Isolation of Blood-vessel-derived Multipotent Precursors from Human Skeletal Muscle

Abstract

Since the discovery of mesenchymal stem/stromal cells (MSCs), the native identity and localization of MSCs have been obscured by their retrospective isolation in culture. Recently, using fluorescence-activated cell sorting (FACS), we and other researchers prospectively identified and purified three subpopulations of multipotent precursor cells associated with the vasculature of human skeletal muscle. These three cell populations: myogenic endothelial cells (MECs), pericytes (PCs), and adventitial cells (ACs), are localized respectively to the three structural layers of blood vessels: intima, media, and adventitia. All of these human blood-vessel-derived stem cell (hBVSC) populations not only express classic MSC markers but also possess mesodermal developmental potentials similar to typical MSCs. Previously, MECs, PCs, and ACs have been isolated through distinct protocols and subsequently characterized in separate studies. The current isolation protocol, through modifications to the isolation process and adjustments in the selective cell surface markers, allows us to simultaneously purify all three hBVSC subpopulations by FACS from a single human muscle biopsy. This new method will not only streamline the isolation of multiple BVSC subpopulations but also facilitate future clinical applications of hBVSCs for distinct therapeutic purposes.

Cultured Human Adipose Tissue Pericytes and Mesenchymal Stromal Cells Display a Very Similar Gene Expression Profile

“…This view could explain the establishment of MSC cultures by AT3G5Cs as shown here and by CD146 + CD56 -perivascular cells in various organs [8], including skeletal muscle [44].…”

Development of mRuby2-Transfected C3H10T1/2 Fibroblasts for Musculoskeletal Tissue Engineering

“…As previously mentioned, MSCs are self-renewing multipotent stem cells that can differentiate into cells of the musculoskeletal system and secrete paracrine factors to promote tissue regeneration [ 3 – 5 , 8 – 11 , 13 , 14 , 16 – 20 , 22 , 24 , 26 – 28 , 30 , 37 – 39 ]. While their origin is still subject to ongoing investigations, some studies have indicated that MSCs are derived from pericyte-like cells [ 3 , 40 ] and are associated with blood vessels throughout the body in tissues such as muscle [ 5 ].…”

“…As previously mentioned, MSCs are self-renewing multipotent stem cells that can differentiate into cells of the musculoskeletal system and secrete paracrine factors to promote tissue regeneration [ 3 – 5 , 8 – 11 , 13 , 14 , 16 – 20 , 22 , 24 , 26 – 28 , 30 , 37 – 39 ]. While their origin is still subject to ongoing investigations, some studies have indicated that MSCs are derived from pericyte-like cells [ 3 , 40 ] and are associated with blood vessels throughout the body in tissues such as muscle [ 5 ]. As a result, these cells can be isolated from multiple tissues including adipose[ 27 ], amniotic fluid [ 13 ], bone marrow [ 10 , 20 ], cord blood [ 9 ], teeth [ 18 ], heart [ 4 ], neuroglia [ 16 ], placenta [ 14 ], synovium [ 8 ], skeletal muscle [ 5 , 26 ] as well as skin [ 26 ].…”

“…While their origin is still subject to ongoing investigations, some studies have indicated that MSCs are derived from pericyte-like cells [ 3 , 40 ] and are associated with blood vessels throughout the body in tissues such as muscle [ 5 ]. As a result, these cells can be isolated from multiple tissues including adipose[ 27 ], amniotic fluid [ 13 ], bone marrow [ 10 , 20 ], cord blood [ 9 ], teeth [ 18 ], heart [ 4 ], neuroglia [ 16 ], placenta [ 14 ], synovium [ 8 ], skeletal muscle [ 5 , 26 ] as well as skin [ 26 ]. In addition to their broad availability, MSCs can differentiate into cells of adipose [ 3 , 20 ], bone [ 3 , 20 ], cartilage [ 3 , 20 ], heart [ 17 ], liver [ 19 ], neural [ 22 ], skeletal muscle [ 24 ] and tendon [ 11 ] lineages.…”

Regenerative Translation of Human Blood-Vessel-Derived MSC Precursors

“…Based on the cell surface marker expression identified by immunohistochemistry, we discovered a unique combination of surface antigens for each subset of hBVSCs that allows one to purify these cells to homogeneity through FACS: MECs (CD34+/56+/144+/45−), PCs (CD146+/34−/45−/56−), and ACs (CD34+/31−/45−/56−/146−) [ 16 , 18 , 28 ]. The isolation and purification of hBVSC subpopulations have been well established [ 46 ]. The workflow of hBVSC purification from fresh human skeletal muscle biopsy is illustrated in Figure 1 .…”