Whole genome analysis in A549 cells treated for short incubation periods with various concentrations of triptolide
ABSTRACT: The purpose of this study was to identify the genes primarily affected by a short treatment of A549 cells with grading concentrations of the natural compound triptolide. The cells were exposed to this molecule before RNA was extracted, then used for whole human genome microarray experiments. Overall design: 12 samples, corresponding to untreated cells plus three concentrations (0.05, 0.15, 0.45 µM) of tripolide for each time point (1, 2, and 4 hours). Each sample was analysed in triplicate, i.e. three microarray for each condition, making a total of 36 microarrays.
INSTRUMENT(S): PF - Agilent-014850 Whole Human Genome Microarray 4x44K
Project description:The purpose of this study was to identify the genes primarily affected by a short treatment of A549 cells with grading concentrations of the natural compound triptolide. The cells were exposed to this molecule before RNA was extracted, then used for whole human genome microarray experiments. 12 samples, corresponding to untreated cells plus three concentrations (0.05, 0.15, 0.45 µM) of tripolide for each time point (1, 2, and 4 hours). Each sample was analysed in triplicate, i.e. three microarray for each condition, making a total of 36 microarrays.
Project description:We compared miRNA expression profiles among 5 groups of HepG2 cells treated with various concentrations (0,100,200 nM) of triptolide for 12 h or 24 h. Two-condition experiment, triptolide intervention vs. without intervention. Biological replicates: 3 control, 3 treated, independently grown and harvested. One replicate per array.
Project description:Purpose: Compare the transcriptional profile of staurosporine-treated cells in Neurospora crassa wild type cells grown with different availabilities of calcium. Methods: Conidial suspensions were obtained and 1 x 106 cells/ml incubated in minimal medium (with the indicated concentrations of calcium) for 6 hours (26ºC, 140 rpm, constant light) followed by the addition of staurosporine (or DMSO) and growth for 1 more hour. For the experiments in 0 Ca2+ or 20 mM CaCl2 media, the concentration of KH2PO4 in the 50x Vogel’s stock solution was limited to 10 mM to avoid precipitation with supplemental calcium. Cells were harvested using 0.45 μm filters and immediately frozen in liquid nitrogen. Total RNA was isolated by the Trizol-Phenol-Chloroform method. After digestion of 25 μg RNA with TURBO DNAse (Life Technologies), mRNA was purified using Dynabeads oligo(dT) magnetic beads (Life Technologies). The mRNA was chemically fragmented using the Ambion RNA fragmentation kit (Life Technologies). First and second strand cDNA synthesis was achieved using appropriate kits (Life Technologies). The illumina TruSeq kit was employed to generate the cDNA libraries with indexing adapters essentially following the manufacturer’s protocol. After purification of the libraries with AMPure XP beads (Roche), the quality of the libraries was analyzed in a Agilent 2100 Bioanalyzer. The cDNA libraries were sequenced in a illumina HiSeq2000 and single reads of 50 bp were obtained. Sequencing data was handled essentially with Tophat, Cufflinks and Cuffdiff. Expression levels are presented as Fragments Per Kilobase of transcript per Million mapped reads (FPKM). Results: Differences in the concentration of extracellularly available Ca2+ result in distinct transcriptional programs. Conclusions: Our transcriptomic analysis provides a reference dataset for future investigations on the role of Ca2+ in fungal biology. Transcriptional profile of staurosporine-treated Neurospora crassa wild type cells growing with different concentrations of calcium was compared using illumina HiSeq2000 and single reads of 50 bp.
Project description:Chronic inflammation plays important role in lung cancer development. Recently, we found that anti-inflammation drugs aspirin and triptolide when combined showed synergistic effect in suppressing lung cancer development. In this study, we aim to use gene microarray to define the genes and pathways that are affected by aspirin, triptolide individually or in combination. Overall design: We will isolate lung tissues from mice of untreated control, aspirin, triptolide, or aspirin/triptolide. Total RNA are isolated and subjected to microarray analysis to define the gene expression profiles. Gene expression data are analyzed by IPA analysis to define the pathways and upstream regulators that are significantly inhibited or stimulated by aspirin, triptolide, or a combination of aspirin and triptolide.
Project description:B. cenocepacia J2315 was exposed to heat stress and to stress form reactive oxygen species. <br>To expose the cultures to heat stress, cells were grown at 37ºC to an O.D. of 0.4 to 0.45 and then transferred into a different shaking incubator at 42.5ºC, incubated for 1 hour at 150 rpm and harvested.<br>To expose cultures to oxidative stress by reactive oxygen species, cells were grown at 37ºC to an OD of 0.5. Then t butyl hydroperoxide or hydrogen peroxide solution were added to the culture at 0.001% and 0.15% final concentration. The culture was further incubated for 15 min and then harvested. <br>The expression profiles were compared to cells grown in LB medium without exposure to stress.<br>
Project description:Pancreatic cancer is characterized by heavy desmoplasia. Triptolide and its water-soluble pro-drug Minnelide are extremely efficient against pancreatic cancer in animal models. However, the effects of triptolide on pancreatic cancer stromal cells are largely unknown. The aim of this project is to indentify potential cellular functions that are affected by triptolide in pancreatic cancer associated fibroblasts. Overall design: Cancer associated fibroblasts were isolated from pancreatic tumor of KPC mouse model. Cells were either untreated or treated with 100nM triptolide for 6h or 24h before RNA isolation. The RNA was quality tested using a Bioanalyzer 2100 (Agilent Technologies, CA, USA). cDNA was created by reverse transcription of oligo-dT purified polyadenylated RNA and fragmented, blunt-ended, and then ligated to barcoded adaptors. Then, the library was size selected, and the selection process was validated and quantified by capillary electrophoresis and qPCR, respectively. Samples were load on the HiSeq 2500 (Illumina Inc., CA, USA) to generate around 25 million paired-end 50bp reads for each sample.
Project description:6 timepoints: Day 0 (normal controls), progressively developing neointimal vascular proliferation and pulmonary hypertension in vehicle treated animals (Days 14, 21, 28 and 35) and triptolide-treated animals at Day 35. Replicates: 6 for Day 0 (normal) 2 for Daty 14 3 each for Days 21, 28, 35 and Triptolide -treated at day 35 (T)
Project description:Expression data from treatment of actinomycin D (2.5uM) and triptolide (500 nM) on MCF7 cells for 2, 4 and 6 hours. We used microarrays to explore the mechanism of triptolide action. Overall design: MCF7 cells were treated with actinomycin D or triptolide for 2, 4 and 6 hours, RNA was extracted and hybridization on Affymetrix microarrays. Data were normalized by RMA.
Project description:Anticancer drug sagopilone in lung cancer line A549. We performed gene expression analysis of lung cancer cell line A549 treated with sagopilone and paclitaxel. The aim was to analyse gene expression differences between the two drugs in two different concentrations. A549 cells were treated with medium containing either 2.5 nM sagopilone, 40 nM sagopilone, 4 nM paclitaxel or 40 nM paclitaxel, respectively, vehicle (ethanol 0.1%), or were left untreated for 18 hours. The two different concentrations were chosen according the phenotypes they have caused in A549 cells. Treatment with 2.5 nM sagopilone, 4 nM paclitaxel induced an aneuploid cell population, whereas treatment with 40 nM sagopilone and 40 nM paclitaxel induced mitotic arrest.
Project description:Sulfolobus solfataricus P2 was grown aerobically, with O2 concentrations ranging from 1.5 to 26 % (v/v; gas phase). To gain some insight in control of the respiratory system, transcriptomes of the strain cultivated in different O2 concentrations (1.5 % vs 21 %, 1.5 % vs 26 %) were compared using a DNA microarray approach. Overall design: Two-condition experiments: 1.5% O2 vs two other O2 concentrations (21 or 26% O2). Biological replicates independently grown. One replicate per array. Dye swaps.