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Biomedical subjects

P Stanley

Publications and source records attributed to P Stanley.

At least 37 records · Page 2Linked to original sources
PubMed2000

Fringe is a glycosyltransferase that modifies Notch.

Notch receptors function in highly conserved intercellular signalling pathways that direct cell-fate decisions, proliferation and apoptosis in metazoans. Fringe proteins can positively and negatively modulate the ability of Notch ligands to activate the Notch receptor. Here we establish the biochemical mechanism of Fringe action. Drosophila and mammalian Fringe proteins possess a fucose-specific beta1,3 N-acetylglucosaminyltransferase activity that initiates elongation of O-linked fucose residues attached to epidermal growth factor-like sequence repeats of Notch. We obtained biological evidence that Fringe-dependent elongation of O-linked fucose on Notch modulates Notch signalling by using co-culture assays in mammalian cells and by expression of an enzymatically inactive Fringe mutant in Drosophila. The post-translational modification of Notch by Fringe represents a striking example of modulation of a signalling event by differential receptor glycosylation and identifies a mechanism that is likely to be relevant to other signalling pathways.

Animals↗
PubMed2000

New evidence for an extra-hepatic role of N-acetylglucosaminyltransferase III in the progression of diethylnitrosamine-induced liver tumors in mice.

N-acetylglucosaminyltransferase III (GlcNAc-TIII) is encoded by the Mgat3 gene and catalyzes the addition of the bisecting GlcNAc to the core of N-glycans. Mice lacking GlcNAc-TIII due to the insertion mutation Mgat3tmlPst (termed Mgat3neo), exhibit retarded progression of liver tumors induced by diethylnitrosamine (DEN; M. Bhaumik et al, Cancer Res., 58: 2881-2887, 1998). This phenotype seemed to be due to a reduction, in activity or amount, of a circulating glycoprotein(s) that enhances DEN-induced liver tumor progression. Here, we provide new evidence to support this hypothesis. First, we show that mice with a deletion mutation of the Mgat3 gene coding exon (Mgat3tmlJxm, termed Mgat3delta) also exhibit retarded progression of DEN-induced liver tumors. At 7 months there was a significant decrease in liver weight (approximately 27%; P < 0.01), reflecting reduced tumor burden in Mgat3delta/delta mice. In addition, tumors were generally fewer and smaller, and histological changes were less severe in Mgat3delta/delta livers. Therefore, tumor progression is retarded in mice with two different null mutations in the Mgat3 gene. Second, we show that the development of DEN-induced tumors is unaltered by high levels of GlcNAc-TIII in the liver of transgenic mice. The Mgat3 gene coding exon under the control of the major urinary protein (MUP) promoter was used to generate transgenic mice that express GlcNAc-TIII in liver. Following DEN injection and phenobarbitol treatment, however, no significant differences were observed between MUP/Mgat3 transgenic and control mice in either tumor numbers or liver weight. The combined data provide strong evidence that retarded progression of tumors in mice lacking GlcNAc-TIII is due to the absence of the bisecting GlcNAc residue on N-glycans of a circulating glycoprotein(s) from a tissue other than liver.

Alleles↗
PubMed2000

alpha(1,3)fucosyltransferases expressed by the gain-of-function Chinese hamster ovary glycosylation mutants LEC12, LEC29, and LEC30.

Gain-of-function glycosylation mutants provide access to glycosylation pathways, glycosylation genes, and mechanisms that regulate expression of a glycotype. Previous studies have shown that the gain-of-function Chinese hamster ovary (CHO) mutants LEC12, LEC29, and LEC30 express an N-ethylmaleimide-resistant alpha(1, 3)fucosyltransferase (alpha(1,3)Fuc-T) activity that is not detected in CHO cells and that generates the Lewis(X) but not the sialyl-Lewis(X) determinant. The three mutants differ, however, in lectin resistance properties, expression of fucosylated antigens, and in vitro alpha(1,3)Fuc-T activities. In this paper we show that each mutant expresses Fuc-TIX, but only LEC30 cells express Fuc-TIV. Using genomic PCR and reverse-transcriptase (RT)-PCR strategies, we isolated coding portions of the CHO Fut4 and Fut9 genes. Each gene is present in a single copy in the CHO and mutant genomes. The Fut4 gene is expressed only in LEC30 cells, while all three mutants express the Fut9 gene. Interestingly, the fucosylation phenotypes of LEC12 and LEC29 cells do not correlate with the relative abundance of their Fut9 gene transcripts (LEC29 >> LEC12). Compared to LEC29 cells, LEC12 cells have an approximately 40-fold higher in vitro alpha(1,3)Fuc-T activity and bind the VIM-2 monoclonal antibody, whereas LEC29 cells do not bind VIM-2. Mixing experiments did not detect Fuc-TIX inhibitory activity in LEC29 cell extracts, and CHO cells expressing a transfected Fut9 gene behaved like LEC12 cells. Therefore, it seems that LEC29 cells may not translate their more abundant Fut9 gene transcripts efficiently or may not synthesize appropriate acceptors for internal alpha(1,3)fucosylation. Alternatively, LEC12 cells may possess, in addition to Fuc-TIX, a novel alpha(1,3)Fuc-T activity.

Amino Acid Sequence↗
PubMed2000

External and internal-external biliary drainage in children with malignant obstructive jaundice.

BACKGROUND: Obstructive jaundice is an uncommon but important clinical problem in children with cancer. Percutaneous transhepatic biliary drainage is widely used to relieve malignant biliary obstruction in adults, but its use in children has not been well described. MATERIALS AND METHODS: Six patients aged between 1 and 17 years underwent external or internal-external biliary drainage to relieve malignant obstructive jaundice. Biochemical, haematological and microbiological parameters were measured before the procedure and repeated 7-9 days later. RESULTS: External or internal-external biliary drainage was technically successful in all patients. No patient developed clinically significant biliary sepsis. Asymptomatic duodenal perforation occurred in one patient with bulky duodenal tumour following conversion from external to internal-external drainage. CONCLUSIONS: External biliary drainage may be successfully performed in children with malignant obstructive jaundice.

Adolescent↗
PubMed2000

Low-stage medulloblastoma: final analysis of trial comparing standard-dose with reduced-dose neuraxis irradiation.

PURPOSE: To evaluate prospectively the effects on survival, relapse-free survival, and patterns of relapse of reduced-dose (23.4 Gy in 13 fractions) compared with standard-dose (36 Gy in 20 fractions) neuraxis irradiation in patients 3 to 21 years of age with low-stage medulloblastoma, minimal postoperative residual disease, and no evidence of neuraxis disease. PATIENTS AND METHODS: The Pediatric Oncology Group and Children's Cancer Group randomized 126 patients to the study. All patients received posterior fossa irradiation to a total dose of 54 Gy in addition to the neuraxis treatment. Patients were staged postoperatively with contrast-enhanced cranial computed tomography, myelography, and CSF cytology. Of the registered patients, 38 were ineligible. RESULTS: The planned interim analysis that resulted in closure of the protocol showed that patients randomized to the reduced neuraxis treatment had increased frequency of relapse. In the final analysis, eligible patients receiving standard-dose neuraxis irradiation had 67% event-free survival (EFS) at 5 years (SE = 7.4%), whereas eligible patients receiving reduced-dose neuraxis irradiation had 52% event-free survival at 5 years (SE = 7.7%) (P =.080). At 8 years, the respective EFS proportions were also 67% (SE = 8.8%) and 52% (SE = 11%) (P =.141). These data confirm the original one-sided conclusions but suggest that differences are less marked with time. CONCLUSION: Reduced-dose neuraxis irradiation (23.4 Gy) is associated with increased risk of early relapse, early isolated neuraxis relapse, and lower 5-year EFS and overall survival than standard irradiation (36 Gy). The 5-year EFS for patients receiving standard-dose irradiation is suboptimal, and improved techniques and/or therapies are needed to improve ultimate outcome. Chemotherapy may contribute to this improvement.

Adolescent↗
PubMed2000

Feasibility and toxicity of chemoembolization for children with liver tumors.

PURPOSE: To determine the feasibility, toxicity, and efficacy of hepatic arterial chemoembolization (HACE) in pediatric patients with refractory primary malignancies of the liver. PATIENTS AND METHODS: Six patients with hepatoblastoma (HB), three with hepatocellular carcinoma (HCC), and two with undifferentiated sarcoma of the liver were treated with HACE every 2 to 4 weeks until their tumors became surgically resectable or they showed signs of disease progression. All but one newly diagnosed patient with HCC had previously received systemic chemotherapy. RESULTS: All patients with HB and HCC responded to HACE, as measured by imaging studies and alpha-fetoprotein levels. Surgical resection (complete or microscopic residual disease) was feasible in five of 11 patients, and three patients remain alive with no evidence of disease. Elevated liver transaminase and bilirubin levels were seen after each one of the 46 courses of HACE. Other toxicities included fever, pain, nausea, vomiting, and transient coagulopathy. CONCLUSION: HACE is feasible, well tolerated, and effective in inducing surgical resectability of primary hepatic tumors in children.

Adolescent↗
PubMed2000

Aspirin--the first hundred years.

'Take two aspirin and call me in the morning', may be an old cliché deriding the medical profession, but the story of the first 100 years of aspirin suggests that this drug may be of benefit for a far greater range of ailments than those who synthesised it 100 years ago could have imagined.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1999

The gain-of-function Chinese hamster ovary mutant LEC11B expresses one of two Chinese hamster FUT6 genes due to the loss of a negative regulatory factor.

The LEC11 Chinese hamster ovary (CHO) gain-of-function mutant expresses an alpha(1,3)fucosyltransferase (alpha(1,3)Fuc-T) activity that generates the LeX, sialyl-LeX, and VIM-2 glycan determinants and has been extensively used for studies of E-selectin ligand specificity. In order to identify regulatory mechanisms that control alpha(1,3)Fuc-T expression in mammals, mechanisms of FUT gene expression were investigated in LEC11 cells and two new, independent mutants, LEC11A and LEC11B. Northern and ribonuclease protection analyses, using probes that span the coding region of a cloned CHO FUT gene, detected transcripts in each LEC11 mutant but not in CHO cells or other gain-of-function CHO mutants that express a different alpha(1,3)Fuc-T activity. Coding region sequence analysis and alpha(1,3)Fuc-T acceptor specificity comparisons with recombinant human Fuc-TV and Fuc-TVI showed that the cloned FUT gene is orthologous to the human FUT6 gene. Southern analyses identified two closely related FUT6 genes in the Chinese hamster, whose evolutionary relationships are discussed. The blots showed that rearrangements had occurred in LEC11A and LEC11 genomic DNA, consistent with a cis mechanism of FUT6 gene activation in these mutants. By contrast, somatic cell hybrid analyses revealed that LEC11B cells express FUT6 gene transcripts due to the loss of a trans-acting, negative regulatory factor. Sequencing of reverse transcriptase-polymerase chain reaction products identified unique 5'- and 3'-untranslated region sequences in FUT6 gene transcripts from each LEC11 mutant. Northern and Southern analyses with gene-specific probes showed that LEC11A cells express only the cgFUT6A gene (where cg is Cricetulus griseus), whereas LEC11 and LEC11B cells express only the cgFUT6B gene. In LEC11A x LEC11B hybrid cells, the cgFUT6A gene was predominantly expressed, as predicted if a trans-acting negative regulatory factor functions to suppress cgFUT6B gene expression in CHO cells. This factor is predicted to be a cell type-specific regulator of FUT6 gene expression in mammals.

Amino Acid Sequence↗
PubMed1999

Etoposide with or without mannitol for the treatment of recurrent or primarily unresponsive brain tumors: a Children's Cancer Group Study, CCG-9881.

PURPOSE: This study was undertaken to evaluate the response of recurrent brain tumors to intravenous etoposide and to evaluate the efficacy of mannitol in augmenting etoposide's tumoricidal effect. PATIENTS AND METHODS: Ninety-nine children between one and 21 years of age with recurrent brain tumors were randomly assigned to treatment with intravenous etoposide 150 mg/M2, with or without mannitol 15 gm/M2, daily for five days every three weeks for one year or until disease progression or death. Computerized tomographic (CT) or magnetic resonance image (MRI) scans, obtained after three cycles of therapy, were compared with pre-therapy scans. Scans were centrally reviewed. RESULTS: Of 87 evaluable patients, 12 (13.8%) were determined to have had an objective response by the institutional radiologist. On central review, 7/66 (10.6%) responses were documented. Responses in centrally reviewed patients were observed in 2/12 (16.7%) low grade astrocytomas, 4/26 (15.4%) medulloblastoma or primitive neuroectodermal tumors (PNET), 1/13 (7.7%) high grade astrocytomas and 0/15 (0%) brain stem gliomas. Survival at one year was 53% (SE 12%) for low grade astrocytomas, 38% (SE 7%) for medulloblastoma or PNET, 28% (SE 10%) for high grade astrocytomas and 9% (SE 5%) for brain stem gliomas. An effect of mannitol was not observed. CONCLUSION: Intravenous etoposide has a low level of activity in the treatment of recurrent low grade astrocytomas and medulloblastoma or PNET. The efficacy of this agent was not enhanced by the coincident intravenous administration of mannitol.

Adolescent↗
PubMed1999

An ordered reaction mechanism for bacterial toxin acylation by the specialized acyltransferase HlyC: formation of a ternary complex with acylACP and protoxin substrates.

The 110 kDa haemolysin protoxin (proHlyA) is activated in the Escherichia coli cytosol by acyl carrier protein-dependent fatty acylation of two internal lysine residues, directed by the co-synthesized protein HlyC. Using an in vitro maturation reaction containing purified protoxin peptides and acylACP, we show unambiguously that HlyC possesses an apparently unique acyltransferase activity fully described by Michaelis-Menten analysis. The Vmax of HlyC at saturating levels of both substrates was approximately 115 nmol acyl group min-1 mg-1 with KMacylACP of 260 nM and KMproHlyA of 27 nM, kinetic parameters sufficient to explain why in vivo HlyC is required at a concentration equimolar to proHlyA. HlyC bound the fatty acyl group from acylACP to generate an acylated HlyC intermediate that was depleted in the presence of proHlyA, but enriched in the presence of proHlyA derivatives lacking acylation target sites. HlyC was also able to bind in vivo 4'-phosphopantetheine. Substitution of conserved amino acids that could act as putative covalent attachment sites did not prevent binding of the fatty acyl or 4'-phosphopantetheine groups. These data and substrate variation analyses suggest that the unique acylation reaction does not involve covalent attachment of fatty acid to the acyltransferase, but rather that it proceeds via a sequential ordered Bi-Bi reaction mechanism, requiring the formation of a non-covalent ternary acylACP-HlyC-proHlyA complex.

Acyl Carrier Protein↗
PubMed1999

A mouse model for mucopolysaccharidosis type III A (Sanfilippo syndrome).

Mucopolysaccharidosis type III A (MPS III A, Sanfilippo syndrome) is a rare, autosomal recessive, lysosomal storage disease characterized by accumulation of heparan sulfate secondary to defective function of the lysosomal enzyme heparan N- sulfatase (sulfamidase). Here we describe a spontaneous mouse mutant that replicates many of the features found in MPS III A in children. Brain sections revealed neurons with distended lysosomes filled with membranous and floccular materials with some having a classical zebra body morphology. Storage materials were also present in lysosomes of cells of many other tissues, and these often stained positively with periodic-acid Schiff reagent. Affected mice usually died at 7-10 months of age exhibiting a distended bladder and hepatosplenomegaly. Heparan sulfate isolated from urine and brain had nonreducing end glucosamine- N -sulfate residues that were digested with recombinant human sulfamidase. Enzyme assays of liver and brain extracts revealed a dramatic reduction in sulfamidase activity. Other lysosomal hydrolases that degrade heparan sulfate or other glycans and glycosaminoglycans were either normal, or were somewhat increased in specific activity. The MPS III A mouse provides an excellent model for evaluating pathogenic mechanisms of disease and for testing treatment strategies, including enzyme or cell replacement and gene therapy.

Animals↗
PubMed1999

Induction for senior house officers. Part I: The hospital programme.

The study was set up to examine how an induction programme can be designed to meet the needs and expectations of senior house officers (SHOs) beginning a new post in a hospital. A total of 63 SHOs, in five hospitals in the Anglia region, participated in standardised structured interviews up to 3 months after starting their post, by sorting a list of 38 possible induction topics in order of priority. Trainees' early induction needs were found to centre on day-one service activities, i.e., information that enables them to undertake their service work efficiently and effectively, is particularly valued and required early. Information related to clinical education and training is also valued, but many trainees prefer it to be provided after they have been in post for a week or two. Much induction information can also be effectively conveyed in writing, with little or no need for additional formal presentation. We conclude that first-day induction programmes which strive to satisfy the range of trainee needs in one session fail to take account of the different priorities that trainees attach to induction information. Trainees do not want all information at once. Information of prime concern to clinical tutors, namely education, training and generic skills, is generally favoured after a week or two. To ensure that trainees' hospital induction needs are met in an efficient and effective way, programmes must be flexible and timely, and supported by comprehensive written information.

Education, Medical, Graduate↗
PubMed1999

Induction for senior house officers. Part II: The departmental programme.

This study was designed to examine the content and usefulness of departmental induction programmes to senior house officers (SHOs) and to explore perceptions of the usefulness of a range of topics to these trainees. A total of 64 SHOs, in five hospitals in the Anglia region, participated in semi-structured interviews up to 3 months after starting their post. The interviews investigated the content of any induction received and also examined what the trainees would have liked to have received. Almost half (29) of the SHOs also completed a questionnaire which examined the perceived usefulness of various induction topics for a trainee starting a new post. The results showed that, although a departmental induction programme is considered important and highly valued, a substantial minority of SHOs had not received one. Of this minority, 75% would have welcomed an induction. Where an induction had taken place, the focus was primarily upon timetables, tours and meeting people, however, a quarter of the questionnaire sample had not received a service timetable, a third had not met any consultants, two-thirds had received no introduction to clinical management, and two-thirds had received no information about how their consultants manage their patients. Thus, although many trainees receive an induction, important information may not be covered. A staged approach to departmental induction is recommended, using a short, high-quality, and comprehensive induction programme. Flexible and timely programmes, supported by comprehensive written information, will contribute to meeting the needs of trainees in an efficient and effective way.

Education, Medical, Graduate↗
PubMed1999

Intraoperative angiography in the management of pediatric vascular disorders.

We reviewed our experience with intraoperative angiography in the management of 7 cerebral aneurysms and 15 arteriovenous malformations (AVMs) in children over the past 5 years. The patients ranged from 4 months to 18 years of age with a mean age of 9 years. In 4 of the 22 cases, angiography provided information that led to changes during surgery. In 3 cases, the intraoperative angiogram revealed residual AVMs that were then completely resected. In 1 patient with an anterior circulation aneurysm, intraoperative studies revealed findings which resulted in clip repositioning. Intraoperative angiography added a mean of 43 (+/-19) min to the operative time, with a mode of 40 minutes. There was 1 complication, a groin hematoma that readily resolved. Intraoperative angiography prolongs to operative course; however, it is a useful adjunct to the management of vascular disorders in children. We now routinely use and recommend intraoperative angiography to assist with the surgical management of vascular malformations in children.

Adolescent↗
PubMed1999

Metastasis stage, adjuvant treatment, and residual tumor are prognostic factors for medulloblastoma in children: conclusions from the Children's Cancer Group 921 randomized phase III study.

PURPOSE: From 1986 to 1992, "eight-drugs-in-one-day" (8-in-1) chemotherapy both before and after radiation therapy (XRT) (54 Gy tumor/36 Gy neuraxis) was compared with vincristine, lomustine (CCNU), and prednisone (VCP) after XRT in children with untreated, high-stage medulloblastoma (MB). PATIENTS AND METHODS: Two hundred three eligible patients with an institutional diagnosis of MB were stratified by local invasion and metastatic stage (Chang T/M) and randomized to therapy. Median time at risk from study entry was 7.0 years. RESULTS: Survival and progression-free survival (PFS) +/- SE at 7 years were 55%+/-5% and 54%+/-5%, respectively. VCP was superior to 8-in-1 chemotherapy, with 5-year PFS rates of 63%+/-5% versus 45%+/-5%, respectively (P = .006). Upon central neuropathology review, 188 patients were confirmed as having MB and were the subjects for analyses of prognostic factors. Children aged 1.5 to younger than 3 years had inferior 5-year estimates of PFS, compared with children 3 years old or older (P = .0014; 32%+/-10% v 58%+/-4%, respectively). For MB patients 3 years of age or older, the prognostic effect of tumor spread (MO v M1 v M2+) on PFS was powerful (P = .0006); 5-year PFS rates were 70%+/-5%, 57%+/-10%, and 40%+/-8%, respectively. PFS distributions at 5 years for patients with M0 tumors with less than 1.5 cm2 of residual tumor, versus > or = 1.5 cm2 of residual tumor by scan, were significantly different (P = .023; 78%+/-6% v 54%+/-11%, respectively). CONCLUSION: VCP plus XRT is a superior adjuvant combination compared with 8-in-1 chemotherapy plus XRT. For patients with M0 tumors, residual tumor bulk (not extent of resection) is a predictor for PFS. Patients with M0 tumors, > or = 3 years with < or = 1.5 cm2 residual tumor, had a 78%+/-6% 5-year PFS rate. Children younger than 3 years old who received a reduced XRT dosage had the lowest survival rate.

Adolescent↗
PubMed1999

Fibrovascular ingrowth in porous ocular implants: the effect of material composition, porosity, growth factors, and coatings.

PURPOSE: Fibrovascular ingrowth into various porous ocular implants as a function of implant material composition, porosity, growth factors, and coatings was investigated in a pilot study in an animal model. METHODS: Eighty-one New Zealand white rabbits underwent unilateral enucleation and implantation with ocular implants composed of the following materials: coralline hydroxyapatite (HA) with 200-microm pores (HA200) or 500-microm pores (HA500), synthetic HA (synHA), and high-density porous polyethylene (PP). The HA200, HA500, and PP implants were implanted untreated or after treatment with recombinant human basic fibroblast growth factor (Rh-bFGF). Nine HA500 implants were implanted after coating with calcium sulfate (plaster of Paris) to provide a smooth outer surface. Implants were harvested at 1-, 2-, 4-, or 8-week intervals and were examined histologically. RESULTS: A significant difference was found between untreated HA500 and PP, with PP showing better ingrowth. There was no significant difference between untreated HA and PP, nor between untreated HA500 and synHA. Significant increases in ingrowth were found in HA200 compared with HA500, and in Rh-bFGF-treated implants compared with untreated controls. The calcium sulfate-coated implants showed less vascularization compared with the uncoated implants, although the difference was not significant. CONCLUSIONS: Fibrovascular ingrowth occurred earlier in HA200 implants than in HA500 implants, and was enhanced when implants were treated with Rh-bFGF.

Animals↗