title: Flow cytometry measurement of bone marrow perfusion in the mouse and sorting of progenitors and stems cells according to position relative to blood flow in vivo.
abstract: Identification of the precise location, where hematopoietic stem cells (HSCs) reside in the bone marrow, has made a great leap forward with the advance of live time-lapse video 2-photon fluorescent microscopy. These studies have shown that HSCs preferentially resides in the endosteal region of the BM, at an average of two cell diameters from osteoblasts covering endosteal bone surfaces. However, this equipment is very sophisticated and only a very few laboratories can perform these studies. To investigate functional attributes of these niches, we have developed a flow cytometry technique in which mice are perfused with the cell-permeable fluorescent dye Hoechst33342 in vivo before bone marrow cells are collected and antibody stained. This method enables to position phenotypic HSC, multipotent and myeloid progenitors, as well as BM nonhematopoietic stromal cells relative to blood flow in vivo. This technique enables prospective isolation of HSCs based on the in vivo perfusion of the niches in which they reside.
authors.affiliation: Stem Cell and Cancer Group, Mater Medical Research Institute, South Brisbane, QLD, Australia.
authors.forename: Valérie
authors.initials: V
authors.lastname: Barbier
authors.forename: Ingrid G
authors.initials: IG
authors.lastname: Winkler
authors.forename: Robert
authors.initials: R
authors.lastname: Wadley
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Lévesque
mesh_terms: D000818:Animals; D001562:Benzimidazoles / Q000008:administration & dosage; D001853:Bone Marrow / Q000098:blood supply*; D001854:Bone Marrow Cells / Q000378:metabolism; D042783:Endothelial Cells / Q000378:metabolism; D005434:Flow Cytometry / Q000379:methods*; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D016130:Immunophenotyping; D007962:Leukocytes / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000378:metabolism; D051379:Mice; D057507:Mice, 129 Strain; D022423:Myeloid Cells / Q000378:metabolism; D010006:Osteoblasts / Q000378:metabolism; D012039:Regional Blood Flow; D013194:Staining and Labeling / Q000379:methods; D055153:Stem Cell Niche
chemical_list: D001562:Benzimidazoles; C017807:bisbenzimide ethoxide trihydrochloride