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Wake Forest University Health Sciences

6 ARTICLES PUBLISHED IN JoVE

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Neuroscience

DiOLISTIC Labeling of Neurons from Rodent and Non-human Primate Brain Slices
Gail K. Seabold 1, James B. Daunais 2, Andrew Rau 3, Kathleen A. Grant 3, Veronica A. Alvarez 1
1Section on Neuronal Structure, Laboratory for Integrative Neuroscience, NIAAA, NIH, 2Department Physiology and Pharmacology, Wake Forest University Health Sciences, 3Oregon National Primate Research Center, Division of Neuroscience, Oregon Health and Science University

We demonstrate the use of the gene gun to introduce fluorescent dyes, such as DiI, into neurons in brain slices from rodents and non-human primates of different ages. In this particular case, we use adult mice (3-6 months old) and adult cynomologus monkeys (9-15 years old). This technique, originally described by the laboratory of Dr. Lichtman (Gan et al., 2000), is well suited for the study of dendritic branching and dendritic spine morphology and can be combined with traditional immunostaining, if detergents are kept at a low concentration.

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Medicine

Generation of Alginate Microspheres for Biomedical Applications
Omaditya Khanna 1, Jeffery C. Larson 2, Monica L. Moya 3, Emmanuel C. Opara 4, Eric M. Brey 2,5
1Department of Chemical and Biological Engineering, Illinois Institute of Technology, 2Department of Biomedical Engineering, Illinois Institute of Technology, 3Department of Biomedical Engineering, University of California at Irvine, 4Wake Forest Institute for Regenerative Medicine and Department of Biomedical Engineering, Wake Forest University Health Sciences, 5Research Service, Hines Veterans Administration Hospital

In the following sections, we outline procedures for the preparation of alginate microspheres for use in biomedical applications. We specifically illustrate a technique for creating multilayered alginate microspheres for the dual purpose of cell and protein encapsulation as a potential treatment for type 1 diabetes.

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Biology

Measuring Respiratory Function in Mice Using Unrestrained Whole-body Plethysmography
Rebecca Lim 1,2, Marcus J. Zavou 1, Phillipa-Louise Milton 3, Siow Teng Chan 1, Jean L. Tan 1, Hayley Dickinson 1,2, Sean V. Murphy 4, Graham Jenkin 1,2, Euan M. Wallace 1,2
1The Ritchie Centre, Monash Institute of Medical Research, 2Department of Obstetrics and Gynaecology, Monash Medical Centre, 3Animal Resource Centre, Perth, Australia, 4Wake Forest Institute for Regenerative Medicine

The assessment of respiratory physiology has traditionally relied upon techniques, which require restraint or sedation of the animal. Unrestrained whole-body plethysmography, however, provides precise, non-invasive, quantitative analysis of respiratory physiology in animal models. In addition, the technique allows repeated respiratory assessment of mice allowing for longitudinal studies.

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Medicine

Isolation, Cryopreservation and Culture of Human Amnion Epithelial Cells for Clinical Applications
Sean V. Murphy 1, Amritha Kidyoor 1, Tanya Reid 1, Anthony Atala 1, Euan M. Wallace 2, Rebecca Lim 2
1Wake Forest Institute for Regenerative Medicine, Wake Forest University Health Sciences, 2The Ritchie Centre, Monash Institute of Medical Research, Monash University

We describe a protocol to isolate and culture human amnion epithelial cells (hAECs) using animal product-free reagents in accordance with current good manufacturing practices (cGMP) guidelines.

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Immunology and Infection

Methods to Evaluate Cytotoxicity and Immunosuppression of Combustible Tobacco Product Preparations
Subhashini Arimilli 1, Brad E. Damratoski 1, Prasad G.L. 2
1Department of Microbiology and Immunology, Wake Forest University Health Sciences, 2R&D Department, R.J. Reynolds Tobacco Company

Using optimized human peripheral blood mononuclear cell (PBMC) ex vivo assays, we showed that a combustible tobacco product preparation markedly suppresses receptor-mediated intracellularly secreted cytokines and cytolytic ability of effector PBMCs. These rapid assays may be useful in product evaluation and understanding the potential long-term effects of tobacco exposure.

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Bioengineering

Bioprinting Cellularized Constructs Using a Tissue-specific Hydrogel Bioink
Aleksander Skardal 1, Mahesh Devarasetty 1, Hyun-Wook Kang 1, Young-Joon Seol 1, Steven D. Forsythe 1, Colin Bishop 1, Thomas Shupe 1, Shay Soker 1, Anthony Atala 1
1Wake Forest Institute for Regenerative Medicine, Wake Forest Univeristy Health Sciences

We describe a set of protocols that together provide a tissue-mimicking hydrogel bioink with which functional and viable 3-D tissue constructs can be bioprinted for use in in vitro screening applications.

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