Fig. 6

Knockdown of ATF6 using the siRNA approach inhibits chondrocyte hypertrophy, as revealed by collagen II, collagen X, and MMP13 expression. a, e siRNA against ATF6 mRNA efficiently inhibited expression of endogenous ATF6 in both ATDC5 (a) and C3H10 cells (e). Cells were infected with either Ad-ATF6 siRNA or control adenovirus (CTR), and total RNA was collected for real-time PCR. Expression of ATF6 was normalized against the GAPDH endogenous control. The normalized values were then calibrated against the control value, here set as 1. *p < 0.05. b, f Repression of ATF6 clearly abolished BMP2-induced Col II expression in ATDC5 (b) and C3H10 (f) cells. Transcript levels of Col II were detected by real-time PCR analysis of RNA isolated from micromass cultures of ATDC5 (b) or C3H10 (f) cells infected with siATF6 (MOI = 50) or control adenovirus in the presence of 300 ng/ml BMP2 at various time points, as indicated. *p < 0.05. c, g Repression of ATF6 obviously inhibits BMP2-induced Col X expression in ATDC5 (c) and C3H10 (g) cells. Transcript levels of Col X were detected by real-time PCR analysis of RNA isolated from micromass cultures of ATDC5 (c) or C3H10 (g) cells infected with siATF6 (MOI = 50) or control adenovirus in the presence of 300 ng/ml of BMP2 at various time points, as indicated. *p < 0.05. d, h Repression of ATF6 largely abolished BMP2-induced MMP13 expression in ATDC5 (d) and C3H10 (h) cells. Transcript levels of MMP13 were detected by real-time PCR analysis of RNA isolated from micromass cultures of ATDC5 (d) or C3H10 (h) cells infected with siATF6 (MOI = 50) or control adenovirus in the presence of 300 ng/ml BMP2 at various time points, as indicated. *p < 0.05. Error bars, SD