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October, 2006
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Diabetes Mellitus: A heterogeneous group of disorders characterized by Hyperglycemia and Glucose intolerance.
 JoVE Medicine

A Zebrafish Model of Diabetes Mellitus and Metabolic Memory

1Dr. William M. Scholl College of Podiatric Medicine, Rosalind Franklin University of Medicine and Science, 2Chicago Medical School, Rosalind Franklin University of Medicine and Science

JoVE 50232

Metabolic memory is the phenomenon by which diabetic complications persist and progress unimpeded even after euglycemia is achieved pharmaceutically. Here we describe a diabetes mellitus zebrafish model which is unique in that it allows for the examination of the mitotically transmissible epigenetic components of metabolic memory in vivo.

 JoVE Medicine

Experimental and Imaging Techniques for Examining Fibrin Clot Structures in Normal and Diseased States

1Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology & Emory University School of Medicine, 2Parker H. Petit Institute for Bioengineering & Bioscience, Georgia Institute of Technology, 3George W. Woodruff School of Mechanical Engineering, Georgia Institute of Technology

JoVE 52019

In this manuscript, experimental techniques, including blood preparation, confocal microscopy, and lysis rate analysis, to examine the morphological differences between normal and abnormal clot structures due to diseased states are presented.

 JoVE Biology

A Method for Mouse Pancreatic Islet Isolation and Intracellular cAMP Determination

1Department of Nutrional Sciences, University of Wisconsin-Madison, 2Department of Medicine, Division of Endocrinology, Diabetes, and Metabolism, University of Wisconsin-Madison, 3School of Pharmacy, University of Waterloo

JoVE 50374

Assaying in vitro β-cell function using isolated mouse islets of Langerhans is an important component in the study of diabetes pathophysiology and therapeutics. While many downstream applications are available, this protocol specifically describes the measurement of intracellular cyclic adenosine monophosphate (cAMP) as an essential parameter determining β-cell function.

 JoVE Medicine

Ultrasound Assessment of Endothelial-Dependent Flow-Mediated Vasodilation of the Brachial Artery in Clinical Research

1Department of Surgery, University of California, San Francisco, 2Department of Surgery, Veterans Affairs Medical Center, San Francisco, 3VipeRx Lab, Veterans Affairs Medical Center, San Francisco

JoVE 52070

Endothelial dysfunction is associated with numerous disease states and is predictive of adverse cardiovascular events in humans. Flow-mediated vasodilation (FMD) is a non-invasive ultrasound method of evaluating endothelial function. Methodological choices and operator experience may affect results. A systematic approach to FMD in human studies is discussed here.

 JoVE Medicine

Isolation of Human Islets from Partially Pancreatectomized Patients

1Department of GI-, Thorax- and Vascular Surgery, University Hospital Carl Gustav Carus, University of Technology Dresden, 2Molecular Diabetology, Paul Langerhans Institute Dresden, 3Department of Pathology, University Hospital Carl Gustav Carus, University of Technology Dresden

JoVE 2962

The supply of type 2 diabetic islets for research is insufficient. Here we share our protocol for isolating islets from patients undergoing partial pancreatectomy. This approach represents a unique venue for obtaining islets from type 2 diabetic and clinically matched non-diabetic subjects in adequate numbers for basic and clinical studies.

 JoVE Medicine

A Method for Murine Islet Isolation and Subcapsular Kidney Transplantation

1Molecular and Cellular Biochemistry, Center for Molecular Neurobiology, The Ohio State University, 2Integrated Biomedical Science Graduate Program, The Ohio State University, 3Comprehensive Cancer Center, The Ohio State University

JoVE 2096

Transplantation of isolated islets has been proposed to be a potential treatment for type 1 diabetes. Here we describe a method to isolate islets from mouse pancreata and transplant them to the subcapsular space of the kidney.

 JoVE Biology

A High-throughput Method for Measurement of Glomerular Filtration Rate in Conscious Mice

1Division of Nephrology-Hypertension, Department of Medicine, University of California, San Diego, 2San Diego VA Healthcare System

JoVE 50330

Measurement of glomerular filtration rate (GFR) is the gold standard for kidney function assessment. Here we describe a high-throughput method which allows the determination of GFR in conscious mice by using a single bolus injection, determination of fluorescein isothiocyanate (FITC)-inulin in plasma and calculation of GFR by a two-phase exponential decay model.

 JoVE Immunology and Infection

Accelerated Type 1 Diabetes Induction in Mice by Adoptive Transfer of Diabetogenic CD4+ T Cells

1Department of Microbiology & Immunology, Pennsylvania State University College of Medicine

JoVE 50389

We provide a reproducible method to induce type 1 diabetes (T1D) in mice within two weeks by the adoptive transfer of islet antigen-specific, primary CD4+ T cells.

 JoVE Medicine

Murine Model of Wound Healing

1The Heart Research Institute, 2Sydney Medical School, University of Sydney, 3Cardiology Department, Royal Prince Alfred Hospital

JoVE 50265

A murine model of cutaneous wound healing that can be used to assess therapeutic compounds in physiological and pathophysiological settings.

 JoVE Medicine

Near Infrared Optical Projection Tomography for Assessments of β-cell Mass Distribution in Diabetes Research

1Umeå Centre for Molecular Medicine, Umeå University, 2Cell Transplant Center, Diabetes Research Institute, University of Miami,, 3EMBL-CRG Systems Biology Program, Centre for Genomic Regulation, Catalan Institute of Research and Advanced Studies, 4Dept. of Computing Science, Umeå University

JoVE 50238

We describe the adaptation of optical projection tomography (OPT)1 to imaging in the near infrared spectrum, and the implementation of a number of computational tools. These protocols enable assessments of pancreatic β-cell mass (BCM) in larger specimens, increase the multichannel capacity of the technique and increase the quality of OPT data.

 JoVE Medicine

Leprdb Mouse Model of Type 2 Diabetes: Pancreatic Islet Isolation and Live-cell 2-Photon Imaging Of Intact Islets

1School of Biomedical Sciences, The University of Queensland

JoVE 52632

We present here a protocol for the isolation of islets from the mouse model of type 2 diabetes, Leprdb and details of a live-cell assay for measurement of insulin secretion from intact islets that utilizes 2 photon microscopy.

 JoVE Biology

Protein WISDOM: A Workbench for In silico De novo Design of BioMolecules

1Department of Chemical and Biological Engineering, Princeton University

JoVE 50476

We developed computational de novo protein design methods capable of tackling several important areas of protein design. To disseminate these methods we present Protein WISDOM, an online tool for protein design ( Starting from a structural template, design of monomeric proteins for increased stability and complexes for increased binding affinity can be performed.

 JoVE Medicine

A Simplified Technique for Producing an Ischemic Wound Model

1Department of Surgery, University of Louisville

JoVE 3341

We have developed a minimally invasive technique to create a rabbit ischemic ear wound model by dividing the central artery and nerve and the cranial neurovascular bundle. A subcutaneous tunnel then cuts all subcutaneous tissues. This procedure causes minimal skin disruption and can be safely used in diabetic animals.

 JoVE Medicine

Hyperinsulinemic-Euglycemic Clamp in the Conscious Rat

1Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Calgary, 2Faculty of Kinesiology, University of Calgary

JoVE 2432

The hyperinsulinemic-euglycemic clamp is the "gold standard" for the assessment of insulin action. Insulin is infused at a constant rate stimulating glucose uptake. The amount of exogenous glucose infused to counter this drop is indicative of insulin sensitivity. Here the procedure is performed on a conscious, unrestrained rat.

 JoVE Medicine

Murine Spinotrapezius Model to Assess the Impact of Arteriolar Ligation on Microvascular Function and Remodeling

1Department of Biomedical Engineering, University of Virginia, 2Department of Biomedical Engineering, California Polytechnic State University, 3Office of Animal Welfare, University of Virginia, 4Department of Biomedical Engineering & Institute for Computational Medicine, Johns Hopkins University

JoVE 50218

We demonstrate a novel arterial ligation model in murine spinotrapezius muscle, including a step-by-step procedure and description of required instrumentation. We describe the surgery and relevant outcome measurements relating to vascular network remodeling and functional vasodilation using intravital and confocal microscopy.

 JoVE Medicine

Assessing Replication and Beta Cell Function in Adenovirally-transduced Isolated Rodent Islets

1Department of Pediatrics, Indiana University School of Medicine, 2Department of Cellular & Integrative Physiology, Indiana University School of Medicine

JoVE 4080

This protocol allows one to identify factors that modulate functional beta cell mass to find potential therapeutic targets for the treatment of diabetes. The protocol consists of a streamlined method to assess islet replication and beta cell function in isolated rat islets following manipulation of gene expression with adenoviruses.

 JoVE Medicine

Improved Protocol For Laser Microdissection Of Human Pancreatic Islets From Surgical Specimens

1Molecular Diabetology, Paul Langerhans Institute Dresden, 2Department of GI-, Thoracic- and Vascular Surgery, University Hospital Carl Gustav Carus, University of Technology Dresden, 3Department of Endocrinology and Metabolism, Metabolic Unit University of Pisa, 4Labs DC0522, Lilly Corporate Center, 5Genomics, Faculty of Medicine Imperial College London, 6Vital-IT, SIB Swiss Institute of Bioinformatics, 7Clinical Biochemistry, Hannover Medical School, 8Cell Physiology and Metabolism, Medical School, University of Geneva, 9Department of Pathology, University Hospital Carl Gustav Carus, University of Technology Dresden, 10R&D DIAB Division / Translational Medicine, Sanofi-Aventis

JoVE 50231

Laser microdissection is a technique that allows the recovery of selected cells from minute amounts of parenchyma. Here we describe a protocol for acquiring human pancreatic islets from surgical specimens to be used for transcriptomic studies. Our protocol improves the intrinsic autofluorescence of human beta cells, thus facilitating their collection.

 JoVE Medicine

Transplantation into the Anterior Chamber of the Eye for Longitudinal, Non-invasive In vivo Imaging with Single-cell Resolution in Real-time

1Diabetes Research Institute, University of Miami Miller School of Medicine, 2Department of Surgery, University of Miami Miller School of Medicine, 3Department of Medicine, University of Miami Miller School of Medicine, 4Department of Physiology & Biophysics, University of Miami Miller School of Medicine, 5The Rolf Luft Research Center for Diabetes and Endocrinology, Karolinska Institutet

JoVE 50466

A new approach combining intraocular transplantation and confocal microscopy enables longitudinal, non-invasive real-time imaging with single-cell resolution within grafted tissues in vivo. We demonstrate how to transplant pancreatic islets into the anterior chamber of the mouse eye.

 JoVE Medicine

Encapsulation Thermogenic Preadipocytes for Transplantation into Adipose Tissue Depots

1Department of Human Sciences, The Ohio State University, 2Department of Minimally Invasive Surgery, The First Affiliated Hospital of Soochow University, 3NSF Nanoscale Science and Engineering Center for Affordable Nanoengineering of Polymeric Biomedical Devices, The Ohio State University

JoVE 52806

Here, we present a protocol for encapsulation of catabolic cells, which consume lipids for heat production in intra-abdominal adipose tissue and increase energy dissipation in obese mice.

 JoVE Medicine

Human Brown Adipose Tissue Depots Automatically Segmented by Positron Emission Tomography/Computed Tomography and Registered Magnetic Resonance Images

1Chemical and Physical Biology Program, Vanderbilt University, 2Department of Physical Medicine and Rehabilitation, Vanderbilt University School of Medicine, 3Radiology & Radiological Sciences, Vanderbilt University Medical Center, 4Department of Pharmacology, Vanderbilt University

JoVE 52415

The method presented here uses 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography/computed tomography (PET-CT) and fat-water separated magnetic resonance imaging (MRI), each scanned following 2 hr exposure to thermoneutral (24 °C) and cold conditions (17 °C) in order to map brown adipose tissue (BAT) in adult human subjects.

 JoVE Biology

An Assay for Lateral Line Regeneration in Adult Zebrafish

1Department of Biomedical Sciences, Dr. William M Scholl College of Podiatric Medicine, Rosalind Franklin University of Medicine and Science, 2Department of Cell Biology and Anatomy, Rosalind Franklin University of Medicine and Science

JoVE 51343

Because many zebrafish models of neurological and non-neurological diseases are studied in the adult fish rather than the embryo/larvae, we developed a quantitative lateral line regenerative assay that can be applied to adult zebrafish disease models. The assay involved resolution at the 1) neuromast and 2) individual hair cell levels.

 JoVE Neuroscience

Rapid Determination of the Thermal Nociceptive Threshold in Diabetic Rats

1Department of Pharmacology and Toxicology, Wright State University, 2Human Movement Laboratory, Universidade São Judas Tadeu

JoVE 3785

Here, we describe a rapid reliable and simple procedure to determine the lowest temperature at which rats or mice show nocifensive behavior, i.e. the thermal nociceptive threshold (TNT). This method applies a slowly increasing thermal stimulus allowing precise and reproducible estimation of TNTs with minimum, if any, stress to the animals.

 JoVE Immunology and Infection

Methods to Assess Beta Cell Death Mediated by Cytotoxic T Lymphocytes

1Departments of Pathology, Immunology and Laboratory Medicine, College of Medicine, University of Florida

JoVE 2724

Cell-mediated lymphocytotoxicity (CML) assays can be used to test autoreactive responses and study mechanisms of cell death in vitro. However, using live-cell confocal microscopic imaging techniques with fluorescent dyes, the type and kinetics of cell death as well as the pathways utilized can be studied in greater detail.

 JoVE Neuroscience

Membrane Potential Dye Imaging of Ventromedial Hypothalamus Neurons From Adult Mice to Study Glucose Sensing

1Department of Pharmacology and Physiology, Rutgers New Jersey Medical School, 2Center for Taste and Feeding Behavior, Universite de Bourgogne

JoVE 50861

The activity of single neurons from adult-aged mice can be studied by dissociating neurons from specific brain regions and using fluorescent membrane potential dye imaging. By testing responses to changes in glucose, this technique can be used to study the glucose sensitivity of adult ventromedial hypothalamic neurons.

 JoVE Immunology and Infection

Isolation of Adipose Tissue Immune Cells

1Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine

JoVE 50707

Adipose tissue (AT) is a site of intense immune cell activation and interaction. Almost all cells of the immune system are present in AT and their ratios are altered by obesity. Proper isolation, quantification, and characterization of AT immune cell populations are critical for understanding their role in immunometabolic disease.

 JoVE Bioengineering

Controlled Microfluidic Environment for Dynamic Investigation of Red Blood Cell Aggregation

1Department of Mechanical Engineering, University of Ottawa

JoVE 52719

The protocol described details an experimental procedure to quantify Red Blood Cell (RBC) aggregates under a controlled and constant shear rate, based on image processing techniques. The goal of this protocol is to relate RBC aggregate sizes to the corresponding shear rate in a controlled microfluidic environment.

 JoVE Medicine

Isolation, Cryopreservation and Culture of Human Amnion Epithelial Cells for Clinical Applications

1Wake Forest Institute for Regenerative Medicine, Wake Forest University Health Sciences, 2The Ritchie Centre, Monash Institute of Medical Research, Monash University

JoVE 52085

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.

 JoVE Medicine

Cholesterol Efflux Assay

1Baker IDI Heart and Diabetes Institute

JoVE 3810

The cholesterol assay is designed to quantitate the rate of cholesterol efflux from cultured cells and the capacity of plasma acceptors to accept cholesterol released from cells. The assay consists of labelling cells with cholesterol, equilibration of cholesterol among intracellular pools and release of cholesterol to an extracellular acceptor.

 JoVE Medicine

Quantification of Atherosclerotic Plaque Activity and Vascular Inflammation using [18-F] Fluorodeoxyglucose Positron Emission Tomography/Computed Tomography (FDG-PET/CT)

1Division of Cardiovascular Medicine, University of Pennsylvania, Perelman School of Medicine, 2Department of Radiology, University of Pennsylvania, Perelman School of Medicine, 3Department of Dermatology, University of Pennsylvania, Perelman School of Medicine

JoVE 3777

There is great need to identify atherosclerosis non-invasively, and here we demonstrate how FDG-PET/CT can be used to detect and quantify atherosclerotic plaque activity and vascular inflammation.

 JoVE Biology

Metabolic Profile Analysis of Zebrafish Embryos

1Metabolic Research Unit & Molecular & Medical Research SRC, Geelong, Australia, School of Medicine, Deakin University

JoVE 4300

Zebrafish represent a powerful vertebrate model that has been under-utilised for metabolic studies. Here we describe a rapid way to measure the in vivo metabolic profile of developing zebrafish that allows the comparison of different mitochondrial function parameters between genetically or pharmacologically manipulated embryos, thereby increasing the applicability of this organism.

 JoVE Bioengineering

Quantitative and Temporal Control of Oxygen Microenvironment at the Single Islet Level

1Department of Mechanical Engineering, University of Michigan-Dearborn, 2Department of Surgery/Transplant, University of Illinois at Chicago, 3Department of Bioengineering, University of Illinois at Chicago

JoVE 50616

Microfluidic oxygen control confers more than just convenience and speed over hypoxic chambers for biological experiments. Especially when implemented via diffusion through a membrane, microfluidic oxygen can provide simultaneous liquid and gas phase modulations at the microscale-level. This technique enables dynamic multi-parametric experiments critical for studying islet pathophysiology.

 JoVE Immunology and Infection

Detecting Glycogen in Peripheral Blood Mononuclear Cells with Periodic Acid Schiff Staining

1Department of Biology, Centre for Structural and Functional Genomics, PERFORM Centre, Concordia University, 2Department of Chemistry and Biochemistry, Centre for Structural and Functional Genomics, PERFORM Centre, Concordia University, 3Department of Exercise Science, Centre for Structural and Functional Genomics, PERFORM Centre, Concordia University

JoVE 52199

Periodic acid Schiff staining is a technique that visualizes the polysaccharide content of tissues. This article demonstrates periodic acid Schiff staining protocol adapted for use on peripheral blood mononuclear cells purified from human venous blood. Such samples are enriched for lymphocytes and other white blood cells of the immune system.

 JoVE Biology

Human Pancreatic Islet Isolation: Part I: Digestion and Collection of Pancreatic Tissue

1Department of Surgery, University of Illinois, Chicago

JoVE 1125

Achieving high quality and appropriate quantity of human islets is one of the prominent prerequisites for successful islet transplantation. In this video, we describe step by step the procedures for human pancreatic islet isolation (part I: digestion and collection of pancreatic tissue) using a modified automated method.

 JoVE Bioengineering

Tracking Hypoxic Signaling within Encapsulated Cell Aggregates

1Biomedical Engineering Program, University of South Carolina, 2Chemical Engineering Department, University of South Carolina

JoVE 3521

A method for photo-encapsulation of cells in a crosslinked PEG hydrogel is described. Hypoxic signaling within encapsulated murine insulinoma (MIN6) aggregates is tracked using a fluorescent marker system. This system allows serial examination of cells within a hydrogel scaffold and correlation of hypoxic signaling with changes in cell phenotype.

 JoVE Biology

Human Pancreatic Islet Isolation: Part II: Purification and Culture of Human Islets

1Department of Surgery, University of Illinois, Chicago

JoVE 1343

Achieving high quality and appropriate quantity of human islets is one of the prominent prerequisites for successful islet transplantation. In this video, we describe step by step the procedures for human pancreatic islet isolation (part II: purification and culture of human islets) using a modified automated method.

 JoVE Medicine

Measuring Frailty in HIV-infected Individuals. Identification of Frail Patients is the First Step to Amelioration and Reversal of Frailty

1Division of Infectious Diseases, University of Arizona, 2Arizona Center on Aging, University of Arizona

JoVE 50537

Frailty syndrome is commonly seen in the aged and reflects multi-system physiological change. However, with reduced functional reserve and resilience frailty is also known to be common in the HIV infected population. This study outlined an easily administered screening test to identify HIV patients with frailty. When significant components of frailty are identified, clinicians will be able to focus on amelioration of the problem and promote reversion to the pre-frail state.

 JoVE Medicine

The α-test: Rapid Cell-free CD4 Enumeration Using Whole Saliva

1Department of Medicine, Weill Cornell Medical College, 2Department of Oral Biology, University of Missouri-Kansas City-School of Dentistry, 3Department of Pharmacology and Toxicology, University of Missouri Kansas City- School of Pharmacy, 4Regional Hospital, Bamenda, NWP, Cameroon, 5Mezam Polyclinic HIV/AIDS Treatment Center, Cameroon, 6Institute for Human Genetics and Biochemistry

JoVE 3999

A CD4 enumeration method, the α-test, is described which uses whole saliva to provide rapid and accurate CD4 counts. The α-test costs pennies and eliminates the need for technical training, costly reagents such as monoclonal antibodies, instrumentation, refrigeration, transport of samples, as well as collection and handling of blood.

 JoVE Biology

Endothelial Cell Co-culture Mediates Maturation of Human Embryonic Stem Cell to Pancreatic Insulin Producing Cells in a Directed Differentiation Approach

1Department of Bioengineering, University of Pittsburgh, 2Department of Chemical Engineering, University of Pittsburgh

JoVE 3759

The current study describes a directed differentiation approach in inducing pancreatic differentiation of human embryonic stem cells. Of great significance is the finding that endothelial cell co-culture mediates maturation of human embryonic stem cell derived pancreatic progenitors into insulin expressing cells.

 JoVE Bioengineering

Alginate Microcapsule as a 3D Platform for Propagation and Differentiation of Human Embryonic Stem Cells (hESC) to Different Lineages

1Stem Cell Lab, School of Psychiatry, Faculty of Medicine, The University of New South Wales, 2Siriraj Center of Excellence for Stem cell Research, Faculty of Medicine Siriraj Hospital, Mahidol University, 3Neuropsychiatric Institute, Prince of Wales Hospital

JoVE 3608

We have optimized a microencapsulation technique as an effective 3D platform for propagation and differentiation of embryonic stem cells to endoderm and dopaminergic (DA) neurons. It also provides an opportunity for immune-isolation of cells from the host during transplantation. This platform can be adapted for other cell types.

 Science Education: Essentials of Biology 2

Zebrafish Maintenance and Husbandry

JoVE Science Education

The zebrafish (Danio rerio) is a powerful vertebrate model system for studying development, modeling disease, and screening for novel therapeutics. Due to their small size, large numbers of zebrafish can be housed in the laboratory at low cost. Although zebrafish are relatively easy to maintain, special consideration must be given to both diet and water quality to in order to optimize fish health and reproductive success. This video will provide an overview of zebrafish husbandry and maintenance in the lab. After a brief review of the natural zebrafish habitat, techniques essential to recreating this environment in the lab will be discussed, including key elements of fish facility water recirculation systems and the preparation of brine shrimp as part of the zebrafish diet. Additionally, the presentation will include information on how specific zebrafish strains are tracked in a laboratory setting, with specific reference to the collection of tail fin samples for DNA extraction and genotyping. Finally, experimental modifications of the zebrafish environment will be discussed as a means to further our understanding of these fish, and in turn, ourselves.

 JoVE Biology

Comet Assay as an Indirect Measure of Systemic Oxidative Stress

1Department of Ophthalmology, University of Basel, 2Department of Biomedicine, University of Basel

JoVE 52763

Comet assay measures DNA breaks, induced by different factors. If all factors (except oxidative stress) causing DNA damage are kept constant, the amount of DNA damage is a good indirect parameter of oxidative stress. The goal of this protocol is to use comet assay for indirect measurement of oxidative stress.

 JoVE Medicine

Improving IV Insulin Administration in a Community Hospital

1Wyoming Medical Center

JoVE 3705

When compared to the previous paper protocol, implementation of a computerized glucose management system results in a substantial increase in blood glucose concentration measurements within the target range. Using a computerized glucose management system to monitor blood glucose levels, decreases in severe hypoglycemia (<40 mg/dL), clinical hypoglycemia (<70 mg/dL), and hyperglycemia (>180 mg/dL) also can be observed.

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