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David J Halsall

Publications and source records attributed to David J Halsall.

10 recordsLinked to original sources
PubMed2006

Looking at long molecules in solution: what happens when they are subjected to Couette flow?

Knowing the structure of a molecule is one of the keys to deducing its function in a biological system. However, many biomacromolecules are not amenable to structural characterisation by the powerful techniques often used namely NMR and X-ray diffraction because they are too large, or too flexible or simply refuse to crystallize. Long molecules such as DNA and fibrous proteins are two such classes of molecule. In this article the extent to which flow linear dichroism (LD) can be used to characterise the structure and function of such molecules is reviewed. Consideration is given to the issues of fluid dynamics and light scattering by such large molecules. A range of applications of LD are reviewed including (i) fibrous proteins with particular attention being given to actin; (ii) a far from comprehensive discussion of the use of LD for DNA and DNA-ligand systems; (iii) LD for the kinetics of restriction digestion of circular supercoiled DNA; and (iv) carbon nanotubes to illustrate that LD can be used on any long molecules with accessible absorption transitions.

Absorption↗
PubMed2005

Reduced vitamin D in acute stroke.

BACKGROUND AND PURPOSE: Stroke leads to a reduction in bone mineral density, altered calcium homeostasis, and an increase in hip fractures. Vitamin D deficiency is well documented in long-term stroke survivors and is associated with post-stroke hip fractures. Less is known regarding levels in acute stroke. METHODS: We compared the serum 25-dihydroxyvitamin D levels of 44 patients admitted to an acute stroke unit with first-ever stroke with results obtained by measuring 96 healthy ambulant elderly subjects every 2 months for 1 year. Statistical Z scores of serum vitamin D were then calculated after seasonal adjustment for the month of sampling. RESULTS: The mean Z score of vitamin D in acute stroke was -1.4 SD units (95% CI, -1.7, -1.1), with 77% of patients falling in the insufficient range. CONCLUSIONS: Reduced vitamin D was identified in the majority of patients with acute stroke throughout the year and may have preceded stroke. Vitamin D is a potential risk marker for stroke, and the role of vitamin D repletion in enhancing musculoskeletal health after stroke needs to be explored.

Aged↗
PubMed2005

Validation of new microvolume Couette flow linear dichroism cells.

Long molecules such as fibrous proteins are particularly difficult to characterise structurally. We have recently designed a microvolume Couette flow linear dichroism (LD) cell whose sample volume is only 20-40 microL in contrast to previous cells where the volume of sample required has typically been of the order of 1000-2000 microL. This brings the sample requirements of LD to a level where it can be used for biological samples. Since LD is the difference in absorption of light polarised parallel to an orientation direction and perpendicular to that direction, it is the ideal technique for determining relative orientations of subunits of e.g. fibrous proteins, DNA-drug systems, etc. For solution phase samples, Couette flow orientation, whereby the sample is sandwiched between two cylinders, one of which rotates, has proved to be the optimal technique for LD experiments in many laboratories. Our capillary microvolume LD cell has been designed using extruded quartz rods and capillaries and focusing and collecting lenses. We have developed applications with PCR products, fibrous proteins, liposome-bound membrane proteins, as well as DNA-dye systems. Despite this range of applications, to date there is nothing reported in the literature to enable one to validate the performance of Couette flow LD cells. In this paper we establish validation criteria and show that the data from the microvolume cells are reproducible, vary by less than 1% with sample reloading, follow the Beer-Lambert law, and have signals linear in voltage over a wide voltage range. The microvolume cell data are consistent with those from the large-volume cells for DNA samples. Surprisingly, upon extending the wavelength range by adding the intercalator ethidium bromide, the spectra in the microvolume and large-volume cells differ by a wavelength dependent orientation parameter. This wavelength variation was concluded to be the result of Taylor-vortices in the large-volume cells which have inner rotating cylinders in our laboratory. Thus the microvolume LD cells can be concluded to provide better data than our large-volume LD cells, though the latter are still to be preferred for titration series as it is extremely difficult to add sample to the capillary cells without introducing artefacts.

DNA↗
PubMed2005

Primary iron overload with inappropriate hepcidin expression in V162del ferroportin disease.

Ferroportin disease (hemochromatosis type 4) is a recently recognized disorder of human iron metabolism, characterized by iron deposition in macrophages, including Kupffer cells. Mutations in the gene encoding ferroportin 1, a cellular iron exporter, are responsible for this iron storage disease, inherited as an autosomal dominant trait. We present clinical, histopathological, and radiological findings in a family with the most common ferroportin mutation, V162del. In the index case, the disorder is characterized by abundant deposition of hemosiderin in all tissues investigated (mesenteric lymph node, liver, gastric and duodenal mucosa, and also in squamous cell carcinoma of the lung). The radiological findings indicated the presence of excess iron in bone marrow and spleen. Despite a significant burden of iron, no features of chronic liver disease were found in affected members of the family, including individuals aged up to 80 years. Hyperferritinemia greater than 1,000 microg/L was a penetrant biochemical finding before the second decade in life and was associated with significantly increased serum concentrations of pro-hepcidin that correlated positively with urinary hepcidin concentrations. In conclusion, the systemic iron burden in ferroportin disease is not a sufficient cause for chronic liver disease. In patients with most, but not all, ferroportin mutations, retention of iron in macrophages of the liver and other organs may protect against damage to parenchymal cells. Finally, macrophage iron storage in ferroportin disease is associated with elevated serum pro-hepcidin levels.

Adolescent↗
PubMed2004

Protein fiber linear dichroism for structure determination and kinetics in a low-volume, low-wavelength couette flow cell.

High-resolution structure determination of soluble globular proteins relies heavily on x-ray crystallography techniques. Such an approach is often ineffective for investigations into the structure of fibrous proteins as these proteins generally do not crystallize. Thus investigations into fibrous protein structure have relied on less direct methods such as x-ray fiber diffraction and circular dichroism. Ultraviolet linear dichroism has the potential to provide additional information on the structure of such biomolecular systems. However, existing systems are not optimized for the requirements of fibrous proteins. We have designed and built a low-volume (200 microL), low-wavelength (down to 180 nm), low-pathlength (100 microm), high-alignment flow-alignment system (couette) to perform ultraviolet linear dichroism studies on the fibers formed by a range of biomolecules. The apparatus has been tested using a number of proteins for which longer wavelength linear dichroism spectra had already been measured. The new couette cell has also been used to obtain data on two medically important protein fibers, the all-beta-sheet amyloid fibers of the Alzheimer's derived protein Abeta and the long-chain assemblies of alpha1-antitrypsin polymers.

Actins↗
PubMed2004

Micro-volume couette flow sample orientation for absorbance and fluorescence linear dichroism.

Linear dichroism (LD) can be used to study the alignment of absorbing chromophores within long molecules. In particular, Couette flow LD has been used to good effect in probing ligand binding to DNA and to fibrous proteins. This technique has been previously limited by large sample requirements. Here we report the design and application of a new micro-volume Couette flow cell that significantly enhances the potential applications of flow LD spectroscopy by reducing the sample requirements for flow linear dichroism to 25 microL (with concentrations such that the absorbance maximum of the sample in a 1-cm pathlength cuvette is approximately 1). The micro-volume Couette cell has also enabled the measurement of fluorescence-detected Couette flow linear dichroism. This new technique enables the orientation of fluorescent ligands to be probed even when their electronic transitions overlap with those of the macromolecule and conversely. The potential of flow-oriented fluorescence dichroism and application of the micro-volume Couette LD cell are illustrated by the collection of data for DNA with minor groove and intercalating ligands: DAPI, Hoechst, and ethidium bromide. As with conventional fluorescence, improved sensitivity compared with absorbance LD is to be expected after instrumentation optimization.

Animals↗
PubMed2003

Typical type 2 diabetes mellitus and HFE gene mutations: a population-based case - control study.

Diabetes mellitus is a recognized consequence of hereditary haemochromatosis. Whether the common HFE mutations, that associate with this condition and pre-dispose to increases in serum iron indices, are over-represented in diabetic populations remains controversial. We present data from the largest case-control study of the C282Y and H63D HFE allele frequencies in typical type 2 diabetes mellitus, as defined by an age of onset greater than 30 years and no requirement for insulin in the first year post-diagnosis. We also present a meta-analysis of all similar studies to date. We see no evidence for over-representation of iron loading HFE alleles in type 2 diabetes mellitus, suggesting that screening for HFE mutations in this population is of no value.

Age of Onset↗
PubMed2003

Rapid genetic testing for Gaucher disease by reverse hybridization.

BACKGROUND: We evaluated a reverse hybridization method for the simultaneous detection of nine mutations that are associated with Gaucher disease. RESULTS AND CONCLUSION: Results were in agreement with those obtained by a polymerase chain reaction-restriction digestion method. The hybridization method produced results more quickly and with less operator input.

Gaucher Disease↗
PubMedDate not supplied

Hemochromatosis--neonatal and young subjects.

Juvenile hemochromatosis (JH) is an autosomal recessive disease causing iron overload before age 30 in both sexes. JH is characterised by hypogonadism, growth retardation and cardiomyopathy. Linkage of JH to chromosome lq is established in pedigrees throughout Europe. Studies of 29 patients in 20 families of diverse ethnic origin confirm early-onset iron overload. Neonatal hemochromatosis (NH) is a syndrome of unknown origin characterized by congenital cirrhosis or fulminant hepatitis with hepatic and extra-hepatic iron deposits. We assessed 40 infants from 27 families and identified 3 patterns of disease transmission. In 12 of the 27 there was >1 affected infant and in 5 families all infants were affected by NH. In 19 families unaffected children were also born. In 4 families there was bacterial or viral maternal infection associated with NH. In two families, antibodies to DNA or ribonuclear proteins were identified. In 12 families, unaffected children were born to the same parents in the absence of maternal antibodies or infection and without indications of maternal transmission. Consanguinity was observed in 1 family with 4 affected offspring (1 stillbirth + 3 neonatal deaths). Sequence analysis of HFE, beta2M, and both human heme oxygenase genes failed to identify any causal mutations in nuclear NH families but our study points to the existence of a cohort of patients likely to suffer from an autosomal recessive trait. A genome wide scanning study is underway to identify the putative locus.

Adult↗