dataesr/scientific-paragraphs-categorization huggingface dataset:
Need transparency and better documentation of the decision making process Develop new test methods for step 1 (hazard identification) of risk assessment: assays that reflect endocrine endpoints and critical windows Enhance available assays: endocrine endpoints, critical windows (extended 1 generation study may be a good first step) Address data gaps in our knowledge of exposures at vulnerable periods (fetal) Take into account allometry in administered doses (where appropriate) to choose ''low doses'' Integrate existing classification system with recognized endocrine endpoints Establish an agreed upon definition of ''low dose'' or ask all to be precise in their language Develop new test methods Recognize that processes need to proceed in parallel, and information needs to be continually integrated Studies that show an effect at low dose which disappears at high dose should not be dismissed Continue to use a tiered approach to exclude chemicals that are unlikely to cause endocrine disruption Use of QSAR and in vitro multiple dose screening to have indications for further testing Develop or elaborate on guidance which allows evaluation of exploratory research, to make it more useful to Risk Assessment Use of assays developed by exploratory research in risk assessment: aid risk assessors to understand and interpret them Consider using test methods with consistent results even if not formally validated a Points taken from breakout group reports, some wording changes for clarity and to avoid overlap.
A workshop to develop a consensus on the definition of low dose and a consensus on the significance of the 'low dose' issue followed by guidance on how and when to include 'low doses' in the design of toxicology studies, considering the need for not only adding more doses to testing protocols but to start dose selection at ''low doses'' rather than from high doses downward. A knowledge base of findings in vivo and in vitro showing NMDR and low dose effects in a secure database as a resource, which may include raw data, was also proposed.
Acknowledgements The Pew Charitable Trusts supported the workshop facilitation. The U. S. -based non-profit organization RESOLVE provided the facilitators with two from the  US  and three from Europe. Meeting organizers specifically thank  Gilbert Schoenfelder 's team at  Charite Universitatmedizin  for the logistical and local arrangements and the  Oak Foundation ,  Forsythia Foundation ,  NIEHS ,  European Commission  and  UBA  for their financial support. This article may be the work of an employee of the  National Institute of Environmental Health Sciences (NIEHS) ,  NIH : however the statements,opinions or conclusions contained therein do not necessarily represent the statements, opinions or conclusions of the  NIEHS ,  NIH  or the  U. S. Government. ** Planning committee members:  Jerrold Heindel ,  Sharon Munn ,  Jason Aungst ,  Claire Beausoleil ,  John Bucher ,  Anna Federica Castoldi ,  Alexandre Feigenbaum ,  Andreas Gies ,  Ulla Hass ,  Marie Louise Holmer ,  Dennis Keefe ,  John Peterson Myers ,  Pia Juul Nielsen ,  Gregory Noonan ,  Jean-Nicolas Ormsby ,  Bernd Schaefer ,  Gilbert Schoenfelder ,  Thaddeus Schug ,  Wolfgang Straff ,  Laura Vandenberg  and  R.
0-4. 4 ppm for the glycerol backbone and the peak at 0. 9 ppm for the methyl group as an internal standard, and using the peak intensity ratio between the peak at 2. 8-3. 2 and 4. 0-4. 4 ppm  [56]. This number was used to represent the sample code of the ESO. The degree of epoxidation (DOE) or percentage of conversion from double bonds to epoxide groups was determined from Equation (1) and the number of starting double bonds of the soybean oil used in the present study was 4. 60: (1) We assumed that DOE (Degree of epoxidation) of Vikoflex ® 7170 should be very high, i. e. ! 100%. However, there was no data on the starting soybean oil; therefore, it was not possible to determine its DOE in the present study. ESOs containing 50 and 75% DOE were prepared, and Vikoflex ® 7170 was a representative of 100% DOE.
We identify the producing locations from national and international data on feed production and trade. The N food-print is then calculated as total nitrogen losses from entire livestock systems using data on total N fertilization, atmospheric deposition and manure management per crop and livestock farm. The N food-print is expressed per capita (kg N cap -1 yr -1 ) and per unit of agricultural land (kg N ha -1 ) in order to make results comparable with other products, consumers and production systems. Emissions of Nr in consequence of feed, milk and carcass transportation and processing are not accounted for. Nr is hereafter referred to as N.
Meat and fresh milk consumption We used data on national average apparent consumption. Apparent consumption (also known as food availability) is calculated from national food supply balances as domestic production, plus imports, minus exports and it includes, therefore, retail and household food waste. Apparent food consumption (hereafter referred to as consumption) links resource use in agriculture with total food production. In France, consumption of milk, pork and beef is respectively 52 L cap yr -1 , 34 kg cap -1 yr -1 , 26 kg cap -1 yr -1  (Agreste, 2006). Table  1  shows the chemical composition of milk  (CNVA, 2006 ), beef (NRC, 2000; Wulf, 1999; Hoch and Agabriel, 2004)  and pork  (Lange et al. , 2003; NRC, 1998)  and the annual per capita consumption of each product in terms of nitrogen (kg N cap yr -1 ) and gross energy (Mcal cap -1 yr -1 -energy in lipids, proteins and lactose is 9.
Roughages (including annual fodder) are typically produced on livestock farms because they are bulky and therefore difficult to transport. The type of roughages used in cattle farming is derived from agricultural statistics for each region (Statistique agricole annuelle, 2006, Ministry of Agriculture, www. agreste. agriculture. gouv. fr). In general, natural and semi-natural grasslands and meadows are dominant in regions specialized in beef meat production while maizeforage is dominant in regions specialized in dairy production  (Agreste, 2008c). As opposed to roughages, concentrate feeds are often purchased by the livestock farms. The ingredients used in the production of concentrate feeds are derived from datasets of agro-industries per livestock sector. Major ingredients are cereal grains, cereal by-products and soy and rape meals  (Agreste, 2008a).
N budgets of livestock systems and the N food-print of products Figure  3  gives a generic representation of a livestock system and of input and output N flows among its segments. The N inputs per livestock system are computed per region and feed crop from statistical data except for soybean. Data on soybean fertilisation in the countries exporting to France were not available. N inputs in soybean agrosystems are estimated indirectly, assuming global average NUE of 50 %  (Cassman et al. , 2002). Agricultural yields for soybean are taken from  FAO (2004)  and average (in terms of nitrogen content) 125 kg N ha -1 in Brazil, 120 kg N ha -1 in Argentina, 155 kg N ha -1 in the USA and 120 kg N ha -1 on global average. For crops other than soybean, we calculate N inputs and nitrogen use efficiencies (NUE) from specific data on total fertilization and crop yields (expressed in nitrogen) per region and crop  (Agreste, 2006).
All data are for 2006. Data on chemical fertilizer application per region and crop are taken from  Agreste (2008b). For atmospheric deposition we used simulation data from EMEP (2006). N deposition rates for France vary between 3 to 6 kg N ha -1 /y. For BNF, we used commonly accepted values from literature: 250 kg N ha -1 yr -1 for alfalfa, 35 kg N ha -1 yr -1 for pasture (assuming 15 % legumes) and 5 kg N ha -1 yr -1 for fallow  (Smil, 1999 , Peoples et al. , 1995). In crop rotations, green fertilizers are mainly used in association with maize in half-year rotations. We assume an annual BNF rate of 125 kg N ha -1 in these rotations. For manure application, we assume a uniform rate of 170 kg N ha -1 , which is the prevailing upper limit for manure application in the European Commission Nitrate Directive, 1991/676/CEE.
Certainly, not all crop farmers fertilize at the allowance rate, but, lacking precise data obliged us to adopt this simplification. We note that on livestock farms, manure N is an internal N flow but additional N is imported as feed (cf. Fig. 3 ). N inputs in feed are calculated in the previous step. N inputs to farming systems are calculated per livestock product and sum up to the gross N food-print of that product. N leaves the farming systems in the form of manure towards crop agriculture, in the form of live animals and milk, in the form of N 2 through denitrification, and in oxidized and reduced forms through volatilization and leaching, which directly enter the N cascade. N output in the form of animal biomass sums the N in fresh milk, pork and beef (cf. Table  1 ) plus the N in slaughter waste for beef and swine.
, 2009). Chronic ghrelin treatment also modifies the expression of dopaminergic receptors in Acc, more specially D1 and D3  (Skibicka et al. , 2012) , which are described to be involved in obesity, food reward (D1 receptors) and inhibition of reward behavior (D3 receptors). Similar data were obtained from human imaging studies that emphasize the role of ghrelin in food reward. Indeed, intravenous ghrelin injection to human subjects increases activity in brain areas involved in the evaluation of the reward value attributed to food and food cues including the striatum, amygdala, insula, and orbitofrontal cortex  (Malik et al. , 2008).
Acknowledgements This study was supported by grants from the  Conseil Général de l'Eure , the  Communauté d'Agglomération d'Evreux and Europe  ( FEDER n°31970 ). LMSM is member of the  PFT Normandie Sécurité Sanitaire, of the Institute for Research and Innovation in Biotechnologies (IRIB)  and of the first perfumery-cosmetic worldwide network,  Cosmetic Valley. AD was a recipient from the  French Ministry of Research (MRE). The authors are grateful to  Christine Farmer  for linguistic support.
A Loss of Genetic Diversity in Western European Cattle Polled In an independent attempt to fine map the Polled locus, we genotyped a panel of 51 animals (15 homozygous polled and nine paternal halfsib designs with a heterozygous founder) from nine different breeds, using the Illumina bovine 777K SNP beadchip. After phasing, we identified a large segment of 79 markers located at the beginning of BTA01 that was shared by all polled animals (ranging from position 1,693,164 to 2,018,403, according to the UMD3. 1 bovine genome assembly  [27] ). To test the significance of this result, we then calculated the frequency of the same haplotype in 17 Western European horned breeds, based on analogous genotyping data from 4,843 animals. Surprisingly, the frequency of the ''polled'' haplotype was extremely high in all horned breeds (ranging from 44.
Accurate Mapping of the Polled Charolais, Based on Observed Recombination Events To avoid any bias in mapping the Polled locus, we decided to investigate the French genomic selection dataset for recombination events. This dataset comprised haplotyping data from 3,349 Charolais and 76,851 Holstein animals (out of which 535 and 1,493, respectively, were born to at least one polled parent) that were genotyped with the Illumina BovineSNP50 beadchip. Analyzing BTA01 haplotypes from five Charolais and eight Holstein polled founders, we identified four different large haplotypes (two in each breed) that were never found among 189 and 1,233 horned founders, respectively. These consisted of 160, 191, 59 and 410 consecutive marker blocks (7. 5, 9. 8, 2. 5 and 24. 5 MB) encompassing previously reported locations of the Polled locus (not shown)  [16].
Whole Genome Sequencing Data Confirm Allelic To exhaustively identify candidate causative mutations, we then sequenced the complete genome of two animals that were homozygous for the most frequent polled haplotype in their respective breeds (i. e. , Cha-PH1 and Hol-PH1) using 100-bp paired-end reads. The average sequence depth of non-repetitive sequences within the corresponding polled intervals was 15. 3x and 25. 2x for Charolais and Holstein homozygous polled animals, respectively. From these data, we identified 1,610 and 1,225 putative DNA sequence variants in each polled interval, based on a comparison with the UMD3. 1 genome assembly (which was obtained by sequencing a horned Hereford cow  [28] ). Filtering for variants found among analogous sequencing data from 55 horned bulls (one Charolais, eight Montbeliarde, nine Normande and 37 Holstein), we reduced the number of candidate causative mutations to 12 for Cha-PH1 and 47 for Hol-PH1 (Table  S1 ).
EMT No Longer Plays a Role in Horn Ontogenesis at 90 dpc Meanwhile, we investigated the role of other functional candidate genes in the same dataset to obtain better insights into the mechanism involved in normal and pathological horn bud differentiation. These comprised FOXL2, RXFP2, TWIST1 and ZEB2 (gene mapping to loci involved in horn defect syndromes in Bovidae), TWIST2 and FOXC2 (involved in distichiasis in model species), and E-Cadherin, N-Cadherin, Occludin and Vimentin (four markers of EMT). With the exception of FOXC2 (Fig. 5 ), none of the master regulators of EMT (i. e. , TWIST1, TWIST2 and ZEB2) and none of the markers of this process showed differential expression in the tissues studied at this stage (not shown). From our point of view, these results do not rule out a possible role of EMT in horn ontogenesis per se, but focus instead on an earlier involvement of this process in the differentiation of horn bud precursor cells, as suggested by the key role played by TWIST1 and ZEB2 (whose loss of heterozygosity is associated with T2SS and PMS horn  development defects) in the differentiation of several types of precursor cells from the cranial neural crest  [40] [41] [42] [43] [44].
Blood was collected by veterinarians or by agricultural technicians licensed by the French Departmental Breeding Establishments (Etablissements Departementaux de l'Elevage (EDE)) during routine blood sampling for paternity testing, annual prophylaxis, or genomic selection purpose. Sperm was obtained from semen straws collected by approved commercial artificial insemination stations as part of their regular semen collection process. All the samples and data analyzed in the present study were obtained with the permission of breeders, breeding organizations and research group providers.
IBD Mapping of the Polled Locus Fifty-one animals were genotyped with the Illumina bovine 777K SNP beadchip for this project and 4,843 within the framework of other projects led by the INRA GABI unit (minimum call rate per sample: 0. 95). Markers with call rates lower than 0. 90, deviating from the Hardy-Weinberg equilibrium within breeds, or which were monomorphic over the whole dataset were discarded. Quality control also included parentage checking in the small paternal half-sib design and elimination of remaining Mendelian inconsistencies. A total of 1,046 markers located within the first three megabases of BTA01 were included in the study. Haplotype phasing was then performed within each breed using BEAGLE 3. 3. 2 software  [77] [78]  and setting the scale and shift parameters to 2 and 0.
1, respectively. Finally, haplotypes were analyzed as described in the Results/Discussion section. Accurate Mapping of the Polled Locus using Illumina BovineSNP50 Beadchip Haplotyping Data from the French Genomic Selection Database Illumina BovineSNP50 beadchip haplotyping data were extracted from the French database for Holstein and Charolais genomic selection. Briefly, the French genomic selection procedure  [79]  includes: (i) elimination of markers with minor allele frequency lower than 3% in each breed, deviating from the Hardy-Weinberg equilibrium or without a known position on the UMD3. 1 genome assembly; (ii) parentage checking and elimination of remaining Mendelian inconsistencies; and (iii) phasing and imputation of some missing genotypes with DualPhase, part of the PhaseBook software  [80].
1-3 (GATK)  [83] , whereas the discovery of larger indels was achieved with Pindel  [84]. Finally, detection of Copy Number Variation was performed according to Medugorac et al. [19]. Polymorphisms identified in each animal were then independently filtered for polymorphisms found in whole genome sequencing data from 55 horned bulls (one Charolais, eight Montbeliarde, nine Normande and 37 Holstein), and processed with the same tools within the framework of other projects led by the INRA GABI unit. The average sequence depth of non-repetitive sequences within the polled intervals ranged from 5. 9x to 39. 8x for these horned animals. Genome assembly gaps and regions with null coverage mapping to the restrained Polled intervals were PCR amplified and sequenced using the Sanger method, as described in the following section.
Genotyping of the Candidate Causative Mutations To refine the localization of the recombination point in recombining Friesian haplotypes, eight polymorphisms among the candidates were genotyped based on a dichotomous approach. The seven SNP (g. 1654405G. A, g. 1655463C. T, g. 1684055G. C, g. 1764239T. C, g. 1768587C. A, g. 1855898G. A and g. 2028953C. G) were genotyped by PCR-Sequencing. PCR primers were designed from the UMD3. 1 bovine genome assembly with Primer3 software  [85]. The list of all the primers used in this study is available in Table  S4. PCR reactions were performed using Go-Taq Flexi DNA Polymerase (Promega), according to the manufacturer's instructions, on a Mastercycler pro thermocycler (Eppendorf). The resulting amplicons were purified and bidirectionally sequenced by Eurofins MWG (Germany) using conventional Sanger sequencing.
The other 2,101 animals were genotyped for the Celtic mutation, the Friesian 80-kb duplication, SNP g. 1768587C. A, and two other polymorphisms (g. 1654405G. A and g. 1655463C. T) showing strong linkage disequilibrium with the Friesian allele, as described in Medugorac et al. [19]. Genotypes for candidate SNP g. 1764239T. C and g. 1855898G. A were inferred from genotypes at the other SNP. Of note, in both datasets, no recombination was observed within the Friesian haplotype between the Friesian candidate mutations or mutations showing strong linkage disequilibrium with them. In addition, neither the 80-kb duplication nor the SNP alleles associated with the Polled Friesian allele were found in p/p animals.
Acknowledgments The authors would like to thank the numerous cattle breeders, breeding companies, insemination centers and research groups for generously providing samples, phenotypic information and occasional assistance in this research. In particular, we thank  B. Boiret , the  Humeau  family and  T. Baudouin  ( Charolais breeders ),  B. Theil  ( Limousin breeder ), X. David (UNCEIA),  P. Milon ,  G. Michel ,  B. Raimbaud ,  P. Doradoux J. Thomin ,  T. de Bretagne  and  M. Philippe  ( Evolution, Rennes, France ),  A. Valle  ´e,  M. -T. Blanchon  and  S. Landemaine  ( Genes Diffusion, Douai, France ),  P. Chevalier  ( GENESIA, Lempdes, France ),  J. -M. Cazillac  ( UALC, Naves, France ),  D. Boichon  and  L. Izard  ( MIDATEST, Soual, France ),  J. -M. Champagne  and  J. Negrerie  ( COOP'EVOLIA, Fontaines, France ),  J.
Detection of mRNA by real-time PCR (RT-PCR) Total RNA was extracted from CD36-positive TBC by using TRIzol (Invitrogen) and subjected to DNase treatment using the RNAse-free DNAse Set (QIAGEN). One microgram of total RNA was reverse transcribed with SuperScript II RNAse H reverse transcriptase using oligo(dT) according to the manufacturer's instructions (Invitrogen). RT-PCR was performed on the iCycler iQ real-time detection system, and amplification was undertaken by using SYBR Green I detection. Primers against the genes of interest were designed using the available mRNA sequence information in NCBI Genbank. Basic local alignment search tool (Blast, NCBI, available at http://www. ncbi. nlm. nih. gov/blast. cgi) was used for the verification of primers. Because the genes studied have not yet been sequenced for P.
obesus, we designed primers based on the conserved regions in mRNA of the three wellcharacterized species: Homosapiens, Rattus norvegicus, and Mus musculus  [25]. The sequences of the genes used are as follows: b-Actin (Forward: 59-AGAGGGAAATCGTGCGTGAC-39, Reverse: 59-CAATAGTGATGACCTGGCCGT-39); CD36 (Forward: 59-ACTCTCTCCTCGGATGGCTAGCTG -39, Reverse: 59-CCACAGTTCCGATCGCAGCCC -39); a-gustducin (Forward 59-GTTGGCTGAAATAAT TAAACG-39, Reverse: 59-ATCTCTGGCCACCTACATC-39). The amplification was carried out in a total volume of 25 ml, which contained 12. 5 ml SYBR Green Supermix buffer (50 mM KCl; 20 mM Tris-HCl [pH 8. 4]; 3 mM MgCl 2 ; 0. 2 mM of each dNTP, 0. 63 U iTaq DNA polymerase, and SYBR Green 1. 0 nM fluorescein) and 12. 5 ml (0. 3 mM) of each primer and diluted cDNA. Results were evaluated by iCycler iQ software and relative quantification of mRNA in different groups was determined.
Measurement of Ca 2+ signaling The CD36-positive TBC (2610 6 /ml) were cultured on Will-Codish wells with a glass bottom and loaded with Fura-2/AM (1 mM) for 60 min at 37uC in loading buffer which contained the following: 110 mM NaCl; 5. 4 mM KCl; 25 mM NaHCO 3 ; 0. 8 mM MgCl 2 ; 0. 4 mM KH 2 PO 4 ; 20 mM Hepes-Na; 0. 33 mM Na 2 HPO 4 ; 1. 2 mM CaCl 2 , pH 7. 4. After loading, the cells were washed three times and remained suspended in the identical buffer. The changes in intracellular Ca 2+ (F 340 /F 380 ) were monitored under the Nikon microscope (TiU) by using S-fluor 406 oil immersion objective. The planes were taken at Z intervals of 0. 3 mm, and NIS-Elements software was used to deconvolve the images. The microscope was equipped with EM-CCD (Lucas) camera for real time recording of 16-bit digital images.
The dual excitation fluorescence imaging system was used for studies of individual cells. The changes in [Ca 2+ ]i were expressed as DRatio, which was calculated as the difference between the peak F 340 /F 380 ratios. The data were summarized from the large number of individual cells (20-40 cells in a single run, with 3-9 identical experiments done in at least three cell preparations). For experiments in Ca 2+ -free medium, CaCl 2 was replaced by EGTA (2 mM).
As shown in Fig. 2A  , fat-1 mouse tumor tissues exhibit a marked decrease of NF-B protein expression compared SP2001) for 15 min twice. Then sections were incubated with primary antibody (anti-p-HER3; Cell Signaling, 4791, 1:100 ) for 1 h. The sections were then washed and incubated with secondary antibody linked to biotin (Southern Biotech, 6440-08, 1:500) for 45 min. After three washes, sections were incubated in streptavidin peroxidase (Invitrogen, T20934, 1:100) and revealed in tyramide-AlexaFluor 568 (Invitrogen, T20934, 1:100). Sections were counterstained with DAPI (Sigma, D9542, 2 g/ml) for 10 min and rinsed. Negative controls (primary antibody omission) were included. After processing, the sections were imaged by the Cell Observer station (Zeiss). This station is composed of an inverted motorized microscope, a mercury lamp for the fl uorescence, a CCD camera (Zeiss HRm), a computer, and the AxioVision software which controls the station.
Images were acquired using 10× objective; each image represents about 0. 6 mm 2. About three images were made by level (six images by tumor). Image analysis was carried out using Visilog software. For the p-HER3 surface, images were threshold, binarized, and surface measurement of the binarized images was done. These surfaces were divided by the number of nuclei present in each image. This number was found by determining the mean area of a nucleus.