MicroRNA profiling of mouse thymus from diabetic and pre-diabetic NOD (non obese diabetic) animals
Ontology highlight
ABSTRACT: The current project is within the range of molecular immunogenetic auto immune diseases and refers to the comparative study of promiscuous gene expression of tissue-specific antigens (TSAs) in the thymus of NOD mice line (non obese diabetic) who plays the auto-immune diabetes mellitus type 1, during the transition from state pre-diabetics to diabetics. Use the technology of oligo arrays to investigate the expression of miRNAs and cDNA microarrays to investigate the expression of genes encoding the messenger RNAs including TSAs (tissue specific antigens).
Project description:The time course experiment was designed as follow: (1) Mesenchymal stem cell (indifferentiated cells)/ time 0h; (2) After 24 h of osteogenic induction; (3) After 48 h of osteogenic induction (4) After 7 days of osteogenic induction.
Project description:Umbilical cord mesenchymal stem cells were stimulated to osteogenesis by 24h, 48h and 7 days. Undifferentiated and treated cells have their total RNA extracted and 25 ug were used to extract micro RNAs by Flash PAGE Fractionator (Ambion). These miRNA were labeled and used to hybridize to 662 oligo arrays corresponding to miRNA sequences from human, rat and mouse. The hybridized slides were scanned in the Amersham Automatic Slide Processor, quantified by TIGR Spotfinder software and submitted to stathiscal treatment in TIGR MEV software.
Project description:Small interfering RNA ( siRNA) was used to knockdown Autoimmune regulator (Aire) gene by in vivo electroporation of the thymus of BALB-c mice. In this set of data we include control and Aire-knockdown mTEC cells isolated from thymus of BALB-c mice.
Project description:miRNA array comparing the transcription profile of control rats and rats after intra-hippocampal pilocarpine-induced Status Epilepticus (PILO-SE).
Project description:Hypoxia results in the changes in expression of many genes, the majority of which are mediated via the transcriptional activity of the hypoxia inducible factor (HIF) complex. However, other mechanisms of gene regulation by hypoxia are likely and include control of mRNA stability, regulation of mRNA translation and regulation mediated by micrornas. The aim of this study is to identify microRNAs which expression is regulated by hypoxia. We chose the breast cancer line MCF7 for study as we had previously characterised the expression of the components of the HIF system in that cell line and undertaken an extensive study of the gene expression profile in response to hypoxia, a prolyl hydroxylase inhibitor dimethyloxalylglycine and HIF-1a isoform manipulations (Eldvidge. G.P. et al. (2006) JBC, vol. 281, 22, 15215-15226).
Project description:Folate deficiency, arsenic exposure, and gamma-IR are known developmental toxicants and have carcinogenic effects. The mechanism by which IR is known, but the effect of arsenic or folate deficiency remains unclear. Their effect may be mediated by epigenetic alterations, leading to miRNA expression changes, and this experiment examined this hypothesis We used microarrays to detail the miRNA expression profiles of TK6 cells treated with 2 uM sodium arsenite for 6 days, folate-deficient media for 6 days, or 2.5 Gy IR exposure either acutely at 4 hours post exposure or long-term at 6 days post exposure, as well as requiste controls, all in biological triplicate. Keywords: exposure differences TK6, were cultured in standard RPMI 1640 (Invitrogen Inc., Gaithersburg, MD) or folate-deficient RPMI 1640 (Invitrogen). Growth media was supplemented with 10% fetal bovine serum (Invitrogen) and 1% penicillin-streptomycin; dialyzed fetal bovine serum (Invitrogen) was added to the folate-deficient medium in order to eliminate folic acid in the serum. For controls, folate-deficiency, arsenic exposure, and 6-day ?-IR exposure groups, 106 cells were diluted into 50ml of appropriate growth media. For the 4-hour post-?-IR exposure group, 107 cells were diluted into 50ml of growth media. For the arsenic exposed group, sodium arsenite was added to the media to a concentration of 2 ?M. For the ?-IR exposure groups, cells were diluted and allowed to incubate for 4 hours prior to irradiation treatment, and were exposed at a dose rate of 86.76 rad/min to a final dose of 2.5 Gy using a Philips MGC-40 X-ray source. Following exposure, cells were returned to the incubator. After fours hours, the short-term post-??-IR exposure group was collected for RNA isolation, as well as a mock (control) group. All experimental and control conditions were performed in triplicate. For all other groups, cells were cultured for 6 days, with the media changed and renewed, with the appropriate treatment, on day 3.
Project description:specific pathogen-free female BALB/c mice aged 7-8 weeks (Animal Resources Centre, Perth, Western Australia) were systemically sensitised by intraperitoneal injection of 50 M-5g of alum-precipitated chicken egg OVA (Grade V, ?98% pure, Sigma Australia) 21 and 7 days before inhalational challenge, then exposed to aerosolised OVA in a whole body inhalation exposure chamber (Unifab Corporation, Kalamazoo, MI). Chronic low-level challenge involved exposure to ?3 mg/m3 aerosolised OVA for 30 minutes/day on 3 days/week for up to 6 weeks. Particle concentration within the chamber was continuously monitored using a DustTrak 8520 instrument (TSI, St Paul, MN). All experimental procedures complied with the requirements of the Animal Care and Ethics Committee of the University of New South Wales (reference numbers: 06/119B and 08/09B). Mice were sacrificed after 1,2,4 and 6 weeks of OVA exposure. Control groups included naM-ove mice and mice that were not sensitised but were challenged for 6 weeks with aerosolised OVA.
Project description:Defects in stress responses are important contributors in many chronic conditions including cancer, cardiovascular disease, diabetes, and obesity-driven pathologies like non-alcoholic steatohepatitis (NASH). Specifically, endoplasmic reticulum (ER) stress is linked with these pathologies and control of ER stress can ameliorate tissue damage. MicroRNAs have a critical role in regulating diverse stress responses including ER stress. Here we show that miR-494-3p plays a functional role during ER stress. ER stress inducers (tunicamycin and thapsigargin) robustly increase the expression of miR-494 in vitro in an ATF6 dependent manner. Surprisingly, miR-494 pretreatment dampens the induction and magnitude of ER stress in response to tunicamycin in endothelial cells. Conversely, inhibition of miR-494 increases ER stress de novo and amplifies the effects of ER stress inducers. Using Mass Spectrometry (TMT-MS) we identified many proteins that are downregulated by both tunicamycin and miR-494 in cultured human umbilical vein endothelial cells (HUVECs). Among these, we found 6 transcripts which harbor a putative miR-494 binding site. Our data indicates that ER stress driven miR-494 may act in a feedback inhibitory loop to dampen downstream ER stress signaling. We propose that RNA-based approaches targeting miR-494 or its targets may be attractive candidates for inhibiting ER stress dependent pathologies in human disease.
Project description:The current project is within the range of molecular immunogenetic auto immune diseases and refers to the comparative study of promiscuous gene expression of tissue-specific antigens (TSAs) in the thymus of NOD mice line (non obese diabetic) who plays the auto-immune diabetes mellitus type 1, during the transition from state pre-diabetics to diabetics. Use the technology of oligo arrays to investigate the expression of miRNAs and cDNA microarrays to investigate the expression of genes encoding the messenger RNAs including TSAs (tissue specific antigens). Use the technology of oligo arrays to investigate the expression of miRNAs and cDNA microarrays to investigate the expression of genes encoding the messenger RNAs including TSAs (tissue specific antigens). Female NOD mice were born in specific pathogen free (SPF) conditions at the CEMIB-UNICAMP animal facility (University of Campinas, SP, Brazil) and maintained in SPF mini-isolators in our laboratory at the University of S�o Paulo, Campus of Ribeir�o Preto, Brazil. We studied both pre-diabetic (8�2 week-old) and diabetic (20�2 week-old) animals. Diabetes was confirmed by blood glucose levels (?250 mg glucose/dL) using the Accu Check � Active Kit (Roche Diagn�stica Brasil, S�o Paulo, Brazil). The thymic stroma was separated from the whole thymus, as previously described (Gray et al. 2002). The central idea is to trace signatures of differential gene expression of the thymus at different stages (transition from state pre-diabetics to diabetics) and, using bioinformatics programs, applied to the analysis of data from arrays [Cluster & Tree View (for signatures of expression), SAM (for statistical analysis of the miRNAs and differentially expressed genes from TSAs), GenMiR++ and Cytoscape (to establish networks between miRNAs genes and genes of TSAs)].
Project description:To examine the effect of DEAD-box RNA helicase subunits on the primary miRNAs processing by Drosha complex, we made knockout mice of p72, DEAD-box RNA helicase, a component of Drosha complex. And we compare the miRNA expression profiles derived from whole mice embryo between wild-type and p72 KO mice.