Biomedical subjects

Steven J Compton

Publications and source records attributed to Steven J Compton.

15 recordsLinked to original sources

PubMed2006

Inflammatory mediators modulate thrombin and cathepsin-G signaling in human bronchial fibroblasts by inducing expression of proteinase-activated receptor-4.

Human lung fibroblasts express proteinase-activated receptor-1 (PAR1), PAR2 and PAR3, but not PAR4. Because PAR2 has inflammatory effects on human primary bronchial fibroblasts (HPBF), we asked 1) whether the inflammatory mediators TNF-alpha and LPS could modify HPBF PAR expression and 2) whether modified PAR expression altered HPBF responsiveness to PAR agonists in terms of calcium signaling and cell growth. TNF-alpha and LPS induced PAR4 mRNA expression (RT-PCR) at 6 h and 24 h, respectively. TNF-alpha and LPS also upregulated PAR2 mRNA expression with similar kinetics but had negligible effect on PAR1 and PAR3. Flow cytometry for PAR1, PAR2, and PAR3 also demonstrated selective PAR2 upregulation in response to TNF-alpha and LPS. Intracellular calcium signaling to SLIGKV-NH2 (a selective PAR2-activating peptide; PAR2-AP) and AYPGQV-NH2 (PAR4-AP) revealed that TNF-alpha and LPS induced maximal responses to these PAR agonists at 24 h and 48 h, respectively. Upregulation of PAR2 by TNF-alpha heightened HPBF responses to trypsin, while PAR4 induction enabled cathepsin-G-mediated calcium signaling. Cathepsin-G also disarmed PAR1 and PAR2 in HPBF, while tryptase disarmed PAR2. Induction of PAR4 also enabled thrombin to elicit a calcium signal through both PAR1 and PAR4, as determined by a desensitization assay. In cell growth assays the PAR4 agonists cathepsin-G and AYPGQV-NH2 reduced HPBF cell number only in TNF-alpha-treated HPBF. Moreover, the mitogenic effect of thrombin (a PAR1/PAR4 agonist) but not the PAR1-AP TFLLR-NH2, was ablated in TNF-alpha-treated HPBF. These findings point to an important mechanism, whereby cellular responses to thrombin and cathepsin-G can be modified during an inflammatory response.

Bronchi↗

PubMed2006

Proteinase-activated receptor2 agonists upregulate granulocyte colony-stimulating factor, IL-8, and VCAM-1 expression in human bronchial fibroblasts.

Proteinase-activated receptors (PARs) are a novel family of G-protein-coupled receptors. PAR2 has been implicated in inflammatory airways disease. Although fibroblasts are pathologically important in the airways, the proinflammatory role of PAR2 in these cells remains unknown. We assessed PAR expression and functionality in human primary bronchial fibroblasts (HPBFs) before assessing PAR2-mediated HPBF proliferation, cytokine production, and adhesion molecule expression. RT-PCR and flow cytometry demonstrated that HPBFs express hPAR1, hPAR2, and hPAR3, but not hPAR4. Intracellular calcium signaling in HPBFs in response to PAR agonists showed that only hPAR1 and hPAR2 were functional receptors. We used the MTT assay to assess HPBF proliferation. Of the PAR2 agonist proteinases or selective PAR2-activating peptides (PAR2-APs) tested, none stimulated HPBF proliferation, whereas thrombin was a HPBF growth factor. mRNA for IL-8 and granulocyte colony-stimulating factor (G-CSF) was upregulated after addition of SLIGKV-NH2 when assessed by RT-PCR. No significant increase in G-CSF or IL-8 protein was detected. Trypsin stimulated IL-8 and G-CSF release from HPBF in a time- and dose-dependent manner. Leupeptin and soya trypsin inhibitor abrogated trypsin-stimulated cytokine release, indicating a requirement for trypsin's proteolytic activity. Trypsin and SLIGKV-NH2 stimulated an increase in VCAM-1 expression at 12 h after treatment, which declined thereafter. PAR2-driven upregulation of VCAM-1 cell surface expression and the release of IL-8 and G-CSF from bronchial fibroblasts may be important in promoting neutrophilic airways inflammation.

Bronchi↗

PubMed2006

Continuous template collection and updating for electrogram morphology discrimination in implantable cardioverter defibrillators.

INTRODUCTION: Electrogram morphology analysis improves discrimination of supraventricular tachycardias (SVTs) from ventricular tachycardias (VTs) in implantable cardioverter defibrillators (ICDs), but electrogram morphology may change with lead maturation, drugs, or disease progression. We report the clinical performance of an automatic algorithm that creates and updates templates from non-paced, slow rhythm and continuously checks the quality of the template used for arrhythmia discrimination. METHODS AND RESULTS: We studied this algorithm in 193 patients with single-chamber ICDs (Marquis VR, Medtronic Inc., Minneapolis, MN, USA). Of the 112 patients who completed 6-month follow-up, 99.1% of the patients had > or =1 automatic template created. Match scores between template and ongoing rhythm are computed using Haar Wavelets. Of the 435 automatic templates evaluated at follow-up, 423 (97.2%) had a median match score > or =70%. Intrinsic rhythm at 1 month had significantly higher match scores (P < 0.001) with automatic templates (90.3 +/- 7.0%) than with manual templates (85.7 +/- 10.9%) generated at pre-hospital discharge (PHD). The percentage of appropriately rejected SVTs was slightly higher with the automatic template (280/339 episodes) than with the manual template at PHD (272/339 episodes) while the Wavelet detection of VT was the same (218/220 episodes). CONCLUSIONS: In patients receiving ICDs, the automatic templates were successfully created during a 6-month follow-up period, and consistently matched the patients' intrinsic rhythm at the nominal match threshold. Both early (<1 month postimplant) and late (1- to 3-month follow-up period) changes in electrogram morphology were identified, confirming the need for automatic template updating.

Algorithms↗

PubMed2006

Reduction of RV pacing by continuous optimization of the AV interval.

BACKGROUND: In patients requiring permanent pacing, preservation of intrinsic ventricular activation is preferred whenever possible. The Search AV+ (SAV+) algorithm in Medtronic EnPulsetrade mark dual-chamber pacemakers can increase atrioventricular (AV) intervals to 320 ms in patients with intact or intermittent AV conduction. This prospective, multicenter study compared the percentage of ventricular pacing with and without AV interval extension. METHODS: Among 197 patients enrolled in the study, the percentage of ventricular-paced beats was evaluated via device diagnostics at the 1-month follow-up. Patient cohorts were defined by clinician assessment of conduction via a 1:1 AV conduction test at the 2-week follow-up. The observed percentage of ventricular pacing with SAV + ON and the predicted percentage of ventricular pacing with SAV + OFF were determined from the SAV + histogram data for the period between the 2-week and 1-month follow-up visits. RESULTS: Of 197 patients, 110 (55.8%) had intact 1:1 AV conduction, of which 109 had 1-month data. SAV + remained ON in 99/109 patients; 10 patients had intrinsic A-V conduction intervals beyond SAV + nominal and therefore SAV + disabled. The mean percentage of ventricular pacing in the 109 patients was SAV+ ON = 23.1% (median 3.7%) versus SAV + OFF = 97.2% (median 99.7%). In 87 patients without 1:1 AV conduction, SAV + was programmed OFF in 6, automatically disabled in 52, and remained ON in 29. In 8 of these patients, 80-100% reduction in ventricular pacing was observed with SAV + ON. CONCLUSION: The Search AV+ algorithm in the EnPulse pacemaker effectively promotes intrinsic ventricular activation and substantially reduces unnecessary ventricular pacing.

Aged↗

PubMed2006

Multi-center clinical experience with a lumenless, catheter-delivered, bipolar, permanent pacemaker lead: implant safety and electrical performance.

PURPOSE: Reduced lead diameter and reliability can be designed into transvenous permanent pacing leads through use of redundant insulation and removal of the stylet lumen. The model 3830 lead (Medtronic Inc., Minneapolis, MN, USA) is a bipolar, fixed-screw, steroid-eluting, lumenless, 4.1-Fr pacing lead. Implantation can be performed in a variety of right heart sites using a deflectable catheter (Model 10600, Medtronic). Lead performance and safety were studied. METHODS: Two prospective trials of 338 implanted subjects from 56 global sites were conducted. Electrical and safety data were obtained at implant, pre-discharge, and up to 18 months post-implant. Leads were implanted at traditional and alternate right heart sites. RESULTS: The study enrolled 338 subjects (204 males, 70.6 +/- 11.6 years) followed-up for a mean of 10.2 months (range, 0-21.6). Mean P-wave amplitudes ranged from 3.2 mV at 3 months to 2.9 mV at 18 months, while mean atrial pulse width thresholds at 2.5 V ranged from 0.07 ms at 3 months to 0.09 ms at 18 months. Mean R-wave amplitudes ranged from 11.3 mV to 11.1 mV and mean ventricular pulse width thresholds at 2.5 V ranged from 0.10 ms to 0.14 ms. There were 22 ventricular and 12 atrial lead complications within 3 months post-implant. Survival from lead-related complications improved to a clinically acceptable rate in the cohort of patients when revised implant techniques were employed. CONCLUSIONS: With the use of recommended implant techniques, the study results support the electrical efficacy and safety of a catheter-delivered, lumenless lead in traditional or alternate right atrium or right ventricle sites through 18 months post-implant.

Aged↗

PubMed2005

Characterization of a novel protein from Proatheris superciliaris venom: proatherocytin, a 34-kDa platelet receptor PAR1 agonist.

Many toxins from viperid venoms have been characterised as powerful activators of platelets. Here, the venom from the East African Lowland viper, Proatheris superciliaris, was investigated for its effect on platelets and the coagulation system. Whole P. superciliaris venom stimulated platelet shape change and aggregation; however, the stimulation of platelet activation was unaffected by the structurally distinct Src family kinase inhibitors PP1 and PD0173952, suggesting that platelet activation was mediated by a G protein-coupled receptor. A platelet reactive 34-kDa protein was isolated from P. superciliaris venom which we have designated proatherocytin. This protein induced Src kinase-independent aggregation of both human and mouse platelets that was inhibited by the serine protease inhibitor AEBSF. Proatherocytin did not clot bovine or human fibrinogen, degrade fibrinogen or hydrolyse the serine protease substrate benzoyl-FVR-pNA. It activated human PAR1 on stably transfected rat kidney epithelial cells, whereas no activation of the trypsin receptor PAR2 was shown. Surprisingly, Edman degradation of proatherocytin revealed sequence identity with existing disintegrin-like domains of snake venom metalloproteinases. These results suggest that proatherocytin is a highly selective PAR1 agonist that also causes mouse platelet aggregation, probably through cleavage of PAR4.

Amino Acid Sequence↗

PubMed2005

Expression and characterization of the intracellular vanilloid receptor (TRPV1) in bronchi from patients with chronic cough.

TRPV1 is a modulator of noxious stimuli known to be important in the cough reflex. We have compared the expression of TRPV1 in normal human airways and those from patients with chronic cough and found that there is up regulation in airways smooth muscle in disease. This increased expression appears to be intracellular and we have therefore examined the role of intracellular rat and human TRPV1 activity was found using intracellular calcium signalling with human intracellular TRPV1 being located in a thapsigargin insensitive compartment. Increase in TRPV1 activity may have a role in the airway hypersensitivity seen in chronic cough.

Animals↗

PubMed2005

Expression of proteinase-activated receptor 2 on human primary gastrointestinal myofibroblasts and stimulation of prostaglandin synthesis.

It is known that subepithelial myofibroblast-derived prostaglandin (PG)E2 can regulate intestinal epithelial cell functions, and that proteinase-activated receptor-2 (PAR2) is abundantly expressed in the gastrointestinal tract. Since PAR2 activation has previously been associated with stimulation of PGE2 synthesis, we hypothesized that PAR2 expressed on primary human gastrointestinal myofibroblasts regulates PGE2 synthesis via cyclooxygenase (COX)-1 and (or) COX-2, and associated PGE synthases. Primary human myofibroblasts were isolated from the resection tissue of the esophagus, small intestine, and colon. Expression of functional PAR2 was determined by RT-PCR and by calcium mobilization in Fura-2/AM-loaded cells. Trypsin and the selective PAR2-activating peptide (PAR2-AP) SLIGRL-NH2 stimulated PGE2 synthesis in a concentration-dependent manner, as measured by enzyme immunoassay. Selective COX inhibition showed PAR2-induced PGE2 synthesis to be COX-1 dependent in esophageal myofibroblasts and both COX-1 and COX-2 dependent in colonic cells, consistent with the distribution of COX-1 and COX-2 expression. Although both cytosolic and microsomal PGE synthases were expressed in cells from all tissues, microsomal PGE synthases were expressed at highest levels in the colonic myofibroblasts. Activation of PAR2 on gastrointestinal myofibroblasts stimulates PGE2 synthesis via different pathways in the colon than in the esophagus and small intestine.

Blotting, Western↗

PubMed2004

Optical calcium sensors: development of a generic method for their introduction to the cell using conjugated cell penetrating peptides.

Probes Encapsulated By Biologically Localised Embedding (PEBBLEs) are optical sensors with nanometer dimensions fabricated by microemulsion polymerisation. The most beneficial characteristic of these sensors is the protection offered by the sensor matrix which decreases interaction between the fluorophore and the cell. These sensors have been introduced to the cell by a number of methods; however this paper discusses the development of a generic method to facilitate inclusion of this type of sensor in the cell by a simple incubation step. This was achieved by covalent linkage of a synthetic Cell Penetrating Peptide (CPP) based on the Human Immuno-deficiency Virus (HIV) -1 Tat, to the external sensor matrix. Calcium sensors were used to demonstrate this approach to incorporate the sensors within the cell. Characterisation revealed the calcium sensors were approximately 30 +/- 7 nm in diameter with a slightly negative zeta potential. The sensors demonstrated a linear range of 0-50 microM with negligible interference from a range of cellular ions and protein. Leaching of entrapped dyes from the calcium sensors was determined as 3% in a 24 h period, while photobleaching of the entrapped dye was minimal over a 40 min period. The sensors ability to cross the cell membrane using the covalently attached synthetic Tat peptide is demonstrated. Cellular inclusion of the sensors occurred within a 30 min incubation period.

Animals↗

PubMed2004

Remote monitoring of implantable cardioverter defibrillators: a prospective analysis.

A prospective study evaluating the functionality and ease of use of the Medtronic CareLink Network, "CareLink," was conducted at ten investigational sites. This internet-based remote monitoring service allows clinicians to remotely manage their patients' implantable cardioverter defibrillators (ICDs) and chronic diseases. The network is comprised of a patient monitor, a secure server, and clinician and patient websites. Under clinician direction, patients interrogated their ICDs at home, and transmitted data to secure servers via a standard telephone line. Comprehensive device data and a 10-second presenting rhythm electrogram were captured by the monitor and available for access and review on the clinician website. The information could also be printed using a standard desktop computer with internet access. During this study, patients were asked to transmit device data twice, at least 7 days apart, as scheduled by the clinic. Monitor functionality was assessed, and ease of using the system components was evaluated via questionnaires completed by patients and clinicians following each data transmission and review. Fifty-nine patients (64 +/- 14 years, range 22-85 years) completed 119 transmissions with only 14 calls to the study support center. Clinician review of data transmissions revealed several clinically significant findings, including silent AF discovery, assessment of antiarrhythmic drug efficacy in a previously diagnosed AF patient, previously unobserved atrial undersensing, and ventricular tachycardia. ICD patients found the monitor easy to use. Clinicians were pleased with the performance of the network and the quality of the web-accessed data, and found it comparable to an in-office device interrogation. CareLink is a practical tool for routine device management and may allow timely identification of clinically important issues.

Adult↗

PubMed2003

A new oral therapy for long QT syndrome: long-term oral potassium improves repolarization in patients with HERG mutations.

OBJECTIVES: We sought to determine whether oral potassium supplementation safely increases serum K(+) and results in sustained improvement of repolarization parameters in long QT syndrome type 2 (LQT2) subjects. BACKGROUND: Mutations in HERG (LQT2), the gene encoding the rapid delayed rectifier K(+) current I(Kr), account for a significant proportion of congenital long QT syndrome (LQTS). The magnitude of I(Kr) is paradoxically increased by an increase in extracellular K(+). We tested the hypothesis that long-term oral potassium supplementation results in a mild, sustainable increase in serum K(+) that improves repolarization abnormalities in subjects with LQT2. METHODS: After an initial evaluation consisting of electrocardiography, electrolytes, blood urea nitrogen, and creatinine, escalating doses of potassium chloride (KCl) and spironolactone were administered to eight subjects with six distinct HERG mutations. Medications were continued for four weeks, at which time, the final evaluation was undertaken. Beta-adrenergic blocking therapy was maintained. RESULTS: The subjects ranged in age from 11 to 52 years. The average daily KCl and spironolactone dose was 3.3 +/- 1.5 mEq/kg and 3.5 +/- 1.2 mg/kg, respectively, and this regimen resulted in an increase in serum K(+) from 4.0 +/- 0.3 to 5.2 +/- 0.3 mEq/l. There were no serious complications associated with therapy. The increase in serum K(+) resulted in a decrease in the corrected QT interval from 526 +/- 94 to 423 +/- 36 ms (mean +/- SD; lead V(2)). Both QT dispersion and T-wave morphology improved in most subjects. CONCLUSIONS: Long-term oral potassium administration increases serum K(+) in patients with LQT2. This can be achieved safely and results in improvement in repolarization. Further studies are warranted to determine whether this will reduce the incidence of life-threatening events in LQTS patients.

Administration, Oral↗

PubMed2003

Colitis induced by proteinase-activated receptor-2 agonists is mediated by a neurogenic mechanism.

Proteinase-activated receptor-2 (PAR2) activation induces colonic inflammation by an unknown mechanism. We hypothesized that PAR2 agonists administered intracolonically in mice induce inflammation via a neurogenic mechanism. Pretreatment of mice with neurokinin-1 and calcitonin-gene-related peptide (CGRP) receptor antagonists or with capsaicin showed attenuated PAR2-agonist-induced colitis. Immunohistochemistry demonstrated a differential expression of a marker for the type-1 CGRP receptor during the time course of PAR2-agonist-induced colitis, further suggesting a role for CGRP. We conclude that PAR2-agonist-induced intestinal inflammation involves the release of neuropeptides, which by acting on their receptors cause inflammation. These results implicate PAR2 as an important mediator of intestinal neurogenic inflammation.

Animals↗

PubMed2002

Glycosylation of human proteinase-activated receptor-2 (hPAR2): role in cell surface expression and signalling.

We have analysed the role of N-linked glycosylation in regulating human proteinase-activated receptor-2 (hPAR(2)) expression and function. Epitope-tagged wild-type hPAR(2) (wt-hPAR(2)) or hPAR(2) that lacked glycosylation sequons (following site-directed mutagenesis) in either the N-terminus [hPAR(2)N30A (Asn(30)-->Ala)], extracellular loop 2 [ECL2; hPAR(2)N222Q (Asn(222)-->Gln) or hPAR(2)N222A (Asn(222)-->Ala)] or both (hPAR(2)N30A,N222A or hPAR(2)N30A,N222Q) were expressed in the Chinese-hamster ovary (CHO) fibroblast cell line, Pro5. Western blot analysis of wt-hPAR(2) showed mature wt-hPAR(2) to have a molecular mass of 55-100 kDa, and 33-48 kDa following N -glycosidase F deglycosylation. FACS analysis and immunocytochemistry of the wt-hPAR(2) and PAR(2) mutant cell lines revealed that removal of both glycosylation sequons decreases (50% of wt-hPAR(2)) cell surface expression. Western blot analysis indicated that both N-linked sites are glycosylated. In functional studies, hPAR(2)N30A displayed a selective and significant increase in sensitivity towards tryptase. Interestingly, hPAR(2)N222A displayed a loss in sensitivity towards all PAR(2) agonists tested. However, further analysis revealed receptor sensitivity to alanine mutations in this domain, as the more conservative substitution hPAR(2)N222Q displayed no change in response to PAR(2) agonists. hPAR(2)N30A,N222Q displayed increased sensitivity towards tryptase, but a loss in sensitivity towards trypsin and the synthetic peptide SLIGRL-NH(2), although this loss in sensitivity towards trypsin and SLIGRL-NH(2) was secondary to changes in cell-surface expression. Finally, expression of sialic-acid-deficient wt-hPAR(2) in the CHO Lec2 glycosylation-deficient mutant cell line, showed a 40 kDa loss in molecular mass, in addition to a marked and selective increase in sensitivity towards tryptase. We conclude that hPAR(2) N-linked glycosylation and sialylation regulates receptor expression and/or signalling.

Alanine↗

PubMed2002

International Union of Pharmacology. XXVIII. Proteinase-activated receptors.

Proteinase-activated receptors (PARs) represent a unique subclass of G-protein-coupled receptors of which four family members have now been cloned from a number of species. The novel mechanism of receptor activation involves the proteolytic unmasking of a cryptic N-terminal receptor sequence that, remaining tethered, binds to and triggers receptor function. In addition, short (five to six amino acids) synthetic peptides, based on the proteolytically revealed motif, can activate PARs without the unmasking of the tethered ligand. This article summarizes the experiments leading to the pharmacological characterization and cloning of the four PAR family members and provides a rationale for their designation by the acronym "PAR". The ability to distinguish among the PARs pharmacologically 1) with selective proteinase activators, 2) with receptor-selective peptide agonists, and 3) with peptide and nonpeptide antagonists is discussed, as are the molecular mechanisms of receptor activation and desensitization/internalization. Finally, the potential physiological roles of the PARs, which are widely distributed in many organs in the settings of tissue injury, repair, and remodeling, including embryogenesis and oncogenesis are discussed, and the newly appreciated roles of proteinases as signaling molecules that can act as either functional agonists or antagonists are highlighted.

Humans↗

PubMed2002

Restricted ability of human mast cell tryptase to activate proteinase-activated receptor-2 in rat aorta.

We investigated the potential of human mast cell tryptase to induce relaxation of rat aorta. Trypsin and the selective PAR2-activating peptide (PAR2-AP) SLIGRL-NH2 stimulated robust relaxation of phenylephrine-precontracted rat aortic rings. However, human lung tryptase (1-100 nM) either in the presence or absence of heparin failed to induce any significant relaxation. Notwithstanding, incubation of the aorta with tryptase (100 nM), following the addition of a peptide corresponding to the cleavage/activation sequence of rat PAR2 (rPAR2), resulted in relaxation of precontracted tissue due to the proteolytic release of the PAR2-AP SLIGRL/ from the parent peptide. Thus, tryptase was enzymatically active in the bioassay system. Preincubation of aorta with neuraminidase to remove cell-surface sialic acid unmasked the ability of tryptase to induce relaxation of the aorta, but had no effect on relaxation induced by trypsin, SLIGRL-NH2, or acetylcholine (Ach). Like trypsin and SLIGRL-NH2, the tryptase-induced relaxation was inhibited by either removal of the endothelium or pretreatment of the tissue with NG-nitro-L-arginine methyl ester (L-NAME), suggesting an endothelium-derived nitric oxide mechanism. Interestingly, tryptase in the presence of heparin failed to induce relaxation of precontracted neuraminidase-treated rat aorta. We conclude that tryptase-induced relaxation of rat aorta, most likely via PAR2, is tightly regulated by heparin and cell-surface sialic acid.

Animals↗