Research and Clinical Center for Infertility, Shahid Sadoughi University of Medical Sciences of Yazd
Abstract
Background: Since embryonic stem (ES) cells have the dual ability to
proliferate indefinitely and differentiate into multiple tissue types,
ES cells could potentially provide an unlimited cell supply for human
transplantation. Objective: In order to study the differentiation of
mouse embryonic stem (mES) cells, they were cultured in suspension by
using ES media without Leukemia Inhibitory Factor (LIF) to induce
spontaneous differentiation. Cellular morphology of differentiated
derivatives was then evaluated. Materials and Methods: Undifferentiated
mES from our laboratory were cultured in three different settings by
using ES media containing 0.1% / 1mM trypsin/EDTA and removing LIF; in
the absence of murine embryonic fibroblast (MEF) feeder cells (group
1), in the presence of MEF feeder cells with a density of 0.5×105
cells/ml (group 2), and 0.5×106 cells/ml (group 3). Five days
after the initiation of cell culture, and inducing mES cells to form
embryoid bodies (EBs), they were removed from dish by centrifugation,
and then they were cultured on collagen coated dishes for 20 days. The
dishes were fixed and stained by Wright-Gimsa method at the end of the
study period. Results: In group 1, mES cells showed spontaneous
differentiation to all derivatives of three germ cells, including:
epithelia like, fibroblast like and neron-like cells. In group 2,
almost all ES cells were found to be differentiated into granular
progenitor cells including hematopoietic cell lineages. In group 3,
various morphologies including nerve cell lineages and fibroblast-like
cells were detected. Conclusion: Differentiation of mES cells can be a
dose response process, depending on the factors that may be released
from MEF feeder layer to ES media in a coculture system. Our results
indicated that in the presence of low numbers of MEF cells, mES cells
can spontaneously differentiate into hematopoeitic cell lineages