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Biomedical subjects

A M Taylor

Publications and source records attributed to A M Taylor.

At least 19 recordsLinked to original sources
PubMed2006

Muscle cell injury, haemolysis and dark urine in children with falciparum malaria in Papua New Guinea.

During a prospective study of red cell variants and severe malaria in children, a surprising observation was the occurrence of dark urine. Children were grouped according to urine findings: 22 had dark urine that contained a haem protein (Group I), 93 had urine of normal colour that contained a haem protein (Group II) and 236 had normal urine (Group III). To investigate the cause of dark urine, haemolysis and muscle cell injury were assessed. Intravascular haemolysis was greater in Group I than in Groups II and III. However, anaemia was more severe in Group III and is likely to have resulted mainly from extravascular haemolysis. Median plasma myoglobin concentrations were greater in Groups I and II than Group III (P = 0.00060). Plasma myoglobin was greater in children with cerebral malaria, hyperlactataemia and those who died but was not associated with acidosis. Urine myoglobin was greater in Group I than Groups II and III (P = 0.00054). It is likely that both haemoglobin and myoglobin contributed to dark urine. The association between muscle cell injury and coma suggests sequestration of parasitized red cells as a common underlying pathology. In malaria, hyperlactataemia may result directly from breakdown of muscle protein as well as tissue hypoxia.

Anemia, Hemolytic↗
PubMed2006

Assessment of differential branch pulmonary blood flow: a comparative study of phase contrast magnetic resonance imaging and radionuclide lung perfusion imaging.

OBJECTIVES: To test whether magnetic resonance (MR) imaging can be used to assess differential lung blood flow as accurately as isotope lung perfusion studies in patients investigated for congenital heart disease. METHODS AND RESULTS: Radionuclide lung perfusion and MR imaging were performed in 12 children with suspected unilateral branch pulmonary artery stenosis (mean age 12.1 (5.9) years, range 3.1-17.2 years). A non-breath hold, fast gradient echo phase contrast MR sequence was used to measure flow in the pulmonary trunk and one pulmonary artery to calculate differential flow. Good agreement was shown between the two imaging methods by Bland-Altman analysis. There was excellent correlation between the radionuclide and MR phase contrast calculated total lung blood flow (r = 0.98, p < 0.0001). CONCLUSION: MR phase contrast is an accurate method for measuring differential total right and left lung blood flow. If MR imaging is performed to assess the branch pulmonary arteries, differential lung blood flow can be also measured, avoiding the need for an additional radionuclide lung perfusion scan and reducing the overall radiation burden to this group of patients.

Adolescent↗
PubMed2001

Molecular analysis of single colonies reveals a diverse origin of initial clonal proliferation in B-precursor acute lymphoblastic leukemia that can precede the t(12;21) translocation.

The pathogenesis of pediatric B-precursor acute lymphoblastic leukemia is largely unknown, and even with nonrandom chromosomal translocations present, the precise order of clonal molecular events is undefined. We developed an in vitro system using cytokines interleukin (IL)-3, IL-7, IL-10, and FMS-like tyrosine kinase 3 ligand with CD40 ligand-expressing fibroblasts to obtain single blast colonies from which clonal immunoglobulin heavy chain (IgH), T-cell receptor delta gene rearrangements, and, in t(12;21)-positive cases, TEL-AML1 fusion transcripts could be simultaneously PCR amplified. The proliferation of early tumor progenitors increased subclone detection enabling us, in seven diagnostic samples, to determine the stage of differentiation at which each leukemia occurred. Four were derived from the stage before initiation of IgH rearrangement, one during recombination of variable, joining, and diversity segments of the heavy chain gene VDJ(H), and two after completion of IgH rearrangement. Furthermore, analysis of a t(12;21)-positive leukemia with unusually late onset, identified both TEL-AML1-positive and -negative colonies carrying a clonal T-cell receptor delta rearrangement, inferring the presence of clonal expansion before the occurrence of the t(12;21). In contrast, in a typical, early onset t(12;21)-positive leukemia, the t(12;21) appeared to be the first clonal event. In both leukemias, the t(12;21) occurred before recombination of variable, joining and diversity segments of the heavy chain gene VDJ.

B-Lymphocytes↗
PubMed2001

Cysteine-directed cross-linking localizes regions of the human erythrocyte anion-exchange protein (AE1) relative to the dimeric interface.

The human erythrocyte anion-exchanger isoform 1 (AE1) is a dimeric membrane protein that exchanges chloride for bicarbonate across the erythrocyte plasma membrane. Crystallographic studies suggest that the transmembrane anion channel lies at the interface between the two monomers, whereas kinetic analysis provides evidence that each monomer contains an anion channel. We have studied the structure-function relationship of residues at the dimeric interface of AE1 by cysteine-directed cross-linking. Single cysteine mutations were introduced in 16 positions of putative loop regions throughout AE1. The ability of these residues to be chemically cross-linked to their partner within the dimeric protein complex was assessed by mobility of the protein on immunoblots. Introduced cysteine residues in extracellular loops (ECs) 1-4 and intracellular loop 1 formed disulphide cross-linked dimers. Treatment with homobifunctional maleimide cross-linkers of different lengths (6, 10 and 16 A; 1 A identical with 0.1 nm) also cross-linked AE1 with introduced cysteines in EC5 and close to the start of transmembrane segment (TM) 1. On the basis of these data, tentative positional constraints of TMs 1-4 and 6 relative to the dimeric interface are proposed. Neither disulphide- nor maleimide-mediated cross-linking perturbed AE1 transport function, suggesting that loop-loop contacts across the dimeric interface are not primarily responsible for allosteric interactions between monomers within the functional dimeric protein complex.

Amino Acid Sequence↗
PubMed2001

p53 dysfunction in B-cell chronic lymphocytic leukemia: inactivation of ATM as an alternative to TP53 mutation.

The well-established association between TP53 mutations and adverse clinical outcome in a range of human cancers reflects the importance of p53 protein in regulating tumor-cell growth and survival. Although it is theoretically possible for p53 dysfunction to arise through mechanisms that do not involve TP53 mutation, such a phenomenon has not previously been demonstrated in a sporadic tumor. Here, we show that p53 dysfunction in B-cell chronic lymphocytic leukemia (CLL) can occur in the absence of TP53 mutation and that such dysfunction is associated with mutation of the gene encoding ATM, a kinase implicated in p53 activation. Forty-three patients with CLL were examined for p53 dysfunction, as detected by impaired up-regulation of p53 and of the p53-dependent protein p21(CIP1/WAF1) after exposure to ionizing radiation (IR). Thirty (70%) patients had normal p53 responses and underwent progressive IR-induced apoptosis. In 13 (30%) patients, p21 up-regulation was markedly impaired, indicating p53 dysfunction. Six (14%) of these patients with p53 dysfunction had increased baseline levels of p53, were found to have TP53 mutations, and were completely resistant to IR-induced apoptosis. In the other 7 (16%) patients with p53 dysfunction, IR-induced p53 up-regulation and apoptosis were markedly impaired, but baseline levels of p53 were not increased, and no TP53 mutations were detected. Each of these patients was found to have at least one ATM mutation, and a variable reduction in ATM protein was detected in all 4 patients examined. This is the first study to provide a direct demonstration that p53 dysfunction can arise in a sporadic tumor by a mechanism that does not involve TP53 mutation. (Blood. 2001;98:814-822)

Ataxia Telangiectasia Mutated Proteins↗
PubMed2001

Residual ataxia telangiectasia mutated protein function in cells from ataxia telangiectasia patients, with 5762ins137 and 7271T-->G mutations, showing a less severe phenotype.

We have assessed several ataxia Telangiectasia mutated (ATM)-dependent functions in cells derived from ataxia telangiectasia patients, carrying either an ATM 5762ins137 splice site or a 7271T-->G missense mutation, with a less severe phenotype compared with the classical disorder. ATM kinase in vitro, from 5762ins137 cells, showed the same specific activity as ATM in normal cells, but the protein was present at low levels. In contrast, mutant ATM kinase activity in the 7271T-->G cells was only about 6% that of the activity in normal cells, although the level of mutant protein expressed was similar to normal cells. Phosphorylation of the DNA double strand break repair proteins Nbs1 and hMre11, following DNA damage, was observed in normal and 7271T-->G cells but was almost absent in both 5762ins137 and classical ataxia telangiectasia cells. The kinetics of p53 response was intermediate between normal and classical ataxia telangiectasia cells in both the 7271T-->G and 5762ins137 cells, but interestingly, c-Jun kinase activation following DNA damage was equally deficient in cell lines derived from all the ataxia telangiectasia patients. Our results indicate that levels of ATM kinase activity, but not induction of p53 or c-Jun kinase activity, in these cells correlate with the degree of neurological disorder in the patients.

Ataxia Telangiectasia↗
PubMed2001

hMRE11: genomic structure and a null mutation identified in a transcript protected from nonsense-mediated mRNA decay.

We showed recently that mutation of the hMRE11 gene identified a new ataxia telangiectasia-like disorder (ATLD). In this report we describe the genomic organization of the hMRE11 gene and the analysis of a promoter region that appears to direct the divergent transcription of hMRE11 and the adjacent gene. The characterization of the genomic organization of the hMRE11 gene allowed us to determine the basis of an apparent null hMRE11 allele present in the mother and two patients in one of our two ATLD families. Polymorphic markers in the hMRE11 gene, including the promoter region, provided evidence that the mutated maternal allele was not deleted. An exon by exon search revealed the presence of a missense mutation in exon 15, the effect of which was to create a premature termination codon. Transcripts derived from the mutant allele were found to be subject to nonsense-mediated mRNA decay (NMD). Therefore, this allele was effectively null, because little if any mRNA from it was available for translation. The ATLD patients carrying this protein-truncating hMRE11 mutation have survived because the null allele they inherited from their mother is present with a missense mutation inherited from their father, which is expressed as normal levels of partially functional MRE11 protein. The mutation in the maternal hMRE11 allele of family 2 was also identified in a further unrelated Italian family with ATLD and also found to be subject to NMD.

Alleles↗
PubMed2001

Mutations in the sterol-sensing domain of Patched suggest a role for vesicular trafficking in Smoothened regulation.

The tumor suppressor gene patched (ptc) encodes an approximately 140 kDa polytopic transmembrane protein [1-3] [corrected] that binds members of the Hedgehog (Hh) family of signaling proteins [4-6] [corrected] and regulates the activity of Smoothened (Smo), a G protein-coupled receptor-like protein essential for Hh signal transduction [7-9] [corrected]. Ptc contains a sterol-sensing domain (SSD) [10, 11] [corrected], a motif found in proteins implicated in the intracellular trafficking of cholesterol [12] [corrected], and/or other cargoes [13-15] [corrected]. Cholesterol plays a critical role in Hedgehog (Hh) signaling by facilitating the regulated secretion and sequestration of the Hh protein [16] [corrected], to which it is covalently coupled. In addition, cholesterol synthesis inhibitors block the ability of cells to respond to Hh [18, 19] [corrected], and this finding points to an additional requirement for the lipid in regulating downstream components of the Hh signaling pathway. Although the SSD of Ptc has been linked to both the sequestration of, and the cellular response to Hh [16, 20, 21] [corrected], definitive evidence for its function has so far been lacking. Here we describe the identification and characterization of two missense mutations in the SSD of Drosophila Ptc; strikingly, while both mutations abolish Smo repression, neither affects the ability of Ptc to interact with Hh. We speculate that Ptc may control Smo activity by regulating an intracellular trafficking process dependent upon the integrity of the SSD.

Amino Acid Sequence↗
PubMed2001

Ameloblastic fibrosarcoma of the mandible: report of two cases and review of the literature.

Ameloblastic fibrosarcoma (AFS) is a rare malignant odontogenic tumour. To date, 60 well-documented cases have been published in the English-language literature. Two additional cases located in the posterior region of the mandible are reported. The relevant clinicopathological features of these cases, as well as those previously reported are discussed. Approximately two-thirds of AFS are malignant tumors de novo, with an average age of the affected patients being 22.9 years. This age is lower than the one observed in AFS developing from a pre-existent benign lesion (mean 33 years). Although regional and distant metastases are very infrequent, AFS is a locally aggressive lesion, with 23 (37%) of the reported cases having at least one recurrence; 12 patients (19.3%) died of the disease. Wide surgical excision with long-term follow-up remains the treatment of choice for this neoplasm.

Adult↗
PubMed2001

Detailed characterization of cysteine-less P-glycoprotein reveals subtle pharmacological differences in function from wild-type protein.

1. Subtle alterations in the coupling of drug binding to nucleotide hydrolysis were observed following mutation of all seven endogenous cysteine residues to serines in the human multidrug resistance transporter, P-glycoprotein. Wild-type (wt) and the mutant (cys-less) forms of P-gp were expressed in Trichoplusia ni (High Five) cells and purified by metal affinity chromatography in order to undertake functional studies. 2. No significant differences were observed in substrate ([(3)H]-azidopine) binding to wt or cys-less P-gp. Furthermore, neither the transported substrate vinblastine, nor the modulator nicardipine, differed in their respective potencies to displace [(3)H]-azidopine from the wt or cys-less P-gp. These results suggest that respective binding sites for these drugs were unaffected by the introduced cysteine to serine substitutions. 3. The Michaelis-Menten characteristics of basal ATP hydrolysis of the two isoforms of P-gp were identical. The maximal ATPase activity in the presence of vinblastine was marginally reduced whilst the K(m) was unchanged in cys-less P-gp compared to control. However, cys-less P-gp displayed lower overall maximal ATPase activity (62%), a decreased K(m) and a lower degree of stimulation (76%) in the presence of the modulator nicardipine. 4. Therefore, the serine to cysteine mutations in P-gp may suggest that vinblastine and nicardipine transduce their effects on ATP hydrolysis through distinct conformational pathways. The wt and cys-less P-gp isoforms display similarity in their fundamental kinetic properties thereby validating the use of cys-less P-gp as a template for future cysteine-directed structure/function analysis.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2001

Experience with laparoscopic ultrasonography for defining tumour resectability in carcinoma of the pancreatic head and periampullary region.

BACKGROUND: Computed tomography (CT) is currently the most widely available staging investigation for pancreatic tumours. However, the accuracy of CT for determining tumour resectability is variable and can be poor. Laparoscopic ultrasonography (LUS) is potentially a more accurate method for disease staging. The authors' experience of LUS for staging carcinoma of the pancreatic head and periampullary region is described. METHODS: Fifty-one patients with potentially resectable pancreatic tumours defined at CT underwent further investigation with LUS. Twenty-seven patients subsequently had an open laparotomy. The evaluations of tumour resectability at CT and LUS were compared with the operative findings. RESULTS: At LUS, 24 patients were considered to have resectable tumours, 21 non-resectable tumours and six patients were shown to have no pancreatic tumour mass. Twenty-two patients deemed to have a resectable tumour at LUS underwent surgery. Twenty patients were confirmed to have resectable disease and two patients had non-resectable disease. A further five patients underwent surgery. In all five the ultrasonographic diagnosis was confirmed at surgery (four patients with non-resectable disease and no pancreatic tumour in one patient). LUS prevented unnecessary extensive surgery in 53 per cent of patients. For the 22 patients who underwent surgery for potentially resectable disease, the positive predictive value of LUS for defining tumour resectability was 91 per cent. CONCLUSION: LUS is an accurate additional investigation for defining tumour resectability and directing management in patients with potentially resectable carcinoma of the pancreatic head or periampullary region.

Adult↗
PubMed2001

Chromosome instability syndromes.

The chromosome instability syndromes, ataxia telangiectasia (A-T), Fanconi anaemia (FA) and Bloom syndrome (BS) have been known for many years. More recently Nijmegen breakage syndrome (NBS) and ataxia telangiectasia-like disorder (ATLD) have been identified. A-T, ATLD and NBS form a group of disorders all of which show very similar cellular features that result from the consequences of increased sensitivity to ionizing radiation (IR). They also share some clinical features, particularly A-T and ATLD, and all show an immunodeficiency. A-T and NBS both show a predisposition to lymphoid tumours. Fanconi anaemia can be caused by mutations in eight different genes, although the majority of mutations are accounted for by FANCA and FANCC. The very rare Bloom syndrome is caused by mutation in a single gene, BLM. An important feature which all of these disorders have in common is that the genes identified are involved in aspects of recombination repair of DNA damage.

Animals↗
PubMed2001

trans-dominant inhibition of connexin-43 by mutant connexin-26: implications for dominant connexin disorders affecting epidermal differentiation.

Dominant mutations of GJB2-encoding connexin-26 (Cx26) have pleiotropic effects, causing either hearing impairment (HI) alone or in association with palmoplantar keratoderma (PPK/HI). We examined a British family with the latter phenotype and identified a new dominant GJB2 mutation predicted to eliminate the amino acid residue E42 (DeltaE42) in Cx26. To dissect the pathomechanisms that result in diverse phenotypes of dominant GJB2 mutations, we studied the effect of three Cx26 mutants (DeltaE42, D66H and R75W) identified in individuals with PPK/HI, and another (W44C) present in individuals with non-syndromic HI on gap junctional intercellular communication. We expressed mutant Cx26 alone and together with the epidermal connexins Cx26, Cx37 and Cx43 in paired Xenopus oocytes, and measured the intercellular coupling by dual voltage clamping. Homotypic expression of each connexin as well as co-expression of wild-type (wt) Cx26/wtCx43 and wtCx26/wtCx37 yielded variable, yet robust, levels of channel activity. However, all four Cx26 mutants were functionally impaired and failed to induce intercellular coupling. When co-expressed with wtCx26, all four mutants suppressed the wtCx26 channel activity consistent with a dominant inhibitory effect. However, only those Cx26 mutants associated with a skin phenotype also significantly (P<0.05) inhibited intercellular conductance of co-expressed wtCx43, indicating a direct interaction of mutant Cx26 units with wtCx43. These results demonstrate, for the first time, a trans-dominant negative effect of Cx26 mutants in vitro. Furthermore, they support a novel concept suggesting that the principal mechanism for manifestation of dominant GJB2 mutations in the skin is their dominant interference with the function of wtCx43. This assumption is further corroborated by our finding that Cx26 and Cx43 focally colocalize at gap junctional plaques in affected skin tissue of two carriers of DeltaE42.

Adolescent↗
PubMed2001

Response to radioimmunotherapy correlates with tumor pO2 measured by EPR oximetry in human tumor xenografts.

The efficacy of radiation treatment depends upon local oxygen concentration. We postulated that the variability in responsiveness of tumor xenografts to a fixed dose of radioimmunotherapy might be related to the tumor pO2 at the time that radioimmunotherapy was administered. We evaluated the growth of xenografts of CALU-3 tumors, a non-small cell lung carcinoma, in response to an 8.9-MBq dose of 131I-RS-7-anti-EGP-1 and correlated tumor growth rate with initial tumor pO2 measured by EPR oximetry. The greatest growth delay in response to radioimmunotherapy had the highest initial pO2, and the fastest-growing tumors had the lowest initial pO2. We then determined the dynamic effect of radioimmunotherapy on tumor pO2 by serial measurements of pO2 for 35 days after radioimmunotherapy. This information could be important for ascertaining the likelihood that a tumor will respond to additional doses as part of a multiple dose scheme. Serial tumor pO2 measurements may help identify a window of opportunity when the surviving tumor regions will be responsive to a second round of radioimmunotherapy or a second therapeutic modality such as chemotherapy or an anti-vascular agent. After radioimmunotherapy, there was an increase in tumor pO2 followed by a decrease below initial levels in most mice. Thus defined times may exist when a tumor is more or less radiosensitive after radioimmunotherapy.

Animals↗