Activation of autophagy decreased osteogenic responses. (A) Activation of autophagy by rapamycin-reduced osteogenic responses in diseased AVICs. Diseased AVICs were treated by rapamycin for 24 h at indicated dose, autophagy indicator LC3-II, and osteogenic factors ALP and BMP-2 are examined by immunoblotting. Results are expressed as mean ± standard error of the mean; n = 5; *P < 0.05. #Significant with each other; &significant with each other for ALP; $significant with each other for LC3-II. (B) Overexpression (OE) of Atg7 decreased osteogenic responses in diseased AVICs. Diseased AVICS were infected with adAtg7 or adbgal (Control, Ctrl) for 48 h; Atg7, LC3-II, BMP-2, and ALP protein levels were analyzed by immunoblotting. Bar graphs indicate densitometry data of LC3-II, BMP-2, and ALP changes. Results are expressed as mean ± standard error of the mean; n = 5; *P < 0.05 (OE compared with Ctrl); &P < 0.05 (OE compared with Ctrl for ALP). (Color version of figure is available online.)