Figure 5.

SIRT3 deacetylates SOD2 to maintain mitochondrial function and osteogenic differentiation. (a) mRNA levels of sirtuin family members in MC3T3-E1 cells under differentiation conditions for 48 h. (b) SIRT3 protein expression in cells under differentiation conditions for the indicated period. (c) SOD2 acetylation and specific K68 acetylation after 7 days of differentiation was analyzed by immunoprecipitation. (d) SOD2 acetylation of primary osteoblasts from wild-type and Sirt3−/− mice after 7 days of differentiation was analyzed by immunoprecipitation. (e) SOD2 activity of SIRT3-knockdown cells. (f) Mitochondrial superoxide levels were visualized by fluorescence staining. (g) OCR analysis of SIRT3-knockdown cells. (h) mRNA levels of Runx2, Osterix, ALP, and BSP from SIRT3-knockdown cells that were differentiated for 2 days. (i) ARS and ALP staining of SIRT3-knockdown cells that were differentiated for 7 days. (j) mRNA levels of osteogenic markers in SIRT3-knockdown cells after differentiation for 2 days with or without NAC. (k) mRNA levels of osteogenic markers in SOD2-knockdown cells after differentiation for 2 days with or without NAC. Data are presented as the mean±S.E.M. from at least three independent experiments. *P<0.05, **P<0.01 versus relative control