Search PubMed⌕ Search

Biomedical subjects

D A Cox

Publications and source records attributed to D A Cox.

At least 55 records · Page 3Linked to original sources
PubMed1995

Is nitric oxide involved in 5-HT2B receptor-mediated contraction in the rat stomach fundus?

Although contraction of the rat stomach fundus by 5-HT is known to be mediated by the 5-HT2B receptor, the second messenger pathways involved in this response remain unclear. Since nitric oxide (NO) has been associated with contraction of certain gastrointestinal smooth muscle, the purpose of this study was to determine if NO is involved in 5-HT-induced contraction in the rat stomach fundus. The arginine analogs L-NAME and L-NMMA, at a concentration (100 microM) established to inhibit NO synthase in the rat stomach fundus by inhibiting depolarization-induced relaxation in this tissue, had no effect on 5-HT contraction. Furthermore, the NO donors sodium nitroprusside and SNAP did not contract rat stomach fundus under basal tone, whereas 5-HT was a potent contractile agonist. These data do not support a role for NO in 5-HT2B receptor-mediated contraction in the rat stomach fundus.

Animals↗
PubMed1995

Transforming growth factor-beta 3.

Transforming Growth Factor-Beta (TGF-beta) is the general name for a family of naturally-occurring polypeptides which have multiple regulatory effects on cell proliferation and differentiation. Over the last decade it has become apparent that TGF-betas can be produced by most cell types and exert a wide range of effects in a context-dependent autocrine, paracrine or endocrine fashion via interactions with distinct receptors on the cell surface. This review summarizes current knowledge concerning the molecular and cellular biology of TGF-beta 3, the most recently described mammalian isoform, and focuses on those physiological actions which may lead to clinical applications, particularly in the indication areas of wound healing and chemoprotection.

Animals↗
PubMed1995

5-Hydroxytryptamine2B receptor signaling in rat stomach fundus: role of voltage-dependent calcium channels, intracellular calcium release and protein kinase C.

The rat stomach fundus is enriched with the 5-hydroxytryptamine (5-HT)2B receptor, the newest subtype of the 5-HT2 receptor family to be cloned. Although the 5-HT2A and 5-HT2C receptor subtypes couple to phosphatidylinositol hydrolysis, such a coupling has not been established for the 5-HT2B receptor in tissues. Thus, the purpose of this study was to characterize further the signal transduction mechanism of the 5-HT2B receptor in rat stomach fundus. Nitrendipine (1 microM) inhibited the maximal contraction to 5-HT (1 microM) by approximately 50%. Removal of extracellular calcium did not inhibit 5-HT contraction to a greater extent than that produced by nitrendipine, indicating that calcium influx through voltage-dependent calcium channels was predominantly responsible for the dependence of the 5-HT contraction on extracellular calcium. Depletion of both extracellular calcium and intracellular calcium stores abolished 5-HT contraction. Ryanodine (30 microM), a compound which inhibits calcium release from intracellular stores, significantly inhibited the maximal contraction to carbamylcholine (3 microM). In contrast, ryanodine (30 microM) did not inhibit the maximal contraction to 5-HT (1 microM) in the absence of nitrendipine. However, ryanodine (30 microM) did significantly inhibit the nitrendipine-insensitive 5-HT contraction, suggesting that this component of the contraction was due in part to calcium release from a ryanodine-sensitive store. Bisindolylmaleimide (5 microM), a specific inhibitor of protein kinase C (PKC), inhibited 5-HT contraction in either the absence or presence of nitrendipine, suggesting that activation of PKC is also involved.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Age-related alterations in sensitivity of muscarinic and serotonergic responses of guinea pig trachea to removal of extracellular Ca2+.

Contractile responses to carbamylcholine and 5-hydroxytryptamine (5-HT) in the presence and absence of extracellular Ca2+ were compared in tracheal rings from 3-4-week-old and 7-8-week-old guinea pigs. Contraction to carbamylcholine and 5-HT did not differ in potency or maximal response between age groups in the presence of extracellular Ca2+. However, removal of extracellular Ca2+ inhibited maximal contraction mediated by both agonists to a greater extent in younger versus older guinea pigs. These data suggest that carbamylcholine- and 5-HT-induced contraction of guinea pig trachea from younger animals is more dependent on extracellular Ca2+ as compared to mature animals, and may be related to an age-dependent maturation of sarcoplasmic reticulum function in smooth muscle.

Aging↗
PubMed1994

Effect on outcome of the presence or absence of chest pain at initiation of recombinant tissue plasminogen activator therapy in acute myocardial infarction. The Thrombolysis in Myocardial Infarction Investigators.

To ascertain whether the outcome of patients with suspected myocardial infarction differs when chest pain is still present at initiation of thrombolytic therapy, participants in the Thrombolysis in Myocardial Infarction Phase II study, all of whom presented within 4 hours of symptoms onset, were retrospectively divided into 2 groups: (1) those with chest pain present at onset of intravenous thrombolysis, n = 3,000; and (2) those who were free of chest pain on beginning intravenous thrombolytic therapy, n = 337. Patients free of chest pain were older (58 vs 57 years, p = 0.01), more often women (23 vs 17%, p = 0.01), had fewer electrocardiographic leads with ST elevation (3.8 vs 4.1, p < 0.001), and the presenting event was confirmed less often as myocardial infarction than as chest pain without infarction (88 vs 96%, p < 0.001). There were no significant differences between the 2 groups for in-hospital death, reinfarction, recurrent ischemic events, stroke, overall hemorrhagic complications, coronary angioplasty or bypass surgery. At 6-weeks follow-up, more pain-free patients had resting ejection fraction > 0.55 (35 vs 31%, p = 0.001) and fewer developed congestive heart failure (12 vs 20%). At 1-year follow-up, fewer pain-free patients developed congestive heart failure (15 vs 21%, p = 0.009), but no differences existed between the 2 groups in frequency of death, reinfarction, coronary angioplasty, bypass surgery or anginal class. Thus, there are several observations in patients who were free of chest pain at onset of lytic therapy. (1) The majority developed enzymatic or electrocardiographic evidence of acute myocardial infarction.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1994

Contractile serotonin-2A receptor signal transduction in guinea pig trachea: importance of protein kinase C and extracellular and intracellular calcium but not phosphoinositide hydrolysis.

Serotonin (5-HT) contracts the guinea pig trachea through stimulation of the 5-HT2A receptor, a receptor generally linked with phosphoinositide (PI) hydrolysis. However, previous limited evidence suggested that 5-HT did not increase PI hydrolysis in guinea pig trachea. The present studies confirmed that the 5-HT2A receptor is not coupled to PI hydrolysis and investigated the calcium source and involvement of protein kinase C (PKC) in 5-HT-induced contraction in guinea pig trachea. In vitro experiments, which used an enriched tracheal muscle preparation, confirmed the inability of 5-HT (10(-10) to 10(-2) M) to increase PI hydrolysis. Short incubations (1-60 min) of the trachea with 5-HT (10(-4) M) to minimize possible 5-HT2A receptor desensitization did not increase PI hydrolysis, whereas carbamylcholine (10(-7) to 10(-3) M) and histamine (10(-7) to 10(-4) M) did. These results demonstrate that, unlike most other 5-HT2A receptors, the 5-HT2A receptor in guinea pig trachea is not coupled to PI hydrolysis. The L-type calcium channel antagonists nitrendipine (10(-6) and 10(-5) M) and diltiazem (5 x 10(-5) M) significantly blocked maximal tracheal contraction to 5-HT (45-60%) inhibition) but not to carbamylcholine. The maximal response to 5-HT in calcium-free buffer (0 calcium, 0.05 mM EGTA) was also inhibited by 56%. The residual contraction to 5-HT in the absence of extracellular calcium suggested that at least a portion of the nitrendipine-insensitive 5-HT contraction was due to the release of intracellular calcium. In support of this idea, ryanodine (3 x 10(-5) M), a compound known to deplete intracellular calcium stores, depressed maximal 5-HT contraction in the presence of either nitrendipine or diltiazem. Neither calphostin C (4 x 10(-8) and 10(-6) M) or staurosporine (10(-8) M), both putative PKC inhibitors, affected tracheal contraction to 5-HT. However, the PKC inhibitor bisindolylmaleimide (5 x 10(-6) M), which abolished contraction to phorbol 12,13-dibutyrate (10(-6) M), unlike calphostin C, inhibited contraction to 5-HT in both the absence and presence of nitrendipine. This finding suggests that PKC activation is involved in 5-HT contraction. Thus, the tracheal 5-HT2A receptor is unique in that activation of the receptor does not result in PI hydrolysis but increases calcium influx through L-type voltage-dependent calcium channels, calcium release from the sarcoplasmic reticulum and activation of a bisindolylmaleimide-sensitive PKC.

Alkaloids↗
PubMed1993

A role for the mitochondrial Na(+)-Ca2+ exchanger in the regulation of oxidative phosphorylation in isolated heart mitochondria.

The role of the Na(+)-Ca2+ exchanger of heart mitochondria in cellular functioning is not yet clear. The objectives of this study were to investigate the effects of stimulation and inhibition of the Na(+)-Ca2+ exchanger on the matrix free Ca2+ concentration in isolated heart mitochondria and to determine the consequences of these changes on the rate of NADH production via Krebs cycle turnover and the oxidative phosphorylation rate (OPR) supported by alpha-ketoglutarate dehydrogenase, a Ca(2+)-regulated matrix enzyme. Activation of Na(+)-Ca2+ exchange by increasing extramitochondrial Na+ concentration was found to decrease the matrix free [Ca2+] in a concentration-dependent manner. Inhibitors of mitochondrial Na(+)-Ca2+ exchange activity inhibited the decrease in matrix free [Ca2+] mediated by Na+. Increasing concentrations of Na+ were also found to inhibit both the rate of NADH production and OPR. Inhibitors of mitochondrial Na(+)-Ca2+ exchange also blocked the effects of Na+ on the rate of NADH production and OPR in a similar concentration range. The results indicate that alterations in matrix free [Ca2+] induced by changes in mitochondrial Na(+)-Ca2+ exchange activity are translated into changes in the rate of NADH production and the overall rate of oxidative phosphorylation.

Animals↗
PubMed1993

Clinically guided closure of femoral arterial pseudoaneurysms complicating cardiac catheterization and coronary angioplasty.

Femoral artery pseudoaneurysm formation is a significant problem in patients undergoing cardiac catheterization and interventional cardiac procedures. It is especially more common with the use of anticoagulant and antiplatelet therapy and the use of intracoronary stents. We describe our initial experience with clinically guided bedside compression of femoral pseudoaneurysms in such patients. Eleven patients, 10 undergoing coronary angioplasty (including 3 with intracoronary stents) and 1 undergoing diagnostic cardiac catheterization, developed a femoral pseudoaneurysm. All patients had a femoral bruit and 9 had an expansile groin hematoma. The diagnosis was confirmed in each case by Doppler ultrasound. Seven patients were receiving heparin while 4 were on oral anticoagulants at the time of detection of the pseudoaneurysm. These patients underwent clinically guided graded external compression to close the pseudoaneurysm neck while maintaining femoral arterial flow. External compression for 104.1 +/- 63 min resulted in successful clinical resolution of pseudoaneurysm in all patients without complications. The results were confirmed by Doppler ultrasound at least 12 hr later. Bedside compression of femoral pseudoaneurysms guided by clinical clues is simple and appears to be an effective and safe technique to manage this iatrogenic problem.

Aged↗
PubMed1993

Modulation of intramitochondrial free Ca2+ concentration by antagonists of Na(+)-Ca2+ exchange.

Evidence has accumulated in the past decade suggesting that Ca2+ acts as a second messenger not only in the cytosol of the heart to regulate contractility, but also within the mitochondria to regulate the rate of oxidative ATP synthesis. Just as elucidation of the second messenger pathways for Ca2+ in the cytosol has led to the development of pharmacological interventions that alter mechanical functioning of the heart, understanding the role of Ca2+ as a second messenger within the mitochondria and the mechanisms by which this organelle transports and regulates Ca2+ has exciting potential for developing pharmacological interventions that alter myocardial energy metabolism. In this article, David Cox and Mohammed Matlib discuss the potential consequences of pharmacologically increasing the intramitochondrial Ca2+ concentration on myocardial energy metabolism, and suggest some pathological conditions in which such an effect may be beneficial.

Adenosine Triphosphate↗
PubMed1993

Selectivity of inhibition of Na(+)-Ca2+ exchange of heart mitochondria by benzothiazepine CGP-37157.

The objective was to determine if the benzothiazepine compound CGP-37157 selectively inhibits the Na(+)-Ca2+ exchanger of cardiac mitochondria without affecting the L-type voltage-dependent calcium channel, the Na(+)-Ca2+ exchanger, or the Na(+)-K(+)-ATPase of the cardiac sarcolemma, or the Ca(2+)-ATPase of the cardiac sarcoplasmic reticulum. Mitochondrial Na(+)-Ca2+ exchange activity was determined by monitoring intramitochondrial free [Ca2+] in isolated heart mitochondria loaded with the Ca(2+)-sensitive fluorophore fura-2. CGP-37157 inhibited the activity of mitochondrial Na(+)-Ca2+ exchange in a dose-dependent manner (IC50 0.36 microM). Calcium currents were recorded by whole-cell voltage clamp in isolated neonatal ventricular myocytes. Diltiazem was able to block the recorded current completely, thus confirming the current to be exclusively L-type. CGP-37157 had no effect on the calcium current recorded under identical conditions. CGP-37157, at concentrations < or = 10 microM, had no effect on the activities of the Na(+)-Ca2+ exchanger and Na(+)-K(+)-ATPase in isolated cardiac sarcolemmal vesicles or on activity of the Ca(2+)-ATPase in isolated cardiac sarcoplasmic reticulum vesicles. The data suggest that CGP-37157 is a potent, selective, and specific inhibitor of mitochondrial Na(+)-Ca2+ exchange at concentrations < or = 10 microM.

Animals↗
PubMed1992

Crystallization and preliminary X-ray analysis of recombinant human transforming growth factor beta 2.

Recombinant human transforming growth factor beta 2 (TGF-beta 2) was cloned and expressed in E. coli. The protein was isolated from inclusion bodies, renatured and purified to a single component as judged by reversed-phase HPLC. The recombinant TGF-beta 2 was shown to have a biological activity equal to that of native TGF-beta 2 in a fibroblast migration assay. Pure, active recombinant TGF-beta 2 has been crystallized from polyethylene glycol 400. The trigonal crystals of spacegroup P3(1)21 or P3(2)21 have unit cell dimensions of a=b=60.6 A, c=75.2 A and diffract beyond 2.0 A.

3T3 Cells↗
PubMed1992

Wound healing in aged animals--effects of locally applied transforming growth factor beta 2 in different model systems.

Local application of a growth factor which could stimulate cell turnover, extracellular matrix synthesis and blood vessel formation in the skin should improve and accelerate wound healing processes which are often impaired in old age. We demonstrate the effects of TGF-beta 2 in promoting wound repair in old animals where normal healing responses are shown to be naturally slower. The potential use of TGF-beta s for the treatment of wound injuries, including chronic non-healing ulcers in the elderly, is discussed.

Allantois↗
PubMed1991

Isolation and characterisation of milk growth factor, a transforming-growth-factor-beta 2-related polypeptide, from bovine milk.

A strategy was developed for the purification of a biologically active polypeptide growth and migration factor from skimmed bovine milk. This 25-kDa dimeric molecule, termed milk growth factor (MGF), was isolated by a method consisting of a combination of strong cation-exchange chromatography, low-pressure hydrophobic-interaction chromatography, hydrophobic-interaction HPLC and size-exclusion HPLC steps, which separated the protein according to its properties of charge, hydrophobicity and size, respectively. On average, a total purification of 10(6)-10(7)-fold and a yield of approximately 115 +/- 78 ng/MGF/milk was obtained using the method described. All purification steps were performed with novel combinations of ethanol and volatile acidic salt (ammonium acetate) solutions in order to retain biological activity of the protein. These conditions, together with the easy removal of salt by lyophilization, facilitated the detection of biological activity in fractions collected at each step of the purification by means of a sensitive in vitro fibroblast-migration assay in which the half-maximal activity was obtained at a concentration of approximately 17 +/- 4 pg/ml (i.e. approximately 1 pM) of the pure protein. Biological activity of the dimeric protein was unaffected by heat treatment or exposure to acid (pH 2.0), but was lost upon reduction to its monomeric form. Amino acid composition and sequence analyses demonstrate that MGF is related to transforming growth factor type beta 2.

Amino Acid Sequence↗
PubMed1991

Separation, purification, and sequence identification of TGF-beta 1 and TGF-beta 2 from bovine milk.

Cox and Bürk (Eur. J. Biochem., 1991) reported the partial characterization of Milk Growth Factor (MGF) which stimulated the migration of fibroblasts. We have fractionated the partially purified sample by RP-HPLC and obtained the separation of two peaks of activity. The two active components were isolated as pure MGF-a and MGF-b by RP-HPLC and preparative SDS-PAGE. The purified MGF-a, consisting of a single band by gel electrophoresis and a single peak on an HPLC reversed-phase C-4 column, has the same specific activity as TGF-beta 2 in the fibroblast migration assay. MGF-a was digested by endoprotease Asp-N and the cleaved peptides were analyzed by Edman degradation and plasma desorption mass spectrometry (PDMS). The whole sequence of MGF-a determined by automated sequenator and PDMS of S-pyridylethylated protein and selected fragments was found to be identical to that of TGF-beta 2. MGF-b protein mixture separated by SDS-PAGE was electrophoretically transferred onto a Biometra Glassybond membrane, and the blotted MGF-b protein was directly sequenced on an automated sequenator. The identified 29 amino acids sequence of MGF-b was identical to the amino-terminal sequence of TGF-beta 1. Our study demonstrates that MGF is composed of both TGF-beta 1 and TGF-beta 2. TGF-beta 2 (85%) is the predominant form.

Amino Acid Sequence↗
PubMed1991

Identification of a 17-kDa protein associated with the peripheral-type benzodiazepine receptor in vascular and other smooth muscle types.

The existence of the peripheral-type benzodiazepine receptor (PBR) in vascular smooth muscle has been demonstrated in this laboratory. The present study utilized the photoaffinity ligand [3H]PK14105 to identify the protein subunit in rat aortic and other smooth muscle types to which high affinity ligands for the PBR bind. [3H]PK14105 bound to mitochondrial fractions isolated from rat aortic smooth muscle with high affinity (Kd = 7.0 +/- 0.5 nM) and high density (Bmax = 10.1 +/- 1.5 pmol/mg protein). The rank order of potency of a series of PBR ligands displacing the binding was PK11195 approximately equal to Ro5-4864 greater than protoporphyrin IX greater than flunitrazepam greater than diazepam much greater than clonazepam. [3H]PK14105 bound with comparable affinity and density to mitochondria isolated from rat myometrium and gastric smooth muscle as well. With ultraviolet irradiation, [3H]PK14105 specifically labeled a single protein of approximately 17 kDa in all three smooth muscle types examined. This protein was identical in size to that identified by [3H]PK14105 in rat adrenal gland. In adrenal gland an additional, minor protein of approximately 43 kDa was also specifically labeled by [3H]PK14105. Utilizing a probe designed from the known nucleotide sequence of the PBR in rat adrenal gland, an mRNA transcript of approximately 0.8 kilobases in size was identified in rat aortic smooth muscle by Northern blot analysis. These data indicate that a protein subunit of approximately 17 kDa comprises, at least in part, the PBR not only in vascular smooth muscle, but also in other smooth muscle types and adrenal gland as well.

Affinity Labels↗
PubMed1990

Endothelium-dependent dilation of the coronary microvasculature is impaired in dilated cardiomyopathy.

Dilator reserve of the coronary microvasculature is diminished in patients with dilated cardiomyopathy. Although increased extravascular compressive forces, tachycardia, and increased myocardial mass can explain some impairment, recent evidence suggests the possibility of intrinsic microvascular disease. We tested the hypothesis that impairment of endothelium-dependent dilation of the microvasculature could be a contributing mechanism. We infused the endothelium-dependent dilator acetylcholine (Ach) (10(-8) to 10(-6) M) and the smooth muscle vasodilator adenosine (AD) (10(-6) to 10(-4) M) into the left anterior descending coronary artery in eight patients with dilated cardiomyopathy (mean ejection fraction, 28%) and seven controls (atypical chest pain). Small vessel resistance was assessed by measuring coronary blood flow (CBF) at constant arterial pressure with a Doppler velocity catheter (corrected for cross-sectional area by angiography). With Ach, control patients increased CBF 232 +/- 40% (mean +/- SEM), whereas CBF did not significantly change in cardiomyopathy patients (41 +/- 24%) (p less than 0.0001, control vs. cardiomyopathy). With AD, control patients increased CBF 422 +/- 56% and cardiomyopathy patients increased CBF 268 +/- 43% (p = 0.13). An index of the proportion of coronary flow reserve attributable to endothelium-dependent vasodilation was obtained by standardizing each patient's Ach dose response to his maximal AD flow response. In seven control patients receiving both Ach and AD, 56 +/- 9% of the maximal AD flow response was attained with the endothelium-dependent vasodilator Ach, whereas in seven cardiomyopathy patients receiving both Ach and AD, only 23 +/- 14% of the maximal AD response was attained (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1990

Early evidence of endothelial vasodilator dysfunction at coronary branch points.

Intracoronary acetylcholine produces endothelium-dependent dilation of normal coronary arteries and paradoxical constriction of atherosclerotic vessels. Regional differences in endothelium-dependent vasomotion, however, have not been studied in relation to the nonuniform development of atherosclerosis. We compared the vasomotor response to increasing doses of acetylcholine of angiographically smooth coronary artery segments prone to atherosclerosis (coronary branch points) with segments remote from branch points (straight segments). In patients with entirely smooth coronary arteries and a dilator response to acetylcholine (group 1, n = 7), branch points and straight segments demonstrated equal and significant dose-dependent dilation to acetylcholine (14.7 +/- 8.9% and 12.3 +/- 12.7%, respectively; p identical to NS). In patients with early atherosclerosis as manifest by luminal coronary irregularities, the lowest dose of acetylcholine (10(-8) M) produced constriction at branch points and slight dilation at straight segments (-6.3 +/- 7.4% vs. +2.2 +/- 7.3%, p less than 0.05). At higher doses of acetylcholine, both branch point and straight segments constricted, but constriction remained more pronounced at branch points. Both branch point and straight segments, however, retained the ability to dilate to the non-endothelium-dependent agent, nitroglycerin. In a third group of patients with angiographically entirely smooth coronary arteries but without dilation to acetylcholine, constriction to acetylcholine again occurred first at branch points. Thus, coronary branch points demonstrate increased sensitivity to acetylcholine-induced constriction in patients with angiographic evidence of early coronary atherosclerosis and in middle-aged patients with smooth coronary arteries. These segments, however, retain the ability to dilate to nitroglycerin. Whether this early evidence of defective endothelium-dependent vasodilation predicts the later development of occlusive atherosclerosis is not yet known.

Acetylcholine↗