Methods of Adipose Tissue Biology Part B - 1st Edition - ISBN: 9780128002803, 9780128003367

Methods of Adipose Tissue Biology Part B, Volume 538

1st Edition

Methods of Adipose Tissue Biology

Serial Volume Editors: Ormond Macdougald
eBook ISBN: 9780128003367
Hardcover ISBN: 9780128002803
Imprint: Academic Press
Published Date: 1st February 2014
Page Count: 344
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Table of Contents

Contributors
Preface
Chapter One: Preparation and Differentiation of Mesenchymal Stem Cells from Ears of Adult Mice
Abstract

1 Introduction

2 EMSC Isolation, Culture, Cryopreservation, and Differentiation

3 Conclusions

Acknowledgments

Chapter Two: 3-D Adipocyte Differentiation and Peri-adipocyte Collagen Turnover
Abstract

1 Introduction

2 Materials and Methods

Acknowledgment

Chapter Three: Differentiation of White and Brown Adipocytes from Human Pluripotent Stem Cells
Abstract

1 Introduction

2 Human Pluripotent Stem Cells

3 Differentiation into Mesenchymal Progenitor Cells

4 Differentiation into Adipocytes

Chapter Four: Optimal Protocol for the Differentiation and Metabolic Analysis of Human Adipose Stromal Cells
Abstract

1 Introduction

2 Isolation of ASCs from AT

3 Protocols for Proliferation, Subculture, and Freezing Down

4 Protocols to Differentiate Human ASCs to Adipocytes

5 Methods to Use Newly Differentiated Adipocytes for Metabolic Studies

Acknowledgments

Chapter Five: Human Adipose-Derived Stromal/Stem Cell Isolation, Culture, and Osteogenic Differentiation
Abstract

1 Introduction

2 Methods for Cell Culture

3 Methods for Scaffold Preparation

4 Material Characterization

5 Cell Loading and Scaffold Culture

6 Cell Viability and Proliferation

7 Assessing Osteogenic Differentiation

Chapter Six: Analysis of Adipose Tissue Lipid Using Mass Spectrometry
Abstract

1 Introduction

2 Neutral Lipids

3 Phospholipids

4 Summary

Acknowledgments

Chapter Seven: Measurement of Long-Chain Fatty Acid Uptake into Adipocytes
Abstract

1 Introduction

2 Adipocyte Sources

3 Uptake Assay with Radioactive Fatty Acids

4 Uptake Assay with Flow Cytometer

5 Quencher-Based LCFA Uptake Assay

6 Bioluminescent Fatty Acid Uptake Assays

7 In vivo Imaging of BAT Fatty Acid Uptake Rates

Acknowledgments

Chapter Eight: Measurement of the Unfolded Protein Response to Investigate Its Role in Adipogenesis and Obesity
Abstract

1 Introduction

2 Methods to Measure UPR Induction in Adipose Tissues

3 Methods to Investigate Role of UPR in Metabolic Phenotypes of Adipocytes

Acknowledgment

Chapter Nine: Application of Activity-Based Protein Profiling to Study Enzyme Function in Adipocytes
Abstract

1 Introduction

2 Technical Aspects

3 Discussion

Chapter Ten: Measurement of Lipolysis
Abstract

1 Introduction

2 Methodological Description of Activity-Based Assays

3 Conclusion

Acknowledgments

Chapter Eleven: Measurement of Lipolysis Products Secreted by 3T3-L1 Adipocytes Using Microfluidics
Abstract

1 Introduction

2 Methods

3 Discussion

4 Design and Operation Limitations

5 Continued Improvements

6 Notes

Chapter Twelve: Methods for Performing Lipidomics in White Adipose Tissue
Abstract

1 Introduction

2 Tissue Extraction of Adipose Tissue and Adipocytes

3 GC–MS of Total Fatty Acids

4 LC–MS of Intact Lipids

5 Conclusions

Chapter Thirteen: Measuring Respiratory Activity of Adipocytes and Adipose Tissues in Real Time
Abstract

1 Introduction

2 Basis of the Methods and the XF24

3 Cell Culture and Tissue Isolation

4 Preparing the XF24 Analyzer Setup

5 Important Remarks

Acknowledgment

Chapter Fourteen: Detecting Protein Carbonylation in Adipose Tissue and in Cultured Adipocytes
Abstract

1 Introduction

2 Methods to Analyze Protein Carbonylation

3 Results

4 Discussion

Chapter Fifteen: Use of Fluorescence Microscopy to Probe Intracellular Lipolysis
Abstract

1 Introduction

2 Methods to Image and Colocalize Endogenous Lipolytic Effectors in Adipocytes

3 Imaging Protein Trafficking and Protein–Protein Interactions in Live Model Cells

4 Imaging Intracellular Fatty Acid Production

Acknowledgments

Chapter Sixteen: Evaluation of Protein Phosphorylation During Adipogenesis
Abstract

1 Introduction

2 Phosphoproteomic Analysis

3 Western Blot Analysis

4 Phosphorylation Analysis by In Vitro Radiolabeling

5 Enzyme-Linked Immunosorbent Assay

6 Mutation of Phosphorylation Sites

Acknowledgments

Author Index
Subject Index


Description

This book is a must-have for anyone interested in obesity or the physiology of white or brown adipose tissues. It contains state-of-the-art methods from researchers that are world leaders in this field. Detailed lab protocols range from methods to visualize adipocytes and adipose tissues in humans and experimental models, to convert stem cells into white and brown adipocytes in vitro, to evaluate aspects of adipocyte metabolism, to inducibly knock out genes in adipose tissues, and to evaluate transcriptional control of adipogenesis on a global scale.

Key Features

  • The study of adipose tissue goes hand in hand with our global effort to understand and reverse the epidemic of obesity and associated medical complications
  • Contributors include leading researchers who have made tremendous contributions to our ability to investigate white and brown adipose tissues
  • The wide variety of experimental approaches detailed within this volume: including the evaluation of adipose tissue biology at the molecular, biochemical, cellular, tissue, and organismal levels

Readership

Investigators and students whose research interests include the development, physiology, or pathophysiology of adipose tissues


Details

No. of pages:
344
Language:
English
Copyright:
© Academic Press 2014
Published:
Imprint:
Academic Press
eBook ISBN:
9780128003367
Hardcover ISBN:
9780128002803

About the Serial Volume Editors

Ormond Macdougald Serial Volume Editor

Ormond A. MacDougald, Ph.D. is the John A. Faulkner Collegiate Professor in Physiology, and a Professor of Molecular & Integrative Physiology, and Internal Medicine in the University of Michigan Medical School. He currently is a Fulbright Scholar at Pembroke College and the Institute of Metabolic Science, University of Cambridge.

MacDougald is an internationally recognized investigator for his work on adipocyte differentiation and metabolism. Specifically, his research explores the signals that act on mesenchymal stem cells to influence fat tissue development and metabolism, including effects on insulin sensitivity. This research provides important insight into the problems of obesity and type 2 diabetes. MacDougald’s bibliography reflects more than 100 peer-reviewed journal articles, reviews, and book chapters. MacDougald serves on numerous editorial boards and provides extensive peer-review service for journals and funding agencies. He’s a highly sought-after speaker at national and international meetings.

In 2005, MacDougald received the UM Medical School’s Achievement in Basic Science Research Award. The same year, he earned the Henry Pickering Bowditch Award, one of the American Physiological Society’s highest honors, given to “a distinguished young physiologist less than 42 years of age who has made original and outstanding contributions in physiology.” More recently he was elected Fellow of the American Association for the Advancement of Science.

In addition to his research accomplishments, MacDougald is highly regarded for his dedication and commitment to education. He has served in numerous educational roles as director, co-director and lecturer in departmental courses, and has been a member of more than 80 preliminary examination and graduate dissertation committees. He served as director of the Molecular & Integrative Physiology graduate program, and he initiated and directs a summer research program for undergraduate students. He received the Rackham Distinguished Graduate Mentoring Award, and was elected to the League of Excellence in Education at the University of Michigan.

Ormond received his undergraduate degree from the University of Guelph in Ontario, Canada, in 1986. From Michigan State University he received his master’s degree in 1988 and a doctorate from the Department of Physiology in 1992. He pursued postdoctoral training from 1992-96 in the Department of Biological Chemistry at Johns Hopkins University, where he began his studies on adipocyte biology with M. Daniel Lane, Ph.D.

MacDougald joined the U-M faculty in 1996 as an Assistant Professor of Physiology and was promoted to Associate Professor in 2002 and to Professor in 2006.

Affiliations and Expertise

John A. Faulkner Collegiate Professor in Physiology, and a Professor of Molecular & Integrative Physiology, and Internal Medicine, University of Michigan Medical School