Korean J Reprod Med.  2009 Mar;36(1):23-34.

Characterization of Umbilical Cord-derived Stem Cells during Expansion in Vitro

• Park S
• Kang HM
• Heo J
• Yoon J
• Kim H

Abstract

OBJECTIVES
Mesenchymal stem cells (MSC) comprise a promising tool for cellular therapy. It is known that long-term in vitro culture of human bone marrow and adipose tissue derived-MSCs lead to a reduction of life span and a change of stem-like characters. The aim of our study was to examine whether stem cell properties of human umbilical cord-derived stem cells (HUC) could be affected by in vitro expansion.
METHODS
HUC were isolated from human umbilical cord and cultured for 10 passages in vitro. Morphology and population doubling time (PDT) were investigated, and changes of stem cell properties were examined using RT-PCR and immunocytochemistry during serial subcultures.
RESULTS
Morphology and PDT of HUC began to change slightly from the 7th passage (p7). Expression level of nestin and vimentin mRNAs increased along with the culture period from p4 until p10. In contrast, expression level of SCF mRNA decreased during the same culture period. Expression level of Oct-4 and HNF-4alpha mRNAs was not significantly changed throughout the culture period until p10. Expression level of BMP-4, FGF-5, NCAM and HLA-ABC mRNAs appeared to increase as the culture continued, however, the difference was not significant. Immunocytochemical studies showed that HUC at p3, p6 and p9 positively were stained with antibodies against SSEA-3 and SSEA-4 proteins. Interestingly, staining intensity of HUC for ICAM-1 and HLA-ABC gradually increased throughout the culture period. Intensity against thy-1 and fibronectin antibodies increased at p9 while that against TRA-1-60 and VCAM-1 antibodies began to decrease at p6 until p9.
CONCLUSIONS
These results suggest that HUC change some of their stem cell characteristics during in vitro culture. Development of culture system might be needed for the maintenance of characteristics.

Keyword

Umbilical cord-derived stem cells; Stem cell characteristics; RT-PCR; Immunocytochemistry

MeSH Terms

Adipose Tissue
Antibodies
Antigens, Tumor-Associated, Carbohydrate
Bone Marrow
Fibronectins
Humans
Immunohistochemistry
Intercellular Adhesion Molecule-1
Intermediate Filament Proteins
Mesenchymal Stromal Cells
Nerve Tissue Proteins
Neural Cell Adhesion Molecules
Proteins
RNA, Messenger
Stage-Specific Embryonic Antigens
Stem Cells
Triazenes
Umbilical Cord
Vascular Cell Adhesion Molecule-1
Vimentin
Antibodies
Antigens, Tumor-Associated, Carbohydrate
Fibronectins
Intercellular Adhesion Molecule-1
Intermediate Filament Proteins
Nerve Tissue Proteins
Neural Cell Adhesion Molecules
Proteins
RNA, Messenger
Stage-Specific Embryonic Antigens
Triazenes
Vascular Cell Adhesion Molecule-1
Vimentin