Tulane University View Institution's Website 20 articles published in JoVE Bioengineering Preparation and Structural Evaluation of Epithelial Cell Monolayers in a Physiologically Sized Microfluidic Culture Device Eshan B. Damle1, Eiichiro Yamaguchi1, Joshua E. Yao1, Donald P. Gaver III1 1Department of Biomedical Engineering, School of Science and Engineering, Tulane University The presented protocol describes the development and use of a phalloidin-based filamentous-actin staining technique with confocal laser scanning microscopy (CLSM) to visualize adherent cell layer structure in microfluidic dynamic-culture channels and traditional fixed-well static-culture chambers. This approach aids in evaluating cell layer confluency, monolayer formation, and layer-thickness uniformity. Cancer Research Modeling Breast Cancer in Human Breast Tissue using a Microphysiological System Loren M. Brown1, Katherine L. Hebert2, Rakesh R. Gurrala3, C. Ethan Byrne4, Matthew Burow5, Elizabeth C. Martin4, Frank H. Lau1 1Department of Surgery, Louisiana State University Health Sciences Center, 2Department of Bioinnovation, Tulane University, 3Tulane University School of Medicine, 4Department of Biological and Agricultural Engineering, Louisiana State University, 5Department of Medicine, Tulane University School of Medicine This protocol describes the construction of an in vitro microphysiological system for studying breast cancer using primary human breast tissue with off the shelf materials. Behavior A Novel Pavlovian Fear Conditioning Paradigm to Study Freezing and Flight Behavior Chandrashekhar D. Borkar1,2, Jonathan P. Fadok1,2 1Department of Psychology, Tulane University, 2Tulane Brain Institute, Tulane University Defensive behavioral responses are contingent upon threat intensity, proximity, and context of exposure. Based on these factors, we developed a classical conditioning paradigm that elicits clear transitions between conditioned freezing and flight behavior within individual subjects. This model is crucial for the understanding the pathologies involved in anxiety, panic, and post-traumatic stress disorders. Medicine A Rat Model of Pressure Overload Induced Moderate Remodeling and Systolic Dysfunction as Opposed to Overt Systolic Heart Failure Antoine H. Chaanine1, L. Gabriel Navar2, Patrice Delafontaine1,2,3 1Department of Medicine/Heart and Vascular Institute, Tulane University, 2Department of Physiology, Tulane University, 3Department of Pharmacology, Tulane University We describe the creation of a rat model of pressure overload induced moderate remodeling and early systolic dysfunction where signal transduction pathways involved in the initiation of the remodeling process are activated. This animal model will aid in identifying molecular targets for applying early therapeutic anti-remodeling strategies for heart failure. Bioengineering Biaxial Basal Tone and Passive Testing of the Murine Reproductive System Using a Pressure Myograph Shelby E. White1, Cassandra K. Conway1, Gabrielle L. Clark1, Dylan J. Lawrence1, Carolyn L. Bayer1, Kristin S. Miller1 1Department of Biomedical Engineering, Tulane University This protocol utilized a commercially available pressure myograph system to perform pressure myograph testing on the murine vagina and cervix. Utilizing media with and without calcium, the contributions of the smooth muscle cells (SMC) basal tone and passive extracellular matrix (ECM) were isolated for the organs under estimated physiological conditions. Immunology and Infection Phenotypic Analysis of Rodent Malaria Parasite Asexual and Sexual Blood Stages and Mosquito Stages Ahmed S.I. Aly1,2, Gozde Deveci1, Ilknur Yilmaz1, Amanah Abraham2, Aneesa Golshan2, Robert J. Hart2 1Beykoz Institute of Life Sciences and Biotechnology, Bezmialem Vakif University, Istanbul, Turkey, 2School of Public Health and Tropical Medicine, Department of Tropical Medicine, Tulane University Due to the striking similarities of the life cycle and biology of rodent malaria parasites to human malaria parasites, rodent malaria models have become indispensable for malaria research. Herein, we standardized some of the most important techniques used in the phenotypic analysis of wild-type and transgenic rodent malaria species. Genetics RNA Next-Generation Sequencing and a Bioinformatics Pipeline to Identify Expressed LINE-1s at the Locus-Specific Level Tiffany Kaul1, Maria E. Morales1, Emily Smither1, Melody Baddoo1,2, Victoria P. Belancio1,3, Prescott Deininger1,4 1Tulane Cancer Center, Tulane University, 2Department of Pathology, Tulane University, 3Department of Structural and Cellular Biology, Tulane University, 4Department of Epidemiology, Tulane University Here, we present a bioinformatic approach and analyses to identify LINE-1 expression at the locus specific level. Chemistry Characterization of Synthetic Polymers via Matrix Assisted Laser Desorption Ionization Time of Flight (MALDI-TOF) Mass Spectrometry Molly E. Payne1, Scott M. Grayson1 1Department of Chemistry, Tulane University A protocol for the matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) characterization of synthetic polymers is described including the optimization of sample preparation, spectral acquisition, and data analysis. Bioengineering A Microcontroller Operated Device for the Generation of Liquid Extracts from Conventional Cigarette Smoke and Electronic Cigarette Aerosol Chastain A. Anderson1, Rachael E. Bokota1, Andrew E. Majeste1, Walter L. Murfee2, Shusheng Wang3 1Cell & Molecular Biology, Tulane University, 2Biomedical Engineering, Tulane University, 3Cell & Molecular Biology & Ophthalmology, Tulane University Here, we describe a programmable laboratory device that can be used to create extracts of conventional cigarette smoke and electronic cigarette aerosol. This method provides a useful tool for making direct comparisons between conventional cigarettes and electronic cigarettes, and is an accessible entry point into electronic cigarette research. Developmental Biology An Ex Vivo Method for Time-Lapse Imaging of Cultured Rat Mesenteric Microvascular Networks Mohammad S. Azimi1, Jessica M. Motherwell1, Walter L. Murfee1 1Biomedical Engineering, Tulane University Angiogenesis involves multi-cell, multi-system interactions that need to be investigated in a physiologically relevant environment. The objective of this study is to demonstrate the ability of the rat mesentery culture model to make time-lapse comparisons of intact microvascular networks during angiogenesis. Medicine An Alkali-burn Injury Model of Corneal Neovascularization in the Mouse Chastain Anderson*1, Qinbo Zhou*1, Shusheng Wang1,2 1Department of Cell and Molecular Biology, Tulane University, 2Department of Ophthalmology, Tulane University Neovascularization (NV) of the cornea can complicate multiple visual pathologies. Utilizing a controlled, alkali-burn injury model, a quantifiable level of corneal NV can be produced for mechanistic study of corneal NV and evaluation of potential therapies for neovascular disorders. Bioengineering Nonhuman Primate Lung Decellularization and Recellularization Using a Specialized Large-organ Bioreactor Ryan W. Bonvillain1,2, Michelle E. Scarritt1, Nicholas C. Pashos1, Jacques P. Mayeux1, Christopher L. Meshberger1, Aline M. Betancourt1,3, Deborah E. Sullivan1,3, Bruce A. Bunnell1,2,4 1Center for Stem Cell Research and Regenerative Medicine, Tulane University School of Medicine, 2Division of Regenerative Medicine, Tulane National Primate Research Center, 3Department of Microbiology and Immunology, Tulane University School of Medicine, 4Department of Pharmacology, Tulane University School of Medicine Whole-organ decellularization produces natural biological scaffolds that may be used for regenerative medicine. The description of a nonhuman primate model of lung regeneration in which whole lungs are decellularized and then seeded with adult stem cells and endothelial cells in a bioreactor that facilitates vascular circulation and liquid media ventilation is presented. Immunology and Infection Feeding of Ticks on Animals for Transmission and Xenodiagnosis in Lyme Disease Research Monica E. Embers1, Britton J. Grasperge1, Mary B. Jacobs1, Mario T. Philipp1 1Division of Bacteriology & Parasitology, Tulane National Primate Research Center, Tulane University Health Sciences Center Lyme disease is the most commonly-reported vector-borne disease in North America. The causative agent, Borrelia burgdorferi is a spirochete bacterium transmitted by Ixodid ticks. Transmission and detection of infection in animal models is optimized by the use of tick feeding, which we describe here. Biology Rat Mesentery Exteriorization: A Model for Investigating the Cellular Dynamics Involved in Angiogenesis Ming Yang1, Peter C. Stapor1, Shayn M. Peirce2, Aline M. Betancourt3, Walter L. Murfee1 1Department of Biomedical Engineering, Tulane University, 2Department of Biomedical Engineering, University of Virginia, 3Center for Stem Cell Research and Regenerative Medicine, Tulane University This article describes a simple model for stimulating angiogenesis in the rat mesentery. The model produces dramatic increases in capillary sprouting, vascular area and vascular density over a relatively short time course in a tissue that allows en face visualization of entire microvascular networks down to the single cell level. Neuroscience The Mouse Forced Swim Test Adem Can1, David T. Dao2, Michal Arad1, Chantelle E. Terrillion3, Sean C. Piantadosi1, Todd D. Gould1,3,4 1Department of Psychiatry, University of Maryland School of Medicine, 2Tulane University School of Medicine, 3Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine, 4The Program in Neuroscience, University of Maryland The forced swim test is validated as an experimental approach to assess potential antidepressant efficacy in rodents. Experimental animals are placed in a tank of water and escape-related mobility behavior is quantified. The common procedures for the mouse version of this test are described. Neuroscience The Tail Suspension Test Adem Can*1, David T. Dao*1,2, Chantelle E. Terrillion3, Sean C. Piantadosi1, Shambhu Bhat1, Todd D. Gould1,3,4 1Department of Psychiatry, University of Maryland School of Medicine, 2Tulane University School of Medicine, 3The Program in Neuroscience, University of Maryland, 4Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine The tail-suspension test is validated as an experimental procedure to assess antidepressant efficacy of drug treatments in mice. Mice are suspended by their tails for six minutes and escape-related behaviors are assessed. We describe procedures used in conducting the tail suspension test. Bioengineering Rotating Cell Culture Systems for Human Cell Culture: Human Trophoblast Cells as a Model Kevin J. Zwezdaryk*1, Jessica A. Warner*1,2, Heather L. Machado3, Cindy A. Morris1, Kerstin Höner zu Bentrup1 1Department of Microbiology and Immunology, Tulane University Medical School, 2Physician/Scientist Program, Tulane University Medical School, 3Department of Molecular and Cellular Biology, Baylor College of Medicine Traditional, two dimensional cell culture techniques often result in altered characteristics with respect to differentiation markers, cytokines and growth factors. Three-dimensional cell culture in the rotating cell culture system (RCCS) reestablishes expression of many of these factors as shown here with an extravillous trophoblast cell line. Immunology and Infection Recognition of Epidermal Transglutaminase by IgA and Tissue Transglutaminase 2 Antibodies in a Rare Case of Rhesus Dermatitis Karol Sestak1, Kaushiki Mazumdar1, Cecily C. Midkiff2, Jason Dufour3, Juan T. Borda2, Xavier Alvarez2 1Division of Microbiology, Tulane National Primate Research Center, 2Division of Comparative Pathology, Tulane National Primate Research Center, 3Division of Veterinary Medicine, Tulane National Primate Research Center Dermatitis herpetiformis (DH) is a chronic inflammatory condition characterized by an autoimmune reaction between IgA and epidermal transglutaminase (eTG). DH develops in a very small portion of gluten-sensitive and/or celiac patients. The results of this study indicate that DH can also develop in a rhesus monkey host with symptoms of idiopatic dermatitis. Bioengineering Fabrication of Micropatterned Hydrogels for Neural Culture Systems using Dynamic Mask Projection Photolithography J. Lowry Curley1, Scott R. Jennings1, Michael J. Moore1 1Biomedical Engineering, Tulane University Simple techniques are described for the rapid production of microfabricated neural culture systems using a digital micromirror device for dynamic mask projection lithography on regular cell culture substrates. These culture systems may be more representative of natural biological architecture, and the techniques described could be adapted for numerous applications. Medicine An Orthotopic Model of Murine Bladder Cancer Georgina L. Dobek1, W. T. Godbey2 1Department of Comparative Medicine, Tulane University, 2Department of Chemical and Biomolecular Engineering, Tulane University Bladder tumors are established in female mice in a minimally invasive fashion through catheterization, local cauterization, and subsequent adhesion of carcinoma cells to the burn sites.