MicroRNAs (miRNAs) are short noncoding RNA molecules regulating the expression of mRNAs. Target identification of miRNAs is computationally difficult due to the relatively low homology between miRNAs and their targets. We present here an experimental approach to target identification where the cartilage-specific miR-140 was overexpressed and silenced in cells it is normally expressed in separate experiments. Expression of mRNAs was profiled in both experiments and the intersection of mRNAs repressed by miR-140 overexpression and derepressed by silencing of miR-140 was identified. The intersection contained only 49 genes, although both treatments affected the accumulation of hundreds of mRNAs. These 49 genes showed a very strong enrichment for the miR-140 seed sequence implying that the approach is efficient and specific. 21 of these 49 genes were predicted to be direct targets based on the presence of the seed sequence. Interestingly, none of these were predicted by the published target prediction methods we used. One of the potential target mRNAs, Cxcl12, was experimentally validated by Northern blot analysis and a luciferase reporter assay.
Experimental identification of microRNA-140 targets by silencing and overexpressing miR-140.
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View SamplesAntagonism of microRNA-122 in mice by systemically administered LNA-antimiR leads to up-regulation of a large set of predicted target mRNAs in the liver
Antagonism of microRNA-122 in mice by systemically administered LNA-antimiR leads to up-regulation of a large set of predicted target mRNAs in the liver.
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View SamplesAnalysis of integrin alpha7 transgenic mice skeletal muscle transcription profiles comparing to wild type controls. Integrin alpha7 is the major laminin binding integrin in muscle cells. Enhancing its expression has been demonstrated to alleviate pathology in a murine model of Duchenne muscular dystrophy. Results of this study provide insights into the effects of increasing integrin alpha7 expression on skeletal muscle transcription and physiology in vivo. This analysis also evaluates any potential possible side effects associate with enhancing integrin alpha7 in skeletal muscle.
Increasing alpha 7 beta 1-integrin promotes muscle cell proliferation, adhesion, and resistance to apoptosis without changing gene expression.
Sex, Age, Specimen part
View SamplesThe 24R,25-dihydroxyvitamin D metabolite (24R,25D) has long been suspected of participating to bone fracture repair. We used Cyp24a1-deficient mice, unable to produce 24R25D, to observe gene expression in callus tissue compared to that of control littermates.
Optimal bone fracture repair requires 24R,25-dihydroxyvitamin D3 and its effector molecule FAM57B2.
Age, Specimen part, Treatment, Time
View SamplesImmune privileged Sertoli cells (SC) survive when transplanted across immunological barriers and prolong the survival of co-transplanted allogeneic and xenogeneic cells in rodent models. However, the mechanism for this survival and protection remains unresolved. We have recently identified a mouse Sertoli cell line (MSC-1) that lacks some of the immunoprotective abilities associated with primary SC. The objective of this study was to compare the survival and gene expression profiles of primary SC and MSC-1 cells to identify factors or immune-related pathways potentially important for SC immune privilege. Primary SC or MSC-1 cells were transplanted as allografts to the renal subcapsular area of nave BALB/c mice and cell survival was analyzed by immunohistochemistry. Additionally, transcriptome differences were investigated by microarray and pathway analyses. While primary SC were detected within the grafts with 100% graft survival throughout the 20-day study, MSC-1 cells w ere rejected between 11 and 14 days with 0% graft survival at 20 days post-transplantation. Microarray analysis identified 3198 genes that were differentially expressed with a 4-fold or higher level in primary SC. Cluster and pathway analyses indicate that the mechanism of SC immune privilege is likely complex with multiple immune modulators being involved such as immunosuppressive cytokines and complement inhibitors, lipid mediators for controlling inflammation, and junctional molecules that control leukocyte movement in and out of the immune privileged space. Further study of these immune modulators will increase our understanding of SC immune privilege and in the long-term lead to improvements in transplantation success.
Immunoprotective properties of primary Sertoli cells in mice: potential functional pathways that confer immune privilege.
Specimen part, Cell line
View SamplesMouse LT-HSC were sorted and cultured in mScf, mTpo, mFlt3L, hIGFBP2 and Angptl5 for 2 days. These expression values were related to insertions of gamma-retroviral, lentiviral or alpharetroviral vectors carrying GFP which were retrieved after serial murine BM transplantation. The relation between gene expression in the cells responsible for long-term hematopoiesis and location of vector integration was investigated.
Alpharetroviral self-inactivating vectors: long-term transgene expression in murine hematopoietic cells and low genotoxicity.
Specimen part
View SamplesIre1 conditional null or control mice of 3-months old were injected intraperitoneally with TM or vehicle.
The unfolded protein response transducer IRE1α prevents ER stress-induced hepatic steatosis.
Specimen part
View SamplesIdentification of genes differentially regulated after treatment of zebrafish embryos from 50% epiboly to 24hpf with 6.5uM leflunomide A six chip study comparing expression levels of zebrafish embryos treated with leflunomide 6.5uM
DHODH modulates transcriptional elongation in the neural crest and melanoma.
Specimen part, Treatment, Subject
View SamplesThe goal of this study was to identify the molecular characteristics and putative markers distinguishing IL-10eGFP+CD138hi and IL-10eGFP-CD138hi plasmocytes. To this end, IL-10eGFP B-green mice were challenged intravenously with Salmonella typhimurium (strain SL7207, 10e7 CFU), and IL-10eGFP+CD138hi as well as IL-10eGFP-CD138hi plasmocytes were isolated from the spleen on the next day. For this, single cell suspensions were prepared, cells were treated with Fc block (10 g/ml, anti-CD16/CD32, clone 2.4G2), and then stained with an antibody against CD138 conjugated to PE (1/400; from BD Pharmingen) followed by incubation with anti-PE microbeads (Miltenyi Biotech). CD138+ cells were then enriched on Automacs (Miltenyi Biotech) using the program possel_d2. Cells were then stained with anti-CD19-PerCP, anti-CD138-PE, and antibodies against CD11b, CD11c, and TCR conjugated to APC as a dump channel to exclude possible contaminants. DAPI was added to exclude dead cells. Live IL-10eGFP+CD138hi and IL-10eGFP-CD138hi cells were subsequently isolated on a cell sorter. The purity of the samples was always above 98%. This led to the identification of LAG-3 as a cell surface receptor specifically expressed on IL-10eGFP+CD138hi cells but not on IL-10eGFP-CD138hi cells.
LAG-3 Inhibitory Receptor Expression Identifies Immunosuppressive Natural Regulatory Plasma Cells.
Sex, Specimen part
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