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Biomedical subjects

R G Smith

Publications and source records attributed to R G Smith.

At least 19 recordsLinked to original sources
PubMed2004

Increased lipid peroxidation in sera of ALS patients: a potential biomarker of disease burden.

BACKGROUND: Markers of oxidative stress and immune activation are significantly elevated in postmortem ALS CNS tissue, although the relevance to pathogenesis is unclear. OBJECTIVE: To determine the degree and distribution of oxidative stress and immune activation in living ALS patients and whether these levels correlate with the rate of progression or extent of disease. METHOD: Serum and CSF samples from sporadic ALS (sALS) patients were assayed for 4-hydroxy-2,3-nonenal (HNE), a lipid peroxidation product, and monocyte chemoattractant protein-1alpha (MCP-1alpha), a beta-chemokine, by high-performance liquid chromatography and ELISA and compared with levels measured in disease and normal control subjects by one-way analysis of variance. SALS serum levels were analyzed in relation to rate of progression, stage of disease, and drug therapy. RESULTS: HNE levels were significantly elevated in the sera and spinal fluid of sALS patients compared with control populations and positively correlated with extent of disease but not rate of progression. MCP-1alpha levels were also elevated in the sera of sALS patients, with the exception of the neurodegenerative disease control subjects, but decreased with advancing disease. CSF MCP-1alpha levels were not different between the sampled populations. There was no correlation between serum HNE and MCP-1alpha levels in sALS patients and extent of disease. However, an inverse relationship between HNE and MCP-1alpha was demonstrable in vitro. Low levels of HNE stimulated release of MCP-1alpha from cultured human macrophages, whereas high levels inhibited release of MCP-1alpha. CONCLUSIONS: These data confirm the presence of increased oxidative stress and immune activation in ALS patients. HNE is also suggested as a possible biomarker of disease.

Adult↗
PubMed2004

Orexigenic action of peripheral ghrelin is mediated by neuropeptide Y and agouti-related protein.

Ghrelin, a stomach-derived orexigenic hormone, has stimulated great interest as a potential target for obesity control. Pharmacological evidence indicates that ghrelin's effects on food intake are mediated by neuropeptide Y (NPY) and agouti-related protein (AgRP) in the central nervous system. These include intracerebroventricular application of antibodies to neutralize NPY and AgRP, and the application of an NPY Y1 receptor antagonist, which blocks some of the orexigenic effects of ghrelin. Here we describe treatment of Agrp(-/-);Npy(-/-) and Mc3r(-/-);Mc4r(-/-) double knockout mice as well as Npy(-/-) and Agrp(-/-) single knockout mice with either ghrelin or an orally active nonpeptide ghrelin agonist. The data demonstrate that NPY and AgRP are required for the orexigenic effects of ghrelin, as well as the involvement of the melanocortin pathway in ghrelin signaling. Our results outline a functional interaction between the NPY and AgRP pathways. Although deletion of either NPY or AgRP caused only a modest or nondetectable effect, ablation of both ligands completely abolished the orexigenic action of ghrelin. Our results establish an in vivo orexigenic function for NPY and AgRP, mediating the effect of ghrelin.

Agouti-Related Protein↗
PubMed2004

Estimation of parameters for a mathematical model of growth hormone secretion.

Here, we describe partial calibration of a parsimonious mathematical model of growth hormone (GH) secretion. From first principles, we derived a model of the effects on GH secretion from pituitary somatotrophs of stimulation by GH-releasing factor (GRF) or GH secretagogue, and of inhibition by somatostatin. We obtained a concise model by collapsing the many processes of the signal transduction cascade into a single step broadly reflecting the initial binding of GRF to its receptors. In the model, GH secretion is proportional to the rate of binding of GRF to activatable receptors. Desensitization occurs because of reduction of free receptors/available effector units, and resensitization occurs as those lost are replaced. This replacement is speeded up in the presence of somatostatin, which also inhibits GH secretion by reducing the constant of proportionality between the rate of GH secretion and the rate of GRF binding. We derived simple mathematical equations for the rate of GH secretion and cumulative secretion. Using these, we tested the model against data obtained from experiments performed in vitro, and made it quantitative using rigorous statistical approaches to optimize parameter estimates. The behaviour of the calibrated model matches experimental observations closely.

Algorithms↗
PubMed2003

Effects of noise on the spike timing precision of retinal ganglion cells.

Information in a spike train is limited by variability in the spike timing. This variability is caused by noise from several sources including synapses and membrane channels; but how deleterious each noise source is and how they affect spike train coding is unknown. Combining physiology and a multicompartment model, we studied the effect of synaptic input noise and voltage-gated channel noise on spike train reliability for a mammalian ganglion cell. For tonic stimuli, the SD of the interspike intervals increased supralinearly with increasing interspike interval. When the cell was driven by current injection, voltage-gated channel noise and background synaptic noise caused fluctuations in the interspike interval of comparable amplitude. Spikes initiated on the dendrites could cause additional spike timing fluctuations. For transient stimuli, synaptic noise was dominant and spontaneous background activity strongly increased fluctuations in spike timing but decreased the latency of the first spike.

Adaptation, Physiological↗
PubMed2002

A new method of plaque scoring: a laboratory comparison with other plaque indices.

AIM: The aim of this study was to propose a simple, clear and as sensitive as possible numerical plaque index for the use in clinical oral hygiene trials known as the New Method of Plaque Scoring (NMPS), and to evaluate this index for simplicity, clarity and examiner agreement in comparison with the Distal Mesial Plaque Index, Proximal Marginal Index and a modification to the modified Navy plaque Index. MATERIAL AND METHODS: The scoring system proposed for the NMPS is based on a numerical range from 0 to 10 representing the total stained plaque on the facial or lingual tooth surfaces with the greatest weight of scores placed at the gingival and proximal regions of the surface. For scoring purposes, a horizontal boundary is imagined on the smooth facial or lingual tooth surfaces between the gingival third (A) and coronal two/thirds, the latter being subdivided vertically into thirds, mesial (B), middle (D) and distal (C). Depending on the extent of plaque coverage, whole number scores 0-3 are assigned to A, B, C and 0 or 1 for D. Ten examiners scored four selected tooth surfaces on an acrylic model according to the NMPS and the three comparative indices. The time required for each examiner to score according to each index was recorded. The scores obtained from this trial were analyzed for examiner agreement and correlation between the four indices. RESULTS: The time required for scoring according to the NMPS was found to be significantly less than the three comparative indices. The variability between examiners was found to be least for the NMPS compared to the three comparative indices. A strong correlation was found between all pairs of indices. CONCLUSION: The proposed new method of plaque scoring seems to be simpler, clearer and more reproducible than the three comparative indices. However, clinical testing is warranted.

Analysis of Variance↗
PubMed2001

Generation and refinement of peptide mimetic ligands for paratope-specific purification of monoclonal antibodies.

Paratope-specific purification of antibodies has distinct advantages over conventional methods of antibody purification with respect to its capacity to isolate product of high purity and immunoreactivity. The present report addresses the problems of identifying peptide ligands for the purification of antibodies reactive with nonprotein antigens. Using an anti-steroid antibody as the model, a lead sequence that bound antibody was identified from a peptide phage display library. The minimum binding unit in this sequence was deduced using a series of truncated peptides synthesized on the heads of polyethylene pins. Replacement Net analysis of the minimum binding unit identified peptides with increased affinity for the antibody. The affinity-matured peptide mimotope bound antibody in solution. By molecular modeling the peptide was superimposable onto estrone-3-glucuronide localized in the crystal structure of the antibody binding pocket. In order to resolve problems of presentation posed by the reversal of orientation of the peptide on the affinity matrix compared with the pins, the mimotope peptide was synthesized in reverse sequence using d-amino acids. The resulting affinity matrix was effective for the purification of antibody. Eluted product demonstrated molecular homogeneity and high immunoreactivity. It is concluded that the combination of biological and chemical library techniques described provides a method for the generation and affinity maturation of mimotopes for antibodies against nonprotein antigens.

Animals↗
PubMed2001

Heterocyclic derivatives of 2-(3,5-dimethylphenyl)tryptamine as GnRH receptor antagonists.

A series of heterocyclic 2-(3,5-dimethylphenyl)tryptamine derivatives was prepared and evaluated on a rat gonadotropin releasing hormone receptor assay. The carbon tether length and heterocyclic ring attached to the amino group of 2-(3,5-dimethylphenyl)tryptamine were varied. Several of these derivatives were potent GnRH antagonists with the most potent compound having an IC50 of 16 nM.

Animals↗
PubMed2001

2-(3,5-Dimethylphenyl)tryptamine derivatives that bind to the GnRH receptor.

A series of 2-(3,5-dimethylphenyl)tryptamine derivatives was prepared and evaluated on a rat gonadotropin releasing hormone receptor assay. Some para-substituents on the 4-phenylbutyl side chain attached to the tryptamine nitrogen led to compounds with potent GnRH receptor binding. The study has helped define structural requirements for GnRH receptor binding for the 2-aryltryptamine GnRH antagonists.

Animals↗
PubMed2001

A potent, nonpeptidyl 1H-quinolone antagonist for the gonadotropin-releasing hormone receptor.

Extensive development of the structure-activity relationships of a screening lead determined three important pharmacophores for gonadotropin-releasing hormone (GnRH) receptor antagonist activity. Incorporation of the 3,4,5-trimethylphenyl group at the 3-position, 2-(2(S)-azetidinyl)ethoxy group at the 4-position, and N-4-pyrimidinylcarboxamide at the 6-position of the quinolone core resulted in the identification of 4-(2-(azetidin-2(S)-yl)ethoxy)-7-chloro-2-oxo-3-(3,4,5-trimethylphenyl)-1,2-dihydroquinoline-6-carboxylic acid pyrimidin-4-ylamide (1) as a potent antagonist of the GnRH receptor. A 10(4)-fold increase in in vitro binding affinity is observed for the GnRH receptor as compared to the initial screening lead. Compound 1 exhibits nanomolar binding activity and functional antagonism at the human receptor and is 7-fold less active at the rhesus receptor. Intravenous administration of compound 1 to rhesus monkeys results in a significant decrease of the serum levels of downstream hormones, luteinizing hormone (79% decrease in area under the curve) and testosterone (92% decrease in area under the curve), at a dose of 3 mg/kg. Quinolone 1 is a potent nonpeptidyl antagonist for the human GnRH receptor that is efficacious for the suppression of luteinizing hormone and testosterone in primates.

Animals↗
PubMed2001

Initial structure-activity relationship of a novel class of nonpeptidyl GnRH receptor antagonists: 2-arylindoles.

A nonpeptidyl GnRH receptor antagonist (1), with a unique 2-arylindole core, was identified through the Merck in-house screening for binding affinity on the rat GnRH receptor. SAR studies directed toward the alkoxy-ethanolamine and 2-aryl groups resulted in a simpler lead structure with improved activity. This compound 50 exhibits a 60-fold improvement in binding activity over our initial lead 1.

Animals↗
PubMed2001

Bridge to transplantation with a Thoratec left ventricular assist device in a 17-kg child.

A Thoratec left ventricular assist device (LVAD) was used to support a 7-year-old 17-kg boy with viral cardiomyopathy for 23 days before heart transplantation. The boy is still living more than 1 year posttransplant, and functional except for some spastic paresis of the left hand, a residual from a stroke during device support. He is the smallest person to be supported with this device. We discuss techniques for using the Thoratec in children.

Body Weight↗
PubMed2001

Comparison of the CardioWest total artificial heart, the novacor left ventricular assist system and the thoratec ventricular assist system in bridge to transplantation.

BACKGROUND: Device selection has historically been supported by minimal comparative data. Since 1994, we have implanted 43 patients with the CardioWest Total Artificial Heart (CW), 23 with the Novacor Left Ventricular Assist System (N), and 26 with the Thoratec Ventricular Assist System (T). This experience provides a basis for our device selection criteria. METHODS: We reviewed retrospectively the results for survival, stroke, and infection in the CW, N, and T groups. Statistical methods included the Student's t-test, chi2 analysis, and Kaplan-Meier actuarial survival curves. RESULTS: The T group patients were younger and smaller sized than the CW or N group. The CW group had the highest mean central venous pressure (CVP) and lowest mean cardiac index. Survival to transplantation was 75% for CW, 57% for N, and 38% for T. Multiple organ failure postimplant caused most deaths in the CW and T groups. Right heart failure and stroke caused most N deaths. Linearized stroke rates (event/patient-month) were 0.03 for CW, 0.28 for N, and 0.08 for T. Serious infections were found in 20% of CW, 30% of N, and 8% of T patients, but linearized rates showed little difference and death from infection was rare. CONCLUSIONS: The N device should be used in "stable" patients with body surface area (BSA) greater than 1.7 m2 and with minimal right heart failure. Unstable patients with biventricular failure should receive a CW if the BSA is greater than 1.7 m2 or a T if they are smaller.

Adolescent↗
PubMed2001

Synthetic membrane neo-pericardium facilitates total artificial heart explantation.

BACKGROUND: In the past, explantation of the Cardio West total artificial heart (TAH) has been technically challenging because of the presence of dense adhesions and extremely thickened pericardium. To prevent this, we constructed a synthetic neo-pericardium in 14 patients. METHODS: Using expanded polytetrafluoroethylene (e-PTFE) membrane, we constructed a pericardium within the pericardium, or "neo-pericardium," completely covering the Cardio West TAH separating the native atria from the native pericardium, and wrapping the ascending aorta from the outflow conduit distally for about 5 to 7 cm. RESULTS: Of the 14 patients, 9 were transplanted and could be evaluated, 3 died on device support, and 2 are currently on device support. In each case, we attained faster (by 25 minutes) and easier reentry through the sternum. Surgical planes around the aorta, over the right and left atria, and throughout the pericardial space became apparent immediately after e-PTFE membrane removal. The pericardium and related tissues although slightly thickened (<2 mm) were pliable compared with our previous 36 patients, with very thick adherent pericardium over the device and native atria. CONCLUSIONS: The plastic materials forming the ventricular housing and drivelines of the Cardio West TAH and the Dacron outflow conduits have in the past caused profound local inflammatory reactions, resulting in extremely dense adhesions and thickened adherent pericardium. Using e-PTFE membrane to fashion a complete neo-pericardium and to wrap the ascending aorta at the time of Cardio West implantation dramatically reduces adhesions and pericardial thickening and facilitates explantation.

Adult↗
PubMed2001

Parvalbumin overexpression alters immune-mediated increases in intracellular calcium, and delays disease onset in a transgenic model of familial amyotrophic lateral sclerosis.

Intracellular calcium is increased in vulnerable spinal motoneurons in immune-mediated as well as transgenic models of amyotrophic lateral sclerosis (ALS). To determine whether intracellular calcium levels are influenced by the calcium-binding protein parvalbumin, we developed transgenic mice overexpressing parvalbumin in spinal motoneurons. ALS immunoglobulins increased intracellular calcium and spontaneous transmitter release at motoneuron terminals in control animals, but not in parvalbumin overexpressing transgenic mice. Parvalbumin transgenic mice interbred with mutant SOD1 (mSOD1) transgenic mice, an animal model of familial ALS, had significantly reduced motoneuron loss, and had delayed disease onset (17%) and prolonged survival (11%) when compared with mice with only the mSOD1 transgene. These results affirm the importance of the calcium binding protein parvalbumin in altering calcium homeostasis in motoneurons. The increased motoneuron parvalbumin can significantly attenuate the immune-mediated increases in calcium and to a lesser extent compensate for the mSOD1-mediated 'toxic-gain-of-function' in transgenic mice.

Age of Onset↗
PubMed2001

Regulation of Pit-1 expression by ghrelin and GHRP-6 through the GH secretagogue receptor.

GH secretagogues are an expanding class of synthetic peptide and nonpeptide molecules that stimulate the pituitary gland to secrete GH through their own specific receptor, the GH-secretagogue receptor. The cloning of the receptor for these nonclassical GH releasing molecules, together with the more recent characterization of an endogenous ligand, named ghrelin, have unambiguously demonstrated the existence of a physiological system that regulates GH secretion. Somatotroph cell-specific expression of the GH gene is dependent on a pituitary-specific transcription factor (Pit-1). This factor is transcribed in a highly restricted manner in the anterior pituitary gland. The present experiments sought to determine whether the synthetic hexapeptide GHRP-6, a reference GH secretagogue compound, as well as an endogenous ligand, ghrelin, regulate pit-1 expression. By a combination of Northern and Western blot analysis we found that GHRP-6 elicits a time- and dose-dependent activation of pit-1 expression in monolayer cultures of infant rat anterior pituitary cells. This effect was blocked by pretreatment with actinomycin D, but not by cycloheximide, suggesting that this action was due to direct transcriptional activation of pit-1. Using an established cell line (HEK293-GHS-R) that overexpresses the GH secretagogue receptor, we showed a marked stimulatory effect of GHRP-6 on the pit-1 -2,500 bp 5'-region driving luciferase expression. We truncated the responsive region to -231 bp, a sequence that contains two CREs, and found that both CREs are needed for GHRP-6-induced transcriptional activation in both HEK293-GHS-R cells and infant rat anterior pituitary primary cultures. The effect was dependent on PKC, MAPK kinase, and PKA activation. Increasing Pit-1 by coexpression of pCMV-pit-1 potentiated the GHRP-6 effect on the pit-1 promoter. Similarly, we showed that the endogenous GH secretagogue receptor ligand ghrelin exerts a similar effect on the pit-1 promoter. These data provide the first evidence that ghrelin, in addition to its previously reported GH-releasing activities, is also capable of regulating pit-1 transcription through the GH secretagogue receptor in the pituitary, thus giving new insights into the physiological role of the GH secretagogue receptor on somatotroph cell differentiation and function.

Animals↗
PubMed2001

Growth hormone secretagogue receptor family members and ligands.

We have previously reported the cloning and characterization of a new orphan G-protein-coupled receptor (GPC-R), the growth hormone secretagogue receptor (GHS-R), and shown that this receptor mediates the activity of the growth hormone-releasing peptides (GHRPs) and nonpeptide ligands such as L-692,429 and MK-0677. Because the GHS-R obviously does not belong to any of the known GPC-R subfamilies, we searched for GHS-R family members by screening a human genomic library using low-stringency hybridization and screening a Pufferfish genomic library. The Pufferfish was selected because of its compact genome. From the human genomic library, a homolog, GPR38, with 52% identity to the GHS-R was isolated. From the Pufferfish library, three family members were isolated. The Pufferfish gene having 58% identity to the GHS-R, on expression in HEK293 cells, was activated with GHRP-6 and MK-0677. These results indicate that the GHS-R has been conserved for at least 400 million years and that the Pufferfish genome is appropriate for isolation of GHS-R family members. In our search for endogenous ligands for the orphan receptors GHS-R and GPR38, we showed that adenosine is a partial agonist of the GHS-R and that motilin is the endogenous ligand for GPR38. We also confirmed that the endogenous ligand ghrelin is a full agonist of the GHS-R.

Adenosine↗