DEPARTAMENTO DE CIÊNCIAS DA VIDA ... - Estudo Geral

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hydrogels of 1 and 7 kPa (for cells originally isolated on hydrogels and on plastic) and
on TCPs (for cells originally isolated on plastic), in order to check if the hard stiffness
from plastic was affecting the cells isolated even at low passages (P2) when compared
to cells that never had contact with hard stiff substrates. All cells were treated with
mitomycin C and let on the aforementioned substrates for 7 days with proliferation
medium (PM) after plating and then stained for BIII tubulin, Nestin, GFAP and O4. The
results (Figure 22 and 23) showed that for Nestin there is not much expression in any
of the conditions but there is a slight increase in the cells isolated and cultured on the
PA hydrogels only (7kPA hydrogels and 1kPa hydrogels) when compared to those that
had contact with the plastic (Plastic, 7kPa hydrogel/TCPs and 1kPA hydrogel/TCPs),
being the 7 kPa hydrogels the condition with highest staining. Expression of Beta-III
tubulin (BIII) seems higher in the cells cultured on 1 kPa hydrogels, both the ones
cultured exclusively on the hydrogels as the ones that were on TCPs (plastic) for 2
passages and then cultured on hydrogels. The expression of BIII tubulin seems to be
lower in cells plated exclusively on plastic, with an increase that can be observed when
the differentiation took place on hydrogels. In terms of O4, it seems that the cells
plated exclusively on the hydrogels (7kPa and 1kPa hydrogels) have more expression
when compared to the other cells that had contact with plastic (Plastic, 7kPa and 1kPa
Hydrogel/TCPs), being the cells plated exclusively on plastic the ones that show lower
O4 expression. Finally, GFAP expression seems to be very similar in all hydrogels being
probably the cells plated exclusively on plastic the ones with less expression of GFAP.
These results suggest that the expression of the oligodendroglial marker O4 is
increased when MSCs are isolated and differentiated on soft substrates (PA hydrogels).
This was also observed, to a lesser extent, regarding the expression of nestin. The
expression of Beta-III tubulin seems to be enhanced when differentiation takes place
on PA hydrogels (soft substrates), but the effect seems to be more independent of the
platform used to isolate the cells.