Use of osmium tetroxide staining with microcomputerized tomography to visualize and quantify bone marrow adipose tissue in vivo.
Adipocytes reside in discrete, well-defined depots throughout the body. In addition to mature adipocytes, white adipose tissue depots are composed of many cell types, including macrophages, endothelial cells, fibroblasts, and stromal cells, which together are referred to as the stromal vascular fraction (SVF). The SVF also contains adipocyte progenitors that give rise to mature adipocytes in those depots. Marrow adipose tissue (MAT) or marrow fat has long been known to be present in bone marrow (BM) but its origin, development, and function remain largely unknown. Clinically, increased MAT is associated with age, metabolic diseases, drug treatment, and marrow recovery in children receiving radiation and chemotherapy. In contrast to the other depots, MAT is unevenly distributed in the BM of long bones. Conventional quantitation relies on sectioning of the bone to overcome issues with distribution but is time-consuming, resource intensive, inconsistent between laboratories and may be unreliable as it may miss changes in MAT volume. Thus, the inability to quantitate MAT in a rapid, systematic, and reproducible manner has hampered a full understanding of its development and function. In this chapter, we describe a new technique that couples histochemical staining of lipid using osmium tetroxide with microcomputerized tomography to visualize and quantitate MAT within the medullary canal in three dimensions. Imaging of osmium staining provides a high-resolution map of existing and developing MAT in the BM. Because this method is simple, reproducible, and quantitative, we expect it will become a useful tool for the precise characterization of MAT.
Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan, USA.
Erica L
EL
Scheller
Department of Orthopaedics and Rehabilitation, Yale University School of Medicine, New Haven, Connecticut, USA.
Nancy
N
Troiano
Department of Internal Medicine, Endocrinology, Yale University School of Medicine, New Haven, Connecticut, USA.
Joshua N
JN
Vanhoutan
Center for Advanced Orthopedic Studies, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts, USA.
Mary A
MA
Bouxsein
Department of Orthopaedics and Rehabilitation, Yale University School of Medicine, New Haven, Connecticut, USA.
Jackie A
JA
Fretz
Department of Orthopaedics and Rehabilitation, Yale University School of Medicine, New Haven, Connecticut, USA.
Yougen
Y
Xi
Department of Orthopaedics and Rehabilitation, Yale University School of Medicine, New Haven, Connecticut, USA.
Tracy
T
Nelson
Department of Orthopaedics and Rehabilitation, Yale University School of Medicine, New Haven, Connecticut, USA.
Griffin
G
Katz
Section of Comparative Medicine, Yale University School of Medicine, New Haven, Connecticut, USA.
Ryan
R
Berry
Section of Comparative Medicine, Yale University School of Medicine, New Haven, Connecticut, USA.
Christopher D
CD
Church
Maine Medical Center Research Institute, Scarborough, Maine, USA.
Casey R
CR
Doucette
Section of Comparative Medicine, Yale University School of Medicine, New Haven, Connecticut, USA.
Matthew S
MS
Rodeheffer
Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan, USA.
Ormond A
OA
Macdougald
Maine Medical Center Research Institute, Scarborough, Maine, USA.
Clifford J
CJ
Rosen
Department of Orthopaedics and Rehabilitation, Yale University School of Medicine, New Haven, Connecticut, USA. Electronic address: mark.horowitz@yale.edu.
Mark C
MC
Horowitz
D050156:Adipogenesis / Q000235:genetics; D052436:Adipose Tissue, White / Q000254:growth & development; D001853:Bone Marrow / Q000254:growth & development; D002454:Cell Differentiation; D006801:Humans; D009993:Osmium Tetroxide / Q000737:chemistry*; D013194:Staining and Labeling / Q000379:methods*; D017154:Stromal Cells / Q000166:cytology; D055114:X-Ray Microtomography / Q000379:methods*
D009993:Osmium Tetroxide
Bone marrow fat; Imaging bone marrow adipose tissue; In vivo fat measurement; Mapping of bone marrow adipose tissue; Marrow adipose tissue; Microcomputerized tomography; Osmium tetroxide; Yellow marrow