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Accelerated and Safe Proliferation of Human Adipose-derived Stem Cells in Medium Supplemented with Human Serum

Fonny Josh¹, Kyoko Kobe², Morikuni Tobita¹, Rica Tanaka¹, Koji Suzuki³, Kasumi Ono³, Hiko Hyakusoku² and Hiroshi Mizuno^1,2

¹Department of Plastic and Reconstructive Surgery, Juntendo University School of Medicine
²Department of Plastic and Reconstructive Surgery, Nippon Medical School
³R&D Central Research Laboratory, JMS Co. Ltd., Hiroshima

Adipose-derived stem cells (ASCs) are a promising cell source and are being investigated for a variety of therapeutic applications. However, standard expansion protocols use fetal bovine serum (FBS) as a growth factor supplement, which is a potential source of undesirable xenogeneic pathogens. For clinical safety, autologous human serum (HS) would be more appropriate. This study compared FBS-supplemented and HS-supplemented media for their enhancement of the proliferation and differentiation potential of human ASCs (hASCs). HS was obtained from the blood of 8 healthy volunteers using collection devices specially designed to derive growth factors from platelets. Growth factors in HS or FBS were measured with enzyme-linked immunosorbent assays. The hASCs were isolated with an established protocol from discarded human fat tissues obtained during a medical procedure and cultured in a medium supplemented with either 10% HS or 10% FBS. The hASCs were collected at several time points for the proliferation assays. The capacity for differentiation into the osteogenic, chondrogenic and adipogenic lineages was assessed qualitatively with the histochemical stains von Kossa, Alcian blue, and Oil red O, respectively, and quantitatively with the qualitative reverse transcriptase polymerase chain reaction. Differences in cell surface marker expression between the HS-supplemented and FBS-supplemented cultures were examined with flow cytometric analysis. Proliferation assays showed that the growth of hASCs was more rapid in HS-supplemented medium than in FBS-supplemented medium. All cells grown in each medium expressed similar patterns of cell surface markers. The ASCs cultured in the HS-supplemented medium proliferated more rapidly than those cultured in the FBS-supplemented medium and retained their differentiation capacity and immunophenotype. These results support the establishment of a safe and rapid expansion protocol with autologous serum for cell-based therapies, such as tissue engineering and regenerative medicine, using hASCs.

J Nippon Med Sch 2012; 79: 444-452

Keywords
adipose-derived stem cells, human serum, mesenchymal stromal cells, fetal bovine serum, platelet-derived factors

Correspondence to
Hiroshi Mizuno, MD, PhD, Department of Plastic and Reconstructive Surgery, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421, Japan
hmizuno@juntendo.ac.jp

Received, September 11, 2012
Accepted, October 18, 2012