Skip to main content
. 2019 May 29;11(6):747. doi: 10.3390/cancers11060747

Figure 4.

Figure 4

HDACis decrease U87MG directional cell migration towards FBS in two different transwell migration systems. (A) Representative western blot analysis of histonic cell extracts obtained from U87 MG cells treated with indicated compounds at 50 nM for 24 h. Immunoblotting was performed analysing the acetylated H3K9K14. H4 was used as loading control (B) Quantitative analysis of directional cell migration towards FBS using Boyden chamber migration assay. U87MG cells, suspended in serum-free media, were treated with 50 nM HDACis or DMSO 0.1% and were plated in the upper compartment of the Boyden chambers. 10% FBS-DMEM was added in the lower compartment as a chemoattractant. The chambers were incubated (37 °C, 5% CO2) and migrated cells were counted after 3 h. The number of migrated cells, in treated and control groups, is expressed as a percentage of the no treated migrated cells (NT) in the absence of chemo-attractant (basal migration), taken as 100%. (C) Quantitative migration analysis using xCELLigence® Real-Time Cell Analyzer. U87 MG cells, suspended in serum-free media, were treated with the indicated compound at 50 nM and were plated into CIM-plate 16 with 8 μm pores; cells were seeded into the upper chamber with the lower chamber containing 10% FBS-DMEM as a chemoattractant. Experiments were carried out using the RTCA DP instrument, which was placed in a humidified incubator maintained at 37 °C with 5% CO2. The electronic sensors provided a continuous and quantitative measurement of the cell index (reflecting impedence, which depends on the number of attached cells) in each well. The CIM-plate 16 was monitored every 15 min for 24 h and the graph represents the percentage of cell index of treated groups respect to control group (DMSO), at 24 h. As shown, MS275, MC1568 and TSA significantly reduce the chemotactic response to FBS, in U87MG cells, as compared to cells migration in the presence of DMSO, the HDACis molecules solvent. Data show mean values from at least three independent measurements. Error bars show standard deviations. * p-value < 0.05.