Microarray analysis of gene expression profiles in human neuroblastoma SH-SY5Y cells exposed to norephedrine
ABSTRACT: To reveal the molecular mechanism underling necrotic neuronal cell death caused by norephedrine, we examined alteration of gene expression profile during norephedrine exposure in human neuroblastoma SH-SY5Y cells. The alteration of gene expression during norephedrine exposure (3 mM, 0,2 and 6 hours) in differentiated SH-SY5Y cells was examined.
Project description:Multiple genetic and environmental factors play a role in the development and progression of Parkinson’s disease (PD). The main neuropathological hallmark of PD is the degeneration of dopaminergic (DAergic) neurons in the substantia nigra pars compacta (SN). To study genetic and molecular contributors to the disease process, there is a great need for readily accessible cells with prominent DAergic features that can be used for reproducible in vitro cellular screening. Here, we investigated the molecular phenotype of retinoic acid (RA) differentiated SH-SY5Y cells using genome wide transcriptional profiling combined with gene ontology, transcription factor and molecular pathway analysis. We demonstrated that RA induces a general neuronal differentiation program in SH-SY5Y cells and that these cells develop a predominantly mature DAergic-like neurotransmitter phenotype. This phenotype is characterized by increased dopamine levels together with substantial suppression of other neurotransmitter phenotypes, such as those for noradrenaline, acetylcholine, glutamate, serotonin and histamine. In addition, we show that RA differentiated SH-SY5Y cells express dopamine and noradrenalin neurotransmitter transporters that are responsible for uptake of MPP(+), a well known DAergic cell toxicant, and that MPP(+) treatment alters mitochondrial activity according to its proposed cytotoxic effect in DAergic neurons. Taken together, RA differentiated SH-SY5Y cells have a DAergic-like phenotype, and provide a good cellular screening tool to find novel genes or compounds that affect cytotoxic processes that are associated with PD. SH-SY5Y cell differentiation process was assessed by comparing RA differentiated and noRA differentiated cells in 8 days of culture. Cells were compared at 6 different time points.
Project description:This SuperSeries is composed of the following subset Series:; GSE16656: Transcriptome analysis identifies molecular effectors of unconjugated bilirubin in human neuroblatoma SH-SY5Y cells: 24h; GSE16766: Transcriptome analysis identifies molecular effectors of unconjugated bilirubin in human neuroblastoma SH-SY5Y cells: 1h; GSE16767: Transcriptome analysis identifies molecular effectors of unconjugated bilirubin in human neuroblastoma SH-SY5Y cells: 4h Experiment Overall Design: Refer to individual Series
Project description:Recombinant PGRN and GRN peptides were used to treat fully differentiated SH-SY5Y cells. Post-treatment cells were lysed and used for RNA Seq using the ion-Torrent proton platform. Results were validated using real time PCR.
Project description:BACKGROUND: SH-SY5Y cells exhibit a neuronal phenotype when treated with all-trans retinoic acid (RA), but the molecular mechanism of activation in the signalling pathway mediated by phosphatidylinositol 3-kinase (PI3K) is unclear. To investigate this mechanism, we compared the gene expression profiles in SK-N-SH cells and two subtypes of SH-SY5Y cells (SH-SY5Y-A and SH-SY5Y-E), each of which show a different phenotype during RA-mediated differentiation. FINDINGS: SH-SY5Y-A cells differentiated in the presence of RA, whereas RA-treated SH-SY5Y-E cells required additional treatment with brain-derived neurotrophic factor (BDNF) for full differentiation. After exposing cells to a PI3K inhibitor, LY294002, we identified 386 genes and categorised these genes into two clusters dependent on the PI3K signalling pathway during RA-mediated differentiation in SH-SY5Y-A cells. Transcriptional regulation of the gene cluster, including 158 neural genes, was greatly reduced in SK-N-SH cells and partially impaired in SH-SY5Y-E cells, which is consistent with a defect in the neuronal phenotype of these cells. Additional stimulation with BDNF induced a set of neural genes that were down-regulated in RA-treated SH-SY5Y-E cells but were abundant in differentiated SH-SY5Y-A cells. CONCLUSION: We identified gene clusters controlled by PI3K- and TRKB-mediated signalling pathways during the differentiation of two subtypes of SH-SY5Y cells. The TRKB-mediated bypass pathway compensates for impaired neural function generated by defects in several signalling pathways, including PI3K in SH-SY5Y-E cells. Our expression profiling data will be useful for further elucidation of the signal transduction-transcriptional network involving PI3K or TRKB.
Project description:Many physiological functions of hydrogen sulfide (H2S) have been reported in mammalian cells over the last 20 years. These physiological effects have been ascertained through in vitro treatment of cells with Na2S or NaHS, both of which are precursors of H2S. Since H2S exists as HS- in a neutral solution, a disulfide compound such as cystine could react with HS- in culture medium as well as in the cell. This study demonstrated that after the addition of Na2S solution into culture medium, HS- was transiently generated and disappeared immediately through the reaction between HS- and cystine to form cysteine persulfides and polysulfides in the culture medium (bound sulfur mixture: BS-Mix). Furthermore, we found that the addition of Na2S solution resulted in an increase of intracellular cysteine persulfide levels in SH-SY5Y cells. This alteration in intracellular persulfide was also observed in cystine-free medium. Considering this reaction of HS- as a precursor of BS-Mix, we highlighted the cytoprotective effect of Na2S on human neuroblastoma SH-SY5Y cells against methylglyoxal (MG)-induced toxicity. BS-Mix produced with Na2S in cystine-containing medium provided SH-SY5Y cells significant protective effect against MG-induced toxicity. However, the protective effect was attenuated in cystine-free medium. Moreover, we observed that Na2S or BS-Mix activated the Keap1/Nrf2 system and increased glutathione (GSH) levels in the cell. In addition, the activation of Nrf2 is significantly attenuated in cystine-free medium. These results suggested that Na2S protects SH-SY5Y cells from MG cytotoxicity through the activation of Nrf2, mediated by cysteine persulfides and polysulfides that were generated by Na2S addition.
Project description:Cold exposure leads to alteration in the structure of the male sex chromosome of the mutant strain In(1)BM2(reinverted). Genome wide expression profiling was used to identify candidate genes involved in the expression of this phenotype. Adult male flies were exposed to cold shock at 12±1°C for 4 hours and the differentially expressed genes in the strain was compared to similarly exposed wild type Oregon R males. The microarray data was further validated using real time PCR. Two-condition experiment, RT vs. CS flies. Biological replicates: 2 control replicates, 2 replicates exposed to cold shock.
Project description:?-Synuclein is highly associated with some neurodegeneration and malignancies. Overexpressing wild-type or mutant ?-synuclein promotes neuronal death by mitochondrial dysfunction, the underlying mechanisms of which remain poorly defined. It was recently reported that ?-synuclein expression could directly lead to mitochondrial fragmentation in vitro and in vivo, which may be due to ?-synuclein localization on mitochondria. Here, we applied a double staining method to demonstrate mitochondrial morphogenetic changes in cells overexpressed with ?-synuclein. We show that mitochondrial localization of ?-synuclein was increased following its overexpression in three distinct cell lines, including HeLa, SH-SY5Y, and PC12 cells, but no alteration in mitochondrial morphology was detected. However, ?-synuclein knockdown prevents MPP(+)-induced mitochondrial fragmentation in SH-SY5Y and PC12 cells. These data suggest that ?-synuclein protein levels hardly affect mitochondrial morphology in normal cell lines, but may have some influence on that under certain environmental conditions.
Project description:Whole-genome profiling of SH-SY5Y cells was done on neuroblastoma SH-SY5Y stably transfected with cDNAs coding for SOD1WT or the mutant SOD1(G93A) protein. Five wt SOD versus five mutant SOD