Search PubMed⌕ Search

Biomedical subjects

F Gannon

Publications and source records attributed to F Gannon.

At least 73 records · Page 4Linked to original sources
PubMed1995

Salmon HNF1: cDNA sequence, evolution, tissue specificity and binding to the salmon serum albumin promoter.

cDNA clones coding for the transcription factor HNF1 have been isolated from Atlantic salmon (Salmo salar L.). The 559 amino acid residue long encoded protein shows high conservation, with respect to other species, of the domains necessary for DNA-binding: the HNF1 atypical homeodomain, the POU related sequence and the dimerisation domain. Alignment with rat HNF1 protein reveals that the transcription activation domains ADI and ADIII are relatively conserved in the fish sequence whereas ADII is not. Phylogenetic analysis indicates that higher vertebrate HNF1s and the related variant HNF1s (vHNF1s) are more closely related to each other than any of them is to Salmon HNF1, suggesting that the duplication event from which HNF1 and vHNF1 genes arose occurred after the divergence of the tetrapod and teleost ancestors. Northern blot analysis show a single transcript, of about 2.6 kb, which is not exclusive to liver but is also present in intestine, kidney and spleen. Using polymerase chain reaction (PCR) we have isolated the salmon albumin gene promoter which contains, upstream of the TATA box, a potential binding site for HNF1. The salmon HNF1 protein synthesized by in vitro transcription-translation of the full-length cDNA is able to bind specifically with equivalent affinities to either the rat or salmon albumin promoter.

Amino Acid Sequence↗
PubMed1995

Thickness of patellofemoral articular cartilage as measured on MR imaging: sequence comparison of accuracy, reproducibility, and interobserver variation.

OBJECTIVE: Since the thickness of cartilage is an important indicator of the status, progression and response to therapy of articular disorders, assessment of it is desirable. This study was undertaken to assess the accuracy, precision, and reliability of magnetic resonance (MR) measurements of articular cartilage. METHODS: Fifteen cadaveric patellas were imaged in the axial plane at 1.5 T. Gradient echo and fat-suppressed FSE, T2-weighted, proton density, and T1-weighted sequences were performed. We measured each 5-mm section separately at three standardized positions, giving a total of 900 measurements. These findings were correlated with independently performed measurements of the corresponding anatomic sections. A hundred random measurements were also evaluated for reproducibility and interobserver variation. RESULTS: Although all sequences were highly accurate (range r = 0.78-0.82), the T1-weighted images were the most accurate, with a mean difference of 0.25 mm and a correlation coefficient of 0.85. All sequences were also highly reproducible (mean difference between -0.09 and 0.05 mm) with little inter-observer variation (mean difference -0.04 and 0.11 mm). In an attempt to improve the accuracy of the MR measurements further, we retrospectively evaluated all measurements with discrepancies greater than 1 mm from the specimen. All these differences were attributable to focal defects causing exaggeration of the thickness on MR imaging. CONCLUSION: MR imaging is accurate, precise, and reliable as a basis for measuring articular cartilage and may potentially be usable to monitor progression of articular disorders. Care must be taken not to overestimate the thickness of areas with surface defects.

Adult↗
PubMed1995

Evaluation of a PCR assay for detection of Mycobacterium tuberculosis in clinical specimens.

A polymerase chain reaction (PCR) assay for detection of M. tuberculosis was optimized for application to clinical specimens, which were prepared for amplification by boiling in buffer. The buffer contained a synthetic DNA fragment to determine if DNA amplification from the individual prepared specimens was subject to inhibition because of substances present in the specimen, or by the process of specimen preparation or storage. The PCR test was less sensitive than direct microscopy (75% as against 87.5%) and had a specificity of 97%. Invalid results due to inhibition of amplification occurred in 12% of specimens. Incorporation of the internal standard into the specimen preparation buffer ensures that any step in the process which inhibits DNA amplification is detected in the failure of amplification of the internal standard. The use of internal standard in this way should be considered in developing diagnostic protocols.

Base Sequence↗
PubMed1995

The sensitive detection of fluorescently labelled PCR products using an automated detection system.

The polymerase chain reaction plays a central role in many detection assays and methods to improve the sensitivity and specificity of these detection systems are constantly being explored. In this study we investigated the use of an automated laser fluorescent system (ALF) in the context of DNA-based diagnostics for pathogenic bacteria. PCR products were generated using species-specific primer sets, one of which was labelled with a 5' fluorescein. PCR products with a fluorescent label were detected on line with an ALF DNA sequencer and the sensitivity of detection was found to be comparable to that for DNA probe hybridization with a radioactive probe. The technology was successfully applied to the detection of Mycobacterium tuberculosis supplemented into sputum samples and to the detection of listeria in paraffin-embedded tissue samples.

Base Sequence↗
PubMed1995

Comparison of five cardiac markers in the detection of reperfusion after thrombolysis in acute myocardial infarction.

OBJECTIVE: To investigate and compare the clinical usefulness of serial measurements of five cardiac marker proteins, namely creatine kinase (CK), CK-MB mass, myoglobin, troponin T, and myosin light chain 1, in the early detection of reperfusion after thrombolytic treatment. METHOD: Serial blood samples were taken from 26 patients presenting with acute myocardial infarction. Concentrations of the five markers were assayed in each sample. Thrombolytic treatment was given to the patients who were divided into those who reperfused (n = 17, group A) and those who failed to reperfuse (n = 9, group B) on the basis of clinical signs and angiography within 24 h. RESULTS: The release profiles of CK, CK-MB mass, myoglobin, and troponin T for patients in group A differed from those of patients in group B. No difference was observed in the release profile of myosin light chain 1 between the two groups. The time to peak concentration of CK, CK-MB mass, myoglobin, and troponin T occurred significantly earlier in patients of group A than in those of group B, with myoglobin peaking earlier than the other markers. An index, defined as the ratio of the concentration of each marker immediately before and 2 h after the start of thrombolytic treatment, was calculated for each marker in groups A and B. The 2 h myoglobin and troponin T indices were significantly different between groups A and B. The diagnostic efficiency of the myoglobin index, however, was best at 85%. CONCLUSIONS: These studies suggest that myoglobin has greater potential than the other markers examined in the detection of reperfusion after thrombolytic treatment.

Adult↗
PubMed1995

Study of osteomyelitis: utility of combined histologic and microbiologic evaluation of percutaneous biopsy samples.

PURPOSE: To determine the utility of combined histologic and microbiologic evaluation of percutaneous biopsy samples in cases of suspected osteomyelitis. MATERIALS AND METHODS: Twenty-five patients with suspected osteomyelitis were prospectively studied over a 1-year period. With usual trephine techniques under radiologic guidance, core and aspiration biopsy samples were obtained in each patient for both histologic and microbiologic analyses. RESULTS: Sixteen biopsy specimens demonstrated histologic evidence of osteomyelitis. Of these, eight were also culture positive. Seven of the eight culture-negative, histologically positive cases were interpreted as chronic osteomyelitis. In no patient were cultures positive and histologic findings negative. Three of 19 patients with proved osteomyelitis had negative histologic and microbiologic findings. The sensitivity of culture in the diagnosis of osteomyelitis in our study was 42%; the sensitivity of both culture and histologic findings was 84%. CONCLUSION: Although the volume of a biopsy specimen is a major determinant of culture yield, a portion of a percutaneous biopsy sample should be histologically evaluated for possible osteomyelitis.

Acute Disease↗
PubMed1995

Does the use of lidocaine affect the culture of percutaneous bone biopsy specimens obtained to diagnose osteomyelitis? An in vitro and in vivo study.

OBJECTIVE: When percutaneous bone biopsy is done by radiologists, local anesthetics such as lidocaine are routinely used. Although percutaneous bone biopsy of neoplasms is well accepted, it has been suggested that this procedure not be used to diagnose osteomyelitis because of a reported bactericidal effect of lidocaine and related drugs on certain organisms. The purposes of this study were to determine if lidocaine is bactericidal in vitro and to determine if it has an effect on the culture of bacteria in specimens obtained by percutaneous bone biopsy in vivo. SUBJECTS AND METHODS: The minimal inhibitory concentration and minimal bactericidal concentration of 1% lidocaine hydrochloride (10 mg/ml preserved with methylparaben) were determined in vitro for seven bacteria known to be frequent causes of osteomyelitis by using conventional clinical microbiologic methods. Percutaneous core bone biopsy for suspected osteomyelitis was done in 28 patients: 21 with and seven without the use of lidocaine. Sites sampled included vertebrae (14); calcanei, pubis, and ischia (two each); and intervertebral disks (eight). Six of the 21 patients who had percutaneous biopsy with lidocaine also had an open surgical biopsy without lidocaine. The results of cultures of the specimens were compared. Histologic evaluation and radiographic follow-up were used to identify false-negative results. RESULTS: The minimal inhibitory and the minimal bactericidal concentrations, respectively, of lidocaine (in milligrams per milliliter) were as follows: Klebsiella pneumoniae, 5.0 and > 5.0; group B streptococci, 2.5 and 5.0; Staphylococcus aureus, > 5.0; and > 5.0; methicillin-resistant S. aureus, > 5.0 and > 5.0; Escherichia coli, 2.5 and > 5.0; Pseudomonas aeruginosa, 5.0 and 5.0; Salmonella species, 5.0 and > 5.0. We found no difference in bacterial growth and the number of false-negative results between patients who had biopsies with and those who had biopsies without lidocaine. Fifty percent of patients who had growth on cultures of specimens from percutaneous biopsies done with lidocaine had no growth on cultures of specimens from surgical biopsies done without lidocaine. This likely occurred because the surgical specimens were not obtained under cross-sectional imaging guidance. CONCLUSION: Up to a 50% mixture of lidocaine has no significant effect in vitro on the bacterial growth of the seven organisms that cause osteomyelitis most frequently, and no inhibitory effect on bacterial growth was seen in biopsies done with lidocaine in vivo. The inhibitory effect of lidocaine therefore occurs at a greater concentration than is used clinically. We conclude that lidocaine used for biopsy does not interfere with the diagnosis of osteomyelitis.

Anesthesia, Local↗
PubMed1995

A novel line of transgenic mice (RSV/LTR-bGH) expressing growth hormone in cardiac and striated muscle.

In order to further investigate the deleterious effects of GH overexpression, we generated a novel line of transgenic mice featuring stable and specific expression of bovine GH in the heart and striated muscle. A DNA construct, containing a region with promoter activity from the Long Terminal Repeat of Rous Sarcoma Virus (RSV-LTR) and the entire structural gene of bovine GH (bGH), was microinjected by standard techniques in male pronuclei of fertilized mice eggs. Transgenic mice expressed bGH mRNA in the heart and striated muscle starting at 5-6 weeks of age. They featured circulating levels of a 22 kDa form of bGH up to 700 ng/ml and enhanced growth starting at 6 weeks of age. No pathologic changes of the myocardium and striated muscle fibers, other than hypertrophy, were noticed, although severe glomerulosclerosis and liver alteration occurred in older mice. Future studies on this new line of transgenic GH mice and integration with the existing data might improve our understanding of the molecular mechanism underlying the detrimental effects of elevated GH levels on various organs and functions.

Animals↗
PubMed1994

The impact of the PCR plateau phase on quantitative PCR.

The quantitative use of the polymerase chain reaction (PCR) is often compromised by the variability of the amplification. The most useful system for quantitation by PCR involves the use of controls which are almost identical to the target and which can be amplified using the same primers as the sequences of interest. In this paper, we use a model system consisting of differently sized targets amplifiable with varying primers to demonstrate the effects of the plateau phase of PCR on quantitation by PCR. This model confirms two commonly observed results: (i) when varying amounts of a single target are amplified, a constant maximum level of product is obtained and (ii) coamplification of different concentrations of different targets results in retention of the initial proportions. The inherent contradiction in these results is examined by replacement of the key elements of the reaction including enzyme, dNTPs or primers, none of which have an effect on the plateau. Pyrophosphate is found to exert no inhibitory effect on the reaction, nor does the exonuclease action of the enzyme cause the plateau. Levels of amplification attained during amplification are both theoretically and empirically defined as being insufficient to lead to the plateau due to competition between self-annealing of product DNAs and primer binding. We conclude that, pending further biochemical enquiry into the enzyme(s) used in the PCR, none of reasons conventionally proposed for the plateau phase of the PCR are sufficient to explain the phenomenon. This being so, we define the plateau as being a feature of the reaction as a whole and, since the onset of this phase is simultaneous for all amplicons, quantitation using the internal control system need not require exponential amplification. This therefore greatly simplifies the quantitative application of PCR.

Animals↗
PubMed1994

Periarticular hyperostosis and renal disease in six black lemurs of two family groups.

Proliferative periosteal disease was identified in 6 black lemurs (Eulemur macaco macaco) of 2 family groups. Bilaterally symmetric formation of periosteal new bone at the metaphyseal regions of major long bones was first detected at the stifle and tarsal areas and was detected later at the carpal areas. Bony changes were accompanied by progressive renal disease. The syndrome progressed for 6 to 16 months before the lemurs were euthanatized because of debility. Necropsy revealed changes confined to the skeleton and kidneys. Formation of new bone was detected at all affected joints, and chronic renal disease was evident in each lemur. A specific cause was not identified. Although indistinguishable histologically from hypertrophic osteoarthropathy, several important differences were apparent. Distribution of the periosteal new bone was in the metaphyseal rather than diaphyseal areas. Thoracic or gastrointestinal lesions, typically seen with hypertrophic osteoarthropathy, were not detected, and substantial renal disease was evident. A genetic component may be involved in the development of this condition.

Animals↗
PubMed1994

Isolation and functional analysis of the promoter of the bovine serum albumin gene.

The bovine serum albumin (bSA) promoter has been cloned from bovine genomic DNA using the polymerase chain reaction. In common with other albumin promoters, this promoter functions efficiently in the differentiated rat hepatoma cell line H4II and not in the its dedifferentiated derivative, H5. Analysis of 5' deletions of the bSA promoter after transient transfection into H4II has revealed that a short construct containing the HNF1 binding site and TATA box functions efficiently but requires the presence of the more upstream sequences to achieve full activity Footprint analysis of the promoter reveals seven sites of DNA protein interaction extending from -31 to -213. One of these sites, extending from -170 to -236, whose deletion results in a four fold increase in promoter activity. This site has not previously been reported in other albumin promoters and is bound by the C/EBP-like family of proteins.

Animals↗
PubMed1994

Tail-to-tail orientation of the Atlantic salmon alpha- and beta-globin genes.

We report the cloning of a cDNA and two corresponding beta-globin genes of the Atlantic salmon (Salmo salar L.) as well as two genes for alpha-globins. Nucleotide sequence analysis of the cDNA shows that the predicted beta-globin peptide comprises 148 amino acids with a calculated molecular mass of 16,127 Da and an overall amino acid similarity of 40-50% to higher vertebrates and 60-90% to fish sequences. The study of the genomic organization of alpha- and beta-globin genes shows that, as is the case in Xenopus, the salmon genes are adjacent. Two sets of linked alpha- and beta-globin genes were isolated and restriction-enzyme polymorphisms indicate that they belong to two distinct loci, possibly as a result of the salmon tetraploidy. In each locus the alpha- and beta-globin genes are oriented 3' to 3' relative to each other with the RNA coding sequences located on opposite DNA strands. This is the first evidence for this type of arrangement found for globin genes. Moreover, while the linkage found in salmon and Xenopus supports the hypothesis of an initial tandem duplication of a globin ancestor gene, our results raise the question of the actual original orientation of the duplicated genes.

Amino Acid Sequence↗
PubMed1994

The genomic organisation, sequence and functional analysis of the 5' flanking region of the chicken estrogen receptor gene.

The cDNA of many members of the nuclear receptor superfamily has been cloned. Recently more effort has been expended on the analysis of these genes at the genomic level and on the factors controlling their expression. The genomic organization of the chicken estrogen receptor gene is presented and compared to the other members of the superfamily of hormone receptor genes with emphasis on the relationship to the functional domains. The results show that the gene is divided into eight exons and that the position of the intron/exon boundaries are as in the human gene but different to the trout estrogen receptor gene. Primer extension and cDNA clone isolation was used to determine the transcription start site and 3.0 kb of 5' flanking sequence was generated. There is striking sequence homology to the human estrogen receptor promoter and there is a well positioned "typical" TATA sequence, with potential candidate CAAT box sequences close to the start site of transcription. In transient transfection assays, subfragments of this region drove CAT expression in chicken embryo fibroblasts, and the level was increased further with the addition of forskolin, but not phorbol myristate acetate. Including sequences more distal to the cap site in promoter constructs, completely abolished the promoter activity and forskolin inducibility, indicating the presence of strong silencing activity.

Animals↗
PubMed1994

The ribosomal intergenic spacer region: a target for the PCR based diagnosis of tuberculosis.

SETTING: DNA Diagnostics, The National Diagnostics Centre and University College Hospital, Galway, Republic of Ireland. OBJECTIVE: To investigate the possibility of developing a DNA probe to distinguish the members of the Mycobacterium tuberculosis complex and develop an assay for the detection of M. tuberculosis in sputum. DESIGN: The ribosomal intergenic spacer regions from the members of the M. tuberculosis complex were sequenced and analysed for the ability of this region to provide sequence diversity for DNA probe development. RESULTS: The 100% sequence homology in this region precluded the development of a probe to distinguish the individual members of the M. tuberculosis complex. However it was possible to develop a DNA probe that could specifically detect only the members of the M. tuberculosis complex. CONCLUSION: A specific DNA probe was developed for the detection of the M. tuberculosis complex and its application to M. tuberculosis detection in sputum was demonstrated.

Base Sequence↗
PubMed1994

Cloning and sequencing of the Atlantic salmon (Salmo salar) cytochrome c oxidase subunit III gene (coxIII) and analysis of coxIII expression during parr-smolt transformation.

Smoltification is the process whereby salmon alter their metabolism in preparation for movement from freshwater to seawater. Differential screening of a cDNA library prepared from post-smolt salmon liver mRNA led to the selection of a smoltification-induced sequence. Analysis of this cDNA revealed that it partially encoded subunit III of the enzyme cytochrome c oxidase. The complete coxIII sequence was amplified from salmon genomic DNA using consensus oligonucleotides based on ATPase 6 and tRNA(GLY) sequences from Pacific salmonid species. Cytochrome c oxidase subunit III liver mRNA levels were found to be significantly increased in salmon smolts. Northern blot analysis revealed a coxIII transcript of approximately 750 bp in all salmon tissues tested except blood. The DNA sequence of coxIII employs the mammalian mitochondrial genetic code and is strongly conserved when compared with that of other species.

Amino Acid Sequence↗