Figure 3

Runx2 regulates ST2 and Runx3 expressions in ATDC5. The evaluation of mRNA expression level of Runx2 (A), ST2L (B) and sST2 (C) in Runx2 silenced ATDC5 or Runx2 (D), ST2L (E) and sST2 (F) in Runx2-overexpressing ATDC5 cells (Runx2 cDNA stable transfection) by q-PCR (n = 5-6). (G and H) Representative immunoblotting and quantification of Runx2, ST2L and sST2 protein expressions in ATDC5 chondrocytes silenced by two different Runx2 siRNA sets. Transfection period was 72 hrs. (I and J) Representative immunoblotting and quantification of Runx2, ST2L and sST2 protein expressions in ATDC5 chondrocytes stably transfected with empty vector (pCMV6) or mouse Runx2 cDNA vector (pCMV6-Runx2) (n = 5-6). Figure I is the lane 1 and lane 4 of an immunoblot and full blot is shown in Supplemental Fig. 8. The qPCR evaluation of Runx3 mRNA level in siRunx2 ATDC5 (K) or Runx2 cDNA transfected (pCMV6-Runx2) ATDC5 cells (L) (n = 5-6). (M–P) The Runx3 protein expression analysis (representative immunoblotting and quantification) in Runx2 silenced (M and N) and Runx2-overexpressing ATDC5 chondrocytes (O–P). Runx2 knockdown was performed by two different Runx2 siRNA sets. Analysis was performed 48 to 72 hrs after siRNA or cDNA transfection. (Q–S) The qPCR of Runx3, ST2L and sST2 expressions in Runx3 silenced cells. The expression level of each gene was normalized to GAPDH. The protein quantifications are relative to their control and normalized to the respective β-actin level. *P ≤ 0.05, **P < 0.01.