Fig. 4.

FN III 13–14 mediate aggrecanase activity via TLR4. a Hip explants from 6-week-old C57B6 wild-type and TLR 4-/- null mice were dissected and rested in serum containing medium for 2 days. After this, explants were washed 3 times in serum-free medium and subjected to treatment with control medium (no additives), FN III 13–14 (1 μM), IL-1α (10 ng/ml) and LPS (10 ng/ml). After 2 days’ culture, conditioned medium was harvested and equal volumes of conditioned medium (50 μl) were deglycosylated using chondroitinase and keratanase. The samples were subjected to Western blotting using the anti-ALGS neoepitope antibody. The signal intensity of the samples (from triplicate wells, n = 3) was measured using Phoretics software and expressed as a mean and SEM. b Hip explants from Myd88-/- mice were treated with the same factors as described in a. Following 2 days’ culture, conditioned medium was harvested, deglycosylated and subjected to Western blotting as described in a with the anti-ALGS antibody.