Wayne State University School of Medicine 16 articles published in JoVE Cancer Research An Ex Vivo Model of Ovarian Cancer Peritoneal Metastasis Using Human Omentum Terrence Wong1, Roslyn Tedja2, Hussein Chehade2, Robert Morris1, Ayesha B. Alvero2, Gil Mor2 1Department of Gynecologic Oncology, Karmanos Cancer Institute, 2C.S. Mott Center for Human Growth and Development, Department of Obstetrics and Gynecology, Wayne State University School of Medicine This protocol describes the establishment of a three-dimensional (3D) ex vivo model of cancer cell-omentum interaction. The model provides a platform for elucidating pro-tumor mechanisms within the adipose niche and for testing novel therapies. Behavior Using Looming Visual Stimuli to Evaluate Mouse Vision Christina C. Koehler1, Leo M. Hall1, Chase B. Hellmer1, Tomomi Ichinose1 1Department of Ophthalmology, Visual and Anatomical Sciences, Wayne State University School of Medicine To examine mouse vision, we conducted a looming test. Mice were placed in a large square arena with a monitor on its ceiling. The looming visual stimulus consistently evoked freezing or flight reactions in the mice. Immunology and Infection An Immunohistopathologic Study to Profile the Folate Receptor Beta Macrophage and Vascular Immune Microenvironment in Giant Cell Arteritis Shirley Albano-Aluquin1, Jozef Malysz2, Michal Kidacki3, Manohar Ratnam4, Nancy J. Olsen1 1Department of Medicine, Penn State University, 2Department of Pathology, Penn State University, 3Penn State College of Medicine, 4Department of Oncology, Wayne State University School of Medicine The protocol illustrates the use of histopathologic examination and immunohistochemistry to profile the folate receptor beta macrophage and its relationship with the total immune cell infiltrate in temporal artery biopsies in giant cell arteritis. Bioengineering Comprehensive Evaluation of the Effectiveness and Safety of Placenta-Targeted Drug Delivery Using Three Complementary Methods Baozhen Zhang1, Zhilong Chen1,2, Jinyu Han1,3, Mengxia Li1, Nihar R. Nayak4, Xiujun Fan1 1Laboratory for Reproductive Health, Institute of Biomedicine and Biotechnology, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, 2College of Veterinary Medicine, Hunan Agricultural University, 3Key Laboratory of Chemical Engineering Process and Technology for High-efficiency Conversion, College of Chemistry and Material Sciences, Heilongjiang University, 4Department of Obstetrics and Gynecology, Wayne State University School of Medicine We describe a system that utilizes three methods to evaluate the safety and effectiveness of placenta-targeted drug delivery: in vivo imaging to monitor nanoparticle accumulation, high-frequency ultrasound to monitor placental and fetal development, and HPLC to quantify drug delivery to tissue. Immunology and Infection Quantifying Vibrio cholerae Colonization and Diarrhea in the Adult Zebrafish Model Dhrubajyoti Nag1, Kristie Mitchell1, Paul Breen1, Jeffrey H. Withey1 1Department of Microbiology, Immunology, and Biochemistry, Wayne State University School of Medicine Zebrafish are a natural Vibrio cholerae host and can be used to recapitulate and study the entire infectious cycle from colonization to transmission. Here, we demonstrate how to assess V. cholerae colonization levels and quantify diarrhea in zebrafish. Developmental Biology CARIP-Seq and ChIP-Seq: Methods to Identify Chromatin-Associated RNAs and Protein-DNA Interactions in Embryonic Stem Cells Benjamin L. Kidder1,2 1Department of Oncology, Wayne State University School of Medicine, 2Karmanos Cancer Institute, Wayne State University School of Medicine Here, we describe methods to perform ChIP-Seq and CARIP-Seq, including library preparation for next-generation sequencing, to generate global epigenomic and chromatin-associated RNA maps in ES cells. Neuroscience Focal Macropatch Recordings of Synaptic Currents from the Drosophila Larval Neuromuscular Junction Alexander Vasin1, Maria Bykhovskaia1,2 1Department of Neurology, School of Medicine, Wayne State University, 2Department of Anatomy and Cell Biology, School of Medicine, Wayne State University Synaptic currents can be recorded focally from visualized synaptic boutons at the Drosophila third instar larvae neuromuscular junction. This technique enables monitoring the activity of a single synaptic bouton. Medicine Strategic Endothelial Cell Tube Formation Assay: Comparing Extracellular Matrix and Growth Factor Reduced Extracellular Matrix Daniel Xie1, Donghong Ju2, Cecilia Speyer2, David Gorski2,3, Mary A. Kosir2,3 1Center for Molecular Medicine and Genetics (CMMG), Wayne State University School of Medicine, 2Department of Surgery, Wayne State University School of Medicine, 3Department of Oncology, Wayne State University School of Medicine This manuscript describes a tube formation assay to quantify the effects of a given compound or condition on angiogenesis by using endothelial cell tube formation in a controlled environment. Biology Methods to Discover Alternative Promoter Usage and Transcriptional Regulation of Murine Bcrp1 Karthika Natarajan1,2, Yi Xie1,3, Takeo Nakanishi4, Rebecca S. Moreci5,6, Pancharatnam Jeyasuria7, Arif Hussain1,3,8,9, Douglas D. Ross1,3,8,9,10,11 1Greenebaum Cancer Center, University of Maryland School of Medicine, 2Pharmaceutical Sciences, University of Maryland School of Pharmacy, 3Baltimore VA Medical Center, 4Membrane Transport and Biopharmaceutics, School of Pharmaceutical Sciences, Kanazawa University, 5Obstetrics, Gynecology and Reproductive Science, University of Pittsburgh, 6 With the murine ABC transporter Bcrp1 (Abcg2) as an example, in-silico protocols are presented to detect alternative promoter usage in genes expressed in mouse tissues, and to evaluate the functionality of the alternative promoters identified using reporter assays. Medicine Rapid Fractionation and Isolation of Whole Blood Components in Samples Obtained from a Community-based Setting Amy Weckle1,2, Allison E. Aiello3, Monica Uddin1,4, Sandro Galea5, Rebecca M. Coulborn6, Richelo Soliven7, Helen Meier6, Derek E. Wildman1,2 1Carl R. Woese Institute for Genomic Biology, University of Illinois at Urbana-Champaign, 2Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, 3Department of Epidemiology, Gillings School of Global Public Health, University of North Carolina, 4Department of Psychology, University of Illinois at Urbana-Champaign, 5Department of Epidemiology, Mailman School of Public Health, Columbia University, 6Department of Epidemiology, University of Michigan School of Public Health, 7Center for Molecular Medicine and Genetics, Wayne State University School of Medicine We outline a methodology for the processing of whole blood to obtain a variety of components for further analysis. We have optimized a streamlined protocol that enables rapid, high-throughput simultaneous processing of whole blood samples in a non-clinical setting. Immunology and Infection Isolation of Leukocytes from the Murine Tissues at the Maternal-Fetal Interface Marcia Arenas-Hernandez1, Elly N. Sanchez-Rodriguez1, Tara N. Mial1, Sarah A. Robertson2, Nardhy Gomez-Lopez1,3,4 1Department of Obstetrics & Gynecology, Wayne State University School of Medicine, 2School of Paediatrics and Reproductive Health, Research Centre for Reproductive Health, the Robinson Research Institute, The University of Adelaide, 3Department of Immunology & Microbiology, Wayne State University School of Medicine, 4Perinatology Research Branch, NICHD/NIH/DHHS Described herein is a protocol to isolate and analyze the infiltrating leukocytes of tissues at the maternal-fetal interface (uterus, decidua, and placenta) of mice. This protocol maintains the integrity of most cell surface markers and yields enough viable cells for downstream applications including flow cytometry analysis. Immunology and Infection Isolation of Leukocytes from the Human Maternal-fetal Interface Yi Xu1, Olesya Plazyo1, Roberto Romero1,2,3,4, Sonia S. Hassan1,5, Nardhy Gomez-Lopez1,5,6 1Perinatology Research Branch, NICHD/NIH/DHHS, 2Department of Obstetrics and Gynecology, University of Michigan, 3Department of Epidemiology and Biostatistics, Michigan State University, 4Department of Molecular Obstetrics and Genetics, Wayne State University, 5Department of Obstetrics and Gynecology, Wayne State University School of Medicine, 6Department of Immunology and Microbiology, Wayne State University School of Medicine Described herein is a protocol to isolate and further study the infiltrating leukocytes of the decidua basalis and decidua parietalis - the human maternal-fetal interface. This protocol maintains the integrity of cell surface markers and yields enough viable cells for downstream applications as proven by flow cytometry analysis. Neuroscience Recording Light-evoked Postsynaptic Responses in Neurons in Dark-adapted, Mouse Retinal Slice Preparations Using Patch Clamp Techniques Chase B. Hellmer1, Tomomi Ichinose1,2 1Anatomy and Cell Biology, Wayne State University School of Medicine, 2Ophthalmology, Wayne State University School of Medicine We will demonstrate how to prepare retinal slices from the mouse eye and record light responses in retinal neurons. The entire procedure is conducted in dark-adapted conditions. Medicine A Novel Model of Mild Traumatic Brain Injury for Juvenile Rats Richelle Mychasiuk1, Allyson Farran1, Mariana Angoa-Perez2, Denise Briggs2, Donald Kuhn2, Michael J. Esser1 1Alberta Children's Hospital Research Institute, University of Calgary, 2Department of Psychiatry and Behavioral Neurosciences, Wayne State University School of Medicine & John D. Dingell VA Medical Center The modified weight-drop technique is an easy, cost-effective procedure used for the induction of mild traumatic brain injury in juvenile rats. This novel technique produces clinically relevant symptomology that will advance the study of mild traumatic brain injury (mTBI) and concussion. Behavior Marble Burying and Nestlet Shredding as Tests of Repetitive, Compulsive-like Behaviors in Mice Mariana Angoa-Pérez1, Michael J. Kane1, Denise I. Briggs1, Dina M. Francescutti1, Donald M. Kuhn1 1Research and Development Service, John D. Dingell VA Medical Center and Department of Psychiatry & Behavioral Neurosciences, Wayne State University School of Medicine Disease, head injury, genetic modifications, and treatment of mice with drugs can have profound effects on behavior. Utilizing well-characterized and validated approaches such as marble burying and nestlet shredding, compulsive-like behaviors can be documented accurately in mice as models of human obsessive-compulsive disorder and autism spectrum disorder. Medicine 2-Vessel Occlusion/Hypotension: A Rat Model of Global Brain Ischemia Thomas H. Sanderson1,2,3, Joseph M. Wider2,3 1Department of Emergency Medicine, Wayne State University School of Medicine, 2Cardiovascular Research Institute, Wayne State University School of Medicine, 3Department of Physiology, Wayne State University School of Medicine Bilateral carotid occlusion coupled with systemic hypotension produces global brain ischemia in the rat, resulting in damage to the hippocampus with reproducible severity. Animal subjects are impaired with predictable patterns of brain damage, they recover expediently, and mortality rates are comparatively low.