|
|
|
|
|
|
|
|
|||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | |||||
|
Fig. 1. Blockade of integrin-mediated adhesion does not prevent cyclin D1 gene expression in MCF10A cells. (A) Serum-starved MCF10A cells were trypsinized, reseeded on dishes coated with collagen or agarose, and stimulated with 10% FBS and growth factor cocktail. Cell lysates were analyzed by western blotting for cyclin D1, Cdk6, phosphoY397-FAK, and FAK. (B) The experiment in A was repeated except that collected cells were analyzed for cyclin D1 mRNA by QPCR and western blotting using antibodies to dually phosphorylated ERK and total ERK. QPCR results show mean ± s.e.m. of two experiments after normalizing the levels of cyclin D1 mRNA to the level in serum-starved cells. (C) Quiescent MCF10A cells were preincubated in suspension with vehicle or AIIB2 β1-integrin-blocking antibody and RGD peptide. The cells were then plated on agarose-coated dishes and incubated for 3 and 9 hours with either 10% FBS and growth factor cocktail (GF), growth factor cocktail in serum-free medium, or growth factor cocktail in serum-free medium with AIIB2 and RGD. The suspended cells were collected by attachment to poly-L-lysine-coated coverslips and visualized by phase-contrast microscopy. Bar, 210 µm. (D) Attachment of MCF10A cells to collagen (COLL), vitronectin (VN), fibronectin (FN) or laminin-1 (LM) in the absence and presence of AIIB2 and/or RGD as described in the Materials and methods. The results are plotted as percentage maximal attachment relative to the positive control (no inhibitors) for each matrix protein.
|
