Two steps to functional mesenchymal stromal cells for clinical application
2007; Wiley; Volume: 47; Issue: 8 Linguagem: Inglês
10.1111/j.1537-2995.2007.01219.x
ISSN1537-2995
AutoresChristina Bartmann, Eva Rohde, Katharina Schallmoser, P. Pürstner, Gerhard Lanzer, Werner Linkesch, Dirk Strunk,
Tópico(s)Neonatal Respiratory Health Research
ResumoBACKGROUND: Ex vivo expansion of multipotent mesenchymal stromal cells (MSCs) is a prerequisite for evaluating their therapeutic potential in ongoing clinical trials. Even large volumes of starting material and extended culture periods, however, do not necessarily produce 2 × 10 6 MSCs per kg per adult patient. A new two‐step procedure has been devised to propagate more than 1 × 10 8 MSCs from small marrow volumes within fewer than 4 weeks. STUDY DESIGN AND METHODS: The influence of log fold decreased MSC seeding (2500, 250, 25, 2.5/cm 2 ) on clinical‐scale expansion, MSC phenotype, and immunomodulatory function combined with multiplex cytokine display was analyzed. Maintenance of MSC characteristics was tested in fibroblast colony‐forming unit and differentiation assays. RESULTS: Reduced seeding density boosted MSC propagation. Low‐density expanded MSCs were CD29+, CD73+, CD90+, CD105+, CD14–, CD34–, CD45–, HLA‐DR–; retained their differentiation potential; and inhibited lymphocyte proliferation. This was accompanied by deregulated cytokine production. Seeding 0.7 × 10 6 to 1 × 10 6 MSCs derived from a 10‐ to 13‐day primary culture at a low density of 28 to 40 per cm 2 permitted propagation of 1.5 × 10 8 to 3.7 × 10 8 functional MSCs within a 13‐ to 15‐day secondary expansion step. CONCLUSION: Primary seeding of only 10‐mL marrow aspirates on approximately 0.2‐m 2 culture area (Step 1) followed by expansion on 2.5 m 2 (Step 2) is sufficient to consistently generate at least 1.5 × 10 8 MSCs in fetal bovine serum–supplemented medium within less than 4 weeks. The efficiency of this two‐step procedure for clinical‐scale MSC propagation may facilitate rational clinical testing of MSC‐based therapies.
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