title: A xenogeneic-free bioreactor system for the clinical-scale expansion of human mesenchymal stem/stromal cells.
abstract: The large cell doses (>1 × 10(6)  cells/kg) used in clinical trials with mesenchymal stem/stromal cells (MSC) will require an efficient production process. Moreover, monitoring and control of MSC ex-vivo expansion is critical to provide a safe and reliable cell product. Bioprocess engineering approaches, such as bioreactor technology, offer the adequate tools to develop and optimize a cost-effective culture system for the rapid expansion of human MSC for cellular therapy. Herein, a xenogeneic (xeno)-free microcarrier-based culture system was successfully established for bone marrow (BM) MSC and adipose tissue-derived stem/stromal cell (ASC) cultivation using a 1L-scale controlled stirred-tank bioreactor, allowing the production of (1.1 ± 0.1) × 10(8) and (4.5 ± 0.2) × 10(7) cells for BM MSC and ASC, respectively, after 7 days. Additionally, the effect of different percent air saturation values (%Airsat ) and feeding regime on the proliferation and metabolism of BM MSC was evaluated. No significant differences in cell growth and metabolic patterns were observed under 20% and 9%Airsat . Also, the three different feeding regimes studied-(i) 25% daily medium renewal, (ii) 25% medium renewal every 2 days, and (iii) fed-batch addition of concentrated nutrients and growth factors every 2 days-yielded similar cell numbers, and only slight metabolic differences were observed. Moreover, the immunophenotype (positive for CD73, CD90 and CD105 and negative for CD31, CD80 and HLA-DR) and multilineage differentiative potential of expanded cells were not affected upon bioreactor culture. These results demonstrated the feasibility of expanding human MSC from different sources in a clinically relevant expansion configuration in a controlled microcarrier-based stirred culture system under xeno-free conditions. The further optimization of this bioreactor culture system will represent a crucial step towards an efficient GMP-compliant clinical-scale MSC production system.
authors.affiliation: Department of Bioengineering and IBB-Institute for Biotechnology and Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Lisboa, Portugal.
authors.forename: Francisco
authors.initials: F
authors.lastname: Dos Santos
authors.forename: Andrew
authors.initials: A
authors.lastname: Campbell
authors.forename: Ana
authors.initials: A
authors.lastname: Fernandes-Platzgummer
authors.forename: Pedro Z
authors.initials: PZ
authors.lastname: Andrade
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: Gimble
authors.forename: Yuan
authors.initials: Y
authors.lastname: Wen
authors.forename: Shayne
authors.initials: S
authors.lastname: Boucher
authors.forename: Mohan C
authors.initials: MC
authors.lastname: Vemuri
authors.forename: Cláudia L
authors.initials: CL
authors.lastname: da Silva
authors.forename: Joaquim M S
authors.initials: JM
authors.lastname: Cabral
mesh_terms: D019149:Bioreactors; D018929:Cell Culture Techniques / Q000379:methods; D049109:Cell Proliferation; D006801:Humans; D016130:Immunophenotyping; D059630:Mesenchymal Stem Cells / Q000502:physiology*; D017154:Stromal Cells / Q000502:physiology*
keywords: bioreactor; expansion; mesenchymal stem/stromal cells; microcarriers; scale-up; xenogeneic-free