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17 ARTICLES PUBLISHED IN JoVE

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Medicine

Combined In vivo Optical and µCT Imaging to Monitor Infection, Inflammation, and Bone Anatomy in an Orthopaedic Implant Infection in Mice
Nicholas M. Bernthal 1, Brad N. Taylor 2, Jeffrey A. Meganck 2, Yu Wang 3, Jonathan H. Shahbazian 3, Jared A. Niska 1, Kevin P. Francis 2, Lloyd S. Miller 3,4
1Orthopaedic Hospital Research Center, Orthopaedic Hospital Department of Orthopaedic Surgery, David Geffen School of Medicine at University of California, Los Angeles (UCLA), 2PerkinElmer, 3Department of Dermatology, Johns Hopkins University School of Medicine, 4Department of Medicine, Division of Infectious Diseases, Department of Orthopaedic Surgery, Johns Hopkins University School of Medicine

Combined optical and μCT imaging in a mouse model of orthopaedic implant infection, utilizing a bioluminescent engineered strain of Staphylococcus aureus, provided the capability to noninvasively and longitudinally monitor the dynamics of the bacterial infection, as well as the corresponding inflammatory response and anatomical changes in the bone.

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Bioengineering

Construction of Defined Human Engineered Cardiac Tissues to Study Mechanisms of Cardiac Cell Therapy
Timothy J. Cashman 1, Rebecca Josowitz 2, Bruce D. Gelb 2, Ronald A. Li 1,3, Nicole C. Dubois 2, Kevin D. Costa 1
1Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, 2The Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, 3Stem Cell & Regenerative Medicine Consortium, LKS Faculty of Medicine, University of Hong Kong

This manuscript describes the creation of defined engineered cardiac tissues using surface marker expression and cell sorting. The defined tissues can then be used in a multi-tissue bioreactor to investigate mechanisms of cardiac cell therapy in order to provide a functional, yet controlled, model system of the human heart.

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Bioengineering

Modeling Ovarian Cancer Multicellular Spheroid Behavior in a Dynamic 3D Peritoneal Microdevice
Shan-Shan Li *1, Carman K. M. Ip *1, Matthew Y. H. Tang *2, Samuel K. H. Sy 2, Susan Yung 3, Tak-Mao Chan 3, Mengsu Yang 4, Ho Cheung Shum 2, Alice S.T. Wong 1
1School of Biological Sciences, University of Hong Kong, 2Department of Mechanical Engineering, University of Hong Kong, 3Department of Medicine, University of Hong Kong, 4Department of Biomedical Sciences, Key Laboratory of Biochip Technology, Biotech and Health Centre, Shenzhen Research Institutes of City University of Hong Kong

To study ovarian tumor progression in a physiologically relevant model, multicellular spheroids were cultured in a microdevice under simulated fluid flow. This dynamic 3D model emulates the intraperitoneal environment with the cellular and mechanical components where ovarian cancer metastasis occurs.

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Developmental Biology

Defined and Scalable Generation of Hepatocyte-like Cells from Human Pluripotent Stem Cells
Yu Wang 1, Sharmin Alhaque 1, Kate Cameron 1, Jose Meseguer-Ripolles 1, Baltasar Lucendo-Villarin 1, Hassan Rashidi 1, David C. Hay 1
1MRC Centre for Regenerative Medicine, University of Edinburgh

The method presented here describes a scalable and good manufacturing practice (GMP)-ready differentiation system to generate human hepatocyte-like cells from pluripotent stem cells. It serves as a cost-effective and standardized system to generate human hepatocyte-like cells for basic and applied human liver research.

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JoVE Core

Scalable Solution-processed Fabrication Strategy for High-performance, Flexible, Transparent Electrodes with Embedded Metal Mesh
Arshad Khan 1, Sangeon Lee 2, Taehee Jang 3, Ze Xiong 4, Cuiping Zhang 1,5, Jinyao Tang 4, L. Jay Guo 2,3, Wen-Di Li 1,5
1Department of Mechanical Engineering, University of Hong Kong, 2Department of Mechanical Engineering, University of Michigan, 3Department of Electrical Engineering and Computer Science, University of Michigan, 4Department of Chemistry, University of Hong Kong, 5HKU-Shenzhen Institute of Research and Innovation

This protocol describes a solution-based fabrication strategy for high-performance, flexible, transparent electrodes with fully-embedded, thick metal mesh. Flexible transparent electrodes fabricated by this process demonstrate among the highest reported performances, including ultra-low sheet resistance, high optical transmittance, mechanical stability under bending, strong substrate adhesion, surface smoothness, and environmental stability.

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Developmental Biology

A Familial Hypercholesterolemia Human Liver Chimeric Mouse Model Using Induced Pluripotent Stem Cell-derived Hepatocytes
Jiayin Yang *1,2,6, Lai-Yung Wong *2, Xiao-Yu Tian *3, Rui Wei 1,2, Wing-Hon Lai 2, Ka-Wing Au 2, Zhiwei Luo 4,5, Carl Ward 4,5, Wai-In Ho 2, David P. Ibañez 4,5, Hao Liu 4,5, Xichen Bao 4,5, Baoming Qin 4,5, Yu Huang 3, Miguel A. Esteban 4,5,7, Hung-Fat Tse 1,2,6,7
1Department of Medicine, University of Hong Kong-Shenzhen Hospital, 2The Cardiology Division, Department of Medicine, Li Ka Shing Faculty of Medicine, University of Hong Kong, 3School of Biomedical Sciences, Institute of Vascular Medicine, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong, 4Key Laboratory of Regenerative Biology of the Chinese Academy of Sciences, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health and Guangzhou Medical University, 5Laboratory of RNA, Chromatin, and Human Disease, CAS Key Laboratory of Regenerative Biology and Guangdong Provincial Key Laboratory of Stem Cells and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, 6Research Centre of Heart, Brain, Hormone, and Healthy Ageing, Li Ka Shing Faculty of Medicine, University of Hong Kong, 7Hong Kong-Guangdong Stem Cell and Regenerative Medicine Research Centre, University of Hong Kong and Guangzhou Institutes of Biomedicine and Health

Here, we present a protocol to generate a human liver chimeric mouse model of familial hypercholesterolemia using human induced pluripotent stem cell-derived hepatocytes. This is a valuable model for testing new therapies for hypercholesterolemia.

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Biology

Laser Capture Microdissection of Highly Pure Trabecular Meshwork from Mouse Eyes for Gene Expression Analysis
Caleb Sutherland *1, Yu Wang *2, Robert V. Brown *1, Julie Foley 2, Beth Mahler 2, Kyathanahalli S. Janardhan 2,3, Ramesh C. Kovi 2,4, Anton M. Jetten 1
1Immunity, Inflammation, and Disease Laboratory, Division of Intramural Research, National Institute of Environmental Health Sciences, NIH, 2Cellular and Molecular Pathology Branch, Division of the National Toxicology Program, National Institute of Environmental Health Sciences, NIH, 3Integrated Laboratory Systems Inc., 4Experimental Pathology Laboratories Inc.

Here, we describe a protocol for a reproducible laser capture microdissection (LCM) for isolating trabecular meshwork (TM) for downstream RNA analysis. The ability to analyze changes in gene expression in the TM will help in understanding the underlying molecular mechanisms of TM-related ocular diseases.

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Developmental Biology

Semi-automated Production of Hepatocyte Like Cells from Pluripotent Stem Cells
Jose Meseguer-Ripolles 1, Baltasar Lucendo-Villarin 1, Yu Wang 1, David C. Hay 1
1MRC Centre for Regenerative Medicine, University of Edinburgh

This protocol describes a semi-automated approach to produce hepatocyte-like cells from human pluripotent stem cells in a 96 well plate format. This process is rapid and cost-effective, allowing the production of quality assured batches of hepatocyte-like cells for basic and applied human research.

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Establishment of Liver Cancer Patient-derived Xenograft Models
Chuan Xing Wu 1,2, Jia Yi Chen 1, Kun Dong Zhang 1, Dong Wang Yan 1, Xiao Qi Wang 2, Zheng Jun Qiu 1
1Department of General Surgery, Shanghai General Hospital, School of Medicine, Shanghai Jiaotong University, 2Department of Surgery, University of Hong Kong

Here, we present a protocol to establish liver cancer patient-derived xenograft models for the preclinical study of novel anticancer drugs.

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Biochemistry

Robust Comparison of Protein Levels Across Tissues and Throughout Development Using Standardized Quantitative Western Blotting
Yu-Ting Huang 1,2, Dinja van der Hoorn *1,2, Leire M. Ledahawsky *1,2, Anna A. L. Motyl *1,2, Crispin Y. Jordan 1, Thomas H. Gillingwater 1,2, Ewout J. N. Groen 1,2
1Centre for Discovery Brain Sciences, University of Edinburgh, 2Euan MacDonald Centre for Motor Neurone Disease Research, University of Edinburgh

This method describes a robust and reproducible approach for the comparison of protein levels in different tissues and at different developmental timepoints using a standardized quantitative western blotting approach.

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Biology

Assessment of Vascular Tone Responsiveness using Isolated Mesenteric Arteries with a Focus on Modulation by Perivascular Adipose Tissues
Daniels Konja 1, Cuiting Luo 1, Wai Yan Sun 1, Kangmin Yang 1, Andy W.C. Man 1, Aimin Xu 1, Paul M. Vanhoutte 1, Yu Wang 1
1The State Key Laboratory of Pharmaceutical Biotechnology and the Department of Pharmacology and Pharmacy, University of Hong Kong

The protocol describes the use of wire myography to evaluate the transmural isometric tension of mesenteric arteries isolated from mice, with special consideration of the modulation by factors released from endothelial cells and perivascular adipose tissues.

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Developmental Biology

Serum Free Production of Three-dimensional Human Hepatospheres from Pluripotent Stem Cells
Balta Lucendo-Villarin 1, Hassan Rashidi 1,2, Sharmin Alhaque 1,3, Lena Fischer 1,4, Jose Meseguer-Ripolles 1, Yu Wang 1, Cliona O'Farrelly 4, Michael Themis 3, David C. Hay 1
1MRC Centre for Regenerative Medicine, University of Edinburgh, 2UCL Great Ormond Street Institute of Child Health, University College London, 3Division of Biosciences, Department of Life Sciences, College of Health and Life Sciences, Brunel University London, 4School of Biochemistry and Immunology, Trinity Biomedical Sciences Institute, Trinity College Dublin

This protocol describes an approach to produce hepatospheres from human pluripotent stem cells using a defined culture system and cell self-assembly. This protocol is reproducible in a number of cell lines, cost effective and allows the production of stable human hepatospheres for biomedical application.

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Medicine

Establishing a Swine Model of Post-myocardial Infarction Heart Failure for Stem Cell Treatment
Sijia Sun *1, Yu Jiang *1, Zhe Zhen 1,2, Wing-Hon Lai 1,2, Songyan Liao 1,2, Hung-Fat Tse 1,2
1Cardiology Division, Department of Medicine, Queen Mary Hospital, University of Hong Kong, 2Shenzhen Institutes of Research and Innovation, University of Hong Kong

We sought to establish a swine model of heart failure induced by left circumflex artery blockage and rapid pacing to test the effect and safety of intramyocardial administration of stem cells for cell-based therapies.

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JoVE Core

Asymmetric Thermoelectrochemical Cell for Harvesting Low-grade Heat under Isothermal Operation
Kaiyu Mu *1, Xun Wang *1, Ka Ho Li 1, Yu-Ting Huang 1, Shien-Ping Feng 1
1Electrochemical Nanoengineering Group, Department of Mechanical Engineering, University of Hong Kong

Low-grade heat is abundant, but its efficient recovery is still a great challenge. We report an asymmetric thermoelectrochemical cell using graphene oxide as a cathode and polyaniline as an anode with KCl as the electrolyte. This cell works under isothermal heating, exhibiting a high heat-to-electricity conversion efficiency in low-temperature regions.

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Genetics

Introducing Point Mutations into Human Pluripotent Stem Cells Using Seamless Genome Editing
Yu Wang 1, Andrew J. H. Smith 1, David C. Hay 1
1MRC Centre for Regenerative Medicine, University of Edinburgh

Here, we describe a detailed method for seamless gene editing in human pluripotent stem cells using a piggyBac-based donor plasmid and the Cas9 nickase mutant. Two point mutations were introduced into exon 8 of the hepatocyte nuclear factor 4 alpha (HNF4α) locus in human embryonic stem cells (hESCs).

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Biology

Hepatic Progenitor Specification from Pluripotent Stem Cells using a Defined Differentiation System
Jose Meseguer-Ripolles 1, Yu Wang 1, Agnes Sorteberg 1, Aishwariya Sharma 2, Nan-Linda Ding 2, Baltasar Lucendo-Villarin 1, Philipp Kramer 2, Charis-Patricia Segeritz 2, David C. Hay 1
1MRC Centre for Regenerative Medicine, University of Edinburgh, 2Research & Development, STEMCELL Technologies Inc

The goal of this article is to provide a standardized approach to induce human hepatic progenitor differentiation from pluripotent stem cells. The development of this procedure with ready-to-use media formulations offer the user a facile system to generate human liver cells for biomedical research and translation.

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Medicine

Acupuncture in a Rat Model of Asthma
Dong-Dong Zhou 1, Guang-Quan Zhang 1, Qing-Yi Zhao 1, Mi Cheng 1, Jin Lu 1, Yu Wang 1, Yu-Dong Xu 1, Yan-Jiao Chen 1, Yong-Qing Yang 1, Lei-Miao Yin 1,2
1Laboratory of Molecular Biology, Shanghai Research Institute of Acupuncture and Meridian, Shanghai University of Traditional Chinese Medicine, 2Shanghai Innovation Center of Traditional Chinese Medicine Health Service

A high platform can fix rats without restriction and completely expose the acupoints on the back during acupuncture manipulation. This article describes methods for the fabrication of the high platform, establishes a rat model of asthma and measures changes in respiratory function using a noninvasive and real-time whole-body plethysmography (WBP) system.

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