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Biomedical subjects

E M Lee

Publications and source records attributed to E M Lee.

At least 19 recordsLinked to original sources
PubMed2006

The mitochondrial genome of the Korean hairstreak, Coreana raphaelis (Lepidoptera: Lycaenidae).

We determined the complete nucleotide sequences of the mitochondrial genome (mitogenome) of the Korean hairstreak, Coreana raphaelis (Lepidoptera: Lycaenidae). The entire mitochondrial DNA (mtDNA) molecule was 15,314 bp long. The C. raphaelis genes were in the same order and orientation as the completely sequenced mitogenomes of other lepidopteran species, except for the presence of an extra copy of tRNA(Ser)(AGN). High similarity in primary sequence and secondary structure between the two tandemly located copies of the tRNA(Ser)(AGN) suggest a recent duplication of an original single tRNA(Ser)(AGN). The DHU arm of the two copies of tRNA(Ser)(AGN) formed a simple loop as seen in many other metazoan mt tRNA(Ser)(AGN). The putative initiation codon for the C. raphaelis COI gene appears to be a tetranucleotide, TTAG, found commonly in the sequenced lepidopterans. ATPase8, ATPase6, ND4L and ND6 genes, which are next to another protein-coding gene at their 3' end all had the sequences potential to form a hairpin structure, suggesting the importance of such a structure for precise cleavage of the mature protein-coding genes.

AT Rich Sequence↗
PubMed2005

Investigation of occupational radiation exposures to NORM at an Irish peat-fired power station and potential use of peat fly ash by the construction industry.

Annually, approximately 15% of Ireland's electricity requirement is provided through the combustion of 3 x 10(6) tonnes of peat. While the literature on coal-fired power generation is quite abundant, studies on the peat-fired power generation industry from a radiological point of view are scarce. A study of the largest Irish peat-fired power plant was initiated to review the potential occupational radiation exposures arising from the occurrence of naturally occurring radioactive material (NORM) at different stages of the industrial process and to investigate any radiological health consequences that may arise should peat fly ash be used as a component of building materials. Ambient gamma dose rate measurements, radon measurements, quantification of the occupational exposure from inhalation of airborne particles (personal air sampling) and gamma spectrometry analysis of peat, peat ash and effluent samples from the ash ponds were undertaken. The results indicate that the radiation dose received by any worker involved in the processing of the peat and the handling of the ash resulting from peat combustion does not exceed 150 microSv per annum. Regulatory control of the peat-fired power generation is therefore unnecessary according to the Irish legislation with regards to NORM. The potential use of peat fly ash as a by-product in the building industry was also found to have a negligible radiological impact for construction workers and for members of the public.

Air Pollutants, Radioactive↗
PubMed2005

The prevalence of abnormal cervical cytology in a sexually transmitted diseases clinic.

Women seeking sexually transmitted disease (STD) services are at high risk of human papillomavirus infections. Cervical cytological screening with Papanicolau staining (Pap smear) is not consistently offered at public STD clinics. We reviewed Pap smear results on a series of 1000 female STD clinic attendees, abstracted demographics, risk behaviours and STD diagnosis from the clinical record and tested for associations with abnormal Pap smear. In all, 5.7% of the satisfactory specimens (56/993) were abnormal; increasing age category, genital warts, and chlamydia infections were independently associated with an abnormal Pap smear in multivariate analysis. Routine Pap smear screening provided satisfactory results in the STD clinic and, where population-based programmes are not available, should be fully integrated into public STD care, (particularly in settings serving younger women).

Adult↗
PubMed2004

Natural radioactivity in building materials in the Republic of Ireland.

This paper presents the findings of a study undertaken to determine the natural radioactivity present in building materials in the Republic of Ireland. A total of 70 samples of commonly used building materials were collected from various manufacturers and suppliers throughout the country and analyzed using a HPGe gamma ray spectrometer. The specific activities of 226Ra, 232Th, and 40K were measured in all samples, and results ranged from minimum values of <1, <1, and 4 Bq kg(-1) to maximum values of 139, 57, and 1,977 Bq kg(-1), with mean values of 32, 18, and 353 Bq kg(-1), respectively. Results obtained were compared with relevant national, EU, and international legislation and guidance, and with the results of similar studies undertaken in other countries. These activities would suggest that the use of such building materials in the construction of domestic dwellings or workplaces in Ireland is unlikely to give rise to any significant radiation exposure to the occupants. In general, these activities were comparable to the results of similar studies undertaken in other countries. 137Cs activity was also detected in some samples.

Air Pollution, Indoor↗
PubMed2003

Liposome-complexed adenoviral gene transfer in cancer cells expressing various levels of coxsackievirus and adenovirus receptor.

PURPOSE: Loss of coxsackievirus and adenovirus receptor (CAR) is frequently observed in malignant cancer, hampering adenoviral gene therapy approaches. Complexing adenovirus with cationic liposomes can increase adenoviral transgene expression, particularly in cells with CAR-deficiency. We investigated whether other factors such as lipid composition might be involved in determining the efficiency of liposome-complexed adenoviral gene transfer in cancer cells. MATERIAL AND METHODS: Human cancer cell lines with different expression levels of CAR were infected with a GFP transgene. The efficiency of transgene expression was assessed by determining GFP expression using FACS analysis. RESULTS: The efficiency of liposome-complexed adenoviral gene transfer was dependent on the lipid composition constituting liposomes. Polyethylene glycol (PEG)-containing liposomes were most effective in increasing liposome-complexed adenoviral gene transfer. In CAR-deficient cells, use of PEG-containing liposomes enhanced adenoviral gene transfer, whereas in CAR-expressing cells enhancement varied depending on cell type. In some CAR-expressing cells, the effect of liposome complexing was even comparable to that in CAR-deficient cells. Increased adenoviral transgene expression following complexing with PEG-containing liposomes correlated with liposome uptake in cancer cells. CONCLUSIONS: Liposome-complexed adenoviral gene transfer appears to depend on lipid composition and the level of liposome uptake by cancer cells, in addition to CAR levels. Our study suggest that these multiple factors should be considered in designing liposome-complexed adenoviral vectors to improve outcomes of current adenoviral cancer gene therapies.

Adenoviridae↗
PubMed2003

Effects of pathophysiological concentrations of albumin on NHE3 activity and cell proliferation in primary cultures of human proximal tubule cells.

The progression of renal disease correlates strongly with hypertension and the degree of proteinuria, suggesting a link between excessive Na+ reabsorption and exposure of the proximal tubule to protein. The present study investigated the effects of albumin on cell growth and Na+ uptake in primary cultures of human proximal tubule cells (PTC). Albumin (1.0 mg/ml) increased cell proliferation to 134.1 +/- 11.8% (P < 0.001) of control levels with no change in levels of apoptosis. Exposure to 0.1 and 1.0 mg/ml albumin increased total 22Na+ uptake to 119.1 +/- 6.3% (P = 0.005) and 115.6 +/- 5.3% (P < 0.006) of control levels, respectively, because of an increase in Na+/H+ exchanger isoform 3 (NHE3) activity. This was associated with an increase in NHE3 mRNA to 161.1 +/- 15.1% (P < 0.005) of control levels in response to 0.1 mg/ml albumin. Using confocal microscopy with a novel antibody raised against the predicted extracellular NH2 terminus of human NHE3, we observed in nonpermeabilized cells that exposure of PTC to albumin (0.1 and 1.0 mg/ml) increased NHE3 at the cell surface to 115.4 +/- 2.7% (P < 0.0005) and 122.4 +/- 3.7% (P < 0.0001) of control levels, respectively. This effect was paralleled by significant increases in NHE3 in the subplasmalemmal region as measured in permeabilized cells. These albumin-induced increases in expression and activity of NHE3 in PTC suggest a possible mechanism for Na+ retention in response to proteinuria.

Albumins↗
PubMed2001

RANTES and MIP-1beta mRNA expression in human peripheral blood mononuclear cells: transcript quantification using NASBA technology.

The importance of chemokines in the immune response, as well as in a range of specific disease states, is becoming increasingly apparent. The role of CC- (or beta-) chemokines and their receptors in the pathology and mechanisms of HIV-1 infection has served to intensify interest in these factors. Although the functionality of these factors resides in their protein forms, assays for the detection and quantification of these protein factors in clinical samples are not readily available. Consequently, we designed NASBA-based assays for the quantification of the mRNA encoding two members of the CC-chemokine family: RANTES and MIP-1beta. The NASBA-based assays are extremely sensitive, accurate, and reproducible across a dynamic range of at least four orders of magnitude. Inter-assay performance is comparable to intra-assay performance. We applied these methods to the analysis of normal human PBMC and PBMC from HIV-1 infected individuals. Although MIP-1beta mRNA levels are higher than RANTES levels in both populations, RANTES levels in HIV-1+ patients are higher than in normal individuals. The utility of these assays in longitudinal studies of specific subpopulations of cells, as well as their potential use in clinical diagnostics, is discussed.

Blood Circulation↗
PubMed2001

Macrophage-derived chemokine gene expression in human and macaque cells: mRNA quantification using NASBA technology.

Macrophage-derived chemokine (MDC) is a CC-chemokine that inhibits infection by both macrophage- and T cell-tropic strains of HIV-1. This suppressor activity has led to great interest in fully characterizing the role of MDC in the pathogenesis of HIV-1 infection. Methods for the quantitation of constitutive levels of MDC protein in vivo are lacking. In this report, we describe the development and performance of a NASBA-based assay for the quantification of MDC mRNA expression in human and macaque cells. Although the constitutive in vivo levels of MDC mRNA in macaque and human T lymphocytes were low, in vitro activation of these cells greatly increased MDC transcription. Levels in the human and macaque cells were comparable under all conditions tested. Positive correlations between MDC transcription and protein expression were observed. The results indicate that this assay is extremely sensitive and reproducible over a five log dynamic range, and effectively quantifies MDC mRNA in resting and activated T cells. This assay may therefore permit characterization of the role of MDC in HIV-1/SIV pathogenesis, and in vaccine-induced immune responses.

Animals↗
PubMed2001

Polymorphisms in the TNF-alpha and TNF-receptor genes in patients with coronary artery disease.

BACKGROUND: Coronary artery disease (CAD) is both multifactorial and polygenic in nature. Atheroma formation, the pathological hallmark of CAD, is an inflammatory process, with pro-inflammatory cytokines, such as tumour necrosis factor-alpha (TNF-alpha), having a major role in its pathogenesis. We have therefore investigated whether polymorphisms in the TNF-alpha (- 238 and - 308), TNF receptor 1 (position - 609 and + 10, intron 6) and TNF receptor 2 (position + 422, codon 198) genes show an association with CAD. MATERIALS AND METHODS: Patients with angiographically proven single vessel (n = 58) and multivessel (n = 122) CAD were compared to patients with angiographically proven normal coronary arteries (n = 79) and volunteers without clinical evidence of CAD (n = 250). Genotyping was performed by PCR-RFLP analysis. For the TNF-alpha polymorphisms, a meta-analysis of all published studies was also undertaken. RESULTS: No significant differences in allele or genotype frequencies were found between the normal coronary artery group or healthy volunteers and patients with CAD for any of the polymorphisms. There was also no difference in allele frequency between patients with single- and multivessel disease. For the - 308 and - 238 TNRalpha gene polymorphisms, a meta-analysis of our data and previously published studies failed to demonstrate any significant association with CAD. CONCLUSIONS: Polymorphisms in the TNF-alpha promoter region and TNF-receptor genes are not associated with the development of CAD.

Adult↗
PubMed2001

Chemokine receptor CCR5 Delta 32 genetic analysis using multiple specimen types and the NucliSens Basic Kit.

Resistance to HIV-1 infection and delayed disease progression have been associated with a 32-bp deletion (Delta32) in the gene encoding the CCR5 chemokine receptor. In the present study we describe the modification of a nucleic acid sequence-based amplification (NASBA)-based CCR5 genotyping assay for a NucliSens Basic Kit (Organon Teknika, Durham, N.C.) format using a new target-specific sandwich oligonucleotide detection methodology. The new method permitted the use of generic electrochemiluminescent probes supplied in the NucliSens Basic Kit, whereas the original NASBA method required expensive target-specific ruthenium detection probes. The Basic Kit CCR5 Delta32 genotypic analysis was in 100% concordance with both the original NASBA assay and DNA PCR results. This study also evaluated the use of multiple specimen types, including peripheral blood mononuclear cells (PBMC), whole blood, dried blood spots, buccal scrapings, and plasma, for CCR5 genotype analysis. The sensitivities of the three assays were comparable when PBMC or whole blood was the specimen source. In contrast, when dried blood spots, buccal scrapings, or plasma was used as the sample source, the sensitivity of DNA PCR was 80.95, 42.8, or 0%, respectively, compared to 100% sensitivity obtained with the original NASBA and Basic Kit NASBA assays. Our study indicates that the NucliSens Basic Kit NASBA assay is very sensitive and specific for CCR5 Delta32 genotyping using multiple sample types.

Base Sequence↗
PubMed2001

Detection of dengue viral RNA using a nucleic acid sequence-based amplification assay.

Faster techniques are needed for the early diagnosis of dengue fever and dengue hemorrhagic fever during the acute viremic phase of infection. An isothermal nucleic acid sequence-based amplification (NASBA) assay was optimized to amplify viral RNA of all four dengue virus serotypes by a set of universal primers and to type the amplified products by serotype-specific capture probes. The NASBA assay involved the use of silica to extract viral nucleic acid, which was amplified without thermocycling. The amplified product was detected by a probe-hybridization method that utilized electrochemiluminescence. Using normal human plasma spiked with dengue viruses, the NASBA assay had a detection threshold of 1 to 10 PFU/ml. The sensitivity and specificity of the assay were determined by testing 67 dengue virus-positive and 21 dengue virus-negative human serum or plasma samples. The "gold standard" used for comparison and evaluation was the mosquito C6/36 cell culture assay followed by an immunofluorescent assay. Viral infectivity titers in test samples were also determined by a direct plaque assay in Vero cells. The NASBA assay was able to detect dengue viral RNA in the clinical samples at plaque titers below 25 PFU/ml (the detection limit of the plaque assay). Of the 67 samples found positive by the C6/36 assay, 66 were found positive by the NASBA assay, for a sensitivity of 98.5%. The NASBA assay had a specificity of 100% based on the negative test results for the 21 normal human serum or plasma samples. These results indicate that the NASBA assay is a promising assay for the early diagnosis of dengue infections.

Animals↗
PubMed2000

The distribution of P2X receptor clusters on individual neurons in sympathetic ganglia and their redistribution on agonist activation.

The distribution of P2X receptors on neurons in rat superior cervical ganglia and lability of P2X receptors on exposure to agonists were determined. Antibody labeling of each P2X subtype P2X(1)-P2X(7) showed neurons isolated into culture possessed primarily P2X(2) subunits with others occurring in order P2X(7) > P2X(6) > P2X(3) > P2X(1) > P2X(5) > P2X(4). Application of ATP and alpha,beta-meATP to neurons showed they possessed a predominantly nondesensitizing P2X receptor type insensitive to alpha,beta-meATP, consistent with immunohistochemical observations. P2X(1)-green fluorescent protein (GFP) was used to study the time course of P2X(1) receptor clustering in plasma membranes of neurons and internalization of receptors following prolonged exposure to ATP. At 12-24 h after adenoviral infection, P2X(1)-GFP formed clusters about 1 microm diameter in the neuron membrane. Application of ATP and alpha,beta-meATP showed these neurons possessed a predominantly desensitizing P2X receptor type sensitive to alpha,beta-meATP. Infection converted the major functional P2X receptor type in the membrane to P2X(1). Exposure of infected neurons to alpha,beta-meATP for less than 60 s led to the disappearance of P2X(1)-GFP fluorescence from the cell surface that was blocked by monensin, indicating the chimera is normally endocytosed into these organelles on exposure to agonist.

Adenosine Triphosphate↗
PubMed2000

Improving the environment of mice by using synthetic gauze pads.

We conducted a study to evaluate the use of synthetic gauze pads for improving the environment of mice. To evaluate differences in clinical and pathology parameters, we used two treatment groups of mice, which were housed with or without gauze pads. The mice were assigned to the study at 5 to 7 weeks of age, and the study lasted 1 year. The mice were housed individually in stainless-steel ventilated cages with wire-mesh floors. Clinical observations, body weights, and food consumption were recorded frequently during the study. A complete necropsy, with histopathologic evaluation of tissues and collection of blood for clinical pathology, was performed at completion of the study. The mice with gauze pads preferred to rest on them. In addition, these mice showed a statistically significant reduction in food consumption, but their body weights and weight gains did not differ from those of animals without gauze pads. Synthetic gauze pads provide an improved environment for mice housed in cages with wire floors and may produce reduced food consumption. Gauze pads in the cages of mice do not seem to influence body weight gain, clinical signs, clinical pathology, or morphologic pathology.

Animal Welfare↗
PubMed1999

Peptide vaccination using nonionic block copolymers induces protective anti-viral CTL responses.

High molecular weight nonionic block copolymers have been developed as vaccine adjuvants. We employed these adjuvants in water-in-oil emulsion and multiple emulsion formulations with a synthetic peptide-based antigen vaccine to test their ability to prime anti-viral CD8(+) T cell responses. Vaccines were made using the H-2(d)-restricted immunodominant peptide from lymphocytic choriomeningitis virus (LCMV), NP118-126, and administered to BALB/c ByJ (H-2(d)) mice. Peptide-containing emulsions were able to induce NP118-126 specific CTL and IFN-gamma secreting CD8(+) T cells in the vaccinated mice and these responses were maintained for at least 90 days post immunization. At all times, the responses induced by the copolymer formulations were equal to, or better than, formulations based on incomplete Freund's adjuvant (IFA). In addition, the responses induced by prophylactic vaccination using the multiple emulsion formulation resulted in accelerated viral clearance following infection with a strain of LCMV (clone 13) that causes a persistent infection in naïve adult mice. These results indicate that peptide vaccination using a formulation based on high molecular weight nonionic block copolymer in a simple water-in-oil or a multiple emulsion format can induce virus-specific CD8(+) T cell responses and confer protection sufficient enough to prevent the establishment of a persistent infection.

Adjuvants, Immunologic↗
PubMed1999

Genotyping of the CCR5 chemokine receptor by isothermal NASBA amplification and differential probe hybridization.

The human CCR5 chemokine receptor functions as a coreceptor with CD4 for infection by macrophage-tropic isolates of human immunodeficiency virus type 1 (HIV-1). A mutated CCR5 allele which encodes a protein that does not function as a coreceptor for HIV-1 has been identified. Thus, expression of the wild-type and/or mutation allele is relevant to determining the infectability of patient peripheral blood mononuclear cells (PBMC) and affects disease progression in vivo. We developed a qualitative CCR5 genotyping assay using NASBA, an isothermal nucleic acid amplification technology. The method involves three enzymes and two oligonucleotides and targets the CCR5 mRNA, which is expressed in PBMC at a copy number higher than 2, the number of copies of DNA present encoding the gene. The single oligonucleotide set amplifies both alleles, and genotyping is achieved by separate hybridizations of wild-type- and mutation-specific probes directly to the single-stranded RNA amplification product. Assay sensitivity and specificity were demonstrated with RNAs produced in vitro from plasmid clones bearing the DNA encoding each allele. No detectable cross-reactivity between wild-type and mutation probes was found, and 50 copies of each allele were readily detectable. Analysis of patient samples found that 20% were heterozygous and 1% were homozygous for the CCR5 mutation. Thus, NASBA is a sensitive and specific means of rapidly determining CCR5 genotype and provides several technical advantages over alternative assay systems.

Alleles↗
PubMed1998

Conservative operation for infective endocarditis of the mitral valve.

BACKGROUND: Subvalvular preservation is necessary to maintain left ventricular function, but accidental retention of infected tissue could cause postoperative endocarditis. METHODS: We examined 71 consecutive patients who underwent operation for mitral endocarditis. Endocarditis was uncontrolled and active in 24 patients, partially treated (unfinished antibiotic course) in 17, and healed in 30. RESULTS: Valves were repaired in 17% versus 59% versus 63% and replaced with subvalvular preservation in 25% versus 6% versus 3% of the uncontrolled active, partially treated, and healed groups, respectively. Thirty-day mortality was 29% versus 0% versus 3.3% (p=0.003), total mortality was 46% versus 18% versus 17% (p=0.035), and complications-related mortality was 38% versus 11% versus 13% (p=0.054), respectively. There was a trend toward lower complications-related mortality with subvalvular preservation than without. Postoperative endocarditis occurred in 3 of 30 patients without and 1 of 41 patients with subvalvular preservation. CONCLUSIONS: Postoperative mortality in uncontrolled active mitral endocarditis remains high, but results are good with partially treated or healed endocarditis. Subvalvular preservation improves outcome, does not increase postoperative endocarditis rates, and should be performed whenever feasible.

Anti-Bacterial Agents↗
PubMed1998

Transcatheter closure of a residual postmyocardial infarction ventricular septal defect with the Amplatzer septal occluder.

Acute ventricular septal rupture following myocardial infarction carries a high mortality. Early surgery improves survival but long term outcome depends on residual shunting and left ventricular function. Residual shunting is common despite apparently successful closure and may require reoperation. Transcatheter closure is an established method of treating selected congenital defects but clinical experience of transcatheter closure in postinfarction ventricular septal rupture is minimal. Transcatheter closure of a residual ventricular septal defect was successfully done using a new device, the Amplatzer septal occluder, in a 50 year old Indian man who had previously undergone emergency surgical repair for postinfarction acute ventricular septal rupture. The technique is described and its potential as a treatment in postinfarction ventricular septal rupture, its possible complications, and the important aspects of case selection and device design are discussed.

Cardiac Catheterization↗