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Wenzhou Medical University

21 ARTICLES PUBLISHED IN JoVE

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Biology

Collection, Isolation and Enrichment of Naturally Occurring Magnetotactic Bacteria from the Environment
Zachery Oestreicher 1, Steven K. Lower 1,2, Wei Lin 3, Brian H. Lower 2
1School of Earth Sciences, The Ohio State University, 2School of Environment & Natural Resources, The Ohio State University, 3Institute of Geology and Geophysics, Chinese Academy of Sciences

We demonstrate a method to collect magnetotactic bacteria (MTB) that can be applied to natural waters. MTB can be isolated and enriched from sediment samples using a relatively simple setup that takes advantage of the bacteria's natural magnetism. Isolated MTB can then be examined in detail using both light and electron microscopy.

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Biology

Confocal Imaging of Single Mitochondrial Superoxide Flashes in Intact Heart or In Vivo
Guohua Gong 1, Wang Wang 1
1Mitochondria and Metabolism Center, Department of Anesthesiology and Pain Medicine, University of Washington

Confocal scanning microscopy is applied for imaging single mitochondrial events in perfused heart or skeletal muscles in live animal. Real-time monitoring of single mitochondrial processes such as superoxide flashes and membrane potential fluctuations enables the evaluation of mitochondrial function in a physiologically relevant context and during pathological perturbations.

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Developmental Biology

Rapid Acquisition of 3D Images Using High-resolution Episcopic Microscopy
Haochuan Zhang *1,2,3, JunGang Huang *2,3,4, Xin Liu 2,3, Ping Zhu 4, Zhongrong Li 1, Xue Li 2,3
1Department of Pediatric Surgery, The Second Affiliated Hospital & Yuying Children's Hospital, Wenzhou Medical University, 2Departments of Urology, Boston Children's Hospital, 3Department of Surgery, Harvard Medical School, 4Department of Cardiovascular Surgery, Guangdong Cardiovascular Institute, Guangdong General Hospital, Guangdong Academy of Medical Sciences

We describe a detailed protocol using high-resolution episcopic microscopy to acquire three-dimensional (3D) images of mouse embryos. This improved protocol utilizes a modified tissue preparation method to enhance penetration of the fluorescent dye, thereby permitting morphometric analysis of both small and large-sized specimens.

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Developmental Biology

In Vitro Growth of Mouse Preantral Follicles Under Simulated Microgravity
Shen Zhang 1, Yonggen Wu 1, Yimin Weng 2, Zhihui Xu 1, Wenmin Chen 3, Dahan Zheng 4, Wei Lin 5, Jun Liu 6, Ying Zhou 1,7
1Reproductive Medicine Center, The First Affiliated Hospital of Wenzhou Medical University, 2Department of Orthopaedics, The Second Affiliated Hospital of Wenzhou Medical University, 3Department of Obstetrics, The First Affiliated Hospital of Wenzhou Medical University, 4School of Laboratory Medicine and Life Science, Wenzhou Medical University, 5School of Pharmaceutical Science, Wenzhou Medical University, 6Stem Cells and Genetic Engineering Group, AgriBioscience Research Centre, Department of Economic Development, Jobs, Transport and Resources, 7Department of Histology and Embryology, Wenzhou Medical University

A highly promising technique to generate tissue constructs without using matrix is to culture cells in a simulated microgravity condition. Using a rotary culture system, we examined ovarian follicle growth and oocyte maturation in terms of follicle survival, morphology, growth, and oocyte function under the simulated microgravity condition.

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Medicine

Surgical Treatment for Benign Prostatic Hyperplasia: Holmium Laser Enucleation of the Prostate (HoLEP).
Huan Xu *1, Xiang Wan *1, Meng Gu 1, Yanbo Chen 1, Qiling Shi 1, Qi Chen 1, Zhong Wang 1
1Department of Urology, Shanghai 9th People's Hospital, Shanghai Jiaotong University School of Medicine

Here we present a safe and efficient protocol, holmium laser enucleation of the prostate, to treat benign prostatic hyperplasia.

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Biology

A Method for Islet Transplantation to the Omentum in Mouse
Ying Lu *1, Shangyou Zou *1, Suzanne Bertera *2, Rita Bottino 2, David K.C. Cooper 3, Zhengzhao Liu 1, Yi Huang 1, Chengjun Wang 1, Chungu Hong 1, Tian He 1, Hancheng Zhang 1, Qin Huo 4, Hongxing Fu 5, Zhiming Cai 1, Lisha Mou 1
1Shenzhen Xenotransplantation Medical Engineering Research and Development Center, Institute of Translational Medicine, Shenzhen Second People's Hospital, First Affiliated Hospital of Shenzhen University, Shenzhen University School of Medicine, Shenzhen University Health Science Center, 2Institute for Cellular Therapeutics, Allegheny-Singer Research Institute, 3Xenotransplantation Program/Department of Surgery, The University of Alabama at Birmingham, 4College of Life Science and Oceanography, Shenzhen University, 5School of Pharmcy, Wenzhou Medical College

A method for the omental transplantation of islets in a mouse is introduced. The isolated islets are mixed with hydrogel and the mixture is placed into the omental pouch of a diabetic mouse. Then, the blood glucose is monitored, and immuno-histochemical analysis is performed.

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Immunology and Infection

Evaluation of Zika Virus-specific T-cell Responses in Immunoprivileged Organs of Infected Ifnar1-/- Mice
Yongli Zhang *1,2, Hangjie Zhang *2, Wenqiang Ma *3, Kefang Liu 1,2, Min Zhao 4, Yingze Zhao 2, Xuancheng Lu 5, Fuping Zhang 6, Xiangdong Li 3, George F. Gao 1,2,4,6, William J. Liu 1,2
1School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, 2NHC Key Laboratory of Medical Virology and Viral Diseases, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, 3State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, 4Research Network of Immunity and Health (RNIH), Beijing Institutes of Life Science, Chinese Academy of Sciences, 5Laboratory Animal Center, Chinese Center for Disease Control and Prevention, 6CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences

A protocol to evaluate antigen-specific T-cell responses in the immunoprivileged organs of the Ifnar1-/- murine model for the Zika virus (ZIKV) infection is described. This method is pivotal for investigating the cellular mechanisms of the protection and immunopathogenesis of ZIKV vaccines and is also valuable for their efficacy evaluation.

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Medicine

Inguinal Subcutaneous White Adipose Tissue (ISWAT) Transplantation Model of Murine Islets
Yuanzheng Peng *1, Zhicheng Zou *1, Jiao Chen 1, Hancheng Zhang 1, Ying Lu 1, Rito Bittino 2, Hongxing Fu 3, David K. C. Cooper 4, Shan Lin 1, Mengtao Cao 1, Yifan Dai 5, Zhiming Cai 1, Lisha Mou 1
1Shenzhen Xenotransplantation Medical Engineering Research and Development Center, Institute of Translational Medicine, Shenzhen Second People's Hospital, First Affiliated Hospital of Shenzhen University, Health Science Center, 2Institute for Cellular Therapeutics, Allegheny-Singer Research Institute, 3School of Pharmaceutical Sciences, Wenzhou Medical University, 4Xenotransplantation Program/Department of Surgery, University of Alabama at Birmingham, 5Jiangsu Key Laboratory of Xenotransplantation, Nanjing Medical University

In this protocol, a method of murine islet isolation and transplantation into the inguinal subcutaneous white adipose tissue is described. Isolated syngeneic murine islets are transplanted into a murine recipient using a basement membrane hydrogel. The blood glucose level of the recipients is monitored, and histology analysis of the islet grafts is performed.

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Immunology and Infection

Stability and Structure of Bat Major Histocompatibility Complex Class I with Heterologous β2-Microglobulin
Di Zhang *1,2, Kefang Liu *3,4, Dan Lu *2,5, Pengyan Wang 1,2, Qingxu Zhang 1,2, Peipei Liu 2, Yingze Zhao 2, Yan Chai 4, Jianxin Lyu 1, Jianxun Qi 4, William J. Liu 1,2
1School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, 2NHC Key Laboratory of Biosafety, National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, 3Faculty of Health Sciences, University of Macau, 4CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, 5Savaid Medical School, University of Chinese Academy of Sciences

The protocol describes experimental methods to obtain stable major histocompatibility complex (MHC) class I through potential β2-microglobulin (β2m) substitutions from different species. The structural comparison of MHC I stabilized by homologous and heterologous β2m were investigated.

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Neuroscience

In Vivo Methods to Assess Retinal Ganglion Cell and Optic Nerve Function and Structure in Large Animals
Qian Ye *1, Zhonghao Yu *1, Tian Xia *1, Shengjian Lu 1, Jiaying Sun 1, Mengyun Li 1, Yu Xia 1, Si Zhang 1, Wencan Wu 1, Yikui Zhang 1
1The Eye Hospital, School of Ophthalmology & Optometry, Wenzhou Medical University

Here we demostrate several in vivo tests (flash visual evoked potential, pattern electroretinogram and optic coherence tomography) in goat and rhesus macaque to understand the structure and function of the optic nerve and its neurons.

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A Collection of Biomaterials and Biomechanical Experiments and Analysis Methods
Feng-Xun Li 1, Zhen-Zhe Li 2, Rui Jiang 2, Mei-Ling Zhang 3
1Ulsan Ship and Ocean College, Ludong University, 2College of Mechanical and Electrical Engineering, Wenzhou University, 3School of Pharmacy, Wenzhou Medical University

A Collection of Biomaterials and Biomechanical Experiments and Analysis Methods

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Genetics

Investigating the Pathogenesis of MYH7 Mutation Gly823Glu in Familial Hypertrophic Cardiomyopathy using a Mouse Model
Yu Xia *1,2, Jinlin Hu *3, Xiang Li 4, Shuang Zheng 4, Ge Wang 1,2, Songtao Tan 1,2, Zengxiao Zou 1,2, Qiong Ling 2,5, Fenghua Yang 4, Xiaoping Fan 1,2
1Department of Cardiovascular Surgery, Guangdong Provincial Hospital of Traditional Chinese Medicine, the Second Affiliated Hospital of Guangzhou University of Chinese Medicine, 2The Second Clinical College of Guangzhou University of Chinese Medicine, 3Department of Cardiovascular Surgery, The First Affiliated Hospital, Jinan University, 4Guangdong Provincial Key Laboratory of Laboratory Animals, Guangdong Laboratory Animals Monitoring Institute, 5Department of Anesthesiology, Guangdong Provincial Hospital of Chinese Medicine, the Second Affiliated Hospital of Guangzhou University of Chinese Medicine

Based on the familial hereditary cardiomyopathy family found in our clinical work, we created a C57BL/6N mouse model with a point mutation (G823E) at the mouse MYH7 locus through CRISPR/Cas9-mediated genome engineering to verify this mutation.

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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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Medicine

Image-Guided Optical Coherence Tomography to Assess Structural Changes in Rodent Retinas
Huiyi Zeng *1, Qiaoli Yang *1, Jingrun Chen 1, Liujun Ding 1, Fengqin Rao 1, Jineng Lv 1, Bintao Xie 1, Shengjin Xiang 1, Huanyun Yu 1, Xuejiao Chen 1, Kunchao Wu 1,2, Qi Chen 1, Lue Xiang 1
1State Key Laboratory of Ophthalmology, Optometry, and Vision Science, Eye Hospital, Wenzhou Medical University, 2Department of Ophthalmology, The First People's Hospital of Guiyang

The protocol presented here details the procedures of data collection and data analysis for image-guided optical coherence tomography (OCT) and demonstrates its application in multiple rodent models of ocular diseases.

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Neuroscience

Targeted Knockdown of Genes in the Choroid Plexus
Yuwen Yang *1, Chenxing Qi *1, Lanxin Hu 1, Cheng Zheng 1, Xuhang Li 1, Wu Zheng 1, Yiyun Weng 2, Haiyan Lin 3,4
1State Key Laboratory of Ophthalmology, Optometry and Visual Science, Eye Hospital, Wenzhou Medical University, 2Department of Neurology, the First Affiliated Hospital of Wenzhou Medical University, 3Rehabilitation Medicine Center, The Second Affiliated Hospital of Wenzhou Medical University, 4Integrative & Optimized Medicine Research Center, China-USA Institute for Acupuncture and Rehabilitation, Wenzhou Medical University

Here, we describe a method to selectively alter gene expressions in the choroid plexus while avoiding any impact in other brain areas.

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Biology

Modified In Vivo Matrix Gel Plug Assay for Angiogenesis Studies
Zixian Lu *1,2, Mei Yi *2, Taoli Chen *2, Yue He *2, Xia Fan *2, Hui Chen 2, Yinli Huang 1, Jianlou Niu 1,2, Xiaoqing Yan 1,2
1Pingyang Affiliated Hospital of Wenzhou Medical University, 2School of Pharmaceutical Sciences, Wenzhou Medical University

The method presented here can evaluate the effect of reagents on angiogenesis or vascular permeability in vivo without staining. The method uses dextran-FITC injection via the tail vein to visualize neo-vessels or vascular leakage.

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Medicine

Cognitive Function and Upper Limb Rehabilitation Training Post-Stroke Using a Digital Occupational Training System
Zhao Yao 1, Tianfang Zhang 2, Fenfen Chen 1,3, Weiyi Shi 2, Jinjin zheng 2, Ziwei Zhang 2, Zuobing Chen 1,2
1The Second Affiliated Hospital and Yuying Children's Hospital, Wenzhou Medical University, 2Department of Rehabilitation Medicine, The First Affiliated Hospital, School of Medicine, Zhejiang University, 3Department of Rehabilitation Medicine, Taizhou Hospital Affiliated to Wenzhou Medical University

The current protocol outlines how the VR-based digital occupational training system enhances the rehabilitation of patients with cognitive impairment and upper limb dysfunction following a stroke.

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Neuroscience

Investigating Retinal Circuits and Molecular Localization by Pre-Embedding Immunoelectron Microscopy
Fenglan Wang *1,2, Wenhui Zhong *1,2, Jun Zhang 1,2
1State Key Laboratory of Ophthalmology, Optometry and Visual Science, Eye Hospital, Wenzhou Medical University, 2Laboratory of Retinal Physiology and Disease, School of Ophthalmology and Optometry, Wenzhou Medical University

This protocol outlines the detailed steps of pre-embedding immunoelectron microscopy, with a focus on exploring synaptic circuits and protein localization in the retina.

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Neuroscience

Preparation of Retinal Samples for Volume Electron Microscopy
Wenna Zhao *1,2, Qingwen Yang *1,2, Xiaoqian Lai 1,2, Jun Zhang 1,2
1State Key Laboratory of Ophthalmology, Optometry and Visual Science, Eye Hospital, Wenzhou Medical University, 2Laboratory of Retinal Physiology and Disease, School of Ophthalmology and Optometry, Wenzhou Medical University

This protocol describes the detailed steps for preparing retinal samples for volume electron microscopy, focusing on the structural features of retinal photoreceptor terminals.

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Neuroscience

Preparing Retinal Organoid Samples for Transmission Electron Microscopy
Xiaoqing Liu *1,2, Bilin Rao *1,2, Qingyang Lin 1,2, Meiling Gao 1,2, Jun Zhang 1,2
1State Key Laboratory of Ophthalmology, Optometry and Visual Science, Eye Hospital, Wenzhou Medical University, 2Laboratory of Retinal Physiology and Disease, School of Ophthalmology and Optometry, Wenzhou Medical University

This protocol provides an optimized and elaborate preparation procedure for retinal organoid samples for transmission electron microscopy. It is suitable for applications that involve the analysis of synapses in mature retinal organoids.

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Medicine

Enhancing Upper Limb Function and Motor Skills Post-Stroke Through an Upper Limb Rehabilitation Robot
Tianfang Zhang 1, Zhao Yao 1,2, Fenfen Chen 2,3, Jie Wang 4, Weiyi Shi 1, Jinjin Zheng 1, Ziwei Zhang 1, Zuobing Chen 1,2,3
1Department of Rehabilitation Medicine, The First Affiliated Hospital, School of Medicine, Zhejiang University, 2The Second Affiliated Hospital and Yuying Children's Hospital, Wenzhou Medical University, 3Department of Rehabilitation Medicine, Taizhou Hospital Affiliated to Wenzhou Medical University, 4Affiliate Hospital of Shandong University of Traditional Chinese Medicine

This protocol describes an upper limb rehabilitation robot that provides intelligent feedback through four modes. These modes enhance upper limb function and flexibility, thereby improving patients' quality of life.

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