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Biomedical subjects

J Wharton

Publications and source records attributed to J Wharton.

At least 19 recordsLinked to original sources
PubMed2005

Phosphodiesterase type 5 and high altitude pulmonary hypertension.

BACKGROUND: This study explored phosphodiesterase type 5 (PDE5) inhibition as a strategy for treating high altitude pulmonary arterial hypertension (HAPH). METHODS: 689 subjects (313 men) of mean (SD) age 44 (0.6) years living above 2500 m were screened for HAPH by medical examination and electrocardiography, and 188 (27%) met the criteria for right ventricular hypertrophy. 44 underwent cardiac catheterisation and 29 (66%) had a resting mean pulmonary artery pressure (PAP) above 25 mmHg. 22 patients with a raised mean PAP were randomised to receive sildenafil (25 or 100 mg) or matching placebo taken 8 hourly for 12 weeks. RESULTS: At 3 months, patients on sildenafil 25 mg 8 hourly (n = 9) had a significantly (p = 0.018) lower mean PAP (-6.9 mmHg) at the end of the dosing interval than those on placebo (n = 8) (95% CI -12.4 to -1.3). The treatment effect for sildenafil 100 mg 8 hourly (n = 5) compared with placebo was -6.4 mm Hg (95% CI -12.9 to 0.1). Both doses improved 6 minute walk distance, the lower dose by 45.4 m (95% CI 11.5 to 79.4; p = 0.011) and the higher dose by 40.0 m (95% CI 0.2 to 79.8; p = 0.049). Sildenafil was well tolerated. Necroscopic lung specimens from three subjects with HAPH showed abundant PDE5 in the muscular coat of remodelled pulmonary arterioles. CONCLUSIONS: PDE5 is an attractive drug target for the treatment of HAPH and a larger study of the long term effects of PDE5 inhibition in HAPH is warranted.

3',5'-Cyclic-GMP Phosphodiesterases↗
PubMed2002

Developments in therapeutics for pulmonary arterial hypertension.

For many years, the management of pulmonary hypertension has been frustrated by an inadequate understanding of its pathology and limited therapeutic options, but this is changing rapidly. Recently, novel insight into the pathogenesis of primary pulmonary hypertension (PPH) has been provided by the demonstration of mutations in BMPR2 and ALK-1 genes in a significant number of patients with the condition. These genes encode members of the TGF-b receptor superfamily and their integrity is important in the maintenance of normal pulmonary vascular structure and function. At the same time, there has been a major advance in the treatment of the condition due to development of 2 orally active pharmacological agents, bosentan and sildenafil, which demonstrate some selectivity for the pulmonary vasculature. This review examines how the management of PPH and severe pulmonary hypertension in associated diseases has changed and looks at exciting future developments.

Antihypertensive Agents↗
PubMed2001

Adrenomedullin expression and growth inhibitory effects in distinct pulmonary artery smooth muscle cell subpopulations.

The vasodilator peptide adrenomedullin is elevated in patients with pulmonary hypertension and has been implicated in the inhibition of vascular remodeling. We questioned whether adrenomedullin is released by human pulmonary artery smooth muscle cells (PASMCs) and inhibits PASMC growth and release of endothelin, a known smooth muscle cell mitogen. The majority of PASMCs isolated from proximal pulmonary arteries and all PASMCs from distal pulmonary arteries released adrenomedullin, although at differing rates (mean, 177 +/- 28 and 62 +/- 11 fmol/10(5) cells/24 h, respectively). These cells were designated ADM+. However, some proximal PASMC isolates did not release adrenomedullin, designated ADM-. Northern blot analysis confirmed adrenomedullin expression in proximal ADM+ but not ADM- isolates. ADM- and distal ADM+ PASMCs proliferated faster in serum than did proximal ADM+ cells. Adrenomedullin potently and dose-dependently (mean EC(50) = 2.2 +/- 0.5 nM) increased intracellular cyclic adenosine monophosphate (cAMP) in ADM- isolates via specific adrenomedullin receptors. In contrast, both adrenomedullin and calcitonin gene-related peptide modestly elevated cAMP in 50% of ADM+ isolates. Adrenomedullin dose-dependently inhibited platelet-derived growth factor-stimulated [3H]thymidine incorporation and endothelin release in ADM- cells but did not affect [3H]thymidine uptake in ADM+ isolates. We conclude that distinct subpopulations of human PASMCs release and respond to adrenomedullin. The heterogeneity of adrenomedullin release and the inhibition of PASMC DNA synthesis and endothelin release suggest that adrenomedullin may function as a paracrine mediator in the inhibition of pulmonary vascular remodeling.

Adrenomedullin↗
PubMed2001

Differential adrenomedullin release and endothelin receptor expression in distinct subpopulations of human airway smooth-muscle cells.

Although adrenomedullin (ADM) is implicated in the control of airway tone, regulation of ADM release from airway smooth-muscle cells (ASMCs) has not been explored. Preliminary experiments have indicated that human ASMC populations were heterogeneous in their rate of ADM release and expression of endothelin (ET)(A) and ET(B) receptors. We isolated these phenotypically distinct ASMCs from explants derived from the same airway segment. ASMCs possessing exclusively ET(A) receptors appeared smaller and proliferated faster than ET(A)/ET(B) isolates. Macroautoradiographic analysis confirmed the presence of both receptors in human bronchi. ADM release and messenger RNA expression was greater in ET(A)/ET(B) isolates compared with ET(A) isolates. No measurable ET release was detected from ASMCs. Exogenous ET-1 (1 to 100 nM) more potently stimulated the release of ADM from ET(A)/ET(B) compared with ET(A) isolates. In addition, ET-3 (1 to 100 nM) stimulated ADM release only from ET(A)/ET(B) isolates, implicating the ET(B) receptor in this response. Exogenous ET-1 potentiated platelet- derived growth factor-stimulated [3H]thymidine uptake in ET(A)/ ET(B) but not ET(A) isolates. ET-3 did not affect [3H]thymidine uptake in either cell type. Possession of ET(A)/ET(B) receptors is associated with higher rates of ADM release and slower proliferation, but a capacity for ET-1 stimulated DNA synthesis via ET(A) receptors. These results support a paracrine role for the regulation of ADM release predominantly via the ET(B) receptor in human ASMCs.

Adrenomedullin↗
PubMed2000

Prostacyclin analogues differentially inhibit growth of distal and proximal human pulmonary artery smooth muscle cells.

BACKGROUND: Prostacyclin has proved to be a beneficial treatment for patients with severe pulmonary hypertension. We postulated that the response may reflect, at least in part, inhibition of pulmonary artery smooth muscle cell (PASMC) growth. METHODS AND RESULTS: Human PASMCs were derived from distal (<1-mm external diameter, n=8) and proximal (>8-mm external diameter, n=12) pulmonary arteries obtained at transplant surgery and pneumonectomy. The effects of the stable prostacyclin analogues on [methyl-(3)H]thymidine incorporation and cell proliferation were investigated by using immunohistochemically characterized cells. Distal cells proliferated faster than did proximal PASMCs and displayed a distinct sensitivity to cicaprost and iloprost. Both analogues inhibited thymidine uptake over 24 hours (20% to 60%, P<0.001; n=8) and abolished stimulation of DNA synthesis by platelet-derived growth factor-BB (10 ng/mL) in distal but not proximal cells. The inhibitory effect of cicaprost was mimicked by isoproterenol (10(-5) mol/L), forskolin (10(-5) mol/L), and dibutyryl cAMP (5x10(-4) mol/L) and was potentiated by the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (5x10(-5) mol/L). Cicaprost (10(-10) to 10(-6) mol/L) inhibited the proliferation of PASMCs, which had been stimulated with either platelet-derived growth factor-BB or serum, and increased cAMP production. These effects were potentiated by 3-isobutyl-1-methylxanthine and attenuated by the adenylyl cyclase inhibitor 2',5'-dideoxyadenosine (10(-5) to 10(-4) mol/L). CONCLUSIONS: ++Cicaprost and iloprost inhibit DNA synthesis and proliferation to a greater extent in distal compared with proximal human PASMCs, acting at least in part via a cAMP-dependent mechanism. The results are consistent with the hypothesis that prostacyclin analogues inhibit vascular remodeling in pulmonary hypertension and demonstrate heterogeneity among human PASMCs.

Adult↗
PubMed2000

Expression and function of angiotensin converting enzyme, chymase, and angiotensin II in the human radial artery and internal thoracic artery.

BACKGROUND: The potential role of the local renin-angiotensin system to differentially affect radial artery and internal thoracic artery graft performance has not been examined. METHODS: Contractile responses to angiotensin I and II in the radial artery and the internal thoracic artery were examined in vitro. The expression function, and localization of angiotensin receptors, angiotensin converting enzyme, and chymase were studied in radial artery and internal thoracic artery segments. RESULTS: Angiotensin I and II contractions were significantly greater (p < 0.05) in the radial artery compared to the internal thoracic artery. In both arteries, angiotensin II responses were mediated via the AT1 receptor. Messenger RNA transcripts for angiotensin-converting enzyme and chymase were detected in both arteries. Angiotensin-converting enzyme was localized to luminal and vaso vasorum endothelial cells and smooth muscle cells in both vessels, while chymase was colocalized with mast cells in adventitial and medial layers. An angiotensin converting enzyme or a chymase inhibitor singularly had no effect on angiotensin I contractions, however, when combined, a marked inhibition of the angiotensin I response was observed in both vessels. CONCLUSIONS: Our results illustrate the complexities which exist within the local renin angiotensin system and suggest that clinical trials which may modulate the system are warranted.

Aged↗
PubMed2000

Angiotensin converting enzyme in human synovium: increased stromal [(125)I]351A binding in rheumatoid arthritis.

OBJECTIVE: To determine whether tissue angiotensin converting enzyme (ACE) is increased in synovia from patients with rheumatoid arthritis, osteoarthritis or chondromalacia patellae. METHODS: Sections of synovia from patients with rheumatoid arthritis (n = 7), osteoarthritis (n = 7) or chondromalacia patellae (n = 6) were tested for immunoreactivity for ACE, and for binding of the ACE inhibitor [(125)I]351A. The amount of ACE was measured with computer assisted image analysis as the proportion of synovial section area occupied by ACE-immunoreactive cells, and the density of [(125)I]351A binding. RESULTS: [(125)I]351A binding sites had characteristics of ACE and colocalised with ACE-like immunoreactivity to microvascular endothelium and fibroblast-like stromal cells in inflamed and non-inflamed human synovium. Stromal [(125)I]351A binding densities (B(eq)) and the fraction of synovial section area occupied by ACE-immunoreactivity (fractional area) were higher in synovia from patients with rheumatoid arthritis (B(eq) 28 amol/mm(2), fractional area 0.21) than from those with osteoarthritis (B(eq) 9 amol/mm(2), fractional area 0.10) or chondromalacia patellae (B(eq) 9 amol/mm(2), fractional area 0.09)(p < 0.05). Density of [(125)I]351A binding to stroma was similar to that to blood vessels in rheumatoid arthritis, but less dense than vascular binding in chondromalacia patellae and osteoarthritis. Increases in [(125)I]351A binding densities were attributable to increases in the numbers of binding sites, and were consistent with an increase in the density of ACE bearing stromal cells. CONCLUSION: ACE is upregulated in synovial stroma in rheumatoid arthritis. Increased tissue ACE may result in increased local generation of the vasoconstrictor and mitogenic peptide angiotensin II and thereby potentiate synovial hypoxia and proliferation in rheumatoid arthritis.

Adult↗
PubMed1999

Reciprocal changes in endothelial and inducible nitric oxide synthase expression following carotid angioplasty in the pig.

OBJECTIVE: Nitric oxide produced by nitric oxide synthase appears to have an important role in the regulation of arterial tone, platelet adhesion and smooth muscle cell proliferation. Our aim was to investigate the effects of balloon angioplasty on expression of endothelial NO synthase (cNOS) and inducible NO/synthase (iNOS) in the pig carotid artery and to relate any changes in expression to the processes of reendothelialisation and vascular repair. METHODS: Pigs were sacrificed at various time points to follow NOS expression in the neointima, media and regenerated endothelium. Immunocytochemical staining was used to localize cNOS and iNOS expression in the vessel wall. Relative amounts of cNOS were measured using quantitative in vitro alitoradiography. cNOS mRNA and iNOS mRNA was quantified by competitive PCR based on the sequenced cDNA of porcine cNOS and iNOS. RESULTS: Uninjured carotid arteries exhibited dense uniform luminal endothelial staining for cNOS. Balloon angioplasty caused denudation of cNOS immunoreactive cells and a marked reduction of cNOS gene expression but a complete recovery was noted by day 35. In normal uninjured carotid arteries no evidence of iNOS immunoreactivity was demonstrable but 24 h after injury, marked homogeneous iNOS immunoreactivity was detected in medial vascular smooth muscle cells. By 5 days, staining was evident in cells within the forming neointimal layer with no evidence of iNOS immunoreactivity in the media. iNOS immunoreactivity persisted in cells at the luminal surface at 7 days and iNOS gene expression appeared to be sustained in some animals with ruptured internal elastic lamina at 21 days. CONCLUSION: Balloon injury is associated with de-endothelialisation and a marked reduction in cNOS gene expression and activity. iNOS is induced throughout the arterial media within VSMC soon after balloon injury and persists for up to 21 days. These observations imply an important regulatory role for locally generated NO in the pathophysiological response to balloon injury.

Amino Acid Sequence↗
PubMed1999

Angiotensin II activates MAPK and stimulates growth of human pulmonary artery smooth muscle via AT1 receptors.

To determine a potential role for the renin-angiotensin system in the growth of human pulmonary artery (PA) smooth muscle, we studied the localization of angiotensin (ANG) II-receptor subtypes by autoradiography in sections of human PA and in cultured PA smooth muscle cells (PASMCs) and examined the growth responses to ANG II in vitro. Specific 125I-labeled [Sar1,Ile8]ANG II binding was demonstrated within the pulmonary arterial media, but binding to cultured cells varied between isolates. Binding in tissues and cells was inhibited by the ANG II type 1 (AT1) receptor antagonist losartan but not by the type 2 (AT2) receptor antagonist PD-123319. Microautoradiographic studies indicated that cultured PASMCs exhibit heterogeneity with regard to ANG II binding sites. Addition of ANG II to serum-deprived PASMCs, exhibiting a relatively high level of 125I-[Sar1,Ile8]ANG II binding, led to a dose-dependent stimulation of DNA synthesis at 24 h and protein synthesis at 48 h. ANG II led to an increase in cell size without an increase in cell number. These effects were inhibited by losartan but not by PD-123319. In addition, ANG II led to rapid activation of mitogen-activated protein kinase (MAPK), and ANG II-stimulated DNA synthesis was inhibited by the specific inhibitor of MAPK PD-98059. We conclude that the AT1 receptor is expressed by human PASMCs in vivo and in vitro and is coupled to activation of MAPK and increased DNA and protein synthesis in vitro. These results are consistent with the hypothesis that ANG II may be involved in human pulmonary vascular remodeling.

Angiotensin II↗
PubMed1998

Expression of functional angiotensin-converting enzyme and AT1 receptors in cultured human cardiac fibroblasts.

BACKGROUND: Angiotensin II (Ang II) has been implicated in the development of cardiac fibrosis. The aims of the present study were to examine expression and activity of ACE and of angiotensin receptors in human cardiac fibroblasts cultured from dilated cardiomyopathic and ischemic hearts. The effects of Ang II on fibroblasts were also investigated. METHODS AND RESULTS: Human cardiac fibroblasts were cultured from ventricular and atrial myocardium and characterized immunohistochemically. Expression of ACE and the angiotensin AT1 receptor was demonstrated in cardiac fibroblasts by reverse transcriptase-polymerase chain reaction and radioligand binding. Functional ACE activity, measured by radiolabeled substrate conversion assay, was detected in both ventricular (Vmax. Km-1. mg-1, 0.031+/-0.010; n=13) and atrial (0. 034+/-0.012; n=6) fibroblasts. Fibroblast ACE activity was increased after 48 hours of treatment with basic fibroblast growth factor, dexamethasone, and phorbol ester. Ang II did not affect DNA synthesis but stimulated [3H]proline incorporation in cardiac fibroblasts (20.0+/-4.0% increase above control by 10 micromol/L; P<0.05, n=7), which was abolished by losartan 10 micromol/L but not PD123319 1 micromol/L. Ang II also stimulated a rise in intracellular calcium (basal, 56+/-1 nmol/L; Ang II, 355+/-24 nmol/L) via the AT1 receptor, as shown by complete inhibition with losartan. CONCLUSIONS: We have demonstrated expression and activity of ACE and AT1 receptor in cultured human cardiac fibroblasts. In addition, cardiac fibroblasts respond to Ang II with AT1 receptor-mediated collagen synthesis. The presence of local ACE and AT1 receptors in human fibroblasts suggests their involvement in the development of cardiac fibrosis.

Calcium↗
PubMed1998

Differential distribution of angiotensin AT2 receptors in the normal and failing human heart.

Cardiac expression of angiotensin II (Ang II) AT1 and AT2 receptor subtypes is species dependent, and changes in their relative proportion may influence myocardial hypertrophy and fibrosis. Regional differences in the distribution of Ang II receptors in the normal and failing human heart were assessed using 125I-(Sar1,Ile8)Ang II binding and quantitative autoradiography. Receptor subtypes were distinguished by their affinity for selective nonpeptide antagonists (losartan and PD123319) and sensitivity to dithiothreitol. Ventricular and atrial tissues displayed a heterogeneous distribution of ligand binding sites. AT2 receptors predominated, representing 70% to 77% of the sites in normal and noninfarcted myocardium. Endocardial, interstitial, perivascular and infarcted regions in the ventricles of patients with end-stage ischemic heart disease or dilated cardiomyopathy exhibited a significantly greater density (P < .001) of high affinity AT2 binding sites (Kd = 0.57 nmol/liter) compared with adjacent noninfarcted myocardium. Regions displaying the relative increase in AT2 binding sites corresponded to areas of fibroblast proliferation and collagen deposition, shown by picrosirius red staining. AT1 binding sites were localized to nerves, occurred at relatively low density in coronary vessels and represented only 23% to 29% of myocardial 125I-(Sar1,Ile8)Ang II binding sites. The border zone between infarcted and noninfarcted myocardium characteristically contained numerous microvessels, exhibiting perivascular AT2 receptors and endothelial angiotensin converting enzyme activity, as demonstrated by binding of 125I-351A. Specific myocardial AT2 receptor mRNA transcripts (approximately 3 kb) were identified and exhibited alternative splicing of untranslated 5' exons. The differential distribution of cardiac Ang II receptor subtypes and selective increase in binding to AT2 sites in the diseased heart suggest that cells bearing the AT2 receptor represent a significant target for Ang II, possibly contributing to its growth-related actions.

Adolescent↗
PubMed1997

Downregulation of natriuretic peptide C-receptor protein in the hypertrophied ventricle of the aortovenocaval fistula rat.

OBJECTIVES: This study examined the expression of the C-type receptor for the natriuretic peptide family (NPR-C) in the ventricles of normal and aortovenocaval (AV)-fistula rats, the latter a model of cardiac volume overload producing hypertrophy of both ventricles. METHODS: Western blotting with a rabbit anti-NPR-C antibody was used to quantify NPR-C levels in ventricular membranes. NPR-C expression was localised anatomically and measured in frozen sections of cardiac tissue by histochemistry and in vitro autoradiography. RESULTS: Western blot analysis revealed a single band (approximately 120 kDa) in ventricular membranes which was reduced to approximately 60 kDa after treatment with beta-mercaptoethanol. NPR-C immunoreactivity and [125I]rat ANP1-28 binding (displaceable by the NPR-C-specific ligand C-ANP 4-23) were localised to the endocardium. NPR-C protein levels, as measured by all three techniques, were reduced significantly in the hypertrophied ventricles of AV-fistula rats compared to sham-operated animals. CONCLUSIONS: Volume-induced cardiac hypertrophy in the AV-fistula rat is associated with downregulation of endocardial NPR-C. This may be one mechanism by which the endocardium regulates the myocardial response to changes in haemodynamic load.

Animals↗
PubMed1997

Sequential development of angiotensin receptors and angiotensin I converting enzyme during angiogenesis in the rat subcutaneous sponge granuloma.

1. The vasoconstrictor peptide antiotensin II (AII) can stimulate angiogenesis, an important process in wound healing, tumour growth and chronic inflammation. To elucidate mechanisms underlying AII-enhanced angiogenesis, we have studied a subcutaneous sponge granuloma model in the rat by use of 133Xe clearance, morphometry and quantitative in vitro autoradiography. 2. When injected directly into the sponge, AII (1 nmol day-1) increased 133Xe clearance from, and fibrovascular growth in sponge granulomas, indicating enhanced angiogenesis 6 to 12 days after implantation. This AII-enhanced angiogenesis was inhibited by daily doses (100 nmol/sponge) of the specific but subtype non-selective AII receptor antagonist (Sar1, Ile8)AII, and by the selective non-peptide AT1 receptor antagonists losartan and DuP 532. In contrast, AII-enhanced neovascularization was not inhibited by the AT2 receptor antagonist PD123319, nor was it mimicked by the AT2 receptor agonist CGP42112A (each at 100 nmol/sponge day-1). 3. AI (1 nmol/sponge day-1), the angiotensin converting enzyme (ACE) inhibitors captopril (up to 100 micrograms/sponge day-1) and lisinopril (40 micrograms/sponge day-1), or AII receptor antagonists did not affect angiogenesis in the absence of exogenous AII. 4. [125I]-(Sar1, Ile8)AII binding sites with characteristics of AT1 receptors were localized to microvessels and to non-vascular cells within the sponge stroma from 4 days after implantation, and were at higher density than in skin throughout the study. 5. [125I]-(Sar1, Ile8)AII binding sites with characteristics of AT2 receptors were localized to non-vascular stromal cells, were of lower density and appeared later than did AT1 sites. 6. The ACE inhibitor [125I]-351A bound to sites with characteristics of ACE, 14 days after sponge implantation. [125I]-351A bound less densely to sponge stroma than to skin. 7. We propose that AII can stimulate angiogenesis, acting via AT1 receptors within the sponge granuloma. AT1 and AT2 receptors and ACE develop sequentially during microvascular maturation, and the role of the endogenous angiotensin system in angiogenesis will depend on the balanced local expression of its various components. Pharmacological modulation of this balance may provide novel therapeutic approaches in angiogenesis-dependent diseases.

1-Sarcosine-8-Isoleucine Angiotensin II↗