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Keio University School of Medicine

13 ARTICLES PUBLISHED IN JoVE

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Biology

Isolation of Basal Cells and Submucosal Gland Duct Cells from Mouse Trachea
Ahmed E. Hegab 1, Vi Luan Ha 1, Yasser S. Attiga 1, Derek W. Nickerson 1, Brigitte N. Gomperts 1
1Department of Pediatrics, David Geffen School of Medicine at UCLA

Here we demonstrate our protocol for isolation of basal and submucosal gland duct cells from mouse tracheas. We also demonstrate the method of injecting stem cells into the dorsal mouse fat pad to create an in vivo model of submucosal gland regeneration.

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Medicine

Performing Subretinal Injections in Rodents to Deliver Retinal Pigment Epithelium Cells in Suspension
Peter D. Westenskow 1,2, Toshihide Kurihara 1, Stephen Bravo 1, Daniel Feitelberg 1, Zack A. Sedillo 2, Edith Aguilar 1, Martin Friedlander 1,2
1Department of Cell and Molecular Biology, The Scripps Research Institute, 2Lowy Medical Research Institute

Here we present a community accepted protocol in multimedia format for subretinally injecting a bolus of RPE cells in rats and mice. This approach can be used for determining rescue potentials, safety profiles, and survival capacities of grafted RPE cells upon implantation in animal models of retinal degeneration.

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Developmental Biology

Generation of Induced Pluripotent Stem Cells from Human Peripheral T Cells Using Sendai Virus in Feeder-free Conditions
Yoshikazu Kishino 1, Tomohisa Seki 2, Shinsuke Yuasa 1, Jun Fujita 1, Keiichi Fukuda 1
1Department of Cardiology, Keio University School of Medicine, 2Department of Emergency and Critical Care Medicine, Keio University School of Medicine

This protocol describes how to generate induced pluripotent stem cells (iPSCs) from human peripheral T cells in feeder-free conditions using a combination of matrigel and Sendai virus vectors containing reprogramming factors.

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Medicine

Using Micro-computed Tomography for the Assessment of Tumor Development and Follow-up of Response to Treatment in a Mouse Model of Lung Cancer
Ahmed E. Hegab 1, Naofumi Kameyama 1, Aoi Kuroda 1, Shizuko Kagawa 1, Yongjun Yin 2, David Ornitz 2, Tomoko Betsuyaku 1
1Division of Pulmonary Medicine, Keio University School of Medicine, 2Department of Developmental Biology, Washington University School of Medicine

We describe a method for the detection of tumor nodule development in the lungs of an adenocarcinoma mouse model using micro-computed tomography and its use for monitoring changes in nodule size over time and in response to treatment. The accuracy of the assessment was confirmed with end-point histological quantification.

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Developmental Biology

Generation of Induced Pluripotent Stem Cells from Human Melanoma Tumor-infiltrating Lymphocytes
Hidehito Saito 1,2, Kumiko Iwabuchi 3, Noemi Fusaki 4,5, Fumito Ito 3,6
1Department of Surgery, University of Michigan, 2Department of Biochemistry II, Kanazawa Medical University, 3Center for Immunotherapy, Roswell Park Cancer Institute, 4DNAVEC Corporation, 5Department of Ophthalmology, Keio University School of Medicine, 6Department of Surgical Oncology, Roswell Park Cancer Institute

The goal of this protocol is to show the protocol for reprogramming melanoma tumor-infiltrating lymphocytes into induced pluripotent stem cells.

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Biology

Dissection of the Auditory Bulla in Postnatal Mice: Isolation of the Middle Ear Bones and Histological Analysis
Ayako Sakamoto 1, Yukiko Kuroda 1, Sho Kanzaki 2, Koichi Matsuo 1
1Laboratory of Cell and Tissue Biology, Keio University School of Medicine, 2Department of Otolaryngology Head and Neck Surgery, Keio University School of Medicine

We present a protocol to isolate the auditory bulla, capsule, and ossicles from postnatal mice for whole mount and histological analysis.

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Medicine

A Surgical Procedure for the Administration of Drugs to the Inner Ear in a Non-Human Primate Common Marmoset (Callithrix jacchus)
Sho Kurihara *1,2, Masato Fujioka *3, Tomohiko Yoshida *1,2, Makoto Koizumi 4, Kaoru Ogawa 3, Hiromi Kojima 2, Hirotaka James Okano 1
1Division of Regenerative Medicine, Jikei University School of Medicine, 2Department of Otorhinolaryngology, Jikei University School of Medicine, 3Department of Otorhinolaryngology, Head and Neck Surgery, Keio University School of Medicine, 4Laboratory Animal Facilities, Jikei University School of Medicine

We report a surgical method to administrate drugs to the inner ear of a non-human primate, the common marmoset (Callithrix jacchus), via the round window membrane.

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Neuroscience

Inducement and Evaluation of a Murine Model of Experimental Myopia
Xiaoyan Jiang 1,2, Toshihide Kurihara 1,2, Shin-ichi Ikeda 1,2, Hiromitsu Kunimi 1,2, Kiwako Mori 1,2, Hidemasa Torii 1,2, Kazuo Tsubota 2
1Laboratory of Photobiology, Keio University School of Medicine, 2Department of Ophthalmology, Keio University School of Medicine

In this protocol, we describe the full process of experimental myopia inducement in mice using newly designed eyeglasses and the technic needed for achieving stable and reproducible results in ocular parameter measurements.

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Neuroscience

Chronic Implantation of Whole-cortical Electrocorticographic Array in the Common Marmoset
Misako Komatsu 1, Takaaki Kaneko 2,3, Hideyuki Okano 2,3, Noritaka Ichinohe 1,4
1Laboratory for Molecular Analysis of Higher Brain Function, RIKEN Center for Brain Science, 2Laboratory for Marmoset Neural Architecture, RIKEN Center for Brain Science, 3Department of Physiology, Keio University School of Medicine, 4Department of Ultrastructural Research, National Center of Neurology and Psychiatry

We have developed a whole-cortical electrocorticographic array for the common marmoset that continuously covers almost the entire lateral surface of cortex, from the occipital pole to the temporal and frontal poles. This protocol describes a chronic implantation procedure of the array in the epidural space of the marmoset brain.

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Bioengineering

Four-Dimensional CT Analysis Using Sequential 3D-3D Registration
Satoshi Oki 1,2, Kazuya Kaneda 1, Yoshitake Yamada 3, Minoru Yamada 3, Yutaro Morishige 1, Kengo Harato 1, Noboru Matsumura 1, Takeo Nagura 1, Masahiro Jinzaki 3
1Department of Orthopedic Surgery, Keio University School of Medicine, 2Department of Orthopedic Surgery, Saiseikai Utsunomiya Hospital, 3Department of Radiology, Keio University School of Medicine

We analyzed joint kinematics from four-dimensional computed tomography data. The sequential 3D-3D registration method semiautomatically provides the kinematics of the moving bone with respect to the subject bone from four-dimensional computed tomography data.

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Immunological Elimination of Undifferentiated Human Pluripotent Stem Cells Using Pluripotent State-Specific Antigen, Glypican-3
Marina Okada 1, Shugo Tohyama 1, Jun Fujita 1, Tetsuya Nakatsura 2, Keiichi Fukuda 1
1Department of Cardiology, Keio University School of Medicine, 2Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center Japan

Regenerative therapies using human induced pluripotent stem cells (hiPSCs) have recently attracted much attention. In this study, we use anticancer immunotherapy with peptide vaccination to prevent hiPSC-derived tumorigenesis. Our results show that glypican-3 (GPC3) works as a pluripotent state-specific immunogenic antigen in hiPSCs.

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Medicine

A Murine Model of Ischemic Retinal Injury Induced by Transient Bilateral Common Carotid Artery Occlusion
Deokho Lee *1,2, Yukihiro Miwa *1,2,3, Heonuk Jeong 1,2, Shin-ichi Ikeda 1,2, Yusaku Katada 1,2, Kazuo Tsubota 1,2,4, Toshihide Kurihara 1,2
1Laboratory of Photobiology, Keio University School of Medicine, 2Department of Ophthalmology, Keio University School of Medicine, 3Animal eye care, Tokyo Animal Eye Clinic, 4Tsubota Laboratory, Inc.

Here, we describe a mouse model of retinal ischemia by transient bilateral common carotid artery occlusion using simple sutures and a clamp. This model can be useful for understanding the pathological mechanisms of retinal ischemia caused by cardiovascular abnormalities.

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Neuroscience

Time-Lapse Imaging of Migrating Neurons and Glial Progenitors in Embryonic Mouse Brain Slices
Hidenori Tabata 1,2, Koh-ichi Nagata 2, Kazunori Nakajima 1
1Department of Anatomy, Keio University School of Medicine, 2Department of Molecular Neurobiology, Institute for Developmental Research, Aichi Developmental Disability Center

During the development of the cerebral cortex, neurons and glial cells originate in the ventricular zone lining the ventricle and migrate toward the brain surface. Many genes are involved in this process. This protocol introduces the technique for the time-lapse imaging of migrating neurons and glial progenitors.

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