FIG. 4.

Estrogen enhances the migration of thyroid cells. (A) About 2.5 × 10⁴ cells were resuspended in 500 μL of Rosswell Park Memorial Institute (RPMI) with 1% FBS ± E₂ and fulvestrant and seeded on upper chamber of BD Biocoat Control Inserts (8-μm pore membrane filters). Seven hundred and fifty microliters of Rosswell Park Memorial Institute (RPMI) containing 5% FBS was added to the bottom chamber as chemoattractant. After 18 hours, cells that migrated and adhered on the lower surface of the membrane were fixed, stained, and counted in 10 × field. The groups are as follows: untreated (white bars), E₂ treated (black bars), and E₂ + fulvestrant treated (gray bars). MCF-7 cells were used as a positive control for migration studies. Data are expressed as numbers of cells counted (migrated cells) per 10 × field micrograph for each sample well and normalized to the untreated control. The asterisk denotes statistically significant differences (p < 0.05) between experimental and control groups. (B) Scratch wound assay for Nthy-ori 3-1. (C) Scratch wound assay for BCPAP. About 5 × 10⁵ cells were plated and were allowed to grow to semiconfluent cell monolayers when three vertical wounds were caused per well using a pipette tip, and the cells were allowed to migrate in the presence of E₂ and/or fulvestrant. The cells were observed under 5 × field every 3 hours until the cells completely migrated from one end of scratch to other end.