| 1. |
- Russom, Aman, et al.
(author)
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Microfluidic leukocyte isolation for gene expression analysis in critically ill hospitalized patients
- 2008
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In: Clinical Chemistry. - : Oxford University Press (OUP). - 0009-9147 .- 1530-8561. ; 54:5, s. 891-900
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Journal article (peer-reviewed)abstract
- BACKGROUND: Microarray technology is becoming a powerful tool for diagnostic, therapeutic, and prognostic applications. There is at present no consensus regarding the optimal technique to isolate nucleic acids from blood leukocyte populations for subsequent expression analyses. Current collection and processing techniques pose significant challenges in the clinical setting. Here, we report the clinical validation of a novel microfluidic leukocyte nucleic acid isolation technique for gene expression analysis from critically ill, hospitalized patients that can be readily used on small volumes of blood. METHODS: We processed whole blood from hospitalized patients after burn injury and severe blunt trauma according to the microfluidic and standard macroscale leukocyte isolation protocol. Side-by-side comparison of RNA quantity, quality, and genome-wide expression patterns was used to clinically validate the microfluidic technique. RESULTS: When the microfluidic protocol was used for processing, sufficient amounts of total RNA were obtained for genome-wide expression analysis from 0.5 mL whole blood. We found that the leukocyte expression patterns from samples processed using the 2 protocols were concordant, and there was less variability introduced as a result of harvesting method than there existed between individuals. CONCLUSIONS: The novel microfluidic approach achieves leukocyte isolation in <25 min, and the quality of nucleic acids and genome expression analysis is equivalent to or surpasses that obtained from macroscale approaches. Microfluidics can significantly improve the isolation of blood leukocytes for genomic analyses in the clinical setting. (c) 2008 American Association for Clinical Chemistry.
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| 2. |
- Finnerty, N., et al.
(author)
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Increased brain nitric oxide levels following ethanol administration
- 2015
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In: Nitric oxide. - : Elsevier BV. - 1089-8603 .- 1089-8611. ; 47, s. 52-57
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Journal article (peer-reviewed)abstract
- Nitric oxide is a ubiquitous messenger molecule, which at elevated concentrations has been implicated in the pathogenesis of several neurological disorders. Its role in oxidative stress, attributed in particular to the formation of peroxynitrite, proceeds through its high affinity for the superoxide radical. Alcoholism has recently been associated with the induction of oxidative stress, which is generally defined as a shift in equilibrium between pro-oxidant and anti-oxidant species in the direction of the former. Furthermore, its primary metabolite acetaldehyde, has been extensively associated with oxidative damage related toxic effects following alcohol ingestion. The principal objective of this study was the application of long term in vivo electrochemistry (LIVE) to investigate the effect of ethanol (0.125, 0.5 and 2.0 g kg-1) and acetaldehyde (12.5, 50 and 200 mg kg-1) on NO levels in the nucleus accumbens of freely moving rats. Systemic administrations of ethanol and acetaldehyde resulted in a dose-dependent increases in NO levels, albeit with very differing time courses. Subsequent to this the effect on accumbal NO levels, of subjecting the animal to different drug combinations, was also elucidated. The nitric oxide synthase inhibitor L-NAME (20 mg kg-1) and acetaldehyde sequestering agent D-penicillamine (50 mg kg-1) both attenuated the increase in NO levels following ethanol (1 g kg-1) administration. Conversely, the alcohol dehydrogenase inhibitor 4-methylpyrazole (25 mg kg-1) and catalase inhibitor sodium azide (10 mg kg-1) potentiated the increase in NO levels following ethanol administration. Finally, dual inhibition of aldehyde dehydrogenase and catalase by cyanamide (25 mg kg-1) caused an attenuation of ethanol effects on NO levels. Taken together these data highlight a robust increase in brain NO levels following systemic alcohol administration which is dependent on NO synthase activity and may involve both alcohol- and acetaldehyde-dependent mechanisms. © 2015 Elsevier Inc. All rights reserved.
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| 3. |
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| 4. |
- Finnerty, N. J., et al.
(author)
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Brain nitric oxide: Regional characterisation of a real-time microelectrochemical sensor
- 2012
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In: Journal of Neuroscience Methods. - : Elsevier BV. - 0165-0270. ; 209:1, s. 13-21
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Journal article (peer-reviewed)abstract
- A reliable method of directly measuring endogenously generated nitric oxide (NO) in real-time and in various brain regions is presented. An extensive characterisation of a previously described amperometric sensor has been carried out in the prefrontal cortex and nucleus accumbens of freely moving rats. Systemic administration of saline caused a transient increase in signal from baseline levels in both the prefrontal cortex (13 +/- 3 pA, n = 17) and nucleus accumbens (12 +/- 3 pA, n = 8). NO levels in the prefrontal cortex were significantly increased by 43 +/- 9 pA (n = 9) following administration of L-arginine. A similar trend was observed in the nucleus accumbens, where an increase of 44 +/- 9 pA (n = 8) was observed when compared against baseline levels. Systemic injections of the non-selective NOS inhibitor L-NAME produced a significant decrease in current recorded in the prefrontal cortex (24 +/- 6 pA, n = 5) and nucleus accumbens (17 +/- 3 pA, n = 6). Finally it was necessary to validate the sensors functionality in vivo by investigating the effect of the interferent ascorbate on the oxidation current. The current showed no variation in both regions over the selected time interval of 60 min, indicating no deterioration of the polymer membrane. A detailed comparison identified significantly greater affects of administrations on NO sensors implanted in the striatum than those inserted in the prefrontal cortex and the nucleus accumbens. (c) 2012 Elsevier B.V. All rights reserved.
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