Firearm suicide in Queensland

The purpose of this article is to examine firearm suicide in Queensland. In 2006, statistical data were gathered from all closed paper coronial files for the 12-month period of December 2003—December 2004. Of the 567 people who committed suicide in Queensland during this period, 48 (8.5%) used firearms. The following results emerge from this data: first, gun suicides are continuing to decrease in Queensland, most likely as a function of ongoing gun controls, a decrease accompanied by a lesser increase in other methods of suicide, thereby providing little support for substitution theory; second, men continue to be more likely to shoot themselves, particularly elderly men; third, firearms are more likely to be used in rural settings, and by those with no known history of mental illness or previous suicide attempts. Finally, in spite of otherwise very high suicide rates, Aborigines rarely employ firearms, using instead the culturally significant method of hanging.

The factor V HR2 haplotype: Prevalence and association of the A4070G and A6755G polymorphisms

Recently, a polymorphism was identified in exon 25 of the factor V gene that is possibly a functional candidate for the HR2 haplotype. This haplotype is characterized by a single base substitution named R2 (A4070G) in the B domain of the protein. A mutation (A6755G; 2194Asp→Gly) located near the C terminus has been hypothesized to influence protein folding and glycosylation, and might be responsible for the shift in factor V isoform (FV1 / FV2) ratio. This study investigated the prevalence of these two factor V HR2 haplotype polymorphisms in a cohort of normal blood donors, patients with osteoarthritis and women with complications during pregnancy, and in families of factor V Leiden individuals. A high allele frequency for the two polymorphisms was found in the blood donor group (6.2% R2, 5.6% A6755G). No significant difference in allele frequency was observed in the clinical groups (obstetric complications and osteoarthritis, 4.1-4.9% for the two polymorphisms) when compared with that of healthy blood donors. We confirm that the factor V A6755G polymorphism shows strong linkage to the R2 allele, although it is not exclusively inherited with the exon 13 A4070G variant and can occur independently. © 2001 Lippincott Williams & Wilkins.

Multiple analysis of three common genetic alterations associated with thrombophilia

We have previously reported the use of a novel mini-sequencing protocol for detection of the factor V Leiden variant, the first nucleotide change (FNC) technology. This technology is based on a single nucleotide extension of a primer, which is hybridized immediately adjacent to the site of mutation. The extended nucleotide that carries a reporter molecule (fluorescein) has the power to discriminate the genotype at the site of mutation. More recently, the prothrombin 20210 and thermolabile methylene tetrahydrofolate reductase (MTHFR) 677 variants have been identified as possible risk factors associated with thrombophilia. This study describes the use of the FNC technology in a combined assay to detect factor V, prothrombin and MTHFR variants in a population of Australian blood donors, and describes the objective numerical methodology used to determine genotype cut-off values for each genetic variation. Using FNC to test 500 normal blood donors, the incidence of Factor V Leiden was 3.6% (all heterozygous), that of prothrombin 20210 was 2.8% (all heterozygous) and that of MTHFR was 10% (homozygous). The combined FNC technology offers a simple, rapid, automatable DNA-based test for the detection of these three important mutations that are associated with familial thrombophilia. (C) 2000 Lippincott Williams and Wilkins.

Use of first nucleotide change technology to determine the frequency of factor V Leiden in a population of Australian blood donors

Activated protein C resistance (APCR), the most common risk factor for venous thrombosis, is the result of a G to A base substitution at nucleotide 1691 (R506Q) in the factor V gene. Current techniques to detect the factor V Leiden mutation, such as determination of restriction length polymorphisms, do not have the capacity to screen large numbers of samples in a rapid, cost- effective test. The aim of this study was to apply the first nucleotide change (FNC) technology, to the detection of the factor V Leiden mutation. After preliminary amplification of genomic DNA by polymerase chain reaction (PCR), an allele-specific primer was hybridised to the PCR product and extended using fluorescent terminating dideoxynucleotides which were detected by colorimetric assay. Using this ELISA-based assay, the prevalence of the factor V Leiden mutation was determined in an Australian blood donor population (n = 500). A total of 18 heterozygotes were identified (3.6%) and all of these were confirmed with conventional MnlI restriction digest. No homozygotes for the variant allele were detected. We conclude from this study that the frequency of 3.6% is compatible with others published for Caucasian populations. In addition, the FNC technology shows promise as the basis for a rapid, automated DNA based test for factor V Leiden.

Generation of profluorescent isoindoline nitroxides using click chemistry

Novel profluorescent nitroxides bearing a triazole linker between the coumarin fluorophore and an isoindoline nitroxide were prepared in good yields using the coppercatalyzed azide�alkyne 1,3-dipolar cycloaddition reaction (CuAAC). Nitroxides containing 7-hydroxy and 7-diethylamino substitution on their coumarin rings displayed significant fluorescence suppression, and upon reaction with methyl radicals, normal fluorescence emission was returned. The fluorescence emission for the 7-hydroxycoumarin nitroxide and its diamagnetic analogue was found to be strongly influenced by pH with maximal fluorescence emission achieved in basic solution. Solvent polarity was also shown to affect fluorescence emission. The significant difference in fluorescence output between the nitroxides and their corresponding diamagnetic analogues makes these compounds ideal tools for monitoring processes involving free-radical species.

Cloning and characterization of a novel human gene related to vascular endothelial growth factor

This paper describes the cloning and characterization of a new member of the vascular endothelial growth factor (VEGF) gene family, which we have designated VRF for VEGF-related-factor. Sequencing of cDNAs from a human fetal brain library and RT-PCR products from normal and tumor tissue cDNA pools indicate two alternatively spliced messages with open reading frames of 621 and 564 bp, respectively. The predicted proteins differ at their carboxyl ends resulting from a shift in the open reading frame. Both isoforms show strong homology to VEGF at their amino termini, but only the shorter isoform maintains homology to VEGF at its carboxyl terminus and conserves all 16 cysteine residues of VEGF165. Similarity comparisons of this isoform revealed overall protein identity of 48% and conservative substitution of 69% with VEGF189. VRF is predicted to contain a signal peptide, suggesting that it may be a secreted factor. The VRF gene maps to the D11S750 locus at chromosome band 11q13, and the protein coding region, spanning approximately 5 kb, is comprised of 8 exons that range in size from 36 to 431 bp. Exons 6 and 7 are contiguous and the two isoforms of VRF arise through alternate splicing of exon 6. VRF appears to be ubiquitously expressed as two transcripts of 2.0 and 5.5 kb; the level of expression is similar among normal and malignant tissues.

Analysis of the CDKN2A, CDKN2B and CDK4 genes in 48 Australian melanoma kindreds

Germline mutations within the cyclin-dependent kinase inhibitor 2A (CDKN2A) gene and one of its targets, the cyclin dependent kinase 4 (CDK4) gene, have been identified in a proportion of melanoma kindreds. In the case of CDK4, only one specific mutation, resulting in the substitution of a cysteine for an arginine at codon 24 (R24C), has been found to be associated with melanoma. We have previously reported the identification of germline CDKN2A mutations in 7/18 Australian melanoma kindreds and the absence of the R24C CDK4 mutation in 21 families lacking evidence of a CDKN2A mutation. The current study represents an expansion of these efforts and includes a total of 48 melanoma families from Australia. All of these families have now been screened for mutations within CDKN2A and CDK4, as well as for mutations within the CDKN2A homolog and 9p21 neighbor, the CDKN2B gene, and the alternative exon 1 (E1beta) of CDKN2A. Families lacking CDKN2A mutations, but positive for a polymorphism(s) within this gene, were further evaluated to determine if their disease was associated with transcriptional silencing of one CDKN2A allele. Overall, CDKN2A mutations were detected in 3/30 (10%) of the new kindreds. Two of these mutations have been observed previously: a 24 bp duplication at the 5' end of the gene and a G to C transversion in exon 2 resulting in an M531 substitution. A novel G to A transition in exon 2, resulting in a D108N substitution was also detected. Combined with our previous findings, we have now detected germline CDKN2A mutations in 10/48 (21%) of our melanoma kindreds. In none of the 'CDKN2A-negative' families was melanoma found to segregate with either an untranscribed CDKN2A allele, an R24C CDK4 mutation, a CDKN2B mutation, or an E1beta mutation. The last three observations suggest that these other cell cycle control genes (or alternative gene products) are either not involved at all, or to any great extent, in melanoma predisposition.

High frequency of BRAF mutations in nevi

To evaluate the timing of mutations in BRAF (v-raf murine sarcoma viral oncogene homolog B1) during melanocytic neoplasia, we carried out mutation analysis on microdissected melanoma and nevi samples. We observed mutations resulting in the V599E amino-acid substitution in 41 of 60 (68%) melanoma metastases, 4 of 5 (80%) primary melanomas and, unexpectedly, in 63 of 77 (82%) nevi. These data suggest that mutational activation of the RAS/RAF/MAPK pathway in nevi is a critical step in the initiation of melanocytic neoplasia but alone is insufficient for melanoma tumorigenesis.

Microarray expression profiling in melanoma reveals a BRAF mutation signature

We have used microarray gene expression profiling and machine learning to predict the presence of BRAF mutations in a panel of 61 melanoma cell lines. The BRAF gene was found to be mutated in 42 samples (69%) and intragenic mutations of the NRAS gene were detected in seven samples (11%). No cell line carried mutations of both genes. Using support vector machines, we have built a classifier that differentiates between melanoma cell lines based on BRAF mutation status. As few as 83 genes are able to discriminate between BRAF mutant and BRAF wild-type samples with clear separation observed using hierarchical clustering. Multidimensional scaling was used to visualize the relationship between a BRAF mutation signature and that of a generalized mitogen-activated protein kinase (MAPK) activation (either BRAF or NRAS mutation) in the context of the discriminating gene list. We observed that samples carrying NRAS mutations lie somewhere between those with or without BRAF mutations. These observations suggest that there are gene-specific mutation signals in addition to a common MAPK activation that result from the pleiotropic effects of either BRAF or NRAS on other signaling pathways, leading to measurably different transcriptional changes.

Diels-Alder reactions as an efficient route to high purity cyclic polymers

A simple and efficient route for the synthesis of cyclic polymer systems is presented. Linear furan protected α-maleimide-ω-cyclopentadienyl functionalized precursors (poly(methyl methacrylate) and poly(tert-butyl acrylate)) were synthesized via atom transfer radical polymerization (ATRP) and subsequent substitution of the bromine end-group with cyclopentadiene. Upon heating at high dilution, deprotection of the dieneophile occurs followed by an intramolecular Diels–Alder reaction yielding a high purity cyclic product.

Raman spectroscopy of selected tsumcorite Pb(Zn,Fe3+)2(AsO4)2(OH,H2O) minerals –implications for arsenate accumulation

The presence of arsenic in the environment is a hazard. The accumulation of arsenate by a range of cations in the formation of minerals provides a mechanism for the accumulation of arsenate. The formation of the tsumcorite minerals is an example of a series of minerals which accumulate arsenate. There are about twelve examples in this mineral group. Raman spectroscopy offers a method for the analysis of these minerals. The structure of selected tsumcorite minerals with arsenate and sulphate anions were analysed by Raman spectroscopy. Isomorphic substitution of sulphate for arsenate is observed for gartrellite and thometzekite. A comparison is made with the sulphate bearing mineral natrochalcite. The position of the hydroxyl and water stretching vibrations are related to the strength of the hydrogen bond formed between the OH unit and the AsO43- anion. Characteristic Raman spectra of the minerals enable the assignment of the bands to specific vibrational modes.