Y-39983 Dihydrochloride

Y-39983 opposes ROCK-dependent phosphorylation of MYPT1 predominantly at Thr853 with a corresponding decrease in MLC phosphorylation. It reduces the phosphorylation at Thr696 and Thr853 and leads to reduced actomyosin contraction. Y-39983 shows an IC50 of ~200 nM for dephosphorylation at Thr696 and only 15 nM for dephosphorylation at Thr853. The IC50 for dephosphorylation of MLC is 14 nM^[3].

Cell cultures of an immortalized human glaucomatous trabecular meshwork (GTM3) cell line are grown at 37℃ in 5% CO2 in Dulbecco's Minimum Essential Medium (DMEM + Glutamax) supplemented with 10% fetal bovine serum and 10 μg/ml of gentamicin. Upon reaching confluence, cells are divided using 0.05% trypsin. Cells of passages 13 to 20 are used in experiments. Cells are treated with 5 μM of the specific ROCK inhibitors, Y-27632 and Y-39983, for 1 h in a serum-rich medium. Cell lysate is used for western blotting to test effect of ROCK inhibitors on the phosphorylation of MYPT1 and MLC.

Y-39983 significantly suppresses clinical symptoms of EAE and prevents its relapse while increasing the amount of myelin proteins. Its treatment results in reduced demyelination with no significant change in axonal damage, which may be due to the inactivation of ROCK substrates, including phosphorylated (p)-MLC, LIMK2 and CRMP-2, which are important for neurite outgrowth, growth cone collapse and remyelination of oligodendrocytes^[2].