Studies on Multifunctional Effect of All-Trans Retinoic Acid (ATRA) on Matrix Metalloproteinase-2 (MMP-2) and Its Regulatory Molecules in Human Breast Cancer Cells (MCF-7)
Figure 5
(a) Cell Adhesion Assay: MCF-7 cells (300,000 cells/1.5 mL) were grown in absence (Control) and in presence of 30 M ATRA for 24 hours in 10% MEM. Microtitre wells were coated with various concentrations of fibronectin and kept at 37°C for 1.5 hour. 1% BSA solution was used to block the nonspecific sites. Control and ATRA-treated cells were collected by trypsinization and 50,000 cells were added to each well. After 1 hour of incubation wells were washed thoroughly, number of bound cells was counted on a haemocytometer, and the % of cells adhered to the ligand was calculated. (b) RT-PCR of 5 & 1: MCF-7 (300,000 cells/5 mL) cells were grown in absence (lane C) and in presence of 30 M ATRA (lane E) for 24 hours in SFCM. Two steps RT-PCR (Retroscript, Ambion) was done as before with equal amounts of total RNA using specific primers for 5 & 1. GAPDH was used as internal control. (c) ELISA of E-cadherin: ELISA of E-cadherin was performed with anti-E-cadherin antibody in control (C) and ATRA treated (E) MCF-7 cells. () indicates that the difference in E-cadherin expression between control and ATRA treated MCF-7 cell is statistically significant.
