TY - JOUR
T1 - Development of large-scale manufacturing of adipose-derived stromal cells for clinical applications using bioreactors and human platelet lysate
AU - Haack-Sørensen, Mandana
AU - Juhl, Morten
AU - Follin, Bjarke
AU - Harary Søndergaard, Rebekka
AU - Kirchhoff, Maria
AU - Kastrup, Jens
AU - Ekblond, Annette
PY - 2018/7
Y1 - 2018/7
N2 - In vitro expanded adipose-derived stromal cells (ASCs) are a useful resource for tissue regeneration. Translation of small-scale autologous cell production into a large-scale, allogeneic production process for clinical applications necessitates well-chosen raw materials and cell culture platform. We compare the use of clinical-grade human platelet lysate (hPL) and fetal bovine serum (FBS) as growth supplements for ASC expansion in the automated, closed hollow fibre quantum cell expansion system (bioreactor). Stromal vascular fractions were isolated from human subcutaneous abdominal fat. In average, 95 × 106 cells were suspended in 10% FBS or 5% hPL medium, and loaded into a bioreactor coated with cryoprecipitate. ASCs (P0) were harvested, and 30 × 106 ASCs were reloaded for continued expansion (P1). Feeding rate and time of harvest was guided by metabolic monitoring. Viability, sterility, purity, differentiation capacity, and genomic stability of ASCs P1 were determined. Cultivation of SVF in hPL medium for in average nine days, yielded 546 × 106 ASCs compared to 111 × 106 ASCs, after 17 days in FBS medium. ASCs P1 yields were in average 605 × 106 ASCs (PD [population doublings]: 4.65) after six days in hPL medium, compared to 119 × 106 ASCs (PD: 2.45) in FBS medium, after 21 days. ASCs fulfilled ISCT criteria and demonstrated genomic stability and sterility. The use of hPL as a growth supplement for ASCs expansion in the quantum cell expansion system provides an efficient expansion process compared to the use of FBS, while maintaining cell quality appropriate for clinical use. The described process is an obvious choice for manufacturing of large-scale allogeneic ASC products.
AB - In vitro expanded adipose-derived stromal cells (ASCs) are a useful resource for tissue regeneration. Translation of small-scale autologous cell production into a large-scale, allogeneic production process for clinical applications necessitates well-chosen raw materials and cell culture platform. We compare the use of clinical-grade human platelet lysate (hPL) and fetal bovine serum (FBS) as growth supplements for ASC expansion in the automated, closed hollow fibre quantum cell expansion system (bioreactor). Stromal vascular fractions were isolated from human subcutaneous abdominal fat. In average, 95 × 106 cells were suspended in 10% FBS or 5% hPL medium, and loaded into a bioreactor coated with cryoprecipitate. ASCs (P0) were harvested, and 30 × 106 ASCs were reloaded for continued expansion (P1). Feeding rate and time of harvest was guided by metabolic monitoring. Viability, sterility, purity, differentiation capacity, and genomic stability of ASCs P1 were determined. Cultivation of SVF in hPL medium for in average nine days, yielded 546 × 106 ASCs compared to 111 × 106 ASCs, after 17 days in FBS medium. ASCs P1 yields were in average 605 × 106 ASCs (PD [population doublings]: 4.65) after six days in hPL medium, compared to 119 × 106 ASCs (PD: 2.45) in FBS medium, after 21 days. ASCs fulfilled ISCT criteria and demonstrated genomic stability and sterility. The use of hPL as a growth supplement for ASCs expansion in the quantum cell expansion system provides an efficient expansion process compared to the use of FBS, while maintaining cell quality appropriate for clinical use. The described process is an obvious choice for manufacturing of large-scale allogeneic ASC products.
KW - Adipose Tissue/cytology
KW - Adult
KW - Bioreactors
KW - Blood Platelets/metabolism
KW - Cell Culture Techniques/methods
KW - Cell Differentiation
KW - Cell Proliferation
KW - Female
KW - Genomic Instability
KW - Humans
KW - Lactates/metabolism
KW - Male
KW - Mesenchymal Stem Cells/cytology
KW - Middle Aged
KW - Phenotype
KW - Time Factors
U2 - 10.1080/00365513.2018.1462082
DO - 10.1080/00365513.2018.1462082
M3 - Journal article
C2 - 29661028
VL - 78
SP - 293
EP - 300
JO - Scandinavian Journal of Clinical and Laboratory Investigation
JF - Scandinavian Journal of Clinical and Laboratory Investigation
SN - 0036-5513
IS - 4
ER -