Fig. 4. Cdc42 silencing upregulates MMP-1 but not MMP-2 or ₁I mRNA steady-state levels. (A) Representative quantitative RT-PCR analysis of MMP-1, MMP-2 and ₁I mRNA levels, and of the 28S rRNA in total RNA extracted from HSFs cultured in DMEM containing 10% FCS 72 hours after transfection with calcium phosphate alone (CaP), an irrelevant siRNA (siScr) or an siRNA targeting RhoA (siRhoA), Rac1 (siRac1) or Cdc42 (siCdc42). Sample-to-sample variations in RT-PCR efficiency are controlled by adding a known copy number of synthetic RNA co-transcribed and co-amplified with the same primers, to generate a product of larger size (*). (B) Densitometric analysis of A. Results are the means±s.d. of three independent experiments. (C) Western-blot analysis of whole-cell lysates of HSFs transfected with calcium phosphate alone (CaP), with 20 nM siRNA targeting RhoA (siRhoA), Rac1 (siRac1) or Cdc42 (siCdc42), or with 20 nM irrelevant siRNA (siScr). 72 hours after transfection, the cells were lysed in SDS-PAGE loading buffer and analysed by immunoblotting with specific antibodies to Stat1 and ERK1/2.