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AIM: colesevelam is indicated to lower low density lipoprotein cholesterol (LDL-C) in hyperlipidaemia and improve glycaemic control in adults with type 2 diabetes. This short-term pilot study evaluates its effects in type 1 diabetes. METHODS: this double-blind, randomized, investigator-initiated, single-centred, 12-week pilot study evaluated 40 adults (age = 36.4 ± 9.4 years) with type 1 diabetes (duration = 20.4 ± 8.5 years) and hyperlipidaemia. It was powered to show a treatment difference of >10% LDL-C reduction. Subjects received 3.75 g/day colesevelam (n = 20) or placebo (n = 20) for 12 weeks. LDL-C and haemoglobin A1c (A1c) levels were assessed at screening (week 2), baseline (week 0) and every 4 weeks throughout the treatment duration. Glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP) levels were measured during 4-h meal (Boost Plus, Nestle HealthCare Nutrition Inc., Florham Park, New Jersey, USA) challenge tests (MCT) at baseline and 12 weeks. RESULTS: colesevelam treatment resulted in a significant reduction in LDL-C values at 4 weeks [-12.1% (95% CI: -20.1 to -4.1), p = 0.004] which was sustained for the study duration (p = 0.005 at 12 weeks). The treatment group also showed a significant change in A1c from baseline at week 4; however, this was not significant for the study duration. There was a significant median increase in GLP-1 levels during the first 2 h of the baseline MCT in the treated group but no difference at 12 weeks. CONCLUSIONS: during this short-term pilot study, colesevelam treatment effectively lowered LDL-C in patients with type 1 diabetes. Improvements in A1c seen at week 4 were not sustained. Effects on glycaemic control in subjects with type 1 diabetes may be related to a postprandial rise in GLP-1 levels and require further clinical study.

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The grazing trial at Kidston Gold Mine, North Queensland, was aimed specifically to assess the uptake of metals from the tailing and the potential for unacceptable contamination of saleable meat. Further aims included estimating metal dose rates and identifying potential exposure pathways including plant uptake of heavy metals, mine tailings adhered to plants and direct ingestion of mine tailing. It was found that of the 11 metals analysed (As, Zn, Co, Cd, Cr, Sn, Pb, Sb, Hg, Se and Ni) in the animal's liver, muscle and blood during the 8-month trial period, only accumulation of arsenic and zinc occurred. A risk assessment including these two metals was conducted to determine the potential for chronic metal toxicity and long-term contamination, using the estimates of metal dose rate. It was concluded that no toxicity or long-term contamination in cattle was likely at this site. Management procedures were therefore not required at this site; however, the results highlight percent ground cover and standing dry matter (DM) as important factors in decreasing metal exposure from direct ingestion of tailings and dust adhered to plants.

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A prototype occupational exposure database was developed as part of a study to retrospectively collect chemical exposure data from U.K. industry. The data dictionary for the database was constructed using existing recommendations on core data elements developed by working groups from the ACGIH and the European Union. The study also made use of existing job and workplace coding schemes. The practicalities of gathering the data by voluntary donation, its storage in a database, and the transfer of suitably anonymised data to the U.K. Health and Safety Executive's National Exposure Database system were investigated and assessed. Prior to the development, several existing exposure database systems were evaluated for their suitability to store the data from the study. Though of high quality, these were found to be insufficiently flexible for the diversity of datasets encountered and so the prototype exposure database was constructed using a leading database development package. The database was successfully used to gather data and forward it in a suitable format to the U.K. Health and Safety Executive. The published recommendations on occupational exposure databases and the associated coding schemes provided a very useful foundation for designing and implementing the prototype database. However, as data collection proceeded it became clear that the existing recommendations often were poorly understood and misinterpreted, or at least interpreted differently, by different database designers, data collectors, and other users of occupational exposure data. It is suggested that several items in the ACGIH and European Union core recommendations are ambiguous and need to be clarified. Once agreed, the improved database design criteria need to be widely promoted to foster a common understanding and to encourage their use by all those involved in collecting occupational exposure data. Beyond this, recommendations for exposure databases should be augmented to facilitate easy exchange of data between organizations.

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The microsomal triglyceride transfer protein (MTP) and apolipoprotein B (apoB) belong to the vitellogenin (VTG) family of lipid transfer proteins. MTP is essential for the intracellular assembly and secretion of apoB-containing lipoproteins, the key intravascular lipid transport proteins in vertebrates. We report the predicted three-dimensional structure of the C-terminal lipid binding cavity of MTP, modeled on the crystal structure of the lamprey VTG gene product, lipovitellin. The cavity in MTP resembles those found in the intracellular lipid-binding proteins and bactericidal/permeability-increasing protein. Two conserved helices, designated A and B, at the entrance to the MTP cavity mediate lipid acquisition and binding. Helix A (amino acids 725-736) interacts with membranes in a manner similar to viral fusion peptides. Mutation of helix A blocks the interaction of MTP with phospholipid vesicles containing triglyceride and impairs triglyceride binding. Mutations of helix B (amino acids 781-786) and of N780Y, which causes abetalipoproteinemia, have no impact on the interaction of MTP with phospholipid vesicles but impair triglyceride binding. We propose that insertion of helix A into lipid membranes is necessary for the acquisition of neutral lipids and that helix B is required for their transfer to the lipid binding cavity of MTP.

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The microsomal triglyceride transfer protein (MTP) complexed to protein disulphide isomerase (PDI) is obligatory for the assembly of chylomicrons and very-low-density lipoproteins. The determination of the atomic structure of the MTP-PDI heterodimer has important implications for the treatment of those forms of hyperlipidaemia associated with the overproduction of very-low-density lipoproteins, which predispose to premature coronary heart disease. To perform structural studies of the human MTP-PDI complex it was necessary to produce milligram quantities of pure protein. We chose the baculovirus expression system for this purpose. Insects cells were co-infected with recombinant viruses encoding FLAG-tagged MTP and His-tagged PDI; the resulting heterodimer was purified by affinity chromatography. From 5 litres of insect cells, 4-6 mg of more than 95% pure recombinant protein was obtained. CD and attenuated total reflection Fourier-transform infrared spectroscopy indicate that the purified protein has around 34% alpha-helical and 33% beta-structure content. The recombinant protein had a comparable triglyceride transfer activity to that of bovine MTP-PDI. The production of polyclonal antibodies raised against the MTP and PDI subunits of the purified protein is described. The present study demonstrates the feasibility of expressing two proteins at high levels in insect cells and describes a transferable methodology for the purification of the resulting protein complex.

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The assembly of triglyceride-rich lipoproteins requires the formation in the endoplasmic reticulum of a complex between apolipoprotein B (apoB), a microsomal triglyceride transfer protein (MTP), and protein disulfide isomerase (PDI). In the MTP complex, the amino-terminal region of MTP (residues 22-303) interacts with the amino-terminal region of apoB (residues 1-264). Here, we report the identification and characterization of a site on apoB between residues 512 and 721, which interacts with residues 517-603 of MTP. PDI binds in close proximity to this apoB binding site on MTP. The proximity of these binding sites on MTP for PDI and amino acids 512-721 of apoB was evident from studies carried out in a yeast two-hybrid system and by co-immunoprecipitation. The expression of PDI with MTP and apoB16 (residues 1-721) in the baculovirus expression system reduced the amount of MTP co-immunoprecipitated with apoB by 73%. The interaction of residues 512-721 of apoB with MTP facilitates lipoprotein production. Mutations of apoB that markedly reduced this interaction also reduced the level of apoB-containing lipoprotein secretion.

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The assembly of atherogenic lipoproteins requires the formation in the endoplasmic reticulum of a complex between apolipoprotein (apo)B, a microsomal triglyceride transfer protein (MTP) and protein disulphide isomerase (PDI). Here we show by molecular modelling and mutagenesis that the globular amino-terminal regions of apoB and MTP are closely related in structure to the ancient egg yolk storage protein, vitellogenin (VTG). In the MTP complex, conserved structural motifs that form the reciprocal homodimerization interfaces in VTG are re-utilized by MTP to form a stable heterodimer with PDI, which anchors MTP at the site of apoB translocation, and to associate with apoB and initiate lipid transfer. The structural and functional evolution of the VTGs provides a unifying scheme for the invertebrate origins of the major vertebrate lipid transport system.

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An immunohistochemical method that exploits the rapid light-evoked expression of Fos, the protein product of the immediate early gene, c-fos, to visualize eye-related columns in the visual cortex, has been used to provide preliminary data on the relative innervation of the cortex by the two eyes of monocularly deprived kittens and the speed of the changes that occur afterward during reverse occlusion. In contrast to conventional anatomical techniques, the method allows both cellular resolution and documentation of the dimensions of eye-related columns through the depth of the cortex. In kittens monocularly deprived from near birth, Fos-immunoreactive neurones were observed in oval or circular patches, the size of which decreased as the duration of deprivation was increased from 4 to 6 weeks. Following reverse occlusion at 5 weeks of age, the size of the patches increased rapidly so that after 4 days their area had approximately tripled. In addition to providing possible insights into the anatomical underpinnings of the puzzling behavioural effects that occur following termination of short periods of reverse occlusion, the method can be used to investigate the temporal order of the anatomical effects of monocular deprivation in different cortical layers.

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An integrated curriculum was implemented in an upper division baccalaureate nursing program which required letter grades for clinical courses. The second semester clinical course involved student evaluation in four different settings. The original evaluation tool lacked both the discrimination needed for letter grades and the competencies common to all the settings. A computerized evaluation tool was developed to identify behaviors which could be evaluated and assigned a letter grade in all four clinical settings, and to provide formative and summative evaluation of performance for students throughout the semester. The tool focuses on the four areas of the nursing process, each of which is weighted according to the conceptual framework of the curriculum and the ability of a student at this level. In each of these four areas, entry level behaviors were identified and then built upon by progression from fundamental skills to more complex and independent behaviors. Students are rated on a criterion-referenced rating scale. A statistician verified that the tool was mathematically sound. Once developed, the tool was placed on computer where both students and faculty evaluate student clinical performance every two to four weeks. Because the tool was placed on computer, students receive immediate feedback, which facilitates formative evaluations. Trends in student performance can be identified easily, and faculty paperwork is decreased. The tool also promotes objectivity in evaluation and student awareness of expected behaviors. Finally, faculty and students have become more familiar and more comfortable with computers.

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