bluuebunny/crossref_metadata_2025_split huggingface dataset:
Selenium‐dependent Resolution of Inflammation in Macrophages is mediated through Δ12PGJ2‐CRTH2 Signaling Axis
Authors: Bastihalli‐Tukaramrao Diwakar , Nettleford Shaneice , Prabhu K. Sandeep 
Published: November 2023

Selenium in the form of selenoproteins is known to mitigate inflammation by differential regulation of cyclooxygenase (COX)‐derived prostaglandin (PG) synthesis in endotoxin lipopolysaccharide (LPS)‐treated macrophages as well as in other models of inflammation. During late phase of inflammation, MΦs supplemented with selenium produce elevated levels of pro‐resolution cyclopentenone prostaglandins (CyPGs), such as Δ12‐PGJ2 that downregulate inflammatory signalling pathways. MΦs express the chemoattractant receptor‐homologous molecule, a cell surface receptor, expressed on Th2 cells (CRTH2/DP2) that exhibits high affinity for Δ12‐PGJ2. In this study, we investigated the role of CRTH2 signalling during LPS‐induced inflammation in murine macrophages cultured in the presence or absence of selenium. Deletion of Crth2 in MΦs, using the CRISPR/ Cas9 system, resulted in significant increase in the expression of genes required for PG biosynthesis (COX‐2 and microsomal PGE2 synthase‐1) and pro‐inflammatory mediator such as iNOS, mimicking the effects seen in selenium deficiency. Furthermore, use of specific pharmacological antagonists of CRTH2, inhibition of heterotrimeric G protein downstream of receptor using pertussis toxin, or use of specific inhibitors of enzymes leading to the biosynthesis of Δ12‐PGJ2 all increased expression of proinflammatory genes, indicating a previously underappreciated feedback mechanism of autocrine/paracrine role of Δ12‐PGJ2‐mediated CRTH2 signalling in selenium‐dependent resolution of inflammation.Support or Funding InformationFunded by NIH R01 DK077152 to KSP
Mechanistic Aspects of Selenium Delivery by Selenoprotein P
Authors: Copeland Paul , Shetty Sumangala 
Published: November 2023

Selenoprotein P (SELENOP) is a serum glycoprotein that is required for proper selenium distribution in mammals, particularly in supplying selenium to the brain and testes. As the sole mechanism for providing essential selenium to developing spermatazoa, SELENOP metabolism is central to male fertility in all mammals. In addition, this process is important for proper brain function, especially under conditions of limited dietary selenium. Several specific and nonspecific mechanisms for SELENOP uptake in target tissues have been described but the utilization of the protein as a source of selenium for intracellular selenoprotein production has not been systematically characterized. We have examined the process by which SELENOP delivers selenium to cells using a robust selenium uptake assay that measures selenium utilization in cells fed 75Se‐radiolabeled SELENOP. Using a series of inhibitors and modulators we have identified specific requirements and regulators of the process.Support or Funding InformationThis work was supported by the Found Animals Foundation and NIH GM077073
Folic Acid and Riboflavin Deficiencies in Murine Macrophage RAW Cells Reduce LPS‐induced Nitric Oxide Production
Authors: Rittmann Marijke , Miller Joshua W , Gow Andrew 
Published: November 2023

Methylenetetrahydrofolate reductase (MTHFR) is a flavin adenine dinucleotide (FAD)‐dependent folate metabolizing enzyme that catalyzes the conversion of dihydrobiopterin to tetrahydrobiopterin (BH4). BH4 is an essential cofactor for nitric oxide synthase (NOS) that produces nitric oxide for multiple functions including vasodilation and blood pressure (BP) regulation. Genome‐wide association studies have linked a common polymorphism, C677T, in MTHFR with BP, and riboflavin supplements have been shown to reduce BP in individuals homozygous for the 677TT variant form of the enzyme. We hypothesize that this relationship between MTHFR, riboflavin, and BP is mediated through nitric oxide synthesis. We investigated this hypothesis in vitro in stimulated murine macrophage RAW cells, which express the inflammatory form of NOS, exposed to standard and reduced levels of folic acid and riboflavin. RAW cells were grown in medium with 0.4 mg/L riboflavin and 4.0 mg/L folic acid (control), 0.04 mg/L riboflavin (lowB2), or 0.4 mg/L folic acid (lowFA) for 48 hours, and then exposed to 100 ng/ml or 1000 ng/ml lipopolysaccharide (LPS) for 24 hours. Media was then analyzed for nitric oxide production by chemiluminescence assay using a Nitric Oxide Analyzer. Quantitative PCR was used to analyze gene expression of inducible NOS (iNOS). In all three media conditions, no differences in RAW cell proliferation rates were observed over 48 hours. After LPS exposure, nitric oxide production in the lowB2 and lowFA cells was 30–35% and 35–40% of the control cells, respectively (p ≤ 0.02). Expression of iNOS after LPS induction increased in all three media conditions. LPS‐induced nitric oxide production is reduced in RAW cells grown in either riboflavin or folic acid deficient media independent of iNOS expression. These results demonstrate the importance of the folate cycle in maintaining NOS function, and indicate a potential mechanism for the effects of MTHFR polymorphisms on BP.Support or Funding InformationDepartment of Nutritional Sciences, Rutgers University, New Brunswick, NJ
Modifications to Ascorbate Concentration and pH Result in Faster Deconjugation of Whole Blood Folate Polyglutamates to Monoglutamates for Measurement by Liquid Chromatography‐Tandem Mass Spectrometry
Authors: Stamm Rosemary A , Fazili Zia , Pfeiffer Christine M 
Published: November 2023

ObjectiveFolate in red blood cells is polyglutamylated and requires deconjugation of glutamate residues by the enzyme γ glutamyl hydrolase (GGH) (E.C. 3.4.19.9) during an incubation step prior to analysis. The reported pH optimum for human GGH is 4.5. A lengthy incubation time is required for measurement of whole blood total folate by liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) to ensure all folates are measured as folate monoglutamates. We aimed to reduce the time taken for enzymatic cleavage of folate polyglutamates to monoglutamates.MethodsWe investigated modification of pH and ascorbate concentration to the hemolysate diluent to optimize deconjugation of folate polyglutamates. Preliminary testing was performed on four samples, two with high non‐methyl folate concentrations.ResultsThe conventional dilution of whole blood (1/11 in 1% ascorbic acid solution, pH 2.7) resulted in a mean hemolysate pH of 3.8. Dilution with 0.5% ascorbic acid (pH 2.9) resulted in a mean hemolysate pH of 4.2 and dilution with 1% ascorbic acid adjusted to pH 4.1 resulted in a mean hemolysate pH of 4.4. Both conditions resulted in <1% of 5 methyltetrahydrofolate (5‐methylTHF) remaining as dilglutamate after 3 h incubation at 37°C, while ~2% of 5 methylTHF remained as diglutamate after 4 h incubation in conventionally prepared hemolysates. However, both conditions also resulted in a greater loss of tetrahydrofolate (THF) internal standard compared with the conventional dilution, likely resulting in a corresponding greater overestimation of THF concentration. In a follow up experiment, 2% sodium ascorbate solution offered better protection of THF internal standard compared with 1% sodium ascorbate (both adjusted to achieve a hemolysate pH of 4.3). However, 2% sodium ascorbate resulted in inadequate deconjugation with >4% of 5 methylTHF remaining as diglutamate after 4 h incubation, while <1% remained as 5‐methylTHF diglutamate in 1% sodium ascorbate.ConclusionThe conventional dilution of whole blood results in a pH that is more acidic than the pH optimum of human GGH. Faster and more complete deconjugation of folate polyglutamates can be achieved either by reducing the ascorbic acid concentration or increasing the diluent pH to achieve a hemolysate pH closer to 4.5. Increasing the ascorbate concentration provides additional protection of THF, however, results in slower and less complete deconjugation, thereby resulting in under‐recovery of total folate. An additional means of protecting THF during incubation is required to accurately measure whole blood hemolysate total folate by LC‐MS/MS.
Folic acid intake among U.S. women of childbearing age with suboptimal red blood cell folate concentrations for neural tube defect prevention
Authors: Qi Yan Ping , Tinker Sarah , Berry Robert , Crider Krista 
Published: November 2023

BackgroundA recent study reported that 22.8% of U.S. women of childbearing age have folate insufficiency, defined as red blood cell (RBC) folate below optimal concentrations needed for neural tube defect (NTD) prevention per guidelines established by the World Health Organization. Clinical trials have shown that consuming 400 μg of folic acid daily prevents NTDs.ObjectiveWe assessed the daily usual total folic acid intake and RBC folate status of women of childbearing age. We estimated the proportion of women consuming ≥400 μg folic acid/day, the daily amount recommended for NTD prevention for all women capable of becoming pregnant.MethodsWe used 2007–2012 data on non‐pregnant women aged 12–49 years (n = 4783) from the National Health and Nutrition Examination Survey (NHANES) and estimated daily usual total folic acid intake from all fortified foods and dietary supplements. Folate insufficiency was defined as an RBC folate concentration of <748 nmol/L using the NHANES microbiologic assay, which is associated with prevalence of NTDs of ≥9 cases per 10,000 live births.ResultsAmong all women in our study, 21.2% (95% confidence interval [CI]: 18.0, 24.3) had an average total folic acid intake of ≥400 μg /day from fortified foods and supplements. The prevalence of folate insufficiency was 22.5% (95% CI: 20.3, 24.9). Among the subgroup of women with folate insufficiency (n = 1196), only 3.9% (95% CI: 2.0, 5.7) achieved the recommended daily intake of folic acid. Among all women in our study, median usual daily total folic acid intake was highest among non‐Hispanic Whites (264 μg) and lowest among non‐Hispanic Blacks (185 μg). However, among women with folate insufficiency, the median usual daily total folic acid intake did not differ by race/ethnicity. Among women with folate insufficiency, 69.7% (95%CI: 64.8, 74.1) had mandatorily fortified foods (i.e., enriched cereal grain products) as their only source of folic acid; 12.6% (95% CI: 9.7, 16.1) reported taking supplements containing folic acid in the previous 30 days. Among folate‐insufficient women who did not use supplements, less than 1% had usual total folic acid intake ≥400 μg/day.ConclusionsThe majority of U.S. women of childbearing age with RBC folate below the optimal concentration do not consume the amount of folic acid recommended for NTD prevention. Targeted programs and guidance might increase folic acid intake among groups of women at higher risks for folate insufficiency.Support or Funding InformationThis work was supported by the CDC.
Association Between One‐carbon Metabolism Indices and DNA Methylation Status in Maternal and Cord Blood
Authors: Park Hea Jin , Knight Anna K , Fleming Jennifer M , Hausman Dorothy B , Caudill Marie A , Malysheva Olga V , Kauwell Gail PA , Sokolow Andrew , Fisher Susan , Smith Alicia K , Bailey Lynn B 
Published: November 2023

One‐carbon metabolism is a crucial component of metabolism in all cells and tissues supporting DNA methylation and DNA synthesis. It is critically important during pregnancy to ensure normal cell division and growth of fetal and maternal tissues; however, the impact of one‐carbon metabolism indices during pregnancy on DNA methylation has not been investigated. In this study, status of one‐carbon metabolism indices and DNA methylation in maternal blood and cord blood were measured. One‐carbon metabolism indices included serum folate, red blood cell folate, plasma folic acid, plasma 5‐methyltetrahydrofolate (5MTHF), plasma methionine, plasma choline and choline metabolites (dimethylglycine (DMG), betaine, trimethylamine N‐oxide), plasma homocysteine, plasma SAM, SAH and the ratio of SAM/SAH and plasma vitamin B12. Healthy pregnant women (n=28) were recruited at a local midwifery clinic (Athens Regional Midwifery Clinic, Athens, GA), and maternal blood was collected at baseline (<12wk gestation) and delivery. Umbilical cord blood from the mothers was also collected at delivery. DNA methylation in maternal and cord blood were measured across > 485,000 CpG sites using the Infinium Human Methylation 450 BeadChip. Using a linear mixed‐effects model, we examined the relationship between DNA methylation of each CpG site and gestational week to determine CpG sites that change over pregnancy in maternal blood. We identified 993 such CpG sites (FDR<.05), which were then interrogated for associations with one‐carbon metabolism indices. In maternal blood, a CpG site was associated with DMG levels after multiple test correction. Interestingly, in cord blood, methylation of three CpG sites was negatively associated with 5MTHF level (FDR<.05). These CpG sites were annotated to Myc‐associated Zinc finger protein (MAZ), discs large homolog 2 (DLG2), and paraneoplastic Ma antigen (PNMA1), which have previously been associated with cancer progression and neurological function. Results suggest a unique association between one‐carbon metabolism indices and methylation status of specific CpG sites in cord blood. Future research should address changes in methylation of these genes in studies of brain development and neural tube defects.Support or Funding InformationSupport provided by Georgia Experimental Agricultural Station, HATCH #GEO00706 and #GEO00707, Obesity Initiative at the University of Georgia (UGA) and UGA Office of Vice President for Research.
Relationships among one‐carbon nutrients status, DNA methylation at H19 differentially methylated region and the long interspersed nucleotide elements, and serum insulin‐like growth factor levels of pregnant women
Authors: Huang Kwang‐Ta , Shen Yu‐Li , Lee Chien‐Nan , Cheng Chin‐Pao , Ku Wei‐Chi , Huang Rwei‐Fen S. 
Published: November 2023

Kwang‐Ta Huang1,2, Yu‐Li Shen3, Chien‐Nan Lee 4, Chin‐Pao Cheng5, Wei‐Chi Ku6, and Rwei‐Fen S. Huang1,41Ph.D. Program in Nutrition and Food Science, Fu Jen University2LaBor Birth Center, Xinzhuang Dist., New Taipei City, Taiwan3Department of Nutritional Science, Fu Jen University,4Department of Gynecology and Obstetrics, National Taiwan University Hospital, Taipei;5Dietetic Department, National Taiwan University Hospital, Taipei, Taiwan5Department of Medicine, Medicine College, Fu Jen University, TaiwanAberrant methylation at the H19 differentially methylated region (H19 DMR) or imprinting center 1 (IC1) of cluster imprinted genes including insulin‐like growth factor (IGF2) during fetal development was associated with increased expression of IGF2 and increased chronic diseases risks in offspring. The early environmental factors to modify DNA methylation of the H19 DMR and IGF2 expressions remain unclear. In this study, we examined the determinants of maternal DNA methylation at the specific H19 DMR and the long interspersed nucleotide elements (LINE‐1), a one‐carbon nutrients responsive genomic methylation sensor, in PMBC of 116 first‐term pregnant females, and their associations with disease risk markers of serum IGF2. By linear regression analysis, RBC folate (beta = −0.235, P = 0.028) and plasma free choline levels (beta = −0.230, P = 0.026) were the most significant determinants of LINE‐1 methylation status in PMBC of subjects. The LINE‐1 methylation status was the most significant determinant of methylation status at H19 DMR (beta = −0.235, P = 0.028). Degree of PBMC LINE‐1 methylation was positively correlated with methylation at H19 DMR (r = 0.373, P < 0.001) and inversely associated with serum IGF2 levels (r = −0.199, P = 0.045). Serum folate (r = 0.258, P = 0.08) and betaine levels (r = 0.227, P = 0.026) were positively correlated with serum IGF2 levels, with no modifying effect on H19 DMR DMR. Plasma betaine level was the most significant determinant of serum IGF2 levels (beta = 0.223, P = 0.029). The data suggested that maternal H19 DMR corresponded to global LINE‐1 methylation which correlated with expression of serum IGF2 levels, which are modified by serum folate and betaine status of pregnant women. The data support the critical role of maternal one‐carbon nutrient status in modifying risks factors of chronic diseases during pregnancy.
Effect of Marginal Zinc Intake and Repletion on Essential Fatty Acid Metabolism
Authors: Suh Jung , Burke Sarah , Shigenaga Mark , Killilea David , Holland Tai , Shenvi Swapna , Sutherland Barbara , King Janet 
Published: November 2023

ObjectiveTo determine the effects of marginal zinc deficiency and repletion on essential fatty acid, sphingolipid, and lipoprotein metabolism.MethodsSixteen apparently healthy male subjects between ages of 18–45 were subjected to three sequential phases of dietary zinc intake modulation: Phase 1: Low Zinc intake: 6 mg/day with 1.5 g of phytate (2 wks); Phase II: Zinc Repletion: 10 mg/day with no phytate (4 wks); Phase III: Zinc Supplementation: Ad lib diet supplemented with 20 mg zinc (2 wks). Subject weight, and compliance with diet were monitored every 3–4 days during the first two phases. Plasma free fatty acids and sphingolipids were measured by mass spectrometry. FADS1 and FADS2 activities were calculated by g‐linolenic (GLA)/linolenic acid (LA) or arachidonic acid (AA)/dihomo‐g‐linolenic acid (DGLA) ratios, respectively.ResultsPlasma zinc levels remained unchanged during Phases I and II in all subjects and increased only at the conclusion of phase III. FADS1 activity significantly (p=0.005) decreased from 5.2 ± 0.5 to 3.6 ± 0.5 (mean ± SEM) following Zn depletion. In contrast, FADS2 activity was insensitive to effects of low zinc intake. As a consequence of FADS1 activity loss, concentrations of AA‐containing phosphatidylcholine (PC), phosphatidylethanolamine (PE), sphingomyelins and plasmalogen antioxidant lipids decreased significantly. Furthermore, plasma concentrations of C16:0 ceramide increased following Zn depletion (p = 0.06) and was inversely correlated with FADS1 activity (p = 0.04; r2 = 0.6). Zn depletion also increased plasma triglyceride (TG) by 26 ± 15% (p< 0.05) and decreased high‐density lipoprotein (HDL) by 18.5 ± 3%; P<0.05. Interestingly, changes in FADS1 activity observed during the depletion was not normalized by dietary Zn repletion (Phase II) but, it was completely normalized with Zn supplementation (20 mg/day; Phase III).ConclusionDietary Zn depletion (6 mg/day) decreases FADS1 activity and lowered AA incorporation into major lipid sub‐classes. Loss in FADS1 activity was inversely correlated with C16:0 ceramide and was further associated with dyslipidemia as evidenced by significant alterations in plasma TG and HDL. These effects were not corrected with dietary Zn repletion (10 mg/day); supplementation with 20 mg Zn/d for an additional 3 weeks was required. These data suggest that FADS1 activity may be a sensitive biomarker of inadequate zinc intake and they also implicate marginal Zn intake as a novel risk factor for dyslipidemia and insulin resistance.Support or Funding InformationHarvest Plus, NIH S10OD0018070‐01
The effect of zinc deficiency and diet restriction on GATA family gene expression in rat's kidney
Authors: Konomi Aki , Yokoi Katsuhiko 
Published: November 2023

We reported that plasma erythropoietin (EPO) concentration, kidney EPO concentration and kidney EPO mRNA level were changed by moderate zinc deficiency or diet restriction. It is not established how moderate zinc deficiency or diet restriction induces these alterations. We propose a hypothesis that moderate zinc deficiency or diet restriction affects renal GATA family genes expression. Thirty 3‐week‐old male Sprague‐Dawley rats were assigned into 3 dietary groups (n=10 each) and fed on dietary regimens for 4 weeks: control group (AIN‐93G; Control), moderate zinc‐deficient group (4.5 mg Zn/kg; zinc deficiency) and pair‐fed group (AIN‐93G; diet restriction). Kidney was separated into cortex and medulla for RNA isolation. Quantitative real time‐PCR using TaqMan proves was performed for mRNA of GATA family gene (GATA1, GATA2, GATA3, GATA4, GATA5, GATA6, Gatad1, Gatad2a and Gatad2b). Statistical analysis was performed by ANOVA and the multiple comparison test of Tukey‐Kramer. P value less than 0.05 was considered significant. Kidney medulla mRNA expression per tissue weight of GATA4 and GATA5 were significantly decreased by zinc deficiency compared with Control. GATA2, GATA3, Gatad1, Gatad2a and Gatad2b were significantly decreased by zinc deficiency and diet restriction compared with Control. Kidney cortex mRNA expression per tissue weight were not significantly different among the groups for all genes. Cortex and medulla GATA1 and GATA6 mRNA expression per tissue weight were not changed. These results suggest that kidney GATA family gene mRNA expression were decreased by zinc deficiency and diet restriction, especially in medulla.Support or Funding InformationThis work was supported by KAKENHI grant‐in‐aid for Research Activity Start‐up (21800073) and Young Scientists (B) (23700928) and (15K16216).
Wnt/Beta‐catenin and mTOR Signaling in Liver Pathophysiology
Authors: Michael Adeola O Adebayo , Pradhan‐Sundd Tirthadipa , Russell Jacquelyn , Monga Satdarshan (Paul) S. 
Published: November 2023

Two distinct signaling pathways have been implicated in the pathogenesis of hepatocellular carcinoma, the mTOR and Wnt/Beta catenin pathways. At present, there is ample data on the singular role of mTOR activity and Wnt signaling in early HCC development. However, several of these studies conclude that each pathway alone is not sufficient to account for HCC initiation and development, suggesting potential interactive mechanisms between pathways. The aim of this study is to investigate signaling mechanisms of cross‐talk between mTOR signaling and Wnt/Beta catenin signaling in hepatocarcinogenesis. Previous studies in the lab have shown modulation of mTOR and target proteins in point‐mutated beta‐catenin+MET induced tumors. Based on these, we hypothesized that Wnt/Beta‐catenin and mTOR pathways may cooperate and cross‐talk during hepatic pathophysiology. We utilized beta‐catenin‐floxed and LRP5‐6 double floxed mice to interbreed to the Albumin‐Cre transgenic mice to conditionally delete these genes from hepatocytes to examine mTOR. We also used human HCC Hep3B cells either overexpressing or with silenced beta‐catenin. In vivo and in vitro studies show that beta catenin regulates mTOR in the liver. Total mTOR levels remain unaltered, but mTOR phosphorylation along with its downstream targets such as ribosomal protein S6 and 4E‐BP1 are also affected. In conclusion, our studies provide evidence of the interactions between mTOR signaling and Wnt/Beta catenin signaling pathways in both liver homeostasis and in HCC cells. These models will provide a novel opportunity to understand the role and regulation of such interactions and crosstalk.Support or Funding InformationPost‐doctoral Trainee award T32CA186783NIDDK 4R01DK100287‐04NIDDK 5R01DK062277‐13