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M Kumada

Publications and source records attributed to M Kumada.

At least 55 records · Page 3Linked to original sources
PubMed1994

Synergistic stimulation of parathyroid hormone-related peptide gene expression by mechanical stretch and angiotensin II in rat aortic smooth muscle cells.

Repetitive cyclic stretch (60 cycles/min) of rat aortic smooth muscle cells dramatically enhances the effect of angiotensin II (AII) on mRNA expression of the vasorelaxant, parathyroid hormone-related peptide (PTHrP). Thus, combined stimulation of rat aortic smooth muscle cells by cyclic stretch and low concentrations of AII, but not either alone, induces a synergistic, marked increase in the PTHrP mRNA level, in a manner dependent on the strength of stretch. This response is accompanied by a synergistic increase in secretion of PTHrP from smooth muscle cells. Removal of extracellular Ca2+ or addition of Ca2+ channel blockers, including Gd3+ and nitrendipine, does not considerably reduce the combined effects of stretch and AII, indicating that this response is not dependent on stretch-induced Ca2+ influx across the plasma membrane. The combined effect of stretch and AII on PTHrP mRNA expression is strongly attenuated by the protein kinase C (PKC) inhibitor staurosporine or by down-regulation of PKC, suggesting that PKC plays an important role in the synergistic response. However, stretch neither elicits activation of phospholipase C or PKC by itself, nor does it enhance AII-induced activation of these enzymes. These results indicate that in vascular smooth muscle cells mechanical stretch acts together with the vasoconstrictor AII to regulate the expression of the vasodilator PTHrP and suggest the role of PTHrP as a local modulator of myogenic tone.

Alkaloids↗
PubMed1994

A fumagillin derivative angiogenesis inhibitor, AGM-1470, inhibits activation of cyclin-dependent kinases and phosphorylation of retinoblastoma gene product but not protein tyrosyl phosphorylation or protooncogene expression in vascular endothelial cells.

Fumagillin analogue AGM-1470 potently inhibits angiogenesis with a minimal toxicity in vivo and is expected to be of therapeutic use as a powerful antitumor agent (Ingber et al., Nature, 348:555-557, 1990). In the present study, we have investigated the effects and the mechanism of action of AGM-1470 on cultured human umbilical vein endothelial cells. AGM-1470 acts directly on endothelial cells to inhibit growth factor-induced DNA synthesis, with half maximal and maximal effects obtained at approximately 2 x 10(-10) and 5 x 10(-9) M, respectively. AGM-1470 does not inhibit early G1 mitogenic events, such as cellular protein tyrosyl phosphorylation or the expression of immediate early genes c-fos and c-myc, but potently inhibits phosphorylation of RB protein, a tumor suppressor retinoblastoma gene product. The later addition of AGM-1470 up to 3 h after the growth factor stimulation still exerts full inhibitory effects on both DNA synthesis and RB phosphorylation, suggesting that the major site of action of AGM-1470 is located relatively late in the G1 phase. AGM-1470 inhibits growth factor-induced activation of candidate RB kinases cdc2 and cdk2 but fails to inhibit them directly in vitro. AGM-1470 completely abolishes the growth factor-induced mRNA expression of cdc2 and cyclin A and partially inhibits that of cyclin E but has little effect on the mRNA level of cdk2, cdk4, or cyclin D1. These results indicate that angioinhibitory action of AGM-1470 involves suppression of mRNA expression of specific members of cdks and cyclins and of activation of both cdc2 and cdk2 kinases in endothelial cells.

CDC2 Protein Kinase↗
PubMed1994

Molecular cloning of a novel putative G protein-coupled receptor from rat aortic smooth muscle. Downregulation of the mRNA level by the cyclic AMP messenger pathway.

A cDNA clone encoding a novel putative G protein-coupled receptor was isolated from rat aortic vascular smooth muscle cell cDNA library by the polymerase chain reaction (PCR) using degenerate oligonucleotide primers and subsequent hybridization screening with a cloned PCR product. Sequence analysis of this clone (AGR9) shows that it encodes a 483 amino acid protein with seven streches of hydrophobic amino acids, which are presumed to represent transmembrane domains. In addition, AGR9 protein exhibits several structural features characteristic of the G protein-coupled receptor family, which include the existence of potential N-linked glycosylation sites in the amino-terminal region, phosphorylation sites by serine/threonine kinases in the intracellular regions, and a number of well-conserved residues among most of the G protein-coupled receptors. Northern blot analysis indicates abundant expression of a major 3.9 kb AGR9 mRNA in brain, lung, heart, stomach, intestine, cultured rat aortic smooth muscle cells and cardiac myocytes. Treatment of rat aortic smooth muscle cells with the adenylyl cyclase activator forskolin causes a marked and transient decrease in the steady-state level of AGR9 mRNA. Dibutyryl cyclic AMP and the beta-adrenergic agonist isoproterenol mimick the effect of forskolin. The ligand for AGR9 receptor has yet to be identified, however, these results suggest the existence of a novel G protein-coupled receptor expressed in the cardiovascular, central nervous and digestive systems.

Animals↗
PubMed1994

E2F1, B-myb and selective members of cyclin/cdk subunits are targets for protein kinase C-mediated bimodal growth regulation in vascular endothelial cells.

In human umbilical vein endothelial cells, the protein kinase C (PKC) stimulation during the early G1 phase leads to potentiations in growth factor-stimulated DNA synthesis, the activation of cdc2 and cdk2 cyclin-dependent kinases, and the mRNA expression of cdc2, cyclins A, D1 and E, but not cdk2 or cdk4. Conversely, the PKC stimulation in the late G1 phase completely inhibits DNA synthesis, the activation of cyclin-dependent kinases, and the mRNA expression of the same set of molecules except cyclin D1. Further, we found that the PKC stimulation bimodally regulates the message levels of E2F1 and B-myb, which are transcription factors implicated in the control of the mammalian cell cycle progression. These results indicate that the PKC signal transduction pathway, depending on the timing of activation in the G1 phase, either positively or negatively regulates the message level of growth-regulating genes that are crucial for the G1 to S phase progression.

CDC2-CDC28 Kinases↗
PubMed1994

Suppression of neointimal smooth muscle cell accumulation in vivo by antisense cdc2 and cdk2 oligonucleotides in rat carotid artery.

Deendothelializing balloon injury of rat carotid artery results in progressive intimal smooth muscle cell accumulation and luminal stenosis over 14 days after injury. We have found transient rises (approximately 3-fold maximal increases over the uninjured control value) of the kinase activities of both cdc2 and cdk2, key molecules for cell cycle progression, in the injured carotid artery along with the development of intimal proliferation. The topical application of the antisense, but not the sense, cdc2 and cdk2 phosphorothioate oligodeoxynucleotides dissolved in F127 pluronic gel around the freshly injured artery resulted in reductions of the intimal smooth muscle cell accumulation by 47% and 55% respectively, as estimated by an intimal to medial cross-sectional area ratio, with concomitant decreases in cdc2 and cdk2 kinase activities. These results indicate that both cdc2 and cdk2 kinases are involved in intimal smooth muscle cell accumulation after balloon angioplasty and suggest a potential usefulness of the antisense cdc2 and cdk2 oligonucleotide therapy for arterial stenosis.

Angioplasty, Balloon↗
PubMed1994

Tongue displacement: visualization with rapid tagged magnetization-prepared MR imaging.

The authors describe a technique to depict tongue muscle displacement with tagged magnetization-prepared gradient-recalled echo magnetic resonance imaging. Three Japanese volunteers (three men, aged 31-36 years) were studied while they articulated specific sounds. Each image was obtained in less than 2 seconds without need for gating. Displacement and alteration of the tags within the tongue muscle during phonation were clearly visible. The technique may prove useful in the study of phoniatric pathophysiology.

Adult↗
PubMed1994

High calcium diet prevents baroreflex impairment in salt-loaded spontaneously hypertensive rats.

To investigate the role of the sympathetic control mechanism in the antihypertensive effect of dietary calcium supplementation, we examined whether a high calcium diet affected mean arterial pressure, renal sympathetic nerve activity, heart rate, and overall and central properties of the arterial baroreceptor reflex in salt-loaded young spontaneously hypertensive rats (SHR). Six-week-old SHR were fed either a normal (0.66%) or high (8.00%) salt diet with either a normal (1.17%) or high (4.07%) calcium content for 4 weeks. The arterial baroreceptor reflex was elicited with rats under halothane anesthesia by altering mean arterial pressure with nitroprusside or phenylephrine. The overall property of the arterial baroreceptor reflex was assessed by the median mean arterial pressure (MAP50) and maximal gain (Gmax) of the relation between mean arterial pressure and renal sympathetic nerve activity and between mean arterial pressure and heart rate. The central property of the arterial baroreceptor reflex was assessed by reflex inhibition of renal sympathetic nerve activity and heart rate elicited by electrical stimulation of the aortic depressor nerve. Compared with the control group fed a normal salt/normal calcium diet, the high salt/normal calcium group had significantly higher mean arterial pressure and renal sympathetic nerve activity but not heart rate. Moreover, the arterial baroreceptor reflex was impaired in the latter group, as evidenced by an increase in MAP50 and decrease in Gmax of the two relations and an attenuation of reflex inhibition of renal sympathetic nerve activity by aortic depressor nerve stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Protein kinase C-mediated bidirectional regulation of DNA synthesis, RB protein phosphorylation, and cyclin-dependent kinases in human vascular endothelial cells.

In human umbilical vein endothelial cells, activators of protein kinase C (PKC) exert cell cycle-dependent, bidirectional growth regulatory effects. Thus, phorbol 12,13-dibutyrate or 1,2-dioctanoylglycerol potentiates growth factor-induced DNA synthesis up to 3-fold when they act during the early G1 phase, whereas they completely inhibit the initiation of DNA synthesis when they act in the late G1 phase. In addition, the PKC activators induce a rapid inhibition of the ongoing DNA synthesis when they are applied after entry into the S phase. The effects of the PKC activators in both stimulatory and inhibitory directions are abolished in PKC-downregulated cells. The cell cycle-dependent, PKC-mediated bidirectional growth regulation is closely associated with either potentiation or inhibition of RB protein phosphorylation and the histone H1 kinase activity of cyclin-dependent kinases (cdks) cdc2 and cdk2, which normally accumulate along the G1 to the S phase transition. Northern and Western blot analyses of cdc2 and cdk2 have revealed that PKC regulates the cdks at multiple steps in distinct ways. Thus, for cdc2, the levels of mRNA and protein as well as the extent of post-translational modification are all subject to the PKC-mediated regulation. In contrast, the level of mRNA or protein of cdk2 is not affected by PKC stimulation at any phase of the cell cycle. These results demonstrate the existence of a complex array of PKC-cdk signaling pathways, which mediate temporally organized bimodal growth regulation in endothelial cells.

Amino Acid Sequence↗
PubMed1993

Molecular cloning of a novel putative G protein-coupled receptor expressed in the cardiovascular system.

A novel cDNA clone encoding a putative G protein-coupled receptor has been isolated from a rat aortic smooth muscle cDNA library. Sequence analysis of this clone reveals the structural features characteristic of this superfamily, with a high degree of sequence similarity to the Edg-1 receptor previously reported to be expressed in vascular endothelial cells. This novel receptor mRNA is expressed predominantly in the lung and heart. Although the ligand for this receptor has yet to be identified, the results suggest the existence of a new receptor subfamily in the cardiovascular system thus far unrecognized.

Amino Acid Sequence↗
PubMed1993

Ca(2+)-dependent stimulation of retinoblastoma gene product phosphorylation and p34cdc2 kinase activation in serum-stimulated human fibroblasts.

In serum-deprived human fibroblasts IMR-90 and WI-38 cells, the addition of fetal calf serum or basic fibroblast growth factor stimulates DNA synthesis in an extracellular Ca(2+)-dependent manner; the effect of serum on [3H]thymidine incorporation into DNA is 4-16-fold greater at 2.0 mM CaCl2 as compared with that at 0.03 mM CaCl2. By contrast, in SV40 virus-transformed WI-38 (SV-WI-38) cells DNA synthesis is essentially independent of the extracellular calcium concentration ([Ca]out) and serum growth factors. To explore the role of Ca2+ in mitogenic signal transduction through G1 to S phase cell cycle progression, we studied and compared the effect of [Ca]out on phosphorylation of RB protein, the product of a tumor suppressor retinoblastoma gene. In IMR-90 and WI-38 cells, serum or basic fibroblast growth factor induces an increase in the amount of hyperphosphorylated forms of RB protein in a manner strictly dependent on [Ca]out. In sharp contrast, in SV-WI-38 cells, the extent of RB phosphorylation is little affected by [Ca]out or the presence or absence of serum growth factors. In addition, potent calmodulin antagonists W-7 and calmidazolium, but not an inactive analogue W-12 or W-5, strongly inhibit serum-induced increases in DNA synthesis and RB phosphorylation in IMR-90 and WI-38 cells, whereas in SV-WI-38 cells, the inhibitory effect is much more limited. Under the same treatment conditions, we measured histone H1 kinase activity associated with anti-p34cdc2 immunoprecipitate and found that the serum-induced increase in p34cdc2 kinase activity is strongly dependent on [Ca]out and is potently inhibited by the active calmodulin antagonists in IMR-90 and WI-38 cells, but not in SV-WI-38 cells. In IMR-90 cells that have been incubated with serum in 0.03 mM [Ca]out for 24 h, restoration of [Ca]out to 2.0 mM results in initiation of DNA synthesis after 13 h and concomitant increases in RB phosphorylation and p34cdc2 histone H1 kinase activity. These results suggest that in human fibroblasts, Ca2+/calmodulin regulates the signaling cascade leading to cdc2 kinase activation, RB protein phosphorylation, and DNA synthesis and that this Ca(2+)-dependent regulation is abrogated in SV40-transformed cells.

Blood↗
PubMed1993

Action of endothelin-1 on vasomotor neurons in rat rostral ventrolateral medulla.

Intracisternal administration of endothelin-1 (ET-1) elicits sympathetically mediated cardiovascular responses by acting on the ventral surface of the medulla oblongata (VSM) subjacent to the rostral ventrolateral medulla (RVLM). We examined, in urethane-anesthetized rats, whether intracisternal ET-1 affected activity of vasomotor neurons (VMNs) in the RVLM, by acting either directly on the VMNs or indirectly via the VSM. VMNs were identified electrophysiologically. Intracisternal administration of ET-1 altered activity of all the 13 VMNs tested. At a dose of 0.1 pmol, ET-1 invariably caused transient excitation in six VMNs examined, whereas at a dose of 1 pmol in separate experiments all the seven VMNs tested were inhibited with (n = 6) or without (n = 1) preceding excitation. Similarly, topical application of ET-1 (0.1-1 pmol) to the VSM caused inhibition with (n = 3) or without (n = 2) preceding excitation in all the five VMNs tested. Direct iontophoretic application of ET-1 to the VMNs caused excitation in four of seven VMNs examined but did not affect the other three neurons. These results support the view that intracisternally administered ET-1 alters activity of VMNs in the RVLM, by acting directly on neurons themselves and indirectly via the VSM.

Animals↗
PubMed1993

Botulinum toxin treatment for spasmodic dysphonia.

Effective treatment of adductor type spasmodic dysphonia with botulinum toxin injection is presented. Patients showed objective and/or subjective improvement in phonation. The beneficial effect lasted for approximately 3 months. An immediate complication is temporary hoarseness or aphonia, mainly due to diffusion of BT into the adjacent muscles. This is avoided by limiting the injection to one vocal fold only and by keeping the dose at less than 5 units. Insertion technique of the needles, such as percutaneous and laryngoscopically controlled techniques, are discussed.

Adult↗
PubMed1992

Activators of protein kinase C induce p34cdc2 histone H1 kinase stimulation in Swiss 3T3 fibroblasts.

Phorbol-12,13-dibutyrate and 1,2-dioctanoylglycerol, activators of protein kinase C (PKC) that stimulate DNA synthesis in serum-deprived Swiss 3T3 fibroblasts, induce histone H1 kinase activity associated with anti-cdc2 immunoprecipitates after a lag period of 15h, a time point close to G1/S boundary of the cell cycle in these cells. Downregulation of PKC does not affect the basal cdc2 kinase activity, but potently inhibits both phorbol dibutyrate- and dioctanoylglycerol-induced cdc2 kinase activation. Phorbol dibutyrate induces a dramatic increase in the p34cdc2 protein level as well as the appearance of p35-p36 forms of cdc2 on Western blot. In PKC-downregulated cells, the p34 form of cdc2 remains elevated but p35-p36 forms do not appear upon phorbol dibutyrate stimulation. These results demonstrate that PKC activation leads to cdc2 kinase activation in mitogenically responsive Swiss 3T3 cells, and strongly suggest that both expression of p34cdc2 protein and its posttranslational modification(s) are involved in this process. Western blot analysis of PKC isozymes suggests that either PKC alpha, PKC delta or PKC epsilon may be involved in p34cdc2 kinase activation and mitogenesis.

3T3 Cells↗
PubMed1992

Ca2+/calmodulin is involved in growth factor-induced retinoblastoma gene product phosphorylation in human vascular endothelial cells.

In human vascular endothelial cells, both growth factor-induced DNA synthesis and retinoblastoma gene product (RB) phosphorylation are absolutely dependent on extracellular Ca2+, and are potently inhibited by an active calmodulin antagonist, W-7, but not an inactive analogue, W-12. A reduction in the extracellular Ca2+ or an addition of W-7 as late as 8 h after growth factor stimulation still inhibits both RB phosphorylation and DNA synthesis to the full extent. However, once RB phosphorylation occurs 12-16 h after addition of the growth factors, it is not reversed by subsequent Ca2+ reduction or W-7. These results suggest the existence of a Ca2+/calmodulin-dependent process relatively late in the mitogenic signalling cascade, at a step proximal to RB phosphorylation reaction itself.

Blotting, Western↗
PubMed1991

Mechanisms of bombesin-induced arachidonate mobilization in Swiss 3T3 fibroblasts.

A peptide mitogen bombesin, which activates the phospholipase C-protein kinase C signaling pathway, induces a mepacrine-sensitive, dose-dependent increase in the release of [3H]arachidonic acid and its metabolites ([3H]AA) from prelabeled Swiss 3T3 fibroblasts. The effect is temporally composed of two phases, i.e. an initial transient burst that is essentially independent of extracellular Ca2+, and a following sustained phase that is absolutely dependent on the extracellular Ca2+. The initial transient [3H]AA liberation occurs concomitantly with bombesin-induced 45Ca efflux from prelabeled cells: both responses being substantially attenuated by loading cells with a Ca2+ chelator quin2. However, bombesin-induced intracellular Ca2+ mobilization by itself is not sufficient as a signal for the initial transient [3H]AA liberation, since A23187 potently stimulates 45Ca efflux to an extent comparable to bombesin but fails to induce [3H]AA release in the absence of extracellular Ca2+. The second sustained phase of the bombesin-induced [3H]AA release is abolished by reducing extracellular Ca2+ to 0.03 mM, although bombesin effects on phospholipase C and protein kinase C activation are barely affected by the same procedure. A protein kinase C activator phorbol 12,13-dibutyrate induces an extracellular Ca(2+)-dependent, slowly developing sustained increase in [3H]AA release, and markedly potentiates both phases of bombesin-induced [3H]AA release. Down-regulation of cellular protein kinase C completely abolishes all of the effects of phorbol dibutyrate, and partially inhibits the second but not the first phase of bombesin-induced [3H]AA release. These results indicate that bombesin-induced receptor-mediated activation of phospholipase A2 involves multiple mechanisms, including intracellular Ca2+ mobilization for the first phase, protein kinase C activation plus Ca2+ influx for the second phase, and as yet unknown mechanism(s) independent of intracellular Ca2+ mobilization or protein kinase C for both of the phases.

Animals↗
PubMed1991

Neurons in the caudal ventrolateral medulla mediate the arterial baroreceptor reflex by inhibiting barosensitive reticulospinal neurons in the rostral ventrolateral medulla in rabbits.

Participation of the caudal ventrolateral medulla in the arterial baroreceptor reflex was examined in urethane-anesthetized, vagotomized and immobilized rabbits whose aortic nerve was cut bilaterally. The extent of the caudal ventrolateral medulla was mapped by decreases in the renal sympathetic nerve activity and arterial pressure following a local microinjection of a neuroexcitatory amino acid, sodium glutamate (0.075-1.5 nmol). It extended between the levels 1.3 mm rostral and 3.0 mm caudal to the obex. An injection of sodium glutamate into the caudal ventrolateral medulla also diminished spontaneous activity of barosensitive reticulospinal neurons in the rostral ventrolateral medulla. In the 'split medulla preparation' in which the medulla was split along the midsagittal plane to disrupt fiber connections associating both sides, a neurotoxic agent, kainic acid, was injected unilaterally into the rostral ventrolateral medulla. This treatment markedly attenuated responses of renal sympathetic nerve activity and arterial pressure induced by a sodium glutamate injection into the ipsilateral caudal ventrolateral medulla, whereas responses to an injection into the contralateral caudal ventrolateral medulla were totally preserved. In four separate experiments, three to five injections of kainic acid were made unilaterally to cover the whole extent of the caudal ventrolateral medulla. The sympathoinhibitory and depressor responses to stimulation of the ipsilateral aortic nerve were then totally abolished. Simultaneously, the cardiac cycle-related rhythmic fluctuation of renal sympathetic nerve activity, which represented activity of the carotid sinus baroreceptor reflex, was attenuated to the noise level. These results, together with our previous electrophysiological demonstration of barosensitive caudal ventrolateral medulla neurons with axonal projections to the rostral ventrolateral medulla, strongly support the hypothesis that neurons in the caudal ventrolateral medulla mediate the arterial baroreceptor-vasomotor reflex through inhibition of barosensitive reticulospinal neurons in the rostral ventrolateral medulla.

Animals↗
PubMed1991

Role of Ca2+ influx in bombesin-induced mitogenesis in Swiss 3T3 fibroblasts.

In Swiss 3T3 fibroblasts a peptide mitogen bombesin, which acts through the phospholipase C-protein kinase C signaling pathway, stimulates DNA synthesis in a manner strictly dependent on the medium calcium concentration: [3H]thymidine incorporation into DNA in the presence of a saturating concentration of bombesin (10(-8) M) is 4-fold greater at 3.0 mM extracellular calcium as compared with a value obtained at 0.03 mM calcium. In the present study we attempted to identify the site and the mechanism of action of Ca2+ influx along the bombesin-induced mitogenic signaling pathway, by comparing bombesin effects at 0.03 and 3.0 mM of medium calcium. Bombesin induces the same extent of increases in [3H]inositol phosphates after 1 min, and comparable sustained increases in the cellular content of 1,2-diacylglycerol for up to 4 h, at either 0.03 or 3.0 mM calcium. Bombesin induces the same extent of phosphorylation of MARCKS protein, the major cellular substrate for protein kinase C, irrespective of the medium calcium concentration for at least 4 h. Moreover, diverse cellular responses elicited by bombesin, including c-fos expression, activation of microtubule-associated protein 2 kinase and S6 kinase, glucose uptake, and protein synthesis but not the release of arachidonic acid and its metabolites, are induced similarly at either 0.03 or 3.0 mM calcium. Down-regulation of cellular protein kinase C nearly completely abolishes bombesin effects on c-fos expression, S6 kinase activation, glucose uptake, and DNA synthesis. These results suggest that the target of Ca2+ influx in bombesin-induced mitogenic signaling pathway is not located along the phospholipase C-protein kinase C signal transduction system including cellular events in early G1 phase that exist downstream to protein kinase C action.

Animals↗