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R Burgess

Publications and source records attributed to R Burgess.

At least 37 records · Page 2Linked to original sources
PubMed1998

Roles of DnaK and RpoS in starvation-induced thermotolerance of Escherichia coli.

DnaK is essential for starvation-induced resistance to heat, oxidation, and reductive division in Escherichia coli. Studies reported here indicate that DnaK is also required for starvation-induced osmotolerance, catalase activity, and the production of the RpoS-controlled Dps (PexB) protein. Because these dnaK mutant phenotypes closely resemble those of rpoS (sigma38) mutants, the relationship between DnaK and RpoS was evaluated directly during growth and starvation at 30 degrees C in strains with genetically altered DnaK content. A starvation-specific effect of DnaK on RpoS abundance was observed. During carbon starvation, DnaK deficiency reduced RpoS levels threefold, while DnaK excess increased RpoS levels nearly twofold. Complementation of the dnaK mutation restored starvation-induced RpoS levels to normal. RpoS deficiency had no effect on the cellular concentration of DnaK, revealing an epistatic relationship between DnaK and RpoS. Protein half-life studies conducted at the onset of starvation indicate that DnaK deficiency significantly destabilized RpoS. RpoH (sigma32) suppressors of the dnaK mutant with restored levels of RpoS and dnaK rpoS double mutants were used to show that DnaK plays both an independent and an RpoS-dependent role in starvation-induced thermotolerance. The results suggest that DnaK coordinates sigma factor levels in glucose-starved E. coli.

Adaptation, Biological↗
PubMed1997

Effect on cell kill of addition of multidrug resistance modifiers cyclosporin A and PSC 833 to cytotoxic agents in acute myeloid leukaemia.

Multidrug resistance (MDR) mediated by the drug efflux pump P-glycoprotein (Pgp), may cause remission failure and relapse in patients with acute myeloid leukaemia (AML) by extruding cytotoxic agents such as anthracyclines from leukaemic cells thus allowing them to survive. Cell line data suggest that reversal of MDR is possible using modifying drugs such as cyclosporin A (CSA) and its analogue PSC 833. We have investigated the effects on cell kill of the addition of CSA and PSC 833 to daunorubicin, idarubicin, mitozantrone, etoposide and cytarabine in 52 fresh cell samples from AML patients using an MTT assay. Pgp status was determined by using monoclonal antibodies JSB-1 and MRK-16 and by assessment of rhodamine efflux. Although overall each cytotoxic-modifier combination produced significant improvements in cell kill compared to cytotoxic alone (P values ranged from P < 0.001 to P = 0.017), modifiers also produced significant cytotoxicity in their own right, and no consistent difference was seen between responses in Pgp-positive and negative groups. Up to one in three Pgp-positive samples failed to show any improvement in cell kill with the addition of CSA or PSC 833, possibly owing to co-expression of alternative resistance mechanisms not affected by the MDR modifiers. The best responses were seen when PSC 833 was added to idarubicin, with 7 out of 22 Pgp-positive cases (32%) showing five-fold improvements in cell kill or better compared to idarubicin alone. Comparison of equimolar concentrations of the two modifiers in the Pgp positive group failed to show a significant difference in cell kill, though PSC 833 was markedly superior to CSA in a minority of highly responsive samples which demonstrated clear evidence of MDR reversal. Our in vitro data suggest that MDR modifiers such as CSA and PSC 833 could play an important role in the therapy of AML and indicate the need for prospective randomised trials to assess their clinical efficacy.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

Protection of sensitive patient-connected medical devices from the subversive use of mobile telecommunication equipment.

The UK Department of Health has recently advised caution with respect to the uncontrolled use of private mobile telecommunication equipment in designated sensitive hospital areas. This advice, which has clear implications for patient safety and well being, was issued following reports of the interference of automatic drug delivery devices, infusion pumps and other sensitive systems. The difficulty of monitoring relatives, visitors and, occasionally, staff who use such mobile telephones in the sensitive areas has led to an initiative by The Princess Margaret Hospital to automatically and continuously monitor the frequencies used by the mobile telephones and to initiate an alarm should such phones be used within a designated area. This paper describes a protocol for the use of these monitors developed in the Princess Margaret Hospital which is designed to minimize the number of monitoring units while providing optimum departmental cover.

Artifacts↗
PubMed1997

The effect of cardiopulmonary bypass on circulating megakaryocytes.

Megakaryocytes (Mks) are found in the lungs and the blood stream as well as in the bone marrow. We modified a whole blood filtration method for Mks by immunostaining for CD61 using biotin streptavidin, and used this technique to study Mks and their morphology in the central venous and arterial circulations before, during and after cardiopulmonary bypass (CPB) in haematologically normal patients undergoing routine cardiac surgery. Blood samples were taken immediately after the insertion of central venous (V) and arterial (A) catheters and after thoracotomy, immediately before bypass. Further samples were taken after 60-90 min on-CPB and 180-240 min post-bypass. In comparison with the steady state before bypass, circulating Mk levels in blood on bypass increased dramatically, from (V) 10.93 +/- 3.94/ml (mean +/- SD) to 36.48 +/- 11.52/ml and from (A) 8.37 +/- 4.39/ml to 38.65 +/- 20.68/ml. This effect was still present, to a lesser extent, 180-240 min post-bypass. Circulating levels of Mks were consistently lower in the arterial circulation than in the venous circulation off bypass, but levels in the two circulations were comparable during CPB, confirming previous suggestions that the lungs are net removers of Mks from the circulation. Type 4 Mks, the largest and most normal morphologically, were rarely seen in arterial blood, but increased significantly during CPB, indicating that the lungs selectively remove large Mks. The lungs appear to play an active role in the regulation of Mk levels. This is lost during CPB and despite the extracorporeal 40 microm arterial line filter, large Mks enter the systemic circulation. More effective extracorporeal filtration of large Mks might reduce the neurological impairment seen in some patients who have undergone CPB.

Adult↗
PubMed1997

The in vitro effect of pegylated recombinant human megakaryocyte growth and development factor (PEG rHuMGDF) on megakaryopoiesis in normal subjects and patients with myelodysplasia and acute myeloid leukaemia.

Mpl ligand is a recently cloned haemopoietic growth factor that stimulates megakaryopoiesis in vitro and in vivo. We describe the in vitro effect of a truncated form of Mpl ligand, recombinant human megakaryocyte growth and development factor (rHuMGDF), on megakaryopoiesis in bone marrow from normal subjects and patients with myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML). We used both semi-solid and suspension culture techniques to assess the effect of pegylated (PEG) rHuMGDF on megakaryocyte colony growth (CFU-Mk) and on the production of CD61+ cells in 7d suspension cultures. PEG rHuMGDF increased CFU-Mk growth and CD61+ cell production in a dose-dependent fashion in all normal marrows tested. Normal CFU-Mk growth was increased threefold with the addition of 10 ng/ml PEG rHuMGDF to cultures and CD61+ cells were increased 8-10-fold by the same dose. Although increased CFU-Mk growth was only seen in 1/10 AML and 6/16 MDS marrows, CD61+ cell numbers in suspension culture were increased in 9/13 AML and 12/15 MDS samples, responses ranged from very limited to normal magnitude. There was no correlation between platelet count and CFU-Mk number, CD61+ cell number or response to PEG rHuMGDF. We did not find any increased CFU-GM colony or cluster growth in response to PEG rHuMGDF and the CD61+ cells produced in suspension culture had features of megakaryocytic differentiation. These data suggest that PEG rHuMGDF can enhance megakaryocyte proliferation in some patients with MDS and AML, and may have a role in the treatment of thrombocytopenia in these patients.

Bone Marrow Cells↗
PubMed1997

Colony counting is a major source of variation in CFU-GM results between centres.

The results for colony forming unit granulocyte-macrophage (CFU-GM) assays vary substantially between centres. It is possible that colony counting is largely responsible for this discrepancy. In order to examine this exclusively from the many factors that make up the CFU-GM assay, we performed a colony counting exercise involving 11 laboratories. Two-way analysis of variance showed a highly significant difference (P = 0.0001) in the counts obtained from the centres. One centre was found to score consistently high and two others scored consistently low numbers of colonies. This suggests that identification of colonies is a major source of variation between centres.

Analysis of Variance↗
PubMed1997

Usher's syndrome type IC: clinical studies and fine-mapping the disease locus.

Usher's syndrome type I is a heterogeneous group of diseases characterized by severe to profound sensorineural hearing loss, absent vestibular function, and progressive pigmentary retinopathy. Other identifying clinical features have not been documented. In this study, we examined olfactory acuity, plasma levels of polyunsaturated fatty acids and sarcosine, and cilia ultrastructure in a homogeneous cohort of patients with Usher's syndrome type IC. The normal age-dependent decline in olfactory acuity was observed, and normal plasma levels of polyunsaturated fatty acids and sarcosine were found. However, the incidence of compound cilia in biopsies from the inferior turbinate was significantly higher than that reported in control populations. By reconstructing haplotypes in affected persons. D11S902 and D11S1310 were identified as flanking markers over an interval that contains a candidate gene, KCNC1. No mutations in the coding sequence of this gene could be demonstrated in affected persons.

Adolescent↗
PubMed1996

Requirement of the paraxis gene for somite formation and musculoskeletal patterning.

The segmental organization of the vertebrate embryo is first apparent when somites form in a rostrocaudal progression from the paraxial mesoderm adjacent to the neural tube. Newly formed somites appear as paired epithelial spheres that become patterned to form vertebrae, ribs, skeletal muscle and dermis. Paraxis is a basic helix-loop-helix transcription factor expressed in paraxial mesoderm and somites. Here we show that in mice homozygous for a paraxis null mutation, cells from the paraxial mesoderm are unable to form epithelia and so somite formation is disrupted. In the absence of normal somites, the axial skeleton and skeletal muscle form but are improperly patterned. Unexpectedly, however, we found that formation of epithelial somites was not required for segmentation of the embryo or for the establishment of somitic cell lineages. These results demonstrate that paraxis regulates somite morphogenesis, and that the function of somites is to pattern the axial skeleton and skeletal muscles.

Animals↗
PubMed1995

Paraxis: a basic helix-loop-helix protein expressed in paraxial mesoderm and developing somites.

During vertebrate embryogenesis, cells from the paraxial mesoderm coalesce in a rostral-to-caudal progression to form the somites. Subsequent compartmentalization of the somites yields the sclerotome, myotome, and dermatome, which give rise to the axial skeleton, axial musculature, and dermis, respectively. Recently, we cloned a novel basic helix-loop-helix (bHLH) protein, called scleraxis, which is expressed in the sclerotome, in mesenchymal precursors of bone and cartilage, and in connective tissues. Here we report the cloning of a bHLH protein, called paraxis, which is nearly identical to scleraxis within the bHLH region but diverges in its amino and carboxyl termini. During mouse embryogenesis, paraxis transcripts are first detected at about Day 7.5 postcoitum within primitive mesoderm lying posterior to the head and heart primordia. Subsequently, paraxis expression progresses caudally through the paraxial mesoderm, immediately preceding somite formation. Paraxis is expressed at high levels in newly formed somites before the first detectable expression of the myogenic bHLH genes, and as the somite becomes compartmentalized, paraxis becomes downregulated in the myotome. Paraxis and scleraxis are coexpressed in the sclerotome, but paraxis expression declines soon after sclerotome formation, whereas scleroaxis expression increases in the sclerotome and its derivatives. The sequential expression of paraxis and scleraxis in the paraxial mesoderm and somites suggests that these bHLH proteins may comprise part of a regulatory pathway involved in patterning of the paraxial mesoderm and in the establishment of somitic cell lineages.

Amino Acid Sequence↗
PubMed1995

PICK1: a perinuclear binding protein and substrate for protein kinase C isolated by the yeast two-hybrid system.

Protein kinase C (PKC) plays a central role in the control of proliferation and differentiation of a wide range of cell types by mediating the signal transduction response to hormones and growth factors. Upon activation by diacylglycerol, PKC translocates to different subcellular sites where it phosphorylates numerous proteins, most of which are unidentified. We used the yeast two-hybrid system to identify proteins that interact with activated PKC alpha. Using the catalytic region of PKC fused to the DNA binding domain of yeast GAL4 as "bait" to screen a mouse T cell cDNA library in which cDNA was fused to the GAL4 activation domain, we cloned several novel proteins that interact with C-kinase (PICKs). One of these proteins, designated PICK1, interacts specifically with the catalytic domain of PKC and is an efficient substrate for phosphorylation by PKC in vitro and in vivo. PICK1 is localized to the perinuclear region and is phosphorylated in response to PKC activation. PICK1 and other PICKs may play important roles in mediating the actions of PKC.

Amino Acid Sequence↗
PubMed1995

Immunostaining of whole agar cultures by APAAP.

We have adapted the alkaline phosphatase-anti alkaline phosphatase (APAAP) technique to demonstrate cell antigen distributions in intact agar culture. The method facilitates batch processing and is no less convenient to perform than standard APAAP procedures. Myeloid and lymphoid antigens generally demonstrated strong staining intensity. However, staining at day 0 consistently produced no antigen expression for two monoclonals (CD11c and CD34) in contrast to positivity in parallel cytospins. CD11c showed rapidly increasing antigen expression over subsequent days of culture whereas the expression of CD34 could not be shown in conventional agar culture at any time from day 0 to day 14. Positivity was only restored in CD34-positive leukaemic cells using a modified culture technique in which cells were cultured as pre-formed small aggregates. Assessment of these aggregates extended to cell cycle analysis using anti-bromodeoxyuridine. CD71 positivity in normal culture samples correlated with colony configuration (whether clones were 'spread' or 'tight' in appearance). CD38 staining of normal bone marrow culture at day 7 showed asymmetrical staining of cells in a small number of micro-groups. The clonal detection of aberrant antigens (CD7, CD2) for assessment of minimal residual disease in AML was a disappointment due to the relative frequency of positive clones in normal culture.

Agar↗
PubMed1995

Propagation of interictal epileptic activity in temporal lobe epilepsy.

We recorded interictal spikes with closely spaced scalp electrodes and sphenoidal electrodes in four patients with temporal lobe epilepsy. We used multiple dipole modeling to study the number, three-dimensional intracerebral location, time activity, and functional relationship of the neuronal sources underlying the epileptic spike complexes. In all patients, we found two significant sources generating the interictal spikes which showed considerable overlap in both space and time. Source 1 was located in the mesiobasal temporal lobe and generated a restricted negativity at the ipsilateral sphenoidal electrode and a widespread positivity over the vertex. Source 2 could be attributed to the lateral temporal neocortex and was associated with a relatively restricted negativity at the ipsilateral temporal electrodes and a more widespread positivity over the contralateral hemisphere. The sources were well separated in space, with an average distance of 45 mm between them. The time activities of both sources showed similar biphasic patterns, with the mesial source leading the lateral source by approximately 40 msec, suggesting propagation of interictal epileptic activity from the mesiobasal to the lateral temporal lobe.

Adult↗
PubMed1994

P-glycoprotein expression on acute myeloid leukaemia blast cells at diagnosis predicts response to chemotherapy and survival.

P-glycoprotein (Pgp) expression, which is associated with the multi-drug resistance (MDR) phenotype, has been reported to be a useful predictor of treatment outcome in acute leukaemia. We have examined the expression of Pgp on acute myeloid leukaemia (AML) cells in 54 newly diagnosed patients, using a novel streptavidin-biotin complex (ABC) technique. 55% of patients at diagnosis were positive for Pgp with JSB-1, a monoclonal antibody that binds to an internal epitope of Pgp. All patients received intensive induction chemotherapy. Post-remission treatment consisted of further chemotherapy +/- bone marrow transplantation. Complete remission (CR) rates were significantly lower in the Pgp positive group than in the Pgp negative group (60% v 92%; P = 0.02). The overall survival for Pgp-positive patients was significantly shorter (329 v 534d, P = 0.004), disease-free survival was also reduced but the difference was not statistically significant (median 277 v 522d, P = 0.16). In this study CD34 expression was not predictive of response to chemotherapy nor was it associated with Pgp expression. Our results confirm the prognostic value of Pgp expression in AML at diagnosis and we suggest that Pgp could be a useful therapeutic target for reversing multi-drug resistance. Furthermore, our simple and sensitive method of detecting Pgp should enable widespread testing to be performed.

ATP Binding Cassette Transporter, Subfamily B, Mem↗