Project description:Carcinomas of unknown primary origin constitute 3-5% of all newly diagnosed metastatic cancers, of which the primary source is difficult to classify with current histological methods. Effective cancer treatment depends on early and accurate identification of the tumor, which is why patients with metastases of unknown origin have poor prognosis and short survival. Because microRNA expression is highly tissue specific, the microRNA profile of a metastasis may be used to identify its origin. As a first step to realize this goal, we evaluated the potential of microRNA profiling for identification of the primary tumor of known metastases. 208 formalin-fixed paraffin-embedded samples representing 15 different histologies were profiled on an LNA-enhanced microarray platform, which allows for highly sensitive and specific detection of microRNA. Based on these data, we developed and cross-validated a novel classification algorithm, LASSO (Least Absolute Shrinkage and Selection Operator), which had an overall accuracy of 85%. When the classifier was applied on an independent test set of 48 metastases, the primary site was correctly identified in 42 cases (88% accuracy). Our findings suggest that microRNA expression profiling on paraffin tissue can efficiently predict the primary origin of a tumor, and may provide pathologists with a molecular diagnostic tool that can improve their capability to correctly identify the origin of hitherto unidentifiable metastatic tumors, and eventually, enable tailored therapy. 94 samples
Project description:Carcinomas of unknown primary origin constitute 3-5% of all newly diagnosed metastatic cancers, of which the primary source is difficult to classify with current histological methods. Effective cancer treatment depends on early and accurate identification 220 samples
Project description:It is widely accepted that long-term changes in synapse structure and function are mediated by rapid activity-dependent gene transcription and new protein synthesis. A growing amount of evidence suggests that the microRNA (miRNA) pathway plays an important role in coordinating these processes. Despite recent advances in this field, there remains a critical need to identify specific activity-regulated miRNAs as well as their key messenger RNA (mRNA) targets. To address these questions, we used the larval Drosophila melanogaster neuromuscular junction (NMJ) as a model synapse in which to identify novel miRNA-mediated mechanisms that control activity-dependent synaptic growth. First, we developed a screen to identify miRNAs differentially regulated in the larval CNS following spaced synaptic stimulation. Surprisingly, we identified five miRNAs (miRs-1, -8, -289, -314, and -958) that were significantly downregulated by activity. Neuronal misexpression of three miRNAs (miRs-8, -289, and -958) suppressed activity-dependent synaptic growth suggesting that these miRNAs control the translation of biologically relevant target mRNAs. Functional annotation cluster analysis revealed that putative targets of miRs-8 and -289 are significantly enriched in clusters involved in the control of neuronal processes including axon development, pathfinding, and growth. In support of this, miR-8 regulated the expression of a wingless 3M-bM-^@M-^YUTR (wg 3M-bM-^@M-^Y untranslated region) reporter in vitro. Wg is an important presynaptic regulatory protein required for activity-dependent axon terminal growth at the fly NMJ. In conclusion, our results are consistent with a model where key activity-regulated miRNAs are required to coordinate the expression of genes involved in activity-dependent synaptogenesis. larval CNS of UAS-ChR2 x C380-Gal4 following synaptic stimulation
Project description:MicroRNA expression profiling in matched lesional skin samples from 25 patients with psoriasis using the miRNA analysis platform miRCURY LNATM MicroRNA array (v. 11.0) (Exiqon). Aim: To explore the effect of three different preservation methods (Formalin-fixation paraffin-embedding (FFPE), frozen (FS) and OCT-embedding (OCT)) on miRNA expression levels in matched lesional skin samples from 25 patients with psoriasis. Three-condition experiment, FS vs. FFPE, OCT vs. FFPE and FS vs. OCT. Biological replicates: 25 matched samples from patients with psoriasis. One replicate per array.
Project description:Upon muscle injury the high mobility group box 1 (HMGB1) protein is up-regulated and secreted to initiate reparative responses. Here we show that HMGB1 controls myogenesis both in vitro and in vivo, during development and after adult muscle injury. HMGB1 expression in muscle cells is regulated at the translational level: the miRNA miR-1192 inhibits HMGB1 translation and the RNA-binding protein HuR promotes it. HuR binds to a cis-element, HuRBS, located in the 3'UTR of the HMGB1 transcript, and at the same time miR-1192 is recruited to an adjacent seed element. The binding of HuR to the HuRBS prevents the recruitment of Argonaute 2 (Ago2), overriding miR-1192-mediated translation inhibition. Depleting HuR reduces myoblast fusion and silencing miR-1192 re-establishes the fusion potential of HuR-depleted cells. We propose that HuR promotes the commitment of myoblasts to myogenesis by enhancing the translation of HMGB1 and suppressing the translation inhibition mediated by miR-1192. RNA content was extracted following immunoprecipitation of HuR using a monoclonal antibody (3A2) and the levels of mRNA were compared to an IgG control in order to determine which transcripts were enriched in the HuR ribonucleoprotein complex.
Project description:Endothelial-enriched total RNAs were obtained from the suprarenal region of the abdominal aorta which is the murine AAA prone area in AngII-infused C57BL/6 mice. At 12 or 36h post-AngII pump implantation, endothelial-enriched RNAs from four abdominal aortas were pooled to obtain ~30 ng total RNA as one array sample, performed in triplicates. All RNA samples used for miRNA microarray study passed the initial quality control test and each sample was linearly amplified.
Project description:Inflammatory conditions can contribute to tumor formation. However, any clear marker predicting progression to cancer are still lacking. The aim of our study was to analyze microRNA modulations accompanying inflammation-induced tumor development to determine whether these microRNA may jointly affect the expression of genes involved in cancer. For this purpose, we used the well-established azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced mouse model of colitis-associated cancer. We performed a microRNA microarray to establish microRNA expression profiles in mouse whole colon at early and late time points during inflammation and/or tumor growth. Chronic inflammation and carcinogenesis were associated with distinct changes in microRNA expression. Nevertheless, prediction algorithms of microRNA-mRNA interactions and computational analyses based on ranked microRNA lists consistently identified putative target genes that play essential roles in tumor growth or belong to key carcinogenesis-related networks or signaling pathways. Hence, inflammation, through microRNA, may affect unexpected genes or signaling pathways, thereby contributing to carcinogenesis. The present method can lead to the identification of novel genes or signaling pathways involved in cancer development. miRNA microarray profiling in whole mouse colon at 4 time points during AOM/DSS treatment. Controls : PBS, DSS alone or AOM alone, at two time points; 10 experimental conditions, 5 replicates per experimental conditions, one replicate per array hybridized in dual color with a commercial reference (Universal Reference, Miltenyi Biotec GmbH)