Project description:Short-read NGS technology (SOLIDTM, Life Technologies) was used to establish a comprehensive repertoire of miRNA expressed in either equine cartilage or subchondral bone. Undamaged cartilage and subchondral bone samples from 10-month Anglo-Arabian foals affected by osteochondrosis (OC) were analyzed and compared with samples from healthy foals. Samples were also subjected or not to an experimental mechanical loading to evaluate the role of miRNAs in the regulation of mechano-transduction pathways. Epiphyseal cartilage and subchondral bone miRNome were defined, including about 300 new miRNAs. Differentially expressed miRNAs were identified between bone and cartilage from healthy and OC foals, as well as after the experimental mechanical loading, suggesting that miRNAs play a role in equine OC physiopathology and in the cellular response to biomechanical stress in cartilage and bone.
Project description:Transcriptional profiling of GO-treated (24h) differentiating equine satellite cells (3rd day of differentiation) exposed to hydrogen peroxide (1h; last hour of pre-incubation with GO) compared to control GO-untreated cells. Goal was to determine the effects of GO pre-incubation on miRNA expression in equine satellite cells exposed to hydrogen peroxide.
Project description:Transcriptional profiling of HMB-treated (24h) differentiating equine satellite cells (3rd day of differentiation) exposed to hydrogen peroxide (1h; last hour of pre-incubation with HMB) compared to control HMB-untreated cells. Goal was to determine the effects of HMB pre-incubation on miRNA expression in equine satellite cells exposed to hydrogen peroxide.
Project description:Purpose:The goal of this study was to evalute gene expression patterns of equine chorioallantoic membrane during different stages of the pregnancy Method: mRNA profile of equine chorioallantoic membrane (CAM) from 45days, 4months, 6months and 10months (4 samples for each time points) generated by RNA-sequencing,using a Illumina HiSeq 4000 ( HiSeq 4000 sequencing kit version 1). The sequence reads were trimmed for adapters and quality using TrimGalore Version 0.4.4,and then mapped to EquCab2.0 using STAR-2.5.2b. Final quantification at the gen level was performed by analyzing the BAM files in cufflinks using the Equus_caballus_ENSEMBL_88 gtf file as Guide.
Project description:Identify differentially expressed microRNAs in mild and severe equine distal interphalangeal joint osteoarthritis plasma and synovial fluid samples Determine the effects of selected osteoarthritis-related miRNAs on equine chondrocytes in monolayer culture through the application of miRNA agomirs and antagomirs
Project description:HDMYZ cells were treated with 2ug/ml ActD for 0, 4 and 12 hours. Small RNAs of 15-40 bases were gel-purified from 10 ug total RNA, and subjected to multiplex Illumina small RNA library preparation. Small RNA libraries were sequenced on a HiSeq2000 (Illumina) with 3 samples per lane. To quantify miRNA and isoform abundance, sequence reads were processed by the miRDeep2 package, with the following modifications. First, to remove adaptor sequence, we removed both the main adaptor sequence present in the sequencing reads, as well as the second most abundant adaptor variant. In addition, we did not restrict the size of small RNAs during adaptor removal. Second, we used miRBase v18 for mapping the reads. Third, for quantifying miRNA and isoform frequency, we limited reads to more or equal to 15 bases in length with zero mis-match during mapping. The number of reads that were mapped to known miRNAs was used to normalize read frequencies for each miRNA or each miRNA isoform. For quantification purposes, we only considered miRNAs or isoforms that had frequency >= 1x10e-6 in samples without ActD treatment, which correspond to ~21-30 reads in raw count. These miRNAs or isoforms were referred to as reliably quantifiable.To analyze mapping to the genome, we removed reads that mapped to miRNA precursors. The rest of the reads were then mapped to the genome with Bowtie.
