A, ROCK-dependent increase in nuclear Sox9 after 2 hours of dynamic compression. Alginate-embedded normal human articular chondrocytes were subjected to 2 hours of continuous cyclic dynamic compression from 5–15% strain at 0.5Hz. Cells were fixed, released from alginate, and endogenous Sox9 was quantified by immunofluorescence microscopy. The amount of nuclear Sox9 was quantified by colocalization with the ToPro3 nuclear dye, shown in arbitrary units on the Y-axis. Error bars indicate the 95% confidence interval of the mean, n>300 cells for each group, * indicates p<0.01 compared to control. B, Increased Sox9 phosphorylation and total protein levels by overnight dynamic compression. Agarose-embedded chondrocytes were subjected to 16–18 hours of dynamic compression (DC), or free-swell uncompressed controls (FS). After dynamic compression, the gels were immediately homogenized in SDS-PAGE sample buffer and analyzed by western blot for the amount of phosphorylated Sox9Ser181 with an anti-phospho-Sox9Ser181 antibody. Blots were stripped and re-probed for total Sox9 protein levels. The insets graphically show the image densitometry of the immunoblot, error bars indicate the standard deviation from 3 donors.