dataesr/scientific-paragraphs-categorization huggingface dataset:
Food Consumption Analysis Food intakes of children and women included in this study were extracted from the food consumption data at the household level and the individual consumption of children aged less than 5 years old measured during the 2009 Food Consumption Survey, using the 24-hour recall method combined with controlled food weighing  [30]. To estimate the individual consumption of women of reproductive age (19-50 years old), the energy requirement of an 18-60 year male was set as the reference (weight = 1) and used to weigh the specific energy requirements of other age and gender groups. For women 15 to 17 years of age, a weight of 1. 1 was assigned, 1. 4 for a pregnant or lactating woman and a weight of 0. 9 for women aged 18-60 years old was used during the analyses of the 2000 and 2009 food consumption survey  [31].
Data and Statistical Analysis For biological and anthropometric status, data entry, including quality checks was performed with Excel 2007 TM. For consumption, the data was performed using SPSS software version 19 TM. Data management, including quality checks, and analysis was performed with SAS software version 9. 2 TM (SAS, V9. 2; SAS institute, Cary, NC)  [35]. Anthropometric data were entered through EpiData (version 6. 0, CDC). All analyses took into account characteristics of the cluster sampling design using the appropriate survey procedures of SAS (survey freq and survey means procedures as introduced in SAS Institute Inc. 2008)  [35]. Qualitative variables were expressed as percentages and standard error percentage. Continuous variables were expressed as arithmetic means and standard error of the mean except plasma vitamin D concentration, vitamin D and calcium intakes whose data were not normally distributed.
Limitation of the Study The population analyzed for this article is a subsample of the National Food Consumption Survey sample that was also used for the Nationwide Vitamin and Mineral Deficiencies Study. However, this subsample has not biased the results, as similar findings on dietary calcium intake were found among the 7,980 households surveyed  [53]. Vitamin D deficiency might be season-dependent as shown in Europe  [54] , and the study might not be representative of the vitamin D status during the entire year. In Europe, the 25(OH)D levels were 40% higher in summer than in winter, a reflection of the differences in solar exposition. However, the current study was conducted in summer time (July -September) that is the sunniest period of the year especially in the North Vietnam making it unlikely that the current data overestimates the prevalence of vitamin D deficiency.
Statistical analysis The FA composition and mRNA abundances of each tissue, as well as the plasma metabolites and hormones, were analysed by one-way ANOVA using the MIXED procedure of the Statistical Analysis System software package version 9. 1 (SAS Institute Inc. , Cary, NC, USA). The statistical model included the fixed effect of the treatment. For data on mRNA abundance and major FA in the four tissues, a separate one-way ANOVA was performed including the fixed effects of the tissue in the statistical model. In both cases, the random effect of animal was used as the error term to contrast the effect of either dietary treatment or tissue site. The differences between means were evaluated using the 'pdiff option' of the 'LS means' statement of the MIXED procedure. Pearson correlation coefficients (r) were generated for associations between the tissue mRNA abundance and the concentration of specific FA in the tissues using the CORR procedure of SAS.
Acknowledgements The present study was supported by the  Agence Nationale de la Recherche  ( ANR, France ;  ANR-05-PNRA-No. 5. E. 24 ). P. G. Toral was granted a fellowship from the  Fundacion Alfonso Martı ´n Escudero (Madrid, Spain). The authors gratefully acknowledge the staff of  UniteExperimentale des Ruminants de Theix (INRA, France)  for the diligent care of the experimental animals and assistance in slaughtering, and  J. Rouel ,  D. Bany ,  S. Bes ,  E. Tixier  and  M. Tourret  from  UMR 1213 Herbivores (INRA, France ) for assistance in the management of the animals, sampling and laboratory analysis.
Fe uptake and incorporation into ferritin BMM uninfected or infected with M. avium for 24h were incubated with 2. 5 µM ( 55 Fe) ferric ammonium citrate (Perkin Elmer) for 6h, at 37 °C in a 7% CO 2 atmosphere. Afterwards, cells were washed with cold saline and lysed with 25 mM Tris-HCl buffer pH 7. 4 (Sigma), containing 0. 5% Triton X-100 (Sigma) and 1% proteases inhibitor cocktail P840 (Sigma). Total protein content was measured on soluble homogenates using Bio-Rad DC TM Protein Assay (Bio-Rad) and 18 µg were loaded on a native PAGE (7. 5% acrylamide, 1. 5 mm thick). The gel was dried, subjected to phosphor imaging (Typhoon 8600; Molecular Diagnostics, Amersham Biosciences), and analyzed using the ImageQuant program version 5. 1.
Gene expression Total RNA was extracted using the Micro-to-Midi Total RNA Purification System (Invitrogen) according to the manufacturer's specifications. 2 µg of total RNA was transcribed into cDNA, by a Moloney Murine Leukemia Virus Reverse Transcriptase (Fermentas), using an oligo(dT) 18 primer. The primers used for amplification of cDNA were as follows: Hprt1 (housekeeping) 5'-gtaatgatcagtcaacgggggac -3' (forward) and 5'-ccagcaagcttgcaaccttaacca-3' (reverse); Fth1 5'-ggagttgtatgcctcctac -3' (forward) and 5'-gagatattctgccatgcc -3' (reverse). The primers were shown not to co-amplify genomic DNA. All reactions were performed in a total reaction volume of 20 µL with iQ™ SYBR ® Green Supermix (Bio-Rad) and carried out in the iQ™5 instrument (Bio-Rad). Baseline thresholds were calculated by Bio-Rad iQ5 program and the threshold cycles (Ct) were used in the REST software  [14] , where CT values for target gene were normalized to expression levels of Hprt1.
Acknowledgments We would like to thank everybody who took part in the project, including  Christine Archimbaud  ( Faculty of Pharmacy, Clermont-Ferrand, France ) for help in performing the microbiological experiments and photography, and  Stephanie Tubert  ( Faculty of Dentistry, Clermont-Ferrand, France ) for reading of the note on oral hygiene. Special thanks go to the working group for pertinent discussions during the design and validation of the resource.
Quantitative real time PCR To determine the transcription activity of both transgenic variants of SLC41A1, namely wt and c. 1049C. T, by the quantitative real time PCR (q-RT-PCR) method in induced (+tet) and non-induced (-tet) cells, we used the following primers: hSLC41A1fw, 59-TTGGACGCTCGCCTTGCCTG-39 and hSLC41A1rev, 59-TGGTGTGGAACACCTGCGCC-39. Expression activities of SLC41A2 and SLC41A3 were determined with following primer pairs: hSLC41A2fw, 59-TGGTTATAAGTAG-CATTGGGGGCCT-39 and hSLC41A2rev, 5 9-TCCTGCTAGCCTGAATGGCCA -39; hSLC41A3fw, 59-CA-CAAAGATAGTCGGTATCTGACG-39and hSLC41A3rev, 59-GACCATGGCCAGGATGATT-39. Total RNA isolation, the determination of its integrity, purity, and quantity, cDNA synthesis, and q-RT-PCR were performed as described by Kolisek and colleagues  [20]. Data were evaluated with software application FK-Wolf-01, developed by Katharina Wolf (FU Berlin).
Statistical evaluation was performed with data sets acquired from three biological preparations for each condition loaded in triplicate. Protein detection in tet-inducible HEK293 cell lines HEK293 cells, over-expressing flag-or HA-strep-tagged wt or p. A350V, and the respective uninduced controls were lysed with RIPA buffer for 30 min. Centrifugation was performed to pellet unsolubilized material (14000 rpm, 30 min, 4uC). The total protein concentration was determined with the Bradford protein assay (Biorad). For the flag-tagged wt variant, samples containing 10 mg total protein and, for the HA-strep-tagged wild-type variant and the p. A350V mutant, 30 mg total protein were run on a 10% SDS-polyacrylamide gel and transferred to a polyvinylidene difluoride (PVDF) membrane. Immunoblotting was performed in TBS-TWEEN plus 2.
5% dry milk with antibodies against the flagtag (HRP-conjugated anti-flag M2, Sigma-Aldrich) and the streptag (Qiagen). Anti-mouse IgG linked to horseradish peroxidase (HRP; Cell Signaling Technology) was used as the secondary antibody for the anti-strep antibody. The antibody against RPL19 (Abnova), together with the anti-mouse secondary antibody stated above, was used to detect the loading control. Proteins were visualized by use of the SuperSignal TM West Dura system (Pierce). Image J software (http://rsb. info. nih. gov/ij/) was used for the densitometric analyses.
[Mg 2+ ] i values were calculated from the 340 to 380 nm ratio according to the formula of Grynkiewicz et al. [35]  and as described in Kolisek et al. [20, 27]. SLC41A1-dependent Mg 2+ extrusion from induced SLC41A1 (wt) and p. A350V cells was determined from the [Mg 2+ ] i changes observed in Mg 2+ -loaded cells during recovery in CMF-HBS+ solution and calculated by subtracting the respective values of uninduced cells. If not otherwise stated, data are presented as means 6 SE. All statistical calculations were performed with Sigma-Stat (Jandel Scientific). Significance was determined by Student's t-test or Mann-Whitney rank sum test as appropriate; P#0. 05 was considered to be significant.
Acknowledgments Our gratitude is due to Heike Pro ¨hl ( FBN Dummerstorf ),  Martin Marak , and  Katarina Wolf  (both  FU Berlin ) for competent technical support of the project, to  Dr. Mandana Rezwan  ( Dualsystems Biotech AG ) for cooperation with the construction of the cell lines and to  Dr. Theresa Jones  for linguistic corrections.
Protein content measurement and two-dimensional electrophoresis analysis Protein concentration was measured in duplicates using the Bradford assay where bovine serum albumin (BSA) was used as a standard. 2D electrophoresis was performed as in  Dsamou et al. (2012)  with the following minor modifications. In the first dimension, the protein load per strip was adjusted to 400 lg of proteins. The gels were stained using Sypro Ruby (Bio-Rad), and the images were acquired on a Pharos FX imaging system (Bio-Rad). The image analysis was done using Samespots software (NonLinear Dynamics). For each gel, spot volumes were normalized by the total volume of all spots. Values are expressed in ppm.
1 H-NMR analysis Samples were first thawed and 100 lL aliquots were added to 550 lL of deuterium oxide (D 2 O) containing 0. 25 mM sodium trimethylsilyl-[2,2,3,3-2 H 4 ]-1-propionate (TSP; as a chemical shift reference at 0 ppm). The solutions were centrifuged at 5,0009g for 10 min before being placed in 5 mm NMR tubes. All 1 H NMR spectra were obtained using a Bruker DRX-600 Avance NMR spectrometer operating at 600. 13 MHz for a 1 H resonance frequency and an inverse detection 5 mm 1 H-13 C-15 N cryoprobe attached to a cryoplatform. All 1 H NMR spectra were acquired at 27 °C using the Carr-Purcell-Meiboom-Gill (CPMG) spin-echo pulse sequence with presaturation, as described previously  (Neyraud et al. 2013 ) using 512 transients. All NMR spectra were then data reduced using AMIX (version 3.
0. Kruskal-Wallis test was finally used to determine proteins and metabolites that differed significantly between the two groups considering p \ 0. 05 as the level for significance. Multivariate analysis was performed using SIMCA-P software (V13, Umetrics AB, Umea, Sweden). The overexpression of CA6 and cystatin SN, measured by ELISA, in sensitive? subjects was tested using onetailed Students' t test. This test was done using the Statistica software (StatSoft X). The level of significance was set at 5 %.
1 % trifluoroacetic acid (TFA), 13 lL of ACN for 8 min and 3 lL of ACN. The resulting peptide extract was concentrated by evaporation. For the analysis in MS and MS/MS mode, 1 lL of matrix (3. 5 mg/mL a-cyano-4-hydroxycinnamic acid (HCCA)-ref 201344, Bruker Daltonics-in a solution of 50 % ACN/0. 2 % TFA) and 0. 5 lL of the digest were deposited on a ground steel target plate. Analysis was conducted using a MALDI-TOF/TOF UltrafleXtreme Bruker Daltonics in automatic mode. The database search was performed on SwissProt restricted to human entries. Two missed cleavages were allowed. Carbamidomethyl modification of cysteine was accepted as a stable modification and methionine oxidation as a variable modification. Mass deviation tolerance was set at 30 ppm in MS mode and 0. 6 Da in MS/MS mode. Systematic reinterrogation on the list of unmatched peaks was performed until no new protein was identified.
3% BSA and 0. 03% Triton X-100 PBS, and incubated 18 h at 4 1C. After 3 rinses (0. 01% Tween-20 PBS), cells were incubated 1 h at RT with the secondary antibody Alexa 488-conjugated goat anti rabbit (1:1000, Invitrogen) and anti GFAP antibody (Cy3 ™ conjugated, Sigma-Aldrich). After rinsing, cells were stained for 10 min with DAPI (1:10,000) rinsed again in PBS, then in deionised water and mounted with Fluoromount-G. Acquisitions were collected using the same exposition settings. When the primary antibody was omitted, no signal was detected. After correcting for background intensity, images were processed with Metamorph software (Version 717) by measuring the fluorescence intensities of the cells (ObRb) or the nuclei (P-STAT3 and 5) as determined by the DAPI staining. 2. 7. Immunohistochemistry GFAP-CB 1 -WT and GFAP-CB1-KO mice were deeply anesthetized (Nembutal 50 mg/kg, i.
Statistical analysis All the results are expressed as mean 7 SEM of indicated replicated number of experiments. Statistical analysis and dose-response curves were generated using GraphPad Prism 5. 03 (GraphPad Software Inc. , San Diego, CA, USA). χ 2 test was used to see differences in the percentage of CB 1 receptor expressing astrocytes in the different WT and mutant mice. Student's t-tests were used for the comparison between two groups. When comparing more than two groups, one-way ANOVA followed by Dunnet's or Tukey's post-hoc test (as appropriate for comparisons relative to control or between all groups) was used. Twoway ANOVA followed by Bonferroni's post-hoc test was used to compare results from experiments with two independent variables. p values lower than 0. 05 denote statistical significance.
ACKNOWLEDGMENTS The authors declare no competing financial interests. GM and DC are supported by  INSERM , BB is a post-doctoral fellow from the  National Fund for Scientific Research  (F. N. R. S. ) and was supported by  FNRS  grants  FNRS 1. B144. 10. Thanks to the  Region Aquitaine/INSERM  (GM and DC support and PhD fellowship to MM-L and PC), the  Fondation pour la Recherche Médicale  (GM and MM-L), and the  EMBO post-doctoral Fellowship (LB). LR is in receipt of a post-doctoral Specialization Contract from the  University of the Basque Country UPV/EHU. Supported by  EU-FP7  (REPROBESITY,  HEALTH-F2-2008-223713 , GM),  European Research Council  (ENDOFOOD, ERC-2010-StG-260515, GM),  Fyssen Foundation (ES-G) ,  RTA, I. S. Carlos III  ( RD12/0028/0004 , PG, MJC),  Basque Country Government  ( BCG IT764-13 , PG),  University of the Basque Country   UFI11/41  (PG),  MINECO   BFU2012-33334  (PG).
Statistical Methods. We used the raw SAS Version 9. 1 or higher soware to compute the results. All statistical tests were performed two sided, and a  value less than 5% was considered as statistically signi�cant. Mean changes from baseline to week 12 was calculated with ANOVA. e percentages of subjects reaching 25-hydroxyvitamin D serum concentrations ≥20 ng/mL and ≥30 ng/mL at the end of the study were analysed by means of a chi-square test comparing D-CURE and placebo. For the main continuous baseline characteristics age, and BMI, an ANOVA was performed, and for the binomial baseline variable sex, we used a Cochran-Mantel-Haenszel Test. All safety data obtained in this study was tabulated with descriptive statistics. Comparisons between treatment groups were based on descriptive statistics.
Data Collection Plasma sodium, potassium, Ca, P, total carbon dioxide, urea and creatinine were determined daily for 7 The first blood sample was obtained at around 12 hours of life. Daily diuresis was calculated and a consecutive 8-hour urine collection, starting 4 hours before the blood sampling, was performed using a plastic bag. Urine was analysed for sodium, potassium, Ca, urea and creatinine concentrations. Urine escaping the bag was quantified by weighing nappies. Fluid, energy and nutrient intakes were recorded daily. Oral human milk intakes were considered in fluid and nutrient intake calculations. Plasma and urine electrolytes were measured by an Ortho Clinical Diagnostic analyser (Rochester, NY, USA), which uses direct potentiometry. Ca balance was expressed in mg/kg/day and calculated as the difference between daily intake and daily urinary excretion.
Validation of the microarray expression data by RT-qPCR analysis RT-qPCR was used to validate the expression trends of selected genes identified as being differentially expressed by microarray analysis. Using the same total RNA preparations as for the microarray hybridizations, expression of selected genes, representing the previously defined different functional groups, was analyzed by RT-qPCR. Gene BR1035 of unknown function was used as internal reference for normalization, as its expression rate was constant for both strains in the experiments. The expression ratios obtained from RT-qPCR and microarrays were plotted for comparison (Figure  2 , and Additional file 2: Figure  S1 ). We investigated a total of 40 genes representing most of the previously defined functional groups. Using RT-qPCR, thirty one (78%) of the tested genes were found to be differentially expressed between wild-type and rsh-mutant, when applying a cutoff value of log2(x) > 0.
Data acquisition and microarray data analysis Hybridized microarrays were scanned using a GenePix 4000A dual-channel (635 nm and 532 nm) confocal laser scanner (Axon Instruments). The genes represented on the arrays were adjusted for background and normalized to internal controls using image analysis software (GenePixPro 4. 0; Axon Instruments Inc. ). Genes with fluorescent signal values below background were disregarded in all analyses. Statistical analysis was performed by Seralogix, LLC, Austin, TX, employing their computational tools termed the BioSignature Discovery System (BioSignatureDS) to determine significant gene modulations via a Bayesian z-score sliding window threshold technique and fold change. Normalizations against genomic DNA were performed as previously described  [69]. The microarray data were deposited in the Gene Expression Omnibus (GEO) database at the National Center for Biotechnology Information (Accession #: GSE44688), including the ortholog gene references of B.
RT-qPCR analysis Expression of randomly selected genes from most of the different functional categories (n = 19) that we defined based on microarray analysis, and which were differentially expressed between B. suis wild-type and the rsh mutant, was analyzed by quantitative RT-PCR (RT-qPCR). 800 ng from the same RNA samples used for microarray hybridization were reverse-transcribed using a 6-mer random primers mix, as described earlier  [75] , and quantitative RT-PCR experiments were performed using the Light Cycler 480 with SYBR green chemistry to monitor and quantify the amplification rate (Roche). Primers (Sigma Genosys) were designed using Primer 3 Software (Additional file 3: Table  S2 ) to produce an amplicon length between 150 and 250 bp. For each gene tested, the mean calculated threshold cycles (Ct) of the wild-type and the mutant were averaged and normalized to the Ct of a gene with constant expression identified in the transcriptomic analysis.
Acknowledgements The study was supported by funds from the  Centre National de la Recherche Scientifique (CNRS). N. Hanna  was supported by a fellowship from the  Lebanese CNRS. We thank  V. Jubier-Maurin  for critical reading and helpful comments on the manuscript. We acknowledge  P. Bouhours  for the preparation of bacterial media,  E. Hauser  and  M. Korrichi  for helpful technical assistance, and  P. Clair  ( qPCR Platform, UMS Biocampus, University of Montpellier 2 ) for assistance in RNA analysis and RT-qPCR. We are grateful to the  United States Department of Health and Human Services ,  National Institutes of Health (NIH) ,  Western Regional Center of Excellence (WRCE) Pathogen Expression Core  ( Drs. M. McGee ,  R. Friedberg  and  S. A. Johnston ,  Arizona State University, Tempe, AZ ) for developing and printing the B.
8 M) for 24 h at 37 °C. Western Analysis-Western analysis was performed by established methods using whole cell lysates  (14). The primary antibodies used were anti-Fpn1 (MTP11A; 1:500; Alpha Diagnostics International, St. Antonio, TX), anti-Lrp1 (SC-16168; 1:500; Santa Cruz Biotechnology, Santa Cruz, CA), and anti-␤actin (A5441; 1:10,000; Sigma). Secondary antibodies were antirabbit, anti-goat, and anti-mouse (A0545, A5420, and A9917, respectively; 1:10,000; Sigma). Membranes were washed and developed using ECLϩ Western blot detection reagent (Amersham Biosciences) and exposed to x-ray film. Densitometry was performed using Quantity One software (Bio-Rad). All data were normalized to the loading control, ␤-actin. Chemiluminescent immunodetection by x-ray film demonstrated a linear signal output within 5% error over the protein loading employed and the exposure times assessed.
For experiments examining the lability of hepcidin binding at various pH values, ␣ 2 M/␣ 2 M-MA (or these proteins treated with IAA as described above) were dissolved and incubated in either of the following buffers in 0. 14 M NaCl for 30 min at 37 °C: pH 5 (sodium acetate/acetic acid buffer); pH 6 (Na 2 HPO 4 /NaH 2 PO 4 buffer); pH 7-9 (Tris buffer); and pH 10 (sodium carbonate/sodium bicarbonate buffer). BioHepcidin was then added to the ␣ 2 M/␣ 2 M-MA protein, as described above. Following treatment, samples were concentrated using a vacuum concentrator (Thermo Scientific) and applied dropwise to a nitrocellulose membrane before biotin-streptavidin-HRP detection via chemiluminescence (Chemiluminescence Detection Module, Thermo Scientific). Preliminary studies showed that the biotin label did not interfere with hepcidin binding to ␣ 2 M or ␣ 2 M-MA or its ability to decrease Fpn1 expression in J774 cells (data not shown).
9% saline (vehicle) or this vehicle containing ␣ 2 M, ␣ 2 M-MA, ␣ 2 M⅐hepcidin, or ␣ 2 M-MA⅐hepcidin (proteins, 3. 6 nmol; hepcidin, 0. 9 nmol; complexed at a 4:1 ratio) was injected into the tail vein. After 2 h, blood (600 l) was collected by cardiac puncture from anesthetized mice and placed in Capiject Micro Collection tubes, and serum was isolated. Serum iron was measured using a Konelab Clinical Chemistry Analyzer (Thermo Scientific, Waltham, MA). Statistical Analysis-Results were expressed as mean Ϯ S. D. or S. E. Data were compared using Student's t test. Results were considered significant when p Ͻ 0. 05.
Acknowledgments-We thank  Dr. H. Lok  for outstanding technical assistance in aiding the completion of the mouse experiments. We acknowledge the assistance of the following people for their careful examination and comments on the manuscript before submission:  Dr. Katie Dixon ,  Dr. Zaklina Kovacevic ,  Dr. Darius Lane ,  Angelica Merlot ,  Dr. Vera Richardson ,  Dr. Yu Yu , and  Dr. Daohai Zhang  of the  Molecular Pharmacology and Pathology Program, Dept. of Pathology, University of Sydney. We greatly appreciate the insightful comments of  Dr. Jiri Petrak  ( Dept. of Pathophysiology, Charles University, Prague ) during the preparation of this manuscript.