MicroRNA alterations of pancreatic intraepithelial neoplasias.
ABSTRACT: MicroRNA (miRNA) alterations are likely to contribute to the development of pancreatic cancer and may serve as markers for the early detection of pancreatic neoplasia.To identify the miRNA alterations that arise during the development of pancreatic cancer, we determined the levels of 735 miRNAs in 34 pancreatic intraepithelial neoplasias (PanIN) and 15 normal pancreatic duct samples isolated by laser capture microdissection using TaqMan miRNA microarrays. Differential expression of selected miRNAs was confirmed by FISH analysis and by quantitative real-time reverse transcription PCR (qRT-PCR) analysis of selected candidate miRNAs in an independent set of PanIN and normal duct samples.We identified 107 aberrantly expressed miRNAs in different PanIN grades compared with normal pancreatic duct samples and 35 aberrantly expressed miRNAs in PanIN-3 lesions compared with normal pancreatic duct samples. These differentially expressed miRNAs included those that have been previously identified as differentially expressed in pancreatic ductal adenocarcinomas (PDAC; including miR-21, miR-200a/b/c, miR-216a/b, miR-217, miR-146a, miR-155, miR-182, miR-196b, miR-203, miR-222, miR-338-3p, miR-486-3p, etc.) as well as miRNAs not previously described as differentially expressed in these lesions (miR-125b, miR-296-5p, miR-183*, miR-603, miR-625/*, miR-708, etc.). miR-196b was the most selectively differentially expressed miRNA in PanIN-3 lesions.Many miRNAs undergo aberrant expression in PanIN lesions and are likely to be important in the development of PDAC. The miRNAs, such as miR-196b, whose expression is limited to PanIN-3 lesions or pancreatic cancers could be useful as diagnostic markers.
Project description:The deregulated presence or absence of microRNAs (miRNAs) might play an important role in molecular pathways leading to neoplastic transformation. At present, it is also thought that the approaches to interfere miRNA functions should be helpful for developing novel therapeutic opportunities for human cancer. In this study, we provide evidence that the anticancer agent benzyl isothiocyanate (BITC) has the ability to modulate the level of miRNAs such as miR-221 and miR-375, known to be abnormally expressed in pancreatic cancer patients. Interestingly, ectopic expression of miR-375 or the enforced silencing of miR-221 in cultured pancreatic cancer cells attenuates cell viability and sensitizes antiproliferative action of BITC. We also show that the expression of putative tumor suppressor miR-375 is more abundant in nonpathological mice pancreata than those with Kras(G12D)-driven pancreatic intraepithelial neoplasia (PanIN). To the contrary, the expression of oncogenic miR-221 is significantly elevated in the mouse pancreas with PanIN lesions. Although miR-375 has been shown to be aberrantly expressed in pancreatic cancer patients, there has not been a comprehensive study to investigate the molecular pathways targeted by this miRNA in pancreatic cancer cells. Further analysis by gene expression microarray revealed that IGFBP5 and CAV-1, potential biomarkers of pancreatic cancer, were significantly downregulated in cells transfected with miR-375. Correlatively, elevated expression of IGFBP5 and CAV-1 was evident in the mouse pancreas with preneoplastic lesions in which the expression of miR-375 wanes. Taken together, our findings suggest that anticancer agent BITC might target the expression of miR-221 and miR-375 to switch hyperproliferative pancreatic cancer cells to a hypoproliferative state.
Project description:Pancreatic ductal adenocarcinoma evolves from precursor lesions, the most common of which is pancreatic intraepithelial neoplasia (PanIN). We performed RNA-sequencing analysis of laser capture microdissected PanINs and normal pancreatic duct cells to identify differentially expressed genes between PanINs and normal pancreatic duct, and between low-grade and high-grade PanINs. One of the most highly overexpressed transcripts identified in PanIN is interleukin-2 receptor subunit gamma (IL2RG) encoding the common gamma chain, IL2R?. CRISPR-mediated knockout of IL2RG in orthotopically implanted pancreatic cancer cells resulted in attenuated tumor growth in mice and reduced JAK3 expression in orthotopic tumors. These results indicate that IL2R?/JAK3 signaling contributes to pancreatic cancer cell growth in vivo.
Project description:The molecular mechanisms of lung injury and fibrosis are incompletely understood. MicroRNAs (miRNAs) are crucial biological regulators that act by suppressing their target genes and are involved in a variety of pathophysiological processes. To gain insight into miRNAs in the regulation of lung fibrosis, total RNA was isolated from mouse lungs harvested at different days after bleomycin treatment, and miRNA array with 1,810 miRNA probes was performed thereafter. MiRNAs expressed in lungs with bleomycin treatment at different time points were compared with miRNAs expressed in lungs without bleomycin treatment, resulting in 161 miRNAs differentially expressed. Furthermore, miRNA expression patterns regulated in initial and late periods after bleomycin were identified. Target genes were predicted in silico for differentially expressed miRNAs, including let-7f, let-7g, miR-196b, miR-16, miR-195, miR-25, miR-144, miR-351, miR-153, miR-468, miR-449b, miR-361, miR-700, miR-704, miR-717, miR-10a, miR-211, miR-34a, miR-367, and miR-21. Target genes were then cross-referenced to the molecular pathways, suggesting that the differentially expressed miRNAs regulate apoptosis, Wnt, Toll-like receptor, and TGF-? signaling. Our study demonstrated a relative abundance of miRNA levels in bleomycin-induced lung fibrosis. The miRNAs and their potential target genes identified may contribute to the understanding of the complex transcriptional program of lung fibrosis.
Project description:This study aims to identify the pivotal microRNAs (miRNAs) and genes, and their potential regulatory mechanisms in pancreatic ductal adenocarcinoma (PDAC) through bioinformatics analysis and experimental verification. We comprehensively analyzed two miRNA microarray datasets (GSE32678 and GSE43796) and three gene microarray datasets (GSE28735, GSE41368 and GSE71989), which were downloaded from the Gene Expression Omnibus (GEO) database, and identified the total of 8 differentially expressed miRNAs (DEMs) and 257 differentially expressed genes (DEGs) in common. Next, a new miRNA-mRNA regulatory network was constructed by bioinformatics methods, including 7 miRNAs, 58 putative target genes and 80 interaction pairs of miRNA-mRNA. Scrutinized by OncoLnc and GEPIA, it was found that 3 of 7 miRNAs (miR-21, miR-196b and miR-203) and 20 of 58 genes (MXRA5, EPYC, ECT2, COL12A1, SLC6A14, SLC7A2, BTG2, PDK4, CTNND2, NRP2, PXDN, CD109, TGFBI, LRRN1, ITGA2, DKK1, GREM1, EFNB2, SEMA3C and NT5E) were notably associated with prognosis in patients with PDAC. Furthermore, EFNB2 was significantly upregulated in PDAC compared with normal controls from different public databases. Cellular function experiments demonstrated that EFNB2 knockdown inhibited cell proliferation, migration and invasion in SW1990 cells. Western blot and luciferase reporter assays revealed that miR-557 negatively regulated the expression of EFNB2 by directly binging its 3' UTR. In conclusion, we performed integrated analysis for multiple expression profiles, and provided novel candidate miRNAs and genes to be exploited for functional studies. In addition, our findings suggested that EFNB2 contributes to PDAC progression by acting as the target gene of miR-557. It is useful for uncovering miRNA-based treatments in PDAC.
Project description:<h4>Objectives</h4>Seizure is a common presenting symptom of primary brain tumors. There are no published studies regarding the roles of MicroRNA (miRNA) in tumor-related epilepsy. The authors set out to correlate miR-196b expression in low-grade glioma patients with pre-operative seizures and post-operative seizure control.<h4>Methods</h4>Twenty-three patients with WHO grade II astrocytomas and 83 similar patients for independent validation were included. Follow-up visits regarding seizure prognosis were scheduled at 6 months. MiRNA profiling was used to identify differentially expressed miRNAs. The most important miRNA was determined by quantitative reverse-transcriptase polymerase chain reaction (q-PCR) in the validation cohort. Gene ontology (GO) analysis and whole genome mRNA profiling was performed to investigate the underlying biological processes.<h4>Results</h4>Array results showed that 30 miRNAs were overexpressed and 10 miRNAs were underexpressed (with more than 2 fold change) in patients with pre-operative seizures. MiR-196b was validated in the independent validation cohort. Patients with good seizure prognosis exhibited low levels of miR-196b expression compared with those who had poor seizure prognosis in the group without pre-operative seizures. Biological processes that relate to transcription and cell cycles were over-represented in the miR-196b-associated gene expression signature. MiR-196b-associated gene expression profiling was characterized by enrichment of genes usually involved in cell proliferation.<h4>Conclusions</h4>We have provided the first evidence that expression of miR-196b was associated with the occurrence of pre-operative seizures in low-grade gliomas, and may predict seizure prognosis in patients without pre-operative seizures. Targeted treatments that decrease endogenous levels of miR-196b might represent novel therapeutic strategies.
Project description:Pancreatic cancer (PC), as the leading cause of cancer death worldwide, is one of the deadliest tumors with a very low 5-year survival rate. Therefore, it is urgent to seek new biomarkers of PC for more accurate and reliable treatments. To identify the differentially expressed miRNAs (DEM) in PC tissues, we performed the systematic microarray and qRT-PCR analyses. We found miR-196b was the top dysregulated DEM in PC tissues as compared with the corresponding adjacent tissues, and positively correlated with poor differentiation, tumor size, lymphatic invasion and TNM stage. Furthermore, the late apoptosis rate was significantly reduced, while the cell proliferation was increased in PANC-1 and ASPC-1 cell-lines after treatment with miR-196b mimics. The qRT-PCR and Western blot analysis demonstrated that the level of CADM1 in PANC-1 cells response to the alteration of miR-196b. Moreover, blockade of CADM1 could decrease the late apoptosis in PANC-1 cells as up-regulated by inhibition of miR-196b. Finally, luciferase report assay confirmed that CADM1 was the direct target gene of miR-196b. Overexpression of miR-196b in PC tissues can increase the late apoptosis of pancreatic cancer cells by targeting CADM1. These findings suggested miR-196b is a potential target for diagnosis and therapeutics of human pancreatic cancer.
Project description:Pancreatic cancer is a common malignant tumor with a high incidence and mortality rate. The prognosis of patients with pancreatic cancer is considerably poor due to the lack of effective treatment in clinically. Despite numerous studies have revealed that baicalein, a natural product, is responsible for suppressing multiple cancer cells proliferation, motility and invasion. The mechanism by which baicalein restraining pancreatic cancer progression remains unclear. In this study, we firstly verified that baicalein plays a critical role in inhibiting pancreatic tumorigenesis <i>in vitro</i> and <i>in vivo</i>. Then we analyzed the alteration of microRNAs (miRNAs) expression levels in Panc-1 cells incubated with DMSO, 50 and 100 μM baicalein by High-Throughput sequencing. Intriguingly, we observed that 20 and 39 miRNAs were accordingly up- and down-regulated through comparing Panc-1 cells exposed to 100 μM baicalein with the control group. Quantitative PCR analysis confirmed that miR-139-3p was the most up-regulated miRNA after baicalein treatment, while miR-196b-5p was the most down-regulated miRNA. Further studies showed that miR-139-3p induced, miR-196b-5p inhibited the apoptosis of Panc-1 cells <i>via</i> targeting NOB1 and ING5 respectively. In conclusion, we demonstrated that baicalein is a potent inhibitor against pancreatic cancer by modulating the expression of miR-139-3p or miR-196b-5p.
Project description:Current diagnostic tools for pancreatic cysts fail to reliably differentiate mucinous from nonmucinous cysts. Reliable biomarkers are needed. MicroRNAs (miRNA) may offer insights into pancreatic cysts. Our aims were to (1) identify miRNAs that distinguish benign from both premalignant cysts and malignant pancreatic lesions using formalin-fixed, paraffin-embedded (FFPE) pathology specimens; (2) identify miRNAs that distinguish mucinous cystic neoplasm (MCN) from branch duct-intraductal papillary mucinous neoplasm (BD-IPMN).A total of 69 FFPE pancreatic specimens were identified: (1) benign (20 serous cystadenoma (SCA)), (2) premalignant (10 MCN, 10 BD-IPMN, 10 main duct IPMN (MD-IPMN)), and (3) malignant (19 pancreatic ductal adenocarcinoma (PDAC)). Total nucleic acid extraction was performed followed by miRNA expression profiling of 378 miRNAs interrogated using TaqMan MicroRNA Arrays Pool A and verification of candidate miRNAs. Bioinformatics was used to generate classifiers.MiRNA profiling of 69 FFPE specimens yielded 35 differentially expressed miRNA candidates. Four different 4-miRNA panels differentiated among the lesions: one panel separated SCA from MCN, BD-IPMN, MD-IPMN, and PDAC with sensitivity 85% (62, 97), specificity 100% (93, 100), a second panel distinguished MCN from SCA, BD-IPMN, MD-IPMN, and PDAC with sensitivity and specificity 100% (100, 100), a third panel differentiated PDAC from IPMN with sensitivity 95% (76, 100) and specificity 85% (72, 96), and the final panel diagnosed MCN from BD-IPMN with sensitivity and specificity approaching 100%.MiRNA profiling of surgical pathology specimens differentiates serous cystadenoma from both premalignant pancreatic cystic neoplasms and PDAC and MCN from BD-IPMN.
Project description:MiRNAs regulate gene expression by post-transcriptionally suppressing mRNA translation or by causing mRNA degradation. It has been proposed that unique miRNAs influence specific tumor molecular phenotype. In this paper, we test the hypotheses that miRNA expression differs by tumor molecular phenotype and that those differences may influence prognosis. Data come from population-based studies of colorectal cancer conducted in Utah and the Northern California Kaiser Permanente Medical Care Program. A total of 1893 carcinoma samples were run on the Agilent Human miRNA Microarray V19.0 containing 2006 miRNAs. We assessed differences in miRNA expression between TP53-mutated and non-mutated, KRAS-mutated and non-mutated, BRAF-mutated and non-mutated, CpG island methylator phenotype (CIMP) high and CIMP low, and microsatellite instability (MSI) and microsatellite stable (MSS) colon and rectal tumors. Using a Cox proportional hazard model we evaluated if those miRNAs differentially expressed by tumor phenotype influenced survival after adjusting for age, sex, and AJCC stage. There were 22 differentially expressed miRNAs for TP53-mutated colon tumors and 5 for TP53-mutated rectal tumors with a fold change of >1.49 (or <0.67). Additionally, 13 miRNAS were differentially expressed for KRAS-mutated rectal tumors, 8 differentially expressed miRNAs for colon CIMP high tumors, and 2 differentially expressed miRNAs for BRAF-mutated colon tumors. The majority of differentially expressed miRNAS were observed between MSI and MSS tumors (94 differentially expressed miRNAs for colon; 41 differentially expressed miRNAs for rectal tumors). Of these miRNAs differentially expressed between MSI and MSS tumors, the majority were downregulated. Ten of the differentially expressed miRNAs were associated with survival; after adjustment for MSI status, five miRNAS, miR-196b-5p, miR-31-5p, miR-99b-5p, miR-636, and miR-192-3p, were significantly associated with survival. In summary, it appears that the majority of miRNAs that are differentially expressed by tumor molecular phenotype are MSI tumors. However, these miRNAs appear to have minimal effect on prognosis.
Project description:The molecular mechanisms of lung injury and fibrosis are incompletely understood. microRNAs (miRNAs) are crucial biological regulators by suppression of their target genes and are involved in a variety of pathophysiologic processes. To gain insight into miRNAs in the regulation of lung fibrosis, total RNA was isolated from lung samples harvested at different days after bleomycin treatment, and miRNA array was performed thereafter. miRNAs expressed in lungs with bleomycin treatment at different time points were compared to miRNAs expressed in lungs without bleomycin treatment, resulting in 161 miRNAs differentially expressed. Furthermore, miRNA expression patterns regulated in initial and late periods after bleomycin were identified. Target genes were predicted in silico for differentially expressed miRNAs, including miR-7f, miR-7g, miR-196b, miR-16, miR-195, miR-25, miR-144, miR-351, miR-34a, miR-499, miR-704, miR-717, miR-10a, miR-211, miR-34a, miR-367 and miR-21, and then cross-referenced to molecular pathways including apoptosis, Wnt, Toll-like receptor, and TGF-? signaling, which are involved in different pathological phenotypes such as apoptosis, inflammation, and fibrosis. Our study demonstrated relative abundance of miRNA levels in bleomycin-induced lung fibrosis. The miRNAs and their potential target genes identified herein contribute to the understanding of the complex transcriptional program of lung fibrosis. Under anesthesia, 2.5 U/kg bleomycin dissolved in sterile PBS was administered via trachea as previously described. Lung tissues were harvested at the time point of day 0, 3, 7, 14, and 21 post bleomycin challenges. 3 sample in specific time point, except for day 14 where nday14 = 2.