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Biomedical subjects

M Purcell

Publications and source records attributed to M Purcell.

At least 19 recordsLinked to original sources
PubMed2004

PCT-233, a novel modulator of pro- and anti-inflammatory cytokine production.

Plant extracts have been implicated in various immunoregulatory effects that are poorly understood. Thus, we investigated the modulatory activity of PureCell Complex (PCT)-233, an active molecular complex from mesophyll tissue of Spinacia oleacea on the inflammatory process. Alveolar macrophages (AM) were treated with PCT-233 and/or budesonide, a well-known anti-inflammatory agent, before or after being stimulated with lipopolysaccharides (LPS). Pro- and anti-inflammatory cytokine production, tumour necrosis factor (TNF) and interleukin (IL)-10, respectively, were measured in cell-free supernatants at different times after the treatment. PCT-233 increased unstimulated AM release of both TNF and IL-10, whereas heat- and light-inactivated PCT-233 stimulated only the release of TNF without affecting IL-10 production, suggesting that different mechanisms are involved in the modulation of TNF and IL-10 release by PCT-233. The presence of LPS did not modify PCT-233-stimulated TNF production, but the ratio TNF/IL-10 production by LPS-stimulated AM was reduced significantly in the presence of PCT-233. Pretreatment of AM with PCT-233 and budesonide before LPS stimulation reduced TNF production at both protein and mRNA levels, whereas IL-10 production was increased. Moreover, TNF/IL-10 ratio was reduced further with the combination PCT-233/budesonide. Interestingly, AM treatment with PCT-233 and budesonide 18 h after LPS stimulation did not modulate TNF release significantly but it did increase IL-10 production, and a synergistic effect was observed with the combination PCT-233/budesonide. These exciting data suggest that PCT-233 possesses some anti-inflammatory properties, even when added during the inflammatory process, and could potentiate the effect of other anti-inflammatory agents.

Animals↗
PubMed2001

Interactions of atrazine and 2,4-D with human serum albumin studied by gel and capillary electrophoresis, and FTIR spectroscopy.

The herbicides 6-chloro-N-ethyl-N'-(1-methylethyl)-1,3,5-triazine-2,4-diamine (atrazine) and 2,4-dichlorophenoxyacetic acid (2,4-D) are widely used in agricultural practice to fight dicotyledon weeds mainly in maize, cereals, and lucerne. As a result, these compounds are found not only in the plants, soil, and water, but also in the cultivated ground in the following years as well as in agricultural products such as fruits, milk, butter, and sugar beet. The toxicological effects of herbicides occur in vivo, when transported to the target organ through the bloodstream. It has been suggested that human serum albumin (HSA) serves as a carrier protein to transport 2,4-D to molecular targets. This study was designed to examine the interaction of atrazine and 2,4-D with HSA in aqueous solution at physiological pH with herbicide concentrations of 0.0001-1 mM, and final protein concentration of 1% w/v. Gel and capillary electrophoresis, UV-visible and Fourier transform infrared spectroscopic methods were used to determine the drug binding mode, the drug binding constant, and the protein secondary structure in aqueous solution. Structural analysis showed that different types of herbicide-HSA complexes are formed with stoichiometric ratios (drug/protein) of 3:1 and 11:1 for atrazine and 4.5:1 and 10:1 for 2,4-D complexes. Atrazine showed a weak binding affinity (K=3.50 x 10(4) M(-1)), whereas two bindings (K(1)=2.50 x 10(4) M(-1) and K(2)=8.0 x 10(3) M(-1)) were observed for 2,4-D complexes. The herbicide binding results in major protein secondary structural changes from that of the alpha-helix 55% to 45--39% and beta-sheet 22% to 24--32%, beta-anti 12% to 10--22% and turn 11% to 12--15%, in the drug-HSA complexes. The observed spectral changes indicate a partial unfolding of the protein structure, in the presence of herbicides in aqueous solution.

2,4-Dichlorophenoxyacetic Acid↗
PubMed2001

TRM1, a YY1-like suppressor of rbcS-m3 expression in maize mesophyll cells.

The genes rbcS and rbcL encode, respectively, the small and large subunits of the photosynthetic carbon dioxide fixation enzyme ribulose bisphosphate carboxylase/oxygenase. There is a single rbcL gene in each chloroplast chromosome; a family of rbcS genes is located in the nuclear genome. These two genes are not expressed in mesophyll cells but are in adjacent bundle-sheath cells of leaves of the C4 plant Zea mays. Two regions of the maize gene rbcS-m3 are required for suppressing expression in mesophyll cells. One region is just beyond the translation termination site in the 3' region, and the other is several hundred base pairs upstream of the transcription start site. A binding site for a protein with limited homology to the viral, yeast, and mammalian transcription repressor-activator YY1 (Yin-Yang I), has now been identified in the 3' region. A maize gene for a protein with zinc fingers homologous to those of YY1 has been isolated, characterized, and expressed in Escherichia coli. The gene is designated trm1 (transcription repressor-maize 1). The protein TRM1 binds to the YY1-like site and, in addition, TRM1 binds to two sequence regions in the 5' region of the gene that have no homology to the YY1 site. Mutagenesis or deletion of any of these three sequences eliminates repression of rbcS-m3 reporter genes in mesophyll cells.

Amino Acid Sequence↗
PubMed2001

Evaluation of a model of intervention to create a visually enriched environment.

Within a day resource for people with learning disabilities 'stand-alone' signing training proved ineffectual to sustain lasting change in the formalised gestural behaviours of carers. Consideration was given to setting conditions pre- and post-training and a new model of delivery evolved, designed in partnership with day resource staff. Aims of intervention were to promote the adoption and active use of signing as an aid to communication and move towards a partnership framework which values the role of carers as co-facilitators and lessens dependency on the therapist. This paper describes and evaluates a training model which was found to be successful in translating knowledge into changes in practice and discusses the critical factors which should be built into future interventions.

Caregivers↗
PubMed2000

Interaction of taxol with human serum albumin.

Taxol (paclitaxel) is an anticancer drug, which interacts with microtuble proteins, in a manner that catalyzes their formation from tubulin and stabilizes the resulting structures (Nogales et al., Nature 375 (1995) 424-427). This study was designed to examine the interaction of taxol with human serum albumin (HSA) in aqueous solution at physiological pH with drug concentrations of 0.0001-0.1 mM, and HSA (fatty acid free) concentration of 2% w/v. Gel electrophoresis, absorption spectra and Fourier transform infrared (FTIR) spectroscopy with self-deconvolution and second-derivative resolution enhancement were used to determine the drug binding mode, binding constant and the protein secondary structure in the presence of taxol in aqueous solution. Spectroscopic evidence showed that taxol-protein interaction results into two types of drug-HSA complexes with overall binding constant of K=1.43 x 10(4) M(-1). The molar ratios of complexes were of taxol/HSA 30/1 (30 mM taxol) and 90/1 (90 mM taxol) with the complex ratios of 1.9 and 3.4 drug molecules per HSA molecule, respectively. The taxol binding results in major protein secondary structural changes from that of the alpha-helix 55 to 45% and beta-sheet 22 to 26%, beta-anti 12 to 15% and turn 11 to 16%, in the taxol-HSA complexes. The observed spectral changes indicate a partial unfolding of the protein structure, in the presence of taxol in aqueous solution.

Drug Interactions↗
PubMed2000

A simple PCR method for rapid genotype analysis of Mycobacterium ulcerans.

Two high-copy-number insertion sequences, IS2404 and IS2606, were recently identified in Mycobacterium ulcerans and were shown by Southern hybridization to possess restriction fragment length polymorphism between strains from different geographic origins. We have designed a simple genotyping method that captures these differences by PCR amplification of the region between adjacent copies of IS2404 and IS2606. We have called this system 2426 PCR. The method is rapid, reproducible, sensitive, and specific for M. ulcerans, and it has confirmed previous studies suggesting a clonal population structure of M. ulcerans within a geographic region. M. ulcerans isolates from Australia, Papua New Guinea, Malaysia, Surinam, Mexico, Japan, China, and several countries in Africa were easily differentiated based on an array of 4 to 14 PCR products ranging in size from 200 to 900 bp. Numerical analysis of the banding patterns suggested a close evolutionary link between M. ulcerans isolates from Africa and southeast Asia. The application of 2426 PCR to total DNA, extracted directly from M. ulcerans-infected tissue specimens without culture, demonstrated the sensitivity and specificity of this method and confirmed for the first time that both animal and human isolates from areas of endemicity in southeast Australia have the same genotype.

DNA Transposable Elements↗
PubMed1999

Communications between staff and adults with intellectual disabilities in naturally occurring settings.

Videotapes were made of 43 staff-client dyads in small-scale residential and day service settings. Frequency counts were made of carers' communicative acts, and two experienced speech and language therapists rated these for appropriateness. Recommendations for enhancing communication were also noted. The results showed that clients were presented with few opportunities to engage as equal partners in the conversational interchanges: staff overly relied on verbal acts, even when they were communicating with predominantly non-verbal clients; they tended to favour the use of directives and questions, and the majority of staff failed to adjust their language to the client's level of understanding. The most commonly recommended changes for staff were to use simpler sentences and words, to increase their use of non-verbal signals and open questions, to provide more opportunities for clients to initiate topics, and to increase their responsiveness to client's non-verbal signals. The explanations for staff behaviour are reviewed and the implications for changing practice are discussed.

Adolescent↗
PubMed1999

Interaction of RNase A with VO3- and VO2+ ions. Metal ion binding mode and protein secondary structure.

Some of vanadyl complexes have shown potential to inhibit RNase activity by acting as transition state analogue, while at the same time not inhibiting DNase. To gain an insight into the interaction of protein with vanadate (VO3-) and vanadyl (VO2+) ions, the present study was designed to examine the binding of ribonuclase A (RNase A) with NaVO3 and VOSO4 in aqueous solution at physiological pH with metal ion concentrations of 0.001 mM to 1 mM, and protein concentration of 2% w/v. Absorption spectra and Fourier transform infrared (FTIR) spectroscopy with self-deconvolution and second derivative resolution enhancement were used to determine the cation binding mode, association constant and the protein secondary structure in the presence of vanadate and vanadyl ions in aqueous solution. Spectroscopic results show that an indirect metal ion interaction occurs with the polypeptide C = O, C-N (via H2O) with overall binding constants of K(VO3-) = 3.93x10(2) M(-1) and K(VO2+) = 4.20x10(3) M(-1). At high metal ion concentrations, major protein secondary structural changes occur from that of the alpha-helix 29% (free enzyme) to 23-24%; beta-sheet (pleated and anti) 50% (free enzyme) to 64-66% and turn 21% (free enzyme) to 10-12% in the metal-RNase complexes. The observed structural changes indicate a partial protein unfolding in the presence of high metal ion concentration.

Animals↗
PubMed1998

Host cell killing and bacterial conjugation require overlapping sets of genes within a 22-kb region of the Legionella pneumophila genome.

A 22-kb DNA locus of Legionella pneumophila is described that contains 18 genes, 16 of which are required for macrophage killing (icm genes). In this paper two previously described icm loci were linked by the discovery of five genes located between the two loci. Four of the newly described genes are required for macrophage killing (icmMLKE) and one is dispensable. The 16 icm genes appeared to be organized as six individual genes (icmR, icmQ, icmG, icmC, icmD, and icmF), and four operons (icmTS, icmPO, icmMLKE, and icmJB). Four icm genes (icmP, icmO, icmL, and icmE) show significant sequence similarity to plasmid genes involved in conjugation, whereas the other icm genes were found not to bear any sequence similarity to database entries. We found that L. pneumophila can mediate plasmid DNA transfer at a frequency of 10(-3) to 10(-4) per donor. Strains containing null mutations in two icm genes (icmT and icmR) showed a severe reduction in conjugation frequency and macrophage killing. Strains containing an insertion in four other icm genes (icmF, icmE, icmC, and dotA) were shown to have a less severe defect in conjugation. Mutations in the other 11 icm genes had no effect on conjugation frequency. We currently do not know whether conjugation itself plays a role in macrophage killing. It is possible either that small plasmids can take advantage of an existing secretion system to be mobilized or that DNA transfer is required for human macrophage killing by L. pneumophila.

Base Sequence↗
PubMed1998

The Legionella pneumophila icmGCDJBF genes are required for killing of human macrophages.

Previously, a collection of mutants of Legionella pneumophila that had lost the ability to multiply within and kill human macrophages was generated by Tn903dIIlacZ transposon mutagenesis and classified into DNA hybridization groups. A subset of these mutants was complemented by a plasmid, pMW100, containing a 13.5-kb genomic DNA insert. This plasmid restored the ability to multiply within and produce cytopathic effects on human macrophages to members of DNA hybridization groups II, IV, VI, and XVII. A region of the genomic insert of pMW100 was sequenced, and eight potential genes were identified and named icmE, icmG, icmC, icmD, icmJ, icmB, icmF, and tphA. None of the genes encode potential protein products with significant homology to previously characterized proteins, except for tphA, whose product has significant homology to a family of metabolite/H+ symport proteins from gram-negative bacteria. The positions of the Tn903dIIlacZ insertions within the genes were determined by nucleotide sequencing. No Tn903dIIlacZ insertions mapped to icmG, icmJ, or tphA; therefore, these loci were mutated to test whether they were required for macrophage killing. Complementation analysis was used to evaluate the roles of the potential gene products and provide information on the organization of transcriptional units within the region. The results indicate that all identified open reading frames except tphA are required for killing of human macrophages.

DNA Transposable Elements↗
PubMed1997

Multiple imagings to diagnose the chondrosseous metaplasia within a lipoma near the knee.

A 61-year-old man with a slow-growing, painless mass in the area of the right knee underwent radiographic, computed tomographic (CT), arthrographic, arteriographic, and bone scintigraphic imaging studies. Scintigraphy showed an area of intense uptake in the anterolateral part of the knee; the uptake of the knee was much higher than that of the knee joints, but the area was not connected to the joint. Radiographic findings suggested an osteocartilaginous mass which was seen to contain low-density fatty tissue on the CT exam. Arthrography revealed that there was no connection of the mass to the knee joint. Arteriography showed a mildly vascularized tumor mass. Upon removal, the mass was well encapsulated, measuring 10 x 7 x 7 cm, and consisted of integrated nodules of bone, cartilage, and fat tissue. Microscopic examination confirmed lipoma with osteochondromatous metaplasia. The intense uptake in the lipoma near the bone or joint on the bone scan and multiple osteochondromatous nodules shown on CT may serve as characteristic features of the rare chondrosseous metaplasia within a lipoma.

Arthrography↗
PubMed1997

Detection and comparative analysis of persistent measles virus infection in Crohn's disease by immunogold electron microscopy.

AIMS: To determine the specificity of persistent measles virus infection in intestinal samples from Crohn's disease patients using quantitative immunogold electron microscopy. To compare the results with samples from ulcerative colitis, a granulomatous inflammatory control (tuberculous lymphadenitis), and a positive control. METHODS: Formalin fixed, paraffin embedded intestinal tissue from patients with Crohn's disease was reprocessed and stained with antimeasles nucleocaspid protein primary antibody followed by 10 nm gold conjugated secondary antibody. Tissue samples were taken from granulomatous and non-granulomatous areas of the intestine. Intestinal samples from patients with ulcerative colitis, tuberculous lymphadenitis, or acute mesenteric ischaemia were similarly processed. Brain tissue from a patient with subacute sclerosing panencephalitis (SSPE) was used as the positive control. Duplicate sections of all tissues were processed without the primary antibody. Stained specimens were examined by electron microscopy. RESULTS: In Crohn's disease patients, 8/9 foci of granulomatous inflammation and 0/4 foci of non-specific inflammation were positive for measles virus. Of controls, 0/5 non-inflamed intestinal tissues, 1/8 tuberculous tissues, 1/5 ulcerative colitis tissues, and 1/1 SSPE tissues were positive. Gold grain counts per nuclear field-of-view in both Crohn's disease granulomas (43.29) and SSPE (36.94) were significantly higher than in tissues from patients with ulcerative colitis (13.52) or tuberculous lymphadenitis (15.875), and nongranulomatous areas of Crohn's disease (4.89) (p < 0.001, p < 0.001, p = 0.0006, respectively), with no significant difference between Crohn's disease and SSPE (p > 0.1). In both SSPE and Crohn's disease staining was confined to a small population of cells exhibiting characteristic cytopathology. CONCLUSION: These data support a role for measles virus in the aetiology of Crohn's disease.

Adolescent↗
PubMed1995

Red/far-red and blue light-responsive regions of maize rbcS-m3 are active in bundle sheath and mesophyll cells, respectively.

Leaves of the C4 plant maize have two major types of photosynthetic cells: a ring of five large bundle sheath cells (BSC) surrounds each vascular bundle and smaller mesophyll cells (MC) lie between the cylinders of bundle sheath cells. The enzyme ribulose bisphosphate carboxylase/oxygenase is encoded by nuclear rbcS and chloroplast rbcL genes. It is not present in MC but is abundant in adjacent BSC of green leaves. As reported previously, the separate regions of rbcS-m3, which are required for stimulating transcription of the gene in BSC and for suppressing expression of reporter genes in MC, were identified by an in situ expression assay; expression was not suppressed in MC until after leaves of dark-grown seedlings had been illuminated for 24 h. Now we have found that transient expression of rbcS-m3 reporter genes is stimulated in BSC via a red/far-red reversible phytochrome photoperception and signal transduction system but that blue light is required for suppressing rbcS-m3 reporter gene expression in MC. Blue light is also required for the suppression system to develop in MC. Thus, the maize gene rbcS-m3 contains certain sequences that are responsive to a phytochrome photoperception and signal transduction system and other regions that respond to a UVA/blue light photoperception and signal transduction system. Various models of "coaction" of plant photoreceptors have been advanced; these observations show the basis for one type of coaction.

Gene Expression Regulation, Plant↗
PubMed1994

Localized chemical reactivity in DNA associated with the sequence-specific bisintercalation of echinomycin.

Four complementary footprinting and probing techniques utilizing DNAse I, methidiumpropyl EDTA (MPE).FeII, diethyl pyrocarbonate (DEPC) and KMnO4 as DNA-cleaving or DNA-modifying agents have been applied to investigate the sequence-specific binding to DNA of the antitumour antibiotic echinomycin. A 265 bp EcoRI-PvuII DNA restriction fragment excised from plasmid pBS was used as a substrate. Six regions of protection against DNAase I cleavage were located on the 265-mer: three sites encompass the sequences 5'-TCGA or 5'-GCGT and the three others contain 5'-GpG (CpC) dinucleotide sequences where the inhibition of DNAase I cutting by echinomycin is less pronounced. In contrast, MPE.FeII cleavage allows identification of only three echinomycin-binding sites on the 265-mer: two sites contain the sequence 5'-TCGA and one encompasses the sequence 5'-ACCA. Cleavage of DNA by MPE.FeII in the presence of echinomycin remains practically unaffected at the sequence 5'-GCGT, despite its identification by DNAase I as a strong site for binding the antibiotic, as well as at the two other sequences containing GpG steps. With both DNAase I and MPE.FeII, enhanced DNA cleavage is evident at AT-rich sequences in the presence of echinomycin. Enhanced reactivity towards KMnO4 and DEPC provides clear evidence for sequence-dependent conformational changes in DNA induced by the antibiotic. The experiments reveal that KMnO4 reacts most strongly with thymines located around, but not necessarily adjacent to, an echinomycin-binding site, whereas the carbethoxylation reactions caused by DEPC occur primarily at the adenine residues lying immediately 5' or 3' to the dinucleotide that denotes an echinomycin-binding site. The results reported here demonstrate that DEPC and KMnO4 serve as sensitive probes for different states of the DNA helix. It seems that the reaction with KMnO4 involves transient unstacking events, whereas the carbethoxylation reaction of DEPC requires larger-scale helix opening.

Base Sequence↗