Mitochondrial DNA haplogroups and susceptibility to AD and dementia with Lewy bodies.

The authors analyzed the relationship between nuclear genetic risk factors (apolipoprotein E genotype) and mitochondrial DNA (mtDNA) sequence variants in pathologically proved cases of AD (n = 185), dementia with Lewy bodies (DLB; n = 84), and control subjects (n = 179). Specific European mtDNA haplogroups and the A4336G mutation were not associated with an increased risk of AD. mtDNA haplogroup H was overrepresented in the DLB patients when compared with control subjects.

Investigation of the mechanism by which glucose analogues cause translocation of glucokinase in hepatocytes: evidence for two glucose binding sites.

Glucokinase translocates between the cytoplasm and nucleus of hepatocytes where it is bound to a 68 kDa protein. The mechanism by which glucose induces translocation of glucokinase from the nucleus was investigated using glucose analogues that are not phosphorylated by glucokinase. There was strong synergism on glucokinase translocation between effects of glucose analogues (glucosamine, 5-thioglucose, mannoheptulose) and sorbitol, a precursor of fructose 1-phosphate.

Impact of treatment provision on the epidemiological recording of root caries.

The estimation of root caries prevalence and the identification of risk factors for decay depend upon the successful identification of carious lesions in epidemiological studies. Root surface restorations can either be placed to manage decay or cervical wear/sensitivity. The handling of data for restorations during epidemiological surveys is critical to the accurate assessment of caries prevalence.

Mutagenesis of E477 or K505 in the B' domain of human topoisomerase II beta increases the requirement for magnesium ions during strand passage.

A type II topoisomerase is essential for decatenating DNA replication products, and it accomplishes this task by passing one DNA duplex through a transient break in a second duplex. The B' domain of topoisomerase II contains three highly conserved motifs, EGDSA, PL(R/K)GK(I/L/M)LNVR, and IMTD(Q/A)DXD. We have investigated these motifs in topoisomerase II beta by mutagenesis, and report that they play a critical role in establishing the DNA cleavage-religation equilibrium.

Monoclonal antibodies recognizing CD5, CD10 and CD23 in formalin-fixed, paraffin-embedded tissue: production and assessment of their value in the diagnosis of small B-cell lymphoma.

Aims: Assessment of the expression of antigens CD5, CD10 and CD23 can be of value in the differential diagnosis of small B-cell lymphoma. Correct subclassification is important since optimal treatment regimes differ between the subtypes. The aim of this study was to generate monoclonal antibodies recognizing these antigens in paraffin-embedded tissue and to assess their efficacy using a panel of cases of small B-cell lymphoma of various subtypes.

Peptide nucleic acid delivery to human mitochondria.

Peptide nucleic acids (PNAs) are synthetic polynucleobase molecules, which bind to DNA and RNA with high affinity and specificity. Although PNAs have enormous potential as anti-sense agents, the success of PNA-mediated gene therapy will require efficient cellular uptake and sub-cellular trafficking. At present these mechanisms are poorly understood.

The spectrum of hearing loss due to mitochondrial DNA defects.

Heteroplasmic mitochondrial DNA (mtDNA) defects are an important cause of neurological disease. Although hearing impairment is common in patients with mtDNA defects, the spectrum and pathophysiology of the hearing loss is not well characterized. We therefore studied the relationship between cochlear and brainstem auditory function in 23 patients harbouring a range of different mtDNA mutations.

A family 26 mannanase produced by Clostridium thermocellum as a component of the cellulosome contains a domain which is conserved in mannanases from anaerobic fungi.

Cellulosomes prepared by the cellulose affinity digestion method from Clostridium thermocellum culture supernatant hydrolysed carob galactomannan during incubation at 60 degrees C and pH 6.5. A recombinant phage expressing mannanase activity was isolated from a library of C.

Carbamate analogues of amsacrine active against non-cycling cells: relative activity against topoisomerases IIalpha and beta.

Purpose: Methyl N-(4'-(9-acridinylamino)-phenyl)carbamate hydrochloride (AMCA) and methyl N-(4'-(9-acridinylamino)-2-methoxyphenyl)carbamate hydrochloride (mAMCA) are analogues of the topoisomerase II (topo II) poison amsacrine, and are distinguished from amsacrine by their high cytotoxicity towards non-cycling cells. Since mammalian cells contain two forms (alpha and beta) of topo II and the alpha isoform is down-regulated in non-cycling cells, we have considered whether these carbamate analogues target topo IIbeta selectively.

Methods: A drug permeable yeast strain (JN394 top2-4) was transformed using a shuttle vector containing either human top2alpha, human top2alpha or yeast top2 under the control of a GAL1 promoter.

Mitochondrial DNA analysis: polymorphisms and pathogenicity.

The investigation of mtDNA disease can be relatively straightforward if a person has a recognisable phenotype and if it is possible to identify a known pathogenic mtDNA mutation. The difficulties arise when no known mtDNA defect can be found, or when the clinical abnormalities are complex and not easily matched to those of the more common mitochondrial disorders. We will describe here the difficulties that can be encountered during the identification of pathogenic mtDNA mutations and the approaches that can be used to confirm, or eliminate, a likely pathogenic role, in either single gene diseases or in multifactorial disorders.

Nonrandom tissue distribution of mutant mtDNA.

Heteroplasmic mitochondrial DNA (mtDNA) defects are an important cause of inherited human disease. On a cellular level, the percentage of mutant mtDNA is the principal factor behind the expression of the genetic defect. Marked variation in the level of mutant mtDNA among tissues is thought to be responsible for the diverse clinical phenotypes associated with the same pathogenic mtDNA mutation.

Long-term responses of the green-algal lichen Parmelia caperata to natural CO enrichment.

Acclimation to elevated CO was investigated in Parmelia caperata originating from the vicinity of a natural CO spring, where the average daytime CO concentration was 729 ± 39 μmol mol dry air. Thalli showed no evidence of a down-regulation in photosynthetic capacity following long-term exposure to CO enrichment in the field; carboxylation efficiency, total Ribulose bisphosphate carboxylase/oxygenase (Rubisco) content, apparent quantum yield of CO assimilation, and the light-saturated rate of CO assimilation (measured under ambient and saturating CO concentrations) were similar in thalli from the naturally CO enriched site and an adjacent control site where the average long-term CO concentration was about 355 μmol mol. Thalli from both CO environments exhibited low CO compensation points and early saturation of CO uptake kinetics in response to increasing external CO concentrations, suggesting the presence of an active carbon-concentrating mechanism.

Relaxed replication of mtDNA: A model with implications for the expression of disease.

Heteroplasmic mtDNA defects are an important cause of human disease with clinical features that primarily involve nondividing (postmitotic) tissues. Within single cells the percentage level of mutated mtDNA must exceed a critical threshold level before the genetic defect is expressed. Although the level of mutated mtDNA may alter over time, the mechanism behind the change is not understood.

Modulation of reperfusion injury after single lung transplantation by pentoxifylline, inositol polyanions, and sin-1.

Objective: Previous studies have suggested reductions in lung reperfusion injury with pentoxifylline, inositol polyanions, and the nitric oxide donor, SIN-1, but these agents have never been directly compared to ascertain which is superior. We investigated these agents in a porcine model of left single lung transplantation.

Methods: Donor lungs were preserved with modified Euro-Collins solution for a mean ischemic time of 18.

Human DNA topoisomerase IIbeta binds and cleaves four-way junction DNA in vitro.

We have used gel retardation analysis to show that human DNA topoisomerase IIbeta can bind a 40 bp linear duplex containing a single DNA topoisomerase IIbeta cleavage site. Furthermore, we demonstrate for the first time that human DNA topoisomerase IIbeta binds to four-way junction DNA. This supports previous suggestions that topoisomerase II may be targeted to supercoiled DNA through the recognition of DNA cruciforms, helix-helix crossovers and hairpins.

MELAS and MERRF. The relationship between maternal mutation load and the frequency of clinically affected offspring.

The majority of pathogenic mitochondrial DNA (mtDNA) mutations are heteroplasmic, with both mutant and wild-type alleles present within the same individual. MtDNA is transmitted only from females to their offspring but a single female can bear offspring who harbour different levels of mutant mtDNA and have a variable phenotype. In single families, this complex genetic and phenotypic variability has confounded the identification of any relationship between the level of mutant mtDNA (mutation load) in the mother and the clinical features of her offspring.

Displacement of OmpF loop 3 is not required for the membrane translocation of colicins N and A in vivo.

The pore-forming colicins N and A require the porin, OmpF, in order to translocate across the outer membrane of Escherichia coli. We investigated the hypothesis that in vivo, colicins N and A may traverse the outer membrane through the OmpF channel. In order to accommodate a polypeptide in the pore, the mid-channel constriction loop of OmpF, L3, would need to undergo a conformational change.

Comparative study of electric and manual toothbrushes in patients with fixed orthodontic appliances.

The objective of this three treatment, three period, single blind, cross-over trial was to evaluate the efficacy of three toothbrushes in a cohort of children undergoing fixed appliance orthodontic therapy. The brushes used were: (1) Dental Logic HP550 with regular brush head HP5924 (Philips, U.K.

A novel method for the visualization of the in situ mucus layer in rat and man.

1. The observed thickness of the gastric mucus barrier varies widely, even appearing discontinuous, depending on the methods used. Here we describe the development and application of a modified periodic acid Schiff/Alcian Blue staining technique for use on cryostat sections of gastric mucosa.

Etoposide targets topoisomerase IIalpha and IIbeta in leukemic cells: isoform-specific cleavable complexes visualized and quantified in situ by a novel immunofluorescence technique.

We have shown that both DNA topoisomerase (topo) IIalpha and beta are in vivo targets for etoposide using a new assay which directly measures topo IIalpha and beta cleavable complexes in individual cells after treatment with topo II targeting drugs. CCRF-CEM human leukemic cells were exposed to etoposide for 2 hr, then embedded in agarose on microscope slides before cell lysis. DNA from each cell remained trapped in the agarose and covalently bound topo II molecules from drug-stabilized cleavable complexes remained associated with the DNA.

Hexokinase isoenzymes in normal and cirrhotic human liver: suppression of glucokinase in cirrhosis.

The activities of hexokinase isoenzymes I-IV (EC 2.7.1.

Platinum-DNA adduct formation in leucocytes of children in relation to pharmacokinetics after cisplatin and carboplatin therapy.

Platinum (Pt)-DNA adducts were measured in peripheral blood leucocytes (PBLs) from 24 children with solid tumours after standard cisplatin and/or carboplatin treatment. The relationship between Pt-DNA adduct levels and pharmacokinetics of cisplatin and carboplatin was investigated. Adduct measurements were performed by competitive enzyme-linked immunosorbent assay (ELISA) and plasma unbound Pt concentrations were measured by atomic absorption spectrophotometry (AAS).

Wortmannin is a potent inhibitor of DNA double strand break but not single strand break repair in Chinese hamster ovary cells.

Wortmannin, an inhibitor of p110 PI 3-kinase, also inhibits DNA-dependent protein kinase, which is known to mediate DNA double strand break repair. It was recently demonstrated that wortmannin sensitized cells to ionizing radiation (IR) (Price and Youmell, Cancer Res., 56, 246-250, 1996).

Ultra-specific immunoassays for small molecules: roles of wash steps and multiple binding formats.

New immunometric forms of immunoassay are much more flexible to use than competitive-format immunoassays for small molecular analytes. An example of the utility of this flexibility is the ability to wash the capture antibody after it has been exposed to analyte but before addition of the labeled reagent. This simple maneuver has a large impact on the specificity obtained from already highly specific assays.

The use of an unenclosed field fumigation system to determine the effects of elevated ozone on a grass-clover mixture.

A mixture of Trifolium repens L. (var. Grasslands Huia) and Lolium perenne L.