Y-27632 Dihydrochloride

Y-27632 2HCl inhibits ROCK-II while displaying little activity against PKC, cAMP-dependent protein kinase and myosin light-chain kinase (MLCK) with K_i of 26 μM, 25 μM and > 250 μM, respectively, as well as PKA activated by another Rho-family GTPase member, Cdc42. This compound inhibits smooth-muscle contraction induces by various agonists including phenylephrine, histamine, acetylcholine, serotonin, endothelin, and thromboxane with IC50 of 0.3-1 μM, by selectively inhibiting Ca²⁺ sensitization. It suppresses Rho-induced, p160ROCK-mediated formation of stress fibres in cultured cells. [1]

This chemical treatment blocks both Rho-mediated activation of actomyosin and LPA-stimulated invasive activity of MM1 cells in a concentration-dependent manner. [2]

This compound treatment is not only sufficient to initiate formation of exuberant axonal processes but also facilitates axonal maturation during the very early stages of axonogenesis, while largely sparing axon elongation. [3]

In human embryonic stem (hES) cells, this chemical treatment at 10 μM markedly diminishes dissociation-induced apoptosis even in serum-free suspension (SFEB) culture, increases cloning efficiency (from ~1% to ~27%), facilitates subcloning after gene transfer, and enables SFEB-cultured hES cells to survive and differentiate into Bf1⁺ cortical and basal telencephalic progenitors. [4]

Oral administration of Y-27632 2HCl at 30 mg/kg significantly decreases the blood pressure in a dose-dependent manner in spontaneous hypertensive rats, renal hypertensive rats, as well as deoxycorticosterone acetate (DOCA)-salt hypertensive rats. [1]

When this compound is continuously administered at a rate of 0.55 μL per hour by implanted pumps for 11 days tumor cell invasion (MM1 cells expressing Val14-RhoA in rats) is significantly delayed. [2]

By inhibiting ROCK, this chemical treatment attenuates hypoxia-induced angiogenesis and vascular remodeling in the pulmonary circulation. [5] Pretreatment with this compound has a protective effect against tumor formation in albino mice with Ehrlich ascites carcinoma. [7]

• Phosphorylation reactions

  The p160ROCK is expressed in COS cells as tagged full-length proteins, and immunoprecipitated by the use of anti-tag antibodies. The p160ROCK (30 ng) is incubated with 40 μM [γ-³²P]ATP (3.3 Ci/mmol) and with 3 μg of either histone (HF2A), dephosphorylated casein or MBP in the presence of various concentrations of this compound at 30 °C in a total volume of 31 μL. A 7 μL aliquot is taken at 0, 5, 10, and 20 minutes, mixed with an equal volume of 2 × Laemmli sample buffer, and applied to SDS-PAGE. The gel is stained with Commassie Blue, dried and subjected to analysis by a Bioimage Analyzer BAS2000. The concentration of this chemical required to inhibit p160ROCK activity by 50% (IC50 value) is obtained. K_i value is calculated according to the equation, K_i = IC50/(1 + S/K_m), where S and K_m represent concentrations of and K_m value for ATP.

• Cell lines

  hES Cells

• Concentrations

  10 uM

• Incubation Time

  1 h

• Method

  Y-27632 2HCl was added to culture medium at 10 uM 1 h before detaching the cells from the feeder layer and also upon seeding the cells onto a new MEF layer.

• Animal Models

  Male Wistar rats with spontaneous or induced hypertension; Swiss albino mice with Ehrlich ascites carcinoma

• Dosages

  30 mg/kg/day (Rat); 0-10 mg/kg (mice)

• Administration

  Orally (Rat); i.p. (Mice)