ATDC5 cells were transfected with Sox9 siRNA, Smad2/3 siRNA, or control non-specific (NS) siRNA. Sox9 mRNA expression was significantly reduced in cells expressing Sox9 siRNA (A). Acan mRNA was significantly reduced in the presence of Sox9 siRNA (B). Cells containing NS siRNA or Sox9 siRNA were treated with TGF-β1 and expression of Papss2 mRNA was measured by qPCR (C). In A–C * = p < 0.05 REST, n = 5. Western blots showed Smad2/3 protein levels were reduced in the presence of Smad2/3 siRNA, α-Tubulin was used as a loading control (n = 6) (D). Acan mRNA was significantly down-regulated in the Smad2/3 siRNA-transfected cells compared to cells containing NS siRNA (REST, * = p < 0.05, n = 2) (E). Cells containing NS or Smad2/3 siRNA siRNA were treated with TGF-β1 and Papss2 mRNA was, measured by qPCR (REST, * = p <0.05, n.s = not-significant n = 4) (F).