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Figure 2


Fig. 2. Adhesion of mouse fibroblasts to collagen I after siRNA downregulation of integrin {alpha}2 expression. Subconfluent wild-type mouse fibroblasts were transfected once (A) or twice (B) with 100 nM each siRNA2 and siRNA3 targeting integrin {alpha}2 subunit or an unrelated siRNA as described in the Materials and Methods. Graphs show adhesion assays (30 minutes) to increasing concentrations of collagen I performed 76 hours after a single (A) or two successive (B) transfections. Equal numbers of control (unrelated siRNA; open symbols) and specific siRNA-transfected ({alpha}2-specific; closed symbols) fibroblasts were seeded on the collagen coats. The mean average of adherent cells in triplicate wells and s.d. are shown. Equal aliquots of fibroblasts transfected with control (c) and specific siRNAs (si) were lysed for immunoblotting analysis of integrin {alpha}2 subunit expression. The blots shown to the right of the graphs correspond to the fibroblasts used for the adhesion assays (arrow indicates the position of integrin {alpha}2 subunit). Densitometry measurement of band intensity in the blots shown in A and B indicate that expression of the {alpha}2 subunit is downregulated to 26% and 19%, respectively, of the corresponding controls. Molecular size markers are indicated in kDa on the left of the blots.





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