Ru 486

A typical therapeutic oral dose of Dex is between 4 and 16 mg/day. The predicted concentration of Dex in the brain 6 hours after a 0.5 mg oral dose of Dex peaks at 5x10^-3 μg/ml [19 ]. This is equivalent to a 1.3x10^-8 M in vitro concentration. Therefore, a 4 mg dose is equivalent to an in vitro concentration of 1x10^-7 M. Cells were plated at 70% confluence and incubated for 24 hours, whereupon Dex (Sigma, MO, USA) was added from a 1x10^-3 M stock to a final concentration of 1x10^-7 M. The 49-residue functional upstream domain (FUD) of Streptococcus pyrogenes F1 adhesin has previously been shown to prevent assembly of a FN matrix [20 (link), 21 (link)]. A recombinant FUD (pUR-4) [22 (link)] was obtained from Dr. Jane Sottile through a UBMTA between the University of Rochester and Rutgers-RWJMS. FUD was added to tissue culture medium at a final concentration of 1 μg/ml. Cells were incubated with Dex or Dex+FUD overnight, prior to fixation. RU-486 (Abcam, Cambridge, MA) was added to culture medium at a final concentration of 1 mM weight/volume in DMSO.

The GBM-2 and GBM-3 cell lines used in this study were originally generated with approval of the Rutgers-Robert Wood Johnson Medical School Institutional Review Board under protocol #CINJ 001208, approved on 17 March 2017. Samples were anonymized, and the IRB waived the need for written informed consent. The lines used in this study have previously been published and characterized in [1 (link),47 (link)]. These lines were previously demonstrated to be highly dispersive. Cells were propagated in Earle’s minimal essential medium containing l-glutamine (ThermoFisher Scientific, Grand Island, NY, USA) and 10% fetal bovine serum (Atlanta Biologicals, Flowery Branch, GA, USA). Cells were cultured under standard tissue culture conditions of 37 °C, 5% CO₂ and 95% humidity. Final concentration of 0.1 µM dexamethasone (Cat#D2915, Sigma-Aldrich, St. Louis, MO, USA) and or 1 µM RU-486 (cat#ab120356, Abcam, Cambridge, MA, USA) were used for treatment.