A novel method to derive amniotic fluid stem cells for therapeutic purposes.
BACKGROUND
Human amniotic fluid stem (hAFS) cells have become an attractive stem cell source for medical therapy due to both their ability to propagate as stem cells and the lack of ethical debate that comes with the use of embryonic stem cells. Although techniques to derive stem cells from amniotic fluid are available, the techniques have limitations for clinical uses, including a requirement of long periods of time for stem cell production, population heterogeneity and xeno-contamination from using animal antibody-coated magnetic beads. Herein we describe a novel isolation method that fits for hAFS derivation for cell-based therapy.


RESULTS
With our method, single hAFS cells generate colonies in a primary culture of amniotic fluid cells. Individual hAFS colonies are then expanded by subculturing in order to make a clonal hAFS cell line. This method allows derivation of a substantial amount of a pure stem cell population within a short period of time. Indeed, 108 cells from a clonal hAFS line can be derived in two weeks using our method, while previous techniques require two months. The resultant hAFS cells show a 2-5 times greater proliferative ability than with previous techniques and a population doubling time of 0.8 days. The hAFS cells exhibit typical hAFS cell characteristics including the ability to differentiate into adipogenic-, osteogenic- and neurogenic lineages, expression of specific stem cell markers including Oct4, SSEA4, CD29, CD44, CD73, CD90, CD105 and CD133, and maintenance of a normal karyotype over long culture periods.


CONCLUSIONS
We have created a novel hAFS cell derivation method that can produce a vast amount of high quality stem cells within a short period of time. Our technique makes possibility for providing autogenic fetal stem cells and allogeneic cells for future cell-based therapy.
Phermthai
Tatsanee
T
Stem Cell Research and Development Unit, Department of Obstetrics & Gynecology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10700, Thailand. sitsu@mahidol.ac.th
Odglun
Yuparat
Y
Julavijitphong
Suphakde
S
Titapant
Vitaya
V
Chuenwattana
Prakong
P
Vantanasiri
Chanchai
C
Pattanapanyasat
Kovit
K
D000653:Amniotic Fluid / Q000166:cytology*; D002469:Cell Separation / Q000379:methods*; D043171:Chromosomal Instability; D005260:Female; D005865:Gestational Age; D006801:Humans; D007381:Intermediate Filament Proteins / Q000235:genetics / Q000378:metabolism; D007621:Karyotyping; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D064231:Nestin; D011247:Pregnancy; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D014404:Tubulin / Q000235:genetics / Q000378:metabolism
D007381:Intermediate Filament Proteins; C577484:NES protein, human; D009419:Nerve Tissue Proteins; D064231:Nestin; D014404:Tubulin