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University of Wisconsin School of Medicine and Public Health

14 ARTICLES PUBLISHED IN JoVE

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Medicine

Renal Capsule Xenografting and Subcutaneous Pellet Implantation for the Evaluation of Prostate Carcinogenesis and Benign Prostatic Hyperplasia
Tristan M. Nicholson 1,2, Kristen S. Uchtmann 1, Conrad D. Valdez 1, Ashleigh B. Theberge 1,3, Tihomir Miralem 1, William A. Ricke 1
1Department of Urology, University of Wisconsin-Madison, 2Medical Scientist (MD/PhD) Training Program, University of Rochester School of Medicine & Dentistry, 3Molecular and Environmental Toxicology Center, University of Wisconsin-Madison

We describe the manufacture of compressed hormone pellets, as well as subcutaneous surgical implantation into mice. This strategy can be combined with the growth of cell and tissue xenografts under the renal capsule of athymic nude mice to evaluate hormonal carcinogenesis and regulation of benign prostate growth.

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Medicine

A Murine Model of Arterial Restenosis: Technical Aspects of Femoral Wire Injury
Toshio Takayama 1, Xudong Shi 1, Bowen Wang 1, Sarah Franco 1, Yifan Zhou 1, Daniel DiRenzo 1, Alycia Kent 1, Peter Hartig 1, Joshua Zent 1, Lian-Wang Guo 1
1Department of Surgery, University of Wisconsin School of Medicine and Public Health

The mouse femoral artery wire injury model of restenosis is technically challenging. In this protocol we show the key technical details essential for successfully performing wire injury to induce consistent neointima for studies of restenosis.

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JoVE Journal

Immunostaining to Visualize Murine Enteric Nervous System Development
Amanda J. Barlow-Anacker 1, Christopher S. Erickson 1, Miles L. Epstein 2, Ankush Gosain 1,2,3
1Department of Surgery, University of Wisconsin School of Medicine and Public Health, 2Department of Neurosciences, University of Wisconsin School of Medicine and Public Health, 3Department of Pediatrics, University of Wisconsin School of Medicine and Public Health

The enteric nervous system is formed by neural crest cells that proliferate, migrate and colonize the gut. Neural crest cells differentiate into neurons with markers specific for their neurotransmitter phenotype. This protocol describes a technique for dissecting, fixing and immunostaining of the murine embryonic gastrointestinal tract to visualize enteric nervous system neurotransmitter expression.

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Developmental Biology

Direct Induction of Hemogenic Endothelium and Blood by Overexpression of Transcription Factors in Human Pluripotent Stem Cells
Irina Elcheva 1, Vera Brok-Volchanskaya 1, Igor Slukvin 2
1Primate Research Center, University of Wisconsin-Madison, 2Department of Pathology and Laboratory Medicine, University of Wisconsin School of Medicine and Public Health

This protocol describes the efficient induction of hemogenic endothelium and multipotential hematopoietic progenitors from human pluripotent stem cells via the forced expression of transcription factors.

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Biology

Quantitation of Protein Expression and Co-localization Using Multiplexed Immuno-histochemical Staining and Multispectral Imaging
Tyler M. Bauman 1,2, Emily A. Ricke 2, Sally A. Drew 3, Wei Huang 3,4, William A. Ricke 2,4
1Division of Urologic Surgery, Washington University in St. Louis School of Medicine, 2Department of Urology, University of Wisconsin School of Medicine and Public Health, 3Department of Pathology and Laboratory Medicine, University of Wisconsin School of Medicine and Public Health, 4O’Brien Urology Research Center, University of Wisconsin School of Medicine and Public Health

Immunohistochemistry is a powerful lab technique for evaluating protein localization and expression within tissues. Current semi-automated methods for quantitation introduce subjectivity and often create irreproducible results. Herein, we describe methods for multiplexed immunohistochemistry and objective quantitation of protein expression and co-localization using multispectral imaging.

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Biochemistry

Horizontal Gel Electrophoresis for Enhanced Detection of Protein-RNA Complexes
Megan E. Dowdle 1, Susanne Blaser Imboden 1, Sookhee Park 1, Sean P. Ryder 2, Michael D. Sheets 1
1Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, 2Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School

Native polyacrylamide gel electrophoresis is a fundamental tool for analyzing RNA-protein interactions. Traditionally most experiments have used vertical gels. However, horizontal gels provide several advantages, such as the opportunity to monitor complexes during electrophoresis. We provide a detailed protocol for generating and using horizontal native gel electrophoresis.

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Bioengineering

Two Methods for Decellularization of Plant Tissues for Tissue Engineering Applications
Michal Adamski 1, Gianluca Fontana 2, Joshua R. Gershlak 4, Glenn R. Gaudette 4, Hau D. Le 1, William L. Murphy 2,3
1Department of Surgery, University of Wisconsin-Madison, 2Department of Orthopedics and Rehabilitation, University of Wisconsin School of Medicine and Public Health, 3Department of Biomedical Engineering, University of Wisconsin College of Engineering, 4Department of Biomedical Engineering, Worcester Polytechnic Institute

Here we present, and contrast two protocols used to decellularize plant tissues: a detergent-based approach and a detergent-free approach. Both methods leave behind the extracellular matrix of the plant tissues used, which can then be utilized as scaffolds for tissue engineering applications.

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Bioengineering

A Reliable and Reproducible Critical-Sized Segmental Femoral Defect Model in Rats Stabilized with a Custom External Fixator
Benjamin Kerzner 1, Hannah L. Martin 2, Michael Weiser 1, Gianluca Fontana 1, Nicholas Russell 1, William L. Murphy 1,2, Erik A. Lund 3, Christopher J. Doro 1
1Department of Orthopedics and Rehabilitation, University of Wisconsin-Madison, 2Department of Biomedical Engineering, University of Wisconsin-Madison, 3Summit Orthopedics

In vivo mammalian models of critical-sized bone defects are essential for researchers studying healing mechanisms and orthopedic therapies. Here, we introduce a protocol for the creation of reproducible, segmental, femoral defects in rats stabilized using external fixation.

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Medicine

Ultrasonography of the Adult Male Urinary Tract for Urinary Functional Testing
Teresa T. Liu *1,2, Allison C. Rodgers *3, Tristan M. Nicholson 4, Jill A. Macoska 5,6, Paul C. Marker 6,7, Chad M. Vezina 6,8, Dale E. Bjorling 6,8, Alejandro Roldan-Alzate 2,9,10, Diego Hernando 10,11, Granville L. Lloyd 12, Timothy A. Hacker 3, William A. Ricke 1,6
1Department of Urology, University of Wisconsin-Madison, 2K12 Kure, University of Wisconsin-Madison, 3Cardiovascular Research Center, Department of Medicine, University of Wisconsin-Madison, 4Department of Urology, University of Washington, 5University of Massachusetts Boston, 6U54 George M. O'Brien Center, University of Wisconsin-Madison, 7College of Pharmacy, University of Wisconsin-Madison, 8School of Veterinary Medicine, University of Wisconsin-Madison, 9Department of Mechanical Engineering, University of Wisconsin-Madison, 10Department of Radiology, University of Wisconsin-Madison, 11Department of Medical Physics, University of Wisconsin-Madison, 12Rocky Mountain Regional VA Medical Center, University of Colorado

We describe the use of high frequency ultrasound with contrast imaging as a method to measure bladder volume, bladder wall thickness, urine velocity, void volume, void duration, and urethral diameter. This strategy can be used to assess voiding dysfunction and treatment efficacy in various mouse models of lower urinary tract dysfunction (LUTD).

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Engineering

Sandy Soil Improvement through Microbially Induced Calcite Precipitation (MICP) by Immersion
Shihui Liu 1, Kang Du 1, Kejun Wen 1, Wei Huang 2, Farshad Amini 1, Lin Li 3
1Department of Civil & Environmental Engineering, Jackson State University, 2School of Civil Engineering and Architecture, Chongqing University of Science and Technology, 3Department of Civil & Architectural Engineering, Tennessee State University

Here, microbially induced calcite precipitation (MICP) technology is presented to improve soil properties by immersion.

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Medicine

Establishment of a Severe Dry Eye Model Using Complete Dacryoadenectomy in Rabbits
Robert A. Honkanen 1, Liqun Huang 2,3, Wei Huang 1, Basil Rigas 2,4
1Department of Ophthalmology, Stony Brook University, 2Department of Medicine, Stony Brook University, 3Medicon Pharmaceuticals, Inc, 4Department of Preventive Medicine, Stony Brook University

A novel approach is presented to induce chronic dry eye disease in rabbits by surgically removing all orbital lacrimal glands. This method, distinct from those previously reported, produces a stable, reproducible model of aqueous deficient dry eye well suited to study tear physiology and pathophysiology and ocular therapeutics.

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Medicine

A Doxorubicin-Induced Murine Model of Dilated Cardiomyopathy In Vivo
Yihai Liu 1, Wenfeng Zhang 1, Tingting Hu 1, Jie Ni 1, Biao Xu 1, Wei Huang 1
1Department of Cardiology, Nanjing Drum Tower Hospital, Clinical College of Nanjing Medical University

Described is a protocol to establish a Doxorubicin-induced dilated cardiomyopathy (DCM) model in mice via long-term intraperitoneal injection of Doxorubicin.

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Medicine

Real-Time Dynamic Navigation System for the Precise Quad-Zygomatic Implant Placement in a Patient with a Severely Atrophic Maxilla
Yihan Shen *1, Qinggang Dai *1, Baoxin Tao *1, Wei Huang 2, Feng Wang 2, Kengliang Lan 1, Yuanyuan Sun 1, Xiaowan Ling 1, Lijun Yan 1, Yueping Wang 1, Yiqun Wu 1
1The 2nd Dental Center, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, College of Stomatology, Shanghai Jiao Tong University, National Center for Stomatology, National Clinical Research Center for Oral Diseases, Shanghai Key Laboratory of Stomatology, 2Department of Dental Implantology, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, College of Stomatology, Shanghai Jiao Tong University, National Center for Stomatology, National Clinical Research Center for Oral Diseases, Shanghai Key Laboratory of Stomatology

Here, we present a protocol to achieve precise quad-zygomatic implant placement in patients with severely atrophic maxilla using a real-time dynamic navigation system.

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Bioengineering

High-Throughput Optogenetics Experiments in Yeast Using the Automated Platform Lustro
Zachary P. Harmer 1, Megan N. McClean 1,2
1Department of Biomedical Engineering, University of Wisconsin-Madison, 2University of Wisconsin Carbone Cancer Center, University of Wisconsin School of Medicine and Public Health

This protocol outlines the steps for utilizing the automated platform Lustro to perform high-throughput characterization of optogenetic systems in yeast.

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