siRNA knock down of casein kinase 2 increases force and cross-bridge cycling rates in vascular smooth muscle

Smolock, Elaine M.; Wang, Tanchun; Nolt, Jocelyn; Moreland, Robert S.

doi:10.1152/ajpcell.00343.2006

Cited by 13 publications

(21 citation statements)

References 56 publications

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“…Calcium channel function in organ-cultured smooth muscle tissues and cell-cultured smooth muscle cells have been shown to significantly decrease as time in culture increases (20,26). However, consistent with previous studies from our laboratory using organ culture of vascular smooth muscle tissues (21,48,49), functional voltage-dependent calcium channels are retained. This is based on the observation that both vascular and BSM tissues subjected to organ culture exhibit rapid and robust contractions in response to KCl-PSS (21, 48, 49 and Figs.…”

Section: Quantification Of Rock-i and Pkg In Organ-cultured Bsm Tissuesupporting

confidence: 85%

“…Myosin content, however, in the Hellstrand study (35) was decreased by 50%. This is consistent, in terms of force development, with our previous work showing that a vascular preparation could be subjected to organ culture for up to 9 days in a serum-free media (21,48,49). More recently, Corteling et al (16) again using a vascular preparation, showed no loss of contractility in response to UTP stimulation, no decrease in caldesmon content, but a 50% loss of calponin.…”

supporting

confidence: 89%

“…Organ culture of smooth muscle tissues has already been applied to several smooth muscle types, such as carotid and pulmonary arteries, in conjunction with genetic manipulation of specific smooth muscle-regulatory proteins (23,30,40,48,49). Early studies by Hellstrand's (35) and Karaki's laboratories (66) demonstrated that force could be maintained in an organ-cultured vascular preparation for up to 4 days if cultured in a serum-free media.…”

mentioning

confidence: 99%

“…Specifically, in this study, we determined the phenotype of the organ-cultured tissue in terms of the content of smooth muscle-specific isoforms of contractile proteins, contractility, viability of cells within the tissue, and functional membrane ion channels. Based on our previous studies using vascular smooth muscle subjected to organ culture (48,49), we hypothesized that BSM strips would maintain their contractile phenotype including tissue contractility, expression of smooth muscle-specific protein isoforms, and typical morphology for up to 9 days in organ culture if cultured under stretch in a serum-free media and subjected to daily contraction-relaxation. We have provided a novel and what we believe will be a very valuable tool for the application of molecular biology techniques to study not only the basic regulation of BSM contraction but also the changes that occur in pathophysiological states such as overactive bladder or voiding dysfunction that occurs in response to benign prostatic hyperplasia or diabetic uropathy, using a simple in vitro system which would allow modification of the external milieu.…”

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confidence: 99%

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Bladder smooth muscle organ culture preparation maintains the contractile phenotype

Wang

Kendig

Chang

et al. 2012

American Journal of Physiology-Renal Physiology

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Smooth muscle cells, when subjected to culture, modulate from a contractile to a secretory phenotype. This has hampered the use of cell culture for molecular techniques to study the regulation of smooth muscle biology. The goal of this study was to develop a new organ culture model of bladder smooth muscle (BSM) that would maintain the contractile phenotype and aid in the study of BSM biology. Our results showed that strips of BSM subjected to up to 9 days of organ culture maintained their contractile phenotype, including the ability to achieve near-control levels of force with a temporal profile similar to that of noncultured tissues. The technical aspects of our organ culture preparation that were responsible, in part, for the maintenance of the contractile phenotype were a slight longitudinal stretch during culture and subjection of the strips to daily contraction-relaxation. The tissues contained viable cells throughout the cross section of the strips. There was an increase in extracellular collagenous matrix, resulting in a leftward shift in the passive length-tension relationship. There were no significant changes in the content of smooth muscle-specific α-actin, calponin, h-caldesmon, total myosin heavy chain, protein kinase G, Rho kinase-I, or the ratio of SM1 to SM2 myosin isoforms. Moreover the organ cultured tissues maintained functional voltage-gated calcium channels and large-conductance calcium-activated potassium channels. Therefore, we propose that this novel BSM organ culture model maintains the contractile phenotype and will be a valuable tool for the use in cellular/molecular biology studies of bladder myocytes.

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Section: Quantification Of Rock-i and Pkg In Organ-cultured Bsm Tissuesupporting

confidence: 85%

supporting

confidence: 89%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Bladder smooth muscle organ culture preparation maintains the contractile phenotype

Wang

Kendig

Chang

et al. 2012

American Journal of Physiology-Renal Physiology

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“…RIP-2 phosphorylates p38 MAPK (22), so the inhibition of RIP-2 would simply inhibit p38 MAPK. Casein kinase-2 knockdown using short interfering RNA resulted in an increase in force responses in vascular smooth muscle (38), suggesting that a similar mechanism of force modulation may be present in the myocardium.…”

Section: Discussionmentioning

confidence: 98%

Myocardial adenosine A₁-receptor-mediated adenoprotection involves phospholipase C, PKC-ε, and p38 MAPK, but not HSP27

Fenton

Shea

Doddi

et al. 2010

American Journal of Physiology-Heart and Circulatory Physiology

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Adenosine via an adenosine A1 receptor (A1R) is a negative feedback inhibitor of adrenergic stimulation in the heart, protecting it from toxic effects of overstimulation. Stimulation of the A 1R results in the activation of Gi protein, release of free G␤␥-subunits, and activation/translocation of PKC-ε to the receptor for activated C kinase 2 protein at the Z-line of the cardiomyocyte sarcomere. Using an anti-G␤␥ peptide, we investigated the role of these subunits in the A1R stimulation of phospholipase C (PLC), with the premise that the resulting diacylglycerol provides for the activation of PKC-ε. Inositol 1,4,5-triphosphate release was an index of PLC activity. Chlorocyclopentyl adenosine (CCPA), an A1R agonist, increased inositol 1,4,5-triphosphate production by 273% in mouse heart homogenates, an effect absent in A1R knockout hearts and inhibited by anti-G␤␥ peptide. In a second study, p38 MAPK and heat shock protein 27 (HSP27), found by others to be associated with the loss of myocardial contractile function, were postulated to play a role in the actions of A1R. Isoproterenol, a ␤-adrenergic receptor agonist, increased the Ca 2ϩ transient and sarcomere shortening magnitudes by 36 and 49%, respectively. In the rat cardiomyocyte, CCPA significantly reduced these increases, an action blocked by the p38 MAPK inhibitor SB-203580. While CCPA significantly increased the phosphorylation of HSP27, this action was inhibited by isoproterenol. These data indicate that the activation of PKC-ε by A1R results from the activation of PLC via free G␤␥-subunits released upon A1R-induced dissociation of Gi␣␤␥. Attenuation of ␤-adrenergic-induced contractile function by A1R may involve the activation of p38 MAPK, but not HSP27. G␤␥ subunits; antiadrenergic; rodent; contractility ADENOSINE IS RELEASED ENDOGENOUSLY in the heart that is stressed by hypoxia, ischemia, or adrenergic stimulation (13,15). In the interstitium, adenosine activates cell surface receptors that initiate signal cascades within the cardiomyocyte, ultimately reducing the cardiotoxic manifestations of the initial insult (16,43). For many years, study has focused on the antiadrenergic action of adenosine, or the ability of this nucleoside to protect the heart against excessive stimulation (34) by norepinephrine released during ischemia (36). In manifesting this action, adenosine A 1 receptors (A 1 R) attenuate ␤-adrenergic catecholamine-elicited increases in G s protein cycling (17), adenylyl cyclase activity (33), cAMP formation (5), activation of protein kinase A (PKA) (6), protein phosphorylation (12), myocardial Ca 2ϩ transient magnitude (14), and ventricular contractility (6, 16).Adenosine also reduces the responsiveness of the myocardium to catecholamine stimulation by a mechanism independent of changes in adrenergic-stimulated cyclase activity. This adenoprotection by the A 1 R agonist chlorocyclopentyl adenosine (CCPA) has been demonstrated by us to involve the translocation of PKC-ε to receptor for activated C kinase 2 (RACK2) in rat and mouse myocard...

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Cell rounding in cultured human astrocytes and vascular endothelial cells upon inhibition of CK2 is mediated by actomyosin cytoskeleton alterations

Kramerov

Ahmed

Ljubimov

2012

J of Cellular Biochemistry

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Protein kinase CK2 participates in a wide range of cellular events, including the regulation of cellular morphology and migration, and may be an important mediator of angiogenesis. We previously showed that in the retina, CK2 immunolocalizes mostly to vascular endothelium and astrocytes in association with the cytoskeleton. Additionally, CK2 inhibitors significantly reduced retinal neovascularization and stem cell recruitment in the mouse model of oxygen-induced proliferative retinopathy. We have also shown that CK2 and F-actin co-localized in actin stress fibers in microvascular endothelial cells, and that highly specific CK2 inhibitors caused cell rounding in astrocytes and microvascular endothelial cells, which was alleviated by serum that promotes spreading by Rho/Rho-kinase (RhoK) activation of myosin II. Therefore, we examined a possible role of CK2 in the regulation of actin–myosin II-based contractility. Treatment with CK2 inhibitors correlated with disassembly of actomyosin stress fibers and cell shape changes, including cytoplasmic retraction and process formation that were similar to those occurring during astrocyte stellation. Low doses of specific inhibitors of kinases (RhoK and MLCK) that phosphorylate myosin light chain (MLC) enhanced the effect of suboptimal CK2 inhibition on cell shape. Such striking stellation-like alteration was accompanied by decreased level of phospho-MLC, thus implying a CK2 role in regulation of actomyosin cytoskeleton. Our results suggest an important role of CK2 in the control of cell contractility and motility, which may account for suppressing effect of CK2 inhibition on retinal neovascularization. Together, our data implicate protein kinase CK2 for the first time in stellation-like morphological transformation.

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siRNA knock down of casein kinase 2 increases force and cross-bridge cycling rates in vascular smooth muscle

Cited by 13 publications

References 56 publications

Bladder smooth muscle organ culture preparation maintains the contractile phenotype

Bladder smooth muscle organ culture preparation maintains the contractile phenotype

Myocardial adenosine A₁-receptor-mediated adenoprotection involves phospholipase C, PKC-ε, and p38 MAPK, but not HSP27

Cell rounding in cultured human astrocytes and vascular endothelial cells upon inhibition of CK2 is mediated by actomyosin cytoskeleton alterations

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siRNA knock down of casein kinase 2 increases force and cross-bridge cycling rates in vascular smooth muscle

Cited by 13 publications

References 56 publications

Bladder smooth muscle organ culture preparation maintains the contractile phenotype

Bladder smooth muscle organ culture preparation maintains the contractile phenotype

Myocardial adenosine A1-receptor-mediated adenoprotection involves phospholipase C, PKC-ε, and p38 MAPK, but not HSP27

Cell rounding in cultured human astrocytes and vascular endothelial cells upon inhibition of CK2 is mediated by actomyosin cytoskeleton alterations

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Myocardial adenosine A₁-receptor-mediated adenoprotection involves phospholipase C, PKC-ε, and p38 MAPK, but not HSP27