altered oxygen levels. On‐going research addresses the tissue and oxygen dependence of HIF protein expression, as well as the role of the multiple forms of HIF in fish. Funded by NSF and NIH (Superfund).
Spt4 and Spt5 cooperatively suppress transcription coupled DNA repair through binding to RNA polymerase II in the absence of Rad26
Authors: Ding Baojin , Chen Xuefeng , Ruggiero Christine , LeJeune Danielle , Li Shisheng
Published: June 2021

Rad26, the yeast homolog of human CSB protein, and Rpb9, a nonessential subunit of RNA polymerase II (Pol II), mediate two subpathways of transcription coupled nucleotide excision repair (TC‐NER). However, the deletion of Spt4, a transcription elongation factor, makes both Rad26 and Rpb9 dispensable, suggesting Spt4 works as a suppressor in the modulation of TC‐NER. To examine the molecular mechanism of this modulation, NER activities of different mutants were analyzed by running sequencing gel. The protein‐protein interactions were explored by immunoprecipitation (IP) and Western blot. Here we show that deletion of spt4 increases TC‐NER in rad26 cells but does not affect repair in RAD26+ cells, regardless of the presence of Rpb9. Spt4 protects Spt5 from degradation and enhances the association of Spt5 with Pol II. Overexpression of Spt5 restores the UV sensitivity and partially suppressed NER activity. Moreover, TCR rates in rad26 cells are inversely correlated with the level of Spt5 associated with Pol II. Our results suggest that Spt4 and Spt5 cooperatively suppress TC‐NER only in the absence of Rad26. By binding to Pol II, Spt5 plays a more direct role in the suppression, whereas Spt4 is indirectly involved in the suppression by stabilizing Spt5 and by enhancing the binding of Spt5 to Pol II.

This study was supported by NIH grant ES012718 and NSF grant MCB‐0745229.
The structure of the human tRNALys3 anticodon bound to the HIV genome Loop I is stabilized by modified nucleosides
Authors: Bilbille Yann , Vendeix Franck AP , Guenther Richard , Malkiewicz Andrzej , Ariza Xavier , Vilarrasa Jaume , Agris Paul F
Published: June 2021

Replication of HIV requires base pairing of the reverse transcriptase primer, human tRNALys3, to the viral RNA. Although the major complementary base pairing occurs between the HIV primer binding sequence and the tRNA's 3′‐terminus, an important discriminatory, secondary contact occurs between the viral A‐rich Loop I and the modified, U‐rich anticodon domain of tRNALys3. The thermal stabilities of variously modified tRNA anticodon region sequences bound to the Loop I of viral serosubtypes G and B were analyzed by UV and CD and the structure of one duplex was determined using NMR spectroscopy and restrained molecular dynamics. Thiouridine, s2U34, and pseudouridine, ?39, modifications appreciably stabilized the interaction of the anticodon region with the viral RNAs. The structure of the duplex composed of the HIV‐1 Loop I bound to a doubly modified tRNA's anticodon region resulted in two coaxially stacked A‐form RNA stems separated by two mismatched base pairs, U?? and G?A, that maintained a reasonable helix diameter. Thus, the important interaction of the primer's anticodon region with the viral RNA is strengthened by s2U34 and ?39 modifications that facilitate duplex interaction. This research was supported by a National Institutes of Health grant and National Science Foundation grants.
A UbcH7‐mediated pathway is critical for the conjugation of ubiquitin to picornaviral 3C proteases
Authors: Lawson Thomas Glen , Schlax Peter E.
Published: June 2021

The polyprotein processing 3C proteases of the encephalomyocarditis virus (EMCV) and hepatitis A virus (HAV) are rapidly degraded by the ubiquitin/26S protease system. We have identified three distinct ubiquitin‐conjugating pathways that catalyze 3C protease ubiquitination: one that includes the E2/E3 pair UbcH1/E3α, one requiring UbcH5a, and a UbcH7‐dependent pathway. As part of an effort to understand how these pathways function together to mark the 3C proteases for proteasome‐mediated degradation, we have carried out experiments using reconstituted in vitro ubiquitination reaction systems to evaluate the contribution each pathway makes to the conjugation of polyubiquitin chains to the EMCV and HAV 3C proteases. Our results show the UbcH7‐dependent pathway plays a critical role in the initial conjugation of ubiquitin molecules to the 3C proteases. Initial 3C protease recognition by the UbcH7‐dependent E3 ubiquitin‐protein ligase, or UbcH7/E3 pair, requires that a previously identified ten‐amino acid "destruction signal" in the 3C proteases be intact. The subsequent ubiquitin chain extension process, however, requires the presence of all three complete pathways. Reaction mixtures lacking one or two of the complete individual ubiquitin‐conjugating pathways failed to robustly support 3C protease polyubiquitination. This work was supported NSF grant award MCB‐0210188.
Osmolytes affect the kinetics of Abeta (1‐42) aggregation
Authors: Heldt Caryn L , Sorci Mirco , Belfort Georges
Published: June 2021

The formation of plaques in the brains of Alzheimer's patients has long been thought to be the disease causing agent. Yet, recent data has shown that small oligomers of Abeta that form prior to plaque formation may actually be the toxic disease causing agent. But little is known about these small oligomer species. We have examined the formation of the small oligomers created from Abeta (1‐42) to better understand the formation of these suspected toxic species. Using circular dichroism, the transition of the peptide from a random configuration to an ordered beta sheet oligomer was observed and the kinetic rates were examined to determine how the addition of different osmolytes affected the oligomer formation. With the use of small‐angle scattering, we have examined the rate of change of the size of these small oligomer species under aggregating conditions. By changing the concentration of osmolytes in the solution, we were able to increase and decrease the kinetic rates of the aggregation process and learn more about the chemical environment that causes the transition of Abeta (1‐42) from random peptide to structured oligomer. This information is important to understand the molecular basis of Alzheimer's disease and brings us closer to creating a drug that can slow or stop the progression of this disease. This work was funded by the U.S. Department of Energy, DOE (DE‐FG02‐07ER46429).
Effects of soy vs casein protein diet and obesity on DMBA‐induced mammary tumor development
Authors: Hakkak Reza , Shaaf Saeid , JO Chan‐Hee , Talley Lori , Korourian Soheila
Published: June 2021

We reported that intact obese Zucker rats fed a standard chow diet had increased susceptibility to DMBA‐induced mammary tumors compared to lean Zucker rats. The objective of this study was to investigate the effects of long‐term high‐isoflavone soy (high‐isof soy) vs casein protein intake and obesity on mammary tumor development. Five‐week‐old female Zucker rats (45 obese fa/fa and 54 lean) were used. After one week of acclimation (age 42 days), rats were randomly assigned to the following diet groups; 1) Lean, casein; 2) Obese, casein; 3) Lean, high‐isof soy and 4) Obese, high‐isof soy. Rats had ad lib access to water and diet with either casein or high‐isof soy protein isolate. All rats were orally gavaged at age 50 days with 65 mg/kg DMBA. Rats were weighed and palpated twice weekly and sacrificed 155 days post‐DMBA treatment. Obese soy‐fed rats gain significantly (P<0.001) more weight than obese casein rats. The lean high‐isof soy‐fed rats had slightly higher (P = 0.057) body weight than lean casein‐fed rats. At end of the experiment, 69% of the Lean casein rats developed mammary tumors compared to 50% in lean high‐isof soy group (P<0.08). In the obese group, 76% of the high‐isof soy‐fed rats developed mammary tumors compared to 15% (P<0.05) of obese casein‐ fed rats. These results suggest that intact obese versus lean Zucker female rats fed semi‐purified diets are protected against DMBA‐induced mammary tumor on the casein but not high‐isof soy diet. (Supported by USDA and ABI to RH)
Compositional analysis and apparent macronutrient digestibility of four raw meat diets in domestic cats
Authors: Kerr Katherine R , Beloshapka Alison , Dikeman Cheryl , Simmons Lee G , Swanson Kelly S
Published: June 2021

The popularity of feeding raw diets to pets has increased over the past several years. However, little nutritional or metabolic information has been collected from domestic cats fed such diets. Additionally, meat types and sources may vary greatly in composition. The objective of this study was to evaluate the differences in apparent digestibility of four raw meat based diets: beef (BE; 19% fat, 66% crude protein (CP)); bison (BI; 38% fat, 49% CP); elk (E; 6% fat, 79% CP); and horse (H; 26% fat, 60% CP). Eight intact female cats were fed to maintain BW in a crossover design. Estimated metabolic energy intake was greater (P<0.05) in cats fed BI vs those fed BE, E, and H. Fecal output (g DM) was lower (P<0.05) in cats fed H vs those fed BE and E, and lower (P<0.05) for BI than E. Fecal DM percentage was lower (P<0.05) for cats fed BE vs those fed BI, E, and H. Apparent DM and OM digestibilities were lower (P<0.05) for cats fed BE (84.1%; 87.4%) vs those fed BI (88.1%; 90.4%) and H (87.1%; 89.8%), and lower (P<0.05) for those fed E (84.3%; 87.6%) vs BI. Apparent fat digestibility was lower (P<0.05) for cats fed E (88.7%) vs those fed BE (94.4%), BI (96.8%), and H (96.9%), and was lower (P<0.05) for cats fed BE vs those fed BI and H. Apparent CP digestibility was lower (P<0.05) for cats fed BE (96.6%) vs those fed E (97.3%). Although dietary macronutrient composition was highly variable and statistical differences were noted, all diets were highly digestible.

Grant Funding Source

USDA‐CSREES (project ILLU‐538‐396)
Effect of dietary supplementation with a formulated nutrient mixture together with whey‐based protein on immune response of young and old mice
Authors: Wu Dayong , Onwulata Charles , Ren Zhihong , Pae Munkyong , Pang Hoan‐Jen , Meydani Simin Nikbin
Published: June 2021

Aging is associated with dysregulation of the immune response. Nutrient supplementation has been shown to help maintain a healthy immune system during aging. In this study we fed young (4‐5 mo) and old (23‐24 mo) C57BL/6 mice, a diet supplemented with a formulated nutrient mixture and whey proteins modified by extrusion texturization for enhanced physical functionality (NTWP) for 5 wk. Composition of cell types in spleen cells was not significantly affected by NTWP supplementation. Lymphocyte proliferation in response to T cell mitogen Con A or PHA, or anti‐CD3/CD28 was significantly lower in old mice compared to young mice. Supplementation with NTWP enhanced lymphocyte proliferation in both young and old mice. Splenocytes from old mice produced less IL‐2, more IFN‐γ and IL‐4, and similar amount of IL‐10 compared to those from young mice. Supplementation with NTWP did not significantly affect production of these cytokines, indicating that other factors contributed to NTWP‐induced enhancement of T cell proliferation. Production of inflammatory cytokines IL‐6 and TNF‐α, and T cell suppressive factor prostaglandin E2 tended to be higher in the splenocytes from old than those from young mice fed the control diet. While supplementation with NTWP had no effect on production of these molecules in the splenocytes from young mice, it tended to inhibit their production by those from old mice. These results indicate that consumption of the formulated nutrient mixture together with the whey‐based protein may favorably modulate immune response. Supported by USDA CRIS Projects #58‐1950‐7‐707 and #1935‐41000‐065.
eNOS signaling is essential in GLP‐2‐mediated stimulation of blood flow, but not cell proliferation in the mouse gut
Authors: Guan Xinfu , Wang Yi , Li Xiaojie , Burrin Douglas , Li Defa
Published: June 2021

Through a G protein‐coupled receptor, glucagon‐like peptide‐2 (GLP‐2) stimulates intestinal crypt cell proliferation and mucosal blood flow. GLP‐2 receptor is localized to enteric neurons, endocrine cells, and myofibroblasts, but not enterocytes. However, it is largely unknown how GLP‐2 receptor‐activated mitogenic, cytoprotective, and vasoactive signals are conveyed to the intestinal epithelium. Nitric oxide can function as a cellular signal to regulate cell growth and blood flow. Thus, our objective was to determine whether nitric oxide is a key mediator in the GLP‐2 receptor‐activated enteric signaling network. Endothelial nitric oxide synthase knockout (eNOS KO) and wild‐type (eNOS WT) mice (n=40) were subcutaneously injected with GLP‐2 or saline for 14 d, and with BrdU 2 h prior to euthanization. Mucosal cell proliferation, apoptosis, and blood flow were determined by BrdU‐positive cells, active caspase‐3 abundance, and hemoglobin (Hb) content, respectively. There was no difference between eNOS KO and WT mice in gut growth (indicated by mucosal weight, protein mass, villous height and crypt depth), cell proliferation, apoptosis, or blood flow. GLP‐2 increased gut growth by stimulating crypt cell proliferation in both KO and WT mice. GLP‐2 appeared to decrease apoptosis in the WT mice, but increase it in the KO mice. Importantly, GLP‐2 increased gut blood flow in the WT mice, but not in the KO mice. We conclude that GLP‐2‐induced vasoactive action is mediated by the eNOS signaling, but GLP‐2‐stimulated tropic action is independent of it. (Supported by USDA CRIS 6250‐51000‐043 and NIDDK K01DK75489)
Signal transduction pathways are altered by cellular labile iron
Authors: Hegde Narasimha , Hao Lei , Unger Erica , Han Okhee , Beard John
Published: June 2021

The cellular labile iron pool (LIP), which represents <5% of the total cellular iron is associated with a diverse population of ligands such as organic anions, polypeptides, and surface components of membranes. LIP as a self‐regulatory pool that is sensed by cytosolic iron regulatory proteins (IRPs) and feedback regulated by IRP‐dependent expression of iron import and storage proteins. Variation in the LIP can also contribute to basic cellular signaling events. In these studies we examined the effect of LIP variation on cell surface processes mediated by protein kinase C (PKC) in a model system of neuroblastoma cells. Levels of isoforms of PKC were highly sensitive to LIP while PKC beta isoform being more susceptible to change. Confocal microscopy studies suggest increased ubiqitination and internalization of membrane proteins and are consistent with qRT‐PCR gene array analysis of signaling pathways. Lower LIP levels result in increased ubiqitination, increased phosphorylation, and lower rates of endocytic protein recycling. The proximal "switch" for this activation is being explored. This ongoing study is supported by USDA (# 2007‐35200‐18231).
How accurate are reported energy contents of prepared foods in restaurants and supermarkets in the US?
Authors: Urban Lorien E. , Dallal Gerard E. , Robinson Lisa M. , Ausman Lynne M. , Saltzman Edward , Roberts Susan B.
Published: June 2021

Recent media reports suggest that restaurants may substantially under‐report the energy contents of typical menu items. We examined the accuracy of reported energy contents of 31 items from fast food and chain restaurants and 10 supermarket frozen meals in Massachusetts suitable for weight loss based on their low energy contents posted on websites. Food was analyzed for gross energy (GE) content using a validated bomb calorimetry system accurate to ±1.9% for individual food samples and values were compared to reported information normalized to equivalent GE content. Mean reported energy for restaurant foods and supermarket meals were respectively 16% and 8% higher than reported values, and some individual restaurant items were up to 100% greater than reported energy. When free side dishes were provided and included in the comparison, mean provided energy was 245% of reported values for the purchased items. These findings suggest a tendency for restaurants to under‐report meal energy content, and highlight the major impacts that variability in reporting accuracy and free side dishes have on provided energy. Further studies are needed to determine the extent of underreporting of energy, which has implications for both emerging policy initiatives requiring reporting of food energy at the point of purchase and individual efforts to use provided information for self‐monitoring for weight control.

Grant Funding Source

NIH grant HL069772‐06 and the USDA under agreement no. 58‐1950‐4‐401
Innovative oximes as potential blood‐brain barrier penetrating cholinesterase reactivators
Authors: Campbell Amy J , Olson John , Moorad‐Doctor Deborah , Kim Andrew , Garcia Gregory E.
Published: June 2021

Organophosphate (OP) nerve agents are extremely toxic because they inhibit acetylcholinesterase (AChE). The US Army currently utilizes pralidoxime (2‐PAM) for the reactivation of peripheral OP‐inhibited AChE. However, 2‐PAM does not cross the blood‐brain barrier (BBB) at therapeutically relevant levels; therefore acute OP exposure can lead to status epilepticus (SE) which can cause brain damage. We propose to prevent SE by reactivating CNS AChE. Our oximes will potentially be transported across the BBB because they contain a sugar moiety, hence the terminology 'sugar‐oximes'. This sugar moiety could be recognized by the facilitative transporter GLUT1. Published data has shown that glucose‐oxime conjugates can prevent the body temperature inducing properties of OPs in mice, indicating a CNS effect. We have synthesized these previously reported, but understudied, sugar‐oximes and have developed an in vitro model for evaluating their reactivation of OP inhibited RBC‐ChE. Our results show several sugar‐oximes are good candidates for further studies, including in vivo efficacy studies and in vitro assays to show that the sugar‐oxime interacts with GLUT1. This work is supported by the Defense Threat Reduction Agency (DTRA).