ABSTRACT: Effect of MCPIP1 knockdown on miRNA expression profile. HepG2 cells were transfected with control siRNA or siRNA against MCPIP1, and subjected to miRNA microarray analysis.
Project description:Despite advance in interferon-based treatment for chronic hepatitis C, difficult-to-treat patients remain in existence yet. To identify key genes involved in difficult-to-treat characteristics, gene expression patterns of miRNA and RNA were analyzed by profiling pretreatment liver tissues from five sustained virological responders (SVR), three relapsers (R) and four non-responders (NR). Expression levels of miRNA and mRNA were compared between SVR/R and NR groups by using microarray, respectively. Quantitative real-time reverse-transcriptase polymerase chain reaction and statistical analyses validated genes with significantly differential expression levels in 50 liver tissues: proliferation-, inflammation- and anti-apoptosis-related mRNA expression levels increased significantly in NR, compared to SVR/R. Of miRNA with significantly differential expression levels on microarray, several miRNA were correlated inversely with those significant mRNA. In vitro studies by using miRNA inhibitors and mimics verified the inverse correlation between the miRNA and mRNA. These findings enhance our understanding of the difficult-to-treat molecular mechanism and identification of target molecules for novel treatments. Moderated t-statistics based on the empirical Bayes approach were performed to identify miRNA differentially expressed between SVRs/relapses and NVRs using the M-bM-^@M-^\limmaM-bM-^@M-^] package from BioConductor software (http://www.bioconductor.org/) under R statistical software version 2.12.1 (http://www.r-project.org/). Because of multiple hypothesis testing, p values were adjusted by the Benjamini-Hochberg false discovery rate (FDR) method. Related mRNA expression data are in GSE42697.
Project description:microRNA profiling of comparing control or ETO2 siRNA-treated human K562 cells Two-condition experiment, K562-control siRNA vs. K562-ETO2 siRNA, Biological replicates: 1, 1 control siRNA, 1 ETO2 siRNA, independently.
Project description:Transcriptional profiling of miRNA levels in mammary tumors from 18 [PyMT x AKXD]F1 sublines. The PyMT strain was FVB/N-TgN(MMTV-PyVT)634Mul. Mammary tumor total RNA from mice representing one of 18 AKXD RI strains were pooled to represent each strain and expression profiled using a custom miRNA microarray.
Project description:MicroRNA expression profiling using microRNA microarrays was performed on the porcine developing brain. Two tissues, cortex and cerebellum, were sampled at each of three different developmental stages: gestation day 50 (F50), gestation day 100 (F100) and three months old (named adult), respectively. The aim was to find developmental stage-, as well as tissue-specific, microRNA candidates and validate them with qPCR. RNA samples containing the small RNA fraction were subjected to microarray analysis. Three replicates per developmental stage/tissue.
Project description:We examined the relationship between miRNA expression and the sensitivity of CCA cells to Gem. Two intrahepatic CCA cell lines, HuH28 and HuCCT1 were used. HuCCT1 cells were more sensitive to Gem than were HuH28 cells. The miRNA expression profiles of HuH28 and HuCCT1 were determined by microarray analysis. Eighteen miRNAs were differentially expressed whose ratios over ± 2log2 between HuH28 and HuCCT1. Among these 18 miRNAs, ectopic overexpression of each of three downregulated miRNAs in HuH28 (miR-29b, miR-205, miR-221) restored Gem sensitivity to HuH28. Suppression of one upregulated miRNA in HuH28, miR-125a-5p, inhibited HuH28 cell proliferation independently to Gem treatment. Cell lines and cultures Two human intrahepatic CCA cell lines, HuCCT1 and HuH28, were purchased from Japan Health Science Research Resources Bank (Osaka, Japan). Each cell line was cultured in RPMI-1640 medium (Invitrogen, Life Technologies Corp., CA, USA) that contained 10% fetal bovine serum (Nichirei Bioscience, Tokyo, Japan) and in humidified conditions at 37 ˚C and 5% CO2. Antibiotics were not added to the culture medium when cells were prepared for transfection with miRNA mimics or oligonucleotides. Gem treatment Gem hydrochloride was purchased from Wako (Osaka, Japan). A stock solution was prepared at 1 mmol/L (1 x 10-3 M) and was further diluted to anyone of several different final working concentrations from 1 x 10-4 to 1 x 10-7 M with cell culture medium that lacked antibiotics. Transfection of miRNA mimics, antisense oligonucleotides, or siRNA for miRNA target genes were performed 24 hr before the Gem treatment. All assays were conducted 72 hr after Gem treatment.
Project description:Comparison of miRNA expression profiles in cervical carcinoma cell lines to study the effects of Drosha expression levels Analyzed global miRNA expression profiles from 18 samples (7 samples in duplicate, 4 single sample) representing cervical carcinoma cell lines with either relative Drosha over-expression (Cluster 2) or Drosha under-expression (Cluster 1), due to wild-type expression in those cell lines or manipulation with RNAi.
Project description:MicroRNA expression profiling using microRNA microarrays on NCI-60 cell lines. RNA samples containing small RNA fraction were profilled
Project description:We studied the levels of expression of hundreds of mature miRNA's on an arrray (miRCURY, Exiqon, Cophenhagen, DN) for primary human T cells and NK cells. We found a 4% difference in the level of expression between these two cell types. NK and T cells were purified from peripheral blood mononuclear cells by negative selection from three healthy adult individuals, and total RNA was harvested. Samples reverse transcribed and studied on a microarry (miRCURY, Exiqon)
Project description:Idiopathic pulmonary fibrosis is a chronic devastating disease of unknown etiology. No therapy is currently available. A growing body of evidence supports the role of TGFβ1 as the major player in the pathogenesis of the disease. This study designed novel human- and mouse-specific siRNAs and siRNA/DNA chimeras targeting both human and mouse common sequences and evaluated their inhibitory activity in pulmonary fibrosis induced by bleomycin and lung-specific transgenic expression of human TGFβ1. Selective novel sequences of siRNA and siRNA/DNA chimeras efficiently inhibited pulmonary fibrosis, indicating their applicability as tools for treating fibrotic disease in humans. Total RNA was extracted from lung tissue from mice with bleomycin (BLM)-induced lung fibrosis treated with mouse TGFβ1 siRNAs or vehicle on different days after BLM infusion.
Project description:Dioxin-like chemicals are well-known for their ability to upregulate expression of numerous genes via the AH receptor (AHR). However, recent transcriptomic analyses in several laboratories indicate that dioxin-like chemicals or AHR genotype itself also can downregulate levels of mRNAs encoded by numerous genes. The mechanism responsible for such downregulation is unknown. We hypothesized that microRNAs (miRNAs), which have emerged as powerful negative regulators of mRNA levels in several systems, might be responsible for mRNA downregulation in dioxin/AHR pathways. We used the Exiqon miRNA array platform as well as quantitative RT-PCR to measure miRNA levels in dioxin-sensitive Long-Evans (Turku/AB; L-E) rats. Treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) for 96 hr in vivo caused few changes in miRNA levels in rat livers and those changes that were statistically significant were of modest magnitude. Feed-restricted-control L-E rats were included to ensure that changes in miRNA levels were due to TCDD-treatment per se and not the result of the decreased feed intake which occurs in dioxin-sensitive strains within 96 h after TCDD exposure. Manuscript Submitted: Moffat ID, Boutros PC, Celius T, Pohjanvirta R & Okey AB. Micro-RNAs in rodent liver are refractory to dioxin treatment. Toxicological Sciences May, 2007. Keywords: miRNA expression, response to xenobiotics, feed restriction response A loop design used to profile miRNA levels from dioxin-sensitive L-E AHRWT/WT vehicle-control rats (LC96), those exposed to TCDD for 96 h (LT96), and feed-restricted (LF96) rats.