Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of human MDA-MB-453 breast cancer cell lines treated with actein to assess effects on the growth as a function of time and concentration.

ABSTRACT: Previous studies indicate that the triterpene glycoside actein from the herb black cohosh inhibits growth of human breast cancer cells. This study seeks to identify genes altered in human breast cancer cells by treatment with actein, using gene expression analysis. We treated MDA-MB-453 human breast cancer cells with actein at 2 doses, 20 or 40 Î¼g/mL, for 6 or 24 h. We identified 5 genes that were activated after each of the treatments that are known to play a role in cellular responses to diverse stresses, including the DNA damage and unfolded protein responses. In addition, four genes that mediate the integrated stress response (ISR), including activating transcription factor 4, were induced under at least one of the 4 treatment conditions. We used hierarchical clustering to define clusters comprising patterns of gene expression. Two ISR genes, activating transcription factor 3 (ATF3) and DNA damage- inducible transcript 3, and lipid biosynthetic genes were activated after exposure to actein at 40 Î¼g/mL for 6 h, whereas the cell cycle genes cyclin E2 and cell division cycle 25A were repressed. Our results suggest that actein induces 2 phases of the ISR, the survival phase and the apoptotic phase, depending on the dose and duration of treatment. We confirmed the results of gene expression analysis with real-time RT-PCR for 18 selected genes and Western blot analysis for ATF3. Since actein activated transcription factors that enhance apoptosis, and repressed cell cycle genes, it may be useful in the prevention and therapy of breast cancer. Experiment Overall Design: This study used Affymetrix U133A 2.0 gene chips. We analyzed 3 replicate cultures after treating MDA-MB-453 cells with actein at 20 or 40 mg/mL for 24 h and 2 replicate cultures after treating MDA-MB-453 cells with actein at 20 or 40 mg/mL for 6 h. All samples were normalized to remove chip-dependent regularities using the GCRMA method of Irizarry et al . Chips and controls at each combination of actein concentration and time were normalized together. The statistical significance of differential expression was calculated using the empirical Bayesian LIMMA (LInear Model for MicroArrays) method of Smyth et al. A cut-off B>0 was used for the statistical significance of gene expression.

ORGANISM(S): Homo sapiens

SUBMITTER: Linda Saxe Einbond

PROVIDER: E-GEOD-7848 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

ACCESS DATA

Similar Datasets

Project description:The experiment is aiming to analyze the effect of overexpression of the Arabidopsis At1g06160 AP2/ERF-domain transcription factor on gene expression and the involvement of the At1g06160-regulated genes in the jasmonic acid or jasmonic acid/ethylene signal transduction pathways. The experiment was performed with 12 hybridizations, using 24 samples of species [Arabidopsis thaliana], using 12 arrays of array design [Agilent Arabidopsis 3 Oligo Microarray [G4142A]], producing 12 raw data files and 0 transformed and/or normalized data files. In the first two hybridizations (#1 and #2), two independent transgenic Arabidopsis lines containing a construct carrying the Arabidopsis gene At1g06160 fused to an inducible promoter were used. This promoter is activated after the addition of the chemical estradiol to the plant medium. Addition of estradiol results in overexpression of At1g06160. For each independent line, total RNA from estradiol-treated and -untreated plants were collected and labeled with Cy3 and Cy5, respectively, and hybridization was performed for each At1g06160-inducible line. Two other hybridizations (#3 and #4) were performed using two independent transgenic Arabidopsis control lines containing a construct carrying the GUS gene fused to an inducible promoter. For each independent line, total RNA from estradiol-treated and -untreated plants were collected and labeled with Cy3 and Cy5, respectively and hybridization was performed for each GUS-inducible line. The last 8 hybridizations were performed using wild-type Arabidopsis plants. The aim was analyze the effect of the jasmonic acid hormone or the effect of a combination of the jasmonic acid and ethylene hormones on gene expression in wild-type. Hybridizations #5 and #6: wild-type Arabidopsis seedlings were treated with jasmonic acid (JA) or with the control DMSO and total RNA was collected after 8 hours. Total RNA from JA-treated and DMSO-treated plants were collected and labeled with Cy3 and Cy5, respectively, and hybridization was performed. A second hybridization was performed using independent biological samples that were treated and labeled similarly. Hybridizations #7 and #8: wild-type Arabidopsis seedlings were treated with jasmonic acid (JA) or with the control DMSO and total RNA was collected after 24 hours. Total RNA from JA-treated and DMSO-treated plants were collected and labeled with Cy3 and Cy5, respectively, and hybridization was performed. A second hybridization was performed using independent biological samples that were treated and labeled similarly. Hybridizations #9 and #10: wild-type Arabidopsis seedlings were treated with jasmonic acid (JA) and ethylene or with the control DMSO/Na-phosphate and total RNA was collected after 8 hours. Total RNA from JA/ethylene-treated and DMSO/Na-phosphate-treated plants were collected and labeled with Cy3 and Cy5, respectively, and hybridization was performed. A second hybridization was performed using independent biological samples that were treated and labeled similarly. Hybridizations #11 and #12: wild-type Arabidopsis seedlings were treated with jasmonic acid (JA) and ethylene or with the control DMSO/Na-phosphate and total RNA was collected after 24 hours. Total RNA from JA/ethylene-treated and DMSO/Na-phosphate-treated plants were collected and labeled with Cy3 and Cy5, respectively, and hybridization was performed. A second hybridization was performed using independent biological samples that were treated and labeled similarly.

Project description:Dioxins are ubiquitous environmental poisons that are developmentally toxic. Exposure of mouse E14 tooth germs to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) leads to reduced tooth size and deformation of cuspal morphology implying the induction of a variety of biological responses on both cellular and molecular levels. To verify such responses at the gene level, mouse embryonic cap staged tooth germs were cultured for 24 h with/without 1 ÂµM TCDD. Experiment Overall Design: Organ culture: Mandibular molar tooth regions from E14 mouse embryos (NMRI X NMRI; the day of vaginal plug = E0) were dissected under a stereomicroscope and transferred to an organ culture. The tissue explants were supported by polycarbonate Nuclepore filters [pore size, 0.1 Âµm; Corning Inc., New York] placed on a stainless steel grid. The basal culture medium was Dulbeccoâs modified Eagleâs medium (DMEM) containing Glutamax-1 (Gibco BRL, Paisley, Scotland) supplemented with 10% fetal calf serum (FCS; Gibco BRL). TCDD (from the Department of Chemistry, University of JyvÃ¤skylÃ¤, JyvÃ¤skylÃ¤, Finland) was dissolved in dimethylsulfoxide (DMSO) at a concentration of 50 Âµg/ml. TCDD at the concentration of 1 ÂµM, or DMSO, was added to the basal culture medium from the start of culture. The culture dishes were kept in a humidified incubator at 37 ÂºC in an atmosphere of 5% CO2. The tissue explants were cultured for 24 hours, after which they were snap frozen in liquid nitrogen. Explants from several experiments were collected and kept in liquid nitrogen. In a single experiment, 15â29 explants were included in each experimental group. Experiment Overall Design: Microarray analysis: Experiment Overall Design: Three separate experiments were combined for each replicate sample. About 60 explants each were pooled and homogenized while still frozen in liquid nitrogen. Total RNA was isolated using the RNeasy Mini Kit (Qiagen). cDNA was synthesized from the total RNA by using a GeneChip T7-Oligo(dT)24 Promoter Primer Kit (Affymetrix) and Superscript II Choice System (Invitrogen Life Sciences). The double stranded cDNA was cleaned-up with the GeneChip Sample Cleanup Module (Affymetrix). Biotin-labelled cRNA was synthesized by in vitro transcription usin a RNA labelling kit from Enzo Life Sciences, Inc. The labelling time was 5 h in 37ÂºC waterbath. All syntheses were done according to the manufacturers instructions. Unincorporated NTPs were removed with the GeneChip Sample Cleanup kit and the concentration of the labelled cRNA was mesured by spectrophotometric analysis and quantificated. 15 or 16 Âµg of labelled cRNA was fragmented by incubation at 94 ÂºC for 35 min in the fragmentation buffer supplied in the GeneChip Sample Cleanup kit. The cRNA was hybridized to a murine U74v2A array (Affymetrix) for 16 h at 45 ÂºC, washed and stained with streptavidine-phycoerythrin. The arrays were scanned in a Genechip System confocal scanner (Aligent) and Affymetrix Microarray Suite 5.0 was used to scan and analyze the relative abundance of each gene. The target signal scaling was set to 100.

Dataset Information

Transcription profiling of human MDA-MB-453 breast cancer cell lines treated with actein to assess effects on the growth as a function of time and concentration.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets