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University of Cincinnati

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26 articles published in JoVE

Bioengineering

Reconstituting Cytoarchitecture and Function of Human Epithelial Tissues on an Open-Top Organ-Chip Varone Antonio¹, Adya Panchal², Magdalena Kasendra³, Barrile Riccardo^2,3 ¹Merk Sharp & Dohme LLC, ²Department of Biomedical Engineering, University of Cincinnati, ³ The present protocol describes the capabilities and the essential culture modalities of the Open-Top Organ-Chip for the successful establishment and maturation of full-thickness organ-on-chip cultures of primary tissues (skin, alveolus, airway, and intestine), providing the opportunity to investigate different functional aspects of the human epithelial/mesenchymal and vascular niche interface in vitro.
Neuroscience

Low-intensity Blast Wave Model for Preclinical Assessment of Closed-head Mild Traumatic Brain Injury in Rodents Aric F. Logsdon*^1,2, Brandon P. Lucke-Wold*³, Ryan C. Turner*⁴, Sean M. Collins⁵, Evan L. Reeder⁵, Jason D. Huber⁴, Charles L. Rosen⁶, Matthew J. Robson*⁵, Florian Plattner*⁷ ¹Geriatrics Research Education and Clinical Center, Veterans Affairs, ²Division of Gerontology and Geriatric Medicine, University of Washington, ³Department of Neurosurgery, University of Florida, ⁴Department of Neurosurgery, West Virginia University, ⁵Division of Pharmaceutical Sciences, University of Cincinnati, ⁶Central Illinois Neuro Health Sciences, Bloomington, IL, ⁷Neuro-research, Dallas, TX We present here a protocol of a blast wave model for rodents to investigate neurobiological and pathophysiological effects of mild to moderate traumatic brain injury. We established a gas-driven, bench-top setup equipped with pressure sensors allowing for reliable and reproducible generation of blast-induced mild to moderate traumatic brain injury.
Biology

Isolation, Transfection, and Long-Term Culture of Adult Mouse and Rat Cardiomyocytes Perwez Alam¹, Bryan D. Maliken¹, Malina J. Ivey¹, Shannon M. Jones¹, Onur Kanisicak¹ ¹Department of Pathology and Laboratory Medicine, College of Medicine, University of Cincinnati Here, we present a protocol for the isolation, transfection, and long-term culture of adult mouse and rat cardiomyocytes.
Bioengineering

Preparation of Tunable Extracellular Matrix Microenvironments to Evaluate Schwann Cell Phenotype Specification Zhenyuan Xu¹, Jacob A. Orkwis¹, Greg M. Harris^1,2,3 ¹Department of Chemical and Environmental Engineering, University of Cincinnati, ²Department of Biomedical Engineering, University of Cincinnati, ³Neuroscience Graduate Program, University of Cincinnati College of Medicine This methodology aims to illustrate the mechanisms by which extracellular matrix cues such as substrate stiffness, protein composition and cell morphology regulate Schwann cell (SC) phenotype.
Environment

RNA Interference in Aquatic Beetles as a Powerful Tool for Manipulating Gene Expression at Specific Developmental Time Points Shubham Rathore¹, Jenni Hassert¹, Courtney M. Clark-Hachtel^2,3, Aaron Stahl^1,4, Yoshinori Tomoyasu², Elke K. Bushbeck¹ ¹Department of Biological Sciences, University of Cincinnati, ²Department of Biology, Miami University, ³Department of Biology, University of North Carolina at Chapel Hill, ⁴Department of Neuroscience, The Scripps Research Institute RNA interference is a widely applicable, powerful technique for manipulating gene expression at specific developmental stages. Here, we describe the necessary steps for implementing this technique in the aquatic diving beetle Thermonectus marmoratus, from the acquisition of gene sequences to the knockdown of genes that affect structure or behavior.
Immunology and Infection

Experimental Analysis of Apoptotic Thymocyte Engulfment by Macrophages Yuxuan Zhen¹, Wen-Hai Shao¹ ¹Division of Immunology, Allergy, and Rheumatology, Department of Internal Medicine, College of Medicine, University of Cincinnati Here, we present a protocol to prepare apoptotic thymocytes and peritoneal macrophages and analyze the efficiency of efferocytosis and the specific inhibitor-mediated blocking of apoptotic thymocytes engulfment. This protocol has a broad application in cell-mediated clearance of other particles including artificial beads and bacteria.
Neuroscience

A Bedside, Single Burr Hole Approach to Multimodality Monitoring in Severe Brain Injury Brandon Foreman¹, Daryn Cass², Jonathan Forbes², Laura B. Ngwenya^2,3 ¹Department of Neurology and Rehabilitation Medicine, University of Cincinnati, ²Department of Neurosurgery, University of Cincinnati, ³Neurotrauma Center, University of Cincinnati Gardner Neuroscience Institute A method of recording multimodality monitoring signals in patients with severe brain injuries using a bedside, single burr hole technique is described.
Developmental Biology

Wholemount In Situ Hybridization for Astyanax Embryos Heidi Luc¹, Connor Sears¹, Andrew Raczka¹, Joshua B. Gross¹ ¹Department of Biological Sciences, University of Cincinnati This protocol enables visualization of gene expression in embryonic Astyanax cavefish. This approach has been developed with the goal of maximizing gene expression signal, while minimizing non-specific background staining.
Bioengineering

Novel Process for 3D Printing Decellularized Matrices Stacey M. S. Gruber¹, Paulomi Ghosh², Karl Wilhelm Mueller², Patrick W. Whitlock^1,2,3, Chia-Ying Lin^1,3 ¹Department of Biomedical Engineering, University of Cincinnati, ² This protocol describes the production of polycaprolactone (PCL) filament with embedded polylactic acid (PLA) microspheres which contain decellularized matrices (DM) for 3D printing of structural tissue engineering constructs.
Medicine

Isolation of Primary Mouse Hepatocytes for Nascent Protein Synthesis Analysis by Non-radioactive L-azidohomoalanine Labeling Method Esam S.B. Salem*^1,2, Kazutoshi Murakami*², Toshimasa Takahashi², Elise Bernhard², Vishnupriya Borra², Mridula Bethi², Takahisa Nakamura^2,3,4 ¹Department of Pharmacology and Systems Physiology, College of Medicine, University of Cincinnati, ² Here, we present a protocol for the isolation of healthy and functional primary mouse hepatocytes. Instructions for detecting hepatic nascent protein synthesis by non-radioactive labeling substrate were provided to help understand the mechanisms underlying protein synthesis in the context of energy-metabolism homeostasis in the liver.
Medicine

Collecting Hair Samples for Hair Cortisol Analysis in African Americans Kathy D. Wright¹, Jodi L. Ford¹, Joseph Perazzo², Lenette M. Jones³, Sherrilynn Mahari⁴, Brent A. Sullenbarger^5,6,7, Mark L. Laudenslager⁸ ¹College of Nursing, The Ohio State University, ²College of Nursing, University of Cincinnati, ³School of Nursing, University of Michigan, ⁴Licensed Cosmetologist Consultant, ⁵Clinical Research Manager, The Ohio State University, ⁶Biomedical Core Research Coordinator, The Ohio State University, ⁷College of Nursing Biomedical Core Research Laboratory, The Ohio State University, ⁸Department of Psychiatry, Behavioral Immunology & Endocrinology Laboratory, University of Colorado Denver Anschutz Medical Campus Hair cortisol concentration analysis provides an alternative to traditional measures of cortisol; however, to collect hair samples from African Americans, scientists need to be culturally informed and competent. The purpose of this protocol is to demonstrate a culturally informed technique to collect hair samples for cortisol analysis from African Americans.
Immunology and Infection

Mouse- and Human-derived Primary Gastric Epithelial Monolayer Culture for the Study of Regeneration Emma Teal¹, Nina G. Steele², Jayati Chakrabarti¹, Loryn Holokai³, Yana Zavros¹ ¹Department of Pharmacology and Systems Physiology, University of Cincinnati, ²Department of Cell & Developmental Biology, University of Michigan, ³Department of Molecular Genetics, Biochemistry & Microbiology, University of Cincinnati Here we describe a protocol for establishing and culturing human- and mouse-derived 3-dimensional (3D) gastric organoids, and the method for the transfer of 3D organoids to a 2-dimensional monolayer. The use of the gastric epithelial monolayer as a novel scratch-wound assay for regeneration studies is also described.
Neuroscience

Online Transcranial Magnetic Stimulation Protocol for Measuring Cortical Physiology Associated with Response Inhibition Michael D. Guthrie¹, Donald L. Gilbert², David A. Huddleston², Ernest V. Pedapati^2,3, Paul S. Horn², Stewart H. Mostofsky⁴, Steve W. Wu² ¹College of Medicine, University of Cincinnati, ² We describe an experimental procedure to quantify excitability and inhibition of primary motor cortex during a motor response inhibition task by using Transcranial Magnetic Stimulation throughout the course of a Stop Signal Task.
Medicine

Repeated Measurement of Respiratory Muscle Activity and Ventilation in Mouse Models of Neuromuscular Disease Victoria N. Jensen*¹, Shannon H. Romer*², Sarah M. Turner³, Steven A. Crone³ ¹Neuroscience Graduate Program, University of Cincinnati, ²Department of Biological Sciences, Wright State University, ³ This paper introduces a method for repeated measurements of ventilation and respiratory muscle activity in a freely behaving amyotrophic lateral sclerosis (ALS) mouse model throughout disease progression with whole-body plethysmography and electromyography via an implanted telemetry device.
Behavior

Ultrasound Images of the Tongue: A Tutorial for Assessment and Remediation of Speech Sound Errors Jonathan L. Preston^1,2, Tara McAllister Byun³, Suzanne E. Boyce^2,4, Sarah Hamilton⁴, Mark Tiede², Emily Phillips², Ahmed Rivera-Campos⁴, Douglas H. Whalen^2,5,6 ¹Department of Communication Sciences and Disorders, Syracuse University, ²Haskins Laboratories, ³Department of Communicative Sciences and Disorders, New York University, ⁴Department of Communication Sciences and Disorders, University of Cincinnati, ⁵Program in Speech-Language-Hearing Sciences, City University of New York Graduate Center, ⁶Department of Linguistics, Yale University Ultrasound imaging can be used to display the shape and movements of the tongue in real time during speech. The images can be used to determine the nature of speech sound errors. Visual feedback of the tongue can be used to facilitate improvements in speech sound production in clinical populations.
Immunology and Infection

Developing a Salivary Antibody Multiplex Immunoassay to Measure Human Exposure to Environmental Pathogens Swinburne A. J. Augustine¹, Tarsha N. Eason², Kaneatra J. Simmons¹, Clarissa L. Curioso³, Shannon M. Griffin¹, Malini K. D. Ramudit¹, Trevor R. Plunkett⁴ ¹National Exposure Research Laboratory, U.S. Environmental Protection Agency, ²National Risk Management Research Laboratory, U.S. Environmental Protection Agency, ³Oak Ridge Institute for Science and Education, ⁴Department of Biological Sciences, McMicken College of Arts and Sciences, University of Cincinnati In the current climate of scarce resources, new technologies are emerging that allow researchers to conduct studies cheaper, faster and with more precision. Here we describe the development of a bead-based salivary antibody multiplex immunoassay to measure human exposure to multiple environmental pathogens simultaneously.
Biology

Screening for Functional Non-coding Genetic Variants Using Electrophoretic Mobility Shift Assay (EMSA) and DNA-affinity Precipitation Assay (DAPA) Daniel E. Miller¹, Zubin H. Patel^1,2,3, Xiaoming Lu^1,3, Arthur T. Lynch¹, Matthew T. Weirauch*^1,4, Leah C. Kottyan*¹ ¹Divisions of Biomedical Informatics and Developmental Biology, Cincinnati Children's Hospital We present a strategic plan and protocol for identifying non-coding genetic variants affecting transcription factor (TF) DNA binding. A detailed experimental protocol is provided for electrophoretic mobility shift assay (EMSA) and DNA affinity precipitation assay (DAPA) analysis of genotype-dependent TF DNA binding.
Biology

Genome Editing in Astyanax mexicanus Using Transcription Activator-like Effector Nucleases (TALENs) Johanna E. Kowalko¹, Li Ma², William R. Jeffery³ ¹Genetics, Development and Cell Biology, Iowa State University, ²Department of Biological Sciences, University of Cincinnati, ³Department of Biology, University of Maryland Gene-targeting mutagenesis is now possible in a wide range of organisms using genome editing techniques. Here, we demonstrate a protocol for targeted gene mutagenesis using transcription activator like effector nucleases (TALENs) in Astyanax mexicanus, a species of fish that includes surface fish and cavefish.
Medicine

A Novel Murine Model of Arteriovenous Fistula Failure: The Surgical Procedure in Detail Chun Yu Wong^1,2,3, Margreet R. de Vries^2,3, Yang Wang⁴, Joost R. van der Vorst³, Alexander L. Vahrmeijer³, Anton-Jan van Zonneveld^1,2, Jaap F. Hamming³, Prabir Roy-Chaudhury⁴, Ton J. Rabelink^1,2, Paul H. A. Quax^2,3, Joris I. Rotmans^1,2 ¹Department of Nephrology, Leiden University Medical Center, ²Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, ³Department of Surgery, Leiden University Medical Center, ⁴Division of Nephrology, University of Cincinnati Here we present a murine model of arteriovenous fistula (AVF) failure in which a clinically relevant anastomotic configuration is incorporated. This model can be used to study the pathophysiology and to test possible therapeutic interventions.
Medicine

Quantification of the Immunosuppressant Tacrolimus on Dried Blood Spots Using LC-MS/MS Touraj Shokati¹, Nicholas Bodenberger¹, Holly Gadpaille¹, Björn Schniedewind¹, Alexander A. Vinks², Wenlei Jiang³, Rita R. Alloway⁴, Uwe Christians¹ ¹iC42 Clinical Research and Development, University of Colorado, Anschutz Medical Campus, ² Here we describe a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) assay to quantify the immunosuppressant tacrolimus in dried blood spots using a simple manual protein precipitation step and online column extraction.
Engineering

Phase Diagram Characterization Using Magnetic Beads as Liquid Carriers Nicholas Blumenschein¹, Daewoo Han¹, Andrew J. Steckl¹ ¹Department of Electrical Engineering and Computing Systems, University of Cincinnati Here, we present a protocol to investigate multi-component phase diagrams using externally controlled magnetic beads as liquid carriers in a lab-in-tube approach. This approach can aid in applications that seek to gather further information on phase change in complex liquid systems.
Behavior

Vision Training Methods for Sports Concussion Mitigation and Management Joseph F. Clark¹, Angelo Colosimo², James K. Ellis³, Robert Mangine³, Benjamin Bixenmann⁴, Kimberly Hasselfeld², Patricia Graman⁵, Hagar Elgendy², Gregory Myer⁶, Jon Divine² ¹Neurology and Rehabilitative Medicine, University of Cincinnati, ²Division of Sports Medicine, Department of Orthopaedic Surgery, University of Cincinnati, ³Department of Athletics, University of Cincinnati, ⁴Department of Neurosurgery, University of Cincinnati, ⁵College of Education, Criminal Justice, and Human Services, University of Cincinnati, ⁶ This paper describes a protocol to conduct, quantitatively monitor, and assess the success of vision training initiated as part of a sports medical management program including intervention for concussion prevention and performance enhancement.
Neuroscience

Olfactory Assays for Mouse Models of Neurodegenerative Disease Andrew M. Lehmkuhl¹, Emily R. Dirr^2,3, Sheila M. Fleming^1,2 ¹Department of Psychology, University of Cincinnati, ²Department of Neurology, University of Cincinnati, ³Department of Neuroscience, Cell Biology, and Physiology, Wright State University Impairment in olfactory function is a common feature in many neurodegenerative disorders including Parkinson, Alzheimer, and Huntington diseases. In the present article, we describe a set of tests for assessing olfaction discrimination and detection in mice that can be used to measure olfactory abilities in mouse models of neurodegenerative diseases.
Behavior

Assessment of Sensorimotor Function in Mouse Models of Parkinson's Disease Sheila M. Fleming^1,2, Osunde R. Ekhator^1,2, Valentins Ghisays¹ ¹Department of Psychology, University of Cincinnati, ²Department of Neurology, University of Cincinnati In Parkinson's disease and movement disorders in general, sensitive and reliable behavioral assays are essential for testing novel potential therapeutics. Here, we describe a manageable battery of sensorimotor tests for mice that are sensitive to varying degrees of injury to the nigrostriatal system and useful for preclinical studies.
Biology

Isolation and Primary Culture of Rat Hepatic Cells Ling Shen¹, Allix Hillebrand², David Q.-H. Wang³, Min Liu¹ ¹Department of Pathology and Laboratory Medicine, University of Cincinnati College of Medicine, ²American University in Washington, D.C., ³Department of Internal Medicine, Saint Louis University School of Medicine Primary hepatocytes provide a valuable tool to evaluate biochemical, molecular, and metabolic functions in a physiologically relevant experimental system. We describe a reliable protocol for rat in situ liver perfusion, which consistently generates viable hepatocytes up to 1.0 × 10⁸ cells per preparation with cell viability between 88 ~ 96%.
Immunology and Infection

Obtaining Highly Purified Toxoplasma gondii Oocysts by a Discontinuous Cesium Chloride Gradient Sarah E. Staggs¹, Mary Jean See², J P. Dubey³, Eric N. Villegas^2,4 ¹Dynamac, Inc., ²Department of Biological Sciences, University of Cincinnati, McMicken College of Arts and Science, ³Animal Parasitic Disease Laboratory, Agricultural Research Service, U.S. Department of Agriculture, ⁴National Exposure Research Laboratory, US Environmental Protection Agency This study describes the development of a modified CsCl method that easily purifies T. gondii oocysts from feces of infected cats that are suitable for molecular biological and tissue culture manipulation

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