1477.3.2.3 Chondrogenesis
Cartilage is a specialized connective tissue which consists of chondrocytes
surrounded by an ECM. Chondrocytes produce and maintain an ECM which is
rich in glycosaminoglycans and proteoglycans and interact with collagen and
elastic fibres. The cartilage matrix is composed mainly of type II collagen, pro-
viding tensile strength, and aggrecan, which provides compressive strength.
Cartilage is classified according to its composition. Hyaline cartilage consists
mainly of type II collagen fibres, elastic cartilage of type II collagen and elastic
fibres and fibrocartilage of type I collagen fibres (Naumann et al. 2002 ).
Chondrogenesis involves the recruitment and condensation of MSCs followed
by differentiation into chondroblasts, chondrocyte maturation, and ECM
production.
In vitro chondrogenesis is induced by artificially condensing the cells by cen-
trifugation followed by addition of TGF-β3, dexamethasone and BMP-2 to the
medium. The composition of the chondrogenic induction medium is provided in the
supplementary material. Johnstone and colleagues ( 1998 ) developed a culture sys-
tem to facilitate chondrogenic differentiation for BM-MSCs (Johnstone et al. 1998 ).
This pellet culture system allows for cell-cell interaction similar to those that occur
during embryonic development (Fell 1925 ). It has been shown that a defined
medium, to which dexamethasone and TGF-β1 have been added, is required
(Johnstone et al. 1998 ). Dexamethasone induces a metachromatic staining pattern in
aggregated cells. This indicates the synthesis of a cartilaginous matrix which was
confirmed by collagen type II immunohistochemistry. The use of TGF-β1 either
alone or in combination with dexamethasone induces chondrogenesis in aggregated
cells. It was later found that the addition of BMP-6 further increased chondrogene-
sis (Sekiya et al. 2001 ). Critical roles are played by BMPs in compaction of MSCs
and for the shaping of the condensations (Zuscik et al. 2008 ). Comparisons between
Fig. 7.5 Light microscopy analysis of osteogenesis. (a) Alizarin Red S-stained non-induced cul-
ture (osteogenesis control) showing no visible calcium depositions. (b) Alizarin Red S-stained
osteogenic induced culture showing the red calcium deposits within the bone matrix characteristic
of bone formation
7 Isolation and Characterization of Adipose-Derived Stromal Cells