Figure 6From: Brg-1 mediates the constitutive and fenretinide-induced expression of SPARC in mammary carcinoma cells via its interaction with transcription factor Sp1Analyses of SPARC expression and secretion from mammary tumor cell lines with different metastatic capacities. Cells were seeded into 6-well plates at a density of 1 × 105 cells (4T1 or 168FARN) or 1.5 × 105 cells (67NR) per well and cultured at 37°C. Cell-free medium served as a control. Seventy-two hrs after culture, cell media were collected, and total RNA as well as protein extracts were prepared from the three cell lines. (A) Real-time qRT-PCR was used to analyze the relative expression of SPARC mRNA level. The SPARC mRNA was normalized by GAPDH expression and the relative expression level is represented as a fold of the SPARC mRNA level in 4T1 cells. (Data represented as mean ± SEM, n = 4). *P < 0.01, compared with 4T1; **P < 0.01, compared with 168FARN. (B) Cellular SPARC protein expression was assessed using Western blot analysis. The illustrated result is a representative of three independent experiments. (C) The levels of secreted SPARC protein in the culture media were assessed by Western blot analysis. Cell-free medium (Med) was used as a control. Densitometric quantitation was performed and results were adjusted for total protein content of cell lysate (densitometry/μg cell total protein). The relative level of secreted SPARC in the medium of 4T1 cells is set as 1. Data represented as mean ± SEM (n = 3). *P < 0.01, compared with 4T1; **P < 0.01, compared with 168FARN.Back to article page