Phosphorylation of CPI‐17 and MYPT in rabbit bladder smooth muscle: Role of PKC, Rho kinase, or both

Smooth muscle contraction is regulated by phosphorylation of the myosin light chain (MLC) catalyzed by MLC kinase and dephosphorylation catalyzed by MLC phosphatase. Agonist stimulation of smooth muscle results in the inhibition of MLC phosphatase activity and a net increase in MLC phosphorylation and therefore force. The two pathways believed to be primarily important for inhibition of MLC phosphatase activity are protein kinase C (PKC)-catalyzed CPI-17 phosphorylation and Rho kinase (ROCK)-catalyzed myosin phosphatase-targeting subunit (MYPT1) phosphorylation. The goal of this study was to determine the roles of PKC and ROCK and their downstream effectors in regulating MLC phosphorylation levels and force during the phasic and sustained phases of carbachol-stimulated contraction in intact bladder smooth muscle. These studies were performed in the presence and absence of the PKC inhibitor bisindolylmaleimide-1 (Bis) or the ROCK inhibitor H-1152. Phosphorylation levels of Thr38-CPI-17 and Thr696/Thr850-MYPT1 were measured at different times during carbachol stimulation using site-specific antibodies. Thr38-CPI-17 phosphorylation increased concurrently with carbachol-stimulated force generation. This increase was reduced by inhibition of PKC during the entire contraction but was only reduced by ROCK inhibition during the sustained phase of contraction. MYPT1 showed high basal phosphorylation levels at both sites; however, only Thr850 phosphorylation increased with carbachol stimulation; the increase was abolished by the inhibition of either ROCK or PKC. Our results suggest that during agonist stimulation, PKC regulates MLC phosphatase activity through phosphorylation of CPI-17. In contrast, ROCK phosphorylates both Thr850-MYPT1 and CPI-17, possibly through cross talk with a PKC pathway, but is only significant during the sustained phase of contraction. Last, our results demonstrate that there is a constitutively activate pool of ROCK that phosphorylates MYPT1 in the basal state, which may account for the high resting levels of MLC phosphorylation measured in rabbit bladder smooth muscle.

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Changes in Urinary Bladder Smooth Muscle Contractility and the Role of Rho Kinase and Isoprostane after Partial Bladder outlet Obstruction in rats

Bulletin of Egyptian Society for Physiological Sciences ◽

10.21608/besps.2012.35784 ◽

2012 ◽

Vol 32 (2) ◽

pp. 1-16

Author(s):

Abeer Dief ◽

Adham Mohamed ◽

Gehan Sharara

Keyword(s):

Smooth Muscle ◽

Bladder Outlet Obstruction ◽

Rho Kinase ◽

Outlet Obstruction ◽

Bladder Smooth Muscle ◽

Muscle Contractility ◽

Smooth Muscle Contractility ◽

Partial Bladder Outlet Obstruction ◽

Urinary Bladder Smooth Muscle

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The Role of Signal Transducer and Activator of Transcription 3 (Stat3) in Stretch Injury to Bladder Smooth Muscle Cells

Journal of Pediatric Urology ◽

10.1016/j.jpurol.2007.01.047 ◽

2007 ◽

Vol 3 ◽

pp. S35

Author(s):

Sarel Halachmi ◽

Karen Aitken ◽

Martha Szybowska ◽

Nesrin Sabha ◽

Shariff Dessouki ◽

...

Keyword(s):

Smooth Muscle ◽

Smooth Muscle Cells ◽

Muscle Cells ◽

Signal Transducer ◽

Bladder Smooth Muscle ◽

Stretch Injury ◽

Transcription 3

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Fatty acid profiles in normal and obstructed rabbit bladder smooth muscle and mucosa

Neurourology and Urodynamics ◽

10.1002/(sici)1520-6777(1999)18:6<697::aid-nau20>3.0.co;2-m ◽

1999 ◽

Vol 18 (6) ◽

pp. 697-711 ◽

Cited By ~ 19

Author(s):

Martha A. Hass ◽

Elena Leonova ◽

Robert M. Levin

Keyword(s):

Smooth Muscle ◽

Fatty Acid ◽

Fatty Acid Profiles ◽

Bladder Smooth Muscle ◽

Rabbit Bladder

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PKC‐dependent inhibition of MAP kinase activity and caldesmon phosphorylation in rabbit bladder smooth muscle (865.14)

The FASEB Journal ◽

10.1096/fasebj.28.1_supplement.865.14 ◽

2014 ◽

Vol 28 (S1) ◽

Author(s):

Dongyu Wei ◽

Sarah Sivilich ◽

Robert Moreland

Keyword(s):

Smooth Muscle ◽

Map Kinase ◽

Kinase Activity ◽

Bladder Smooth Muscle ◽

Rabbit Bladder ◽

Dependent Inhibition

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Resveratrol’s Impact on Vascular Smooth Muscle Cells Hyporeactivity: The Role of Rho-Kinase Inhibition

BioMed Research International ◽

10.1155/2020/9012071 ◽

2020 ◽

Vol 2020 ◽

pp. 1-8

Author(s):

Michał Wiciński ◽

Bartosz Malinowski ◽

Paweł Rajewski ◽

Paweł Szychta ◽

Eryk Wódkiewicz ◽

...

Keyword(s):

Smooth Muscle ◽

Protein Kinases ◽

Vascular Smooth Muscle Cells ◽

Kinase Inhibitor ◽

Rho Kinase ◽

Pressure Effects ◽

Kinase Inhibition ◽

Rho Kinase Inhibitor ◽

Dependent Protein

Resveratrol (3,5,4′-trihydroxystilbene) is a chemical compound belonging to the group of polyphenols and flavonoids. The aim of the present study was to determine the influence of resveratrol application along with certain modulating factors, such as 8Br-cGMP-activator of cGMP-dependent protein kinases, HA-1077-Rho-kinase inhibitor, and Bay K8644-calcium channel agonist, on VMSCs constriction triggered by phenylephrine. Resveratrol at a dose of 10 mg/kg/24 h administered for 4 weeks reduced the reactivity of the arteries to the pressure action of catecholamines. Tests performed after four weeks of resveratrol administration showed that 8Br-cGMP at the concentrations of 0.01 mM/l and 0.1 mM/l intensifies this effect. Simultaneous resveratrol and Bay K8644 administration led to a significant decrease in contractility compared to the vessels collected from animals (Res−). This effect was dependent on the concentration of Bay K8644. Resveratrol seems to be counteractive against Bay K8644 by blocking L-type calcium channels. As the concentration of HA-1077 increased, there was a marked hyporeactivity of the vessels to the pressure effects of phenylephrine. The results indicate synergy between resveratrol and Rho-kinase inhibition.

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Activation of Rho signaling contributes to lysophosphatidic acid-induced contraction of intact ileal smooth muscle of guinea-pig

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y00-050 ◽

2000 ◽

Vol 78 (9) ◽

pp. 729-736 ◽

Cited By ~ 15

Author(s):

Mayumi Mori ◽

Hiromi Tsushima

Keyword(s):

Smooth Muscle ◽

Guinea Pig ◽

Lysophosphatidic Acid ◽

Membrane Fraction ◽

Smooth Muscles ◽

Rho Kinase ◽

Specific Inhibitor ◽

High K ◽

Rho A

To elucidate the possible role of Rho A/Rho-kinase on lysophosphatidic acid (LPA)-induced contraction in intact guinea-pig ileal smooth muscle, we examined effects of pretreatment with a specific inhibitor of Rho-kinase (Y-27632) on the LPA-induced contraction and MLC20 phosphorylation. In addition, we investigated whether LPA actually elicits an activation of Rho A by studying subcellular distribution of Rho A in unstimulated and stimulated smooth muscles by LPA. LPA induced a less intense, but sustained, contraction compared with ACh, and was accompanied by significant increases in MLC20 phosphorylation. The effects of LPA on tension and MLC20 phosphorylation were inhibited by Y-27632. The ACh-induced contraction, but not increases in MLC20 phosphorylation, was partially inhibited by Y-27632. High K+-induced contraction was unaffected by the inhibitor. LPA stimulated translocation of Rho A from the cytosol to the membrane fraction of the muscle. Translocation of Rho A was also induced by ACh and high K+. These results suggest that LPA-induced contraction of intact ileal smooth muscle is dominated through activation of Rho A and Rho-kinase and subsequent increases in MLC20 phosphorylation.Key words: lysophosphatidic acid, Rho, Rho-kinase, ileal smooth muscle.

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