Analysis of gene expression in multiple sclerosis lesions using cDNA microarrays

Whitney, Laurie Ward; Becker, Kevin G.; Tresser, Nancy; Caballero-Ramos, Carla I.; Munson, Peter J.; Prabhu, Vinayakumar; Trent, Jeffrey M.; McFarland, Henry F.; Biddison, William E.

doi:10.1002/1531-8249(199909)46:3<425::aid-ana22>3.0.co;2-o

Cited by 222 publications

(118 citation statements)

References 19 publications

(2 reference statements)

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“…Radioactive cDNA Probe Preparation and Microarray Hybridization-cDNA probe preparation and microarray hybridization were performed as described previously (20). Briefly, 5 g of total RNA was reverse-transcribed in a reaction mixture containing 8 l of 5ϫ first strand RT buffer, 1 l of 1 g/l 12-18-mer poly(dT) primer, 4 l of 2 mM dNTPs (ϪdCTP), 4 l of 0.1 M dithiothreitol, 1 l (40 units) of RNase-OUT, 6 l of 3000 Ci/mmol [␣-…”

Section: Methodsmentioning

confidence: 99%

Growth Factor Signals in Neural Cells

Martin

Brenneman

Golden

et al. 2009

Journal of Biological Chemistry

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Individual neurons express receptors for several different growth factors that influence the survival, growth, neurotransmitter phenotype, and other properties of the cell. Although there has been considerable progress in elucidating the molecular signal transduction pathways and physiological responses of neurons and other cells to individual growth factors, little is known about if and how signals from different growth factors are integrated within a neuron. In this study, we determined the interactive effects of nerve growth factor, insulin-like growth factor 1, and epidermal growth factor on the activation status of downstream kinase cascades and transcription factors, cell survival, and neurotransmitter production in neural cells that express receptors for all three growth factors. We document considerable differences in the quality and quantity of intracellular signaling and eventual phenotypic responses that are dependent on whether cells are exposed to a single or multiple growth factors. Dual stimulations that generated the greatest antagonistic or synergistic actions, compared with a theoretically neutral summation of their two activities, yielded the largest eventual change of neuronal phenotype indicated by the ability of the cell to produce norepinephrine or resist oxidative stress. Combined activation of insulin-like growth factor 1 and epidermal growth factor receptors was particularly notable for antagonistic interactions at some levels of signal transduction and norepinephrine production, but potentiation at other levels of signaling and cytoprotection. Our findings suggest that in true physiological settings where multiple growth factors are present, activation of one receptor type may result in molecular and phenotypic responses that are different from that observed in typical experimental paradigms in which cells are exposed to only a single growth factor at a time.Numerous growth factors have been identified that influence the survival and plasticity of neurons, including the neurotrophins (NGF, 2 brain-derived neurotrophic factor, NT-3, and NT-4), basic fibroblast growth factor, insulin-like growth factor 1 (IGF-1), and epidermal growth factor (EGF). These different factors activate receptors with intrinsic tyrosine kinase activity, which then engage downstream kinase cascades, resulting in the activation of transcription factors. For example, NGF activates the TrkA receptor that engages the Raf-mitogen-activated protein kinase kinase (MEK)-extracellular signal-regulated kinase (ERK) pathway resulting in activation of transcription factors, including AP1 (1-3); IGF-1 activates the IGF-1 receptor that is coupled to phosphatidylinositol trisphosphate kinase, Akt kinase, and forkhead transcription factors of the FOXO family (4, 5); and EGF receptors engage the Raf-MEK-ERK pathway (6, 7). Many neural cells express receptors for multiple growth factors, each of which induce similar effects on the survival, differentiation, and function of the cells. Although there has been considerable progress ...

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Section: Methodsmentioning

confidence: 99%

Growth Factor Signals in Neural Cells

Martin

Brenneman

Golden

et al. 2009

Journal of Biological Chemistry

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“…The quantity and quality of RNA were measured by spectrophotometry and electrophoresis on denaturing agarose gel. Radiolabeled cDNA probes were synthesized from 10 g total RNA in the presence of dATP, dTTP, dGTP, 33 P dC TP, polyT, Superscript II RNase H Ϫ Reverse Transcriptase, and RNaseOUT Recombinant Inhibitor, as described previously (Whitney et al, 1999). The probes were purified using a SP-30 column and quantified using a liquid scintillation counter.…”

Section: Drugsmentioning

confidence: 99%

Changes in Gene Expression Linked to Methamphetamine-Induced Dopaminergic Neurotoxicity

Xie¹,

Tong²,

Barrett

et al. 2002

J. Neurosci.

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The purpose of these studies was to examine the role of gene expression in methamphetamine (METH)-induced dopamine (DA) neurotoxicity. First, the effects of the mRNA synthesis inhibitor, actinomycin-D, and the protein synthesis inhibitor, cycloheximide, were examined. Both agents afforded complete protection against METH-induced DA neurotoxicity and did so independently of effects on core temperature, DA transporter function, or METH brain levels, suggesting that gene transcription and mRNA translation play a role in METH neurotoxicity. Next, microarray technology, in combination with an experimental approach designed to facilitate recognition of relevant gene expression patterns, was used to identify gene products linked to METH-induced DA neurotoxicity. This led to the identification of several genes in the ventral midbrain associated with the neurotoxic process, including genes for energy metabolism [cytochrome c oxidase subunit 1 (COX1), reduced nicotinamide adenine dinucleotide ubiquinone oxidoreductase chain 2, and phosphoglycerate mutase B], ion regulation (members of sodium/hydrogen exchanger and sodium/bile acid cotransporter family), signal transduction (adenylyl cyclase III), and cell differentiation and degeneration (N-myc downstreamregulated gene 3 and tau protein). Of these differentially expressed genes, we elected to further examine the increase in COX1 expression, because of data implicating energy utilization in METH neurotoxicity and the known role of COX1 in energy metabolism. On the basis of time course studies, Northern blot analyses, in situ hybridization results, and temperature studies, we now report that increased COX1 expression in the ventral midbrain is linked to METH-induced DA neuronal injury. The precise role of COX1 and other genes in METH neurotoxicity remains to be elucidated. Key words: amphetamines; neurotoxicity; dopamine; neurodegeneration; cytochrome c oxidase; microarrayDespite considerable investigation (Gibb et al., 1994;Lew et al., 1997), the mechanisms underlying the neurotoxic effects of methamphetamine (METH) on brain dopamine (DA) neurons remain unknown. However, a considerable body of data attests to the importance of DA transporter (DAT) function and temperature. The essential role of the DAT in METH-induced DA neurotoxicity has been demonstrated by the observation that DAT inhibitors afford complete neuroprotection (Ricaurte et al., 1984; Marek et al., 1990a,b;Pu et al., 1994) and the finding that DAT knock-out mice are insensitive to METH neurotoxicity (Fumagalli et al., 1998). The importance of temperature has been documented by studies demonstrating that decreases in core temperature protect against METH neurotoxicity, whereas increases in core temperature exacerbate the toxicity (Bowyer et al., 1994;Miller and O'Callaghan, 1994;Albers and Sonsalla, 1995;Ali et al., 1996). Furthermore, there is evidence that at least part of the effect of temperature on METH neurotoxicity is mediated at the level of the DAT (Xie et al., 2000). More recent studies have also i...

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“…Accurate and reproducible data can be obtained, provided a standardized system of brain collection, dissection, storage and case matching is employed in conjunction with the consideration of key ante-and post-mortem factors. Techniques in use in our and other laboratories range from quantitative competitive RT-PCR (Cummings et al 2001;Becker et al 2002;Guan et al 2002;Pandey et al 2002), western blotting (Lewohl et al 2001;Mori et al 2002), cDNA microarray (Whitney et al 1999;Lockhart and Barlow 2001;Goldstine et al 2002;Mayfield et al 2002;Tanwar et al 2002), and proteomic assays of unfixed tissue (Cheon et al 2001;Davidsson et al 2001;Husi and Grant 2001;Pasinetti 2001), through to RT-in situ hybridization histochemistry (Schindler et al 1995;Wakeman et al 1997;Hawkins and Dodd 2000;Goldstine et al 2002) and immunohistochemistry on fixed sections (Sparks et al 1998;Cerf and Raidoo 2000;Cervera et al 2002).…”

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confidence: 99%

Biochemical and molecular studies using human autopsy brain tissue

Hynd

Lewohl

Scott

et al. 2003

Journal of Neurochemistry

221

171

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The use of human brain tissue obtained at autopsy for neurochemical, pharmacological and physiological analyses is reviewed. RNA and protein samples have been found suitable for expression profiling by techniques that include RT-PCR, cDNA microarrays, western blotting, immunohistochemistry and proteomics. The rapid development of molecular biological techniques has increased the impetus for this work to be applied to studies of brain disease. It has been shown that most nucleic acids and proteins are reasonably stable postmortem. However, their abundance and integrity can exhibit marked intra-and intercase variability, making comparisons between case-groups difficult. Variability can reveal important functional and biochemical information. The correct interpretation of neurochemical data must take into account such factors as age, gender, ethnicity, medicative history, immediate ante-mortem status, agonal state and post-mortem and post-autopsy intervals. Here we consider issues associated with the sampling of DNA, RNA and proteins using human autopsy brain tissue in relation to various ante-and postmortem factors. We conclude that valid and practical measures of a variety of parameters may be made in human brain tissue, provided that specific factors are controlled.

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Analysis of gene expression in multiple sclerosis lesions using cDNA microarrays

Cited by 222 publications

References 19 publications

Growth Factor Signals in Neural Cells

Growth Factor Signals in Neural Cells

Changes in Gene Expression Linked to Methamphetamine-Induced Dopaminergic Neurotoxicity

Biochemical and molecular studies using human autopsy brain tissue

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