Project description:Analysis of gene expression change in U2OS cells expression synthetic miR-542-3p mimics. Forty-eight hours after transfection with negative miRNA mimics or miR-542-3p mimics, U2OS cells were subjected to RNA isolation.
Project description:We tested the hypothesis that a panel of placental mammal-specific miRNAs and their targets play important to establish receptivity to implantation and their dysregulated expression may be a feature in women with early pregnancy loss. Relative expression levels of miR-340-5p, −542-3p, and −671-5p all increased following treatment of Ishikawa cells with progesterone (10 μg/ml) for 24 hrs (p < 0.05). RNA sequencing of these P4-treated cells identified co-ordinate changes to 6,367 transcripts of which 1713 were predicted targets of miR-340-5p, 670 of miR-542-3p, and 618 of miR-671-5p. Quantitative proteomic analysis of Ishikawa cells transfected with mimic or inhibitor (48 hrs: n=3 biological replicates) for each of the P4-regulated miRNAs was carried out to identify targets of these miRNAs. Excluding off target effects, mir-340-5p mimic altered 1,369 proteins while inhibition changed expression of 376 proteins (p < 0.05) of which, 72 were common to both treatments. A total of 280 proteins were identified between predicted (mirDB) and confirmed (in vitro) targets. In total, 171 proteins predicted to be targets by mirDB were altered in vitro by treatment with miR-340-5p mimic or inhibitor and were also altered by treatment of endometrial epithelial cells with P4. In vitro targets of miR-542-3p identified 1,378 proteins altered by mimic while inhibition altered 975 a core of 200 proteins were changed by both. 100 protein targets were predicted and only 46 proteins were P4 regulated. miR-671-mimic altered 1,252 proteins with inhibition changing 492 proteins of which 97 were common to both, 95 were miDB predicted targets and 46 were also P4-regulated. All miRNAs were detected in endometrial biopsies taken from patients during the luteal phase of their cycle, irrespective of prior or future pregnancy outcomes Expression of mir-340-5p showed an overall increase in patients who had previously suffered a miscarriage and had a subsequent miscarriage, as compared to those who had infertility or previous miscarriage and subsequently went on to have a life birth outcome. The regulation of these miRNAs and their protein targets regulate the function of transport and secretion, and adhesion of the endometrial epithelia required for successful implantation in humans. Dysfunction of these miRNAs (and therefore the targets they regulate) may contribute to endometrial-derived recurrent pregnancy loss in women.
Project description:Oxaliplatin (oxPt) resistance in colorectal cancers (CRC) is a major unsolved problem. Consequently, predictive markers and a better understanding of resistance mechanisms are urgently needed. To investigate if the recently identified predictive miR-625-3p is functionally involved in oxPt resistance, stable and inducible models of miR-625-3p dysregulation were analyzed. Ectopic expression of miR-625-3p in CRC cells led to increased resistance towards oxPt. The mitogen-activated protein kinase (MAPK) kinase 6 (MAP2K6/MKK6) – an activator of p38 MAPK - was identified as a functional target of miR-625-3p, and, in agreement, was down-regulated in patients not responding to oxPt therapy. The miR-625-3p resistance phenotype could be reversed by anti-miR-625-3p treatment and by ectopic expression of a miR-625-3p insensitive MAP2K6 variant. Transcriptome, proteome and phosphoproteome profiles revealed inactivation of MAP2K6-p38 signaling as a possible driving force behind oxPt resistance. We conclude that miR-625-3p induces oxPt resistance by abrogating MAP2K6-p38 regulated apoptosis and cell cycle control networks.
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.
Project description:Osteoporosis is a significant health concern, and the role of microRNAs (miRNAs) in cell growth and development regulation is well recognized. High-throughput sequencing technology is widely employed in current research. This study aimed to identify and validate miRNAs associated with osteoporosis. Bone specimens were collected from patients with osteoporosis (n=3) and without osteoporosis (n=3). High-throughput sequencing was utilized to screen for miRNAs, followed by analysis using volcano maps, Wayne maps, gene ontology (GO) analysis, and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. The identified miRNAs were further confirmed using qRT-PCR. Sequencing analysis revealed 12 down-regulated and five upregulated miRNAs in osteoporosis. GO and KEGG analysis indicated the association of these miRNAs with bone metabolism. qRT-PCR results demonstrated a significant decrease in miR-140-5p, miR-127-3p, miR-199b-5p, miR-181a-5p, miR-181d-5p, and miR-542-3p (all P<0.05) in osteoporosis compared to controls, while miR-486-3p and miR-486-5p exhibited a significant increase (P<0.05). This study utilized high-throughput sequencing to identify differential miRNA expression in individuals with osteoporosis. Specifically, six miRNAs (miR-140-5p, miR-127-3p, miR-199b-5p, miR-181a-5p, miR-181d-5p, and miR-542) showed decreased expression, whereas two miRNAs (miR-486-3p and miR-486-5p) exhibited increased expression. The differential expression of these miRNAs may serve as predictive indicators, potentially aiding in the prognosis and management of osteoporosis.