Fig. 6. Anxa5-lacZ⁺ cells differentiate into early chondrogenic, osteogenic and adipocyte lineages in vitro. Aggregate cultures of PVCs were cultivated in chondrogenic medium (A-F,M-O). By day 14 (B,C,E,F,M-O) or 21 (A,D), cryosections were performed and strong staining was seen for collagen VI (Col6; C) and collagen IX (Col9; B), whereas collagen II was barely detectable (Col2; A). Aggregate sections were stained for neutral fat deposits using oil Red O staining (red droplets) and Hematoxylin (M) as well as for PPAR2 (N). Additionally, isolated PVCs were cultivated in osteogenic medium for 21 days (G-L). Ca²⁺ deposits in cryosections of these aggregates were detected by Alizarin Red (H) as well as van Kossa staining (I). Collagen I was detected by immunostaining (G). Aggregates of PVCs were cultivated in proliferating medium as negative control for the staining of Ca²⁺ deposits (K,L). Secondary antibodies were used as negative control for the immunostaining (D-F,J,O). Scale bars: 50 µm.