Biomedical subjects
Publications and source records attributed to J Crosby.
The minimal actinorhodin polyketide synthase bearing two point mutations (KSbeta Q161A, ACP C17S) was chemically modified to carry novel C4 to C8 starter units on the ACP: on incubation with an excess of malonyl CoA new 16-carbon polyketides are made, supporting a measuring mechanism.
Patients with myotonic dystrophy frequently suffer from excess daytime sleepiness, which can be a significant cause of disability. Previous studies have indicated that this excess daytime sleepiness is only occasionally due to obstructive sleep apnoea and may be principally of central nervous system origin. Modafinil has been successfully used to treat narcolepsy, a central disorder causing excess daytime sleepiness. We have investigated the use of this drug in myotonic dystrophy patients with excess daytime sleepiness. Patients were recruited from a clinic population on the basis of screening with the Epworth Sleepiness Scale. Patients scoring 10 and above were invited to participate in a randomized double-blind crossover trial of modafinil versus placebo, with four weeks in each arm of the study separated by a 2-week washout period. Patients were assessed by polysomnography at baseline. The primary outcome measures were change in both the Epworth Sleepiness Scale and a modified Maintenance of Wakefulness Test, which were measured at the start of each arm of the trial and in week 3 of each intervention period. In agreement with previous smaller studies, sleepiness is not correlated with CTG expansion size. Treatment with modafinil showed a non-significant reduction in median Epworth Sleepiness Scale. However, the median Maintenance of Wakefulness Test score was prolonged by treatment (31.7-40 min, P=0.006). There were no significant adverse cardiac effects of the drug in this group of patients (resting 12 lead and 24 h ECG monitoring). Selected patients with myotonic dystrophy and excess daytime sleepiness may benefit from modafinil. In this patient group the Epworth Sleepiness Scale may not be the most reliable measure of sleepiness. Despite the potential for cardiac disease in these patients, the drug was well tolerated with no adverse effects.
Endogenous beta-carbolines, such as harmane, are known to occur in mammalian species including humans. Radioligand binding studies have revealed that certain beta-carbolines display high affinity for both I(1) and I(2) imidazoline-binding sites (IBS). Functional studies have shown that the beta-carboline harmane elicits many characteristics expected of an endogenous ligand IBS. This article discusses the evidence relating to beta-carbolines as endogenous ligands and presents a case for harmane and related compounds as endogenous ligands for IBS.
Fertility parameters of boar spermatozoa were evaluated in vitro, after freeze-thawing the semen in three different extenders containing permeable and non-permeable cryoprotectants: A (111.0 mM Tris, 31.4 mM citric acid, 185.0 mM glucose, 20 per cent egg yolk, 3 per cent glycerol and 100 iu/ml penicillin G); B (200 mM Tris; 70.8 mM citric acid, 55.5 mM glucose, 20 per cent egg yolk, three per cent glycerol and 100 iu/ml penicillin G); C (200 mM Tris, 70.8 mM citric acid, 55.5 mM fructose, 20 per cent egg yolk, 3 per cent glycerol and 100 iu/ml penicillin G). The freeze-thawing techniques were the same for each extender. Eight ejaculates from four boars were obtained; the sperm-rich fraction of each ejaculate was extended in each of the three media at a final concentration of 400 x 106 sperm/ml, loaded into 0.5 ml straws and frozen at a rate of 30 degrees C/minute to -196 degrees C. The straws were thawed at 60 degrees C for eight seconds. Sperm motility, acrosomal integrity and in vitro sperm penetration through the zona pellucida of gilt oocytes matured in vitro were evaluated. The motility of unfrozen spermatozoa was 93.1 per cent compared with 60.7 per cent, 48.2 per cent and 35 per cent for sperm frozen in extenders A, B and C respectively; these values were all significantly different (P<0.05). There was no significant decline in sperm motility after incubation for 30 minutes in extender A, but there were significant decreases in sperm motility after 30 minutes of incubation in B and C. The percentage acrosomal integrities were 97.2 per cent for the control and 45.5 per cent, 30.3 per cent and 16.8 per cent for the frozen-thawed spermatozoa in extenders A, B and C respectively. The results of the in vitro penetration assay were 80.7 per cent when using control spermatozoa, and 42.2 per cent, 18.4 per cent and 3.3 per cent when using frozen-thawed spermatozoa in extenders A, B and C respectively
This paper describes a set of learning outcomes that clearly define the abilities of medical graduates from any of the five Scottish medical schools. The outcomes are divided into 12 domains that fit into one of three essential elements for the competent and reflective medical practitioner.
The potential role of airway interleukin-5 (IL-5) expression in eliciting mucus production was demonstrated in a pulmonary IL-5 transgenic mouse model (NJ.1726) in which naive transgenic mice display comparable levels of airway mucus relative to allergen-sensitized and -challenged wild-type mice. The intrinsic mucus accumulation of NJ.1726 was abolished in compound transgenic-gene knockout mice deficient of either CD4(+) cells [NJ.1726/CD4(-/-)] or alphabeta T cell receptor-positive (TCR(+)) cells [NJ.1726/alphabeta TCR(-/-)]. In addition, mucus production in naive NJ.1726 was inhibited by >90% after administration of the soluble anti-IL-4 receptor alpha-subunit antagonist. The loss of mucus production in NJ.1726/CD4(-/-), NJ.1726/alphabeta TCR(-/-), and anti-IL-4 receptor alpha-subunit antagonist-treated mice occurred notwithstanding the significant pulmonary eosinophilia and expansion of airway B cells induced by ectopic IL-5 expression. Furthermore, the loss of mucus accumulation occurred in these mice despite elevated levels of airway and peripheral IL-5, indicating that IL-5 does not directly induce goblet cell metaplasia and mucus production. Thus pulmonary expression of IL-5 alone is capable of inducing CD4(+) T cell-dependent goblet cell metaplasia, apparently mediated by IL-4 receptor alpha-subunit-ligand interactions, and represents a previously unrecognized novel pathway for augmenting allergen-induced mucus production.
A series of beta-carbolines were prepared and their affinities for imidazoline (I(1) and I(2)) sites evaluated. Selected compounds were also examined at alpha(2)-adrenoceptors. Some of the beta-carbolines were found to bind with high affinity to I(2)-sites and this affinity was dependent on both the planarity of the molecule and the presence of the aryl ring substituents. Good I(1)-affinity was observed with two of the compounds but none of the tested compounds bound to alpha(2)-adrenoceptors. The hallucinogenic properties of beta-carbolines have been linked to activity at 5-HT receptors, in particular 5-HT(2), however, it is apparent from this study that many of these compounds display substantially higher affinity for the imidazoline sites. This finding, and those showing modulation of some behavioural effects of morphine by I(2)-ligands, suggests that imidazoline sites may be interesting new targets in drug abuse research.
BACKGROUND: There has been on-going debate and public interest in surgical competence in recent years. METHODS: A Delphi reiterative opinion survey was conducted among master surgeons on selection of surgical trainees, methods of assessment of progress of surgical trainees, and revalidation of established consultant surgeons. RESULTS: Selection-the current methods of trainee selection were considered inadequate and in need of revision. The important attributes recognized by group are cognitive factors, innate dexterity, and personality. Important aspects of personality include decision-making ability, insight, team spirit, and emotional stability. Assessment during training-the majority view was that this should be based on clinical judgement/skills, operative skills, and cognitive ability. Assessment of technical ability should be based on standardized checklists. Research within training programs was encouraged but academic achievement does not reflect surgical competence. There was a majority verdict for an exit clinical examination. Revalidation-the group agreed on the need for competence checks during the professional career of surgeons. These should cover knowledge, clinical, operative, and humanistic skills; but expressed concern on the feasibility of a revalidation system that can reliably assess the range of skills needed for surgical competence. There was a majority vote against an internal appraisal system. External assessment by nationally appointed 'assessors' was considered preferable. CONCLUSIONS: Both selection and assessment of surgical trainees require changes and standardization. Although revalidation is necessary, concern was expressed on the reliability and validity of existing and proposed systems.
Lymphocyte and/or eosinophil recruitment is dependent on the sequential interactions between adhesion molecules expressed on activated endothelial cells and both leukocyte subtypes. Endothelial P- and E-selectins mediate tethering and rolling of leukocytes through interactions with P-selectin glycoprotein ligand-1 (PSGL-1), and diapedesis subsequently occurs by engagement of endothelial vascular cell adhesion molecule-1 and CD49d (alpha(4)-integrins). The anti-inflammatory potential of interfering with these adhesive interactions was assessed with an ovalbumin challenge mouse model of asthma. Administration of a soluble form of PSGL-1 reduced eosinophils (80%) and lymphocytes (50%) in bronchoalveolar lavage fluid without affecting epithelial changes or airway hyperreactivity (AHR). In contrast, although administration of anti-CD49d monoclonal antibodies (PS/2) resulted in similar reductions in eosinophils (75%) and lymphocytes (50%), PS/2 reduced and abolished mucous cell metaplasia and AHR, respectively. Administration of both PSGL-1 and PS/2 had the additive effect of eliminating eosinophils from the airways (96% decrease), with few or no additional reductions (relative to PS/2 administration alone) in lymphocyte recruitment, mucous cell metaplasia, or AHR. These data show that eosinophils and lymphocytes differentially utilize adhesive interactions during recruitment and that the inhibition of AHR is independent of this recruitment.
Overexpression of interleukin (IL)-5 by the airway epithelium in mice using the rat CC10 promoter (NJ.1726 line) leads to several histopathologies characteristic of human asthma, including airway hyperreactivity (AHR). We investigated the contribution of B and T cells, as well as CD4 expression, to the development of AHR in IL-5 transgenic mice. NJ.1726 mice on a T cell or CD4 knockout background, but not on a B cell knockout background, lost intrinsic AHR. These effects occurred without decreases in IL-5 or eosinophils. We further investigated the contribution of alpha(4)-integrin signaling to the development of AHR in IL-5 transgenic mice through the administration of anti-CD49d (alpha(4)-integrin) antibody (PS/2). Administration of PS/2 resulted in immediate (16-h) inhibition of AHR. The inhibition of AHR was not associated with a decrease in airway eosinophils. These studies demonstrate that, despite the presence of increased levels of IL-5 and eosinophils in the lungs of NJ.1726 mice, CD4(+) cells and alpha(4)-integrin signaling are necessary for the intrinsic AHR that develops in IL-5 transgenic mice.
Synthesis of 4-hydroxycinnamic acid 6 and its N-hydroxysuccinimide ester 8 has been carried out in high yield on solid support. Further development allowed the synthesis of 4-hydroxycinnamoyl CoA 1 in excellent overall yield. The utility of solid phase as a method for the synthesis of 4-hydroxycinnamic acid derivatives was demonstrated by the synthesis of a number of compounds including the NMDA receptor antagonists, N-(phenylalkyl)cinnamides 9 and 10.
A crude extract of clonidine-displacing substance (CDS) has previously been extracted from the NG108-15 cell line. This study aimed to purify CDS extracted from this cell line further, by the technique of reverse phase-HPLC (RP-HPLC), and subsequently determine whether this refined CDS bears any similarity to CDS's extracted from other tissues. Crude CDS was extracted from NG108-cells and fractionated by RP-HPLC eluting with a linear gradient of methanol (5-65%; 1 ml min(-1) flow rate) over 50 min., and collected at 1 min. intervals. The pharmacological activities of the CDS fractions were determined by their abilities to displace bound [3H]clonidine to alpha(2)-adrenoceptors in rat brain membranes. RP-HPLC analysis of CDS revealed a pharmacologically active fraction distinct from agmatine, eluting at 24 min, corresponding to an absorbance peak observed at this time. Collectively, these results confirmed that CDS was present in the NG108-15 cell line. However, the RP-HPLC analysis showed the pharmacological activity to elute at a more hydrophobic gradient than previously observed with CDS's extracted from bovine tissues. These results support the notion of the existence of several CDS's.
INTRODUCTION: Integrated teaching and problem-based learning (PBL) are powerful educational strategies. Difficulties arise, however, in their application in the later years of the undergraduate medical curriculum, particularly in clinical attachments. Two solutions have been proposed - the use of integrated clinical teaching teams and time allocated during the week for PBL separate from the clinical work. Both approaches have significant disadvantages. Task-based learning (TBL) is a preferred strategy. In TBL, a range of tasks undertaken by a doctor are identified, e.g. management of a patient with abdominal pain, and these are used as the focus for learning. Students have responsibility for integrating their learning round the tasks as they move through a range of clinical attachments in different disciplines. They are assisted in this process by study guides. METHOD: The implementation of TBL is described in one medical school. One hundred and thirteen tasks, arranged in 16 groups, serve to integrate the student learning as they rotate through 10 clinical attachments. RESULTS: This trans-disciplinary approach to integration, which incorporates the principles of PBL offers advantages to both teachers and students. It recognizes that clinical attachments in individual disciplines can offer rich learning opportunities and that such attachments can play a role in an integrated, as well as in a traditional, curriculum. In TBL, the contributions of the clinical attachments to the curriculum learning outcomes must be clearly defined and tasks selected which will serve as a focus for the integration of the students' learning over the range of attachments.
Antibiotic-producing polyketide synthases (PKSs) are enzymes responsible for the biosynthesis in Streptomyces and related filamentous bacteria of a remarkably broad range of bioactive metabolites, including antitumour aromatic compounds such as mithramycin and macrolide antibiotics such as erythromycin. The molecular basis for the selection of the starter unit on aromatic PKSs is unknown. Here we show that a component of aromatic PKS, previously named 'chain-length factor', is a factor required for polyketide chain initiation and that this factor has decarboxylase activity towards malonyl-ACP (acyl carrier protein). We have re-examined the mechanism of initiation on modular PKSs and have identified as a specific initiation factor a domain of previously unknown function named KSQ, which operates like chain-length factor. Both KSQ and chain-length factor are similar to the ketosynthase domains that catalyse polyketide chain extension in modular multifunctional PKSs and in aromatic PKSs, respectively, except that the ketosynthase domain active-site cysteine residue is replaced by a highly conserved glutamine in KSQ and in chain-length factor. The glutamine residue is important both for decarboxylase activity and for polyketide synthesis.
Crude methanolic clonidine-displacing substance (CDS) extracted from bovine lung competed for radioligand binding from alpha2-adrenoceptors and I2-sites present in rat brain membranes, and from I1-sites present in rat brain and kidney membranes. There was no difference in the competition of [3H]clonidine binding to alpha2-adrenoceptors present in either rat or rabbit brain membranes by the crude CDS extract and therefore either tissue could be used to estimate the number of units of CDS present in extracts. Further purification by reverse phase high performance liquid chromatography (RP-HPLC), with UV detection, of extracts obtained from bovine lung, brain and rat brain exhibited similar three-peak profiles, previously reported. Corresponding fractions competed for radioligand binding to alpha2-adrenoceptors present in rat brain membranes, eluting between 19 and 23 min, which corresponded with the middle peak of the three-peaks. Therefore, we propose the CDS-like material eluting from all these tissues to be similar. Interestingly, CDS extracted from bovine adrenal glands under the same conditions showed a similar three-peak profile, but did not repeat the displacement of binding just at 19-23 min, but at every time point after 4 min. This suggests this tissue could represent a source of CDS in this species.
Plasmodium merozoites are covered with a palisade layer of proteins that are arranged as organized bundles or appear as protruding spikes by electron microscopy. Here we present a third Plasmodium vivax merozoite surface protein, PvMSP-3, which is associated with but not anchored in the merozoite membrane. Serum from a P. vivax immune squirrel monkey was used to screen a lambdagt11 P. vivax genomic DNA (gDNA) library. Plaque-selected antibodies from clone no. 6.1, and rabbit antisera against its encoded protein, produced a pattern in immunofluorescence assays (IFAs) that is consistent with a localization at the surface of mature schizonts and free merozoites. Specific antisera also agglutinated merozoites and recognized a protein of 150 000 Da by SDS-PAGE. The complete msp-3 gene and flanking sequences were cloned from a P. vivax lambda Dash II gDNA library and also partly characterized by RACE (rapid amplification of cDNA ends). The immediate upstream sequence contains non-coding repeats and a putative protein encoding open reading frame (ORF), which are also present on the msp-3 5'RACE gene product. Pvmsp-3 encodes a protein with a calculated mass of 89 573 Da, which has a potential signal peptide and a major central alanine-rich domain (31%) that exhibits largely alpha-helical secondary structure and is flanked by charged regions. The protein does not have a putative transmembrane domain or a consensus sequence for a glycosylphosphatidylinositol (GPI) anchor modification. However, the alanine-rich domain has heptad repeats that are predicted to form coiled-coil tertiary structures, which mediate protein-protein interactions. PvMSP-3 is structurally related to P. falciparum MSP-3 and the 140000 Da MSP of P. knowlesi. Characterization of PvMSP-3, thus, also begins to define a new interspecies family of evolutionarily related Plasmodium merozoite proteins.
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