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J M Connell

Publications and source records attributed to J M Connell.

At least 109 records · Page 6Linked to original sources
PubMed1993

G-proteins in essential hypertension: a study of human platelet plasma membranes.

AIM OF STUDY: Guanine nucleotide regulatory (G) proteins act as key signal transducers for many hormones, growth factors and neurotransmitters, and have been shown to have an important influence on platelet function. As abnormal G-protein levels and activity have been reported in platelets from human non-insulin-dependent diabetics (NIDDM) we studied G-protein function in essential hypertension, a condition which is also associated with insulin resistance and in which abnormal platelet function has been reported. METHODS: G-protein function was deduced from studies of adenylyl cyclase activity in platelet membrane preparations from 14 untreated essential hypertensives and 14 controls matched as far as possible for age and sex. Levels of G-protein subunits (Gs alpha, Gi alpha 2 and beta-subunits) were assessed by immunoblotting, using platelets from 15 subjects with untreated essential hypertension and 15 controls. RESULTS: No changes in levels of G-proteins (Gs alpha, Gi alpha 2 and beta-subunits) were seen. However, in contrast to the observations in NIDDM, the studies of adenylyl cyclase function identified greater prostaglandin E1-stimulated activity in hypertensive platelet membranes than in controls (88.8 verus 72% stimulation, P = 0.018). This may have a physiological basis in protecting cells against a Ca2+ overload. CONCLUSION: These data are in opposition to the theory that a common defect in G-proteins can explain the association between hypertension and NIDDM.

Adenylyl Cyclases↗
PubMed1993

Fetal angiotensin II levels and vascular (type I) angiotensin receptors in pregnancies complicated by intrauterine growth retardation.

OBJECTIVE: To investigate the status of the fetal renin-angiotensin system (RAS) in pregnancies complicated by severe intrauterine growth retardation (IUGR), and its possible relationship to elevated fetoplacental vascular resistance as indicated by abnormal umbilical artery Doppler flow velocity waveforms (FVW). DESIGN: Prospective survey of pregnancies falling into predefined categories and presenting at the Queen Mothers Hospital, Glasgow, over the study period. SUBJECTS: Effects of mode of delivery and gestational age were investigated using uncomplicated term pregnancies delivered vaginally (SVD group, n = 15) or by elective caesarean section (ECS group, n = 9), and normal pregnancies with spontaneous preterm onset of labour (PREM group, n = 6; normal birthweight for gestational age (31 weeks)). These groups were used as controls for the 13 IUGR cases delivered preterm (31 weeks) by caesarean section in the fetal interest. MAIN OUTCOMES MEASURES: Umbilical artery FVW, birthweight, cord venous angiotensin II concentration ([cv ANG II]), fetoplacental vascular ANG II receptor concentration. RESULTS: Cord venous angiotensin II concentration was similar to maternal values in the ECS group (31-101 pmol/l, 95% CI), but was elevated (81-288 pmol/l, P = 0.03) after vaginal delivery. The concentration of ANG II receptors (type AT1, dissociation equilibrium constant, 1.27 nmol/l) in placental primary/secondary stem vascular tissue was lower in the SVD group (18-44 fmol/mg membrane protein, 95% CI), compared with the ECS group (29-122 fmol/mg, P = 0.03) consistent with acute receptor down-regulation by the elevated ANG II levels. No effect of gestational age on receptor number was demonstrable (P = 0.13, PREM (premature delivery) vs ECS group). In the IUGR group, [cv ANG II] (94-378 pmol/l) was markedly elevated compared with the ECS controls (P = 0.001) but receptor concentration (28-84 fmol/mg) was not significantly altered (P = 0.13). No relationships between [cv ANG II] or receptor number and umbilical artery FVW could be identified. No changes in receptor affinity were observed. CONCLUSION: These results indicate activation of the fetal RAS in IUGR and suggest that responsiveness of the fetoplacental vasculature to the peptide is not diminished as would be expected from the elevated plasma ANG II levels. ANG II may contribute to the increased fetoplacental vascular resistance observed in this disorder, but does not apparently account for the abnormal umbilical artery FVW that is observed in a proportion of IUGR cases.

Angiotensin II↗
PubMed1993

Characterization of atrial natriuretic peptide receptors in human fetoplacental vasculature.

Two classes of high-affinity binding sites for atrial natriuretic peptide (ANP) were identified in a microsomal fraction from human placental artery using radioligand binding methods and des[Gln18,Ser19,Gly20,Leu21,Gly22]ANP-(4-2 3) (C-ANP), a partially ring-deleted analogue of ANP, consistent with the presence of ANP-A and ANP-C receptor subtypes in this tissue [dissociation equilibrium constant (Kd) 58 pM, maximum binding capacity (Bmax) 14 fmol/mg membrane protein, and Kd 82 pM, Bmax 28 fmol/mg, respectively]. ANP activated a guanylate cyclase present in a particulate fraction from placental vascular tissue with half-maximal response at 104 pM and a maximal rate of guanosine 3',5'-cyclic monophosphate production of 62 pmol.min-1 x mg protein-1. Human brain natriuretic peptide was 10-fold less effective than ANP in stimulating guanylate cyclase activity, indicating the absence of the ANP-B receptor subtype. C-ANP had no effect on basal or ANP-stimulated enzyme activity. This report demonstrates the presence of functional (guanylate cyclase-coupled) receptors for ANP in the human fetoplacental vasculature, suggesting that ANP may have a role in the regulation of fetoplacental hemodynamics.

5'-Nucleotidase↗
PubMed1993

Guanine nucleotide regulatory proteins in the spontaneously hypertensive rat.

We compared guanine nucleotide regulatory protein (G protein) levels and function in plasma membranes from resistance vessels (mesenteric arteries) isolated from spontaneously hypertensive (SHR) and normotensive Wistar rats. G protein function was deduced from studies of adenylate cyclase activity. Although the basal level of adenylate cyclase activity (+/- Mn2+ ions) was significantly greater in SHR membranes, addition of agents that function via the stimulatory G protein--i.e., NaF (10(-2) M), (-)-isoproterenol (10(-4) M), and prostaglandin E1 (10(-5) M)--resulted in a significantly lower stimulatory response in SHR membranes. Ligands that function via the inhibitory G protein--i.e., adrenaline (10(-5) M)/propranolol (10(-5) M) (this combination being equivalent to an alpha 2-receptor agonist), carbachol (10(-3) M), and serotonin (10(-5) M)--were responsible for only slight inhibitory responses in both SHR and Wistar rat membranes, which were not significantly different. Western blotting identified the presence of Gs, Gi2, and Gi3 alpha-subunits in rat vascular smooth muscle, but there were no differences in the levels of these G protein alpha-subunits found in SHR and Wistar rat plasma membranes. The levels of the beta-subunit in the two sets of membranes were also similar. In conclusion, there is a reduced response in adenylate cyclase activity to agents that function via the stimulatory G protein in SHR membranes. However, this is not a consequence of altered levels of the different G protein subunits.

Adenylyl Cyclases↗
PubMed1993

Atrial natriuretic hormone: a paracrine or endocrine role within the human placenta?

There is evidence to suggest that the human placenta may synthesize atrial natriuretic hormone (ANH). To test this hypothesis we examined specimens of normal term placenta obtained at elective caesarean section for evidence of either storage or synthesis of this peptide. No immunocytochemical or ultrastructural evidence of ANH storage was found. RNA extracted from cultured cytotrophoblast cells, the chorionic or decidual plates, or from placental villi contained no ANH-encoding messenger RNA by Northern blot analysis. Polymerase chain reaction amplification of these RNA extracts using primers directed to the midportion of the ANH gene failed to demonstrate any evidence of ANH-encoding RNA. We conclude that ANH is neither stored nor synthesized within the normal term placenta, and thus a purely endocrine role for this peptide within the fetoplacental compartment is proposed.

Atrial Natriuretic Factor↗
PubMed1993

Fetal endothelin levels and placental vascular endothelin receptors in intrauterine growth retardation.

OBJECTIVE: To examine endothelin 1 (ET-1) in normal pregnancies and those complicated by severe intrauterine growth retardation (IUGR) with normal or increased fetoplacental vascular resistance as assessed by umbilical artery Doppler flow velocimetry. METHODS: Endothelin 1 concentrations were measured by radioimmunoassay in fetal umbilical arterial and venous and maternal venous plasma, and vascular ET-1 receptors were quantified by radioligand binding methods in placental primary and secondary stem villous vessels. RESULTS: Fetal and maternal ET-1 concentrations were similar in normal pregnancy (10-15 pmol/L) and were not influenced by mode of delivery (vaginal versus cesarean). Fetal levels were identical to control values in the growth-retarded fetuses with normal umbilical artery flow, but were markedly elevated (above 50 pmol/L) in two cases with increased fetoplacental vascular resistance. The corresponding maternal levels also appeared elevated, but to a lesser extent. Endothelin receptors were identified in placental vascular tissue. No differences in receptor-binding characteristics were observed between the groups. CONCLUSION: The increased fetoplacental vascular resistance present in a proportion of pregnancies complicated by severe IUGR may be associated with elevated fetal plasma ET-1 concentrations, but with no apparent change in vascular sensitivity to the peptide.

Binding, Competitive↗
PubMed1993

Guanine nucleotide regulatory protein alterations in the Milan hypertensive rat strain.

OBJECTIVE: To examine whether the altered regulation of adenylyl cyclase that has been reported in vascular tissues from spontaneously hypertensive rats is also evident in the Milan hypertensive (MHS) rat strain. DESIGN: The plasma membranes of vascular smooth muscle cells derived from thoracic aortae from adult (60-day-old) MHS and Milan normotensive (MNS) strain rats were studied. METHODS: Guanine nucleotide regulatory protein (G-protein) function was inferred from adenylyl cyclase activity studies, and levels of G-protein subunits were assessed by immunoblotting. beta-Adrenergic receptor number and affinity were measured from the binding of the antagonist [125I]-cyanopindolol. RESULTS: Basal adenylyl cyclase activity was increased significantly in MHS rat cell membranes, and stimulation by 0.1 mmol/l isoproterenol and 0.01 mmol/l prostaglandin E1 was significantly greater in MHS than in MNS rat cell membranes. Forskolin (at 0.1 mmol/l) resulted in a significantly greater stimulatory response in MHS membranes, which was eliminated by 0.01 mol/l NaF. Biphasic effects of GTP on isoproterenol-stimulated membranes demonstrated similar Gi function in MHS and MNS rat cell membranes, although a greater stimulatory GTP response was observed in MHS rat cell membranes. The levels of Gs alpha (both forms), Gi3 alpha and the beta-subunit were reduced in MHS rat cell membranes, whereas the levels of Gi2 alpha and Gq alpha and G11 alpha were unchanged. The number of beta-adrenoceptors was increased significantly in MHS rat cell membranes, whereas receptor affinity for the antagonist was unaltered. CONCLUSIONS: There are differences in adenylyl cyclase stimulatory responses in MHS rat vascular smooth muscle cell membranes. We have found evidence of reduced levels of particular G-protein subunits, altered beta-adrenoceptor-Gs coupling and increased beta-adrenoceptor number.

Adenylyl Cyclases↗
PubMed1993

Metabolic effects of lacidipine: a placebo-controlled study using the euglycaemic hyperinsulinaemic clamp.

1. Twelve healthy male volunteers received lacidipine 4 mg and matching placebo, each for 2 weeks, in a randomised, double-blind crossover study, and attended on 4 study days to evaluate the effects of single and multiple dosing using the euglycaemic hyperinsulinaemic 'clamp'. 2. On each study day, a primed constant-rate infusion of soluble insulin (1.5 mu kg-1 min-1) was administered for 180 min with a variable-rate infusion of 20% dextrose to maintain euglycaemia (5.2 mmol l-1). Whole-body insulin sensitivity was calculated during the past 40 min of the 'clamp'. At frequent intervals, measurements of BP and HR were recorded and venous blood samples collected for serum insulin, C-peptide, potassium, triglyceride (TG) and plasma noradrenaline concentrations. 3. Lacidipine was generally well tolerated and there were no adverse biochemical events. Mean values for insulin sensitivity +/- s.d. were 8.9 +/- 1.6 and 9.1 +/- 2.0 mg kg-1 min-1 after single doses of lacidipine and placebo respectively (95% CI, -1.0, 1.3), and correspondingly 9.6 +/- 2.1 and 9.7 +/- 1.5 mg kg-1 min-1 after 2 weeks (95% CI, -1.0, 1.3). 4. There was a significant reduction in fasting serum TG concentrations after 2 weeks of lacidipine: 0.7 +/- 0.3 mmol l-1 vs 0.9 +/- 0.6 (P < 0.001). However, changes in serum TG and potassium concentrations during the 'clamp' were not significantly different between the 4 study days. 5. Thus, in 'insulin sensitive' volunteers, lacidipine reduces fasting serum TG concentrations but has no effect on insulin-stimulated uptake of glucose, potassium and TG under euglycaemic hyperinsulinaemic conditions.

Adolescent↗
PubMed1992

Guanine nucleotide regulatory protein levels and function in spontaneously hypertensive rat vascular smooth-muscle cells.

We compared G-protein levels and function in membranes from vascular smooth-muscle cells (VSMC) derived from mesenteric arteries from SHR, WKY and Wistar rats. Basal adenylyl cyclase activity was significantly reduced in SHR membranes compared with Wistar, but was similar to WKY. Isoproterenol stimulation (10(-4) M) was significantly lower in SHR membranes compared to WKY, but was similar to that in Wistar, which was also significantly lower than WKY. Forskolin (10(-4) M) and NaF (10(-2) M), resulted in a higher stimulatory response in SHR membranes. Biphasic effects of GTP on isoproterenol-stimulated membranes demonstrated unaltered Gi function in SHR membranes. No significant differences were seen in the levels of Gs alpha (44- and 42-kDa forms), Gi2 alpha and the beta-subunit in immunoblotting studies of the membranes. Amounts of Gq alpha/G11 alpha and Gi3 alpha were also unchanged. In conclusion, there are differences in adenylyl cyclase responses in SHR VSMC membranes which are not a consequence of altered levels of G-proteins, but may reflect genetic differences rather than effects of hypertension.

Adenylyl Cyclases↗
PubMed1992

Glucocorticoid-suppressible hyperaldosteronism results from hybrid genes created by unequal crossovers between CYP11B1 and CYP11B2.

Glucocorticoid-suppressible hyperaldosteronism (GSH) is an autosomal dominant form of familial hypertension. The biochemical abnormalities seen in this disorder may be remedied by administration of dexamethasone, implying that aldosterone synthesis is being abnormally regulated by corticotropin. The final three steps of aldosterone synthesis, 11 beta- and 18-hydroxylation and 18-oxidation, are mediated by a cytochrome P450 in the zona glomerulosa of the adrenal cortex termed CYP11B2. A related isozyme in the zona fasciculata, CYP11B1, is required for cortisol synthesis; this isozyme, which is normally expressed at much higher levels than CYP11B2, only has 11 beta-hydroxylase activity. These isozymes are encoded by genes on human chromosome 8q22. We have now studied four unrelated patients with GSH. We found that each patient has one chromosome that carries three CYP11B genes instead of two. This has presumably been generated by unequal meiotic crossing-over. The extra gene is a hybrid with 5' regulatory and coding regions corresponding to CYP11B1 and 3' coding regions from CYP11B2. The breakpoint is in intron 2 in two cases, intron 3 in one, and exon 4 in one. Cells transfected with hybrid cDNAs containing up to the first three exons of CYP11B1 synthesized aldosterone at levels near that of cells carrying normal CYP11B2, but cells transfected with hybrids containing the first five or more exons of CYP11B1 could not synthesize detectable amounts of aldosterone. These data demonstrate that GSH is caused by expression of a gene that is regulated like CYP11B1 but that encodes a protein able to synthesize aldosterone.

Blotting, Southern↗
PubMed1992

The structure and activity of two cytochrome P450c21 proteins encoded in the ovine adrenal cortex.

The steroidogenic enzyme cytochrome P450c21 (CYP21A1) is synthesized in the adrenal cortex and is essential for cortisol and aldosterone production. We have studied the structure and activity of ovine P450c21 proteins by analysis and expression of the corresponding cDNAs. Two P450c21 mRNAs (2.2 and 1.7 kilobases) were detected in ovine adrenal RNA and corresponded to two types of P450c21 cDNA clones that differed in their 3' region. One clone encoded a protein similar in structure to bovine, murine, and human P450c21 proteins. The other clone contained a 3' deletion of about 500 nucleotides and encoded a P450c21 protein that was truncated by 18 residues at the carboxyl terminus. The boundaries of this deletion suggested that an additional splicing event was responsible for the shortened mRNA sequence. Detailed Southern analysis of ovine genomic DNA indicates that the two mRNAs are derived from one gene even though two P450c21 genes are present in the ovine genome. The activities of the two P450c21 proteins were determined by expressing the respective cDNA clones in COS cells. The complete P450c21 protein was an efficient catalyst of 21-hydroxylation reactions, whereas no 21-hydroxylation activity was detected in cells containing the P450c21 protein with the carboxyl-terminal deletion.

Adrenal Cortex↗
PubMed1992

Candoxatril, an orally active neutral endopeptidase inhibitor, raises plasma atrial natriuretic factor and is natriuretic in essential hypertension.

OBJECTIVE: Candoxatril (UK79,300) is an orally available inhibitor of the neutral endopeptidase (E.C.3.4.24.11) which degrades atrial natriuretic factor (ANF). This study was designed to establish initial safety and efficacy data in essential hypertension for this novel class of drug. DESIGN: A prospective, double-blind, placebo-controlled, single-dose comparison of candoxatril with placebo in a crossover manner. Three doses of candoxatril (10, 50 and 200 mg) were used, with four subjects at each dose level. SETTING: The Medical Research Council Blood Pressure Unit, Western Infirmary, Glasgow, UK (a hospital clinical research unit). PATIENTS: Twelve patients with untreated essential hypertension. Diastolic blood pressure was greater than 95 mmHg on three separate occasions before entry to the study. INTERVENTION: Candoxatril or matching placebo was administered orally in the fasting state. Serial measurements of urinary volume and electrolyte excretion were taken (on each hour, urine volume was replaced with an equivalent volume of water by mouth). Blood pressure and heart rate were recorded for 12 h after drug administration and serial blood samples were taken for measurement of plasma ANF and neurohormone concentrations. MAIN OUTCOME MEASURES: Urine volume and electrolyte concentration; blood pressure; heart rate; plasma atrial natriuretic factor. RESULTS: Plasma ANF concentrations rose significantly in all patients within 2 h of candoxatril administration compared with placebo although peak and integrated ANF levels were similar at all three doses. A significant natriuresis was only seen after 200 mg candoxatril, with a greater cumulative urine sodium excretion over 12 h compared with placebo; this was associated with a greater diuresis over 12 h compared with placebo. After a single oral dose of candoxatril, blood pressure and heart rate remained unchanged. CONCLUSIONS: Candoxatril in a single dose caused no adverse effects in essential hypertension. The drug caused a rise in basal ANF levels at all doses, but natriuresis was only seen with the highest dose used. No change in blood pressure was recorded after acute dosing, and the results of chronic studies with this compound are awaited. Oral inhibitors of ANF degradation may have therapeutic potential in cardiovascular disorders.

Administration, Oral↗
PubMed1992

Maternal and fetal atrial natriuretic peptide levels at delivery from normal and growth retarded pregnancies.

OBJECTIVE: To determine whether circulating fetal levels of the vasodilator atrial natriuretic peptide (ANP) are reduced in pregnancies complicated by intrauterine growth retardation (IUGR). DESIGN: Prospective observational study. SETTING: University teaching hospital and research laboratory. SUBJECTS: 25 normal singleton pregnancies delivered at term by spontaneous vertex delivery (n = 16) or by elective caesarean section (n = 9), and a series of 14 singleton pregnancies complicated by IUGR. INTERVENTION: Measurement of ANP by radio-immunoassay in maternal venous, umbilical artery, and umbilical vein plasma from a series of normal, and IUGR pregnancies. MAIN OUTCOME MEASURES: Comparison of plasma ANP levels between the three groups; relation between fetal ANP, PO2 and pH. RESULTS: Mode of delivery did not influence either maternal, umbilical artery or umbilical vein plasma ANP levels in normal term singleton pregnancies. Umbilical vein ANP levels were significantly higher in the IUGR group when compared with normal pregnancies at term (mean 66 95%, CI 36-122 vs mean 37, 95% CI 29-47 pg/ml, P = 0.03) and were inversely related to umbilical artery pH (R2 = 65%; P = 0.003). CONCLUSIONS: These data suggest that umbilical vein ANP levels are elevated in pregnancies complicated by IUGR, and rise appropriately in response to the stress of acidosis. In the absence of any receptor or second messenger defect within feto-placental vascular smooth muscle, these data suggest that ANP is not directly implicated in the vascular pathophysiology of IUGR.

Atrial Natriuretic Factor↗
PubMed1992

Effects of dexamethasone on G protein levels and adenylyl cyclase activity in rat vascular smooth muscle cells.

Dexamethasone administration in vitro has been shown to increase adenylyl cyclase activity in vascular smooth muscle cells (VSMC) from renal arteries and in non-vascular cell lines. To investigate whether G proteins are involved in this response, cultured VSMC from mesenteric arteries of Sprague-Dawley rats were incubated in the presence and absence of 10 nM dexamethasone for 24 and 48 h. Basal and stimulated adenylyl cyclase activities were increased by approximately 50% after treatment with dexamethasone. The changes were neither specifically associated with ligands which stimulate adenylyl cyclase catalytic unit via Gs (isoproterenol and prostaglandin E1) nor with guanylylimidodiphosphate (0.1 nM), which inhibits the catalytic unit via Gi. This suggests that dexamethasone enhances adenylyl cyclase activity through changes at the level of the catalytic unit, rather than through the G proteins which modulate its activity. No differences were seen in immunoblotting studies of the levels of Gi alpha 2, Gs alpha, Gi alpha 3 and beta subunits. Similarly, dexamethasone had no effect on the expression of mRNA for Gi alpha 2 and Gs alpha. The results indicate that glucocorticoid-induced increases of adenylyl cyclase activity are due to changes at the level of the adenylyl cyclase catalytic unit rather than alteration of the levels or turnover of Gs alpha, Gi alpha 2, Gi alpha 3 and beta subunits in the membranes of VSMC.

Adenylyl Cyclases↗