title: Isolation of myogenic stem cells from cultures of cryopreserved human skeletal muscle.
abstract: We demonstrate that subpopulations of adult human skeletal muscle-derived stem cells, myogenic endothelial cells (MECs), and perivascular stem cells (PSCs) can be simultaneously purified by fluorescence-activated cell sorting (FACS) from cryopreserved human primary skeletal muscle cell  cultures (cryo-hPSMCs). For FACS isolation, we utilized a combination of cell lineage markers: the myogenic cell marker CD56, the endothelial cell marker UEA-1 receptor (UEA-1R), and the perivascular cell marker CD146. MECs expressing all three cell lineage markers (CD56(+)UEA-1R(+)CD146(+)/CD45(-))  and PSCs expressing only CD146 (CD146(+)/CD45(-)CD56(-)UEA-1R(-)) were isolated by FACS. To evaluate their myogenic capacities, the sorted cells, with and without expansion in culture, were transplanted into the cardiotoxin-injured skeletal  muscles of immunodeficient mice. The purified MECs exhibited the highest regenerative capacity in the injured mouse muscles among all cell fractions tested, while PSCs remained superior to myoblasts and the unpurified primary skeletal muscle cells. Our findings show that both MECs and PSCs  retain their high myogenic potentials after in vitro expansion, cryopreservation, and FACS sorting. The current study demonstrates that myogenic stem cells are prospectively isolatable from long-term cryopreserved primary skeletal muscle cell cultures. We emphasize the potential application  of this new approach to extract therapeutic stem cells from human muscle cells cryogenically banked for clinical purposes.
authors.affiliation: Stem Cell Research Center, Department of Orthopaedic Surgery, University of Pittsburgh School of Medicine and Children's Hospital of Pittsburgh of UPMC, Pittsburgh, PA, USA.
authors.forename: Bo
authors.initials: B
authors.lastname: Zheng
authors.forename: Chien-Wen
authors.initials: CW
authors.lastname: Chen
authors.forename: Guangheng
authors.initials: G
authors.lastname: Li
authors.forename: Seth D
authors.initials: SD
authors.lastname: Thompson
authors.forename: Minakshi
authors.initials: M
authors.lastname: Poddar
authors.forename: Bruno
authors.initials: B
authors.lastname: Péault
authors.forename: Johnny
authors.initials: J
authors.lastname: Huard
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000818:Animals; D051929:CD146 Antigen / Q000378:metabolism; D019002:CD56 Antigen / Q000378:metabolism; D002454:Cell Differentiation; D019070:Cell Lineage; D002478:Cells, Cultured; D002648:Child; D002675:Child, Preschool; D015925:Cryopreservation; D042783:Endothelial Cells / Q000166:cytology; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D008297:Male; D051379:Mice; D016513:Mice, SCID; D008875:Middle Aged; D018482:Muscle, Skeletal / Q000166:cytology*; D012038:Regeneration / Q000502:physiology; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism; D055815:Young Adult
chemical_list: D051929:CD146 Antigen; D019002:CD56 Antigen; D014157:Transcription Factors; C471624:UBE2V1 protein, human; D044763:Ubiquitin-Conjugating Enzymes