bluuebunny/crossref_metadata_2025 huggingface dataset:
Identifying the Functional Orthologs of Lipid Transfer Proteins (LTPs) for Developmental Regulation in Flowering Plants
Authors: Caonguyen Sean L. , Patel Swapneel J , Vellanoweth Robert L
Published: June 2021

During the floral transition in Arabidopsis, 13‐HOO‐FA, a possible lipid hydroperoxide precursor for signaling molecules, increases in leaves due to lipoxygenase activity. Results have shown the up‐regulation of two lipid transfer protein (LTP) genes, AtLTPf1 and AtLTPf2, at the floral transition. LTPs may act as signal carriers that transfer lipids, like oxylipins, between cells for developmental events. The purpose of this experiment is to locate orthologs of AtLTPf1 and AtLTPf2 from Arabidopsis (a dicot) in rice (a monocot) using bioinformatics coupled with expression pattern analyses. Our phylogenetic trees show LTP homologs, and we hypothesize that the most closely related genes, OsLTPf1 and OsLTPf2 from rice, mirror AtLTPf1 and AtLTPf2 expression, respectively. Through a comparison of various tissues at different stages of development in both plant species, our data suggests that AtLTPf1 and OsLTPf1 are orthologs. Further analysis of these LTPs may clarify their function and elucidate the signaling pathways involved in floral transition and developmental regulation. These LTPs may provide targets for genetic modification to alter plant development. Supported by NIH‐SCORE GM08101
Expression of Single Chain Antibody Variable Regions against Botulinum Neurotoxin‐A in the Unicellular Green Alga Chlamydomonas reinhardtii
Authors: JEONG SOPHIA , VO ASIA , COHEN AMYBETH
Published: June 2021

Botulinum neurotoxin (BoNT) is produced by the bacterium Clostridium botulinum. As the serotype A form of the toxin is extremely potent, it is ranked by The Centers for Disease Control as one of the top six compounds that could be used as a biological weapon. Thus, three single chain variable fragment (scFv) anti‐BoNT antibodies (S25, C25 and 3D12), which were previously shown to inactivate BoNT‐A in mice (Nowakowski et al, PNAS, 99, 11346, 2002), were expressed in the unicellular green alga Chlamydomonas reinhardtii for detection and neutralization of aerosolized BoNT (Coleman, Kuhnlein, and Cohen, unpublished data). Unfortunately, protein purification yields were low, despite optimizing the codon bias within the scFv genes for C. reinhardtii. Each scFv gene was cloned into a more potent expression vector (fD1‐tD1 pBS), which contains the strong light‐regulated D1 promoter and termination sequences. Furthermore, a 3X M2 epitope tag at the carboxy terminus of each antibody was altered to 1X. The new constructs were co‐transformed with a spectinomycin resistance gene into C. reinhardtii cells using microprojectile bombardment. Growth of spectinomycin‐resistant colonies was observed, and transformants are being screened using Southern blot analysis and PCR in order to evaluate the integration of the C25, S25 or 3D12 genes into the chloroplast genome. Western blot analysis will be carried out to assess protein accumulation, and purified scFv BoNT‐A antibodies will be tested individually, and as a cocktail, for their ability to bind and neutralize BoNT‐A.
Sox transcription factor mediated transcriptional regulation of Robo4 expression and function
Authors: Samant Ganesh Vinayak , Pramanik Kallal , Leigh Noah , Horswill Mark , Beltrame Monica , Ramchandran Ramani
Published: June 2021

Our laboratory has identified Roundabout4 (Robo4) as a bonafide angiogenic receptor. Robo4 knockdown (KD) endothelial cells show decreased migration and zebrafish (zf) embryos show intersomitic vessel (ISV) defects. We have reported that Robo4 affected cell migration results in imbalanced active Cdc42 and disrupted actin organization but precise molecules that interact with Robo4 remain unknown. To identify the proteins that bind and orchestrate Robo4 mediated angiogenesis, we performed Yeast 2 Hybrid (Y2H) screen with cytoplasmic zf Robo4 as bait and zf cDNA as prey. Y2H identified many SOX transcription factor (TF) proteins as potential binding partners. Based on their vascular specificity we focused on Sox18 and Sox7 and immune precipitation confirmed that Robo4 binds to both. In vivo, Robo4 KD in conjunction with Sox18 and Sox7 KD decreased Fli ISV expression, indicating disrupted angiogenesis. Bioinformatics on Robo4 promoter reveals Sox TF binding sites and signifies transcriptional regulation of Robo4 via Sox. Using transactivation assays we have identified that Sox18/7 temporally induce Robo4 promoter. Sox18/7 double KD decreases Robo4 transcription and expression in zf vasculature. In sum, our data indicate that Robo4 and Sox TF interact genetically and Sox TF mediated transcriptional and post‐translational Robo4 expression and function is critical in angiogenesis.

Grant Funding Source: NIH
Endothelial sprout formation during vasculogenesis
Authors: Czirok Andras , Szabo Andras , Kosa Edina , Little Charles D , Lansford Rusty , Rupp Paul A
Published: June 2021

We studied vasculogenic sprouting, the multicellular patterning mechanism of vasculogenesis, with time‐lapse microscopy in avian embryos, and in cultures of endothelial cells invading collagen I gels. Cell‐autonomous motility and tissue movements were distinguished utilizing digitally filtered displacements of various immuno‐labeled extracellular matrix components. Endothelial cell motility was investigated inside the forming vessels through a recently developed TIE1‐YFP transgenic quail line. Displacements of pulse‐labeled fibronectin fibers around the nascent vessels reveal that the fibronectin accumulated is of mesodermal origin, and additional filaments are not assembled de novo during the process. Correlational analysis of endothelial cell movements and the morphology of fibronectin filaments shows that fibronectin does not act as a vascular pre‐pattern to guide multicellular sprouts. Sprout elongation depends on a continuous supply of endothelial cells, streaming along the sprout towards its tip. With computer simulations we explore the conditions when cell‐cell guided active cell motility can result in multicellular streaming and sprouting behavior.

Grant Funding Source: NIH R01HL087136
Staphylococcus Enterotoxin B induced Host Response Signatures in the Presence and Absence of Battlefield‐like Stress
Authors: Muhie Seid , Hammamieh Rasha , Yang David , Jett Marti
Published: June 2021

In previous studies, we identified transcripts and proteins suppressed by battlefield‐like stress that were essential for protective immunity. In this study, pre‐ and post‐training leucocytes, from US Army Ranger Trainees, were exposed in vitro to Staphylococcal enterotoxin B (SEB) to analyze SEB induced regulation of gene expression in the presence and absence of severe battlefield‐like stress. cDNA microarrays and real time PCR arrays were used for transcriptome wide profiling, and computational analyses to identify regulatory modules. Stress‐suppressed immune response genes were remained suppressed in SEB treated post‐training leukocytes. Despite complications due to suppressed immune responses seen at the end of intensive battlefield‐like training, we have identified other transcripts that showed consistent differential expression towards SEB in both pre‐ and post‐training leukocytes even in comparison to dengue virus and Yersinia pestis exposure. Ontological and regulatory network analyses revealed that SEB‐specific transcripts are involved in heparin binding, positive regulation of mitochondrial depolarization, apoptosis, negative regulations of JAK‐STAT cascades and axonogenesis. Identified SEB signatures have the potential to be used as SEB‐exposure markers under battlefield stress.

We are grateful to DTRA for funding.
IMAGEtag (Intracellular MultiAptamer Genetic tag) for Real‐time Imaging of Gene Promoter Activity
Authors: Shin Ilchung , Ilgu Muslum , Haarberg H. Eirik , Gupta Vinayak , Kraus George A , Nilsen‐Hamilton Marit
Published: June 2021

Reporter proteins such as fluorescence proteins and luciferase allow for non‐invasive detection of the products of gene expression in living cells. However, current reporter proteins do not provide for real‐time imaging of promoter activity in living cells. This is because of the long time after transcription required for RNA maturation, protein synthesis and maturation. We have developed an alternative reporter system for imaging real‐time using strings of RNA aptamers that constitute IMAGEtags (Intracellular MultiAptamer Genetic tags), which can be expressed from a promoter of choice. The tobramycin and neomycin RNA aptamers have so far been utilized for this system and expressed in yeast from the GAL1 promoter. For imaging, the cells are incubated with their ligands that are labeled with fluorescent dyes. To increase signal/noise, ligands have been separately conjugated with the FRET (Förster resonance energy transfer) pairs, Cy3 and Cy5. With these constructs, the transcribed aptamers can be imaged after induction by galactose. Initial optimization of IMAGEtags for imaging is done with yeast followed by the application of successful IMAGEtags to detect rapid changes in transcriptional activity in animal and plant cells in culture. This work was supported by the National Institutes of Health and the Biological Systems Science Division, Office of Energy Science Research of the U. S. Department of Energy.
Lentiviral vector expressing Nef as a tool to study HIV neuropathology
Authors: Cruz Emmanuel Antonio , Noel Richard Junior
Published: June 2021

The goal of this work is to use a lentivirus to create a simplified endogeonous expression system producing only Nef to study the unique contributions that this viral neurotoxin makes to HIV‐related neurological impairment. We used PCR to generate the Nef gene flanked by unique restriction enzymes sites(BamH1 and Sal1). Then we digested a lentiviral packaging vector (pLenti) and this PCR product with the same restriction enzymes to generate DNA fragments with complementary ends. Cloning was performed by incubating with DNA ligase and ATP to create recombinant pLenti‐Nef plasmids. Colonies were screened by PCR to identify those containing plasmids with HIV Nef. These pLenti‐Nef plasmids were recovered by plasmid prep and confirmed by sequencing. The pLenti‐Nef plasmid can be used to generate lentiviral particles capable of delivering the HIV‐Nef gene to target cells in a single round of infection a single round of infection. We will infect rat astrocytes in vitro with these viruses followed by implantation into the hippocampus of Sprague Dawley rats in vivo to mimic the levels of Nef that have been reported in HIV‐infected patients. This investigation will help us understand the HIV disease better at a molecular level by developing a system that allows us to study the role of Nef in driving immune cell chemotaxis into the brain and the loss of neuron function that accompanies HIV neuropathology in parallel. Grant support by RR003050 and RR016470.
Construction of the broad range cloning vector pHC60mCherry: Plant‐Microbe interaction applications
Authors: Ramirez Gustavo Antonio , Vozza Nicolas , Zorreguieta Angeles , Tolmasky Marcelo
Published: June 2021

Here we present the construction of a wide range fluorescent protein expression vector that is compatible with other commonly used fluorescence conferring vectors in gram negative bacteria. Cloning the red autofluorescent protein “mCherry” in the wide range vector pHC60 has produced a useful tool for various molecular applications such as multiple fluorescent tagged microscopies. The destination vector is capable of constitutively expressing genes in various genera (Rhizobium, Sinorhizobium, ect.). Additionally, its toxin‐antitoxin origin of replication makes it highly compatible with plasmid vectors with pBBR origins. The pHC60 vector originally contains a green fluorescent protein (GFP) sequence under its constitutive promoter. The GFP coding sequence was liberated from the plasmid by a digestion reaction with the restriction enzyme BglII. A subsequent double digestion with the restriction enzymes KpnI and XhoI allowed the pHC60 plasmid without GFP to become linear. The mCherry coding sequence was also liberated from the pJPO11 pCAGGS Chop‐mCherry‐WPR cassette by digestion with the same restriction enzymes. Compatible ends then facilitated a straight forward ligation reaction of the mCherry fragment with the linear pHC60V (GFP free) target vector. Additionally, we observed, by fluorescent microscopy, Rhizobium leguminosarum biovar viciae labeled with the new pHC60mCherry plasmid in free‐living and symbiotically associated states. A wide range fluorescent protein expression vector that is compatible with other commonly used fluorescence conferring vectors can have various applications particularly in multiple fluorescence tagged microscopy studies.
SAR Studies of Diphenyl Ethers, Potential Anti‐tuberculosis Drugs as InhA Inhibitors
Authors: Pan Pan , Liu Nina , Slayden Richard A , Tonge Peter J
Published: June 2021

Diphenyl ethers were demonstrated as a type of InhA inhibitors, which deactivate Fatty Acid Biosynthetic Pathway II in Mycobacterium tuberculosis and then block the synthesis of cell wall. Some small substituent groups at ortho position of B‐ring of diphenyl ethers introduce stronger binding affinity to InhA and special slow‐onset kinetic properties. A series of diphenyl ethers with different ortho groups on B‐ring were synthesized and tested for in vitro activity. All the compounds are tight binding inhibitors and showed good in vitro activity. The kinetics assay indicated that some compounds are slow‐onset inhibitors and that the sizes and charges of substituent groups affect kinetic properties. This work was supported by NIH Grant AI70383.
Binary selectivity for HCV NS3/4A protease versus host proteases via irreversible inactivation at a non‐catalytic cysteine
Authors: Petter Russell C , Hagel Margit , Niu Deqiang , St Martin Thia , Sheets Michael P , Qiao Lixin , Bernhard Hugues , Karp Russell , Zhu Zhendong , Labenski Matthew T , Chaturvedi Prasson , Nacht Mariana , Westlin William , Singh Juswinder
Published: June 2021

Viral protease inhibitors typically achieve potency via interactions with the catalytic apparatus, but as a result are often poorly selective. Viral protease inhibitors must also block emerging mutants. We have developed a fundamentally different approach to the design of covalent inhibitors by targeting non‐catalytic cysteine residues unique to viral targets but absent in host proteases, enabling extreme, “binary” selectivity. An estimated 170 million people worldwide are infected with hepatitis C virus (HCV). HCV NS3/4A protease (HCVP) is a key target for HCV therapy. We have identified a class of irreversible HCVP inhibitors that bond covalently to the non‐catalytic Cys159 in HCVP. Cys159 is conserved across all HCV genotypes but is absent in host proteases. MALDI‐TOF MS and in vitro assays show that the lead inhibitor bonds to and inhibits HCVP across multiple genotypes and drug‐resistant variants. In replicon assays covalent silencing leads to potent inhibition of HCVP and prolonged duration of action (>24 hrs) after brief exposure to inhibitor. Covalent silencing yields potent inhibition of HCVP in vivo, in a new mouse model. Inhibitors with this profile may profoundly improve hepatitis C patient treatment across all HCV genotypes. This design strategy is also readily applicable to a wide range of therapeutically important proteases. This work was supported by Avila Therapeutics.
Thermal Stability Assay for Glaucoma‐Causing Myocilin Mutants using Maltose Binding Fusion Protein (MBP)
Authors: Harris Julia Louise , Lieberman Raquel , Burns J. Nicole , Orwig Susan
Published: June 2021

Myocilin is a protein closely linked to inherited forms of glaucoma, a neurodegenerative disorder associated with increased intraocular pressure. In myocilin, over 90% of reported glaucoma‐causing missense mutations are localized to the C‐terminal olfactomedin (OLF) domain. To determine the relative stability of wild‐type and mutant OLF, we developed a fluorescence thermal stability assay without removal of MBP. The fluorescence thermal shift assay can accommodate relatively low concentrations of protein (0.25–0.5 mg/ml) and uses a real time PCR (RT‐PCR) instrument to conduct a slow melt while measuring fluorescence. The excitation and emission settings of the RT‐PCR instrument are compatible with Sypro Orange, a dye known to bind to hydrophobic regions of proteins, which become increasingly exposed as the temperature of the protein‐dye solution is raised. Given upper and lower limits of the fluorescence signal, a melting point can be determined as the midpoint of the unfolding transition. We expressed and purified the OLF (amino acids 228–504) domain of myocilin and select glaucoma mutants (D380A, I477N, I477S, K423E) as maltose‐binding fusion (MBP) proteins in E. coli. In comparison to wild‐type, all four MBP‐OLF mutants tested were destabilized but retained a discrete melting transition expected for a folded protein. Our study appears to be a novel application of the fluorescence stability assay to a MBP fusion protein and lays the foundation for the identification of tailored therapeutic molecules to delay the onset of glaucoma. This work was supported by NSF REU #0851780, Glaucoma Research Foundation (Shaffer Fund), and American Health Assistance Foundation.
Cross‐national differences in early lactation success among primiparous women
Authors: Dewey Kathryn G , Nommsen‐Rivers Laurie A , Matias Susana L , Chantry Caroline J , Peerson Janet M , Cohen Roberta J
Published: June 2021

We evaluated early lactation success in two cohorts of primiparas who initiated breastfeeding: 448 in Sacramento, CA and 171 in Lima, Peru. Key outcomes were sub‐optimal infant breastfeeding behavior (SIBB), onset of stage II lactogenesis >72 h postpartum (delayed OL) and excess neonatal weight loss (≥ 10% of birth weight by 70–98 h). All adverse outcomes were significantly more common in California than in Peru: 69% vs. 52% for SIBB on day 0 (p=0.0004), 34% vs. 21% for SIBB on day 3 (p=0.005), 44% vs. 17% for delayed OL (p<0.0001), and 18% vs. 10% for excess weight loss (p=0.03). For SIBB on day 0, 33% of the site difference could be explained by lower breastfeeding frequency on day 0 in California. For SIBB on day 3, 45% of the site difference could be explained by lower breastfeeding frequency on day 0 and greater prevalence of flat or inverted nipples in California. For delayed OL, 27% of the site difference could be explained by greater maternal age and BMI, greater birth weight and lower Apgar scores in California. For excess weight loss, 91% of the site difference could be explained by greater maternal age and BMI in California. We conclude that the higher prevalence of early breastfeeding problems in California is linked to maternal characteristics and lower breastfeeding frequency soon after birth. Supported by grant R40MC04294 from the Department of Health and Human Services, Maternal Child Health Branch.
Causal directions in the relationship between perinatal depressive symptoms and breastfeeding difficulties
Authors: Vitta Bineti Sidi , Nommsen‐Rivers Laurie A , Cohen Roberta J , Chantry Caroline J , Dewey Kathryn G
Published: June 2021

Perinatal depressive symptoms (DS) are associated with shorter breastfeeding duration, but the causal direction is not clear. Among 415 Californian primiparae who initiated breastfeeding, we examined if 1) prenatal DS are associated with greater risk for breastfeeding difficulties, and 2) breastfeeding difficulties are associated with worsening DS between pregnancy and day 14 (d14) postpartum (pp). Adjusting for socio‐demographic variables and prenatal breastfeeding intentions (logistic regression adjusted odds ratio [95% confidence interval]), women with prenatal DS were more likely to experience nipple pain in the first 24h pp (2.06[1.16–3.65]), between 1–2wk pp (2.27[1.14–4.52]) and 2–4wk pp (2.13[1.03–4.42]); to have babies who were suckling poorly on d7 pp (1.93[1.06–3.51]); and to supplement with formula between 1–2mo pp (2.06[1.14–3.73]). Worsening of DS between pregnancy and d14 pp was associated with poor infant suckling on d7 pp (2.08[1.26–3.42]) and formula use between d3–7 pp (1.63[1.06–2.50]). Thus, the relationship between perinatal depressive symptoms and breastfeeding difficulties may be bidirectional with prenatal DS leading to nipple pain, poor infant suckling, and formula supplementation; and poor suckling and early formula supplementation leading to worsening DS.

Supported by a grant from the Department of Health and Human Services, Maternal Child Health Branch

Grant Funding Source: Department of Health and Human Services, Maternal Child Health Branch
Mammalian sperm are dependent on JAM‐A for normal motility: progressing from mice to humans
Authors: Huynh Jean , Aravindan Rolands , Martin‐DeLeon Patricia A
Published: June 2021

Defective sperm motility (asthenozoospermia) is a primary cause of male infertility. Junctional Adhesion Molecule‐A (JAM‐A) has been recently shown to be essential for mouse sperm motility. However, to date it has not been studied in human sperm. The objective of this study is to determine if JAM‐A is present in human sperm and to characterize its expression. In silico analysis reveals 83% similarity in the amino acid sequence of JAM‐A in mice and humans, indicating that the proteins are conserved. This suggests that JAM‐A may also play a role in human sperm motility. Western analysis of human sperm proteins revealed the presence of JAM‐A with a MW of ~32kDa, identical to that of the mouse. Immunocytochemistry was used to localize the expression of JAM‐A in human sperm. Flow cytometry confirmed the Western blot data and revealed the presence of the protein on the sperm membrane in one of two individuals attending an infertility clinic. Once JAM‐A has been fully characterized in human sperm, the next step will be to determine its interacting protein partners by attempting to co‐immunoprecipitate it with candidate proteins. The localization and characterization of JAM‐A in human sperm is expected to increase our understanding of genetic factors leading to human male infertility and subfertility, laying the groundwork for diagnosis of a subset of couples with fertility issues. Funded by NIH‐COBRE grant #5P20RR015588‐07.
Impact of nitric oxide (NO) treatment on the spectroscopic properties and DNA binding activity of the carbon monoxide‐sensing heme transcription factor, CooA
Authors: Jani Edra , Clark Robert W.
Published: June 2021

CooA is a carbon monoxide (CO)‐sensing heme transcription factor that regulates CO metabolism in several bacteria including Carboxydothermus hydrogenoformans (Ch). The goal of this research was to investigate the spectroscopic properties and DNA binding activity of Fe(III) Ch CooA that was reacted with NO. Based on electronic absorbance spectra, addition of NO to Fe(III) CooA resulted in the formation of a 6‐coordinate (6‐C) Fe(III)‐NO intermediate species, which slowly converted to a 5‐coordinate (5‐C) Fe(II)‐NO complex. A fluorescence anisotropy assay revealed appreciable DNA binding activity by the 6‐C Fe(III)‐NO form and lesser activity by the 5‐C adduct. DNA binding by 5‐C Fe(II)‐NO CooA was unexpected because this species has a much different coordination structure than the native 6‐C Fe(II)‐CO form or the 6‐C Fe(III)‐NO adduct. Based on these findings, we propose that the most important factor required for activating CooA's DNA binding function is the presence of a gas molecule (CO or NO) that is bound to the correct (i.e. distal) face of the heme. The authors wish to thank the Valparaiso University College of Arts and Sciences for financial support.
Design of Ultra‐Sensitive Diagnosis for Anthrax Protective Antigen base on Polyvalent Directed Peptide Polymer (PDPP) Using Horizontally Stacked Hexagonal ZnO Platelets
Authors: Yoon Moon Young
Published: June 2021

Protective antigen (PA) is a component of anthrax toxin produced by Bacillus anthracis. Although Several approaches have proposed to treat PA as it is an attractive target for toxin neutralization, high lethality and potential as a bioweapon place ultra‐sensitive diagnostic system on demand. Most of the current detection methods rely on specificity of antibody toward its cognate antigen. Herein, we proposed peptide as a novel diagnostic probe with several advantages over antibodies. We applied a novel technique for ultrasensitive detection of PA83 using an array of zinc oxide nanorods in conjunction with a FITC‐labeled peptide which showed high affinity with PA. Therefore, the use of zinc oxide nanorods as fluorescence enhancing substrates permitted attomolar detection sensitivity of proteins. However, the peptide to use as diagnostic probe gives a limit which the binding affinities of peptide reactivity lower than that of antibody. Consequently, our research introduced the concept of the polyvalent directed peptide polymer (PDPP). The PDPP will be able to overcome that a peptide has low binding affinity because the multiple peptides in polymer relatively increase to affinity that binding in the various regions as exists with many kind peptides, and as the possible of cooperative binding with diagnostic target. Also, the signal amplification effect will be able to expect which the various peptide diagnostic probes binding in the diagnosis polymer chain. The design of PDPP is now viable alternatives to bio‐diagnostic systems, such as antibodies.
Genistein mediates perturbations in one‐carbon metabolism during diet‐induced folate deficiency
Authors: Nonnecke Eric B , Schalinske Kevin L
Published: June 2021

Populations consuming isoflavone‐rich diets exhibit lower prevalence of heart disease and estrogen‐responsive cancers. Genistein, a17β‐estradiol‐like isoflavone, has been shown to modify the fetal epigenome in utero in the agouti mouse model and modulate DNA methylation patterns in estrogen‐responsive tissues. Moreover, it also exhibits antiproliferative action in cancerous cells via suppression of S‐adenosylmethionine –dependent DNA methyltransferases, leading to site‐specific hypomethylation. However, it is not known if genistein modulates one‐carbon metabolism, a process that when disrupted leads to dysregulation of the epigenome. The objective of this study was to characterize the metabolic actions of genistein during moderate dietary folate deficiency. Male Sprague Dawley ratswere fed an ad lib control or folate‐deficient (FD) diet with or without genistein supplementation (300 mg/kg diet) for 8 wk. Plasma homocysteine (Hcy) concentrations in genistein‐supplemented, FD animals (16.7 μM) was lower (P = 0.02) than the Hcy concentration of FD animals not supplemented with genistein (27.9 μM). Hepatic enzyme analysis suggests the ability of genistein to attenuate hyperhomocysteinemia in FD rats is not linked to diminished Hcy production. Clearly, genistein has the ability to modulate Hcy and methyl‐group metabolism; however, the underlying mechanisms remain unknown.

Grant Funding Source: AHA assn
In vivo enrichment of chicken eggs with 14C‐B12 for determining vitamin B12 bioavailability in humans
Authors: Garrod Marjorie G , Miller Joshua W. , Calvert Christopher C. , Buchholz Bruce A. , Green Ralph , Johnson Heidi A. , Allen Lindsay H.
Published: June 2021

Animal source foods such as eggs are the only natural dietary source of vitamin B12 but there is limited information on its bioavailability from these sources. The objective was to test the feasibility of enriching eggs in vivo with sufficient 14C‐B12 for use in human bioavailability studies. 14C‐B12 was biosynthesized using Salmonella enterica, purified and injected into the thigh muscle of a laying hen. Eggs were collected 21 d, pooled, scrambled and cooked. A serving of egg containing ~0.7 μg 14C‐B12 (~30 nCi) was fed to each of 10 healthy humans after an overnight fast. 14C, detected by accelerator mass spectrometry, first appeared in plasma at 3–4 h after dose and peaked at 7–10 h, with 5–40% appearing in urine in the first 24 h. Fecal 14C analyses in progress will enable calculation of % bioavailability of B12. We conclude that this protocol can be used for human B12 bioavailability studies. Funded by the American Egg Board.
Phenotypes in utilization of formate for purine nucleotide biosynthesis de novo in adult humans
Authors: Baggott Joseph E , Gorman Gregory S , Morgan Sarah L
Published: June 2021

Folate coenzymes and their dependent enzymes introduce one‐carbon units at positions 2 (C2) and 8 (C8) of the purine ring during de novo biosynthesis. Formate is naturally occurring in humans and is one source of one‐carbon units. Although much is known about lower organisms, little data exist describing formate utilization for human purine biosynthesis. After IRB approval, mass spectrometric analysis of urinary uric acid, the final purine catabolite, following 1.0 g oral doses of [13C]sodium formate was performed in 6 healthy male adults. Using this method, we are can detect 13C enrichment at C2 and C8 independently. Our data suggested the presence of three phenotypes including: one phenotype where 13C was incorporated into C2 much greater than into C8, and in another, there was little or no 13C incorporation into the C2 or C8 position. In the third phenotype, 13C was incorporated into C8; but, C2 incorporation was greater. In subjects who incorporated [13C]formate into C2, peak enrichment occurred in voids from the 8–12 h period (24‐h clock). Our results are not similar to those in non‐mammalian organisms or cultured animal cells. The presence of these phenotypes may reflect genotypes in the purine biosynthesis pathway.
Selenocysteine abundance was decreased in the hepatic GPX1 protein of SOD1−/− mice
Authors: Wang Shi Kui , Zhang Sheng , Lei Xin Gen
Published: June 2021

We previously reported a 40% loss of specific activity in the GPX1 protein purified from the SOD1−/− mouse liver homogenate compared to that of the wild type mice. This study was to explore structural modifications for that GPX1 activity loss. After liver homogenates (200 μg of protein) of both SOD1−/− and wild type mice were pre‐cleared and incubated with the anti‐GPX1 antibody (Abfrontier, Seoul, South Korea) and the protein G agarose beads, the precipitates were subjected to SDS‐PAGE for separating the GPX1 protein band that was excised and digested in situ. The in‐gel‐extracted tryptic peptides were reconstituted for nanoLC‐MS/MS analysis. The identified peptides covered over 90% GPX1 protein sequence for both genotype samples. Notably, the 47th residue in the tryptic peptide (residues 37–52) of GPX1 exhibited two forms: the projected native selenocysteine and the Se‐lost dehydra‐alanine. Using extracted ion chromatogram and multiple reaction monitoring analyses, we found the ratio between the SOD1−/− and the wild type for selenocysteine and the dehydra‐alanine substitute was 0.65 and 1.3, respectively. There were no genotype differences in several other peptide modifications on the hepatic GPX1 protein. In conclusion, the relative abundance change of the selenocysteine residue into the dehydra‐alanine form in the hepatic GPX1 protein of the SOD1−/− mice matched closely to the decrease of the enzyme specific activity. Likely, the physiological level of SOD1 activity plays an important role in maintaining the integrity of selenocysteine in the hepatic GPX1 protein in vivo. (NIH DK 53018 TO XGL).
Effect of Physiological Levels of Sodium Selenite on the Expression and Regulation of Hypermethylated Tumor Supressor Gene in Human Breast Cancer Cell Line
Authors: Chen Yu‐Hsin , Yang Feili Lo , Kao Cheng‐Fu , Chen Yaling
Published: June 2021

Breast cancer (BC) was the 4th‐leading cause of cancer death in Taiwan with the rising incidence as malignant type in young premenopausal women during the recent decades. The aim of our study was to investigate the effects of physiological levels of selenite on regulation of BC tumor suppression gene through hypermethylation. Exposure of human breast cancer cell line MDA‐MB 231 to 1.5 and 2.5 μM of sodium selenite for 7 days significantly decreased cell invasion and migration ability compared to control. Selenite treatment (2.5 μM) reactivated the anti‐invasion gene CST6 (cystatin E/M), of which the promoter region hypomethylation was confirmed by methylation‐specific PCR. The protein expression of DNA methyltransferase £L (DMNT1) and histone deacetyl transferase I were also significantly decreased by selenite treatment. In conclusion, the suppressed invasion ability of MDA‐MB 231 cells by physiological levels of selenite may be due to the suppressed protein expression of DNMT1 as the result of reactivating the anti‐invasion genes such as CST6, of which the promoter region were hypomethylated. The results of this study are expected to explain the role of selenium in regulating the re‐expression of specific tumor suppressor genes and to be applied in improving the prognosis of breast cancer.(NSC98‐2320‐B‐030‐004)

Grant Funding Source: NSC98‐2320‐B‐030‐004
Hockey stick relationship between 25 OH vitamin D and BMI NHANES
Authors: Arab Lenore , Kim Heeyoung , Ebel Alice
Published: June 2021

Circulating levels of vitamin D are shown to be rapidly decreasing in the US population, with up to 90% of the population and more so in the African American population. A negative association between serum levels of 25 OH vitamin D and body mass index levels has also been consistently documented, suggesting that this fat soluble vitamin may be sequestered in adipose tissue. To examine more closely the true nature of this relationship between body fat and circulating levels of vitamin D we used smoothing methods on NHANES adult Caucasian and African American populations. These findings suggest that the nature of the relationship is not linear and reflects more closely an inverted hockey stick. Individuals whose BMIs are under 20 having lower than expected circulating levels of vitamin D, values rise and then fall again among all individuals with a BMI over 20. This is seen in both genders, African Americans and Caucasians. Among African Americans, the reduction in circulating levels of 25 OH vitamin D per unit of BMI is approximately half that seen in Caucasians. The race, gender and age adjusted regression coefficients of BMI on vitamin D levels demonstrated an intercept for African Americans of 17.7, and a coefficient of −0.169. Among Caucasians the comparable values are 37.7 and −0.339, suggesting that among those with the lowest levels of vitamin D, sequestration of vitamin D into adipose tissue is lower.
Physical exercise practice influences serum concentrations of 25(OH)D and PTH in spinal cord injured men.
Authors: Chain Amina , Koury Josely C , Figueiredo Flávia A , d'Oliveira Giselle LC , Bezerra Flávia F
Published: June 2021

The aim of the study was to evaluate the influence of physical exercise practice on serum concentrations of 25(OH)D and PTH in spinal cord injured adult men (19–56 y, n = 25). Hours of training per week were estimated by using a self‐reported questionnaire. Serum concentrations of 25(OH)D and intact PTH were assessed by a chemiluminescent enzyme‐labeled immunometric assay. Serum 25(OH)D varied in a range between 27.5–107.5 nmol/L, being 15 subjects below 75 nmol/L and 3 subjects below 50 nmol/L. Serum iPTH was on average 44.9 ± 15.0 pg/mL and correlated negatively with 25(OH)D (r= −0.45, P<0.05). Hours of physical exercise practice was on average 9 ± 11 h/week. After controlling for age and time after injury, hours of training per week was positively correlated with serum 25(OH)D (r= 0.54, P<0.05) and negatively correlated with serum iPTH (r= −0.54, P<0.05). Our results indicate that most of the spinal cord injured men studied had serum 25(OH)D concentrations below the proposed cutoff (≤ 75nmol/L) to define insufficiency. Moreover, these results suggest that physical exercise practice may contribute for improving vitamin D status in adult spinal cord injured men. Financial support: FAPERJ (Brazil).
OBESITY PREVALENCE ACCORDING CALCIUM STATUS IN ADOLESCENT WOMEN.
Authors: Rio Maria Esther , Felipoff Ana Lia , Zago Liliana , Piazza Norma
Published: June 2021

Adolescence is a highly vulnerable period of life. Multiple changes occur which often translate into bad eating habits among them, shortcomings in the intake of calcium (Ca), problem which has received special attention since recent investigations have shown an association between high Ca intakes and prevention of adiposity. Here we study the influence of Ca status (CaS) on overweight and obesity in 130 adolescent females 10 to 18 years old. Diagnosis of Normality (N), Overweight (OW) and Obesity (O) was performed by IMC (WHO 2007) (N: zIMC < +1DS and >‐2DS; OW: zIMC > + 1DS; O: zIMC > +2DS) and maturity by Tanner (T). Ca status was assessed in basal urine samples by the Ca/creatinine ratio (Ca/C) using AA spetrophotometer and a commercial kit, respectively. CS was defined as: <0.07, deficiency (D); 0.07–0.15, adequate (A); > 0.15: high (H). The obese adolescents accounted for 41 % of (D) group declining to 35 % for (A) and 14 % for (E). The behavior was reverse in those with adequate BMI: 30 % of the Group (D) increasing to 46 % in (A) and 57 % in (H). Also, the average value of zIMC by range of Ca/C decrease from 1.4 + 1.30 to (D) to 1.00 + 1. 44 to (A) and 0.15 + 1.67 for (H) (global ANOVA, p= 0.0070, Tuckey p<0.01 between (D) and (H). Calcium deficiency is a risk factor for obesity, as seen in D, in addition to the Zemel findings concerning the protective effect of Calcium high intakes, as shown by E group.

Supported by UBACyT B/060.
Antiobesity effects of bioceramic stone water in C57BL/6 mice
Authors: Bak Soon‐Sun , Kim Ji‐Young , Lee Sun‐Mi , Byun Jae‐Man , Park Kun‐Young
Published: June 2021

The antiobesity effects of bioceramic stone (BCS) water investigated under in vivo using C57BL/6 mice. The BCS water was made by dipping a pair of SY bioceramic stone (BCS, diameter: 5 cm, thickness: 1 cm) into 20 L of tap water for 24 h. The C57BL/6 mice had free access to high fat diet pellet (D12451) and either the BCS water (BCS group) or tap water (control group) for 6 month. Weight gain, body weight, and fat tissue weight showed a different trend between BCS and control groups, in spite of similar food intakes. The levels of plasma cholesterol, triglycerides, leptin, and insulin in the BCS group were significantly lower than that of control group (p < 0.05). Protein expression of adipogenesis gene markers such as PPARγ, SREBP1c, and C/EBPα in adipose tissue on BCS group were down‐regulation. PPARα and adiponectin protein expressions (lipolysis gene markers) in the liver of the BCS group were increased compared to the control group. Moreover the BCS group increased aquaporin (AQP)‐7 in adipocyte and AQP‐9 in the liver of C57BL/6 mice compared to the control group. This study suggests that the BCS water has antiobesity effects on C57BL/6 mice and it was associated with adipogenesis, lipolysis, and water transport.
Coffee is the most important contributor to the total antioxidant capacity in Brazilians’ diet
Authors: Torres Taíssa , Farah Adriana
Published: June 2021

The consumption of antioxidant compounds has been associated to a number of potential benefits to human health, including the protection against chronic degenerative diseases. In general, antioxidant compounds with different chemical characteristics function in integrated and complementary ways, rather than individually, to protect the human body against oxidative damage. Therefore, the total antioxidant capacity (TAC) of foods has been considered to be the best tool to monitor this protective effect. The aim of the present study was to evaluate the relative contribution of different food products to the TAC of Brazilian's diet. For this, the most consumed food products in Brazil (n = 41) were selected considering the national survey database from “IBGE”, which ranks the annual per capita food consumption by the population. These products accounted for 76% of all plant food consumed in Brazil and 58% of all food intake. TAC was measured by two methods (FRAP and TEAC). Coffee was the most important contributor to TAC among the analyzed food products, contributing with 85% and 69% of dietary TAC, when evaluated by FRAP and TEAC, respectively. The remaining beverages, including wine, fruit juices, mate tea, among others, contributed with only 3% and 8% of TAC, respectively. Beans contributed with 3% and 8%; cereals and derivatives with 5% and 7%; fruits with 3% and 6%; and vegetables with only 2% and 5%, respectively.
Antioxidative Effects of New Herbal Formula Extracts
Authors: PARK SUNG JIN , KIM NA YOUNG , PARK SUNG HYE , YOON WON BYONG , KIM JONG DAI
Published: June 2021

The validity of herbal prescriptions has been widely accepted by the scientific medical establishment, and their use has been increasing. In this study, we examined the antioxidant capacity of the extracts from new herbal formula(NHF; sprout buckwheat, Chrysanthemum indicum L., Akebia caulis., lxeris dentate Nakai, Plantago asiatica L., Sasamorpha purpurascens(Hackel) Nakai var. borealis(Hackel) Nakai., Oenanthe stolonifera DC.) in vitro using DPPH free radical scavenging assay, reducing power assay, total phenolic compounds, and total flavonoid contents. At the water extract, yield, DPPH, total phenolic compounds, total flavonoid, and reducing power in the extract of NHF were 45.14%, 81.35%, 5.47 mg/mL 2.21 mg/mL and 1.08 mg/mL, respectively. This study demonstrated that NHF extract has significant antioxidant properties.
Development of medicinal food recipes including bitter gourd, fenugreek and guava for type II diabetes
Authors: Syu Kai‐lo , Tam Chick
Published: June 2021

Objective

Using potential health benefits of bitter gourd (Momordica charantia), fenugreek (Trigonella foenum graecum) and guava (Psidium guajava) as medicinal foods for the dietary management of the type II diabetes.

Method

Use internet research, peer‐reviewed published papers and books that deal with antidiabetic components of these fruits and vegetables.

Result

Bitter gourd contains a polypeptide called gurmarin and alkaloid momordicine, the bitter component in the gourd. Fenugreek has unique dietary fiber composition and high saponin content to act as hypoglycemic and hypocholesterolemic agents in both animal and human studies. Guava leaves possess flavonoid glycosides such as pedunculagin, strictinin, and isostrictinin which are the effective constituents that have been used in clinical treatment of diabetes to improve the sensitivity of insulin. Various recipes were developed and subjected to computer nutrient analyses to insure nutritional adequacy.

Conclusion

These antidiabetic components from bitter gourd, fenugreek, and guava can be incorporated into recipes with new possibilities of improve and/or prevent the Type II diabetes and/or its clinical complications.
Effects of Ecklonia cava on plasma and liver lipids, platelet aggregation and RBC membrane stability in rats fed cholesterol diets
Authors: Yeh Wan Ting , Jeon You‐Jin , Kang Young‐Hee , Kang Jung Sook
Published: June 2021

We compared bioactivity of 10% Ecklonia cava (EC) powder, 5% extract (carbohydrase hydrolyzed product) and 3% Dieckol (EC polyphenol) in aspect of antiobese, hypolipidemic, antiplatelet and antioxidant on RBC membrane and TBARS production in rats fed cholesterol diets.

The final weight with low food efficiency ratio in rats fed Dieckol was significantly decreased compared with the control (p<0.05). Plasma total cholesterol was significantly decreased in EC powder (p<0.05) and HDL‐cholesterol was significantly decreased in Dieckol (p<0.05) both compared with the control. Liver total cholesterol significantly decreased in EC powder compared with EC extract (p<0.05) and liver TG was significantly decreased in Dieckol compared with control (p<0.05). Platelet agregation both in initial slope and the maximum was significantly decreased in Dieckol compared with the control (both, p<0.05). Erythrocyte Na leak increase after AAPH treatment was significantly high in EC extract compared with other groups (p<0.05), while PRP TBARS porduction was significantly decreased in Dieckol compared with the control (p<0.05). GOT compared with the control and GPT compared with Diekol were significantly decreased in rats fed EC powder (p<0.05).

In conclusion, polyphenol rich Dieckol from Ecklonia cava seems to have some favorable effects in weight gain, platelet aggregation, RBC membrane stability and lipid peroxidation, but it is not clear how it acts at this point.
Walnut type affects the response to occlusion‐induced vasodilation following a high fat meal in dyslipidemic humans
Authors: Fitschen Peter J , Allen Brian K , Manzy Michelle A , Maher Margaret A
Published: June 2021

Endothelial dysfunction, resulting in impaired vasodilation, has been shown in persons with dyslipidemia. English walnuts (EW) improve such impaired vasodilation following a high fat meal, compared to other fat sources such as olive oil. Our lab has studied the effects of both EW and black walnuts (BW) on plasma lipids, but the effect of BW on endothelial function has not been studied. Thus, we compared the effects of 40 g BW versus EW with a high fat meal on vasodilation. Six dyslipidemic (LDL over 130 mg/dl) subjects were tested on two days at least one week apart. On test days, subjects ate a standardized breakfast at 7:00 am and arrived at noon for vasodilation testing. Baseline ultrasound of the brachial artery before and after 4.5 minute occlusion, was followed by high fat meal consumption. Four hours post‐meal, vasodilation testing was repeated. The high fat meal impaired the vasodilation response in both groups as expected, but the mean pre‐post meal vasodilation difference was significantly (p<0.02) greater (−8.7±2.9) with BW versus EW (−1.5±3.1). Changes in after‐occlusion blood flow rate (cm/s) following the high fat meal were not significantly different between EW and BW. Our results indicate that EW improved endothelial dysfunction compared to BW following a high fat meal in dyslipidemic subjects. Funded by University of Wisconsin‐System, WiscAMP, and NSF‐REU grants.
Royal jelly enhances migration of human dermal fibroblasts with decreased levels of triglycerides and cholesterol in in vitro wound healing model
Authors: Kim Youngae , Yun HJ , Lee JA , Park HM , Kim SY , Cho Y
Published: June 2021

Oral administration of royal jelly (RJ) promotes the wound healing in diabetic mice. Concerns have arisen about the efficacy of RJ on wound healing process of normal skin cells. In this study, a wound was created by scratching normal human dermal fibroblasts, one of major cells involved wound healing process. 0.1, 1.0 or 5 ug/ml RJ were promptly treated up to 48 hrs, and migration was analyzed by the closure of wound margin. Furthermore, altered levels of lipids, which is recently reported to be responsible for wound healing process, were analyzed by HPTLC. Migration of fibroblasts was peaked up to ~24hr after wounding. RJ treatment significantly accelerated migration of fibroblasts in a dose dependent manner at 8hr. Although RJ accelerated migration of fibroblasts at either 20 hr or 24hr, the efficacy of RJ was less portent than at 8hr. Among various lipid classes of fibroblasts, the levels of triglycerol and cholesterol were significantly decreased with 5 ug/ml RJ. In spite of a dose‐dependent increase of sphinganine, no altered level of sphingosine, ceramides and glucosylceramide was noted with any concentration of RJ. We demonstrate that RJ enhances migration of fibroblasts with altered levels of various lipids in the wound healing process.

This work is supported by Biogreen21 grant (20090801‐060‐001‐001‐02‐00) of Rural Development Administration