Isolation of osteocytes from human trabecular bone.
Osteocytes are essential regulators of bone homeostasis. However, they are difficult to study due to their location within the bone mineralised matrix. Although several techniques have been published for the isolation of osteocytes from mouse bone, no such technique has been described for human osteocytes. We have therefore developed a protocol for the isolation of osteocytes from human trabecular bone samples acquired during surgery. The cells were digested from the bone matrix by sequential collagenase and ethylenediaminetetraacetic acid (EDTA) digestions and the cells from later digests displayed characteristic dendritic osteocyte morphology when cultured ex vivo. Furthermore, the cells expressed characteristic osteocyte marker genes, such as E11, dentin matrix protein 1 (DMP1), SOST, matrix extracellular phosphoglycoprotein (MEPE) and phosphate regulating endopeptidase homologue, X-linked (PHEX). In addition, genes associated with osteocyte perilacunar remodelling, including matrix metallopeptidase-13 (MMP13), cathepsin K (CTSK) and carbonic anhydrase 2 (CAR2) were expressed. The cells also responded to parathyroid hormone (PTH) by downregulating SOST mRNA expression and to 1α,25-dihydroxyvitamin D3 (1,25D) by upregulating fibroblast growth factor 23 (FGF23) mRNA expression. Therefore, the cells behave in a similar manner to osteocytes in vivo. These cells represent an important tool in enhancing current knowledge in human osteocyte biology.
Centre for Orthopaedic and Trauma Research, Discipline of Orthopaedics and Trauma, University of Adelaide, Adelaide, SA 5005, Australia. Electronic address: matt.prideaux@adelaide.edu.au.
Matthew
M
Prideaux
Centre for Orthopaedic and Trauma Research, Discipline of Orthopaedics and Trauma, University of Adelaide, Adelaide, SA 5005, Australia; Wakefield Orthopaedic Clinic, Adelaide, SA 5000, Australia.
Christine
C
Schutz
Centre for Orthopaedic and Trauma Research, Discipline of Orthopaedics and Trauma, University of Adelaide, Adelaide, SA 5005, Australia.
Asiri R
AR
Wijenayaka
Centre for Orthopaedic and Trauma Research, Discipline of Orthopaedics and Trauma, University of Adelaide, Adelaide, SA 5005, Australia.
David M
DM
Findlay
Wakefield Orthopaedic Clinic, Adelaide, SA 5000, Australia.
David G
DG
Campbell
Centre for Orthopaedic and Trauma Research, Discipline of Orthopaedics and Trauma, University of Adelaide, Adelaide, SA 5005, Australia.
Lucian B
LB
Solomon
Centre for Orthopaedic and Trauma Research, Discipline of Orthopaedics and Trauma, University of Adelaide, Adelaide, SA 5005, Australia. Electronic address: gerald.atkins@adelaide.edu.au.
Gerald J
GJ
Atkins
D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D016723:Bone Remodeling; D000071556:Cancellous Bone / Q000166:cytology*; D002469:Cell Separation / Q000379:methods*; D048430:Cell Shape; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism; D000089703:Fibroblast Growth Factor-23; D005786:Gene Expression Regulation; D006410:Hematopoiesis; D010011:Osteocytes / Q000166:cytology* / Q000378:metabolism; D014019:Tissue Donors
D015415:Biomarkers; C000717427:FGF23 protein, human; C000717428:Fgf23 protein, mouse; D000089703:Fibroblast Growth Factor-23
Cell morphology; Gene expression; Osteocyte; Trabecular bone