University of Sao Paulo 28 articles published in JoVE Medicine Randomized Controlled Trial to Study the Acute Effects of Strength Exercise on Insulin Sensitivity in Obese Adults Luis Filipe Rocha Silva1, Bruna Caroline Chaves Garcia2, Zach A. Mang3, Fabiano Trigueiro Amorim4, Marco Fabrício Dias-Peixoto1, Fernando Gripp1, Valmor Tricoli5, Flávio de Castro Magalhães1,4 1Department of Physical Education, Federal University of the Jequitinhonha and Mucuri Valleys, 2Laboratory of Exercise Biology and Immunometabolism, Centro Integrado de Pós-Graduação e Pesquisa em Saúde, Programa Multicêntrico de Pós-Graduação em Ciências Fisiológicas, Federal University of the Jequitinhonha and Mucuri Valleys, 3Occupational Safety & Health, Los Alamos National Laboratory, 4Department of Health, Exercise, and Sports Sciences, University of New Mexico, 5School of Physical Education and Sport, University of Sao Paulo This study describes a randomized controlled trial protocol aiming at assessing the acute effects of strength exercise volume on insulin sensitivity in obese individuals. Behavior Exploring Life History Choices: Using Temperature and Substrate Type as Interacting Factors for Blowfly Larval and Female Preferences Vanessa A. S. Cunha*1, Sophie Tandonnet*1, Diniz Lima Ferreira1, Andre V. Rodrigues2, Tatiana Teixeira Torres1 1Department of Genetics and Evolutionary Biology, Institute of Biosciences, University of São Paulo, 2Department of Zoology, Institute of Biosciences, University of São Paulo Herein, two protocols for assessing food source and oviposition preferences in larvae and females of blowflies are detailed. These comprise four choices with two interacting factors: substrate type and temperature. The assays enable the determination of the food source preference of the larvae and the oviposition site preference for the females. Medicine Erosion Identification in Metacarpophalangeal Joints in Rheumatoid Arthritis using High-Resolution Peripheral Quantitative Computed Tomography Yousif Al-Khoury1,2, Stephanie Finzel3, Camille Figueiredo4, Andrew J. Burghardt5, Kathryn S. Stok6, Lai-Shan Tam7, Isaac Cheng7, Justin J. Tse1, Sarah L. Manske1 1Department of Radiology, McCaig Institute for Bone and Joint Health, University of Calgary, 2Department of Biomedical Engineering, University of Calgary, 3University Medical Center Freiburg, 4Department of Rheumatology, University of São Paulo, 5Department of Radiology and Biomedical Imaging, University of California, San Francisco, 6Department of Biomedical Engineering, University of Melbourne, 7Department of Medicine and Therapeutics, The Chinese University of Hong Kong Bone erosions are an important pathological feature of rheumatoid arthritis. The purpose of this work is to introduce a training tool to provide users with guidance on identifying pathological cortical breaks on high resolution peripheral quantitative computed tomography images for erosion analysis. Medicine Robotic Myotomy and Partial Fundoplication for Achalasia Rubens Antonio Aissar Sallum1, Felipe Alexandre Fernandes1, Leonardo Torres Branco1, Luna Serena Arguelho Pereira1, Camila Maria Arruda Vilanova de Câmara1, Ítalo Beltrão Pereira Simões1, José Donizeti de Meira Junior1, Melissa Mello Mazepa1, Leonardo Ervolino Corbi1, Rodrigo Nicida Garcia1, Flávio Roberto Takeda1 1Gastroenterology Department, University of São Paulo Surgical myotomy with a partial fundoplication may be used in selected patients as a definitive treatment for achalasia. This article provides a step-by-step description of a robotic myotomy and partial fundoplication in a 32-year-old patient with megaesophagus. Developmental Biology AMEBaS: Automatic Midline Extraction and Background Subtraction of Ratiometric Fluorescence Time-Lapses of Polarized Single Cells Rafael Badain1, Daniel S. C. Damineli1,2, Maria Teresa Portes3, José Feijó4, Stefano Buratti5, Giorgia Tortora6, Hugo Neves de Oliveira1, Roberto M. Cesar Jr1 1Institute of Mathematics and Statistics, University of São Paulo, 2Center for Mathematics, Computing and Cognition, Federal University of ABC, 3Department of Botany, Institute of Biosciences, University of São Paulo, 4Cell Biology and Molecular Genetics Department, University of Maryland, 5Department of Biosciences, University of Milan, 6Department of Physics, Politecnico di Milano Current methods for analyzing the intracellular dynamics of polarized single cells are often manual and lack standardization. This manuscript introduces a novel image analysis pipeline for automating midline extraction of single polarized cells and quantifying spatiotemporal behavior from time lapses in a user-friendly online interface. Biology Collection and Long-Term Maintenance of Leaf-Cutting Ants (Atta) in Laboratory Conditions Bianca Raissa Nogueira1, Amanda Aparecida de Oliveira1, Janiele Pereira da Silva2, Odair Correa Bueno1 1Institute of Biosciences, Department of General and Applied Biology, Laboratory of Leaf-cutting Ants, São Paulo State University, 2Institute of Psychology, Laboratory Walter Hugo Cunha, São Paulo University Here, a protocol is described to successfully collect and maintain healthy Atta (Hymenoptera: Formicidae) ant colonies in laboratory conditions. Additionally, different nest types and configurations are detailed together with possible experimental procedures. Biochemistry Evaluation of Substrate Ubiquitylation by E3 Ubiquitin-ligase in Mammalian Cell Lysates Patrícia M. S. dos Passos*1, Valentine Spagnol*2, Camila R.S.T.B de Correia1, Felipe R. Teixeira1,2 1Department of Genetic and Evolution, Federal University of São Carlos, 2Department of Biochemistry and Immunology, Faculty of Medicine of Ribeirão Preto, University of São Paulo We provide a detailed protocol for a ubiquitylation assay of a specific substrate and an E3 ubiquitin-ligase in mammalian cells. HEK293T cell lines were used for protein overexpression, the polyubiquitylated substrate was purified from cell lysates by immunoprecipitation, and resolved in SDS-PAGE. Immunoblotting was used to visualize this post-translational modification. Genetics Sample Preparation to Bioinformatics Analysis of DNA Methylation: Association Strategy for Obesity and Related Trait Studies Natália Yumi Noronha*1, Guilherme da Silva Rodrigues*1, Marcela Augusta de Souza Pinhel1, Jean-Baptiste Cazier2,3, Lígia Moriguchi Watanabe1, Albert Nobre Menezes4, Carlos Roberto Bueno5, Carolina Ferreira Nicoletti1, Bruno Affonso Parenti de Oliveira1, Isabelle Mello Schineider6, Isabella Harumi Yonehara Noma7, Igor Caetano Dias Alcarás8, Fernando Barbosa9, Carla Barbosa Nonino6 1Department of Internal Medicine, Ribeirão Preto Medical School, University of São Paulo, 2Centre for Computational Biology, University of Birmingham, 3Institute of Cancer and Genomic Sciences, College of Medical and Dental Sciences, University of Birmingham, 4Cancer Genetics and Evolution Laboratory, Cancer Research UK, Institute of Genetics & Molecular Medicine, The University of Edinburgh, 5Ribeirão Preto School of Physical Education and Sport, University of São Paulo, 6Health Sciences Department, Ribeirao Preto Medical School, University of Sao Paulo, 7Faculty of Pharmaceutical Sciences, University of São Paulo, 8Department of Genetics, Ribeirão Preto Medical School, University of São Paulo, 9Department of Clinical, Bromatological and Toxicological Analysis, Faculty of Pharmaceutical Sciences of Ribeirao Preto, University of São Paulo The present study describes the workflow to manage DNA methylation data obtained by microarray technologies. The protocol demonstrates steps from sample preparation to data analysis. All procedures are described in detail, and the video shows the significant steps. Immunology and Infection High-Throughput Transcriptome Analysis for Investigating Host-Pathogen Interactions André Nicolau Aquime Gonçalves1,2, Vanessa Escolano Maso3, Ícaro Maia Santos de Castro2,3, Amanda Pereira Vasconcelos3, Rodrigo Luiz Tomio Ogava2,3, Helder I Nakaya2,3,4 1Laboratory of Pathology of Infectious Diseases, Department of Pathology, Medical School, University of São Paulo, 2Scientific Platform Pasteur USP, 3Department of Clinical and Toxicological Analyses, School of Pharmaceutical Sciences, University of São Paulo, 4Hospital Israelita Albert Einstein The protocol presented here describes a complete pipeline to analyze RNA-sequencing transcriptome data from raw reads to functional analysis, including quality control and preprocessing steps to advanced statistical analytical approaches. Biology Double-Staining Method to Detect Pectin in Plant-Fungus Interaction João Paulo Rodrigues Marques1, Laura Galvan Nuevo2 1Faculty of Animal Science and Food Engineering, Department of Basic Sciences, University of São Paulo, 2Laboratory of Nuclear Instrumentation, Center of Nuclear Energy in Agriculture, University of São Paulo This protocol describes a microscopical method to detect pectin in coffee-fungus interaction. Genetics Detection of Post-Replicative Gaps Accumulation and Repair in Human Cells Using the DNA Fiber Assay Davi J. Martins*1, Stephanie Tirman*2, Annabel Quinet2, Carlos F. M. Menck1 1Department of Microbiology, Institute of Biomedical Sciences, University of São Paulo, 2Division of Oncology, Department of Internal Medicine, Washington University in St. Louis Here we describe two modifications of the DNA fiber assay to investigate single-stranded DNA gaps in replicating DNA after lesion induction. The S1 fiber assay enables the detection of post-replicative gaps using the ssDNA-specific S1 endonuclease, while the gap-filling assay allows visualization and quantification of gap repair. Immunology and Infection High-throughput Antiviral Assays to Screen for Inhibitors of Zika Virus Replication Rafaela S. Fernandes1, Gabriela D. Noske1, Victor O. Gawriljuk1, Ketllyn I. Z. de Oliveira1, Andre S. Godoy1, Nathalya C. M. R. Mesquita1, Glaucius Oliva1 1São Carlos Institute of Physics, University of Sao Paulo In this work, we describe the protocols used in replicon-based and viral enzyme-based assays to screen for inhibitors of Zika virus replication in a high-throughput screening format. Neuroscience The 6-hydroxydopamine Rat Model of Parkinson's Disease Rayanne Poletti Guimarães1, Danilo Leandro Ribeiro1, Keila Bariotto dos Santos1, Lívea Dornela Godoy1, Mirella Rosine Corrêa1, Fernando Eduardo Padovan-Neto1 1Department of Psychology, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto, University of São Paulo The 6-hydroxydopamine (6-OHDA) model has been used for decades to advance the understanding of Parkinson's Disease. In this protocol, we demonstrate how to perform unilateral nigrostriatal lesions in the rat by injecting 6-OHDA in the medial forebrain bundle, assess motor deficits, and predict lesions using the stepping test. Biology Rating L-DOPA-Induced Dyskinesias in the Unilaterally 6-OHDA-Lesioned Rat Model of Parkinson's Disease Keila Bariotto-dos-Santos*1, Danilo Leandro Ribeiro*1, Rayanne Poletti Guimarães1, Fernando Eduardo Padovan-Neto1 1Department of Psychology, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto, University of São Paulo Rodent models of L-DOPA-induced dyskinesias are invaluable tools to identify therapeutic interventions to attenuate the development or alleviate the manifestations that emerge due to the repeated administration of L-DOPA. This protocol demonstrates how to induce and analyze dyskinetic-like movements in the unilaterally 6-OHDA-lesioned rat model of Parkinson's disease. Medicine Low-Cost Single-Port (LoCoSP) Device for a Transcervical Approach in Minimally Invasive Transhiatal Esophagectomy Flavio Roberto Takeda1, Rubens Antonio Aissar Sallum1, Felipe Alexandre Fernandes1, Ivan Cecconello1 1Gastroenterology Department, University of São Paulo Here, we describe a step-by-step description of transhiatal esophagectomy and the development of a low-cost single-port device for a transcervical approach in a minimally invasive transhiatal esophagectomy. Immunology and Infection Double Labeling Immunofluorescence using Antibodies from the Same Species to Study Host-Pathogen Interactions Camila Gachet-Castro*1, Lays Adrianne Mendonça Trajano-Silva*1, Munira Muhammad Abdel Baqui1 1Laboratory of Cell and Molecular Biology of Trypanosomatids, Department of Cellular and Molecular Biology and Pathogenic Bioagents, Ribeirao Preto Medical School, University of Sao Paulo, Ribeirao Preto, SP, Brazil Here, the protocol describes how to perform double labeling immunofluorescence using primary antibodies raised in the same species to study host-pathogen interactions. Also, it can include the third antibody from a different host in this protocol. This approach can be made in any cell type and pathogens. Medicine Acute Kidney Injury Model Induced by Cisplatin in Adult Zebrafish Camila Morales Fénero*1, Barbara Nunes Padovani*1, Mariana Abrantes do Amaral*1,2, Guilherme José Bottura de Barros1, Izabella Karina Xavier de Oliveira1, Meire Ioshie Hiyane1, Niels Olsen Saraiva Camâra1,2 1Department of Immunology, University of São Paulo, 2Department of Medicine, Nephrology Division, Federal University of São Paulo This protocol describes the procedures to induce Acute Kidney Injury (AKI) in adult zebrafish using cisplatin as a nephrotoxic agent. We detailed the steps to evaluate the reproducibility of the technique and two techniques to analyze inflammation and cell death in the renal tissue, flow cytometry and TUNEL, respectively. Biology Systematic Approach to Identify Novel Antimicrobial and Antibiofilm Molecules from Plants' Extracts and Fractions to Prevent Dental Caries Sabrina M. Ribeiro*1, Érick D. O. Fratucelli*1, Júlia M. Fernandes*2, Paula C. P. Bueno2,3, Alberto José Cavalheiro2, Marlise I. Klein1 1Department of Dental Materials and Prosthodontics, School of Dentistry, São Paulo State University (UNESP), 2Department of Organic Chemistry, Institute of Chemistry, São Paulo State University (UNESP), 3Faculty of Pharmaceutical Sciences of Ribeiraão Preto (FCFRP-USP), Department of Physics and Chemistry, University of São Paulo Natural products represent promising starting points for the development of new drugs and therapeutic agents. However, due to the high chemical diversity, finding new therapeutic compounds from plants is a challenging and time-consuming task. We describe a simplified approach to identify antimicrobial and antibiofilm molecules from plant extracts and fractions. Biochemistry Utilization of Grafix for the Detection of Transient Interactors of Saccharomyces cerevisiae Spliceosome Subcomplexes Felipe A. Carvalho1, Moises R. A. Barros1, Thierry P. F. Girotto1, M. Griselda Perona1, Carla C. Oliveira1 1Department of Biochemistry, Institute of Chemistry, University of Sao Paulo Here, we describe the utilization of Grafix (Gradient Fixation), a glycerol gradient centrifugation in the presence of a crosslinker, to identify interactions between splicing factors that bind transiently to the spliceosome complex. Immunology and Infection In Vivo Infection with Leishmania amazonensis to Evaluate Parasite Virulence in Mice Juliana Ide Aoki1, Ahyun Hong1, Ricardo Andrade Zampieri1, Lucile Maria Floeter-Winter1, Maria Fernanda Laranjeira-Silva1 1Department of Physiology, Institute of Bioscience, University of Sao Paulo Here, we present a compiled protocol to evaluate the cutaneous infection of mice with Leishmania amazonensis. This is a reliable method for studying parasite virulence, allowing a systemic view of the vertebrate host response to the infection. Neuroscience Short-Term Free-Floating Slice Cultures from the Adult Human Brain Artur Fernandes1,2, Niele Dias Mendes1,3, Glaucia Maria Almeida1, Giovanna Orlovski Nogueira1, Carla de Moraes Machado4, Jose de Anchieta de Castro Horta-Junior4, João Alberto Assirati Junior5, Norberto Garcia-Cairasco2, Luciano Neder3, Adriano Sebollela1 1Department of Biochemistry and Immunology, Ribeirão Preto Medical School, University of São Paulo, 2Department of Physiology, Ribeirão Preto Medical School, University of São Paulo, 3Department of Pathology and Forensic Medicine, Ribeirão Preto Medical School, University of São Paulo, 4Department of Anatomy, Institute of Biosciences, São Paulo State University, 5Clinical Hospital at the Ribeirão Preto Medical School, University of São Paulo A protocol to prepare free-floating slice cultures from adult human brain is presented. The protocol is a variation of the widely used slice culture method using membrane inserts. It is simple, cost-effective, and recommended for running short-term assays aimed to unravel mechanisms of neurodegeneration behind age-associated brain diseases. Immunology and Infection Quantification of three DNA Lesions by Mass Spectrometry and Assessment of Their Levels in Tissues of Mice Exposed to Ambient Fine Particulate Matter Tiago Franco de Oliveira*1,2, Antonio Anax Falcão de Oliveira*1, Miriam Lemos3, Mariana Veras3, Paulo Hilário Nascimento Saldiva3,4, Marisa Helena Gennari de Medeiros5, Paolo Di Mascio5, Ana Paula de Melo Loureiro1 1Departamento de Análises Clínicas e Toxicológicas, Faculdade de Ciências Farmacêuticas, Universidade de São Paulo, 2Departamento de Farmacociências, Universidade Federal de Ciências da Saúde de Porto Alegre, 3Laboratório de Poluição Atmosfêrica Experimental - LIM05, Hospital das Clínicas, Faculdade de Medicina, Universidade de São Paulo, 4Instituto de Estudos Avançados, Universidade de São Paulo, 5Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo We describe here methods for sensitive and accurate quantification of the lesions 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), 1,N6-etheno-2'-deoxyadenosine (1,N6-dAdo) and 1,N2-etheno-2'-deoxyguanosine (1,N2-dGuo) in DNA. The methods were applied to the assessment of the effects of ambient fine particulate matter (PM2.5) in tissues (lung, liver and kidney) of exposed A/J mice. Neuroscience Real-time Video Projection in an MRI for Characterization of Neural Correlates Associated with Mirror Therapy for Phantom Limb Pain Faddi G. Saleh Velez1,2, Camila B. Pinto1,3, Emma S. Bailin4, Marionna Münger1, Andrew Ellison5, Beatriz T. Costa1, David Crandell6, Nadia Bolognini7,8, Lotfi B. Merabet4, Felipe Fregni1 1Laboratory of Neuromodulation & Center for Clinical Research Learning, Department of Physical Medicine and Rehabilitation, Harvard Medical School, Spaulding Rehabilitation Hospital, 2University of Chicago Medical Center, Department of Neurology, University of Chicago, 3Department of Neuroscience and Behavior, Psychology Institute, University of Sao Paulo, 4The Laboratory for Visual Neuroplasticity, Department of Ophthalmology, Massachusetts Eye and Ear Infirmary, Harvard Medical School, 5Center for Biomedical Imaging, Department of Anatomy and Neurobiology, Boston University School of Medicine, 6Spaulding Rehabilitation Hospital, Harvard Medical School, 7Department of Psychology & Milan Center for Neuroscience, University of Milano-Bicocca, 8Neuropsychological Laboratory, Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS) Istituto Auxologico Italiano We present a novel combined behavioral and neuroimaging protocol employing real-time video projection for the purpose of characterizing the neural correlates associated with mirror therapy within the magnetic resonance imaging scanner environment in leg amputee subjects with phantom limb pain. Developmental Biology Derivation and Differentiation of Canine Ovarian Mesenchymal Stem Cells Amanda B.T. Hill1,2, Jonathan E.B.T. Hill1, Fabiana F. Bressan3, Maria Angélica Miglino4, Joaquim M. Garcia1 1The School of Agricultural and Veterinary Studies (FCAV), Jaboticabal, São Paulo State University (UNESP), 2CRRF, Université de Montréal, 3FZEA, University of São Paulo, 4FMVZ, University of São Paulo Herein, we describe a method for the isolation, expansion, and differentiation of mesenchymal stem cells from canine ovarian tissue. Medicine Oral Biofilm Formation on Different Materials for Dental Implants Thalisson S. O. Silva1, Alice R. Freitas1, Marília L. L. Pinheiro1, Cássio do Nascimento1, Evandro Watanabe2, Rubens F. Albuquerque1 1Department of Dental Materials and Prosthodontics, School of Dentistry of Ribeirão Preto, University of São Paulo, 2Department of Restorative Dentistry, School of Dentistry of Ribeirão Preto, University of São Paulo Here, we present a protocol to evaluate oral biofilm formation on titanium and zirconia materials for dental prosthesis abutments, including the analysis of bacterial cells viability and morphological characteristics. An in situ model associated with powerful microscopy techniques is used for the oral biofilm analysis. Neuroscience RNA-Seq Analysis of Differential Gene Expression in Electroporated Chick Embryonic Spinal Cord Felipe M. Vieceli1, C.Y. Irene Yan1 1Department of Cell and Developmental Biology, Universidade de São Paulo This video shows the basic steps for performing whole transcriptome analysis on dissected chick embryonic spinal cord samples after transfection with in ovo electroporation. Bioengineering Flying Insect Detection and Classification with Inexpensive Sensors Yanping Chen1, Adena Why2, Gustavo Batista3, Agenor Mafra-Neto4, Eamonn Keogh1 1Department of Computer Science and Engineering, University of California, Riverside, 2Department of Entomology, University of California, Riverside, 3Institute of Mathematics and Computer Sciences, University of São Paulo - USP, 4ISCA Technologies We proposed a system that uses inexpensive, noninvasive pseudo-acoustic optical sensors to automatically and accurately detect, count, and classify flying insects based on their flying sound. Environment Mass Production of Genetically Modified Aedes aegypti for Field Releases in Brazil Danilo O. Carvalho1,2, Derric Nimmo1, Neil Naish1, Andrew R. McKemey1, Pam Gray1, André B. B. Wilke3, Mauro T. Marrelli3, Jair F. Virginio4, Luke Alphey1,5, Margareth L. Capurro2,6 1Oxitec Ltd, 2Departamento de Parasitologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, 3Departamento de Epidemiologia, Universidade de São Paulo, 4Moscamed Brasil, 5Deptartment of Zoology, University of Oxford, 6Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular (INCT-EM) To achieve population suppression of Aedes aegypti using the RIDL® (Release of Insects carrying a Dominant Lethal) system, large numbers of male mosquitoes need to be released. This requires the use of mass rearing techniques and technology to provide reliable systems to obtain the maximum number of high quality male mosquitoes.
