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H Lamb

Publications and source records attributed to H Lamb.

At least 19 recordsLinked to original sources
PubMed2005

Mitochondrial mutation detection using enhanced multiplex denaturing high-performance liquid chromatography.

In this study, we investigated the presence of mutations within the mitochondrial genome in 40 Caucasian subjects using an enhanced multiplex denaturing high-performance liquid chromatography (DHPLC) approach. The enhanced DHPLC approach has increased sensitivity and throughput, and reduced analysis time per individual sample compared to conventional methods. This technique involved amplifying the mitochondrial genome in 18 fragments ranging in size from 300 to 2000 bp using a novel proofreading polymerase (Optimase, Transgenomic Inc., Omaha, NE) with a low misincorporation rate. Fourteen of these fragments underwent subsequent restriction digestion using a combination of five restriction enzymes to enable multiplex DHPLC analysis; the remaining four underwent conventional DHPLC. Using this complete mitochondrial genome-screening approach, we confirmed a number of previously reported mutations and additionally identified a large number of novel mutations using an enhanced DHPLC technique.

Adult↗
PubMed2003

Multi-View Active Appearance Models: application to X-ray LV angiography and cardiac MRI.

This paper describes a Multi-View Active Appearance Model (AAM) for coherent segmentation of multiple cardiac views. Cootes' AAM framework was adapted by considering shapes and intensities from multiple views, while eliminating trivial difference in object pose in different views. This way, the coherence in organ shape and intensities between different views is modeled, and utilized during image search. The method is validated in two substantially different and novel applications: segmentation of combined end-diastolic and end-systolic left ventricular X-ray angiograms, and simultaneous segmentation of a combination of four chamber, two chamber and short-axis cardiac MR views.

Algorithms↗
PubMed2003

Rapid denaturing high-performance liquid chromatography (DHPLC) for mutation scanning of the transforming growth factor beta3 gene using a novel proof-reading polymerase.

We have utilized a novel variation on the conventional denaturing high-performance liquid chromatography (DHPLC) technology, which we term rapid DHPLC, combining changes in instrumentation, cartridge technology and analysis conditions to enable significant increases in throughput to be achieved. In addition, the use of a novel proof-reading polymerase for sample amplification with a low misincorporation rate enables simplification of the DHPLC patterns and hence enhanced mutation detection recognition. This scheme for increasing DHPLC throughput has been tested by scanning the transforming growth factor (TGF) beta3 gene for the presence of mutations for which there is limited published or on-line data available regarding the presence of gene polymorphisms. TGFbeta isoforms have multiple roles in cell division, growth, proliferation, transformation and differentiation. TGFbeta3 is a TGFbeta cytokine isoform, and has an important role in embryogenesis, cell differentiation and wound healing. The TGFbeta3 gene consists of seven exons and six introns spanning 43 000 bp of the human genome on chromosome 14q23-24. The rapid DHPLC approach enabled scanning of all seven exons and part of the promoter region (1000 bp upstream from exon 1 in the 5'-flanking regions) of the TGFbeta3 gene in 95 Caucasian individuals in only 8 days, in comparison to the 17 days it would have previously taken. Mutations were clearly identified in the promoter region of the TGFbeta3 gene but were absent from the exonic regions. Understanding the genetic variations affecting the TGFbeta3 gene is important as this molecule has multiple regulatory functions on a variety of cell types.

Adult↗
PubMed2001

Expression, purification and crystallization of Aspergillus nidulans NmrA, a negative regulatory protein involved in nitrogen-metabolite repression.

The NmrA repressor protein of Aspergillus nidulans was overproduced in Escherichia coli and purified to homogeneity. Gel-exclusion chromatography showed that NmrA was monomeric in solution under the buffer conditions used. The protein was crystallized in three forms, belonging to trigonal, monoclinic and hexagonal space groups. Two of these crystal forms (A and B) diffract to high resolution and thus appear suitable for structure determination. Crystal form A belongs to space group P3((1))21, with unit-cell parameters a = b = 76.8, c = 104.9 A. Crystal form B belongs to space group C2, with unit-cell parameters a = 148.8, b = 64.3, c = 110.2 A, beta = 121.8 degrees.

Aspergillus nidulans↗
PubMed2001

Agonist-induced functional desensitization of recombinant human 5-HT2 receptors expressed in CHO-K1 cells.

The desensitization characteristics of recombinant human 5-HT(2A), 5-HT(2B), and 5-HT(2C) receptors (VSV and INI isoforms) stably expressed in CHO-K1 (Chinese hamster ovary) cells was investigated by calcium fluorimetry. Comparative desensitization characteristics of the agonists 5-HT, m-chlorophenylpiperazine (mCPP), and 2,5-dimethoxy-4-iodoamphetamine hydrobromide (DOI) were performed. Human 5-HT(2C (INI)) receptors exhibited a greater degree of desensitization to all agonists tested than edited 5-HT(2C (VSV)) receptors. A 2-hr exposure to 5-HT resulted in a significantly larger reduction in response upon re-exposure to 5-HT at 5-HT(2C (INI)) receptors, as compared to 5-HT(2C (VSV)) receptors (72% and 47% respectively, P < 0.01). Both receptor isoforms were expressed at similar densities. Human 5-HT(2B) receptors exhibited the most dramatic degree of desensitization, with prior exposure to 5-HT reducing subsequent response to 5-HT by 80%, with an extremely rapid time-course (t(1/2) < 5 min). The response at 5-HT(2A) receptors was reduced by 54%. The partial agonists mCPP and DOI also elicited desensitization, generally in line with their relative efficacies at each receptor, but exhibited more rapid kinetic profiles than 5-HT. Heterologous desensitization of an endogenously expressed G(q/11)-coupled purinergic receptor was also examined following preincubation of the cell lines with 10 microM 5-HT. Only stimulation of 5-HT(2C (VSV)) receptors resulted in a profound attenuation of subsequent ATP mediated responses. These results demonstrate differing degrees of both homologous and heterologous desensitization of 5-HT(2) receptors. Additionally, the different desensitization profiles of 5-HT(2C (INI)) and 5-HT(2C (VSV)) receptor may be due to signal transduction differences caused by RNA editing.

Animals↗
PubMed2001

Characterisation of agonist binding on human 5-HT2C receptor isoforms.

The 5-HT2C receptor is expressed in different isoforms as a result of mRNA editing. Both INI (unedited) and VSV (a fully edited version) isoforms are abundant in rat brain. The VSV isoform lacks the high affinity recognition site for 5-HT, which may be caused by low efficiency coupling to G-proteins. In this study we have investigated the pharmacology of the agonist binding site of these two isoforms of the 5-HT2C receptor. The VSV isoform was expressed in Chinese hamster ovary cells (CHO) and the INI isoform in both Chinese hamster ovary cells and human embryonic kidney cells (HEK-293). Saturation analysis using [3H]5-HT revealed high and low affinity recognition sites on the INI isoform in both cell types whilst the VSV isoform did not have the high affinity binding site for [3H]5-HT. Displacement studies were undertaken using [3H]5-HT to label the receptors. In these studies the affinity of agonists (5-HT, Ro600175 ((S)-2-(6-Chloro-5-fluoroindol-1-yl)-1-methylethylamine), MK212 (6-Chloro-2-(piperazinyl) pyrazine), mCPP (1-(m-chlorophenyl)-piperazine), TfMPP (N-(m-trifluoromethylphenyl)piperazine), DOI (1-(2,5-Dimethoxy-4-iodophenyl)-2-aminopropane), DOB (1-(4-bromo-2,5-dimethoxyphenyl)-2-aminopropane) and 8OH-DPAT (8-hydroxy-2-(di-N-propylamino)tetralin) was higher at the INI isoform, whilst antagonist affinity (ketanserin and mesulergine) did not change between the two receptor isoforms. There were no differences between the INI isoform expressed in the CHO and HEK-293. This suggests that the INI isoform of the 5-HT2C receptor is pharmacologically similar to the VSV form of the 5-HT2C receptor but that it couples more efficiently to G-proteins.

Animals↗
PubMed2001

Current and future potential roles of the platinum drugs in the treatment of ovarian cancer.

The discovery of cisplatin more than two decades ago was the most important therapeutic advance in the treatment of ovarian cancer. Today, cisplatin or carboplatin in combination with paclitaxel is the most commonly used first-line treatment for patients with advanced ovarian cancer. Although platinum drugs remain a critical component of chemotherapy in this type of cancer, cumulative toxicities can limit their use. These toxicities include nephrotoxicity, neurotoxicity and ototoxicity with cisplatin and myelosuppression with carboplatin. Although these adverse events can often be managed, the interventions themselves can complicate and add to the costs of treatment. Importantly, acquired resistance to traditional platinum drugs often develops in patients with ovarian cancer and can limit the usefulness of these drugs. Research into new platinum drugs has focused on identifying compounds with improved tolerability profiles and, importantly, those which can circumvent mechanisms of platinum resistance. New platinum drugs currently under development that are showing promise in ovarian cancer include oxaliplatin, nedaplatin, satraplatin, BBR3464 and ZD0473. If the encouraging in vitro activity shown by new compounds, such as ZD0473 and BBR3464, translates into efficacy in the clinic, they may offer an extended spectrum of activity which includes patients with ovarian cancer resistant to the classical platinum drugs.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed2001

Identification of many crystal forms of Aspergillus nidulans dehydroquinate synthase.

Extensive crystallization trials of Aspergillus nidulans dehydroquinate synthase, a potential novel target for antimicrobial drugs, in complexes with different ligands have resulted in the identification of nine crystal forms. Crystals of unliganded DHQS, binary complexes with either the substrate analogue, carbaphosphonate or the cofactor NADH, as well as the ternary DHQS-carbaphosphonate-cofactor complex, were obtained. The ternary complex crystallizes from ammonium sulfate and CoCl(2) in space group P2(1)2(1)2, with unit-cell parameters a = 133.8, b = 86.6, c = 74.9 A. The binary carbaphosphonate complex crystallizes from PEG 6000 in space group P2(1)2(1)2(1), with a = 70.0, b = 64.0, c = 197.6 A, and the binary cofactor complex crystallizes from PEG 3350 and sodium potassium tartrate in space group P2(1), with a = 83.7, b = 70.4, c = 144.3 A, beta = 89.2 degrees. DHQS in the absence of ligands crystallizes in space group P2(1), with a = 41.0, b = 68.9, c = 137.7 A, beta = 94.8 degrees. Each of these crystal forms are suitable for high-resolution structure determination. Structures of a range of DHQS-ligand complexes will be of value in the structure-based design of novel antimicrobial drugs.

Aspergillus nidulans↗
PubMed2000

Comparison of gated single-photon emission computed tomography with magnetic resonance imaging for evaluation of left ventricular function in ischemic cardiomyopathy.

To perform a head-to-head comparison between magnetic resonance imaging (MRI) and gated single-photon emission computed tomography (SPECT) for the evaluation of left ventricular (LV) function (LV ejection fraction [LVEF], LV volumes, and regional wall motion) in patients with ischemic cardiomyopathy, we studied 22 patients with chronic coronary artery disease and LV dysfunction. Multislice, multiphase echoplanar MRI was performed with Philips Gyroscan ACS-NT15. Image analysis was performed using the MASS software package to determine LV end-systolic volume, LV end-diastolic volume, and LVEF. The same parameters were calculated using quantitative gated SPECT software (QGS, Cedars-Sinai Medical Center). The different parameters were compared using linear regression, and correlation coefficients were calculated. Regional wall motion was also determined from both techniques, according to a 13-segment model and a 3-point scoring system (from 1 = normokinesia to 3 = akinesia or dyskinesia). A summed wall motion score was also calculated for MRI and gated SPECT. Good correlations were found between MRI and gated SPECT for all parameters: (1) summed wall motion scoreMRI versus summed wall motion scoreSPECT: y = 0.74x + 8.0, r = 0.88, p <0.01; (2) LV end-systolic volumeMRI versus LV end-systolic volumeSPECT: y = 0.94x - 12.3, r = 0.87, p <0.01; (3) LV end-diastolic volumeMRI versus LV end-diastolic volumeSPECT: y = 0.93x - 18.4, r = 0.84, p <0.01; and (4) LVEFMRI versus LVEFSPECT: y = 0.97x + 0.68, r = 0.90, p <0.01. For regional wall motion, an exact agreement of 83% was found, with a kappa statistic of 0.77 (95% confidence intervals 0.71 to 0.83, SE 0.03), indicating essentially excellent agreement. Thus, close and significant correlations were observed for assessment of LVEF, LV volumes, and regional wall motion by MRI and gated SPECT in patients with ischemic cardiomyopathy.

Aged↗
PubMed2000

Characterization of iodoacetate-mediated neurotoxicity in vitro using primary cultures of rat cerebellar granule cells.

The neuroprotective efficacy of antioxidant molecules against iodoacetate (IAA) neurotoxicity in rat cerebellar granule cell (CGC) cultures was investigated. Transient exposure to IAA caused a concentration-dependent decrease in cell viability (ED50 = 9.8 microM). Dizocilpine maleate (MK-801), and 1,2,3,4-tetrahydro-6-nitro-2,3-dioxobenzo[f]quinoxaline-7-sulfonamide (NBQX), failed to prevent IAA toxicity. Certain antioxidant molecules were shown to be neuroprotective against IAA when combined with MK-801 but were ineffective when administered alone. (S)-(-)-Trolox, butylated hydroxytoluene (BHT), and U-83836E exhibited EC50 values of 78, 5.9, and 0.25 microM, respectively, in the presence of 10 microM MK-801. IAA also induced an increase in intracellular oxidative stress, which was quenched by the antioxidants (in the presence of MK-801) in cultures loaded with the oxidant sensitive dye 2'7'-dichlorodihydrofluorescein diacetate (DCFH-DA).

Animals↗
PubMed1999

Functional characterization of agonists at recombinant human 5-HT2A, 5-HT2B and 5-HT2C receptors in CHO-K1 cells.

1. The goal of this study was to characterize the agonist pharmacology of human 5-HT2A, 5-HT2B and 5-HT2C (VSV) receptors expressed in CHO-K1 (Chinese hamster ovary) cells. 2. We used a fluorometric imaging plate reader (FLIPR) which allows rapid detection of rises in intracellular calcium levels upon the addition of agonists. 3. Stimulation of all three receptors by 5-HT caused a robust concentration dependent increase in intracellular calcium levels. No such effect was observed from non-transfected control CHO-K1 cells. 4. The rank order of potency of agonists at the different receptor subtypes varied. Tryptamines, BW-723C86, d-norfenfluramine, Ro 60-0175 and LSD exhibited the following rank order of potency; 5-HT2B>5-HT2C>5-HT2A. Piperazines such as m-Chlorophenylpiperazine (mCPP), ORG-12962, MK-212 and also ORG-37684 exhibited a rank order of potency of 5-HT2C>5-HT2B>5-HT2A. The phenylisopropylamines DOI and DOB had a rank order of 5-HT2A>5-HT2B>5-HT2C. 5. Many agonists tested had partial agonist actions when compared to 5-HT, and a wide range of relative efficacies were exhibited, which was cell line dependent. For example, mCPP had a relative efficacy of 65% at 5-HT2C receptors but <25% at either 5-HT2A or 5-HT2B receptors. 6. Interpretation of literature values of functional assays using different cell lines, different receptor expression levels and different receptor isoforms, is complex. Species differences and the previous use of antagonist radioligands to characterize agonist potency in binding assays emphasizes the importance of studying agonists in the same experiment using the same assay conditions and parental cell lines.

Animals↗
PubMed1999

Magnetic resonance imaging of ischemic heart disease: why cardiac magnetic resonance imaging will play a significant role in the management of patients with coronary artery disease.

The role of magnetic resonance imaging in the diagnosis of ischemic heart disease has great potential impact on patient management, because a number of aspects of ischemic heart disease can be evaluated in one imaging session. High resolution coronary magnetic resonance angiography is currently available, although several technical improvements are awaited to make the technique routinely applicable. A major advance will probably include the availability of magnetic resonance blood pool contrast agents to improve vessel visualization. Contrast media, in combination with either first pass or delayed myocardial scanning, will also play an important role in myocardial perfusion imaging. Functional magnetic resonance assessment of regional and global ventricular function is currently a well-established technique and is considered a new gold standard, which may impact on routine cardiology practice. This review summarizes some of the recent magnetic resonance developments for evaluating various aspects of ischemic heart disease, including magnetic resonance coronary angiography, flow imaging, and imaging of myocardial perfusion and function.

Blood Flow Velocity↗
PubMed1998

Dopamine and nicotinic receptor binding and the levels of dopamine and homovanillic acid in human brain related to tobacco use.

Reports of a reduction in the risk of developing Parkinson's disease and Alzheimer's disease in tobacco smokers, together with the loss of high-affinity nicotine binding in these diseases, suggest that consequences of nicotinic cholinergic transmission may be neuroprotective. Changes in brain dopaminergic parameters and nicotinic receptors in response to tobacco smoking have been assessed in this study of autopsy samples from normal elderly individuals with known smoking histories and apolipoprotein E genotype. The ratio of homovanillic acid to dopamine, an index of dopamine turnover, was reduced in elderly smokers compared with age matched non-smokers (P<0.05) in both the caudate and putamen. Dopamine levels were significantly elevated in the caudate of smokers compared with non-smokers (P<0.05). However there was no significant change in the numbers of dopamine (D1, D2 and D3) receptors or the dopamine transporter in the striatum, or for dopamine D1 and D2 receptors in the hippocampus in smokers compared with non-smokers or ex-smokers. The density of high-affinity nicotine binding was higher in smokers than non-smokers in the hippocampus, entorhinal cortex and cerebellum (elevated by 51-221%) and to a lesser extent in the striatum (25-55%). The density of high-affinity nicotine binding in ex-smokers was similar to that of the non-smokers in all the areas investigated. The differences in high-affinity nicotine binding between smokers and the non- and ex-smokers could not be explained by variation in apolipoprotein E genotype. There were no differences in alpha-bungarotoxin binding, measured in hippocampus and cerebellum, between any of the groups. These findings suggest that chronic cigarette smoking is associated with a reduction of the firing of nigrostriatal dopaminergic neurons in the absence of changes in the numbers of dopamine receptors and the dopamine transporter. Reduced dopamine turnover associated with increased numbers of high-affinity nicotine receptors is consistent with attenuated efficacy of these receptors in smokers. A decrease in striatal dopamine turnover may be a mechanism of neuroprotection in tobacco smokers that could delay basal ganglia pathology. The current findings are also important in the interpretation of measurements of nicotinic receptors and dopaminergic parameters in psychiatric conditions such as schizophrenia, in which there is a high prevalence of cigarette smoking.

Age Factors↗
PubMed1998

Apolipoprotein E and alpha-1 antichymotrypsin polymorphism genotyping in Alzheimer's disease and in dementia with Lewy bodies. Distinctions between diseases.

The possibility of gene interactions in Alzheimer's disease (AD) has been suggested by the finding of an association of the AA genotype of the alpha-1 antichymotrypsin (AACT) gene and the apolipoprotein E (apoE) epsilon 4/4 genotype in AD. We tested this possibility by genotyping a large series of clinically and neuropathologically confirmed cases of AD and a series of cases with dementia with Lewy bodies (DLB) with a matched control group for the AACT locus and apoE. ApoE genotyping showed the established finding of an increased frequency of the apoE epsilon 4 allele in AD and in DLB. The AD and DLB groups differed between each other with a higher epsilon 2 allele frequency and a reduced incidence of the epsilon 4/4 genotype in DLB. Differences in the apoE frequencies may account for some of the differences between the two diseases. No association was found for the AACT A allele in AD or DLB in the groups as a whole or when stratified with respect to apoE, with the exception of a trend showing an increased incidence of the apoE epsilon 4/4 AACT AA genotype combination in AD patients (chi 2 = 3.18, p = 0.07), although in DLB this was not apparent (chi 2 = 0.0, p = 1.0). The AACT A allele is not a major risk factor for late-onset AD or DLB.

Aged↗
PubMed1997

No association between a polymorphism in the presenilin 1 gene and dementia with Lewy bodies.

The discovery of mis-sense mutations linking the presenilin-1 (PS-1) gene on chromosome 14 to Alzheimer's disease (AD) has lead to a thorough investigation of this locus. The PS-1 gene contains a polymorphism creating two alleles. The most common allele, allele 1, has been linked with late-onset AD. Given the clinical and pathological overlaps between AD and dementia with Lewy bodies we genotyped 46 pathologically confirmed cases of dementia with Lewy bodies for the PS-1 polymorphism and compared the allelic frequencies with 87 age-matched control cases and 103 age-matched AD cases. No association between dementia with Lewy bodies and PS-1 allele 1 was found either in the group as a whole, or in the group stratified according to dosage of the epsilon4 allele of the apolipoprotein E gene. We suggest that either the presenilin polymorphism has no effect on dementia with Lewy bodies or that any linkage is precluded by another, more influential, locus.

Aged↗
PubMed1997

No association between an intronic polymorphism in the presenilin-1 gene and Alzheimer's disease.

Mutations in the presenilin-1 (PS-1) gene are believed to be responsible for the majority of familial early-onset Alzheimer's disease (AD). The finding of an intronic polymorphism in the PS-1 gene prompted an investigation into its relevance in AD. An association between homozygosity for the most common allele (allele 1) in this intronic polymorphism and late-onset AD has been shown and has been confirmed by others though some studies do not support these findings. We genotyped a large series of sporadic AD cases (n = 120) and age-matched controls (n = 108) for this intronic polymorphism. We then compared both the frequency of allele 1 and allele 1 homozygosity between the AD group as a whole and in early-onset (n = 26) and late-onset (n = 94) groups with age-matched control groups (n = 29 and n = 79, respectively). No increase in the frequency or homozygosity of allele 1 in either the AD group as a whole, or when divided into late- and early-onset cases was found. Increases in the frequency of allele 1 homozygotes and in the number of non-apolipoprotein E epsilon4 carrying allele 1 homozygotes/heterozygotes was demonstrated in the early-onset AD cases although these values did not reach significance. We conclude that there is no relationship between this intronic polymorphism in the PS-1 gene and AD in the homogenous population genotyped in this study.

Alleles↗