Increased expression of RhoA or RhoC GTPAses and or their ROCK1 2 effectors has been reported in several metastatic cancers, and they play important roles in tumour progression selleck catalog and invasion. These mole cules also partake in mechanotransduction of signals in response to external tensional stimuli. This work investigates the role of dense collagen matri ces that resemble the matrix densities of mammary breast cancer tissues in regulating the migration of tumour cells. MTLn3 rat mammary carcinoma cells were observed to maneuver between collagen fibrils during migration into dense matrices utilizing cell contractility. These cells also have significantly higher ROCK1 activity levels in high density compared to low density matrices, indicating matrix dependent regulation.

ROCK1 levels and activity were sensitive to HDAC inhibition by MS 275, which was abrogated when Notch1 was blocked. Inhibition of ROCK1 and metallo proteases by themselves had no effect on cell migration indicating alternation of invasion strategies. However, in the presence of both inhibitors, cell migration was sig nificantly blocked. Results Preparing in vitro collagen matrices with similar collagen content and organization to high mammographically dense tissues Regions of low or high mammographic density in prophylactic invasive ductal carcinoma tissues were macrodissected and processed for imaging and quantitative analyses. Massons Trichrome staining showed that HMT regions contained mostly collagen with isolated clusters of glandular cells whereas low mammographically dense tissues consisted mainly of adipose cells with little presence of collagen.

Collagen concentrations were esti mated using picrosirius red to stain collagen and meas uring dye uptake at 531 nm absorbance wavelength. Compared against known standards, the collagen content in LMT and HMT were measured to be 2. 65 1. 60 and 19. 59 2. 91 mg cm3, respectively. High density collagen matrix was prepared by centrifugation to increase collagen concen trations and polymerisation using vaporised NH4OH. A centrifugation time of 60 min was found to be suitable for preparing HD matrices at 19. 16 0. 74 mg cm3, similar to that for HMT extracts. The fibril densities were comparable between HMT tissue and HD matrix measuring 0. 63 0. 08 and 0. 61 0. 07 mm of fibril mm2, respectively. Similarly, in LMT tissue, the density 0.

19 0. 09, was similar to that of LD matrix which measured 0. 19 0. 07 mm of fibril mm2. Pore sizes between tumour tissue and in vitro matrices are also comparable with HMT and HD pore sizes measuring 0. 025 0. 014 and 0. 017 0. 011, res pectively, while those of LMT and LD measure 0. 678 0. 458 and 0. 799 0. 695, respectively. The colla gen fibril size of AV-951 the HD matrix was very similar to HMT tissue, with 62% of the fibrils lying within 125 225 nm for both matrices.