AP20187 is a cell-permeable molecule used to dimerize FK506-binding protein (FKBP) fusion proteins and initiate biological signaling cascades and gene expression or disrupt protein-protein interactions. Real-time PCR analysis shows that AP20187 (0.5 mg/kg, 2 mg/kg, or 5 mg/kg) treatment significantly increases the levels of CHOP mRNA in the CNS of PLP/Fv2E-PERK mice at PID12. AP20187 treatment significantly alleviates EAE-induced myelin damage in these mice. AP20187 treatment significantly reduces the number of degenerating axons and increases the density of axons in the demyelinating lesions in the lumbar spinal cord of PLP/Fv2E-PERK mice.
For the in vitro study, 16 h after ADV infection, cells are treated
with R1881 (10 nM), AP20187 (10 nM), both, or neither for 8 h. Cells are
then rinsed with PBS and fixed with 4% paraformaldehyde for 1 h at room
temperature. After rinsing with PBS, cells are incubated in ice-cold
permeabilization solution (0.1% Triton X-100, 0.1% sodium citrate) for 2
min at 0°C. Cells are rinsed with PBS and stained with TUNEL reaction
mixture for 60 min at 37°C. After another PBS wash, cells are incubated
with Converter-AP for 30 min at 37°C. Cells are rinsed and incubated
with substrate 5-bromo-4-chloro-3-indolyl phosphate/nitroblue
tetrazolium for 30 min. After a final PBS rinse (repeated twice), cells
are microphotographed.
To activate the transgene Fv2E-PERK in oligodendrocytes, PLP/Fv2E-PERK
transgenic mice are given intraperitoneal injections of AP20187 daily
at a dose of 0.5 mg/kg, 2 mg/kg, or 5 mg/kg. Lyophilized AP20187 is
dissolved in 100% ethanol at a concentration of 62.5 mg/mL stock
solution and stored at −20°C. Injection solutions consist of 4% ethanol,
10% PEG-400, and 2% Tween-20 in water. The transgenic mice receiving
only the vehicle (4% ethanol, 10% PEG-400, 2% Tween-20 in water) served
as controls.
MCE has not independently confirmed the accuracy of these methods. They are for reference only.
