Comment[ArrayExpressAccession] E-GEOD-12161 Public Release Date 2010-03-04 Investigation Title Transcriptome profiling of control and TNFalpha treated HepG2 cells Comment[Submitted Name] Transcriptome profiling of control and TNFalpha treated HepG2 cells Experiment Description The proinflammatory cytokine, TNFalpha is critical in maintaining liver homeostasis since it is a major determiner of hepatocyte life and death. Considering this, gene transcription profiling was examined in control and TNFalpha treated HepG2 cells. Results indicated that TNFalpha could significantly alter the expression of a significant number of genes; most of them were functionally distributed among molecular functions like catalytic activity, binding, molecular transducer activity, transporter activity, translation and transcription regulator activities or enzyme regulator activity. Also, within genes up-regulated by TNFalpha, several GO terms related to lipid and fat metabolism were significantly overrepresented indicating global dysregulation of fat metabolism within the hepatocyte and those within the down-regulated dataset included genes involved in immunoglobulin receptor activity and IgE binding thereby indicating a compromise in immune defense mechanism(s) apart from those involved the DNA binding and protein binding categories. The interacting network of “lipid metabolism, small molecule biochemistry” was derived to be significantly affected that correlated well with the top canonical pathway of “biosynthesis of steroids” and molecular and cellular function of “lipid metabolism”. All these indicate TNFalpha to be significantly altering the transcriptome profiling within HepG2 cells with genes involved in lipid and steroid metabolism being the most favoured. This study suitably addresses the genes that determine TNFalpha mediated alterations within the hepatocyte mainly the phenotypes of hepatic steatosis and fatty liver that are associated with several hepatic pathological states. HepG2 cells were maintained in DMEM supplemented with 10% fetal calf serum with 1% antibiotic-antimycotic. On attaining confluency, cells were serum starved overnight and incubated in the absence (control) and presence of TNFalpha (0.5nM, 12h). On termination of incubation, total RNA was isolated, reverse transcribed to cDNA and subjected to in vitro transcription to produce biotinylated cRNA that was hybridized to the human array chip (Human Genome U133 Plus 2.0, Affymetrix) according to the manufacturer's instructions. Images were scanned using the GeneChip 3000 7G scanner (Affymetrix) and analysed. The experiment was performed with triplicates of each set (Control and TNF alpha treated). This experiment was reloaded in November 2010 after additional curation. Comment[AEExperimentDisplayName] Transcription profiling by array of human hepatocellular carcinoma cell line HepG2 treated with tumor necrosis factor-alpha Comment[AEExperimentType] transcription profiling by array Date of Experiment Person Last Name Datta Munjal Pandey Datta Person First Name Malabika Neha Amit Malabika Person Mid Initials K Person Email mdatta@igib.res.in Person Affiliation Institute of Genomics and Integrative Biology Person Phone Person Fax Person Address Institute of Genomics and Integrative Biology, Mall Road, Delhi, India Person Roles submitter Protocol Name P-GSE12161-2 P-GSE12161-1 P-GSE12161-6 P-GSE12161-5 P-GSE12161-3 P-GSE12161-8 P-GSE12161-4 P-GSE12161-7 Protocol Description ID_REF =
VALUE = MAS5-calculated Signal intensity
ABS_CALL = Presence or Absence of gene transcript in sample
DETECTION P-VALUE = significance value for detection of absolute calls ID_REF =
VALUE = MAS5-calculated Signal intensity
ABS_CALL = This indicates whether a transcript is reliably detected (P) or not detected (A)
DETECTION P-VALUE = 'detection p-value', p-value is the significance level of the detection call The genechips were washed thoroughly after hybridisation and scanned using the Affymetrix GeneChip Scanner 3000 7G. The labelled cRNA obtained was fragmented and 15 microg of cRNA were hybridized to the Affymetrix HG-U133 plus 2.0 chip according to the protocols described by the manufacturer. HepG2 cells were grown to confluency and total RNA was isolated using Trizol (Invitrogen, CA, USA) according to the manufacturers instructions. The RNA so obtained was purified using the RNeasy Mini-kit (Qiagen, Germany) HepG2 cells were grown to confluency and total RNA was isolated using Trizol (Invitrogen, CA, USA) according to the manufacturers instructions. The RNA so obtained was purified using the RNeasy Mini-kit (Qiagen, Germany) Biotinylated cRNA were prepared from 6 microg total RNA using the GeneChip Expression IVT Labelling kit from Affymetrix (USA) according to the manufacturers instructions. The data was normalised with MAS 5.0 using the GCOS Software from Affymetrix using default settings. The normalization target was set to 500. Protocol Software Protocol Hardware Protocol Contact Protocol Type bioassay_data_transformation bioassay_data_transformation image_aquisition hybridization nucleic_acid_extraction nucleic_acid_extraction labeling feature_extraction Protocol Term Source REF Protocol Term Accession Number Experimental Factor Name compound dose time Experimental Factor Type compound dose time Experimental Factor Term Source REF Experimental Factor Term Accession Number Publication Title Gene expression profiling and network analysis reveals lipid and steroid metabolism to be the most favored by TNFalpha in HepG2 cells. Publication Author List Pandey AK, Munjal N, Datta M PubMed ID 20140224 Publication DOI 10.1371/journal.pone.0009063 Publication Status published Publication Status Term Source REF EFO Publication Status Term Accession Number EFO_0001796 Comment[SecondaryAccession] GSE12161 Comment[GEOLastUpdateDate] 2010-03-04 Comment[GEOReleaseDate] 2010-03-04 Comment[ArrayExpressSubmissionDate] 2008-07-17 Term Source Name EFO Term Source File http://www.ebi.ac.uk/efo SDRF File E-GEOD-12161.sdrf.txt