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Hudson institute of Medical Research

11 ARTICLES PUBLISHED IN JoVE

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Biology

Anterior Cervical Discectomy and Fusion in the Ovine Model
Tony Goldschlager 1,2, Jeffrey V. Rosenfeld 2, Ian R. Young 1, Graham Jenkin 1
1Monash Immunology and Stem Cell Laboratories (MISCL), Monash University, 2Department of Surgery, Monash University

This video demonstrates the technique of anterior cervical discectomy and fusion in the ovine model.

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Biology

Undecalcified Bone Preparation for Histology, Histomorphometry and Fluorochrome Analysis
Tony Goldschlager 1, Amany Abdelkader 1, Jeffrey Kerr 2, Ian Boundy 2, Graham Jenkin 1
1Monash Immunology and Stem Cell Laboratories, Monash University, 2Anatomy and Developmental Biology, Monash University

Undecalcified bone histology provides important information for a variety of clinical and research applications. It is technically challenging, particularly with large size specimens. This video illustrates the process of producing good quality sections and demonstrates the technical difficulties and methods with which to overcome them.

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Neuroscience

An Isolated Semi-intact Preparation of the Mouse Vestibular Sensory Epithelium for Electrophysiology and High-resolution Two-photon Microscopy
Victoria W. K. Tung 1, Stefano Di Marco 1, Rebecca Lim 2, Alan M. Brichta 2, Aaron J. Camp 1
1Discipline of Biomedical Science, School of Medical Sciences, Sydney Medical School, University of Sydney, 2School of Biomedical Sciences and Pharmacy, University of Newcastle

Analysis of vestibular hair cell function is complicated by their location deep within the hardest part of the skull, the petrous temporal bone. Most functional hair cell studies have used acutely isolated hair cells. Here we describe a semi-intact preparation of mouse vestibular epithelium for electrophysiological and two-photon microscopy studies.

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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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Biology

Simultaneous Measurement of Mitochondrial Calcium and Mitochondrial Membrane Potential in Live Cells by Fluorescent Microscopy
Matthew McKenzie 1,2, Sze C. Lim 1, Michael R. Duchen 3
1Centre for Genetic Diseases, Hudson Institute of Medical Research, 2The Department of Molecular and Translational Sciences, Monash University, 3Department of Physiology, University College London

Mitochondria can utilize the electrochemical potential across their inner membrane (ΔΨm) to sequester calcium (Ca2+), allowing them to shape cytosolic Ca2+ signaling within the cell. We describe a method for simultaneously measuring mitochondria Ca2+ uptake and ΔΨm in live cells using fluorescent dyes and confocal microscopy.

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Medicine

Ovine Lumbar Intervertebral Disc Degeneration Model Utilizing a Lateral Retroperitoneal Drill Bit Injury
Kai-Zheong Lim *1, Christopher D. Daly *1,2,3, Peter Ghosh 4, Graham Jenkin 3, David Oehme 5, Justin Cooper-White 6,7, Taryn Naidoo 6, Tony Goldschlager 1,8
1Department of Surgery, Monash University, 2Department of Neurosurgery, Monash University, 3The Ritchie Centre, Hudson Institute of Medical Research, 4Proteobioactives, Pty Ltd, 5Department of Neurosurgery, St Vincent's Hospital, 6Australian Institute for Bioengineering and Nanotechnology, University of Queensland, 7School of Chemical Engineering, University of Queensland, 8Department of Neurosurgery, Monash Health

Intervertebral disc degeneration is a significant contributor to back pain and a leading cause of disability worldwide. Numerous animal models of intervertebral disc degeneration exist. We demonstrate an ovine model of intervertebral disc degeneration, utilizing a drill bit, which achieves a consistent disc injury and reproducible level of disc degeneration.

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Immunology and Infection

Mouse Models Of Helicobacter Infection And Gastric Pathologies
Kimberley D'Costa 1, Michelle Chonwerawong 1, Le Son Tran 1, Richard L. Ferrero 1,2
1Centre for Innate Immunity and Infectious Diseases, Hudson Institute of Medical Research, 2Infection and Immunity Program, Monash Biomedicine Discovery Institute and Department of Microbiology, Monash University

Mice represent an invaluable in vivo model to study infection and diseases caused by gastrointestinal microorganisms. Here, we describe the methods used to study bacterial colonization and histopathological changes in mouse models of Helicobacter pylori-related disease.

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Neuroscience

Characterization and Isolation of Mouse Primary Microglia by Density Gradient Centrifugation
Jessica C. Stark *1, Euan Wallace 1,2, Rebecca Lim 1, Bryan Leaw *1
1The Ritchie Centre, Hudson Institute of Medical Research, 2Department of Obstetrics & Gynaecology, Monash University

A protocol for the isolation of primary microglia from murine brains is presented. This technique aids in furthering the current understanding of neurological conditions. Density gradient centrifugation and magnetic separation are combined to produce sufficient yield of a highly pure sample. Furthermore, we outline the steps for characterization of microglia.

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Bioengineering

Automated Counterflow Centrifugal System for Small-Scale Cell Processing
Anqi Li 1,2, Stephen Wilson 3, Ian Fitzpatrick 3, Mehri Barabadi 1,2, Siow Teng Chan 1, Mirja Krause 1,2, Gina Diamanta Kusuma 1,2, David James 3, Rebecca Lim 1,2,4
1The Ritchie Centre, Hudson Institute of Medical Research, 2Department of Obstetrics and Gynaecology, Monash University, 3Scinogy, 4Australian Regenerative Medicine Institute, Monash University

Automation is key to upscaling and cost management in cell manufacturing. This manuscript describes the use of a counterflow centrifugal cell processing device for automating the buffer exchange and cell concentration steps for small-scale bioprocessing.

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Biology

Visualizing Mitophagy with Fluorescent Dyes for Mitochondria and Lysosome
Bilin Liu 1,2, Anqi Li 1, Yuan Qin 3, Lei Chen 1, Meng Gao 1, Guohua Gong 1,2
1Institute for Regenerative Medicine, Shanghai East Hospital, School of Life Sciences and Technology, Tongji University, 2Key Laboratory of Laboratory Medicine, Ministry of Education, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, 3Department of Pharmacy, Shanghai East Hospital, Tongji University

Mitophagy is the primary mechanism of mitochondrial quality control. However, the evaluation of mitophagy in vivo is hindered by the lack of reliable quantitative assays. Presented here is a protocol for the observation of mitophagy in living cells using a cell-permeant green-fluorescent mitochondria dye and a red-fluorescent lysosome dye.

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