| 1. |
- Forte, A, et al.
(author)
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c-Myc antisense oligonucleotides preserve smooth muscle differentiation and reduce negative remodelling following rat carotid arteriotomy
- 2005
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In: Journal of Vascular Research. - : S. Karger AG. - 1423-0135 .- 1018-1172. ; 42:3, s. 214-225
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Journal article (peer-reviewed)abstract
- Objectives: The vascular biology of restenosis is complex and not fully understood, thus explaining the lack of effective therapy for its prevention in clinical settings. The role of c-Myc in arteriotomy-induced stenosis, smooth muscle cell (SMC) differentiation and apoptosis was investigated in rat carotids applying full phosphorothioate antisense ( AS) oligonucleotides (ODNs). Methods: Carotid arteries from WKY rats were submitted to arteriotomy and to local application of ODNs through pluronic gel. Apoptosis ( deoxynucleotidyl transferase-mediated dUTP nick end-labelling), SMC differentiation (SM22 immunofluorescence) and vessel morphology and morphometry ( image analysis) were determined 2, 5 and 30 days after injury, respectively. Results: AS ODNs induced a 60% decrease of target c-Myc mRNA 4 h after surgery in comparison to control sense ( S) and scrambled ODN-treated carotids (p < 0.05). A significant 37 and 50% decrease in SM22 protein in the media of S ODN-treated and untreated carotids was detected when compared to uninjured contralateral arteries (p < 0.05). This reduction in SM22 expression was prevented in AS ODN-treated carotids. Stenosis was mainly due to adventitial constrictive remodelling. Lumen area in AS ODN-treated carotids was 35% greater than in control arteries 30 days after surgery (p < 0.05). TUNEL assay revealed increased apoptosis in AS ODN-treated carotids (p < 0.05). Conclusions: c-Myc AS ODNs reduce arteriotomy-induced negative remodelling. This is accompanied by maintained SMC differentiation and greater apoptosis. The combination of reduced c-Myc-induced proliferation and increased apoptosis may thus underlie the less severe remodelling upon treatment with c-Myc mRNA AS ODN.
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| 2. |
- Kumar, B, et al.
(author)
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Upregulated TRPC1 Channel in Vascular Injury In Vivo and Its Role in Human Neointimal Hyperplasia.
- 2006
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In: Circulation Research. - 0009-7330. ; 98:4, s. 557-563
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Journal article (peer-reviewed)abstract
- Occlusive vascular disease is a widespread abnormality leading to lethal or debilitating outcomes such as myocardial infarction and stroke. It is part of atherosclerosis and is evoked by clinical procedures including angioplasty and grafting of saphenous vein in bypass surgery. A causative factor is the switch in smooth muscle cells to an invasive and proliferative mode, leading to neointimal hyperplasia. Here we reveal the importance to this process of TRPC1, a homolog of Drosophila transient receptor potential. Using 2 different in vivo models of vascular injury in rodents we show hyperplasic smooth muscle cells have upregulated TRPC1 associated with enhanced calcium entry and cell cycle activity. Neointimal smooth muscle cells after balloon angioplasty of pig coronary artery also express TRPC1. Furthermore, human vein samples obtained during coronary artery bypass graft surgery commonly exhibit an intimal structure containing smooth muscle cells that expressed more TRPC1 than the medial layer cells. Veins were organ cultured to allow growth of neointimal smooth muscle cells over a 2-week period. To explore the functional relevance of TRPC1, we used a specific E3-targeted antibody to TRPC1 and chemical blocker 2-aminoethoxydiphenyl borate. Both agents significantly reduced neointimal growth in human vein, as well as calcium entry and proliferation of smooth muscle cells in culture. The data suggest upregulated TRPC1 is a general feature of smooth muscle cells in occlusive vascular disease and that TRPC1 inhibitors have potential as protective agents against human vascular failure.
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| 3. |
- Eussen, Simone R. B. M., et al.
(author)
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Simultaneous intake of oat bran and atorvastatin reduces their efficacy to lower lipid levels and atherosclerosis in LDLr-/- mice
- 2011
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In: Pharmacological Research. - : Elsevier BV. - 1096-1186 .- 1043-6618. ; 64:1, s. 36-43
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Journal article (peer-reviewed)abstract
- The present study aimed to investigate the effects of separate and simultaneous dietary intake of atorvastatin (ATO) and the soluble fiber oat bran on serum and hepatic lipid levels and the degree of atherosclerosis. Ninety female LDL-receptor-deficient (LDLr-/-) mice were fed a Western-type diet containing either low dose (0.0025%), high dose (0.01%) or no ATO, with or without oat bran (27%) (n = 15 per group) for 16 weeks. Both ATO and oat bran were effective in reducing serum total cholesterol levels (low ATO: -5.48, high ATO: -9.12, oat bran: -3.82 mmol/l, compared to control (no ATO/no oat bran), all p < 0.0001). When oat bran was added to a low dose ATO, the cholesterol-lowering effects of this combination were 50% smaller compared to the low dose ATO diet alone (between-group difference: 2.77 mmol/l, p = 0.002), whereas total cholesterol decreased to a similar extent in the groups fed a high dose ATO, with or without oat bran (between-group difference: 1.10 mmol/l, p = 0.21). Serum LDL- and HDL-cholesterol, triglycerides, hepatic lipid levels and atherosclerotic lesion development showed a similar pattern. In conclusion, the efficacy of oat bran and atorvastatin to lower lipid levels and atherosclerosis is reduced after simultaneous intake. We hypothesize that oat bran inhibits the intestinal absorption of atorvastatin, and consequently its cholesterol-lowering effects. The effects are likely dependent on the type of statin and dietary fiber, and on the relative timing of intake of the statin and the dietary fiber. Future studies should focus on these aspects to provide further insight into the exact mechanism of this food-drug interaction. (C) 2011 Elsevier Ltd. All rights reserved.
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| 4. |
- Sparrow, M P, et al.
(author)
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Isoforms of myosin in smooth muscle
- 1987
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In: Progress in Clinical and Biological Research. - 0361-7742. ; 245, s. 67-79
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Journal article (peer-reviewed)
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| 5. |
- Swärd, Karl, et al.
(author)
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Inhibition of Rho-associated kinase blocks agonist-induced Ca2+ sensitization of myosin phosphorylation and force in guinea-pig ileum
- 2000
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In: Journal of Physiology. - 1469-7793. ; 522, s. 33-49
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Journal article (peer-reviewed)abstract
- Ca2+ sensitization of smooth muscle contraction involves the small GTPase RhoA, inhibition of myosin light chain phosphatase (MLCP) and enhanced myosin regulatory light chain (LC20) phosphorylation. A potential effector of RhoA is Rho-associated kinase (ROK). The role of ROK in Ca2+ sensitization was investigated in guinea-pig ileum. Contraction of permeabilized muscle strips induced by GTPgammaS at pCa 6.5 was inhibited by the kinase inhibitors Y-27632, HA1077 and H-7 with IC50 values that correlated with the known Ki values for inhibition of ROK. GTPgammaS also increased LC20 phosphorylation and this was prevented by HA1077. Contraction and LC20 phosphorylation elicited at pCa 5.75 were, however, unaffected by HA1077. Pre-treatment of intact tissue strips with HA1077 abolished the tonic component of carbachol-induced contraction and the sustained elevation of LC20 phosphorylation, but had no effect on the transient or sustained increase in [Ca2+]i induced by carbachol. LC20 phosphorylation and contraction dynamics suggest that the ROK-mediated increase in LC20 phosphorylation is due to MLCP inhibition, not myosin light chain kinase activation. In the absence of Ca2+, GTPgammaS stimulated 35S incorporation from [35S]ATPgammaS into the myosin targeting subunit of MLCP (MYPT). The enhanced thiophosphorylation was inhibited by HA1077. No thiophosphorylation of LC20 was detected. These results indicate that ROK mediates agonist-induced increases in myosin phosphorylation and force by inhibiting MLCP activity through phosphorylation of MYPT. Under Ca2+-free conditions, ROK does not appear to phosphorylate LC20 in situ, in contrast to its ability to phosphorylate myosin in vitro. In particular, ROK activation is essential for the tonic phase of agonist-induced contraction.
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| 6. |
- Di Russo, Jacopo, et al.
(author)
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Endothelial basement membrane laminin 511 is essential for shear stress response
- 2017
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In: EMBO Journal. - : EMBO. - 0261-4189 .- 1460-2075. ; 36:2, s. 183-201
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Journal article (peer-reviewed)abstract
- Shear detection and mechanotransduction by arterial endothelium requires junctional complexes containing PECAM-1 and VE-cadherin, as well as firm anchorage to the underlying basement membrane. While considerable information is available for junctional complexes in these processes, gained largely from in vitro studies, little is known about the contribution of the endothelial basement membrane. Using resistance artery explants, we show that the integral endothelial basement membrane component, laminin 511 (laminin α5), is central to shear detection and mechanotransduction and its elimination at this site results in ablation of dilation in response to increased shear stress. Loss of endothelial laminin 511 correlates with reduced cortical stiffness of arterial endothelium in vivo, smaller integrin β1-positive/vinculin-positive focal adhesions, and reduced junctional association of actin–myosin II. In vitro assays reveal that β1 integrin-mediated interaction with laminin 511 results in high strengths of adhesion, which promotes p120 catenin association with VE-cadherin, stabilizing it at cell junctions and increasing cell–cell adhesion strength. This highlights the importance of endothelial laminin 511 in shear response in the physiologically relevant context of resistance arteries.
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| 7. |
- Grossi, Mario, et al.
(author)
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Pyk2 inhibition promotes contractile differentiation in arterial smooth muscle
- 2017
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In: Journal of Cellular Physiology. - : Wiley. - 0021-9541. ; 232:11, s. 3088-3102
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Journal article (peer-reviewed)abstract
- Modulation from contractile to synthetic phenotype of vascular smooth muscle cells is a central process in disorders involving compromised integrity of the vascular wall. Phenotype modulation has been shown to include transition from voltage-dependent toward voltage-independent regulation of the intracellular calcium level, and inhibition of non-voltage dependent calcium influx contributes to maintenance of the contractile phenotype. One possible mediator of calcium-dependent signaling is the FAK-family non-receptor protein kinase Pyk2, which is activated by a number of stimuli in a calcium-dependent manner. We used the Pyk2 inhibitor PF-4594755 and Pyk2 siRNA to investigate the role of Pyk2 in phenotype modulation in rat carotid artery smooth muscle cells and in cultured intact arteries. Pyk2 inhibition promoted the expression of smooth muscle markers at the mRNA and protein levels under stimulation by FBS or PDGF-BB and counteracted phenotype shift in cultured intact carotid arteries and balloon injury ex vivo. During long-term (24–96 hr) treatment with PF-4594755, smooth muscle markers increased before cell proliferation was inhibited, correlating with decreased KLF4 expression and differing from effects of MEK inhibition. The Pyk2 inhibitor reduced Orai1 and preserved SERCA2a expression in carotid artery segments in organ culture, and eliminated the inhibitory effect of PDGF stimulation on L-type calcium channel and large-conductance calcium-activated potassium channel expression in carotid cells. Basal intracellular calcium level, calcium wave activity, and store-operated calcium influx were reduced after Pyk2 inhibition of growth-stimulated cells. Pyk2 inhibition may provide an interesting approach for preserving vascular smooth muscle differentiation under pathophysiological conditions.
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| 8. |
- Himpens, B, et al.
(author)
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Free cytosolic calcium during spontaneous contractions in smooth muscle of the guinea-pig mesotubarium
- 1990
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In: Pflügers Archiv. - 0031-6768. ; 417:4, s. 404-409
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Journal article (peer-reviewed)abstract
- The free intracellular calcium ion concentration ([Ca2+]i) was measured simultaneously with isometric force in strips of guinea-pig mesotubarium using the Fura-2 technique. During the relaxed period (5-15 min) between spontaneous contractions [Ca2+]i continues to decrease after full mechanical relaxation to reach a minimal level of 86 +/- 8 nM (n = 9) just before the start of the next contraction. During the spontaneous contractions (5-15 min) [Ca2+]i reached a maximum of 211 +/- 19 nM and then oscillated between 155 +/- 16 nM and 194 +/- 9 nM. Increased extracellular Ca2+ concentration to 10 mM from the standard concentration of 1.5 mM caused a decreased frequency of spontaneous contractions and an increase in [Ca2+]i both in the relaxed and contracted states. In 10 mM extracellular Ca2+, addition of AlF4-, as 1 mM NaF + 10 microM AlCl3, caused a sustained increase in [Ca2+]i and maintained force. Addition of verapamil (10 microM) in this situation decreased [Ca2+]i to the resting level. The results suggest that the cyclic appearance of trains of action potentials is related to variation in [Ca2+]i, possibly via inactivation of Ca2(+)-dependent K+ channels.
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| 9. |
- Lydrup, M L, et al.
(author)
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Effect of glibenclamide on membrane response to metabolic inhibition in smooth muscle of rat portal vein
- 1994
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In: Journal of Vascular Research. - 1423-0135. ; 31:2, s. 82-91
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Journal article (peer-reviewed)abstract
- Vascular smooth muscle tone is dependent on oxidative metabolism, a phenomenon of potential importance for the metabolic regulation of blood flow to tissues. The response of the rat portal vein to inhibition of cell respiration by cyanide (0.1-1 mM) is a reduction of its spontaneous myogenic activity. The trains of action potentials triggering phasic contractions are reduced in duration, while the frequency of trains is often somewhat increased as the resting membrane potential in the intervals between spike trains is less negative by 6.5 mV. Glibenclamide (10(-7) M) did not affect the resting membrane potential or spontaneous mechanical activity of oxygenated portal veins, but partly restored the depressed myogenic activity in the presence of cyanide (0.5 mM). The spike trains were longer, while the membrane was depolarized by 3 mV compared with the effects of cyanide alone. Inhibition of both oxidative and glycolytic metabolism by 2 mM NaCN in a medium where glucose was replaced by beta-hydroxybutyrate caused a hyperpolarization which was abolished by 10(-7) M glibenclamide. The relaxing effect of the K+ channel opener cromakalim (5 x 10(-9) to 6.25 x 10(-7) M) was partly antagonized by glibenclamide. Basal cytosolic [Ca2+] was increased by cyanide, while the Ca2+ transients associated with phasic contractions were reduced in duration. This latter effect was partially reversed by glibenclamide. The effect of cyanide on high-K+ contractures, which are associated with sustained membrane depolarization and not dependent on repetitive spike activity, was not influenced by 10(-7) M glibenclamide. The effects of inhibited cell respiration on spontaneous electrical activity seem to reflect a depolarizing drive caused by inhibited active ion exchange mechanisms, modified by a repolarizing drive, possibly from ATP-regulated K+ channels, causing reduced duration of the spike trains. While glibenclamide affects spontaneous activity at all levels of oxidative blockade, glibenclamide-sensitive hyperpolarization is seen only when both oxidative and glycolytic metabolism is inhibited.
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| 10. |
- Lydrup, M L, et al.
(author)
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Effects of extracellular K+ and Ca2+ on membrane potential, contraction and 86Rb+ efflux in guinea-pig mesotubarium
- 1990
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In: Pflügers Archiv. - 0031-6768. ; 415:6, s. 664-670
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Journal article (peer-reviewed)abstract
- The effects of varying extracellular concentrations of K+ and Ca2+ [K+]o and [Ca2+]o on force development and membrane potential were investigated in the guinea-pig mesotubarium. At [K+]o up to 40 mM, spontaneous action potentials were present, while higher [K+]o gave sustained contractures at a stable membrane potential (-24 to -12 mV for [K+]o from 60 to 120 mM). Tension decreased successively with increasing [K+]o from 30 to 120 mM. The relaxing potency of the dihydropyridine Ca2+ antagonist, felodipine, increased as the membrane was depolarized with increasing [K+]o and action potentials ceased. These results are compatible with the existence of Ca2+ channels showing voltage-dependent affinity with dihydrophyridines. Increasing [Ca2+]o from 2.5 to 10 mM caused membrane hyperpolarization by about 11 mV and was accompanied by a lower frequency of spontaneous contractions and a longer duration of the relaxation between contractions. 86Rb+ efflux measurements in 60 mM K+ in the absence and presence of felodipine revealed a Ca2(+)-dependent component of the voltage-activated efflux. In normal solution (5.9 mM K+), efflux in the presence of felodipine was similar to the minimal value during normal spontaneous activity. The results indicate regulation of the permeability of K+ channels by the intracellular Ca2+ concentration ([Ca2+]i) and suggest participation of such channels in the generation of the regularly occurring bursts of action potentials characteristic of spontaneous activity in the mesotubarium.
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