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Nanjing Medical University

18 ARTICLES PUBLISHED IN JoVE

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Biochemistry

Regeneration of Arrayed Gold Microelectrodes Equipped for a Real-Time Cell Analyzer
Zhihui Xu *1, Yiyan Song *1, Huijun Jiang 2, Yan Kong 3, Xiaoming Li 3, Jin Chen 1, Yuan Wu 4
1School of Public Health, Nanjing Medical University, 2School of Pharmacy, Nanjing Medical University, 3State Key Laboratory of Materials-Oriented Chemical Engineering, College of Chemistry and Chemical Engineering, Nanjing Tech University, 4Department of Medical Oncology, Jiangsu Cancer Hospital, Jiangsu Institute of Cancer Research, The Affiliated Cancer Hospital of Nanjing Medical University

This protocol describes a general strategy to regenerate commercial arrayed gold microelectrodes equipped for a label-free cell analyzer aimed at saving on the high running costs ofmicrochip-based assays. The regeneration process includes trypsin digestion, rinsing with ethanol and water, and a spinning step, which enables repeated usage of microchips.

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Environment

Experimental Protocol for Detecting Mitochondrial Function in Hepatocytes Exposed to Organochlorine Pesticides
Qian Liu 1,2,3, Zhaoyan Jiang 3, Aihua Gu 1,2
1State Key Laboratory of Reproductive Medicine, Institute of Toxicology, Nanjing Medical University, 2Key Laboratory of Modern Toxicology of Ministry of Education, School of Public Health, Nanjing Medical University, 3Center of Gallbladder Disease, Shanghai East Hospital, Institute of Gallstone Disease, Tongji University School of Medicine

Understanding the influence of environmental organochlorine pesticides (OCPs) on mitochondrial function in hepatocytes is important in exploring the mechanism of OCPs causing metabolic disorders. This paper presents detailed methods on detecting hepatic mitochondrial function.

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Developmental Biology

An In Vivo Method to Study Mouse Blood-Testis Barrier Integrity
Mengrou Liu *1, Chunsen Zhu *1, Shun Bai *2, Xin Li 1, Kaiqiang Fu 1, Lan Ye 1, Ke Zheng 1
1State Key Laboratory of Reproductive Medicine, Nanjing Medical University, 2Center for Reproductive Medicine, Department of Obstetrics and Gynecology, The First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China

Here, we present a protocol to assess the blood-testis barrier integrity by injecting inulin-FITC into testes. This is an efficient in vivo method to study blood-testis barrier integrity that can be compromised by genetic and environmental elements.

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Behavior

A Behavioral Test Battery for the Repeated Assessment of Motor Skills, Mood, and Cognition in Mice
Ran You 1,2,3, Yan Liu 3, Raymond Chuen-Chung Chang 3
1Nanjing Key Laboratory of Pediatrics, Children's Hospital of Nanjing Medical University, 2Nanjing Key Laboratory of Pediatrics, Nanjing Medical University, 3Laboratory of Neurodegenerative Disease, Li Ka Shing Faculty of Medicine, University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region (HK SAR)

A comprehensive behavioral test battery of motor skills, mood—including social interaction, depression, and anxiety—and cognition is designed for the repeated assessment of neurodegeneration-related behavioral changes in mice.

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Biology

Enhanced Crosslinking Immunoprecipitation (eCLIP) Method for Efficient Identification of Protein-bound RNA in Mouse Testis
Qiushi Xu *1, Caifeng Wang *1, Li Ling *1, Qiuling Yue *1, Mengrou Liu 1, Shuya Zhang 1, Kaiqiang Fu 1, Lan Ye 1, Ke Zheng 1
1State Key Laboratory of Reproductive Medicine, Nanjing Medical University

Here, we present an eCLIP protocol to determine major RNA targets of RBP candidates in testis.

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Genetics

In Vitro Biochemical Assays using Biotin Labels to Study Protein-Nucleic Acid Interactions
Lina Yu *1, Wenxiu He *1, Jie Xie *1, Rui Guo *1, Juan Ni 2, Xia Zhang 3, Quishi Xu 1, Caifeng Wang 1, Qiuling Yue 1, Fangfang Li 1, Mengcheng Luo 4, Bo Sun 3, Lan Ye 1, Ke Zheng 1
1State Key Laboratory of Reproductive Medicine, Nanjing Medical University, 2The Affiliated Hospital of Hangzhou Normal University, 3School of Life Science and Technology, ShanghaiTech University, 4Department of Tissue and Embryology, School of Basic Medical Sciences, Hubei Provincial Key Laboratory of Developmentally Originated Disease, Wuhan University

Presented here are protocols for in vitro biochemical assays using biotin labels that may be widely applicable for studying protein-nucleic acid interactions.

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Medicine

Murine Appendectomy Model of Chronic Colitis Associated Colorectal Cancer by Precise Localization of Caecal Patch
Yaguang Li 1, Junhong Liu 2,3, Gaixia Liu 1, Zhenhong Pan 2,3, Mingxia Zhang 1, Yao Ma 4, Qingxia Wei 1,5, Hongping Xia 1,6, Rui Xue Zhang 1,2, Junjun She 1
1Department of General Surgery, The First Affiliated Hospital, Xi'an Jiaotong University, 2Institute of Medical Research, Northwestern Polytechnical University, 3School of Life Sciences, Northwestern Polytechnical University, 4Department of Medicine, Xi'an Jiaotong University, 5Princess Margaret Cancer Center, University of Health Network, 6Department of Pathology, Nanjing Medical University

The presented protocol describes a facile surgical removal of the appendix (caecal patch) in a mouse followed by the induction of inflammatory bowel disease-associated colorectal cancer. This murine appendectomy model enables investigation of the biological role of the appendix in the pathogenesis of human gastrointestinal disease.

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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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Biology

Quantification of Circulating Pig-Specific DNA in the Blood of a Xenotransplantation Model
Yangyang Deng *1,2, Ming Zhou *1,3, Ying Lu 1, Jiao Chen 1, Zuhui Pu 4, Dongjing Yu 1,5, Yifan Dai 6, Yongqiang Zhan 7, Lisha Mou 1
1Shenzhen Xenotransplantation Medical Engineering Research and Development Center, Institute of Translational Medicine, Shenzhen University Health Science Center, Shenzhen University School of Medicine, First Affiliated Hospital of Shenzhen University, Shenzhen Second People's Hospital, 2Xianning Hospital of Traditional Chinese Medicine, 3Liver-biotechnology (Shenzhen) Co., Ltd., 4Department of Radiology, Shenzhen University Health Science Center, Shenzhen University School of Medicine, First Affiliated Hospital of Shenzhen University, Shenzhen Second People's Hospital, 5Department of Life Sciences, University of Toronto, 6Jiangsu Key Laboratory of Xenotransplantation, Nanjing Medical University, 7Department of Hepatopancreatobiliary Surgery, Shenzhen University Health Science Center, Shenzhen University School of Medicine, First Affiliated Hospital of Shenzhen University, Shenzhen Second People's Hospital

In this protocol, porcine specific primers were designed, plasmids-containing porcine specific DNA fragments were constructed, and standard curves for quantitation were established. Using species-specific primers, cpsDNA was quantified by qPCR in pig-to-mouse cell transplantation models and pig-to-monkey artery patch transplantation models.

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Cancer Research

A Rapid Screening Workflow to Identify Potential Combination Therapy for GBM using Patient-Derived Glioma Stem Cells
Ziyi Hu *1, Tingting Zhou *1, Fangrong Wu 1, Fan Lin 1,2
1Department of Cell Biology, School of Basic Medical Sciences, Nanjing Medical University, 2Institute for Brain Tumors & Key Laboratory of Rare Metabolic Diseases, Nanjing Medical University; Nanjing Medical University Affiliated Cancer Hospital; Key Laboratory of Human Functional Genomics of Jiangsu Province

The glioma stem cells (GSCs) are a small fraction of cancer cells which play essential roles in tumor initiation, angiogenesis, and drug resistance in glioblastoma (GBM), the most prevalent and devastating primary brain tumor. The presence of GSCs makes the GBM very refractory to most of individual targeted agents, so high-throughput screening methods are required to identify potential effective combination therapeutics. The protocol describes a simple workflow to enable rapid screening for potential combination therapy with synergistic interaction. The general steps of this workflow consist of establishing luciferase-tagged GSCs, preparing matrigel coated plates, combination drug screening, analyzing, and validating the results.

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Medicine

Creating Radio-cephalic Arteriovenous Fistula in the Forearm with a Modified No-Touch Technique
Zhang Yuanyuan *1, Wang Xiaohe *1, Liu Zhen 1, Hou Guocun 1
1Department of Nephrology, The Affiliated Suzhou Science & Technology Town Hospital, Nanjing Medical University

We present a modified no-touch technique (MNTT) to create a radio-cephalic arteriovenous fistula (RC-AVF) in which the venous and arterial wall avoid devascularization and the radial artery does not sever.

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Developmental Biology

Isolation and Culture of Three Kinds of Umbilical Cord Mesenchymal Stem Cells
Yanjun Yang *1, Chunxue Zhang *2, Xiaoqiang Sheng 3
1Department of Obstetrics and Gynecology, the Third Affiliated Hospital, Soochow University, 2Department of Obstetrics and Gynecology, Nanjing First Hospital, Nanjing Medical University, 3Center for Reproductive Medicine and Obstetrics and Gynecology, Nanjing Drum Tower Hospital, Nanjing University Medical School

The present protocol describes the isolation and culture of mesenchymal stem cells from the umbilical cord arteries, vein, and Wharton's jelly.

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Developmental Biology

Mitochondrial Transfer from Mouse Adipose-Derived Mesenchymal Stem Cells into Aged Mouse Oocytes
Yanjun Yang 1, Chunxue Zhang 2, Xiaoqiang Sheng 3
1Department of Obstetrics and Gynecology, the Third Affiliated Hospital, Soochow University, 2Department of Obstetrics and Gynecology, Nanjing First Hospital, Nanjing Medical University, 3Center for Reproductive Medicine and Obstetrics and Gynecology, Nanjing Drum Tower Hospital, Nanjing University Medical School

Here, we describe the isolation of mitochondria from mouse adipose-derived mesenchymal stem cells, and then transfer the mitochondria into aged mouse oocytes to improve the quality of the oocytes.

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Developmental Biology

Generation of a Mouse Artificial Decidualization Model with Ovariectomy for Endometrial Decidualization Research
Yang Zhang *1,2,3, Zhe Zhang *1,2,3, Nannan Kang 1,2,3, Xiaoqiang Sheng 1,2,3
1Center for Reproductive Medicine and Obstetrics and Gynecology, The Affiliated Drum Tower Hospital of Nanjing University Medical School, 2State Key Laboratory of Reproductive Medicine, Nanjing Medical University, 3Center for Molecular Reproductive Medicine, Nanjing University

Here, we describe the method of generating an artificial decidualization model using the ovariectomized mouse, a classic endometrial decidualization experiment in the research field of endometrial decidualization.

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Cancer Research

Glioblastoma Relapse Post-Resection Model for Therapeutic Hydrogel Investigations
Shuo Sun *1, Dongyan Shi *1, Jian Liu 1, JinYing Lu 1, Pingnan Dou 1, Zewei Zhou 1, Yun Chen 1,2
1Department of Immunology, Key Laboratory of Human Functional Genomics of Jiangsu Province, Gusu School, Nanjing Medical University, 2Jiangsu Key Lab of Cancer Biomarkers, Prevention and Treatment, Collaborative Innovation Center for Cancer Personalized Medicine, Nanjing Medical University

The present protocol establishes a glioblastoma (GBM) relapse post-resection model using microscopy to investigate the therapeutic effect of an injectable, bioresponsive hydrogel in vivo.

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Cancer Research

Development of a 68Gallium-Labeled D-Peptide PET Tracer for Imaging Programmed Death-Ligand 1 Expression
Lulu Zhang *1, Siqi Zhang *2, Wenyu Wu 1, Xingkai Wang 2, Jieting Shen 2, Dongyuan Wang 4, Kuan Hu *2,3, Ming-Rong Zhang *3, Feng Wang *1, Rui Wang *2
1Department of Nuclear Medicine, Nanjing First Hospital, Nanjing Medical University, 2State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, 3Department of Advanced Nuclear Medicine Sciences, Institute for Quantum Medical Science, National Institutes for Quantum Science and Technology, 4Department of Pharmacy, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology

This study developed a noninvasive and real-time method to evaluate the distribution of programmed death-ligand 1 in the whole body, based on positron emission tomographic imaging of [68Ga] D-dodecapeptide antagonist. This technique has advantages over conventional immunohistochemistry and improves the efficiency of identifying appropriate patients who will benefit from immune checkpoint blockade therapy.

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Biology

Establishment of Facial Nerve Injury Rat Model for Idiopathic Facial Paralysis Research
Xie Yandong 1,2, Ying Tingting 1,2, Liu Liang 3, Liu Mingxi 3, Wang Ran 3, Li Shiting 1,2, Xin Zhang 1,2
1Department of Neurosurgery, Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, 2The Cranial Nerve Disease Center, Shanghai Jiaotong University, 3Department of Neurosurgery, Affiliated Nanjing Brain Hospital, Nanjing Medical University

The present protocol establishes a facial nerve injury rat model using microscopy to investigate the diagnostic and therapeutic mechanisms of idiopathic facial paralysis.

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Engineering

Swin-PSAxialNet: An Efficient Multi-Organ Segmentation Technique
Lei Ma 1, Dian Zhang 1, Zhaoxin Wang 1, Jincong Han 1, Xing Wang 1, Xin Zhou 2, Wenwen Yang 1, Peihua Lu 2
1School of Information Science and Technology, Nantong University, 2The Oncology and Hematology Department of Wuxi People's Hospital of Nanjing Medical University, Wuxi Medical Center, Nanjing Medical University

The present protocol describes an efficient multi-organ segmentation method called Swin-PSAxialNet, which has achieved excellent accuracy compared to previous segmentation methods. The key steps of this procedure include dataset collection, environment configuration, data preprocessing, model training and comparison, and ablation experiments.

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