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Biomedical subjects

K Lewis

Publications and source records attributed to K Lewis.

At least 37 records · Page 2Linked to original sources
PubMed2001

National surveillance of HIV-1 subtypes for England and Wales: design, methods, and initial findings.

The HIV-1 infections detected in an ongoing national unlinked anonymous HIV surveillance program were subtyped and analyzed according to demographic and risk characteristics. Of the 893 anti--HIV-1--positive specimens allocated to an exposure group, 70% could be subtyped. Almost 25% of infections subtyped were non-B, mostly subtypes A, C, and D. Non-B infections were rare in homosexual and bisexual men and in drug injectors. Forty percent of infections in heterosexual men attending genitourinary medicine clinics were non-B subtypes; of these, 25% were subtype A infections and 59% were subtype C infections. For female clinic attendees, 61% were non-B subtype infections, of which 48% were subtype A infections and 42% were subtype C infections. Of the clinic attendees born in the United Kingdom and Europe, 14% of the men and 35% of the women were infected with non-B subtypes. In contrast, 78% of infections in antenatal patients were non-B subtypes, of which 61% were subtype A and 29% were subtype C. Extrapolation to the estimated 29,700 prevalent adult infections in the United Kingdom indicates that approximately 8,100 (27%) such infections are non-B subtypes and that these are found almost entirely in heterosexuals. The findings suggest spread of infection of non-B subtypes to heterosexuals born in the United Kingdom from individuals infected in regions of high prevalence.

Acute Disease↗
PubMed2001

Urocortin II: a member of the corticotropin-releasing factor (CRF) neuropeptide family that is selectively bound by type 2 CRF receptors.

Here we describe the cloning and initial characterization of a previously unidentified CRF-related neuropeptide, urocortin II (Ucn II). Searches of the public human genome database identified a region with significant sequence homology to the CRF neuropeptide family. By using homologous primers deduced from the human sequence, a mouse cDNA was isolated from whole brain poly(A)(+) RNA that encodes a predicted 38-aa peptide, structurally related to the other known mammalian family members, CRF and Ucn. Ucn II binds selectively to the type 2 CRF receptor (CRF-R2), with no appreciable activity on CRF-R1. Transcripts encoding Ucn II are expressed in discrete regions of the rodent central nervous system, including stress-related cell groups in the hypothalamus (paraventricular and arcuate nuclei) and brainstem (locus coeruleus). Central administration of 1-10 microg of peptide elicits activational responses (Fos induction) preferentially within a core circuitry subserving autonomic and neuroendocrine regulation, but whose overall pattern does not broadly mimic the CRF-R2 distribution. Behaviorally, central Ucn II attenuates nighttime feeding, with a time course distinct from that seen in response to CRF. In contrast to CRF, however, central Ucn II failed to increase gross motor activity. These findings identify Ucn II as a new member of the CRF family of neuropeptides, which is expressed centrally and binds selectively to CRF-R2. Initial functional studies are consistent with Ucn II involvement in central autonomic and appetitive control, but not in generalized behavioral activation.

Amino Acid Sequence↗
PubMed2001

Flavonolignan and flavone inhibitors of a Staphylococcus aureus multidrug resistance pump: structure-activity relationships.

Although some progress has been reported on structure-activity relationships (SARs) for inhibitors of mammalian P-glycoprotein MDR efflux pumps, there is almost nothing in the literature regarding SARs for inhibitors of any bacterial efflux pump. Indeed, only a few of these have been described. Our discovery of a potent naturally occurring flavonolignan inhibitor of the NorA MDR pump of Staphylococcus aureus provided a structural foundation upon which SARs could be assessed via synthetic analogues. Several flavonolignans were prepared which proved to have greater potency than the natural isolate, 5'-methoxyhydnocarpin-D, while others showed decreased potency. Surprisingly, some simple alkylated flavones also were quite active MDR pump inhibitors. Variability of activity among the compounds tested was sufficient so that at least some SARs could be postulated and compared with those known for P-glycoprotein.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2001

HIV-1 infected immune competent mononuclear phagocytes influence the pathways to neuronal demise.

Secretory products from HIV-1-infected immune-competent mononuclear phagocytes (MP) damage neuronal dendritic arbor (Zheng et al., 2001). The mechanism behind neuronal injury and whether it is species and/or viral strain dependent is not fully understood. To these ends, we investigated whether HIV-1-infected and lipopolysaccharide (LPS)-activated MDM elicit neuronal injury in primary human neurons. Neuronal damage was compared to that seen in rat neurons. Utilizing a spectrum of HIV-1 strains to infect human monocyte-derived macrophages (MDM), productive viral replication proved necessary, but not sufficient, for neuronal injury. Neuronal demise was induced by virion-free HIV-1-infected and immune-activated MDM culture supernatants. Maximal alterations in glutamate mediated neuronal signaling, resulted from exposure to secretory products from HIV-1-infected and immune-activated MDM. Apoptosis was the predominant mechanism of cell death induced by HIV-1-infected and LPS-treated MDM. Importantly, neuronal injury and increases in calcium influx mediated by HIV-1-infected and immune-activated MDM culture supernatants was partially blocked by the N-methyl D-aspartate (NMDA) receptor antagonist, MK 801. These data support a primary role for immune-activation in MP neurotoxic activities. The upregulation of NMDA receptor sensitive soluble factors and neuronal apoptosis by HIV-1-infected and immune-activated MDM provide unique insights into links between soluble factors, produced as a consequence of MP immunity, and neuronal demise in HAD.

Journal Article↗
PubMed2001

Staphylococcus aureus MDR efflux pump inhibitors from a Berberis and a Mahonia (sensu strictu) species.

Bioactive fractionation, based on multi-drug resistance (MDR) pump inhibition in Staphylococcus aureus, resulted in the isolation of the active inhibitors 5'-methoxyhydnocarpin-D from leaves of Berberis (formerly Mahonia) trifoliolata and pheophorbide a from Berberis fendleri. The hydnocarpin derivative was not found in the latter species. Pheophytin a (the phytol derivative of pheophorbide a) was identified from both species, but it proved to have no MDR pump inhibitory activity. The somewhat uncommon, and inactive, flavonoid tricin was identified from B. trifoliolata. The occurrence of a flavonolignan in Mahonia-tpe species and its absence in Berberis sensu strictu may provide a chemical differentiation between the two groups which are now recombined on the basis of DNA studies. The strong bacterial efflux pump inhibition of pheophorbide a could be of importance as a plant defense against natural pathogens.

Journal Article↗
PubMed2001

Analysis of the factor V Leiden mutation using the READIT Assay.

BACKGROUND: A variety of methods exist for the detection of single-nucleotide polymorphisms (SNPs) present in amplified segments of genomic DNA. We show the application of a novel SNP scoring tool for analysis of the factor V Leiden mutation. METHODS AND RESULTS: We have developed a novel method for analyzing SNPs. The luciferase-based technique, known as the READIT Technology (Promega Corp, Madison, WI), was used to analyze 510 residual human samples sent for factor V Leiden testing from three independent testing laboratories. A blinded retrospective analysis of the factor V Leiden mutation was used to determine the accuracy and throughput capabilities of the technology. One hundred percent concordance was observed between the READIT Assay and genotype assignments made in the testing laboratories. In addition, greater than 6 SDs of separation were observed between the means of wild-type and heterozygote sample populations. Repetitive sample measurements with representative wild-type, heterozygote, and mutant samples showed that greater than 9 SDs separated the means of heterozygote and homozygote sample populations. Confidence intervals based on the means of wild-type, heterozygote, and mutant sample populations were determined. CONCLUSION: Perfect concordance using the READIT Assay showed its effectiveness as a SNP scoring tool. The design of the factor V READIT Assay was straightforward, requiring the design of two unmodified oligonucleotides that differ at the 3' penultimate position to form perfect hybrids with the wild-type or Leiden form of the factor V sequence. The use of previously published amplification primers and conditions minimized the time needed to optimize and validate the assay. The READIT Calculator supplied with the assay allowed automated genotype assignments and statistical analysis from the READIT Assay data. Confidence-interval analysis validated the ability to distinguish between wild-type, heterozygote, and mutant samples using the READIT Assay.

DNA↗
PubMed2001

Transfer of cationic antibacterial agents berberine, palmatine, and benzalkonium through bimolecular planar phospholipid film and Staphylococcus aureus membrane.

Some organic cations are known to be electrophoretically imported into bacterial cells and actively extruded from these cells by multidrug resistance (MDR) pumps. We have studied penetration of plant antimicrobial agents berberine and palmatine and synthetic antiseptic benzalkonium chloride through black planar phospholipid membrane (BLM) and membrane of Staphylococcus aureus cells. Gradients of these cations across BLM generated an electric potential difference. Penetrating anion tetraphenyl borate and phloretin (a plant substance decreasing membrane dipole potential) stimulated this effect. Under optimal conditions, the magnitude of the electric potential was close to theoretical, that is, 60 mV/10-fold cation gradient. Berberine accumulated in S. aureus cells as shown by direct measurement of berberine with a berberine-sensitive electrode. The berberine accumulation was prevented by protonophore CCCP and was stimulated by mutation in the MDR pump NorA. It is concluded that the plant alkaloids and benzalkonium are penetrating cations and substrates of an MDR pump.

Anti-Infective Agents, Local↗
PubMed2001

Factor XIIIA and clot strength after cardiopulmonary bypass.

Reduced factor XIIIA levels and decreased clot strength have been associated with increased bleeding after cardiopulmonary bypass (CPB). The purpose of this study was to evaluate the relationship between hemostatic factors, including factor XIIIA, and clot strength before, during and after CPB. Factor XIIIA antigen, platelet counts, fibrinogen, factor V activity, tissue plasminogen activator and clot strength (by thromboelastograph) were measured at baseline, after 45 min of CPB, at the end of CPB and 4 h post-operatively in 34 patients. Baseline factor XIIIA antigen was 5.2 +/- 1.4 mg/l. On average, factor XIIIA levels dropped to 64% and clot strength to 77% of baseline values after 45 min on CPB and remained below baseline during the immediate post-operative period. Clot strength was significantly correlated (r = 0.81) with platelet count and fibrinogen but not plasma factor XIIIA levels. Addition of 10 mg/l recombinant factor XIII[a2] significantly increased clot strength. Postoperative bleeding at 2 h was inversely correlated with platelet count, factor XIIIA antigen and clot strength measured at the end of CPB. Maintenance of adequate platelet counts and factor XIIIA levels at the end of CPB may play a role in maintaining clot strength and reducing blood loss.

Adult↗
PubMed2001

Biofilms and planktonic cells of Pseudomonas aeruginosa have similar resistance to killing by antimicrobials.

Biofilms are considered to be highly resistant to antimicrobial agents. Strictly speaking, this is not the case-biofilms do not grow in the presence of antimicrobials any better than do planktonic cells. Biofilms are indeed highly resistant to killing by bactericidal antimicrobials, compared to logarithmic-phase planktonic cells, and therefore exhibit tolerance. It is assumed that biofilms are also significantly more tolerant than stationary-phase planktonic cells. A detailed comparative examination of tolerance of biofilms versus stationary- and logarithmic-phase planktonic cells with four different antimicrobial agents was performed in this study. Carbenicillin appeared to be completely ineffective against both stationary-phase cells and biofilms. Killing by this beta-lactam antibiotic depends on rapid growth, and this result confirms the notion of slow-growing biofilms resembling the stationary state. Ofloxacin is a fluoroquinolone antibiotic that kills nongrowing cells, and biofilms and stationary-phase cells were comparably tolerant to this antibiotic. The majority of cells in both populations were eradicated at low levels of ofloxacin, leaving a fraction of essentially invulnerable persisters. The bulk of the population in both biofilm and stationary-phase cultures was tolerant to tobramycin. At very high tobramycin concentrations, a fraction of persister cells became apparent in stationary-phase culture. Stationary-phase cells were more tolerant to the biocide peracetic acid than were biofilms. In general, stationary-phase cells were somewhat more tolerant than biofilms in all of the cases examined. We concluded that, at least for Pseudomonas aeruginosa, one of the model organisms for biofilm studies, the notion that biofilms have greater resistance than do planktonic cells is unwarranted. We further suggest that tolerance to antibiotics in stationary-phase or biofilm cultures is largely dependent on the presence of persister cells.

Animals↗
PubMed2001

An assessment of four solvents for the recovery of 2-chlorobenzylidenemalononitrile and capsaicins from "CS" and "Pepper" type lachrymator sprays, and an examination of their persistence on cotton fabric.

The rising incidence of assaults involving lachrymator sprays has led to an increase in items being submitted to this laboratory for the analysis of the associated chemical residues. The following work was undertaken to identify an efficient solvent with which to extract the compounds of interest from cotton fabric. The persistence and subsequent recovery of such compounds was also examined following protracted exposure to wind and rain. Ethyl acetate was established as the most efficient solvent of those examined for the extraction of 2-chlorobenzylidenemalononitrile and a range of capsaicins from "CS Gas" and '"Pepper" sprays respectively. Controlled experiments undertaken showed that capsaicins were recoverable after 72 h of exposure to the "elements" and 2-chlorobenzylidenemalononitrile was still recoverable after one week.

Journal Article↗
PubMed2001

In search of natural substrates and inhibitors of MDR pumps.

The function of microbial MDRs remains a hotly debated subject. Given the very broad substrate specificities of some MDRs, like the RND pumps that can extrude all classes of amphipathic compounds (cationic, neutral, and anionic), it seems difficult to develop a rationale for pinpointing possible natural substrates of these translocases. At the same time, several clues can be used to guide our search for natural MDR substrates. One is the fact that amphipathic cations appear to be the preferred substrates of MDRs. These substances are extruded by MDRs of all 5 known families and are the almost exclusive substrates of SMR and MF family MDRs. The universal nature of amphipathic cations as MDR substrates suggests that these were the substances that fueled the evolution of MDR pumps. Two factors apparently favored this particular class of molecules for the role of original MDR substrates--need and opportunity. Unlike other substances, amphipathic cations accumulate in the cell driven by the membrane potential, which makes cations potentially the most dangerous toxins. At the same time, amphipathic cations are highly hydrated and do not permeate the membrane as readily as neutral compounds, making it feasible to design a defense based on an efflux pump. The paucity of known cationic (non-basic) antimicrobials might be a result of using MDR-expressing microbial cells for antibiotic discovery. Plant amphipathic cations, the berberine alkaloids, are good MDR substrates. The Berberis plants produce 5'-methoxyhydnocarpin-D, an MDR inhibitor that potentiates the action of berberine. It is suggested that the further evolution of MDR pumps was determined largely by the barrier function of the membrane they reside in. Thus Gram negative bacteria have an outer membrane barrier that slows the penetration of virtually all amphipathic molecules, and transenvelope MDRs of the RND and EmrAB-type extrude their substrates across this barrier. A low permeability of the cytoplasmic membrane of yeast similarly allows for the operation of broad-specificity ABC and MF MDRs. The presence of MDR sensors that regulate the expression of some MDR pumps strongly suggests that defense against external toxins is the function of these MDRs. The BmrR transcriptional activator of the MerR family induces expression of the Bmr pump in B. subtilis and is a sensor specifically designed to recognize amphipathic cations. Similarly, the OacR repressor binds chemically unrelated cations, which leads to the expression of the QacA pump in S. aureus. In E. coli, the EmrR sensor of the MarR repressor family binds unrelated neutral molecules, allowing for expression of the transenvelope EmrAB pump.

Anti-Bacterial Agents↗
PubMed2000

Translocases: a bacterial tunnel for drugs and proteins.

Unrelated translocases extrude proteins or antimicrobial agents across both membranes of the cell envelope in Gram-negative bacteria. The TolC protein links the translocases to the external environment. The recently determined crystal structure of TolC shows how this universal tunnel operates.

Bacterial Outer Membrane Proteins↗
PubMed2000

Cross-sectional study of the prevalence of feather pecking in laying hens in alternative systems and its associations with management and disease.

A cross-sectional study of risk factors for feather pecking in layings hens in alternative systems was carried out in July 1998. A total of 637 questionnaires were sent out to farmers and producer groups and, after two reminders, the final response rate was 51.5 per cent. The outcome variable was feather pecking after point of lay. Over 55 per cent of the farmers reported that feather pecking had occurred in the last depopulated flock. This outcome was compared with the management procedures reported by flock managers by using univariate statistics. Factors associated with feather pecking with a significance < or = 0.05 were then tested in two logistic regression models. In the first model the following factors were associated with an increased risk of feather pecking: less than 50 per cent of the flock using the outdoor area on a fine and sunny day, the occurrence of egg peritonitis and the occurrence of infectious bronchitis. The direction of the association between feather pecking and these infectious diseases was unclear, so in the second model only factors which were consistent throughout the laying period were tested. The following factors were associated with an increased risk of feather pecking: less than 50 per cent of the flock using the outdoor area on a fine and sunny day; three or more changes of diet during lay; the inspection of the flock by one person; an absence of loose litter at the end of lay; a temperature in the hen house of less than 20 degrees C; turning the lights up when the flock was inspected; and the use of bell-drinkers. It is concluded that some of these factors could inhibit foraging and dust-bathing behaviour and others may increase competition or frustration, both of these changes having been shown experimentally to initiate feather pecking behaviour.

Animal Husbandry↗
PubMed2000

Synergy in a medicinal plant: antimicrobial action of berberine potentiated by 5'-methoxyhydnocarpin, a multidrug pump inhibitor.

Multidrug resistance pumps (MDRs) protect microbial cells from both synthetic and natural antimicrobials. Amphipathic cations are preferred substrates of MDRs. Berberine alkaloids, which are cationic antimicrobials produced by a variety of plants, are readily extruded by MDRs. Several Berberis medicinal plants producing berberine were found also to synthesize an inhibitor of the NorA MDR pump of a human pathogen Staphylococcus aureus. The inhibitor was identified as 5'-methoxyhydnocarpin (5'-MHC), previously reported as a minor component of chaulmoogra oil, a traditional therapy for leprosy. 5'-MHC is an amphipathic weak acid and is distinctly different from the cationic substrates of NorA. 5'-MHC had no antimicrobial activity alone but strongly potentiated the action of berberine and other NorA substrates against S. aureus. MDR-dependent efflux of ethidium bromide and berberine from S. aureus cells was completely inhibited by 5'-MHC. The level of accumulation of berberine in the cells was increased strongly in the presence of 5'-MHC, indicating that this plant compound effectively disabled the bacterial resistance mechanism against the berberine antimicrobial.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

2-(Acyloxy)ethylphosphonate analogues of prenyl pyrophosphates: synthesis and biological characterization.

2-(Acyloxy)ethylphosphonate analogues of geranyl, farnesyl, and geranylgeranyl pyrophosphate have been prepared. Horner-Wadsworth-Emmons condensation of different terpene aldehydes with an unsymmetrical bisphosphonate was the key step in syntheses of the phosphonates bearing alpha,beta-unsaturated acyloxy groups. After preparation of the respective phosphonic acids through reaction with TMSBr, both acids and esters were tested for their effects on DNA synthesis in human-derived myeloid and lymphoid leukemia cell lines. The phosphonate esters varied substantially in their ability to impair proliferation of the different cell lines, but testing against one possible target, farnesyl protein transferase (FPTase), revealed little impact at concentrations ranging up to 10 microM. Because the corresponding 2,3-dihydro compounds showed similar biological activity, conjugate addition would not appear to be involved in the toxicity.

Alkyl and Aryl Transferases↗
PubMed2000

5'-Methoxyhydnocarpin-D and pheophorbide A: Berberis species components that potentiate berberine growth inhibition of resistant Staphylococcus aureus.

A new method of bioactivity-directed fractionation, based on multidrug resistant pump (MDR) inhibition in Staphylococcus aureus, was demonstrated. This resulted in the isolation, from berberine-containing Berberis species, of two compounds that are themselves devoid of S. aureus antibacterial activity, but that form potent synergistic couples with a subinhibitory concentration of berberine. The bacterial MDR pump inhibitors were identified as the flavonolignan 2 and the porphyrin 3. Another natural flavonolignan, silybin (8) from Silybum marianum, was also shown to be a bacterial MDR pump inhibitor.

Berberine↗