ORCID Profile
0000-0003-1417-1313
Current Organisation
Massey University - Wellington Campus
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Publisher: Elsevier BV
Date: 02-2012
Abstract: TRIzol is used for RNA isolation but also permits protein recovery. We investigated whether proteins prepared with TRIzol were suitable for two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization mass spectrometry. Proteins from TRIzol-treated SH-SY5Y cells produced 2-DE spot patterns similar to those from an equivalent untreated s le. Subsequent identification of TRIzol-treated proteins using peptide mass fingerprinting was successful. TRIzol exposure altered neither the mass of myoglobin extracted from sodium dodecyl sulfate (SDS) gels nor the masses of myoglobin peptides produced by in-gel trypsin digestion. These findings suggest that proteins isolated with TRIzol remain amenable to proteomic analyses.
Publisher: Elsevier BV
Date: 2012
Publisher: Wiley
Date: 28-04-2009
Abstract: The dinoflagellate metabolite yessotoxin (YTX) is produced by several species of algae and accumulates in marine food chains, leading to concerns about possible affects on aquaculture industries and human health. In mice used for toxicity testing, YTX is lethal by the intraperitoneal route, but is considerably less toxic when orally administered. The mode of action of YTX and its potential effect on humans is unclear and we therefore conducted the first proteomic analysis of the effects of this compound. We used 2-DE to examine protein changes in HepG2 cell cultures exposed to 1.4 microM YTX for 3, 12.5, 18 and 24 h. After selecting proteins that changed more than three-fold after YTX exposure, 55 spots were deemed significantly affected by the toxin (p<0.05). Major groups of affected proteins include members from the heterogeneous nuclear ribonucleoprotein (hnRNP), lamin, cathepsin and heat shock protein families that often are associated with apoptosis. We therefore confirmed apoptosis using Annexin-V-FLUOS staining of phosphatidylserine exposed at the surface of apoptotic cells. Ingenuity pathways analysis also indicated effects on pathways involved in protein processing, cell cycling and cell death.
Publisher: Oxford University Press (OUP)
Date: 08-2007
DOI: 10.1080/14622200701485117
Abstract: The effect of total particulate matter (TPM) from cigarette smoke on the expression and binding properties of nicotinic acetylcholine receptors (nAChRs) was investigated using a human neuroblastoma cell line (SH-SY5Y). TPM but not nicotine on its own inhibited cell growth at nicotine concentrations above 5 microM. To examine effects on nAChR expression, intact cells were incubated with 3H-epibatidine, and a Bmax of 13 fmoles/10(5) cells (7.8 x 10(4) binding sites/cell) was measured in unexposed cells as well as in cells treated with 2 microM nicotine alone or with TPM containing 2 microM nicotine. Using Scatchard analysis, we measured a Kd of 0.3 nM for 3H-epibatidine binding to nAChRs. This Kd was increased to 1.3 nM by addition of nicotine or TPM extract, both at 2 microM nicotine. Bmax, however, was unaffected, suggesting competitive binding of nicotine to its receptor. Short-term and prolonged 3-day exposures of SH-SY5Y cells to either TPM or nicotine at nicotine concentrations ranging from 0.2 microM to 20 microM increased specific binding, suggesting upregulation of nAChR expression. Most significant, binding was consistently greater in cells pretreated with TPM than in cells pretreated with nicotine. We conclude that TPM contains compounds that are toxic to cells at high concentrations (cell growth inhibition) but that do not compete with nicotine for binding to nAChRs (Scatchard analysis). These non-nicotinic compounds are capable of increasing the expression of one or more of the nAChR subunits. Furthermore, our cell culture assay provides a useful in vitro model for assessing the relative addictiveness of different tobacco products, including that of non-nicotine components.
Publisher: SAGE Publications
Date: 27-02-2009
Abstract: Smoking is one of the leading causes of preventable death, where nicotine has been identified as the primary addictive constituent of tobacco. Consequently, there have been extensive investigations into the neuroadaptations that occur as nicotine dependence develops, where numerous neurological systems have been implicated. The focus of this review was on nicotinic acetylcholine receptor neuroadaptations that occur during the development of nicotine dependence. This focus was selected because (1) the nicotinic receptors are the primary binding sites for both nicotine and the most efficacious pharmacological smoking cessation treatments and (2) the receptors are located throughout the brain with considerable neuromodulatory ability. However, there was difficulty associated in outlining the role of nicotinic receptors in the development of nicotine dependence because it comprises a series of stages involving different neurological systems rather than a single state. To address this issue, the review adopts a novel approach and considers the role of nicotinic receptor subtypes at separate stages of the nicotine dependence cycle. This information was then used to examine the nicotinic receptor-related therapeutic mechanisms of three main pharmacological smoking cessation treatments.
No related grants have been discovered for Penelope Truman.