Furthermore, EGF treatment in creased CXCR4 mRNA expression http://www.selleckchem.com/products/SB-203580.html in the control cells but had no effect on the increased CXCR4 expression in the COUP cells. Treatments with EGFR and MEK inhibitors decreased CXCR4 expression, reaching a lower level of expression than under the basal conditions. Indeed, the inhibition of EGFR and MAPK signaling abol ished the stimulation effect of COUP TFI on CXCR4 expression. COUP TFI overexpression modifies cells response to CXCL12 signal The CXCL12/CXCR4 axis plays major roles in breast cancer cell proliferation, migration, and invasion. thus, we analyzed the CXCL12 mediated growth and motility of MCF 7 cells overexpressing COUP TFI. First, we tested the control and COUP cells for a prolif erative response to CXCL12 treatment by exposing the cells to CXCL12 for 7 days and quantifying the total cell number.
No significant differ ences were observed with regard to the basal growth of the control and COUP cells. however, when treated with CXCL12, both cells proliferated significantly more than under the control condition. Furthermore, the COUP cells displayed a significantly higher proliferative response to the CXCL12 treatment than the control cells. The cell migratory behavior was then assayed. We an alyzed the capacity of the control and COUP cells to mi grate through a PET membrane with an 8 um filter pore toward a low serum concentration medium, which rep resented the basal migration, or toward a CXCL12 gra dient, which corresponded to the induced migration. After a 24 h period, the relative number of basal migrant cells was almost twice as high for the COUP cells than the control cells.
Moreover, when CXCL12 was added to the lower chamber, the migration of the control and COUP cells increased versus the basal condition. The relative induced migra tion of the COUP cells was 3 times higher than that of the control cells. Interestingly, the specific CXCR4 in hibitor AMD3100 completely abolished the CXCL12 induced migration observed in Anacetrapib the control and COUP cells. No significant difference in migration capability between the two clones was detected. Next, we tested the hypothesis that the reduction of CXCL12 by the COUP cells is important for their migra tion toward a CXCL12 gradient. We therefore added hu man recombinant CXCL12 to both the upper chamber and the lower chamber. Under these conditions, the migratory behavior of the control and COUP cells was dramatically altered, with both clones displaying an ap proximately similar migration potential. Thus, disruption of the CXCL12 gradient by ectopic CXCL12 added to the upper chamber prior to the migration test hampered the migration of both the control and COUP clones.