Project description:Liposomal transfection reagents are frequenly used in gene deliverytransfer experiments. The validity of these experiments depends on the absence of changes in genes other than the target gene. Here we report that transfection of synthetic microRNA-145 using liposomes, but not electroporation, induced the upregulation of a range of immune-related genes in human mesenchymal stem cells. The immune response was dependent on the binding of microRNA-145 to retinoic acid inducible gene-I, and was independent on endosome functionality and toll-like receptors. Immune response was not observed after transfection of any of nine other small RNAs, suggesting a sequence restricted response. One of the upregulated mRNAs encode interferon β, which presumably in turn induced the upregulation of another group of immune associated genes. Interestingly, exposure of liposomes alone led to the upregulation of some immune genes, one of them retinoic acid inducible gene-I mRNA. Comparison of immune gene expression patterns in BM-MSC donors after transfection of synthetic miR-145.
Project description:Liposomal transfection reagents are frequenly used in gene deliverytransfer experiments. The validity of these experiments depends on the absence of changes in genes other than the target gene. Here we report that transfection of synthetic microRNA-145 using liposomes, but not electroporation, induced the upregulation of a range of immune-related genes in human mesenchymal stem cells. The immune response was dependent on the binding of microRNA-145 to retinoic acid inducible gene-I, and was independent on endosome functionality and toll-like receptors. Immune response was not observed after transfection of any of nine other small RNAs, suggesting a sequence restricted response. One of the upregulated mRNAs encode interferon β, which presumably in turn induced the upregulation of another group of immune associated genes. Interestingly, exposure of liposomes alone led to the upregulation of some immune genes, one of them retinoic acid inducible gene-I mRNA.
Project description:We analyzed the expression profiles of hsa-miR-145-5p or hsa-miR-31-5p-targeting genes relating to invasion or migration after co-overexpression of hsa-miR-145-5p and 31-5p Gene expression profiles of U87 cells after co-transfection with hsa-miR-145-5p and 31-5p mimics, and U87 cells after transfection miR mimic negative control
Project description:miR-145 is a tumor suppressor miRNA in various malignancies including pancreatic cancer. Identification of miR-145 targets can lead to more understanding of the development of pancreatic cancer. Therefore, in this project, we aimed to characterize the changes of transcriptomes caused by miR-145 to identify more miR-145 target transcripts. cDNA microarray was used to compare transcriptomes 48 hr after transfection of miR-145 mimics or control scramble RNA in a pancreatic cancer cell line, Mia-PaCa2.
Project description:miR-145 is a tumor suppressor miRNA in various malignancies including pancreatic cancer. How miR-145 regulates expression of other miRNAs could provide a panoramic view of the development of pancreatic cancer. Therefore, in this project, we aimed to characterize how other miRNAs are affected by miR-145 in hope for understanding proteomic changes that are not explained by miR-145 alone. miRNA microarray was used to compare expression of all miRNAs 48 hr after transfection of miR-145 mimics or control scramble RNA in a pancreatic cancer cell line, Mia-PaCa2.
Project description:We identified a small molecule compound, KIN1148, that directly binds RIG-I to drive IRF3 and NF B activation and expression of innate immune genes, cytokines and chemokines. KIN1148 activates RIG-I in an RNA- and ATP-independent manner and does not induce a canonical antiviral interferon (IFN) gene program traditionally associated with RIG-I activation. When administered in combination with a vaccine against influenza A virus (IAV), KIN1148 induces both neutralizing antibody and broadly cross-protective IAV-specific T cell responses compared to vaccination alone, which induces poor responses. In this study, we demonstrate that KIN1148 directly engages RIG-I to activate IRF3- and NFB-dependent innate immune responses, making it the first small molecule RIG-I agonist to be identified. Biochemical studies show that KIN1148 binds to RIG-I to drive RIG-I self-oligomerization and downstream signaling activation in an RNA- and ATP-independent manner. We further find that transcriptional programs induced by KIN1148 treatment exhibit shared and unique signatures to that induced by other methods of RIG-I activation, including Sendai virus (SeV) infection and PAMP RNA transfection. KIN1148 adjuvants a split virus (SV) vaccine at suboptimal dose to protect mice from lethal challenge with a recombinant highly pathogenic avian H5N1 influenza virus, A/Vietnam/1203/2004.
Project description:It is aimed to reveal overall trancriptional change in prostate cancer PC3 cells after ectopic expression of miR-145 Pre-mir-145 transfected PC3 cells were collected at 8, 16 and 24 hours after transfection and control untrasfected PC3 cells were used.
Project description:To identify putative novel specific targets of miR-143-3p and miR-145-5p, we overexpressed these miRNAs in BRAFV600E mutant human M238P melanoma cells using synthetic mimics or a synthetic “negative” control mimic (miR-Neg). RNA samples were harvested 48 hours post-transfection and 2 independent experiments were carried out.
Project description:To investigate the gene expression in human corneal epithelial overexpressing hsa-miR-145 by transfection , we have employed Whole Human Genome Oligo Microarray (Agilent) as a screening platform to identify gene regulation. We discovered a differential gene expression in HCE cells transfected with hsa-mIR-145 against cells with scrambled sequences. Among them, genes related with corneal development, integrity, differentiation and inflammatory responses were found and this was validated by real-time PCR.
Project description:miR-145 is downregulated in multiple cancers. The introduction of miR-145 could alleviate the tumor burden in the pancreatic cancer mouse model. However, how miR-145 mediates the tumor suppression is still an open question. In this study, we aimed to identify the targets of miR-145 using a SILAC approach.