Project description:Background : In lactating cow, a sunflower oil supplementation modulated the milk composition and the mammary genes expression whose mechanisms of regulations are unclear. Results : This lipid supplementation provoke, in mammary gland, the down regulation of microRNA miR-20a-5p and miR-142-5p, which are predicted to target genes previously determined as differentially expressed including those involved in lipid metabolism. Conclusion : Those two miRNA are good candidates to explain lipogenic genes regulations after sunflower oil supplementation. Significance : The current study clarified the nutriregulation of miRNA in the mammary gland.
Project description:Background : In lactating cow, a sunflower oil supplementation modulated the milk composition and the mammary genes expression whose mechanisms of regulations are unclear. Results : This lipid supplementation provoke, in mammary gland, the down regulation of microRNA miR-20a-5p and miR-142-5p, which are predicted to target genes previously determined as differentially expressed including those involved in lipid metabolism. Conclusion : Those two miRNA are good candidates to explain lipogenic genes regulations after sunflower oil supplementation. Significance : The current study clarified the nutriregulation of miRNA in the mammary gland. MiRNomes obtained by NGS sequencing of mammary gland of Holstein cows at the peak of lactation received a low forage diet supplemented or not with 4% of sunflower oil were compared. MiRNA differentially expressed confirmed by RT-qPCR were related to differentially expressed genes list previously identified in the same samples.
Project description:Animal nutrition considerably affects milk composition that influences its nutritional quality. Milk component synthesis and secretion by the mammary gland involve the expression of a large number of genes whose nutritional regulation remains poorly defined. In this study, 16 lactating goats received 4 experimental diets differing in either forage to concentrate ratio (high forage, HF, or low forage, LF) supplemented, or not, with lipids (whole rapeseeds, RS, or sunflower oil, SO) in a 4 x 4 Latin Square design. To investigate the pathways regulated by nutrition, we examined the effect of these diets on the expression of approximately 8400 genes in caprine mammary gland using a bovine oligonucleotide microarray.
Project description:Animal nutrition considerably affects milk composition that influences its nutritional quality. Milk component synthesis and secretion by the mammary gland involve the expression of a large number of genes whose nutritional regulation remains poorly defined. In this study, 16 lactating goats received 4 experimental diets differing in either forage to concentrate ratio (high forage, HF, or low forage, LF) supplemented, or not, with lipids (whole rapeseeds, RS, or sunflower oil, SO) in a 4 x 4 Latin Square design. To investigate the pathways regulated by nutrition, we examined the effect of these diets on the expression of approximately 8400 genes in caprine mammary gland using a bovine oligonucleotide microarray. Due to the limited quantity of mammary RNA available, equal amounts of total RNA sample from mammary gland of each goat belonging to the same Latin Square group were mixed together before labeling. Each mammary pooled sample (4 by dietary treatment) was then co-hybridized with a standard reference corresponding to a mixture of purified total RNA from several caprine tissues. Each hybridization was repeated in a dye-swap manner for a total of 32 slides (8 slides and 4 independent comparisons by dietary treatment).
Project description:Purpose: In this study, we performed RNA-seq analysis as a screening strategy to identify EV-miRNAs derived from serum of well clinically annotated breast cancer (BC) patients from South of Brazil. Methods: EVs from three groups of samples, healthy controls (CT), luminal A (LA), and triple negative (TNBC), were isolated from serum using a precipitation method and analyzed by RNA-seq (screening phase). Subsequently, four EV-miRNAs (miR-142-5p, miR-150-5p, miR-320a, and miR-4433b-5p) were selected to be quantified by RT-qPCR in individual samples (test phase). Results: A panel composed of miR-142-5p, miR-320a, and miR-4433b-5p discriminated BC patients from CT with an AUC of 0.8387 (93.33% sensitivity, 68.75% specificity). In addition, the combination of miR-142-5p and miR-320a, presented an AUC of 0.941 (100% sensitivity, 93.80% specificity) in distinguishing LA patients from CT. Interestingly, decrease expression of miR-142-5p and miR-150-5p were significantly associated with more advanced tumor grades (grade III), while the decrease expression of miR-142-5p and miR-320a with larger tumor size. Conclusion: These results provide insights into the potential application of EVs-miRNAs from serum as novel specific markers for early diagnosis of BC.
Project description:To identify target genes of miR-142-5p and miR-130a-3p that are involved in M2 polarization, we examined the mRNA expression profile changes after altering miR-142-5p or miR-130a-3p expression in IL-4-treated macrophages.
Project description:MicroRNAs (miRNAs) post-transcriptionally regulate gene expression by inhibiting protein synthesis of target messenger RNAs (mRNAs). MicroRNA-142 (miR-142), which has tumor-suppressive properties, was functionally deleted by CRISPR/Cas9 knockout in cell lines derived from diffuse large B-cell lymphoma (DLBCL), a highly aggressive tumor that represents about 30% of non-Hodgkin lymphoma worldwide. Mutations in miR-142 affect about 20% of all cases of DLBCL. By proteome analyses, the miR-142 knockout resulted in a consistent up-regulation of 52 but also down-regulation of 41 proteins in the GC-DLBCL lines BJAB and SUDHL4. Various mitochondrial ribosomal proteins were up-regulated in line with their pro-tumorigenic properties, while proteins necessary for MHC-I presentation were down-regulated in accordance with the finding that miR-142 knockout mice have a defective immune response. Of the deregulated proteins/genes, CFL2, CLIC4, STAU1, and TWF1 are known targets of miR-142, and we could additionally confirm AKT1S1, CCNB1, LIMA1, and TFRC as new targets of miR-142-3p or -5p. We further show that seed-sequence mutations of miR-142 can be used to confirm potential targets and that miRNA knockout cell lines might thus be used to identify novel targets of miRNAs. Due to the complex contribution of miRNAs within cellular regulatory networks, in particular when a miRNA highly present in the RISC complex is deleted and can be replaced by other endogenous miRNAs, primary effects on gene expression may be covered by secondary layers of regulation
Project description:To identify target genes of miR-142-5p and miR-130a-3p that are involved in M2 polarization, we examined the mRNA expression profile changes after altering miR-142-5p or miR-130a-3p expression in IL-4-treated macrophages. Macrophages were transfected with control ASO, miR-142-5p ASO, control mimics or miR-130-3p mimics using lentiviral vectors. After 24 hr, the cells were treated with IL-4 for 24h.mRNA was purified from total RNA after removal of rRNA (mRNA-ONLY™ Eukaryotic mRNA Isolation Kit, Epicentre). Then, each sample was amplified and transcribed into fluorescent cRNA along the entire length of the transcripts without 3’ bias utilizing a random priming method. The labeled cRNAs were hybridized onto the Human RNA Array v2.0 (8 x 60K, Arraystar). After having washed the slides, the arrays were scanned by the Agilent Scanner G2505B.
Project description:Porcine alveolar macrophages were challenged in vitro with a low virulent strain of the Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) to identify and characterize the population of host genes subjected to miRNA post-transcriptional regulation durign early viral infection. We employed a high-throughput biochemical assay, i.e. the immunoprecipitation of RISCs followed by microarray analysis of the RISC-bound miRNA targets (RIP-Chip). qPCR analyses were carried out to validate the resolution of the microarray and to determine levels of expression of a panel of eight miRNAs (ssc-miR-142-3p, ssc-miR-142-5p, ssc-let-7f, miR-146a-5p, miR-155-5p, ssc-miR-181a, ssc-miR-21 and ssc-miR-335).
Project description:Porcine alveolar macrophages were challenged in vitro with a low virulent strain of the Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) to identify and characterize the population of host genes subjected to miRNA post-transcriptional regulation durign early viral infection. We employed a high-throughput biochemical assay, i.e. the immunoprecipitation of RISCs followed by microarray analysis of the RISC-bound miRNA targets (RIP-Chip). qPCR analyses were carried out to validate the resolution of the microarray and to determine levels of expression of a panel of eight miRNAs (ssc-miR-142-3p, ssc-miR-142-5p, ssc-let-7f, miR-146a-5p, miR-155-5p, ssc-miR-181a, ssc-miR-21 and ssc-miR-335).