dataesr/scientific-paragraphs-categorization huggingface dataset:
Acknowledgements The authors wish to thank the different facilities of Genotoul (Metatoul, Get, and Anexplo) in Toulouse and the  Genomic Technologies Facility of the Center for Integrative Genomics, University of Lausanne , and the germ-free facilities at  Karolinska Institute (Stockholm)  and at  Lee Kong Chian School of Medicine, Nanyang Technological University (Singapore ). We also acknowledge the help of Aurore Desquesnes from Anexplo with the plasma analyses. Work performed at ToxAlim was supported by grants from the  Région Midi-Pyrénées  to HG (Mathusalem) and to WW ( Chaire d'Excellence Pierre de Fermat ). This work was also supported by grants from the  Swiss National Science Foundation  (individual grants to WW), the  Bonizzi-Theler-Stiftung (WW) , the 7th  EU program  TORNADO (WW, SP), and by the  Etat de Vaud  and the  Lee Kong Chian School of Medicine, Nanyang Technological University Start-Up Grants.
d At all times, patient adherence and intranasal device technique mastery should be regarded as potential for lack of treatment effect. Alternatively, a step-down approach can be used, and step 3 treatment should be considered as the first option in patients with a previous treatment failure or resistance to monotherapy. After a few days of achieving complete control, consideration could be given to treatment reduction. However, the step-down approach is based on consensus, and more data are needed. The duration of treatment is determined by the type of rhinitis (intermittent or persistent). In the patient with intermittent rhinitis, treatment should be continued daily for 2 weeks or for the duration of the pollen season or other specific allergen exposure. In the patient with persistent rhinitis, a longer course.
Data Acquisition and Analysis Electrical data were filtered by the Multiclamp 700B amplifier and digitized through a Digidata 1440A DAC converter controlled by pClamp 10. 3 software, which was also used for triggering the mechanical shutter (Uniblitz) of the laser. Short (100 ms) hyperpolarizing steps (5 mV) were used to measure access resistance which ranged from 8 to 20 M and membrane resistances varying from 200 M to 12 G. Evoked photocurrents were triggered by short opening (from 15 to 160 ms) of a mechanical shutter in front of the laser controlled by pClamp after a short baseline period (100 ms). The filter frequency was 10 KHz for resistance steps recordings and 6 KHz for ChR2 current recordings. Sampling frequencies were of 20 KHz and 10 KHz respectively. For analysis, the data were imported into the Igor programming environment (Wavemetrics) and analyzed using home-made analysis routines.
Epi-fluorescence illumination and image acquisitions were controlled by the image acquisition software SlideBook 6 (Intelligent Imaging Innovations). Images were then saved in TIFF format and uploaded into the home made Wavefront Designer IV software for analysis of the level of expression and generation of the correct pattern to be sent to the SLM. The average fluorescence was calculated as the integral over the whole cell divided by its surface. All other analysis and statistics were performed into the Igor programming environment. Results are given as mean ± one standard deviation (SD) or standard deviation of the mean (SEM) as noted in the text.
ACKNOWLEDGMENTS The authors wish to thank  Erwin Neher  for a usefull discussion on opsin biophysics,  Cécile Jouffret  for help with cell culture preparation,  Karl Deisseroth  for kindly providing the plasmids. This work received financial support from the  "Agence Nationale de la Recherche " ( ANR-12-BSV5-0011-01   Neurholog  and  ANR-10-INBS-04-01 ,  France-BioImaging Infrastructure network ), the  FRC  and the  Rotary Club through the program  " Espoir en Tête " 2012 and the  National Institutes of Health  ( NIH 1-U01-NS090501-01 ).
Magnetic resonance imaging image analysis MRI (T 2 W) images were used to calculate the lesion volume. Regions of Interest (ROIs) were manually defined using the Open Source software 3D Slicer image analysis software (Version 3. 6. 3 www. slicer. org) for each rat on the region of increased signal in the ipsilateral hemisphere. The total lesion volume was calculated by summing the area of the infarcted regions of all slices affected by the lesion.
Positron emission tomography scans and data acquisition PET scans were repeatedly performed before (day 0) and at 1, 3, 7, 14, 21 and 28 days after reperfusion using a General Electric eXplore Vista CT camera (GE Healthcare). Scans were performed in rats anaesthetized with 4% isofluorane and maintained by 2-2. 5% of isofluorane in 100% O 2. The tail vein was catheterized with a 24-gauge catheter for intravenous administration of the radiotracer. Animals were placed into a rat holder compatible with the PET acquisition system and maintained normothermic using a water-based heating blanket. Animals were subjected to PET scans to assess system xc -([ 18 F]FSPG) and TSPO binding ([ 18 F]DPA-714) at each time point before and after ischemia onset. The appropriate radiotracer ([ 18 F]FSPG or [ 18 F]DPA-714, ~70 MBq) was injected concomitantly with the start of the PET acquisition and dynamic brain images were acquired (24 frames: 1x5, 1x15, 3x30, 5x60, 5x120, 3x180, 6x300 seconds) in the 400-700 keV energetic window, with a total acquisition time of 57.
Positron emission tomography image analysis PET images were analyzed using PMOD image analysis software (PMOD Technologies Ltd, Zürich, Switzerland). To verify the anatomical location of the signal, PET images were co-registered to the anatomical data of a MRI rat brain template. Two type of Volumes of Interest (VOIs) were established as follows: (i) A first set of VOIs was defined to study the whole brain [ 18 F]FSPG and [ 18 F]DPA-714 PET signal over time. Whole brain VOIs were manually drawn in both the entire ipsilateral and contralateral hemispheres containing the territory irrigated by the middle cerebral artery on slices of a MRI (T 2 W) rat brain template from the PMOD software. (ii) A second set of VOIs was automatically generated in the cortex and the striatum by using the regions proposed by the PMOD rat brain template, to study the evolution of PET signal in these specific regions in both the ipsilateral and contralateral cerebral hemispheres.
Real-time quantitative PCR (qPCR) Frozen tissue was cut into 40 µm slices on a cryostat and a pool of 3-4 different slices of ipsilateral or contralateral hemispheres was used per each condition. Total RNA was isolated from tissue using Trizol (Invitrogen) according to the manufacturer's instructions. Subsequently, cDNA synthesis was conducted using SuperScript III retrotranscriptase (200 U/µL; Invitrogen) and random hexamers as primers (Promega). Primers specific for the rat M1 and M2 markers were designed using Primer Express software (Applied Biosystems) at exon junctions to avoid genomic DNA amplification (see Table  1 ). Data was normalized to HPRT gene (best M coefficient using GeNorm Software v3. 5; Vandesompele et al. , 2002). Real-time quantitative PCR reactions were performed with SYBR-Green using a BioRad CFX96 Real-time PCR detection system as described previously  [30].
Neurological outcome comparisons were performed as follows: animals were subjected to the 9-neuroscore test before MCAO and at 1, 3, 7, 14, 21 and 28 days after cerebral ischemia. The results within each time point and after treatment were averaged and compared with baseline average values using Mann-Whitney U-tests. The effect of treatments in the mRNA levels for M1/M2 microglial phenotypes in sham, treated and non-treated ischemic rats were compared using a one-way ANOVA followed by Bonferroni's multiple-comparison tests for post hoc analysis. Finally, the level of arginase mRNA in treated ischemic rats was compared with non-treated ischemic rats using an unpaired t-test. The level of significance was regularly set at P<0. 05. Statistical analyses were performed with GraphPad Prism version 6 software.
Acknowledgements The authors would like to thank  M González ,  A Leukona ,  M Errasti ,  A Arrieta  and  A Cano  for technical support in the radiosynthesis and technical assistance in the PET studies, and the  Department of Industry of the Basque Government  and  Spanish Ministry of Economy and Competitiveness  through grant  SAF2014-54070-JIN  and  SAF2013-45084-R  for financial support.
Two-electrode voltage clamp in Xenopus oocytes cRNA was created from Slack-B and mutant channel cRNA in pOX oocytes expression vector with a mMessage mMachine T3 kit (Ambion) and aliquoted in sterile water. Oocytes were isolated from Xenopus laevis frogs, injected with 50 -100 ng of cRNA, and incubated in MND96  (Brown et al. , 2010)  at 18° C for up to 7 days before experiments were performed. Whole-oocyte currents were measured by a two-electrode voltage clamp amplifier (OC-725C, Warner Instruments. ) Electrodes were filled with 3 M KCl and had resistance 0. 1 -1 MΩ. Standard bath solution was MND96. Ooctyes were depolarized by 400 ms pulses from a holding potential of -90 mV to test pulses of between -80 and +60 mV in 10 mV increments for 5 seconds. Data recording and analysis were performed using pClamp (Molecular Devices), Excel (Microsoft) and Origin 8.
A density gradient map was generated for both records, and individual cells and the local background selected utilizing BIND Scanner analysis software. The change in the index of refraction for each cell and the local background as a function of the difference between the first and second images was calculated. The change in index of refraction for each cell was normalized to the local background, and a mean value calculated for each well. The change in peak wavelength of refraction was divided by the initial peak wavelength to give a physically relevant change in the index of refraction. For primary cortical neuron experiments, niclosamide addition was normalized to DMSO vehicle alone addition. Results were exported to Microsoft Excel and Origin Statistical Package for analysis.
Statistical Analysis Data recording and analysis were performed using pClamp (Molecular Devices), Excel (Microsoft), Origin (Origin Lab Corporation), and BIND Scanner and BIND View (X-BODY Biosciences). The mean±SEM values are plotted. Where 2 groups were compared, 2tailed student's t-test was performed to compare statistical significance; where 3 or more groups were compared, 1-way ANOVA test was performed. See Supplemental Experimental Procedures for chemicals, cell culture, creation of mammalian expression vectors, animals, yeast two hybrid assay, RNAi against candidate Slack-B binding partners, cell viability assay, protein purification of RNAi samples, coimmunopurification experiments, western blotting, as well as detailed experimental procedures.
Acknowledgments The authors are grateful to  Yangyang Yan  and  Fred Sigworth  at  Yale University  for donation of Slack-B, Slick and BK cell lines and technical assistance with protein purification, and to  Steven Shamah  and  Brandt Binder  at  X-BODY Biosciences  for the usage of the BIND Scanner system and technical assistance. This work is supported by grants from the  NIH  ( HD067517 ) and from  FRAXA  to L. K. K.
Patch-Clamp Recordings Whole-cell voltage clamp was made in neurons seeded on coverslips, placed in RC-25 chamber and connected to a external perfusion system at ≈22 • C. External solution was (in mM): 140 NaCl, 4 KCl, 2 CaCl 2 , 2 MgCl 2 , 10 HEPES, 5 D-glucose, 20 mannitol, pH 7. 4 (adjusted with NaOH). Internal pipette solution was (in mM): 144 KCl, 2 MgCl 2 , 10 HEPES, 5 EGTA, pH 7. 2 (adjusted with KOH). Membrane currents were acquired using EPC10 HEKA Patch amplifier (HEKA Electronics). Cells were held at a RMP of -60 mV, and at a sampling rate of 2. 5 Hz. Patch glass pipettes with OD 1. 5 mm × I. D. 1. 17 mm (Harvard Instruments) were pulled with a Sutter Pippete puller Equipment (Sutter Instruments) to have a 3-6 M resistance. Data acquisition and offline analysis were performed with PatchMaster software (HEKA Electronics).
MEA Measurements Extracellular recordings were made using multiple electrode planar arrays of 60-electrode thin MEA chips, with 30 µm diameter electrodes and 200 µm inter-electrode spacing with an integrated reference electrode (Multichannel Systems GmbH). The electrical activity of primary sensory neurons was recorded by the MEA1060 System (Multi Channel Systems GmbH 1  ), and MC_Rack software version 4. 3. 0 at a sampling rate of 25 kHz. TRPV1-mediated neuronal firing activity was evoked by three repetitive 15 s-applications of 500 nM capsaicin, using continuous perfusion system (2 mL/min). BK (1 µM) in external solution was perfused between P2 and P3 for 8 min. For experiments on BK receptor agonists, selective BK receptor agonists for BKR1 and BKR2 were applied between P2 and P3 for 8 min.
Procedure Participants were comfortably seated in front of a Wacom graphic tablet (Intuos4 XL) positioned on the tabletop and connected (via a USB port) to a portable PC. They had to perform pointing movements with the right arm, by sliding a hand-held non-marking stylus (Wacom, Generation 2 tip sensor) over the surface of the tablet, from a starting position toward a target position. Sliding movements were performed in the anterior-posterior direction, with a combination of shoulder flexion and elbow extension. The trunk position was restricted by the chair-back position and the front edge of the table (see Fig. 1 ). Displacements of the stylus on the tablet were recorded using a customized software (ICE) developed at the laboratory (Institute of Movement Sciences, Marseille) at a sampling frequency of 250 Hz.
Variables and data processing The pen-tip raw displacement data were filtered using a secondorder dual pass (no phase-lag) Butterworth filter with a cut-off frequency of 10 Hz. First, second, and third derivatives of displacement (velocity, acceleration, and jerk, respectively) were then computed in MATLAB (MathWorks, v. 7. 5. 0 R2007b). Movement onset and offset were determined on the basis of velocity profiles using the optimal algorithm of  Teasdale, Bard, Fleury, Young, and Proteau (1993). The critical velocity threshold was obtained by multiplying peak velocity by 0. 05. First, this procedure allowed us to calculate for each trial, in each condition, the movement time (MT) corresponding to the time to reach the target and the effective target width (We). The effective target width (We) was calculated from the standard deviation of movement end points  (Mackenzie, 1992)  using the following formula: We = 2 * 1.
7, 8  FADS is the product of FLAD1 (MIM: 610595), which according to the UCSC Genome Browser encodes two transcripts resulting in proteins with an intact FADS domain. Those two transcripts have been characterized in detail;  [7] [8] [9] [10] [11] [12]  a longer isoform derives from a large exon 1 and has the potential to encode FADS1 with a predicted mitochondrial targeting sequence (MTS). In a shorter isoform, exon 1 is interrupted by an additional intron. This leads to usage of a downstream ATG start codon at position 98 of the longer transcript, resulting in FADS2. 13 Both FADS proteins contain an N-terminal molybdopterin binding (MPTb) domain, which has been shown to have FAD hydrolase activity,  10  and a C-terminal 3 0 -phosphoadenosine 5 0 -phosphosulfate (PAPS) reductase domain. This second domain is sufficient to catalyze FAD synthesis.
For subject S7, a panel of 17 genes (Table  S1 ) was analyzed with a targeted resequencing approach respect to isoform 1 (GenBank: NM_025207. 4). Isoforms 1 and 2 are reported in the UCSC Genome Browser as transcripts possessing an intact and active FADS domain. The protein structure highlights the MPTb domain in violet and the FADS domain in orange. Protein consequences of the identified FLAD1 mutations include the frameshift variants located in the MPTb domain and the two amino acid changes in a region of the FADS domain, which is highly conserved among eukaryotic species. Amino acid residues that are conserved across all species are highlighted in dark blue. by Ion AmpliSeq technology (Life Technologies) on a PGM analyzer (Life Technologies). All identified variants and their segregation in the affected families were validated by Sanger sequencing.
SDS-PAGE and Immunoblotting Liver tissue was homogenized with a potter-type homogenizer on ice in buffer (250 mmol/L sucrose, 50 mmol/L Tris-HCl [pH 7. 4], 5 mmol/L MgCl 2 , EDTA-free protease inhibitor cocktail, and 1% Triton X-100), and frozen fibroblast pellets were lysed in a buffer containing 50 mmol/L Tris-HCL (pH 7. 8), 5 mmol/L EDTA (pH 8. 0), 1 mmol/L DTT, 10 mg/mL Aprotinin (Sigma-Aldrich), 1 mg/mL trypsin inhibitor (Roche) in 10 mL, and 1% Triton X-100. Aliquots of total liver protein (50-100 mg) were loaded on 14% SDS-PAGE gels, and 20 mg fibroblast protein lysate was loaded on Criterion GGX Stain-Free Precast Gels (Bio-Rad) and transferred to polyvinylidene difluoride membranes. Membranes were probed with human anti-FLAD1 antibody (fibroblasts: HPA028563, Sigma-Aldrich; liver: ab95312, Abcam) and anti-GAPDH (sc25778, Santa Cruz).
Detected proteins were visualized with the ECL Plus Western Blotting Detection System (GE Healthcare). Mass Spectrometry to Confirm the Identity of Anti-FADS Gel Bands 80 mg of total protein from control fibroblasts was extracted and separated by SDS-PAGE as described above. The bands of interest (~60, 50, 37, 32, and 27 kDa) were excised and processed by an in-gel trypsin-digestion protocol as previously described. 41 elected reaction monitoring (SRM) was performed as previously described. 42 Peptide selection, method development, and data analysis were performed with the software Skyline. 43 ive FADS peptides passed selection criteria based on detectability and FADS uniqueness (checked in the whole human proteome, UniProt version 2012_06_19, containing 20,202 reviewed sequences), and these five FADS peptides were analyzed by liquid chromatography coupled with the TSQ Vantage Triple Quadrupole Mass Spectrometer (Thermo Fisher Scientific).
44 200 ml of muscle homogenate was mixed with 500 ml methanol and centrifuged, and the supernatant was injected for HPLC analysis (mobile phase: 50 mmol/L NaH 2 PO 4 and 13% acetonitrile [pH 2. 9]; flow rate ¼ 1. 2 mL/min). For measurements in fibroblast cells, the frozen cells were re-suspended in 150 ml lysis buffer (50 mmol/L Tris-HCl [pH 7. 5], 1% Triton X-100, 5 mmol/L b-mercaptoethanol [2-ME], 1 mmol/L NaF, 0. 1 mmol/L PMSF, and 13 protease-inhibitor cocktail) and passed through a 26G needle. After incubation for 30 min on ice, the cell suspension was centrifuged at 13,000 3 g for 10 min at 4 C, and the supernatant was recovered as cell lysate. 6] [47] [48]  Separation was achieved with a Gilson HPLC system, including a model 306 pump and a model 307 pump equipped with a FP-2020 Plus Jasco Fluorescence Detector and uniPoint software.
qRT-PCR and Transcriptome Analysis Total RNA was isolated according to the TRIzol Reagent protocol (Ambion) or with the RNeasy Plus Mini Kit (QIAGEN). The quality of the RNA was determined with the Agilent 2100 BioAnalyzer (RNA 6000 Nano Kit, Agilent). cDNA was synthesized from subject S2's RNA with the iScript cDNA Synthesis Kit (Bio-Rad), and the c. 1588C>T genotype was assessed by sequencing. qRT-PCR analysis (iQ SYBR Green Supermix, iCycler, Bio-Rad) of subject S4a's fibroblast RNA (TRI Reagent, Molecular Research Center), treated with DNase (TURBO DNase, Thermo Fisher), was performed with oligo-dT-amplified cDNA (Maxima Reverse Transcriptase, Thermo Fisher) with a set of primers spanning all coding regions of FLAD1 (Figure  S1 ). Threshold cycles (Ct) were normalized (DCt) to housekeeping genes HPRT (GenBank: NM_ 000194.
RNA The American Journal of Human Genetics 98, 1130-1145, June 2, 2016 1133 libraries were assessed for quality and quantity with the Agilent 2100 BioAnalyzer and the Quant-iT PicoGreen dsDNA Assay Kit (Life Technologies) and sequenced as 100 bp paired-end runs on an Illumina HiSeq 2500 platform. Sashimi plots were used for quantitative visualization of splice junctions of their alignment data in the Integrative Genomics Viewer. 50.
Lentiviral Gene Expression The wild-type (WT) sequence of isoform 2 (GenBank: NM_ 201398. 2) was obtained as a plasmid from DNASU (HsCD00039838). Shorter FLAD1 transcripts coding for FADS proteins starting at methionine 268 (Met268) or methionine 355 (Met355) were amplified from a human control cDNA and cloned into the pLenti6. 3/V5-TOPO TA Cloning Kit (Invitrogen) for production in human fibroblasts. 36 The following primers were used: 5 0 -CGCCATGAAGGGACTATTCC-3 0 (forward Met268), 5 0 -CGCCA TGCCCAACGCTGTGG-3 0 (forward Met355), and 5 0 -TCATGTG CGGGAGTTCCGCT-3 0 (reverse). Biophysical Studies of Recombinant 6His-p. Ser495del and 6His-p. Arg530Cys FADS2.
The immobilized proteins were incubated for 3 hr with a 3,000-fold dilution of a polyclonal antiserum against FADS. 13 The bound antibodies were visualized with the aid of secondary anti-rabbit IgG antibodies conjugated with alkaline phosphatase (1:3,500 dilution). Quantitative evaluation of immuno-reactive protein bands was carried out by densitometric analysis with ImageLab software 5. 1 (Bio-Rad).
Results Metabolic research centers from Australia, Austria, Denmark, England, Finland, France, Germany, Italy, and Turkey independently achieved and subsequently shared results from candidate-gene or WES analysis on families and individuals affected by riboflavin-responsive MADD and/ or a suspected mitochondrial respiratory-chain disorder (Table  1 ). As a result of this collaborative effort, from seven unrelated families (F1-F7 in Figure  1A ) we identified nine affected individuals harboring recessive mutations in one gene, FLAD1 (GenBank: NM_025207. 4), coding for FADS. Detailed case reports are available in the Supplemental Note.
We next investigated the possibility of alternative transcripts by analyzing RNA sequencing data generated from two fibroblast samples (C1 and C3) or a whole-blood RNA sample (C2). Figure  3A  summarizes in Sashimi plots all detected RNA splice variants. In both tissues, FADS2 (comprising exons 1a and 1b) represents the most abundant transcript; the next most-abundant transcript is missing exon 1a but still encodes the same cytosolic protein isoform of 54 kDa. In none of the investigated samples did we find evidence of FADS1, which covers all of exon 1 and retains intron 1a, or isoforms 3 (GenBank: NM_ 00114891) and 4 (GenBank: NM_001184892) reported in RefSeq. These observations are in agreement with the immunoblotting experiments (Figure  2 ). In control fibroblast C3, 27 out of 28 reads starting with exon 1a continued in exon 1b; however, one paired read detected a transcript variant covering exon 1a, including an ATG codon directly connected in frame to the open reading frame in exon 3 (isoform 5 in Figure  3A ).
18 nmol) FADS2. The FAD synthesis catalyzed by WT FADS2 is presented as open symbols. Mutant FADS is presented as closed symbols. The rate of FAD synthesis was measured in the presence of 100 mmol/L ATP and 5 mmol/L MgCl 2. v 0 was measured by the initial rate of fluorescence decrease (excitation at 450 nm and emission at 520 nm) and expressed in nmol FMN $ min À1 $ mg À1 mutant FADS2 (A) and as a percentage of the V max value (set arbitrarily to 100%) (B). Data points are fitted according to the Michaelis-Menten equation. MADD or ethylmalonic and/or adipic aciduria (Tables  1  and S3 ), which argues for a more general impairment in flavinylation of mitochondrial proteins under these conditions. Decreased activity of complex II combined with MADD biochemistry was also observed in a recently reported subject with riboflavin-responsive exercise intolerance due to a defect in mitochondrial FAD transport.