Rapid clinical-scale propagation of mesenchymal stem cells using cultures initiated with immunoselected bone marrow CD105+ cells.
Current clinical protocols used for isolation and purification of mesenchymal stem cells (MSC) are based on long-term cultures starting with bone marrow (BM) mononuclear cells. Using a commercially available immunoselection kit for enrichment of MSC, we investigated whether culture of enriched BM-CD105(+) cells could provide an adequate number of pure MSC in a short time for clinical use in the context of graft versus host disease and graft failure/rejection. We isolated a mean of 5.4 × 10(5) ± 0.9 × 10(5) CD105(+) cells from 10 small volume (10-25 ml) BM samples achieving an enrichment >100-fold in MSC. Seeding 2 × 10(3) immunoselected cells/cm(2) we were able to produce 2.5 × 10(8) ± 0.7 × 10(8) MSC from cultures with autologous serum enriched medium within 3 weeks. Neither haematopoietic nor endothelial cells were detectable even in the primary culture cell product. Expanded cells fulfilled both phenotypic and functional current criteria for MSC; they were CD29(+), CD90(+), CD73(+), CD105(+), CD45(-); they suppressed allogeneic T-cell reaction in mixed lymphocyte cultures and retained in vitro differentiation potential. Moreover, comparative genomic hybridization analysis revealed chromosomal stability of the cultured MSC. Our data indicate that adequate numbers of pure MSC suitable for clinical applications can be generated within a short time using enriched BM-CD105(+) cells.
Spiropoulos
Antonia
A
Stem Cell Transplant Unit, 'Aghia Sophia' Children's Hospital, Athens, Greece.
Theodosaki
Maria
M
Stefanaki
Kalliopi
K
Paterakis
George
G
Tzetis
Maria
M
Giannikou
Krinio
K
Petrakou
Eftichia
E
Dimopoulou
Maria N
MN
Papassotiriou
Ioannis
I
Roma
Eleptheria S
ES
Kanavakis
Emmanuel
E
Graphakos
Stelios
S
Goussetis
Evgenios
E
D000328:Adult; D015703:Antigens, CD / Q000378:metabolism*; D001854:Bone Marrow Cells / Q000166:cytology* / Q000378:metabolism*; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D049109:Cell Proliferation; D002469:Cell Separation; D002478:Cells, Cultured; D002648:Child; D055028:Comparative Genomic Hybridization; D000071063:Endoglin; D006801:Humans; D016130:Immunophenotyping / Q000379:methods*; D007959:Lymphocyte Culture Test, Mixed; D008214:Lymphocytes / Q000166:cytology; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D055815:Young Adult
D015703:Antigens, CD; C064206:ENG protein, human; D000071063:Endoglin; D011956:Receptors, Cell Surface