scholarly journals Thrombin-stimulated events in cultured vascular smooth-muscle cells

1991 ◽  
Vol 274 (3) ◽  
pp. 799-805 ◽  
Author(s):  
B C Berk ◽  
M B Taubman ◽  
K K Griendling ◽  
E J Cragoe ◽  
J W Fenton ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Vascular Tissue ◽  
Muscle Cells ◽  
Maximal Increase ◽  
Cellular Signal ◽  
High Concentrations ◽  
Dimethyl Amiloride

Thrombin is present in high concentrations at sites of clots and may have important post-clotting effects on adjacent vascular tissue. This may be particularly important for vascular smooth-muscle cells (VSMC), whose growth and contractility are altered following atherosclerotic-associated thromboses. To study the cellular signal events by which thrombin exerts its actions, the effects of purified human alpha-thrombin were examined in cultured rat aortic VSMC. alpha-Thrombin stimulated a biphasic change in intracellular pH (pHi), causing an early rapid acidification, followed by a sustained alkalinization. The increase in pHi was dependent on extracellular Na+ and inhibited by 5′-(NN-dimethyl)amiloride, consistent with mediation by Na+/H+ exchange. alpha-Thrombin rapidly increased free intracellular [Ca2+] ([Ca2+]i). The increase in [Ca2+]i was secondary to activation of phospholipase C, as demonstrated by increases in InsP3 (226%) and InsP2 (387%) and decreases in polyphosphoinositides at 15 s. Expression of the mRNA for the proto-oncogene c-fos was induced by alpha-thrombin. Stimulation of c-fos mRNA was not dependent on alterations in pHi, but required a rise in [Ca2+]i. Despite many growth-related signals shared by alpha-thrombin with platelet-derived growth factor, alpha-thrombin failed to stimulate [3H]thymidine incorporation or cell division, although there was a maximal increase of 52% in protein synthesis. The data suggest that there are cellular signal events not activated by alpha-thrombin which are required for proliferation of these aortic VSMC.

Spatial dynamics of intracellular calcium in agonist-stimulated vascular smooth muscle cells

1990 ◽  
Vol 259 (4) ◽  
pp. C675-C686 ◽  
Author(s):  
C. B. Neylon ◽  
J. Hoyland ◽  
W. T. Mason ◽  
R. F. Irvine
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Vascular Tissue ◽  
Spatial Dynamics ◽  
Muscle Cells ◽  
Smooth Muscle Contraction ◽  
The Novel ◽  
Internal Mammary

Vasoconstrictor agonists stimulate smooth muscle contraction by inducing a rise in intracellular free Ca2+. Digital-imaging microscopy of fura-2 fluorescence from single vascular smooth muscle cells cultured from the human internal mammary artery has allowed us to record the subcellular alterations in Ca2+ that occur immediately after stimulation by receptor agonists. The thrombin-induced rise in cytoplasmic free Ca2+ begins in a discrete region typically located close to the end of the cell. Subsequently, this region of elevated Ca2+ expands until Ca2+ is elevated throughout the cell cytoplasm. The rate of spreading in the region of elevated Ca2+ in a linear direction averaged 10.1 microns/s, enabling it to traverse the length of most cells within approximately 5 s, and involved rises in Ca2+ of between 200 and 500 nM. In some cells, the Ca2+ rise began at both ends and collided midway. Similar dynamic changes in the spatial distribution of Ca2+ were recorded in cells stimulated by acetylcholine. The novel observation that vasoconstrictor agonists induce an elevation of Ca2+ in a localized region which subsequently expands throughout the cytoplasm of single smooth muscle cells may provide new insight into the nature of Ca2+ signaling in vascular tissue.

scholarly journals Receptor-dependent prorenin activation and induction of PAI-1 expression in vascular smooth muscle cells

2008 ◽  
Vol 295 (4) ◽  
pp. E810-E819 ◽  
Author(s):  
Jiandong Zhang ◽  
Nancy A. Noble ◽  
Wayne A. Border ◽  
Rick T. Owens ◽  
Yufeng Huang
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Vascular Tissue ◽  
Microvascular Complications ◽  
Muscle Cells ◽  
Diabetic Patients ◽  
Ang Ii ◽  
Pai 1

Although elevated plasma prorenin levels are commonly found in diabetic patients and correlate with microvascular complications, the pathological role of these increases, if any, remains unclear. Prorenin/renin binding to the prorenin/renin receptor [(p)RR] enhances the efficiency of angiotensinogen cleavage by renin and unmasks prorenin catalytic activity. We asked whether plasma prorenin could be activated in local vascular tissue through receptor binding. Immunohistochemical staining showing localization of the (p)RR in the aorta to vascular smooth muscle cells (VSMCs). After cultured rat VSMCs were incubated with 10−7 M inactive prorenin, cultured supernatant acquired the ability to generate ANG I from angiotensinogen, indicating that prorenin had been activated. Activated prorenin facilitated angiotensin generation in cultured VSMCs when exogenous angiotensinogen was added. Small interfering RNA (siRNA) against the (p)RR blocked this activation and subsequent angiotensin generation. Prorenin alone induced dose- and time-dependent increases in mRNA and protein for the profibrotic molecule plasminogen activator inhibitor (PAI)-1, effects that were blocked by siRNA, but not by the ANG II receptor antagonist saralasin. When inactive prorenin and angiotensinogen were incubated with cells, PAI-1 mRNA increased a striking 54-fold, 8-fold higher than the increase seen with prorenin alone. PAI-1 protein increased 2.75-fold. These effects were blocked by treatment with siRNA + saralasin. We conclude that prorenin at high concentration binds the (p)RR on VSMCs and is activated. This activation leads to increased expression of PAI-1 via ANG II-independent and -dependent mechanisms. These data provide a mechanism by which elevated prorenin levels in diabetes may contribute to the progression of fibrotic disease.

scholarly journals Novel transcripts of Nox1 are regulated by alternative promoters and expressed under phenotypic modulation of vascular smooth muscle cells

2006 ◽  
Vol 398 (2) ◽  
pp. 303-310 ◽  
Author(s):  
Noriaki Arakawa ◽  
Masato Katsuyama ◽  
Kuniharu Matsuno ◽  
Norifumi Urao ◽  
Yoshiaki Tabuchi ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Nadph Oxidase ◽  
Smooth Muscle Cells ◽  
Cell Line ◽  
Vascular Smooth Muscle Cells ◽  
Vascular Tissue ◽  
Muscle Cells ◽  
Alternative Promoters ◽  
Phenotypic Modulation

NADPH oxidase is implicated in the pathogenesis of various cardiovascular disorders. In vascular smooth muscle cells (VSMC), expression of NOX1 (NADPH oxidase 1), a catalytic subunit of NADPH oxidase, is low and is induced upon stimulation by vasoactive factors, while it is abundantly expressed in colon epithelial cells. To clarify the regulatory mechanisms underlying such cell-specific expression, the upstream regions directing transcription of the NOX1 gene were explored. In P53LMACO1 cells, a cell line originated from mouse VSMCs, two novel Nox1 mRNA species, the c- and f-type, were isolated. These transcripts contained 5′-untranslated regions that differed from the colon type mRNA (a-type) and encoded an additional N-terminal peptide of 28 amino acids. When these transcripts were fused to the c-myc tag and expressed in human embryonic kidney 293 cells, a fraction of translated proteins demonstrated the size containing the additional peptide. Proteins encoded by the c- and f-type mRNAs exhibited superoxide-producing activities equivalent to the activity of the a-type form. The a-type mRNA was expressed in the colon and in the intact aorta, whereas the c-type mRNA was detected in the primary cultured VSMCs migrated from aortic explants, in vascular tissue of a wire-injury model and in the thoracic aorta of mice infused with angiotensin II. The promoter region of the c-type mRNA exhibited transcriptional activity in P53LMACO1 cells, but not in MCE301 cells, a mouse colon epithelial cell line. These results suggest that expression of the Nox1 gene is regulated by alternative promoters and that the novel c-type transcript is induced under phenotypic modulation of VSMCs.

Fluid Shear Stress-Induced Alignment of Cultured Vascular Smooth Muscle Cells

2001 ◽  
Vol 124 (1) ◽  
pp. 37-43 ◽  
Author(s):  
Ann A. Lee ◽  
Dionne A. Graham ◽  
Sheila Dela Cruz ◽  
Anthony Ratcliffe ◽  
William J. Karlon
Keyword(s):  
Shear Stress ◽  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Fluid Shear Stress ◽  
Vascular Tissue ◽  
Continuous Process ◽  
Muscle Cells ◽  
Clear Understanding

The study objectives were to quantify the time- and magnitude-dependence of flow-induced alignment in vascular smooth muscle cells (SMC) and to identify pathways related to the orientation process. Using an intensity gradient method, we demonstrated that SMC aligned in the direction perpendicular to applied shear stress, which contrasts with parallel alignment of endothelial cells under flow. SMC alignment varied with the magnitude of and exposure time to shear stress and is a continuous process that is dependent on calcium and cycloskeleton based mechanisms. A clear understanding and control of flow-induced SMC alignment will have implications for vascular tissue engineering.

Agonist-induced [Ca2+]i waves and Ca(2+)-induced Ca2+ release in mammalian vascular smooth muscle cells

1992 ◽  
Vol 263 (2) ◽  
pp. H576-H586 ◽  
Author(s):  
L. A. Blatter ◽  
W. G. Wier
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Initial Response ◽  
Cellular Mechanisms ◽  
A7r5 Cells ◽  
Cellular Signal ◽  
Entire Cell

Focal application of vasopressin to cultured vascular smooth muscle cells (A7r5 cells) elicits first a localized increase of intracellular Ca2+ concentration ([Ca2+]i) and then a wave of elevated [Ca2+]i that propagates at constant velocity throughout the cell. The cellular mechanisms of such complex spatiotemporal patterns of [Ca2+]i are of interest because they are involved fundamentally in cellular signal transduction in many types of cells. Vasopressin evoked a [Ca2+]i transient even in the absence of extracellular Ca2+, and intracellular perfusion with heparin completely blocked the response to vasopressin stimulation. Therefore the initial response to vasopressin reflects release of Ca2+ from an intracellular myo-inositol-1,4,5-trisphosphate (IP3)-sensitive Ca2+ store. We tested four hypotheses on how a localized increase in [Ca2+]i propagates as a [Ca2+]i wave throughout the entire cell: the hypotheses distinguished 1) whether IP3 or Ca2+ is the primary intracellular messenger that diffuses, and 2) whether positive feedback on the release of intracellular Ca2+ (Ca2+i) is involved (further release of Ca2+ through activation of phospholipase C by Ca2+ and increased production of IP3 or by Ca(2+)-induced Ca2+ release). The results of various experimental interventions, which included probing Ca2+i stores (heparin, caffeine, and ryanodine), were compared with predictions from mathematical models for intracellular diffusion, release, and uptake of Ca2+. We conclude that in A7r5 smooth muscle cells, which have been stimulated focally with vasopressin, Ca2+ is released initially by IP3. The localized increase in [Ca2+]i then propagates throughout the cell as a [Ca2+]i wave. Ca2+ activates its own release, through Ca(2+)-induced release of Ca2+, by diffusing to distant Ca(2+)-release sites.

Lysophosphatidic acids induce proliferation of cultured vascular smooth muscle cells from rat aorta

1994 ◽  
Vol 267 (1) ◽  
pp. C204-C210 ◽  
Author(s):  
A. Tokumura ◽  
M. Iimori ◽  
Y. Nishioka ◽  
M. Kitahara ◽  
M. Sakashita ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Growth Factor ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Rat Aorta ◽  
Fatty Acyl ◽  
Dependent Manner ◽  
High Concentrations

Lysophosphatidic acids (LPA) with a C18 fatty acyl group accelerated thymidine incorporation into cultured rat aortic vascular smooth muscle cells and stimulated their cell division. LPA acted synergistically with epidermal growth factor and fibroblast growth factor but additively with platelet-derived growth factor. The stimulatory actions of LPA were suggested to be rather specific from the following findings: 1) their stimulation of DNA synthesis increased with an increase in their acyl moiety; 2) lysophosphatidylcholine, a neutral lysophospholipid, had no mitogenic action but was cytotoxic at high concentrations; and 3) LPA induced a rapid external Ca(2+)-independent increase in intracellular Ca2+ concentration ([Ca2+]i) in single fura 2-loaded cells that resembled the receptor-mediated increases in [Ca2+]i triggered by different agonists, whereas lysophosphatidylcholine provoked a slow sustained increase in [Ca2+]i in an external Ca(2+)-dependent manner. These results are discussed in relation to the possible pathophysiological role of LPA.

Histamine-induced calcium transients in vascular smooth muscle cells: effects of verapamil and diltiazem

1989 ◽  
Vol 257 (2) ◽  
pp. H563-H570
Author(s):  
T. Matsumoto ◽  
H. Kanaide ◽  
J. Nishimura ◽  
T. Kuga ◽  
S. Kobayashi ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Rat Aorta ◽  
Calcium Transients ◽  
Inhibitory Effects ◽  
H1 Receptor ◽  
High Concentrations

We investigated the effects of verapamil and diltiazem on histamine-induced Ca2+ transients in vascular smooth muscle. 1) With the use of quin2 microfluorometry, cytosolic Ca2+ concentrations were directly measured in cultured vascular smooth muscle cells of the rat aorta. In the presence of extracellular Ca2+, histamine induced an elevation of cytosolic Ca2+ concentrations of a peak and plateau type. The peak component is due to a release of Ca2+ from cellular store sites, and the plateau component depends on extracellular Ca2+. Verapamil and diltiazem inhibited the plateau component, and the 50% inhibitive concentration (IC50) of verapamil and diltiazem for 10 microM histamine was 0.09 and 0.18 microM, respectively. Only at high concentrations did verapamil (IC50 = 8.7 microM) and diltiazem (IC50 = 95.7 microM) inhibit the Ca2+ release from the cellular store sites, as induced by 10 microM histamine. 2) Histamine, verapamil, and diltiazem competed with [3H]mepyramine for binding to the porcine aortic membranes, the order of potency being verapamil (Ki = 7.1 microM) greater than histamine (Ki = 18 microM) greater than diltiazem (Ki = 114 microM). From these results, we conclude that verapamil and diltiazem strongly inhibit the histamine-mediated, extracellular Ca2+-dependent intracellular [Ca2+] increase. In addition, verapamil and diltiazem seem to inhibit the release of Ca2+ from intracellular store sites, only at high concentrations, and probably by competing with histamine for binding to the H1-receptor. The inhibitory effects of Ca2+ antagonists on the histamine-induced contraction or spasm of vascular smooth muscle may well relate to these mechanisms.

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