Autologous serum for isolation and expansion of human mesenchymal stem cells for clinical use
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Autologous serum for isolation and expansion of human mesenchymal stem cells for clinical use. / Stute, Norbert; Holtz, Katja; Bubenheim, Michael; Lange, Claudia; Blake, Felix; Zander, Axel R.
in: Experimental hematology, Jahrgang 32, Nr. 12, 01.12.2004, S. 1212-25.Publikationen: SCORING: Beitrag in Fachzeitschrift/Zeitung › SCORING: Zeitschriftenaufsatz › Forschung › Begutachtung
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TY - JOUR
T1 - Autologous serum for isolation and expansion of human mesenchymal stem cells for clinical use
AU - Stute, Norbert
AU - Holtz, Katja
AU - Bubenheim, Michael
AU - Lange, Claudia
AU - Blake, Felix
AU - Zander, Axel R
PY - 2004/12/1
Y1 - 2004/12/1
N2 - OBJECTIVE: Mesenchymal stem cells (MSC) are promising candidates for cell-based therapies. One major obstacle for their clinical use is the biosafety of fetal calf serum (FCS), which is a crucial part of all media currently used for the culture of MSC.METHODS: Nine donors each contributed 5 mL of bone marrow aspirate. Isolation of MSC was conducted according to Caplan et al., although for expansion we used low-density seeding with 20 MSC/cm2. Four different media A, B, C, and D were tested, containing 1%, 3%, or 10% autologous serum (AS), or 10% selected FCS, respectively. MSC were cultured on 24-well plates until passage 2 and counted under the microscope at regular intervals. Osteogenic and adipogenic differentiation were induced in vitro by using a modified standard cocktail and were evaluated semi-quantitatively through a microscope.RESULTS: Isolation of MSC after 3 days appeared best in media C with almost always C>D congruent with B>A. Proliferation was exponential with generally C>D>B>A. Morphologically, MSC isolated and expanded in medium C were indistinguishable from those in medium D. Phenotypic markers of MSC grown in medium C were: CD34-, CD45-, CD90+, CD105+, MHC class I+, MHC class II-, similar to MSC isolated and grown in medium D. Moreover, MSC grown in medium C showed more osteogenic potential than those from medium D in all cases: C+++, D++, B+, A 0. Cells retained their immaturity as shown by adipogenic differentiation and it always was: D+++, C++, B+, A 0.CONCLUSIONS: Growth of MSC in a FCS-free medium is feasible without addition of growth factors. Ten percent AS appears at least as good as 10% FCS with regard to both isolation and expansion of human MSC, while 1% and 3% AS appear inferior. With respect to osteogenic differentiation, 10% AS proved superior to the other serum conditions.
AB - OBJECTIVE: Mesenchymal stem cells (MSC) are promising candidates for cell-based therapies. One major obstacle for their clinical use is the biosafety of fetal calf serum (FCS), which is a crucial part of all media currently used for the culture of MSC.METHODS: Nine donors each contributed 5 mL of bone marrow aspirate. Isolation of MSC was conducted according to Caplan et al., although for expansion we used low-density seeding with 20 MSC/cm2. Four different media A, B, C, and D were tested, containing 1%, 3%, or 10% autologous serum (AS), or 10% selected FCS, respectively. MSC were cultured on 24-well plates until passage 2 and counted under the microscope at regular intervals. Osteogenic and adipogenic differentiation were induced in vitro by using a modified standard cocktail and were evaluated semi-quantitatively through a microscope.RESULTS: Isolation of MSC after 3 days appeared best in media C with almost always C>D congruent with B>A. Proliferation was exponential with generally C>D>B>A. Morphologically, MSC isolated and expanded in medium C were indistinguishable from those in medium D. Phenotypic markers of MSC grown in medium C were: CD34-, CD45-, CD90+, CD105+, MHC class I+, MHC class II-, similar to MSC isolated and grown in medium D. Moreover, MSC grown in medium C showed more osteogenic potential than those from medium D in all cases: C+++, D++, B+, A 0. Cells retained their immaturity as shown by adipogenic differentiation and it always was: D+++, C++, B+, A 0.CONCLUSIONS: Growth of MSC in a FCS-free medium is feasible without addition of growth factors. Ten percent AS appears at least as good as 10% FCS with regard to both isolation and expansion of human MSC, while 1% and 3% AS appear inferior. With respect to osteogenic differentiation, 10% AS proved superior to the other serum conditions.
KW - Adipocytes
KW - Animals
KW - Antigens, CD
KW - Cattle
KW - Cell Culture Techniques
KW - Cell Differentiation
KW - Cell Proliferation
KW - Cell- and Tissue-Based Therapy
KW - Cells, Cultured
KW - Colony-Forming Units Assay
KW - Culture Media
KW - Humans
KW - Mesenchymal Stromal Cells
KW - Osteoblasts
KW - Safety
KW - Serum
U2 - 10.1016/j.exphem.2004.09.003
DO - 10.1016/j.exphem.2004.09.003
M3 - SCORING: Journal article
C2 - 15588946
VL - 32
SP - 1212
EP - 1225
JO - EXP HEMATOL
JF - EXP HEMATOL
SN - 0301-472X
IS - 12
ER -