Silencing of AEBP1 in U87MG glial cells and Chip-chIP with AEBP1 antibody
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Ladha Jayashree
PROVIDER: S-ECPF-GEOD-18892 | biostudies-other |
REPOSITORIES: biostudies-other
Similar Datasets
Project description:AEBP1 has been identified as a transcriptional repressor playing a significant role in adipogenesis. Recently, we have found that AEBP1 is specifically upregulated in primary glioblastoma. We have used gene silencing to identify genes perturbed upon AEBP1 downregulation and ChIP-chip technique to identify its genomic targets of binding and regulation to understand the un-annotated/undescribed role of this transcription factor in gliomagenesis. Cells either mock silenced or silenced using siRNA against AEBP1 were harvested at 46hrs post transfection. Gene expression was profiled for control and AEBP1 silenced cells. comparison of AEBP1/Rabbit IgG/ Glial cells
2012-04-29 | E-GEOD-18892 | biostudies-arrayexpress
Project description:AEBP1 has been identified as a transcriptional repressor playing a significant role in adipogenesis. Recently, we have found that AEBP1 is specifically upregulated in primary glioblastoma. We have used gene silencing to identify genes perturbed upon AEBP1 downregulation and ChIP-chip technique to identify its genomic targets of binding and regulation to understand the un-annotated/undescribed role of this transcription factor in gliomagenesis.
2012-04-30 | GSE18892 | GEO
Project description:Tumor cell-derived extracellular vesicles and their cargo of bioactive substances have gradually been recognized as novel biomarkers for cancer diagnosis. Meanwhile, the PD-L1 (Programmed Death-Ligand 1) protein, as an immune checkpoint molecule, is highly expressed on certain tumor cells and holds significant potential in immune therapy. In comparison to PD-L1 monoclonal antibodies, the inhibitory effect of PD-L1 siRNA (small interfering RNA) is more advantageous. In this article, we introduced a microfluidic chip integrating cell cultivation and exosome detection modules, which were intended for the investigation of the gene silencing effect of PD-L1 siRNA. Basically, cells were first cultured with PD-L1 siRNA in the chip. Then, the secreted exosomes were detected via super-resolution imaging, to validate the inhibitory effect of siRNA on PD-L1 expression. To be specific, a "sandwich" immunological structure was employed to detect exosomes secreted from HeLa cells. Immunofluorescence staining and DNA-PAINT (DNA Point Accumulation for Imaging in Nanoscale Topography) techniques were utilized to quantitatively analyze the PD-L1 proteins on HeLa exosomes, which enabled precise structural and content analysis of the exosomes. Compared with other existing PD-L1 detection methods, the advantages of our work include, first, the integration of microfluidic chips greatly simplifying the cell culture, gene silencing, and PD-L1 detection procedures. Second, the utilization of DNA-PAINT can provide an ultra-high spatial resolution, which is beneficial for exosomes due to their small sizes. Third, qPAINT could allow quantitative detection of PD-L1 with better precision. Hence, the combination of the microfluidic chip with DNA-PAINT could provide a more powerful integrated platform for the study of PD-L1-related tumor immunotherapy.
| S-EPMC10781284 | biostudies-literature
Project description:Immunoassay is one of the important applications of microfluidic chips and many methodologies were reported for decreasing sample∕reagent volume, shortening assay time, and so on. Micro-enzyme-linked immunosorbent assay (micro-ELISA) is our method that utilizes packed microbeads in the microfluidic channel and the immunoreactions are induced on the beads surface. Due to the large surface-to-volume ratio and small analytical volume, excellent performances have been verified in assay time and sample∕reagent volume. In order to realize the micro-ELISA, one of the important processes is the immobilization of antibody on the beads surface. Previously, the immobilization process was performed in a macroscale tube by physisorption of antibody, and long time (2 h) and large amount of antibody (or high concentration) were required for the immobilization. In addition, the processes including the reaction and washing were laborious, and changing the analyte was not easy. In this research, we integrated the immobilization process into a microfluidic chip by applying the avidin-biotin surface chemistry. The integration enabled very fast (1 min) immobilization with very small amount of precious antibody consumption (100 ng) for one assay. Because the laborious immobilization process can be automatically performed on the microfluidic chip, ELISA method became very easy. On-demand immunoassay was also possible just by changing the antibodies without using large amount of precious antibodies. Finally, the analytical performance was investigated by measuring C-reactive protein and good performance (limit of detection <20 ng∕ml) was verified.
| S-EPMC2967239 | biostudies-other
Project description:Ouabain (OUA) is a cardiac glycoside that binds to Na+,K+-ATPase (NKA), a conserved membrane protein that controls cell transmembrane ionic concentrations and requires ATP hydrolysis. At nM concentrations, OUA activates signaling pathways that are not related to its typical inhibitory effect on the NKA pump. Activation of these signaling pathways protects against some types of injury of the kidneys and central nervous system. There are 4 isoforms of the alpha subunit of NKA, which are differentially distributed across tissues and may have different physiological roles. Glial cells are important regulators of injury and inflammation in the brain and express the α1 and α2 NKA isoforms. This study investigated the role of α2 NKA in OUA modulation of the neuroinflammatory response induced by lipopolysaccharide (LPS) in mouse primary glial cell cultures. LPS treatment increased lactate dehydrogenase release, while OUA did not decrease cell viability and blocked LPS-induced NF-κB activation. Silencing α2 NKA prevented ERK and NF-κB activation by LPS. α2 NKA also regulates TNF-α and IL-1β levels. The data reported here indicate a significant role of α2 NKA in regulating central LPS effects, with implications in the associated neuroinflammatory processes.
| S-EPMC5501845 | biostudies-literature
Project description:The technique of chromatin immunoprecipitation (ChIP) is a powerful method for identifying in vivo DNA binding sites of transcription factors and for studying chromatin modifications. Unfortunately, the large number of cells needed for the standard ChIP protocol has hindered the analysis of many biologically interesting cell populations that are difficult to obtain in large numbers. New ChIP methods involving the use of carrier chromatin have been developed that allow the one-gene-at-a-time analysis of very small numbers of cells. However such methods are not useful if the resultant sample will be applied to genomic microarrays or used in ChIP-sequencing assays. Therefore, we have miniaturized the ChIP protocol such that as few as 10,000 cells (without the addition of carrier reagents) can be used to obtain enough sample material to analyze the entire human genome. We demonstrate the reproducibility of this MicroChIP technique using 2.1 million feature high-density oligonucleotide arrays and antibodies to RNA polymerase II and to histone H3 trimethylated on lysine 27 or lysine 9.
| S-EPMC2268896 | biostudies-literature
Project description:The Yin Yang 2 (YY2) gene encodes a zinc finger transcription factor that is not well characterized, yet. By using chromatin immunoprecipitations combined with whole-genome human promoter microarray (ChIP-chip) in HEK293 cells, we identified a multiplicity of YY2-bound annotated promoters as well as additional chromosomal regions. Interestingly, gene ontology analyses linked YY2 to fundamental biological pathways associated to cancer and developmental processes. Identification of YY2 target genes in HEK293 cells in vivo
2013-06-16 | E-GEOD-22599 | biostudies-arrayexpress
Project description:The chromatin remodeller p400 has oncogenic properties that promote early steps of colon cancer. Chromatin immunoprecipitation (ChIP) of p400 together with chromatin profiling by ChIP-on-chip analysis demonstrated that p400 directly binds the chromatin at promoters of p400 target genes. Study of p400 binding on promoters of HCT116 cells
2013-11-22 | E-GEOD-45614 | biostudies-arrayexpress
Project description:BackgroundThe quest to understand the neurobiology of schizophrenia and bipolar disorder is ongoing with multiple lines of evidence indicating abnormalities of glia, mitochondria, and glutamate in both disorders. Despite high heritability estimates of 81% for schizophrenia and 75% for bipolar disorder, compelling links between findings from neurobiological studies, and findings from large-scale genetic analyses, are only beginning to emerge.MethodTen publically available gene sets (pathways) related to glia, mitochondria, and glutamate were tested for association to schizophrenia and bipolar disorder using MAGENTA as the primary analysis method. To determine the robustness of associations, secondary analyses were performed with: ALIGATOR, INRICH, and Set Screen. Data from the Psychiatric Genomics Consortium (PGC) were used for all analyses. There were 1,068,286 SNP-level p-values for schizophrenia (9,394 cases/12,462 controls), and 2,088,878 SNP-level p-values for bipolar disorder (7,481 cases/9,250 controls).ResultsThe Glia-Oligodendrocyte pathway was associated with schizophrenia, after correction for multiple tests, according to primary analysis (MAGENTA p = 0.0005, 75% requirement for individual gene significance) and also achieved nominal levels of significance with INRICH (p = 0.0057) and ALIGATOR (p = 0.022). For bipolar disorder, Set Screen yielded nominally and method-wide significant associations to all three glial pathways, with strongest association to the Glia-Astrocyte pathway (p = 0.002).ConclusionsConsistent with findings of white matter abnormalities in schizophrenia by other methods of study, the Glia-Oligodendrocyte pathway was associated with schizophrenia in our genomic study. These findings suggest that the abnormalities of myelination observed in schizophrenia are at least in part due to inherited factors, contrasted with the alternative of purely environmental causes (e.g. medication effects or lifestyle). While not the primary purpose of our study, our results also highlight the consequential nature of alternative choices regarding pathway analysis, in that results varied somewhat across methods, despite application to identical datasets and pathways.
| S-EPMC3933626 | biostudies-literature
Project description:We demonstrate a rapid and highly sensitive all-electronic technique based on the resistive pulse method of particle sizing with a pore to detect the binding of unlabeled antibodies to the surface of latex colloids. Here, we use an on-chip pore to sense colloids derivatized with streptavidin and measure accurately their diameter increase on specific binding to several different types of antibodies. We show the sensitivity of this technique to the concentration of free antibody and that it can be used to perform immunoassays in both inhibition and sandwich configurations. Overall, our technique does not require labeling of the reactants and is performed rapidly by using very little solution, and the pore itself is fabricated quickly and inexpensively by using soft lithography. Finally, because this method relies only on the volume of bound ligand, it can be generally applied to detecting a wide range of ligand-receptor binding reactions.
| S-EPMC298685 | biostudies-literature