Fig. 3.

Laminin (LN) induces differentiation of cells with stem cells properties. (A) Ten-day-old LM05-E mammospheres were supplemented for 48 hours with vehicle (control) or 2 μg/mL LN, then dissociated and seeded at a concentration of 10,000 cells per well in 6-well low attachment culture plates under mammosphere forming conditions. The number of mammospheres was quantified after 6 days. (B) LM05-E cells were pretreated with Dulbecco’s modified Eagle’s medium/F12, 1% charcoal stripped fetal calf serum supplemented with 10 nM estradiol and 1 μM 4-OH-tamoxifen or vehicle (control) for 5 days. Subsequently, cells were washed and cultured in growth medium for 1 week, after which they were treated with vehicle (control) or 2 μg/mL LN for 48 hours. Finally, RNA was extracted to investigate the gene expression of Nanog, Sox 2, Oct-4, and glyceraldehyde 3-phosphate dehydrogenase (GADPH) using reverse transcriptase–polymerase chain reaction. Treatment with LN led to a decrease in the expression levels of genes associated with self-renewal. One of at least three experiments is shown for each case (**p < 0.01).