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cells.^10 Upon further examination of the KLF family, KLF-1, KLF-2,
and KLF-5 have all been found to share KLF-4’s ability to induce and
maintain pluripotency.^11
1.2. Serum Free Culture
Early culture conditions for human ESCs (hESCs) included the
culture medium supplemented with fetal bovine serum. However
the serum was a major source of undefined differentiation factors
and therefore led to the promotion hESC differentiation.
Subsequently, serum replacement was created by Price et al. in
1998, consisting of undisclosed albumins and albumin substitutes.^12
Two years later, Amit et al. reported a successful serum-free media
with the addition of basic fibroblast growth factor (bFGF), also
called FGF2. A nearly four-fold increase in cloning efficiency of
hESCs was observed.^13
FGF2’s exact role in promoting hESC pluripotency was further
investigated. A significant downregulation of Oct3/4 occurred upon
the removal of FGF2’s respective receptor from hESCs, suggesting a
role for FGF2 signaling in regulating Oct3/4 expression.^14 Ding et al.
showed that FGF2 starvation followed by subsequent reapplication
led to rapid activation of PI3-K.^14 In the same study, PI3-K inhibition
with LY294002, resulted in the downregulation of pluripotency
markers Oct3/4, PODXL, and Tra-1-60, emphasizing the impor-
tance of FGF2 and downstream PI3-K signaling in maintaining
pluripotency.^14
1.3. Feeder Free Culture
Along with the culture media, the surface on which hESCs are grown is
equally important for successful stem cell culture. Initial efforts to main-
tain hESCs in vitro required culture on a “feeder layer” made of mouse
embryonic fibroblasts (MEFs).^15 In murine ESC culture, the addition of
LIF to the media activated the LIF/STAT3 pathway, and, as discussed
previously, is sufficient to maintain murine ESCs eliminating the need
for the feeder layer.^16 However, for hESC, LIF/STAT3 is ineffective.
