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F Wolf et al. Cartilage engineering with aggregated chondrocytes
Eurpean Cells and Materials Vol. 16 2008 (pages 92-99)
ISSN 1473-2262
Abstract
In this study, we first aimed at determining whether human
articular chondrocytes (HAC) proliferate in aggregates in
the presence of strong chondrocyte mitogens. We then
investigated if the aggregated cells have an enhanced
chondrogenic capacity as compared to cells cultured in
monolayer. HAC from four donors were cultured in tissue
culture dishes either untreated or coated with 1% agarose
in the presence of TGFβ-1, FGF-2 and PDGF-BB.
Proliferation and stage of differentiation were assessed by
measuring respectively DNA contents and type II collagen
mRNA. Expanded cells were induced to differentiate in
pellets or in Hyaff®-11 meshes and the formed tissues were
analysed biochemically for glycosaminoglycans (GAG) and
DNA, and histologically by Safranin O staining. The
amount of DNA in aggregate cultures increased
significantly from day 2 to day 6 (by 3.2-fold), but did not
further increase with additional culture time. Expression
of type II collagen mRNA was about two orders of
magnitude higher in aggregated HAC as compared to
monolayer expanded cells. Pellets generated by aggregated
HAC were generally more intensely stained for GAG than
those generated by monolayer-expanded cells. Scaffolds
seeded with aggregates accumulated more GAG (1.3-fold)
than scaffolds seeded with monolayer expanded HAC. In
conclusion, this study showed that HAC culture in
aggregates does not support a relevant degree of expansion.
However, aggregation of expanded HAC prior to loading
into a porous scaffold enhances the quality of the resulting
tissues and could thus be introduced as an intermediate
culture phase in the manufacture of engineered cartilage
grafts.
Keywords: articular chondrocytes, chondrogenesis, cell
differentiation, cartilag