A novel method to derive amniotic fluid stem cells for therapeutic purposes

Abstract

BackgroundHuman 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.Methods and ResultsWith 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.ConclusionsWe 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.

Exploring the Human Term Placenta as a Novel Source for Stem Cells and their Application in the Clinic

“…Moreover, it has also been reported these cells have a high proliferative potential with over 250 population doublings without doubling time changes and can be differentiated into endodermal and ectodermal lineages in vitro. Although these properties seem theoretically more advantageous, it is debatable whether amniotic fluid is a practical and reliable source for generating MSC on a regular basis for clinical trials (Tsai et al, 2004, De Coppi et al, 2007, Antonucci et al, 2009, Phermthai et al, 2010, Witkowska-Zimny and Wrobel, 2011.…”

“…It has been estimated that approximately 1% of cells in culture obtained from human amniocentesis are MSC. Only a few studies have successfully isolated single cell-derived MSC clones from amniotic fluid (Tsai et al, 2004, De Coppi et al, 2007, Antonucci et al, 2009, Phermthai et al, 2010, Witkowska-Zimny and Wrobel, 2011.…”

Potential antitumor therapeutic strategies of human amniotic membrane and amniotic fluid-derived stem cells

“…34 Human amniotic fluid-derived stem cells (hAFCs) As human amniotic fluid contains the developing embryo and directly contacts with the amniotic membrane during the gestation period, it has diverse cell types derived from an embryo and the amniotic membrane. 17,35 The presence of hAFCs population in amniotic fluid was firstly identified in 2002 by Prusa et al 36 and since then, their stem cell characteristics have been extensively studied. hAFCs was shown to express OCT4 mRNA 37 and was scored positive for mesenchymal markers such as CD90, CD105 (SH2), CD73 (SH3/4) and CD166, but negative for hematopoietic markers.…”

Human amniotic fluid-derived c-kit+ and c-kit− stem cells: growth characteristics and some differentiation potential capacities comparison

“…Through c-kit, the surface marker. A high purity AF cell population can be sorted using c-kit as a target, and it is considered that a purer cell population is safer for clinical use (Phermthai et al 2010). However, c-kit-based selection procedure may induce a different mesenchymal differentiation capacity from the unselected cell populations (Arnhold et al 2011).…”