Y-27632 (Y-27632) is an orally potent, ATP-competitive inhibitor of ROCK-I and ROCK-II. Y-27632 also inhibits isolation-induced apoptosis in mouse prostate stem or progenitor cells.

METHODS: Human induced pluripotent stem cells, marmoset iPSC, were treated with Y-27632 (5-20 μM) for 7 days and clone formation was detected by AKP.
RESULTS: Y-27632 significantly improved the cloning efficiency of marmoset iPSC. [1]
METHODS: Adult adipose tissue-derived stem cells ADSCs were treated with Y-27632 (5 μmol/L) for 1 h. The morphological changes of ADSCs were detected.
RESULTS: Y-27632 dose-dependently induced neuronal differentiation in ADSCs. the percentage of neuron-like cells in ADSCs treated with 5 μmol/L Y-27632 for 1 h was (93.5±4.7)%. [2]
METHODS: Crab monkey embryonic stem cells cyES were routinely passaged or treated with Y-27632 (1-10 μM) for 24 h. Live-dead staining was performed using the Flow Cytometry method, and BrdU was detected using a kit.
RESULTS: Y-27632 promoted the increase of cyES surviving cells. Y-27632 did not promote cell proliferation, but protected the cells from cell death after single-cell digestion. [3]

METHODS: To investigate the therapeutic potential of Y-27632 in motor neuron disease, Y-27632 (2 or 30 mg/kg in drinking water) was administered orally to SOD1G93A mice in the ALS model for 137 days.
RESULTS: Y-27632 2 mg/kg treatment was ineffective, Y-27632 30 mg/kg treatment improved motor function in male mice, and female mice showed only limited improvement. [4]
METHODS: To investigate the effect of Y-27632 on liver fibrosis, Y-27632 (30 mg/kg) was administered orally to rats with dimethylnitrosamine (DMN)-induced liver fibrosis once a day for four weeks.
RESULTS: Y-27632 treatment significantly reduced the incidence of DMN-induced hepatic fibrosis and lowered the levels of collagen and hydroxyproline as well as the expression of α-SMA in the liver. [5]

Recombinant ROCK-I, ROCK-II, PKN, or citron kinase is expressed in HeLa cells as Myc-tagged proteins by transfection using Lipofectamine, and is precipitated from the cell lysates by the use of 9E10 monoclonal anti-Myc antibody coupled to G protein-Sepharose. Recovered immunocomplexes are incubated with various concentrations of [32P]ATP and 10 mg of histone type 2 as substrates in the absence or presence of various concentrations of either Y-27632 or Y-30141 at 30°C for 30 min in a total volume of 30 μL of the kinase buffer containing 50 mM HEPES-NaOH, pH 7.4, 10 mM MgCl2, 5 mM MnCl2, 0.02% Briji 35, and 2 mM dithiothreitol. PKCa is incubated with 5 μM [32P]ATP and 200 μg/mL histone type 2 as substrates in the absence or presence of various concentrations of either Y-27632 or Y-30141 at 30°C for 10 min in a kinase buffer containing 50 mM Tris-HCl, pH 7.5, 0.5 mM CaCl2, 5 mM magnesium acetate, 25 μg/mL phosphatidyl serine, 50 ng/mL 12-O-tetradecanoylphorbol-13-acetate and 0.001% leupeptin in a total volume of 30 μL. Incubation is terminated by the addition of 10 μL of 43 Laemmli sample buffer. After boiling for 5 min, the mixture is subjected to SDS-polyacrylamide gel electrophoresis on a 16% gel. The gel is stained with Coomassie Brilliant Blue, and then dried. The bands corresponding to histone type 2 are excised, and the radioactivity is measured[1].

Y-27632 is dissolved in water and stored[1]. HeLa cells are plated at a density of 3×104 cells per 3.5-cm dish. The cells are cultured in DMEM containing 10% FBS in the presence of 10 mM Thymidine for 16 h. After the cells are washed with DMEM containing 10% FBS, they are cultured for an additional 8 h, and then 40 ng/mL of Nocodazole is added. After 11.5 h of the Nocodazole treatment, various concentrations of Y-27632 (0-300 μM), Y-30141, or vehicle is added and the cells are incubated for another 30 min[1].