dataesr/scientific-paragraphs-categorization huggingface dataset:
Kinetic parameters for Ro 31-8220-mediated modulation of drug transporter activities Half maximal inhibitory concentrations (IC 50 ) for Ro 31-8220-mediated inhibition of OCT1, MATE1 and MATE2-K activities and half maximal effective concentration (EC 50 ) for Ro 31-8220-mediated stimulation of OCT2 activity were determined using Prism software (GraphPad Software, La Jolla, CA) through nonlinear regression based on the four-parameter logistic equation, as previously described  [35]. Mode of action for Ro 31-8220-mediated inhibition of OCT1-related TEA uptake was evaluated through graphical analysis of Lineweaver-Burke plots of 1/v (v is defined as the specific uptake velocity of TEA) as a function of 1/[TEA], in the presence and absence of the BIM. Non-linear regression according to the Michaelis-Menten equation was finally performed using Prism software in order to fit v as a function of [TEA] in HEK-OCT2 cells, in the absence or presence of Ro 31-8220, and thus to determine kinetic parameters such as K m (Michaelis-Menten constant) and V max (maximum specific uptake) values.
Molecular descriptor generation Molecular descriptors were evaluated using the Dragon 6 software (Talete, Milano, Italy) that provides 4885 molecular descriptors that are divided into 29 blocks. These blocks include 3D descriptors such as WHIM (Weighted Holistic Invariant Molecular) descriptors, which contain information about the whole 3D molecular structure in terms of size, shape, symmetry and atom distribution  [36]. These 3D-WHIM indices are calculated from x,y,z-coordinates of a 3D structure of the molecule, usually from a spatial conformation of minimum energy, within different weighting schemes in a straightforward manner and represent a very general approach to describe molecules in a unitary conceptual framework. BIMs, initially expressed in SMILES format, were converted to 3D format using the MarvinView software (ChemAxon, Budapest, Hungary) before processing by Dragon 6 to obtain 0D constitutional, 1D structural, 2D topological and 3D geometrical descriptors.
Statistical analysis Experimental data were routinely expressed as means ± SEM. Statistical analysis was performed using the Student's t-test, ANOVA followed by a Dunnett's post-hoc test or F-test using Prism software; the criterion of significance was p < 0. 05. Correlation between molecular descriptor indexes and % of OCT1 activity inhibition by chemicals was initially done with Dragon 6 software through Pearson correlation, using a cut-off for Pearson's correlation coefficient of r > 0. 8 (positive correlation) or r < -0. 8 (negative correlation), as recommended by the software user's manual. P-values for Pearson correlations, as well as linear regressions for molecular descriptors exhibiting a high level of correlation with OCT1 activity inhibition (|r| > 0. 9), were next determined using Prism software after confirmation of normality of data distribution by D'Agostino and Pearson omnibus normality test.
Such data consequently indicate that OCT1 inhibition caused by Ro 31-8220 was not reversible, in contrast to that due to verapamil. Mode of action for Ro 31-8220-mediated inhibition of OCT1 activity was next evaluated through graphical analysis of Lineweaver-Burke plots of 1/v versus 1/[TEA] in the presence or absence of the BIM. As shown in Fig  4 , the plots were consistent with a mixed inhibition of OCT1 activity by Ro 31-8220, i. e. , the intercept of the plots with or without the BIM was out of the x-axis and the y-axis. Finally, we investigated whether Ro 31-8220 can trans-stimulate TEA uptake, which may constitute an argument in favor of the transport of the BIM by OCT1  [34]. As shown in Fig  5 , pre-loading with Ro 31-8220 resulted in trans-inhibition of radiolabeled TEA uptake. By contrast, pre-loading with unlabeled TEA led to a trans-stimulation of radiolabeled TEA uptake (Fig 5 ), as expected for an OCT1 substrate like TEA  [34].
Effect of various BIMs on OCT1 activity To determine whether the inhibitory effect of Ro 31-8220 towards OCT1 activity may be shared by structural analogs, the effects of various BIMs and of the PKC inhibitor LY 333531, which also contains a BIM core (Fig 1 ), on OCT1-mediated transport were studied. In addition to Ro 31-8220, several BIMs such as BIM-I, BIM-II, BIM-III, BIM-VI, BIM-VII, BIM-VIII (also known as Ro 31-7549) and LY 333531 significantly inhibited OCT1-mediated TEA accumulation in HEK-OCT1 cells by more than 50% when compared to control TEA uptake In order to identify the specific physico-chemical properties associated to OCT1 inhibition for BIMs, molecular descriptors, including 0D-constitutional, 1D-structural, 2D-topological and 3D-geometrical descriptors, were determined using Dragon 6 software.
Design and study population The present analysis gathers original and published data collected in adults (≥18 years) by 13 different cross-sectional surveys. The surveys were conducted in Latin America (Mexico  [27] , Brazil and Argentina), Europe (Spain  [28] , France, UK  [29] , Germany, Poland and Turkey) and Asia (Iran  [30] , China  [31] , Indonesia and Japan). Data collection of the individual surveys was performed between 2008 and 2014 by public (Iranian National Nutrition and Food Technology Research Institute, NNFTRI; and Chinese Centre for Disease Control, CDC) and private organisations. The primary objective of these surveys was to assess the intake of drinking water and different types of beverages. A detailed analysis of the volume of TFI (sum of drinking water and beverages of all types) of these 13 surveys can be found elsewhere  [32].
A random recruitment of participants was performed in each country either from a database of volunteers for population surveys, or via systematic door-to-door recruitment until the quotas for age, gender, region, habitat and/or socio-economic characteristics in relation to the total country population were met. Individuals working in company advertising, marketing, market research, the media, the manufacture, distribution and/or sale of water and all kind of beverages were excluded from participation as these individuals might be more aware of their intakes of fluids. Individuals who were not able to read and write in the language of the questionnaire were not eligible to participate in the survey. Having a specific diagnosed disease and/or following a medically prescribed diet were additional exclusion criteria in UK, Iran and China.
Classification of the fluid types The fluids recorded were classified into: water (tap and bottled water); milk and milk derivatives; hot beverages (coffee, tea and other hot beverages); juices; regular sweetened beverages (RSB) (carbonated and non-carbonated soft drinks, energy drinks, sports drinks and other sugared soft drinks); diet beverages (diet carbonated soft drinks, diet non-carbonated soft drinks, other diet soft drinks); alcoholic drinks and other beverages. A more detailed classification can be found in supplementary Table  1  of this paper. TFI was defined as the sum of all these categories. In UK, Poland, Indonesia and Japan, the intake of diet beverages was very small, and therefore, they were during the first data treatment included in the RSB category. In Argentina, Iran and Indonesia, only non-alcoholic beverages were recorded.
Statistical analysis Data are presented either as means and 95 % confidence intervals (CI) for continuous variables, or as numbers and percentages for dichotomous variables. The mean intakes are estimated values of all participants, including nonconsumers. We compared the distribution of the selected characteristics between groups Student's t tests for continuous variables. All statistical tests were two-tailed, and the significance level was set at P < 0. 05. A Bonferroni post hoc test was used to correct for multiple comparisons in the online resources 2 and 3. All analyses were performed using the SPSS software version 22. 0 (SPSS Inc. , Chicago, IL).
Therefore, the energy intake from fluids is likely to be underestimated. For the evaluation of the percentage of energy provided by free sugar, total energy intake had to be estimated since no food data were collected. The lack of food data also limited the interpretation of the data on energy intake from fluids. However, evidence suggested that a fluid-specific record might more accurately estimate fluid intake compared with a food and fluid record  [48]. Since the primary aim of all 13 surveys included was to assess fluid intake, the preference was given to record fluids only. Due to the lack of anthropometric data of the participants in certain countries, the calculations had to be based on population means of energy intake. Nevertheless, this assumption was considered to be acceptable in epidemiological studies since the individual surveys aimed at collecting data from a nationally large sample of individuals and also because the energy intake data for the food balance sheets were recorded during the same year of the fluid surveys.
Acknowledgments Data collected were performed by the  Center of Disease Control in China ; the  National Nutrition and Food Technology Research Institute in Iran. We acknowledge  Christine Jean ,  Market Research  and  Consumer Intelligence Danone Waters  and the entire  MRCI Team  for having raised the methodology of fluid intake surveys to the current standard and for having guided IPSOS (in  Mexico, UK and Turkey ),  TNS (in Argentina, France, Germany ,  Poland and Spain) ,  Nielsen  (in Indonesia), and  GFK  (in  Brazil ) for the preparation of the survey. All the participants give their consent prior the inclusion in the study. The protocol of the unpublished surveys was reviewed and approved by the  Institutional Review Board ,  Office of Research Compliance of the University of Arkansas  (IRB Protocol #  14-12-376 ).
Statistical analysis All data are expressed as the means ± standard error. Non-parametric Mann-Whitney U test was used to compare the differences between two groups. Comparison of more than two independent groups was carried out using Kruskal-Wallis test. Statistical analyses were performed using SPSS 16. 0 (Statistical Package for the Social Sciences, SPSS Inc. , Chicago, IL, USA) software. A p-value less than 0. 05 was considered statistically significant.
miRNA Prediction To search for miRNAs that might regulate mouse POMC expression, we used the following public prediction algorithms and database: Targetscan (http://www. targetscan. org/) and miRanda (http://www. microrna. org/microrna/home. do). Using these different algorithms, we selected the miRNAs that are conserved among the vertebrates (Targetscan) and that have a good mirSVR scores (miRanda).
Surgery and Injections For the intraperitoneal injection (i. p. ), ob/ob, and C57BL/6 mice were injected between 1100 and 1200 h with recombinant murine leptin (Peprotech, France) (n = 5, 5 mg/kg) or saline. Mice were sacrified 4 h after i. p. injection. Intracerebroventricular (i. c. v) cannula placement and injections were performed as described previously  (Girardet et al. , 2011). Animals were anesthetized by an i. p. injection of ketamine (100 mg/kg; Imalgen 1000, Merial, France) and xylazine (6 mg/kg; Rompun, Bayer, France), and placed in a digital stereotaxic apparatus (Model 502600, WPI) coupled to the neurostar software  (Neurostar GmbH, Germany). A 26-gauge stainless steel cannula was implanted into the lateral ventricle at the following coordinates: 0. 3 mm posterior to bregma, 1.
1% Triton X-100 (PBT) for 10 min. Slides were incubated in PBT with 20% horse serum for 1 h for blocking. Rabbit polyclonal antibody anti-fluorescein (1:500, Gen Tex, TX, USA) and mouse monoclonal anti-GFP (1:500, Abcam, MA, USA) antibodies in PBT with 20% horse serum were added to sections and incubated for overnight at 4 • C. Slides were washed three times in PBT for 10 min. Alexa 488 fluor-conjugated goat anti-mouse and Alexa 594 fluor-conjugated donkey anti-rabbit (1:500, Life Technologies, France) with PBS 3% horse serum and 0. 3% Triton X-100 were added sequentially to slides for 1. 5 h at room temperature and sections were washed three times with PBS. Sections were finally coverslipped with mounting medium for fluorescence microscope preparation. Sections were observed using a Zeiss LSM 700 confocal microscope (Zeiss, France) associated to ZEN 2012 software and a DXM 1200 Camera (Nikon, France) coupled to ACT-1 software.
The Expression of Selected miRNAs are Restored in the Hypothalamus of ob/ob Mice after Peripheral Leptin Administration The above data suggest that leptin may be involved in the regulation of the expression of mir-383, mir-384-3p, and mir-488 miRNAs in the hypothalamus. To directly test this hypothesis, ob/ob mice were i. p. injected with leptin at dose of 5 mg/kg and the hypothalamic expression of mir-383, mir-384-3p, and mir-488 miRNAs was evaluated by qRT-PCR 4 h later. We observed an up-regulation of mir-383, mir-384-3p, and mir-488 miRNAs in 12-week-old ob/ob animals treated with vehicle compared to C57BL/6 control mice (+42, +32%; P < 0. 05 and +137%; P < 0. 01, respectively) (Figure  4 ). As expected, leptin treatment significantly reduced hypothalamic expression of mir-383, mir-384-3p, and mir-488 miRNAs in 12-week-old ob/ob animals treated with leptin compared to vehicle-treated ob/ob mice (-25, -21%; P < 0.
In accordance with this hypothesis, we observed in the databases that mir-488 can also target PC1 3 ′ UTR. In accordance with this hypothesis, PC1 gene expression is different in the arcuate nucleus of leptintreated obese ob/ob mice  (Nilaweera et al. , 2003). It is also conceivable that miRNAs may control the biosynthesis of agoutirelated protein; the endogenous antagonist of melanocortin receptors MC3R and MC4R  (Ollmann et al. , 1997). Several miRNAs have been reported to be differentially expressed in the hypothalamus of ob/ob and db/db mice, but their precise role in the regulation of the hypothalamus function remains to be determined. Recently, it has been shown that mir-200a, mir-200b, and mir-429 are up-regulated in the hypothalamus of ob/ob and db/db mice  (Crépin et al. , 2014). The same group showed that daily injection of a pegylated leptin antagonist predisposed rats to obesity, promoted leptin resistance and modified the hypothalamic miRNA expression profile  (Benoit et al.
Acknowledgments This research was supported by funding obtained from  Aix-Marseille University , the " Région Provence-Alpes-Côte d'Azur, " the "Conseil Général des Bouches-du-Rhône " (PACA,  CG13 ) and  Benjamin Delessert foundation. We also thank the  Aix-Marseille University Microscopy Center CP2M  for the access to confocal Microscopy and the  AVB platform (Analyse et Valorisation de la Biodiversité ,  iSm2 ,  UMR 7313 ,  Marseille ). AD is the recipient of a doctoral fellowship from the  Ministry of Education. The authors are grateful to  O. Knowles  and  S. Mounien  for critical reading of the manuscript. We thank  D. Chabbert  for expert help with intracerebroventricular injection and  Dr. M. Maresca  for the gift of antibodies.
1::M-oPrP (1-254). The WT HuPrP(90-231), HuPrP(90-231, H96Y), HuPrP(90-231, P102L), HuPrP(90-231, Q212P) and the WT MoPrP (23-231) were expressed, purified and in vitro refolded according to our previous protocols  30, 32. XAS spectra data collection. Samples with 1:1 Cu(II):WT and mutants HuPrP(90-231) ratio were prepared in 25 mM NaOAc pH 5. 5 and 25 mM MOPS buffer pH 7. 0. The Cu(I):WT and mutants HuPrP(90-231) complexes were generated reducing Cu(II) with 40 mM ascorbate as previously described  30. X-ray absorption spectra were recorded at the European Synchrotron Radiation Facility (ESRF) on beam line BM30B FAME  35  , under ring conditions of 6. 0 GeV and 180 mA. The spectra were collected at the Cu K-edge in fluorescence mode using a solid state 30-element Ge detector, with sample orientation at 45° to incident beam.
The X-ray photon beam was vertically focused by a Ni-Pt mirror, and dynamically sagittally focused in the horizontal size. The monochromator was equipped with a Si(111) double crystal, in which the second crystal was elastically bent to a cylindrical cross section. The energy resolution at the Cu K-edge is 0. 5 eV. The spectra were calibrated by assigning the first inflection point of the Cu foil spectrum to 8981. All the spectra were collected at 10 K. For the Cu(II) samples photo reduction was observed and thus the beam was moved to different spots of the sample at each scan. During collection, data were continuously monitored in order to insure sample homogenity across the multiple spots collected from different sample-holder's cells. Complete data sets were collected for samples 1 to 14 (Supplementary Table  1 ).
For each sample, 12 spectra were recorded with a 7 s/point collection statistic and averaged. The collection time was 25 min for each spectrum. EXAFS data analysis. EXAFS data analysis was performed using the GNXAS code, which is based on the decomposition of the EXAFS signal (defined as the oscillation with respect to the atomic background cross-section normalized to the corresponding K-edge channel cross-section) into a summation over n-body distribution functions γ  (n)  , calculated by means of the multiple-scattering (MS) theory  36. The analysis of the EXAFS spectra was carried out starting from the coordination model previously determined  30. In particular, Cu(II) was found to be coordinated to H96, H111, and two additional low Z ligands (oxygen or nitrogen donors) in the inner shell and to a sulphur-donating ligand, which was assigned to M109.
The glycan modifications on PrP C were assessed using Endo-H and PNGase-F enzymes (New England Biolabs) according to the manufacturer instructions. Samples were loaded onto a 10% SDS-PAGE gel and immunoblotted on Immobilion PVDF (Millipore) membranes. Membranes were blocked with 5% (w/v) non-fat milk protein in TBS-T (0. 05% Tween), incubated with 1:1000 anti-PrP antibody 3F4 (Covance), and developed by enhanced chemiluminescence (GE Healthcare). Band intensity was acquired using the UVI Soft software (UVITEC, Cambridge). Total PrP C expression levels in N2a and ScN2a cell lysates were normalized on β -actin value using 1:10,000 anti-β -actin Peroxidase (Sigma-Aldrich). The PrP Sc PK-resistance levels in all the mutants were derived using as reference the PK-resistant band intensity of the ScN2a cells transfected with WT 3F4-MoPrP.
For PrP C cell surface detection, cells were incubated at 4 °C for 15 min and probed with 3F4 antibody. Cells were permeabilized with 0. 2% Triton-X and stained with AlexaFlour-488 secondary antibody. To detect Thioflavin-S (ThS)-positive aggregates, transfected and non-transfected cells were fixed with 4% PFA/4% sucrose/1% Triton X-100 in PBS. For organelle markers, we used anti-Calnexin (ER marker), anti-EEA1 (early endosome marker), anti-Tfn (recycling endosome marker), anti-M6PR (late endosomes marker) and anti-LAMP2 (lysosomes marker) purchased from Abcam. Nuclei were stained with DAPI dye (VECTOR Laboratories). Images were acquired with a DMIR2 confocal microscope equipped with Leica Confocal Software (Leica).
Monitoring the kinetics of in vitro fibril formation. To monitor the formation of ThT-positive fibrils, we used 100 μ g/mL of WT full-length MoPrP stored in acetate buffer (25 mM NaOAc, 6 M GdnHCl, pH 5. 5) or Tris buffer (25 mM Tris-HCl, 6 M GdnHCl, pH 7. 0). To induce protein polymerization in the amyloid seeding assay (ASA) we added a preformed PrP Sc seed to the reaction purified from ScN2a transiently expressing either 3F4-tagged WT MoPrP or 3F4-tagged H96Y MoPrP according to our previous protocols. Data were analyzed and figures were produced using Origin 8. 6 software. Prion formation in N2a cells. N2a cells were transiently transfected with either 3F4-tagged WT or H96Y MoPrP and regularly passaged every 7 days up to passage (P) 8. Subsequently, the protein extracts were analyzed by PK digestion to monitor the presence of PK-resistant PrP Sc levels through passages (see above).
0 (hereafter termed 1His) only H111 was bound to the metal. Starting coordinates of HuPrP(90-231) were obtained from the 1QM1 PDB structure  4. Missing hydrogen atoms were added by MolProbity  38  and protonation states counter-checked with the H ++ code 39. To model the binding of Cu(II) in the active site a set of harmonic springs was used to enforce copper-ligand distances  40. Bond lengths of the springs were based on EXAFS data and set to 1. 98 Å for the N δ imidazole atoms in H96 and H111, to 1. 98 Å for the amide oxygen in Q98 and to 3. 25 Å for the S atom in M109. The systems were modeled using the AMBER99SB-ILDN* all-atom force field  41  implemented in the GROMACS MD package version 4. 6. 5  42. The protein was immersed in a rhombic dodecahedron box with a minimum distance of 2. 5 nm from the box edges; a long distance from the box edges was used to take into proper account the flexibility of HuPrP(90-231).
Two residues were considered to be bound by a hydrogen (H) bond in a given frame if the H-acceptor distance was below 0. 375 nm and the acceptor -donor-H angle was below 40. 0°. Two residues are connected by a salt bridge if the nitrogen-oxygen distance is below 0. 4 nm in a given frame. All analyses were carried out with standard tools present in the GROMACS MD package v. 4. 6. 5 or with in-house written codes, except for secondary structure assignment which was performed with DSSP  48. Figures were produced with Chimera 1. 10 software.
Acknowledgements This work was supported by  European Community 's  Seventh Framework Programme  ( FP7  grant agreement no. 222887  -the  PRIORITY  project, to G. L. ) and by the  University of Rome  " La Sapienza" (Progetto Ateneo 2014 , n. C26A14L7CX, to P. D. ). We thank  Dr Andrea Zitolo  for initial work on EXAFS data collection. We are thankful the  European Synchrotron Radiation Facility  and  Isabelle Kieffer  at  BM30B  FAME for the invaluable support.
Cell cycle (sub-G1 ) evaluation. OVCAR-3 cells were seeded in culture flasks and incubated with PC or aPC at 10 ng/ml for 24 h. Phase distribution (G1, S and G2) was performed using DNA content using flow cytometry and analyzed by Multi Cycle Av software as previously described  (28). The coefficient of variation (CV) of the mean value of DNA-associated fluorescence of the G1 population (width of the peak) is a reflection of the accuracy of the DNA content measurement. Anticoagulant activity of living cells. Activated partial thromboplastin time (aPTT) evaluates intrinsic coagulation pathway efficiency; coagulation of a plasma sample is measured after addition of phospholipid (cephalin) and calcium. we previously described a method, based on aPTT, to estimate EPCR expression by endothelial and cancer cells (29), i.
4B6 ) and rapamycin a raptor-mTor complex inhibitor  (data not shown). The number of cells migrating outside the droplets, in five experiments, is presented in Fig. 4C. Anticoagulant activity induced by OVCAR-3 cells. The anticoagulant property of OVCAR-3 cells was assessed  by measuring the prolongation of aPTT of normal plasma induced by aPC. As shown in Fig. 5A , the effect of OvCAR-3 cells, detached and incubated either with PC or APC, in fibrin polymerization curve of normal plasma is shown. The same experiment was also performed using OVCAR-3 cells in adherent conditions. when both detached (Fig. 5B ) and adherent (Fig. 5C ) cells, were incubated with aPC, aPTT was prolonged as compared to cells incubated with protein C. As documented in Fig. 5D , PC added to the adherent or detached ovarian cancer cells induced a 2-fold increase in cephalin clotting time, whereas aPC under the same conditions induced a 3.