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Biomedical subjects

A D Hughes

Publications and source records attributed to A D Hughes.

At least 19 recordsLinked to original sources
PubMed1996

Activation of endogenous c-Src or a related tyrosine kinase by intracellular (pY)EEI peptide increases voltage-operated calcium channel currents in rabbit ear artery cells.

The effect of activation of endogenous c-Src tyrosine kinase by (pY)EEI peptide was examined on voltage-operated calcium channel (VOC) currents in arterial smooth muscle cells. In single rabbit ear artery cells intracellular application of (pY)EEI peptide increased calcium channel currents. Inactive, non-phosphorylated YEEI peptide had no effect on currents. Peptide-A, a 21 amino acid inhibitor of c-Src inhibited currents and prevented the effect of (pY)EEI peptide on calcium channel currents. These results indicate that activation of intrinsic c-Src increases VOC and support a role for c-Src in the regulation of VOC in vascular smooth muscle cells.

Animals↗
PubMed1996

Platelet-derived growth factor (PDGF): actions and mechanisms in vascular smooth muscle.

1. PDGF is a highly hydrophilic cationic glycoprotein (M(r) 28-35kDa) produced by platelets, monocyte/macrophages, endothelial cells and vascular smooth muscle cells under some conditions. 2. Since its original description, PDGF has attracted much attention and it is currently believed to play a role in atherosclerosis and other vascular pathologies. 3. This review describes the vascular biology of PDGF. It particularly focuses on recent findings regarding the intracellular signals activated by PDGF in the context of vascular smooth muscle cell proliferation, migration and, contraction.

Animals↗
PubMed1996

Effect of angiotension II on the expression of the early growth response gene c-fos and DNA synthesis in human vascular smooth muscle cells.

OBJECTIVES: The aims of this study were to characterize the angiotensin II receptor subtype present on vascular smooth muscle cells from human saphenous vein and to assess the effect of angiotensin II on the expression of the early growth response gene c-fos and on DNA synthesis. METHODS AND RESULTS: Using radioligand binding studies, we have defined the angiotensin II receptors present on these cells as being predominantly of the AT1 subtype. Angiotensin II increased peak intracellular calcium levels by 126 +/- 16 nmol/l (mean +/- SEM) in 17/49 cultures. Angiotensin II induced c-fos expression in a concentration-dependent manner only in cultures that exhibited an intracellular calcium transient in response to stimulation with angiotensin II. The induction of c-fos was inhibited by the selective AT1 antagonist losartan in accordance with the binding studies. Angiotensin II stimulated DNA synthesis with a maximal increase of 66.4% +/- 20.5% over serum-free levels at 1 nmol/l (mean +/- SEM, n = 6, P < 0.05). DNA synthesis declined with increasing angiotensin II concentration, falling to control values at 1 mumol/l, suggesting that a growth-inhibitory influence may counter-balance the stimulatory effect that is observed at lower concentrations. CONCLUSION: Vascular smooth muscle cells from human saphenous vein possess predominantly AT1 receptors and in response to angiotensin II show an induction of c-fos and a modest increase in DNA synthesis.

Angiotensin II↗
PubMed1996

Enhanced tissue polyamine content in the spontaneously hypertensive rat.

1. Endogenous polyamines play a key role in mediating cellular growth and differentiation. Hypertension is associated with structural modifications of the circulatory system, a process that may be facilitated by polyamines. In this study, we examined whether there are elevated polyamine concentrations in the cardiovascular tissues of spontaneously hypertensive rats (SHR) relative to Wistar-Kyoto (WKY) rats. We also determined the chronic effect of 2% difluoromethylornithine (DFMO; a polyamine biosynthesis inhibitor) on tissue polyamines and hypertension. 2. SHR and WKY rats were treated with either 2% DFMO or drug-free drinking water; blood pressure was measured on alternate days and tissue polyamines were analysed at the end of the study. 3. We found that spermidine and spermine concentrations were markedly raised in the ventricles, resistance vessels and liver of the SHR, in comparison with corresponding tissues of WKY rats. DFMO did not affect SHR resistance vessel and liver polyamines, although spermidine in the ventricles was reduced. The blood pressure of neither SHR nor WKY rats was affected by DFMO. 4. In conclusion, this study shows for the first time a raised concentration of polyamines in the resistance vasculature of the SHR, in relation to their normotensive counterparts. The inability of DFMO to significantly reduce tissue polyamines in this study is in contrast to the effectiveness of 2% DFMO in other hypertension models, suggesting that polyamine homeostasis in this model may be stringently regulated.

Animals↗
PubMed1996

Depletion of resistance vessel polyamines attenuates angiotensin II induced blood pressure rise in rats.

Vascular structural changes in hypertension are proposed to contribute to raised peripheral resistance. Endogenous polyamines (putrescine, spermidine and spermine) have an essential role in cellular growth and may be required in vascular restructuring. We have previously shown raised polyamines in resistance vessels in response to angiotensin II (angII) infusion in the rat and here we examined whether polyamine depletion influences the hypertensive process. Wistar-Kyoto rats were infused with either angII or saline by osmotic minipump, and maintained on either 2% difluoromethylornithine (DFMO; polyamine synthesis inhibitor) or water for 12 days. AngII significantly increased tail-cuff blood pressure (bp), resistance vessel spermidine and spermine concentration, and media:lumen ratios. DFMO attenuated both the rise in bp (p < 0.05) and vascular spermidine (p < 0.05) in the angII infused rats but vascular structure was apparently unaffected on day 12. In conclusion, raised concentration of resistance vessel spermidine may be a necessary component of the ang II pressor effect.

Angiotensin II↗
PubMed1996

The role of isoprenoids in vascular smooth muscle: potential benefits of statins unrelated to cholesterol lowering.

Elevated plasma cholesterol is a well recognised risk factor for cardiovascular disease. Numerous studies have shown that a reduction in circulating cholesterol results in a reduced cardiovascular risk. The statins are a group of cholesterol lowering agents which act by inhibition of a key enzyme in cholesterol synthesis; 3-hydroxy 3-methyliglutaryl coenzyme A (HMG CoA) reductase. This enzyme is responsible for the conversion of HMG CoA to mevalonate. In addition to their efficacy as cholesterol lowering agents, statins inhibit the proliferation, migration and signalling of vascular smooth muscle cells. Inhibition of isoprenoid synthesis and resultant reduction in prenylation of key cellular proteins such members of p21ras and p21rho families may underlie some of these effects.

Animals↗
PubMed1995

pp60c-src increases voltage-operated calcium channel currents in vascular smooth muscle cells.

Intracellular application of pp60c-src, a nonreceptor tyrosine kinase present in large amounts in smooth muscle cells increased voltage-operated calcium channel currents in rabbit ear artery cells. Intracellular peptide-A, an inhibitor of pp60c-src, reduced calcium channel currents and abolished the action of pp60c-src. Selective tyrosine kinase inhibitors, tyrphostin-23 and genistein also abolished the effect of pp60c-src, but inhibition of protein kinase C did not prevent the action of pp60c-src. These results suggest that endogenous pp60c-src modulates voltage-operated calcium channels by a mechanism dependent on tyrosine phosphorylation but not involving activation of protein kinase C.

Animals↗
PubMed1995

Comparison of effects of platelet-derived growth factor isoforms on signaling and DNA synthesis of human cultured saphenous vein cells.

We examined the effect of platelet-derived growth factor (PDGF)-AA, PDGF-AB, and PDGF-BB isoforms on DNA synthesis, Ca2+ mobilization, and tyrosine phosphorylation in cultured human saphenous vein cells cultured by an explant technique; confluent cells derived from passage 3 were used for all studies. DNA synthesis was measured by [methyl3H]thymidine uptake, intracellular [Ca2+] ([Ca2+]i) was measured with the Ca(2+)-sensitive indicator fura-2, and tyrosine phosphorylation was measured by Western blotting techniques. All three isoforms of PDGF stimulated [methyl3H]thymidine uptake concentration dependently, with similar potency. PDGF-AB induced significantly greater [methyl3H]-thymidine uptake than PDGF-BB, and PDGF-AA was much less effective than either PDGF-AB or PDGF-BB. The effects of all three isoforms were inhibited by tyrphostin-23, a selective inhibitor of tyrosine kinases PDGF-AB and PDGF-BB increased [Ca2+]i, although the maximum response to PDGF-AB was significantly less than that to PDGF-BB. Both isoforms increased [Ca2+]i by stimulating influx and intracellular release of Ca2+. PDGF-AA had no measurable effect on [Ca2+]i. All three isoforms increased tyrosine phosphorylation of a 170-kDa protein detected by Western blotting. Quantitative densitometry indicated that PDGF-BB induced greater tyrosine phosphorylation than PDGF-AB and both PDGF-BB and PDGF-AB induced markedly more tyrosine phosphorylation than PDGF-AA. PDGF isoforms have differing efficacies in terms of DNA synthesis, Ca2+ mobilization, and tyrosine phosphorylation by human saphenous vein cells.

Blotting, Western↗
PubMed1995

Inhibition of norepinephrine and caffeine-induced activation by ryanodine and thapsigargin in rat mesenteric arteries.

We examined the effects of ryanodine and thapsigargin on changes in cytoplasmic [Ca2+] (Cai) and muscle tension in rat mesenteric resistance arteries induced by norepinephrine (NE) and caffeine. Both ryanodine and thapsigargin markedly inhibited the increase in Cai and contractile responses to caffeine in physiological saline and to NE and caffeine in calcium-free conditions. In contrast, peak responses to potassium depolarisation and NE in physiological saline appeared little affected, although time taken to achieve 50% of peak response after addition of NE was slowed after ryanodine and thapsigargin treatment. Neither ryanodine nor thapsigargin altered resting tone or Cai or the Ca2+ sensitivity of contraction under depolarized conditions. The NE concentration-response relationship was not significantly altered after ryanodine or thapsigargin. Ryanodine and thapsigargin inhibit the release of intracellular Ca2+ stores by NE and caffeine. Inhibition of release of intracellular Ca2+ by NE has only slight effects on contractile responses of mesenteric resistance arteries when extracellular Ca2+ is present.

Animals↗
PubMed1995

Increase in tone and intracellular Ca2+ in rabbit isolated ear artery by platelet-derived growth factor.

1. The effect of platelet-derived growth factor (PDGF-AB) on tone and intracellular Ca2+ ([Ca2+]i) was examined in rabbit isolated ear arteries. Arteries were mounted in a myograph and loaded with the Ca(2+)-sensitive fluorescent indicator, fura-2, for concurrent measurements of isometric force and [Ca2+]i. 2. PDGF-AB contracted rabbit ear artery in a concentration-dependent manner. PDGF-AB induced tone was associated with a rise in [Ca2+]i. In the presence of noradrenaline, PDGF-AB induced a similar rise in [Ca2+]i but contraction in response to PDGF-AB in the presence of noradrenaline was increased compared with PDGF-AB alone. 3. PDGF-AB-induced rise in [Ca2+]i and tone were abolished by removal of extracellular Ca2+ (with addition of BAPTA, a Ca2+ chelator), and by preincubation with a dihydropyridine calcium channel blocker, (-)-202,791. Bistyrphostin, a selective inhibitor of tyrosine kinases, also inhibited PDGF-AB-induced tone, but had no effect on noradrenaline- or potassium-induced tone. 4. PDGF-AB contracts rabbit ear artery by increasing Ca2+ entry through voltage-operated calcium channels. This effect involves activation of a tyrosine kinase.

Animals↗
PubMed1995

Effect of platelet-derived growth factor on voltage-operated calcium channels in rabbit isolated ear artery cells.

1. Platelet derived growth factor (PDGF), AB and BB isoforms (100 pM) increased calcium channel currents measured by whole cell voltage clamp technique in single vascular smooth muscle cells isolated from rabbit ear arteries. 2. Tyrphostin-23 (100 microM) a selective inhibitor of protein tyrosine kinases, reduced calcium channel currents. Pre-incubation with tyrphostin-23 prevented PDGF-AB induced increase in calcium channel currents. However, in these same cells 10 nM (+)-202791, a dihydropyridine calcium channel agonist, did increase calcium channel currents. 3. Bistyrphostin (10 microM), a selective inhibitor of epidermal growth factor (EGF)-kinase also reduced calcium channel currents and inhibited PDGF-AB-induced increases in calcium channel currents. 4. Genistein (100 microM) a selective inhibitor of tyrosine kinases, structurally unrelated to the tryphostins, also inhibited calcium channel currents and pre-incubation with genistein prevented the PDGF-AB-induced rise in calcium channel currents. 5. These results indicate that PDGF increases calcium channel currents in vascular smooth muscle. This action of PDGF probably involves a tyrosine kinase.

Analysis of Variance↗