dataesr/scientific-paragraphs-categorization huggingface dataset:
Parasite counting and schizont size determination using high-content imaging The number, size and shape of the parasites were determined using a CellInsight High Content Screening platform equipped with the Studio HCS software (Ther-moFisher Scientific). Parasite immunostaining was done using a α-Plasmodium HSP70 antibody raised in mice and a secondary anti-mouse antibody coupled to AlexaFluor 488  [23]. Host cell and parasite nuclei were labelled with 4′,6-diamidino-2-phenylindole (DAPI). Thirty-seven images, representing more than 95% of the total bottom surface area of a culture well in 96-well plates, were captured for analysis. The custom script used for counting and measuring parasites first required identification of the objects on the basis of a fluorescence intensity threshold. The object identity was then further validated based on size and morphological (shape) criteria.
The presence of DAPI-associated fluorescence in the selected objects allowed for their final selection and rejection of false positives. Inhibitory concentration 50 (IC 50 ) value is the drug concentration at which a 50% reduction of the exo-erythrocytic forms (EEFs) number was observed, as compared to the control cultures. IC 50 were calculated using ICEstimator software version 1. 2  [24]. The parasite size reduction is calculated on the average object area using the total surface area of each selected object (µm 2 ). Cell confluence for primary hepatocytes was measured by counting the absolute number of host cell nuclei using a script that first requires identification of the objects on the basis of the DAPI fluorescence intensity threshold and then involved a validation step based on size and morphological (shape) criteria.
IVIS Spectrum (Caliper Life Science, Hanover, MD, USA) was used to measure luciferase activity. Prior to analysis the mice were injected ip with D-luciferin (100 mg/kg), anesthetized with isoflurane and imaged 10 min post-injection. Images were analysed using the living Image 3. 0 software (Capiler Life Science, Hanover, MD, USA). For blood stage assessment, luminescence (total flux photons/seconde) and Giemsa-stained blood smears were done on days 3, 4 and 6 pi to evaluate the blood stage prepatent period in mice after sporozoite inoculation.
Miscibility of AGs and PLA Mixtures of PLA and AG were dissolved in 2 mL acetone. The solvent was evaporated in a ventilated oven at 37°C and the obtained solid samples were dried under vacuum for 24 h. The thermal transitions of PLA + AG solid mixtures were characterized by modulated differential scanning calorimetry (mDSC). The mDSC thermogramms (3 cycles) were recorded using a Q2000, TA Instruments apparatus. The heating rate was 5°C/min while the cooling rate was (10°C/min) under nitrogen, and the temperature was varied from -25°C up to 200°C. First MDSC cycle was used to remove all thermal history from the sample. Analysis of the data was performed using Universal ® Analysis 2000 (version 4. 5A) software. where d(0. 9), d(0. 5) and d(0. 1) were the particle diameters at 90%, 50% and 10% cumulative volumes, respectively.
Hence, any potential   [16]. Right panel, human CYP2B6 complexed by amlodipine at 2. 24 Å resolution (pdb entry 4zv8)  [17]. The amino acid sequence identity between the two proteins is 14%. The heme is represented in both enzymes as red sticks and the bound ligand as orange sticks. The heme-liganding cysteine is colored yellow. Protein visualizations were performed by using PyMol (Delano Scientific LLC, San Carlos, CA, USA). The caption under both of the crystal structures represents the single CYP74A substrate (allene oxide) on the left and some of many CYP2B substrates on right. The extraordinary chemical ability of P450 enzymes is mainly based on three features that all of these enzymes share. One is the fact that the active site of a P450 is a cavity that is buried within the protein molecule, and thus, clearly separated from the solvent  [18].
muni. cz/ChannelsDB/), which presents a database of all annotated access channels in enzyme structures, including cytochromes P450. This database contains information about channel positions, geometry, and physicochemical properties that are presented in a clear and interactive manner facilitating interpretation. Fifteen years ago, Wade et al. presented the first review of the different ligand routes that they had identified in both bacterial and mammalian P450 structures with the RAMD approach  [37]. A decade ago, Cojocaru et al. have classified and named the different access channels that were found by CAVER on structurally characterized P450 enzymes in a systematic manner  [38]. The different ligand access channels that were identified in P450 enzymes were each named by a different number (channels 1, 2, 3, 4, W, and S).
This motion was observed both when comparing the crystal structures of the same P450 complexed with different ligands and in silicoby revealing the malleability of this region in P450 enzymes. However, the movements of the different elements forming the F/G region are not always easy to identify or to simulate in molecular dynamics, since, for instance, four of the 24 crystal structures of CYP3A4 available in the Protein Data Bank (PDB) have missing residues in the F and F' helices. In this respect, it is interesting to note that two of the less-studied channels that were seen with CAVER in P450 enzymes, namely channels 3 and 4, have their opening right in-between the F and the G helices, one being very close to the F'/G' helices(channel 4) and the other close to the F/F' loop  [38]. It has been shown that one of them, channel 4, couldbe predominantly a metabolite egress channel in CYP3A4  [67].
PBPK models of some type are used by most pharmaceutical companies to guide the molecular design of inhaled drugs. However, the combination of PBPK models, which provide understanding of tissue and target interactions, with mechanistic models, which describe key processes governing the rate and extent of local drug exposure, is still in its infancy. For example, there is currently only one commercially available PBPK software with a mechanistic regional deposition, dissolution and permeation model designed for pulmonary drug delivery (Gastroplus TM Nasal-Pulmonary Compartmental Absorption and Transit Model, SimulationsPlus Inc. , Rochester, US). Other commercial PBPK software, such as the SimCyp Simulator TM account for pulmonary delivery by reducing dissolution and epithelial permeation into a single first order process in a single pulmonary compartment (https://www.
Nine compounds were further evaluated to test the model's predictive ability. Molecular descriptors associated with permeability and hydrophobicity were found to be positively correlated with pulmonary absorption whereas descriptors for charge, ionization and size were negatively correlated. Such QSAR modelling exercises can generate descriptors which can be used as inputs during the mechanistic inhalation modelling during the drug discovery phase where limited experimental data is available. For example, computational, multiscale, cell-based modelling has been used to explore the relationship between the physicochemical properties and absorptive pharmacokinetics of monobasic molecules in the lungs  (Yu et al 2010). A programme to assess the effect of permeability on lung concentration after pulmonary administration is generating an expanding and consistent dataset for antibiotic compounds absorption from rat lungs in vivo  (Gontijo et al, 2014a , Gontijo et al, 2014b , Marchand et al, 2015 , Marchand et al, 2016).
PK-SIM™ and SimCyp Simulator TM Although Gastroplus is the only commercially available software to incorporate a mechanistic model of deposition, dissolution, non-absorptive clearance and absorptive clearance, non-mechanistic PBPK models are available. The SimCyp Simulator TM (https://www. certara. com/software/physiologically-based-pharmacokinetic-modeling-andsimulation/simcyp-simulator/absorption/) and PK-SIM™ (Computational Systems Biology: Bayer AG, http://www. systems-biology. com/products/PK-Sim. html) allow the user to define pulmonary and gastrointestinal (GI) absorption compartments and model PK following pulmonary administration. For example,  Stass et al, (2008 Stass et al, ( , 2013) )  used PK-SIM to deconvolute PK data obtained in healthy volunteers after inhalation of ciprofloxacin, a locally acting antibiotic, to obtain the relative contribution of oral, tracheobronchial (BB and bb) and alveolar interstitial (AI) deposited drug to the total systemic exposure.
Briefly, 200 mg of LS or 4 g of gel were placed into a dialysis tube (molecular weight cut-off 10,000-12,000; Medi Cell International, London, UK), then put into 40 mL of a receiving phase constituted of SVF/ethanol (80:20, v/v), and shaken in a horizontal shaker (MS1, Minishaker, IKA) at 175 r. p. m. at 37 • C. In addition, the release kinetics of free CLO were investigated, placing in dialysis tubes 2 mg of CLO dispersed in 4 mL of distilled water or in 4 g of xanthan gum gel. Samples of receiving phase were withdrawn at regular time intervals and analyzed by HPLC, as described above. Fresh receiving phase was added to maintain a constant volume. The CLO concentrations were determined six times in independent experiments, and the mean values ± standard deviations were calculated. The experimental data obtained by the release experiments were fitted to the following semiempirical equations, respectively describing Fickian dissolutive (4) and diffusion (5) release mechanisms  [32, 33]  M t /M∞ = K Diss t 0.
Acknowledgments: The authors are grateful to  Maria Rita Bovolenta  for her help in microscopic measurements. Author Contributions:  Elisabetta Esposito  and  Rita Cortesi  conceived and designed the experiments;  Maddalena Sguizzato  and  Christian Bories  performed the experiments;  Claudio Nastruzzi  analyzed the data and helped full discussion;  Elisabetta Esposito  and  Rita Cortesi  wrote the paper.
Skin Delivery The skin extraction recovery from skin samples was 35. 98 ± 3. 23 (9%). This percentage of recovery was used to correct data from the permeability assays. The passage of EA was evaluated to determine the best penetration enhancement conditions, from a cosmetic composition using a short application time and different current profiles. Analysis of the skin profiles suggested that the SPC profile applied during 5 min at 0. 2 mA/cm 2 improved the topical transport of EA compared to that obtained from its passive application. The combined positive DC + PC profile did not show any particular advantage in increasing the penetration of the phenolic compound (p < 0. 05; ANOVA), see Figure  5. EA was stable in the presence of an electric current at 0. 03 mA/cm and 0. 3 mA/cm during 30 min period of the application tested, obtaining a fraction of the initial EA concentration of 100.
Skin Delivery The skin extraction recovery from skin samples was 35. 98 ± 3. 23 (9%). This percentage of recovery was used to correct data from the permeability assays. The passage of EA was evaluated to determine the best penetration enhancement conditions, from a cosmetic composition using a short application time and different current profiles. Analysis of the skin profiles suggested that the SPC profile applied during 5 min at 0. 2 mA/cm 2 improved the topical transport of EA compared to that obtained from its passive application. The combined positive DC + PC profile did not show any particular advantage in increasing the penetration of the phenolic compound (p < 0. 05; ANOVA), see Figure  5. The iontophoretic delivery efficiency using the DC profile for EA delivery from simplex solutions was also investigated in order to understand the behavior of the molecule when present in its different ionized states.
, USA) followed by incubation with goat IgG anti-rabbit conjugated with Alexa Fluor 555 A-21428 (Molecular Probes, USA) for immunofluorescence detection. Nuclei were evidenced by DAPI (4, 6-diamidino-2-phenylindole) staining (Sigma-Aldrich); and slides were mounted with Permafluor (Thermo Scientific, Waltham, MA) and analyzed via the AxiObserver Z1 Colibri microscope. Immunofluorescence was quantified using Open-Source ImageJ1 software (Research Services Branch, National Institutes of Health, Bethesda, MD, USA). Protocol for antigen labeling of eukaryotic cells and detection by flow cytometry available at: dx. doi. org/10. 17504/protocols. io. pg8djzw. Protocol for antigen labeling of eukaryotic cultured cells and detection by fluorescence microscopy available at: dx. doi. org/10. 17504/protocols.
Lipid droplets staining and quantification Schwann cells adhering to coverslips were fixed in 4% paraformaldehyde, and the area occupied by LDs were analyzed using fluorescent Oil red O (ORO, Cat. N˚O0625 SIGMA). Coverslips were mounted, the morphology of the fixed cells was observed, and LDs areas were enumerated in 500 consecutively scanned cells. Nuclei were stained with 2 mM DAPI (Sigma-Aldrich) at room temperature for 5 min. The ORO images (taken with a 40-objective lens) were transformed into black and white pictures and analyzed via Open-Source ImageJ1 software. The spots were determined by automatic spot detection; and the total area and average size of fluorescent LDs was obtained for each field and divided by the number of cells in the respective field. Fluorescence microscopy used the Axio Observer Z1 (Carl Zeiss) microscope and AxioVision Rel.
M. leprae viability M. leprae viability was determined by qRT-PCR using the protocol previously described by Martinez, et al  [78]. Briefly, ST8814 cells infected for 48 h with M. leprae and pre-incubated with mannose were submitted to RNA and DNA extraction with TRIzol (Thermo Fisher Scientific). DNA was removed from the RNA preparations using the DNA-free turbo kit (Ambion). RNA was reverse transcribed via a random primer and GoScript kit following the manufacturer's instructions (Promega). The levels of 16S rRNA were determined and normalized against 16S DNA. PCR efficiency for each experiment was gauged via LinRegPCR software  [79] ; and normalization was performed considering efficiency corrections, as described above  [80]. The same set of primers for the rRNA 16S was used for both cDNA and DNA: sense 5 ' -GCATGTCTTGTGGTGGAAAGC -3 ' and antisense 5 ' -CACCCCACCAACAAGCT GAT -3 '.
Statistical analysis Results are presented as the mean ± standard error of the mean (SEM) of independent experiments at a minimum in triplicate. Data were analyzed using the GraphPad Prism 5 Project (GraphPad Software, La Jolla, CA, USA). Unless otherwise stated, analyses were performed by applying one-way ANOVA and the Bonferroni post-test to compare the different groups. A pvalue <0. 05 was considered statistically significant. BCG WT or BCG PGL I. After 48 h of incubation at 33˚C and MOI 50:1, cell viability was determined by MTT assay. Each result is shown as mean ± SD of three assays. MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (Sigma-Aldrich). (TIF).
Acknowledgments The following reagents were kindly obtained from the  NIH Biodefense and Emerging Infections Research Resources Repository ,  NIAID ,  NIH :  Mycobacterium leprae Phenolic Glycolipid I (PGL I)  ( NR-19342 ) Man-LAM NR-19348 and anti-LAM Rabbit polyclonal  NR1809. The authors are grateful to  Judy Grevan  for editing the text.
Baseline patient characteristics were similar in both patient groups, except for day 0 parasitemia and gametocytemia, which were higher for the DHA/PPQtreated group (Table  1 ). The proportion of infectious blood from P. vivax-infected patients and the median proportion of infected mosquitoes fed on blood collected before the  Each dot represents the parasite transmissibility reduction ratio (i. e. , 100 -[proportion of infected mosquitoes fed with blood samples collected at 9:00 pm after the first dose of treatment × 100/ proportion of infected mosquitoes fed with blood samples collected at patient enrollment before the first dose of treatment]). Only data of infectious patients at enrollment are shown (17/19 patients; Table  1 ). For the chloroquine-treated patient group, a second mosquito blood feeding was performed 24 h after the first dose.
Extraction of nucleic acids Genomic DNA was prepared from liver fragments (0. 3e0. 5 g) and Fao hepatoma cells (13 Â 10 6 cells) by phenol extraction and ethanol precipitation after digestion by proteinase K. Total liver RNA was prepared by a single step procedure as described previously  [12]. DNA and RNA integrity was assessed by electrophoresis on 0. 8 or 1% agarose gels containing 0. 1 mg/ml BET followed by UV visualization. 2. 3. Cloning of fragments of the rat IR gene proximal 5 0 flanking region The À1143/À252 fragment of the proximal 5 0 flanking region of the rat IR gene was generated by PCR amplification of liver genomic DNA using as primers the oligonucleotides 5 0 GCTCCAGACCTTTAAATTGGA-3' (sense) and 5 0 CGTCC TGTGTCCGCAATCGC-3' (antisense). The former corresponds to the À1009/À989 sequence of the mouse IR gene (Genbank accession M28869.
1), which exactly matches the À1143/À1123 sequence of the rat IR gene, and the latter to the À271/À252 sequence of the 347 bp 5 0 untranslated portion of rat IR cDNA  [23]  (Genbank accession M29014. 1). The specificity of the À1143/À252 fragment was verified by Southern blotting using as a probe a 32 P-labeled EcoR1-Eco47III fragment of rat IR gene cDNA (sequence À347/À15). Following cloning and sequencing of two additional fragments (À890/ À514 and À1143/À563), the complete sequence of the À1143/À252 fragment was determined and the À1143/À1 sequence was deposited in Genbank (accession AJ006071). Two 5 0 truncated DNA fragments, encompassing À760/À252 and À464/À252 sequences, were prepared by digestion of the parental À1143/À252 fragment cloned in the PCR TM II vector by digestion with the restriction enzymes Pvu2 and Dra1, respectively.
Sequence analysis of the rat IR gene proximal 5 0 flanking region Database searches and alignments of the 5 0 flanking sequence of the rat IR gene with that of the mouse and human IR genes were performed using the blast program (ncbi. nlm. nih. gov/BLAST). The rat sequence was scanned for potential SINEs using SINE base search (http://sines. eimb. ru) and for potential transcription factor binding sites using the Jaspar software and Jaspar core database (http://jaspar. genereg. net).
Structural characterization of the proximal 5 0 flanking region of the rat IR gene The proximal 5 0 flanking region of the IR gene was first cloned in human  [24]  and mouse  [25]  over 1. 8 and 1. 2 kb upstream the ATG, respectively. Alignment of the À1143/À1 sequence of the rat IR gene (this study, Genbank accession AJ006071) with the 1009/À1 sequence of the mouse IR gene (Genbank accession M28869) reveals a 86% identity at À766/ À1 and À734/À1, and a 83% identity at À1143/À993 and À1009/À858, respectively. In contrast, identity of the rat sequence with the À1823/À1 sequence of the human IR gene (Genbank accession M23100) is only 68% at À558/À1 and À536/À1, respectively, with a maximum (83%) at À523/À407 (rat) and À545/À429 (human). Alignments of rat (NC_005111. 4), mouse (NC_000074. 6) and human (NC_018930.
Statistical analysis All data were expressed as mean AE SD. Results were analyzed by one-way analysis of variance (ANOVA), and signicant differences were determined by post-hoc Tukey test using SPSS 21. 0 soware. #P < 0. 05 compared to control group; ## P < 0. 01 compared to control group; *P < 0. 05 compared to model group; and **P < 0. 01 compared to model group. P value < 0. 05 was considered as signicant.
Structure Elucidation of Compounds 1-4 The molecular formula of compound 1, C 15 H 21 Br 3 O 3 , was determined by HRESIMS and from NMR data (Table  1 , Figures  S1-S7 ). Interrogation of our 13 C NMR database suggested that there were similarities to the previously described sagonenyne (compound 5)  [9]. Interpretation of 1D and 2D NMR spectra confirmed that compound 1 shared the same 10-hydroxy-12-bromo-13-ethyltetrahydropyran ring as sagonenyne. However, the differences between the chemical shifts of both of the molecules (∆δ) were larger than 0. 50 ppm for seven out of 15 carbons, indicating that half of the molecules exhibited different structural features. Furthermore, compound 1 lacked the acetoxyl group and exhibited a bromoallene unit [δ H 6. 11 (1H, dd, J = 5. 7, 1. 4 Hz, H-1) and 5.
1 Hz), as well as the small coupling constants between H-9 and H-10 (1. 1 Hz), and between H-6 and H-7 (3. 0 Hz), led to the same conclusions as for compound 1: except the C-12 substitution orientation, all other substituents exhibited a syn relative orientation. Therefore, compound 2 exhibited the same relative configuration as 1, namely 4R*,6R*,7R*,9R*,10R*,12R*,13S* (Figure  1 ). The molecular formula of compound 3, C20H35BrO2, was established by HRESIMS. An interrogation of our 13 C NMR database provided no suitable candidate for this compound. However, the 13 C chemical shifts of compound 3 recorded in acetone-d6 matched perfectly with those of concinndiol  [11, 12]. Since NMR data of concinndiol has remained incomplete up to this point, we report here its complete and assigned list of 1 H and 13 C NMR data (Table  2 ) for the first time.
Therefore, compound 2 exhibited the same relative configuration as 1, namely 4R*,6R*,7R*,9R*,10R*,12R*,13S* (Figure  1 ). The molecular formula of compound 3, C 20 H 35 BrO 2 , was established by HRESIMS (Figure  S20 ). An interrogation of our 13 C NMR database provided no suitable candidate for this compound. However, the 13 C chemical shifts of compound 3 recorded in acetone-d 6 matched perfectly with those of concinndiol  [11, 12]. Since NMR data of concinndiol has remained incomplete up to this point, we report here its complete and assigned list of 1 H and 13 C NMR data (Table  2 , Figures  S14-S19 ) for the first time. The assignment of OH signals was based on the HMBC spectrum. Long-range correlation cross-peaks were observed between the OH resonance at 3. 04 ppm and carbons C-8, C-9, C-10, and C-11, as well as between the signal at 3.
This kind of skeleton had been previously isolated from the Mediterranean sponge Mycale rotalis  [13]. Indeed, rotalin A is the (9-13)-epoxy analogue of compound 4. 13 C chemical shifts of both molecules were compared and no significant difference was observed (∆δ < 1. 35 ppm). The relative configuration of C-3, C-5, C-8, and C-10 was determined based on NOESY data. NOESY cross-peaks observed from H-19 to H-1a, H-2b, and H-5, along with correlations from H-5 to H-1a and H-7a, as well as from H-17 to H-7a, indicated that H-5 and methyls C-17 and C-19 had a syn orientation. NOESY cross-peaks observed from H-20 to H-1b and H-8 showed that methyl C-20 was positioned on the other side of the molecule. Concerning C-9 and C-13, the relative configuration was established by a comparison of chemical shifts with the data contained in the literatures.
Among them, only two labdane derivatives bear a hydroxyl on C-9 (concinndiol and neoconncindiol hydroperoxyde) and all of them exhibit a bromine substitution on C-3, except laukarlaol and neoconcinndiol hydroperoxyde  [14]. Altogether, the 19(4→3)abeo-labdane skeleton isolated in this study has never been reported in Laurencia, and thus represents a highly original structure to be added to the already extensive chemical diversity of this genus. dimension ( 1 H) and of 220 ppm in F1 dimension ( 13 C), and an accumulation of 64 scans. NOESY spectra were recorded using the Bruker microprogram (PULPROG noesyph, Bruker, Wissembourg, France), a mixing time of 300 ms, 1024 data points in F2 dimension and 256 data points in F1 dimension, a SW of 12 ppm in both dimensions, and an accumulation of 64 scans.