Optimized high-throughput microRNA expression profiling provides novel biomarker assessment of clinical prostate and breast cancer biopsies

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Fedele Vita

2006-06-01

Full Text Available Abstract Background Recent studies indicate that microRNAs (miRNAs are mechanistically involved in the development of various human malignancies, suggesting that they represent a promising new class of cancer biomarkers. However, previously reported methods for measuring miRNA expression consume large amounts of tissue, prohibiting high-throughput miRNA profiling from typically small clinical samples such as excision or core needle biopsies of breast or prostate cancer. Here we describe a novel combination of linear amplification and labeling of miRNA for highly sensitive expression microarray profiling requiring only picogram quantities of purified microRNA. Results Comparison of microarray and qRT-PCR measured miRNA levels from two different prostate cancer cell lines showed concordance between the two platforms (Pearson correlation R2 = 0.81; and extension of the amplification, labeling and microarray platform was successfully demonstrated using clinical core and excision biopsy samples from breast and prostate cancer patients. Unsupervised clustering analysis of the prostate biopsy microarrays separated advanced and metastatic prostate cancers from pooled normal prostatic samples and from a non-malignant precursor lesion. Unsupervised clustering of the breast cancer microarrays significantly distinguished ErbB2-positive/ER-negative, ErbB2-positive/ER-positive, and ErbB2-negative/ER-positive breast cancer phenotypes (Fisher exact test, p = 0.03; as well, supervised analysis of these microarray profiles identified distinct miRNA subsets distinguishing ErbB2-positive from ErbB2-negative and ER-positive from ER-negative breast cancers, independent of other clinically important parameters (patient age; tumor size, node status and proliferation index. Conclusion In sum, these findings demonstrate that optimized high-throughput microRNA expression profiling offers novel biomarker identification from typically small clinical samples such as breast

Epigallocatechin Gallate-Mediated Alteration of the MicroRNA Expression Profile in 5α-Dihydrotestosterone-Treated Human Dermal Papilla Cells.

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Shin, Shanghun; Kim, Karam; Lee, Myung Joo; Lee, Jeongju; Choi, Sungjin; Kim, Kyung-Suk; Ko, Jung-Min; Han, Hyunjoo; Kim, Su Young; Youn, Hae Jeong; Ahn, Kyu Joong; An, In-Sook; An, Sungkwan; Cha, Hwa Jun

2016-06-01

Dihydrotestosterone (DHT) induces androgenic alopecia by shortening the hair follicle growth phase, resulting in hair loss. We previously demonstrated how changes in the microRNA (miRNA) expression profile influenced DHT-mediated cell death, cell cycle arrest, cell viability, the generation of reactive oxygen species (ROS), and senescence. Protective effects against DHT have not, however, been elucidated at the genome level. We showed that epigallocatechin gallate (EGCG), a major component of green tea, protects DHT-induced cell death by regulating the cellular miRNA expression profile. We used a miRNA microarray to identify miRNA expression levels in human dermal papilla cells (DPCs). We investigated whether the miRNA expression influenced the protective effects of EGCG against DHT-induced cell death, growth arrest, intracellular ROS levels, and senescence. EGCG protected against the effects of DHT by altering the miRNA expression profile in human DPCs. In addition, EGCG attenuated DHT-mediated cell death and growth arrest and decreased intracellular ROS levels and senescence. A bioinformatics analysis elucidated the relationship between the altered miRNA expression and EGCG-mediated protective effects against DHT. Overall, our results suggest that EGCG ameliorates the negative effects of DHT by altering the miRNA expression profile in human DPCs.

Differential expression analysis of balding and nonbalding dermal papilla microRNAs in male pattern baldness with a microRNA amplification profiling method.

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Goodarzi, H R; Abbasi, A; Saffari, M; Fazelzadeh Haghighi, M; Tabei, M B; Noori Daloii, M R

2012-05-01

  Male pattern baldness or androgenetic alopecia is a common disorder affecting almost 50% of men throughout their lifetime, with androgens and genetics having significant contributing aetiologies. In contrast to the positive regulatory effect of androgens on body hair growth, they are thought to alter scalp hair follicle behaviour pathophysiologically, leading to male pattern baldness. However, the exact mechanisms of this paradoxical action have not yet been elucidated. The role of microRNAs, a novel group of noncoding RNAs impacting almost every aspect of biology, health and human diseases, has been documented in hair follicle formation. In addition, their deregulation in cancer of the prostate, a target organ of androgens, has also been well established. To investigate the possible contribution of microRNAs in the pathophysiology of male pattern baldness. We initially screened microRNA expression profiles of balding and nonbalding hair follicle papillae with a sensitive microRNA cloning method, microRNA amplification profiling, and statistically analysed significant differentially expressed microRNAs in balding relative to nonbalding dermal papillae, with real-time polymerase chain reaction as a confirmatory method to quantify expression in eight individuals affected with the disorder.   We detected the significant upregulation of miR-221, miR-125b, miR-106a and miR-410 in balding papilla cells.   We found four microRNAs that could participate in the pathogenesis of male pattern baldness. Regarding the strong therapeutic potential of microRNAs and the easy accessibility of hair follicles for gene therapy, microRNAs are possible candidates for a new generation of revolutionary treatments. © 2011 The Authors. BJD © 2011 British Association of Dermatologists.

MicroRNA expression profiles associated with pancreatic adenocarcinoma and ampullary adenocarcinoma

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Schultz, Nicolai A; Werner, Jens; Willenbrock, Hanni

2012-01-01

MicroRNAs have potential as diagnostic cancer biomarkers. The aim of this study was (1) to define microRNA expression patterns in formalin-fixed parafin-embedded tissue from pancreatic ductal adenocarcinoma, ampullary adenocarcinoma, normal pancreas and chronic pancreatitis without using micro-di...

Ionizing Radiation Deregulates the MicroRNA Expression Profile in Differentiated Thyroid Cells.

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Penha, Ricardo Cortez Cardoso; Pellecchia, Simona; Pacelli, Roberto; Pinto, Luis Felipe Ribeiro; Fusco, Alfredo

2018-03-01

Ionizing radiation (IR) is a well-known risk factor for papillary thyroid cancer, and it has been reported to deregulate microRNA expression, which is important to thyroid carcinogenesis. Therefore, this study investigated the impact of IR on microRNA expression profile of the normal thyroid cell line (FRTL-5 CL2), as well as its effect on radiosensitivity of thyroid cancer cell lines, especially the human anaplastic thyroid carcinoma cell line (8505c). The global microRNA expression profile of irradiated FRTL-5 CL2 cells (5 Gy X-ray) was characterized, and data were confirmed by quantitative real-time polymerase chain reaction evaluating the expression of rno-miR-10b-5p, rno-miR-33-5p, rno-miR-128-1-5p, rno-miR-199a-3p, rno-miR-296-5p, rno-miR-328a-3p, and rno-miR-541-5p in irradiated cells. The miR-199a-3p and miR-10b-5p targets were validated by quantitative real-time polymerase chain reaction, Western blot, and luciferase target assays. The effects of miR-199a-3p and miR-10b-5p on DNA repair were determined by evaluating the activation of the protein kinases ataxia-telangiectasia mutated, ataxia telangiectasia, and Rad3-related and the serine 39 phosphorylation of variant histone H2AX as an indirect measure of double-strand DNA breaks in irradiated FRTL-5 CL2 cells. The impact of miR-10b-5p on radiosensitivity was analyzed by cell counting and MTT assays in FRTL-5 CL2, Kras-transformed FRTL-5 CL2 (FRTL KiKi), and 8505c cell lines. The results reveal that miR-10b-5p and miR-199a-3p display the most pronounced alterations in expression in irradiated FRTL-5 CL2 cells. Dicer1 and Lin28b were validated as targets of miR-10b-5p and miR-199a-3p, respectively. Functional studies demonstrate that miR-10b-5p increases the growth rate of FRTL-5 CL2 cells, while miR-199a-3p inhibits their proliferation. Moreover, both of these microRNAs negatively affect homologous recombination repair, reducing activated ataxia-telangiectasia mutated and Rad3-related protein levels

MicroRNA Expression Profiling of Human Respiratory Epithelium Affected by Invasive Candida Infection.

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Syed Aun Muhammad

Full Text Available Invasive candidiasis is potentially life-threatening systemic fungal infection caused by Candida albicans (C. albicans. Candida enters the blood stream and disseminate throughout the body and it is often observed in hospitalized patients, immunocompromised individuals or those with chronic diseases. This infection is opportunistic and risk starts with the colonization of C. albicans on mucocutaneous surfaces and respiratory epithelium. MicroRNAs (miRNAs are small non-coding RNAs which are involved in the regulation of virtually every cellular process. They regulate and control the levels of mRNA stability and post-transcriptional gene expression. Aberrant expression of miRNAs has been associated in many disease states, and miRNA-based therapies are in progress. In this study, we investigated possible variations of miRNA expression profiles of respiratory epithelial cells infected by invasive Candida species. For this purpose, respiratory epithelial tissues of infected individuals from hospital laboratory were accessed before their treatment. Invasive Candida infection was confirmed by isolation of Candia albicans from the blood cultures of the same infected individuals. The purity of epithelial tissues was assessed by flow cytometry (FACSCalibur cytometer; BD Biosciences, Heidelberg, Germany using statin antibody (S-44. TaqMan quantitative real-time PCR (in a TaqMan Low Density Array format was used for miRNA expression profiling. MiRNAs investigated, the levels of expression of 55 miRNA were significantly altered in infected tissues. Some miRNAs showed dramatic increase (miR-16-1 or decrease of expression (miR-17-3p as compared to control. Gene ontology enrichment analysis of these miRNA-targeted genes suggests that Candidal infection affect many important biological pathways. In summary, disturbance in miRNA expression levels indicated the change in cascade of pathological processes and the regulation of respiratory epithelial functions

MicroRNAs expression profile in solid and unicystic ameloblastomas

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Setién-Olarra, A.; Bediaga, N. G.; Aguirre-Echebarria, P.; Aguirre-Urizar, J. M.; Mosqueda-Taylor, A.

2017-01-01

Objectives Odontogenic tumors (OT) represent a specific pathological category that includes some lesions with unpredictable biological behavior. Although most of these lesions are benign, some, such as the ameloblastoma, exhibit local aggressiveness and high recurrence rates. The most common types of ameloblastoma are the solid/multicystic (SA) and the unicystic ameloblastoma (UA); the latter considered a much less aggressive entity as compared to the SA. The microRNA system regulates the expression of many human genes while its deregulation has been associated with neoplastic development. The aim of the current study was to determine the expression profiles of microRNAs present in the two most common types of ameloblastomas. Material & methods MicroRNA expression profiles were assessed using TaqMan® Low Density Arrays (TLDAs) in 24 samples (8 SA, 8 UA and 8 control samples). The findings were validated using quantitative RTqPCR in an independent cohort of 19 SA, 8 UA and 19 dentigerous cysts as controls. Results We identified 40 microRNAs differentially regulated in ameloblastomas, which are related to neoplastic development and differentiation, and with the osteogenic process. Further validation of the top ranked microRNAs revealed significant differences in the expression of 6 of them in relation to UA, 7 in relation to SA and 1 (miR-489) that was related to both types. Conclusion We identified a new microRNA signature for the ameloblastoma and for its main types, which may be useful to better understand the etiopathogenesis of this neoplasm. In addition, we identified a microRNA (miR-489) that is suggestive of differentiating among solid from unicystic ameloblastoma. PMID:29053755

Characterisation of microRNA expression in post-natal mouse mammary gland development

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Karagavriilidou Konstantina

2009-11-01

Full Text Available Abstract Background The differential expression pattern of microRNAs (miRNAs during mammary gland development might provide insights into their role in regulating the homeostasis of the mammary epithelium. Our aim was to analyse these regulatory functions by deriving a comprehensive tissue-specific combined miRNA and mRNA expression profile of post-natal mouse mammary gland development. We measured the expression of 318 individual murine miRNAs by bead-based flow-cytometric profiling of whole mouse mammary glands throughout a 16-point developmental time course, including juvenile, puberty, mature virgin, gestation, lactation, and involution stages. In parallel whole-genome mRNA expression data were obtained. Results One third (n = 102 of all murine miRNAs analysed were detected during mammary gland development. MicroRNAs were represented in seven temporally co-expressed clusters, which were enriched for both miRNAs belonging to the same family and breast cancer-associated miRNAs. Global miRNA and mRNA expression was significantly reduced during lactation and the early stages of involution after weaning. For most detected miRNA families we did not observe systematic changes in the expression of predicted targets. For miRNA families whose targets did show changes, we observed inverse patterns of miRNA and target expression. The data sets are made publicly available and the combined expression profiles represent an important community resource for mammary gland biology research. Conclusion MicroRNAs were expressed in likely co-regulated clusters during mammary gland development. Breast cancer-associated miRNAs were significantly enriched in these clusters. The mechanism and functional consequences of this miRNA co-regulation provide new avenues for research into mammary gland biology and generate candidates for functional validation.

Circulating microRNA Profile throughout the menstrual cycle.

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Kadri Rekker

Full Text Available Normal physiological variables, such as age and gender, contribute to alterations in circulating microRNA (miRNA expression levels. The changes in the female body during the menstrual cycle can also be reflected in plasma miRNA expression levels. Therefore, this study aimed to determine the plasma miRNA profile of healthy women during the menstrual cycle and to assess which circulating miRNAs are derived from blood cells. The plasma miRNA expression profiles in nine healthy women were determined by quantitative real time PCR using Exiqon Human Panel I assays from four time-points of the menstrual cycle. This platform was also used for studying miRNAs from pooled whole blood RNA samples at the same four time-points. Our results indicated that circulating miRNA expression levels in healthy women were not significantly altered by the processes occurring during the menstrual cycle. No significant differences in plasma miRNA expression levels were observed between the menstrual cycle time-points, but the number of detected miRNAs showed considerable variation among the studied individuals. miRNA analysis from whole blood samples revealed that majority of miRNAs in plasma are derived from blood cells. The most abundant miRNA in plasma and blood was hsa-miR-451a, but a number of miRNAs were only detected in one or the other sample type. In conclusion, our data suggest that the changes in the female body during the menstrual cycle do not affect the expression of circulating miRNAs at measurable levels.

Paper-Based MicroRNA Expression Profiling from Plasma and Circulating Tumor Cells.

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Leong, Sai Mun; Tan, Karen Mei-Ling; Chua, Hui Wen; Huang, Mo-Chao; Cheong, Wai Chye; Li, Mo-Huang; Tucker, Steven; Koay, Evelyn Siew-Chuan

2017-03-01

Molecular characterization of circulating tumor cells (CTCs) holds great promise for monitoring metastatic progression and characterizing metastatic disease. However, leukocyte and red blood cell contamination of routinely isolated CTCs makes CTC-specific molecular characterization extremely challenging. Here we report the use of a paper-based medium for efficient extraction of microRNAs (miRNAs) from limited amounts of biological samples such as rare CTCs harvested from cancer patient blood. Specifically, we devised a workflow involving the use of Flinders Technology Associates (FTA) ® Elute Card with a digital PCR-inspired "partitioning" method to extract and purify miRNAs from plasma and CTCs. We demonstrated the sensitivity of this method to detect miRNA expression from as few as 3 cancer cells spiked into human blood. Using this method, background miRNA expression was excluded from contaminating blood cells, and CTC-specific miRNA expression profiles were derived from breast and colorectal cancer patients. Plasma separated out during purification of CTCs could likewise be processed using the same paper-based method for miRNA detection, thereby maximizing the amount of patient-specific information that can be derived from a single blood draw. Overall, this paper-based extraction method enables an efficient, cost-effective workflow for maximized recovery of small RNAs from limited biological samples for downstream molecular analyses. © 2016 American Association for Clinical Chemistry.

MicroRNA expression characterizes oligometastasis(es).

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Lussier, Yves A; Xing, H Rosie; Salama, Joseph K; Khodarev, Nikolai N; Huang, Yong; Zhang, Qingbei; Khan, Sajid A; Yang, Xinan; Hasselle, Michael D; Darga, Thomas E; Malik, Renuka; Fan, Hanli; Perakis, Samantha; Filippo, Matthew; Corbin, Kimberly; Lee, Younghee; Posner, Mitchell C; Chmura, Steven J; Hellman, Samuel; Weichselbaum, Ralph R

2011-01-01

Cancer staging and treatment presumes a division into localized or metastatic disease. We proposed an intermediate state defined by ≤ 5 cumulative metastasis(es), termed oligometastases. In contrast to widespread polymetastases, oligometastatic patients may benefit from metastasis-directed local treatments. However, many patients who initially present with oligometastases progress to polymetastases. Predictors of progression could improve patient selection for metastasis-directed therapy. Here, we identified patterns of microRNA expression of tumor samples from oligometastatic patients treated with high-dose radiotherapy. Patients who failed to develop polymetastases are characterized by unique prioritized features of a microRNA classifier that includes the microRNA-200 family. We created an oligometastatic-polymetastatic xenograft model in which the patient-derived microRNAs discriminated between the two metastatic outcomes. MicroRNA-200c enhancement in an oligometastatic cell line resulted in polymetastatic progression. These results demonstrate a biological basis for oligometastases and a potential for using microRNA expression to identify patients most likely to remain oligometastatic after metastasis-directed treatment.

Changes in Rat Brain MicroRNA Expression Profiles Following Sevoflurane and Propofol Anesthesia

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Yu Lu

2015-01-01

Full Text Available Background: Sevoflurane and propofol are widely used anesthetics for surgery. Studies on the mechanisms of general anesthesia have focused on changes in protein expression properties and membrane lipid. MicroRNAs (miRNAs regulate neural function by altering protein expression. We hypothesize that sevoflurane and propofol affect miRNA expression profiles in the brain, expect to understand the mechanism of anesthetic agents. Methods: Rats were randomly assigned to a 2% sevoflurane group, 600 μg·kg − 1·min − 1 propofol group, and a control group without anesthesia (n = 4, respectively. Treatment group was under anesthesia for 6 h, and all rats breathed spontaneously with continuous monitoring of respiration and blood gases. Changes in rat cortex miRNA expression profiles were analyzed by miRNA microarrays and validated by quantitative real-time polymerase chain reaction (qRT-PCR. Differential expression of miRNA using qRT-PCR among the control, sevoflurane, and propofol groups were compared using one-way analysis of variance (ANOVA. Results: Of 677 preloaded rat miRNAs, the microarray detected the expression of 277 miRNAs in rat cortex (40.9%, of which 9 were regulated by propofol and (or sevoflurane. Expression levels of three miRNAs (rno-miR-339-3p, rno-miR-448, rno-miR-466b-1FNx01 were significantly increased following sevoflurane and six (rno-miR-339-3p, rno-miR-347, rno-miR-378FNx01, rno-miR-412FNx01, rno-miR-702-3p, and rno-miR-7a-2FNx01 following propofol. Three miRNAs (rno-miR-466b-1FNx01, rno-miR-3584-5p and rno-miR-702-3p were differentially expressed by the two anesthetic treatment groups. Conclusions: Sevoflurane and propofol anesthesia induced distinct changes in brain miRNA expression patterns, suggesting differential regulation of protein expression. Determining the targets of these differentially expressed miRNAs may help reveal both the common and agent-specific actions of anesthetics on neurological and physiological

MicroRNA profiling in intraocular medulloepitheliomas.

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Deepak P Edward

Full Text Available To study the differential expression of microRNA (miRNA profiles between intraocular medulloepithelioma (ME and normal control tissue (CT.Total RNA was extracted from formalin fixed paraffin embedded (FFPE intraocular ME (n=7 and from age matched ciliary body controls (n=8. The clinical history and phenotype was recorded. MiRNA profiles were determined using the Affymetrix GeneChip miRNA Arrays analyzed using expression console 1.3 software. Validation of significantly dysregulated miRNA was confirmed by quantitative real-time PCR. The web-based DNA Intelligent Analysis (DIANA-miRPath v2.0 was used to perform enrichment analysis of differentially expressed (DE miRNA gene targets in Kyoto Encyclopedia of Genes and Genomes (KEGG pathway.The pathologic evaluation revealed one benign (benign non-teratoid, n=1 and six malignant tumors (malignant teratoid, n=2; malignant non-teratoid, n = 4. A total of 88 miRNAs were upregulated and 43 miRNAs were downregulated significantly (P<0.05 in the tumor specimens. Many of these significantly dysregulated miRNAs were known to play various roles in carcinogenesis and tumor behavior. RT-PCR validated three significantly upregulated miRNAs and three significantly downregulated miRNAs namely miR-217, miR-216a, miR-216b, miR-146a, miR-509-3p and miR-211. Many DE miRNAs that were significant in ME tumors showed dysregulation in retinoblastoma, glioblastoma, and precursor, normal and reactive human cartilage. Enriched pathway analysis suggested a significant association of upregulated miRNAs with 15 pathways involved in prion disease and several types of cancer. The pathways involving significantly downregulated miRNAs included the toll-like receptor (TLR (p<4.36E-16 and Nuclear Factor kappa B (NF-κB signaling pathways (p<9.00E-06.We report significantly dysregulated miRNAs in intraocular ME tumors, which exhibited abnormal profiles in other cancers as well such as retinoblastoma and glioblastoma. Pathway analysis

MicroRNA expression variability in human cervical tissues.

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Patrícia M Pereira

Full Text Available MicroRNAs (miRNAs are short (approximately 22 nt non-coding regulatory RNAs that control gene expression at the post-transcriptional level. Deregulation of miRNA expression has been discovered in a wide variety of tumours and it is now clear that they contribute to cancer development and progression. Cervical cancer is one of the most common cancers in women worldwide and there is a strong need for a non-invasive, fast and efficient method to diagnose the disease. We investigated miRNA expression profiles in cervical cancer using a microarray platform containing probes for mature miRNAs. We have evaluated miRNA expression profiles of a heterogeneous set of cervical tissues from 25 different patients. This set included 19 normal cervical tissues, 4 squamous cell carcinoma, 5 high-grade squamous intraepithelial lesion (HSIL and 9 low-grade squamous intraepithelial lesion (LSIL samples. We observed high variability in miRNA expression especially among normal cervical samples, which prevented us from obtaining a unique miRNA expression signature for this tumour type. However, deregulated miRNAs were identified in malignant and pre-malignant cervical tissues after tackling the high expression variability observed. We were also able to identify putative target genes of relevant candidate miRNAs. Our results show that miRNA expression shows natural variability among human samples, which complicates miRNA data profiling analysis. However, such expression noise can be filtered and does not prevent the identification of deregulated miRNAs that play a role in the malignant transformation of cervical squamous cells. Deregulated miRNAs highlight new candidate gene targets allowing for a better understanding of the molecular mechanism underlying the development of this tumour type.

Changes in apoptotic microRNA and mRNA expression profiling in Caenorhabditis elegans during the Shenzhou-8 mission

International Nuclear Information System (INIS)

Gao Ying; Li Shuai; Xu Dan; Wang Junjun; Sun Yeqing

2015-01-01

Radiation and microgravity exposure have been proven to induce abnormal apoptosis in microRNA (miRNA) and mRNA expression, but whether space conditions, including radiation and microgravity, activate miRNAs to regulate the apoptosis is undetermined. For that purpose, we investigated miRNome and mRNA expression in the ced-1 Caenorhabditis elegans mutant vs the wild-type, both of which underwent spaceflight, spaceflight 1g-centrifuge control and ground control conditions during the Shenzhou-8 mission. Results showed that no morphological changes in the worms were detected, but differential miRNA expression increased from 43 (ground control condition) to 57 and 91 in spaceflight and spaceflight control conditions, respectively. Microgravity altered miRNA expression profiling by decreasing the number and significance of differentially expressed miRNA compared with 1 g incubation during spaceflight. Alterations in the miRNAs were involved in alterations in apoptosis, neurogenesis larval development, ATP metabolism and GTPase-mediated signal transduction. Among these, 17 altered miRNAs potentially involved in apoptosis were screened and showed obviously different expression signatures between space conditions. By integrated analysis of miRNA and mRNA, miR-797 and miR-81 may be involved in apoptosis by targeting the genes ced-10 and both drp-1 and hsp-1, respectively. Compared with ground condition, space conditions regulated apoptosis though a different manner on transcription, by altering expression of seven core apoptotic genes in spaceflight condition, and eight in spaceflight control condition. Results indicate that, miRNA of Caenorhabditis elegans probably regulates apoptotic gene expression in response to space environmental stress, and shows different behavior under microgravity condition compared with 1 g condition in the presence of space radiation. (author)

MicroRNA expression characterizes oligometastasis(es.

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Yves A Lussier

Full Text Available Cancer staging and treatment presumes a division into localized or metastatic disease. We proposed an intermediate state defined by ≤ 5 cumulative metastasis(es, termed oligometastases. In contrast to widespread polymetastases, oligometastatic patients may benefit from metastasis-directed local treatments. However, many patients who initially present with oligometastases progress to polymetastases. Predictors of progression could improve patient selection for metastasis-directed therapy.Here, we identified patterns of microRNA expression of tumor samples from oligometastatic patients treated with high-dose radiotherapy.Patients who failed to develop polymetastases are characterized by unique prioritized features of a microRNA classifier that includes the microRNA-200 family. We created an oligometastatic-polymetastatic xenograft model in which the patient-derived microRNAs discriminated between the two metastatic outcomes. MicroRNA-200c enhancement in an oligometastatic cell line resulted in polymetastatic progression.These results demonstrate a biological basis for oligometastases and a potential for using microRNA expression to identify patients most likely to remain oligometastatic after metastasis-directed treatment.

Comparative mRNA and microRNA expression profiling of three genitourinary cancers reveals common hallmarks and cancer-specific molecular events.

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Xianxin Li

Full Text Available Genome-wide gene expression profile using deep sequencing technologies can drive the discovery of cancer biomarkers and therapeutic targets. Such efforts are often limited to profiling the expression signature of either mRNA or microRNA (miRNA in a single type of cancer.Here we provided an integrated analysis of the genome-wide mRNA and miRNA expression profiles of three different genitourinary cancers: carcinomas of the bladder, kidney and testis.Our results highlight the general or cancer-specific roles of several genes and miRNAs that may serve as candidate oncogenes or suppressors of tumor development. Further comparative analyses at the systems level revealed that significant aberrations of the cell adhesion process, p53 signaling, calcium signaling, the ECM-receptor and cell cycle pathways, the DNA repair and replication processes and the immune and inflammatory response processes were the common hallmarks of human cancers. Gene sets showing testicular cancer-specific deregulation patterns were mainly implicated in processes related to male reproductive function, and general disruptions of multiple metabolic pathways and processes related to cell migration were the characteristic molecular events for renal and bladder cancer, respectively. Furthermore, we also demonstrated that tumors with the same histological origins and genes with similar functions tended to group together in a clustering analysis. By assessing the correlation between the expression of each miRNA and its targets, we determined that deregulation of 'key' miRNAs may result in the global aberration of one or more pathways or processes as a whole.This systematic analysis deciphered the molecular phenotypes of three genitourinary cancers and investigated their variations at the miRNA level simultaneously. Our results provided a valuable source for future studies and highlighted some promising genes, miRNAs, pathways and processes that may be useful for diagnostic or

Combining miRNA and mRNA Expression Profiles in Wilms Tumor Subtypes

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Nicole Ludwig

2016-03-01

Full Text Available Wilms tumor (WT is the most common childhood renal cancer. Recent findings of mutations in microRNA (miRNA processing proteins suggest a pivotal role of miRNAs in WT genesis. We performed miRNA expression profiling of 36 WTs of different subtypes and four normal kidney tissues using microarrays. Additionally, we determined the gene expression profile of 28 of these tumors to identify potentially correlated target genes and affected pathways. We identified 85 miRNAs and 2107 messenger RNAs (mRNA differentially expressed in blastemal WT, and 266 miRNAs and 1267 mRNAs differentially expressed in regressive subtype. The hierarchical clustering of the samples, using either the miRNA or mRNA profile, showed the clear separation of WT from normal kidney samples, but the miRNA pattern yielded better separation of WT subtypes. A correlation analysis of the deregulated miRNA and mRNAs identified 13,026 miRNA/mRNA pairs with inversely correlated expression, of which 2844 are potential interactions of miRNA and their predicted mRNA targets. We found significant upregulation of miRNAs-183, -301a/b and -335 for the blastemal subtype, and miRNAs-181b, -223 and -630 for the regressive subtype. We found marked deregulation of miRNAs regulating epithelial to mesenchymal transition, especially in the blastemal subtype, and miRNAs influencing chemosensitivity, especially in regressive subtypes. Further research is needed to assess the influence of preoperative chemotherapy and tumor infiltrating lymphocytes on the miRNA and mRNA patterns in WT.

microRNA expression profile in human coronary smooth muscle cell-derived microparticles is a source of biomarkers.

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de Gonzalo-Calvo, David; Cenarro, Ana; Civeira, Fernando; Llorente-Cortes, Vicenta

2016-01-01

microRNA (miRNA) expression profile of extracellular vesicles is a potential tool for clinical practice. Despite the key role of vascular smooth muscle cells (VSMC) in cardiovascular pathology, there is limited information about the presence of miRNAs in microparticles secreted by this cell type, including human coronary artery smooth muscle cells (HCASMC). Here, we tested whether HCASMC-derived microparticles contain miRNAs and the value of these miRNAs as biomarkers. HCASMC and explants from atherosclerotic or non-atherosclerotic areas were obtained from coronary arteries of patients undergoing heart transplant. Plasma samples were collected from: normocholesterolemic controls (N=12) and familial hypercholesterolemia (FH) patients (N=12). Both groups were strictly matched for age, sex and cardiovascular risk factors. Microparticle (0.1-1μm) isolation and characterization was performed using standard techniques. VSMC-enriched miRNAs expression (miR-21-5p, -143-3p, -145-5p, -221-3p and -222-3p) was analyzed using RT-qPCR. Total RNA isolated from HCASMC-derived microparticles contained small RNAs, including VSMC-enriched miRNAs. Exposition of HCASMC to pathophysiological conditions, such as hypercholesterolemia, induced a decrease in the expression level of miR-143-3p and miR-222-3p in microparticles, not in cells. Expression levels of miR-222-3p were lower in circulating microparticles from FH patients compared to normocholesterolemic controls. Microparticles derived from atherosclerotic plaque areas showed a decreased level of miR-143-3p and miR-222-3p compared to non-atherosclerotic areas. We demonstrated for the first time that microparticles secreted by HCASMC contain microRNAs. Hypercholesterolemia alters the microRNA profile of HCASMC-derived microparticles. The miRNA signature of HCASMC-derived microparticles is a source of cardiovascular biomarkers. Copyright © 2016 Sociedad Española de Arteriosclerosis. Publicado por Elsevier España, S.L.U. All rights

MicroRNA expression profiling during the life cycle of the silkworm (Bombyx mori

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Cheng Daojun

2009-09-01

Full Text Available Abstract Background MicroRNAs (miRNAs are expressed by a wide range of eukaryotic organisms, and function in diverse biological processes. Numerous miRNAs have been identified in Bombyx mori, but the temporal expression profiles of miRNAs corresponding to each stage transition over the entire life cycle of the silkworm remain to be established. To obtain a comprehensive overview of the correlation between miRNA expression and stage transitions, we performed a whole-life test and subsequent stage-by-stage examinations on nearly one hundred miRNAs in the silkworm. Results Our results show that miRNAs display a wide variety of expression profiles over the whole life of the silkworm, including continuous expression from embryo to adult (miR-184, up-regulation over the entire life cycle (let-7 and miR-100, down-regulation over the entire life cycle (miR-124, expression associated with embryogenesis (miR-29 and miR-92, up-regulation from early 3rd instar to pupa (miR-275, and complementary pulses in expression between miR-34b and miR-275. Stage-by-stage examinations revealed further expression patterns, such as emergence at specific time-points during embryogenesis and up-regulation of miRNA groups in late embryos (miR-1 and bantam, expression associated with stage transition between instar and molt larval stages (miR-34b, expression associated with silk gland growth and spinning activity (miR-274, continuous high expression from the spinning larval to pupal and adult stages (miR-252 and miR-31a, a coordinate expression trough in day 3 pupae of both sexes (miR-10b and miR-281, up-regulation in pupal metamorphosis of both sexes (miR-29b, and down-regulation in pupal metamorphosis of both sexes (miR-275. Conclusion We present the full-scale expression profiles of miRNAs throughout the life cycle of Bombyx mori. The whole-life expression profile was further investigated via stage-by-stage analysis. Our data provide an important resource for more detailed

MicroRNA expression profiling during the life cycle of the silkworm (Bombyx mori).

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Liu, Shiping; Zhang, Liang; Li, Qibin; Zhao, Ping; Duan, Jun; Cheng, Daojun; Xiang, Zhonghuai; Xia, Qingyou

2009-09-28

MicroRNAs (miRNAs) are expressed by a wide range of eukaryotic organisms, and function in diverse biological processes. Numerous miRNAs have been identified in Bombyx mori, but the temporal expression profiles of miRNAs corresponding to each stage transition over the entire life cycle of the silkworm remain to be established. To obtain a comprehensive overview of the correlation between miRNA expression and stage transitions, we performed a whole-life test and subsequent stage-by-stage examinations on nearly one hundred miRNAs in the silkworm. Our results show that miRNAs display a wide variety of expression profiles over the whole life of the silkworm, including continuous expression from embryo to adult (miR-184), up-regulation over the entire life cycle (let-7 and miR-100), down-regulation over the entire life cycle (miR-124), expression associated with embryogenesis (miR-29 and miR-92), up-regulation from early 3rd instar to pupa (miR-275), and complementary pulses in expression between miR-34b and miR-275. Stage-by-stage examinations revealed further expression patterns, such as emergence at specific time-points during embryogenesis and up-regulation of miRNA groups in late embryos (miR-1 and bantam), expression associated with stage transition between instar and molt larval stages (miR-34b), expression associated with silk gland growth and spinning activity (miR-274), continuous high expression from the spinning larval to pupal and adult stages (miR-252 and miR-31a), a coordinate expression trough in day 3 pupae of both sexes (miR-10b and miR-281), up-regulation in pupal metamorphosis of both sexes (miR-29b), and down-regulation in pupal metamorphosis of both sexes (miR-275). We present the full-scale expression profiles of miRNAs throughout the life cycle of Bombyx mori. The whole-life expression profile was further investigated via stage-by-stage analysis. Our data provide an important resource for more detailed functional analysis of miRNAs in this animal.

MicroRNA Expression Profiles in Cultured Human Fibroblasts in Space

Science.gov (United States)

Wu, Honglu; Lu, Tao; Jeevarajan, John; Rohde, Larry; Zhang, Ye

2014-01-01

Microgravity, or an altered gravity environment from the static 1g, has been shown to influence global gene expression patterns and protein levels in living organisms. However, it is unclear how these changes in gene and protein expressions are related to each other or are related to other factors regulating such changes. A different class of RNA, the small non-coding microRNA (miRNA), can have a broad effect on gene expression networks by mainly inhibiting the translation process. Previously, we investigated changes in the expression of miRNA and related genes under simulated microgravity conditions on the ground using the NASA invented bioreactor. In comparison to static 1 g, simulated microgravity altered a number of miRNAs in human lymphoblastoid cells. Pathway analysis with the altered miRNAs and RNA expressions revealed differential involvement of cell communication and catalytic activity, as well as immune response signaling and NGF activation of NF-kB pathways under simulated microgravity condition. The network analysis also identified several projected networks with c- Rel, ETS1 and Ubiquitin C as key factors. In a flight experiment on the International Space Station (ISS), we will investigate the effects of actual spaceflight on miRNA expressions in nondividing human fibroblast cells in mostly G1 phase of the cell cycle. A fibroblast is a type of cell that synthesizes the extracellular matrix and collagen, the structural framework for tissues, and plays a critical role in wound healing and other functions. In addition to miRNA expressions, we will investigate the effects of spaceflight on the cellular response to DNA damages from bleomycin treatment.

Expression of coding (mRNA) and non-coding (microRNA) RNA in lung tissue and blood isolated from pigs suffering from bacterial pleuropneumonia

DEFF Research Database (Denmark)

Skovgaard, Kerstin; Schou, Kirstine Klitgaard; Wendt, Karin Tarp

2010-01-01

MicroRNAs are small non-coding RNA molecules (18-23 nt), that regulate the activity of other genes at the post-transcriptional level. Recently it has become evident that microRNA plays an important role in modulating and fine tuning innate and adaptive immune responses. Still, little is known about...... the impact of microRNAs in the development and pathogenesis of lung infections. Expression of microRNA known to be induced by bacterial (i.e., LPS) ligands and thus supposed to play a role in the regulation of antimicrobial defence, were studied in lung tissue and in blood from pigs experimentally infected...... with Actinobacillus pleuropneumoniae (AP). Expression differences of mRNA and microRNA were quantified at different time points (6h, 12h, 24h, 48h PI) using reverse transcription quantitative real-time PCR (Rotor-Gene and Fluidigm). Expression profiles of miRNA in blood of seven animals were further studied using mi...

Profiling microRNA expression during multi-staged date palm (Phoenix dactylifera L.) fruit development

KAUST Repository

Xin, Chengqi; Liu, Wanfei; Lin, Qiang; Zhang, Xiaowei; Cui, Peng; Li, Fusen; Zhang, Guangyu; Pan, Linlin; Al-Amer, Ali; Mei, Hailiang; Al-Mssallem, Ibrahim S.; Hu, Songnian; Al-Johi, Hasan Awad; Yu, Jun

2015-01-01

MicroRNAs (miRNAs) play crucial roles in multiple stages of plant development and regulate gene expression at posttranscriptional and translational levels. In this study, we first identified 238 conserved miRNAs in date palm (Phoenix dactylifera) based on a high-quality genome assembly and defined 78 fruit-development-associated (FDA) miRNAs, whose expression profiles are variable at different fruit development stages. Using experimental data, we subsequently detected 276 novel P. dactylifera-specific FDA miRNAs and predicted their targets. We also revealed that FDA miRNAs function mainly in regulating genes involved in starch/sucrose metabolisms and other carbon metabolic pathways; among them, 221 FDA miRNAs exhibit negative correlation with their corresponding targets, which suggests their direct regulatory roles on mRNA targets. Our data define a comprehensive set of conserved and novel FDA miRNAs along with their expression profiles, which provide a basis for further experimentation in assigning discrete functions of these miRNAs in P. dactylifera fruit development.

Profiling microRNA expression during multi-staged date palm (Phoenix dactylifera L.) fruit development

KAUST Repository

Xin, Chengqi

2015-01-29

MicroRNAs (miRNAs) play crucial roles in multiple stages of plant development and regulate gene expression at posttranscriptional and translational levels. In this study, we first identified 238 conserved miRNAs in date palm (Phoenix dactylifera) based on a high-quality genome assembly and defined 78 fruit-development-associated (FDA) miRNAs, whose expression profiles are variable at different fruit development stages. Using experimental data, we subsequently detected 276 novel P. dactylifera-specific FDA miRNAs and predicted their targets. We also revealed that FDA miRNAs function mainly in regulating genes involved in starch/sucrose metabolisms and other carbon metabolic pathways; among them, 221 FDA miRNAs exhibit negative correlation with their corresponding targets, which suggests their direct regulatory roles on mRNA targets. Our data define a comprehensive set of conserved and novel FDA miRNAs along with their expression profiles, which provide a basis for further experimentation in assigning discrete functions of these miRNAs in P. dactylifera fruit development.

Expression of MicroRNA-146a and MicroRNA-155 in Placental Villi in Early- and Late-Onset Preeclampsia.

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Nizyaeva, N V; Kulikova, G V; Nagovitsyna, M N; Kan, N E; Prozorovskaya, K N; Shchegolev, A I; Sukhikh, G T

2017-07-01

We studied the expression of microRNA-146a and microRNA-155 in placental villi from 18 women (26-39 weeks of gestation) of reproductive age with early- or late-onset preeclampsia. The reference group consisted of women with physiological pregnancy and full-term gestation and with preterm birth after caesarian section on gestation week 26-31. MicroRNA-146a and microRNA-155 were detected by in situ hybridization with digoxigenin on paraffin sections. It was found that the expression of microRNA-146a in both syncytiotrophoblast of the intermediate villi and syncytial knots was lower at late-onset preeclampsia than at physiologic pregnancy of full-term period (p=0.037 and p=0.001 respectively). The expression of microRNA-155 in syncytiotrophoblast of intermediate placental villi in early-onset preeclampsia was higher than in group with preterm delivery (p=0.003). However, in syncytiotrophoblast of intermediate villi and in syncytial knots, the expression of microRNA-155 was lower at late-onset preeclampsia in comparison with full-term physiological pregnancy (p=0.005). In addition, the expression of microRNA-146a and microRNA-155 did not increase in the later terms in preeclampsia, while in the reference groups demonstrating gradual increase in the expression of these markers with increasing gestational age. Expression microRNA-146a and microRNA-155 little differed in early- and late-onset preeclampsia. These findings suggest that different variants of preeclampsia are probably characterized by common pathogenetic pathways. Damaged trophoblast cannot maintain of microRNAs synthesis at the required level, which determines the formation of a vicious circle in preeclampsia and further progression of the disease.

mESAdb: microRNA expression and sequence analysis database.

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Kaya, Koray D; Karakülah, Gökhan; Yakicier, Cengiz M; Acar, Aybar C; Konu, Ozlen

2011-01-01

microRNA expression and sequence analysis database (http://konulab.fen.bilkent.edu.tr/mirna/) (mESAdb) is a regularly updated database for the multivariate analysis of sequences and expression of microRNAs from multiple taxa. mESAdb is modular and has a user interface implemented in PHP and JavaScript and coupled with statistical analysis and visualization packages written for the R language. The database primarily comprises mature microRNA sequences and their target data, along with selected human, mouse and zebrafish expression data sets. mESAdb analysis modules allow (i) mining of microRNA expression data sets for subsets of microRNAs selected manually or by motif; (ii) pair-wise multivariate analysis of expression data sets within and between taxa; and (iii) association of microRNA subsets with annotation databases, HUGE Navigator, KEGG and GO. The use of existing and customized R packages facilitates future addition of data sets and analysis tools. Furthermore, the ability to upload and analyze user-specified data sets makes mESAdb an interactive and expandable analysis tool for microRNA sequence and expression data.

In Vitro Assays for Mouse Müller Cell Phenotyping Through microRNA Profiling in the Damaged Retina.

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Reyes-Aguirre, Luis I; Quintero, Heberto; Estrada-Leyva, Brenda; Lamas, Mónica

2018-01-01

microRNA profiling has identified cell-specific expression patterns that could represent molecular signatures triggering the acquisition of a specific phenotype; in other words, of cellular identity and its associated function. Several groups have hypothesized that retinal cell phenotyping could be achieved through the determination of the global pattern of miRNA expression across specific cell types in the adult retina. This is especially relevant for Müller glia in the context of retinal damage, as these cells undergo dramatic changes of gene expression in response to injury, that render them susceptible to acquire a progenitor-like phenotype and be a source of new neurons.We describe a method that combines an experimental protocol for excitotoxic-induced retinal damage through N-methyl-D-aspartate subretinal injection with magnetic-activated cell sorting (MACS) of Müller cells and RNA isolation for microRNA profiling. Comparison of microRNA patterns of expression should allow Müller cell phenotyping under different experimental conditions.

MicroRNA and gene signature of severe cutaneous drug ...

African Journals Online (AJOL)

Purpose: To build a microRNA and gene signature of severe cutaneous adverse drug reactions (SCAR), including Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). Methods: MicroRNA expression profiles were downloaded from miRNA expression profile of patients' skin suffering from TEN using an ...

Circulating, Cell-Free Micro-RNA Profiles Reflect Discordant Development of Dementia in Monozygotic Twins

DEFF Research Database (Denmark)

Mengel-From, Jonas; Rønne, Mette E; Carlsen, Anting L

2018-01-01

We aim to examine if circulating micro-RNA and cytokine levels associate with dementia diagnosis and cognitive scores. To test our hypothesis, we use plasma donated from 48 monozygotic twin pairs in 1997 and 46 micro-RNAs and 10 cytokines were quantified using microfluidic RT-qPCR and multiplex...... solid-phase immunoassays, respectively. Micro-RNA and cytokine profiling were examined for associations with dementia diagnoses in a longitudinal registry study or with cognitive scores at baseline. Thirty-six micro-RNAs and all cytokines were detected consistently. Micro-RNA profiles associate...... with diagnoses and cognitive scores at statistically significant levels while cytokine only showed trends pointing at chronic inflammation in twins having or developing dementia. The most notable findings were decreased miR-106a and miR-210, and increased miR-106b expression in twins with a dementia diagnosis...

Integrated microRNA and gene expression profiling reveals the crucial miRNAs in curcumin anti-lung cancer cell invasion.

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Zhan, Jian-Wei; Jiao, De-Min; Wang, Yi; Song, Jia; Wu, Jin-Hong; Wu, Li-Jun; Chen, Qing-Yong; Ma, Sheng-Lin

2017-09-01

Curcumin (diferuloylmethane) has chemopreventive and therapeutic properties against many types of tumors, both in vitro and in vivo. Previous reports have shown that curcumin exhibits anti-invasive activities, but the mechanisms remain largely unclear. In this study, both microRNA (miRNA) and messenger RNA (mRNA) expression profiles were used to characterize the anti-metastasis mechanisms of curcumin in human non-small cell lung cancer A549 cell line. Microarray analysis revealed that 36 miRNAs were differentially expressed between the curcumin-treated and control groups. miR-330-5p exhibited maximum upregulation, while miR-25-5p exhibited maximum downregulation in the curcumin treatment group. mRNA expression profiles and functional analysis indicated that 226 differentially expressed mRNAs belonged to different functional categories. Significant pathway analysis showed that mitogen-activated protein kinase, transforming growth factor-β, and Wnt signaling pathways were significantly downregulated. At the same time, axon guidance, glioma, and ErbB tyrosine kinase receptor signaling pathways were significantly upregulated. We constructed a miRNA gene network that contributed to the curcumin inhibition of metastasis in lung cancer cells. let-7a-3p, miR-1262, miR-499a-5p, miR-1276, miR-331-5p, and miR-330-5p were identified as key microRNA regulators in the network. Finally, using miR-330-5p as an example, we confirmed the role of miR-330-5p in mediating the anti-migration effect of curcumin, suggesting the importance of miRNAs in the regulation of curcumin biological activity. Our findings provide new insights into the anti-metastasis mechanism of curcumin in lung cancer. © 2017 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.

MicroRNA signature of the human developing pancreas

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Correa-Medina Mayrin

2010-09-01

Full Text Available Abstract Background MicroRNAs are non-coding RNAs that regulate gene expression including differentiation and development by either inhibiting translation or inducing target degradation. The aim of this study is to determine the microRNA expression signature during human pancreatic development and to identify potential microRNA gene targets calculating correlations between the signature microRNAs and their corresponding mRNA targets, predicted by bioinformatics, in genome-wide RNA microarray study. Results The microRNA signature of human fetal pancreatic samples 10-22 weeks of gestational age (wga, was obtained by PCR-based high throughput screening with Taqman Low Density Arrays. This method led to identification of 212 microRNAs. The microRNAs were classified in 3 groups: Group number I contains 4 microRNAs with the increasing profile; II, 35 microRNAs with decreasing profile and III with 173 microRNAs, which remain unchanged. We calculated Pearson correlations between the expression profile of microRNAs and target mRNAs, predicted by TargetScan 5.1 and miRBase altgorithms, using genome-wide mRNA expression data. Group I correlated with the decreasing expression of 142 target mRNAs and Group II with the increasing expression of 876 target mRNAs. Most microRNAs correlate with multiple targets, just as mRNAs are targeted by multiple microRNAs. Among the identified targets are the genes and transcription factors known to play an essential role in pancreatic development. Conclusions We have determined specific groups of microRNAs in human fetal pancreas that change the degree of their expression throughout the development. A negative correlative analysis suggests an intertwined network of microRNAs and mRNAs collaborating with each other. This study provides information leading to potential two-way level of combinatorial control regulating gene expression through microRNAs targeting multiple mRNAs and, conversely, target mRNAs regulated in

MicroRNA expression profiling identifies activated B cell status in chronic lymphocytic leukemia cells.

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Shuqiang Li

2011-03-01

Full Text Available Chronic lymphocytic leukemia (CLL is thought to be a disease of resting lymphocytes. However, recent data suggest that CLL cells may more closely resemble activated B cells. Using microRNA (miRNA expression profiling of highly-enriched CLL cells from 38 patients and 9 untransformed B cells from normal donors before acute CpG activation and 5 matched B cells after acute CpG activation, we demonstrate an activated B cell status for CLL. Gene set enrichment analysis (GSEA identified statistically-significant similarities in miRNA expression between activated B cells and CLL cells including upregulation of miR-34a, miR-155, and miR-342-3p and downregulation of miR-103, miR-181a and miR-181b. Additionally, decreased levels of two CLL signature miRNAs miR-29c and miR-223 are associated with ZAP70(+ and IgV(H unmutated status and with shorter time to first therapy. These data indicate an activated B cell status for CLL cells and suggest that the direction of change of individual miRNAs may predict clinical course in CLL.

Methods for small RNA preparation for digital gene expression profiling by next-generation sequencing

NARCIS (Netherlands)

Linsen, S.E.V.; Cuppen, E.

2012-01-01

Digital gene expression (DGE) profiling techniques are playing an eminent role in the detection, localization, and differential expression quantification of many small RNA species, including microRNAs (1-3). Procedures in small RNA library preparation techniques typically include adapter ligation by

Altered retinal microRNA expression profiles in early diabetic retinopathy: an in silico analysis.

Science.gov (United States)

Xiong, Fen; Du, Xinhua; Hu, Jianyan; Li, Tingting; Du, Shanshan; Wu, Qiang

2014-07-01

MicroRNAs (miRNAs) - as negative regulators of target genes - are associated with various human diseases, but their precise role(s) in diabetic retinopathy (DR) remains to be elucidated. The aim of this study was to elucidate the involvement of miRNAs in early DR using in silico analysis to explore their gene expression patterns. We used the streptozotocin (STZ)-induced diabetic rat to investigate the roles of miRNAs in early DR. Retinal miRNA expression profiles from diabetic versus healthy control rats were examined by miRNA array analysis. Based on several bioinformatic systems, specifically, gene ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, we identified signatures of the potential pathological processes, gene functions, and signaling pathways that are influenced by dysregulated miRNAs. We used quantitative real-time polymerase chain reaction (qRT-PCR) to validate six (i.e. those with significant changes in expression levels) of the 17 miRNAs that were detected in the miRNA array. We also describe the significant role of the miRNA-gene network, which is based on the interactions between miRNAs and target genes. GO analysis of the 17 miRNAs detected in the miRNA array analysis revealed the most prevalent miRNAs to be those related to biological processes, olfactory bulb development and axonogenesis. These miRNAs also exert significant influence on additional pathways, including the mitogen-activated protein and calcium signaling pathways. Six of the seventeen miRNAs were chosen for qRT-PCR validation. With the exception of a slight difference in miRNA-350, our results are in close agreement with the differential expressions detected by array analysis. This study, which describes miRNA expression during the early developmental phases of DR, revealed extensive miRNA interactions. Based on both their target genes and signaling pathways, we suggest that miRNAs perform critical regulatory functions during the early stages of DR

Comparison of the functional microRNA expression in immune cell subsets of neonates and adults

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Hong-Ren Yu

2016-12-01

Full Text Available Diversity of biological molecules in newborn and adult immune cells contributes to differences in cell function and atopic properties. Micro RNAs (miRNAs are reported involve in the regulation of immune system. Therefore, determining the miRNA expression profile of leukocyte sub-populations is important for understanding immune system regulation. In order to explore the unique microRNA profiling that contribute to altered immune in neonates, we comprehensively analyzed the functional miRNA signatures of eight leukocyte subsets (polymorphonuclear cells, monocytes, CD4+ T cells, CD8+ T cells, natural killer cells, B cells, plasmacytoid dendritic cells (pDCs, and myeloid dendritic cells (mDCs from both neonatal and adult umbilical cord and peripheral blood samples, respectively. We observed distinct miRNA profiles between adult and neonatal blood leukocyte subsets, including unique miRNA signatures for each cell lineage. Leukocyte miRNA signatures were altered after stimulation. Adult peripheral leukocytes had higher let-7b-5p expression levels compared to neonatal cord leukocytes across multiple subsets, irrespective of stimulation. Transfecting neonatal monocytes with a let-7b-5p mimic resulted in a reduction of LPS-induced IL-6 and TNF-alpha production, while transfection of a let-7b-5p inhibitor into adult monocytes enhanced IL-6 and TNF-alpha production. With this functional approach, we provide intact differential microRNA expression profiling of specific immune cell subsets between neonates and adults. These studies serve as a basis to further understand the altered immune response observed in neonates and advance the development of therapeutic strategies.

microRNA expression during trophectoderm specification.

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Srinivas R Viswanathan

2009-07-01

Full Text Available Segregation of the trophectoderm from the inner cell mass of the embryo represents the first cell-fate decision of mammalian development. Transcription factors essential for specifying trophectoderm have been identified, but the role of microRNAs (miRNAs in modulating this fate-choice has been largely unexplored. We have compared miRNA expression in embryonic stem cell (ESC-derived trophectoderm and in staged murine embryos to identify a set of candidate miRNAs likely to be involved in trophectoderm specification.We profiled embryonic stem cells (ESCs as they were induced to differentiate into trophectodermal cells by ectopic expression of HRas/Q61L. We also profiled murine embryos at progressive stages of preimplantation development (zygote, 2-cell, 4-cell, 8-cell, morula, and blastocyst, which includes the time window in which the trophectoderm is specified in vivo Q61L/H.We describe miRNA expression changes that occur during trophectoderm specification and validate that our in vitro system faithfully recapitulates trophectoderm specification in vivo. By comparing our in vitro and in vivo datasets, we have identified a minimal set of candidate miRNAs likely to play a role in trophectoderm specification. These miRNAs are predicted to regulate a host of development-associated target genes, and many of these miRNAs have previously reported roles in development and differentiation. Additionally, we highlight a number of miRNAs whose tight developmental regulation may reflect a functional role in other stages of embryogenesis. Our embryo profiling data may be useful to investigators studying trophectoderm specification and other stages of preimplantation development.

Multiple sclerosis: microRNA expression profiles accurately differentiate patients with relapsing-remitting disease from healthy controls.

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Andreas Keller

Full Text Available Multiple sclerosis (MS is a chronic inflammatory demyelinating disease of the central nervous system, which is heterogenous with respect to clinical manifestations and response to therapy. Identification of biomarkers appears desirable for an improved diagnosis of MS as well as for monitoring of disease activity and treatment response. MicroRNAs (miRNAs are short non-coding RNAs, which have been shown to have the potential to serve as biomarkers for different human diseases, most notably cancer. Here, we analyzed the expression profiles of 866 human miRNAs. In detail, we investigated the miRNA expression in blood cells of 20 patients with relapsing-remitting MS (RRMS and 19 healthy controls using a human miRNA microarray and the Geniom Real Time Analyzer (GRTA platform. We identified 165 miRNAs that were significantly up- or downregulated in patients with RRMS as compared to healthy controls. The best single miRNA marker, hsa-miR-145, allowed discriminating MS from controls with a specificity of 89.5%, a sensitivity of 90.0%, and an accuracy of 89.7%. A set of 48 miRNAs that was evaluated by radial basis function kernel support vector machines and 10-fold cross validation yielded a specificity of 95%, a sensitivity of 97.6%, and an accuracy of 96.3%. While 43 of the 165 miRNAs deregulated in patients with MS have previously been related to other human diseases, the remaining 122 miRNAs are so far exclusively associated with MS. The implications of our study are twofold. The miRNA expression profiles in blood cells may serve as a biomarker for MS, and deregulation of miRNA expression may play a role in the pathogenesis of MS.

Systematic Expression Profiling Analysis Identifies Specific MicroRNA-Gene Interactions that May Differentiate between Active and Latent Tuberculosis Infection

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Lawrence Shih-Hsin Wu

2014-01-01

Full Text Available Tuberculosis (TB is the second most common cause of death from infectious diseases. About 90% of those infected are asymptomatic—the so-called latent TB infections (LTBI, with a 10% lifetime chance of progressing to active TB. To further understand the molecular pathogenesis of TB, several molecular studies have attempted to compare the expression profiles between healthy controls and active TB or LTBI patients. However, the results vary due to diverse genetic backgrounds and study designs and the inherent complexity of the disease process. Thus, developing a sensitive and efficient method for the detection of LTBI is both crucial and challenging. For the present study, we performed a systematic analysis of the gene and microRNA profiles of healthy individuals versus those affected with TB or LTBI. Combined with a series of in silico analysis utilizing publicly available microRNA knowledge bases and published literature data, we have uncovered several microRNA-gene interactions that specifically target both the blood and lungs. Some of these molecular interactions are novel and may serve as potential biomarkers of TB and LTBI, facilitating the development for a more sensitive, efficient, and cost-effective diagnostic assay for TB and LTBI for the Taiwanese population.

Systematic expression profiling analysis identifies specific microRNA-gene interactions that may differentiate between active and latent tuberculosis infection.

Science.gov (United States)

Wu, Lawrence Shih-Hsin; Lee, Shih-Wei; Huang, Kai-Yao; Lee, Tzong-Yi; Hsu, Paul Wei-Che; Weng, Julia Tzu-Ya

2014-01-01

Tuberculosis (TB) is the second most common cause of death from infectious diseases. About 90% of those infected are asymptomatic--the so-called latent TB infections (LTBI), with a 10% lifetime chance of progressing to active TB. To further understand the molecular pathogenesis of TB, several molecular studies have attempted to compare the expression profiles between healthy controls and active TB or LTBI patients. However, the results vary due to diverse genetic backgrounds and study designs and the inherent complexity of the disease process. Thus, developing a sensitive and efficient method for the detection of LTBI is both crucial and challenging. For the present study, we performed a systematic analysis of the gene and microRNA profiles of healthy individuals versus those affected with TB or LTBI. Combined with a series of in silico analysis utilizing publicly available microRNA knowledge bases and published literature data, we have uncovered several microRNA-gene interactions that specifically target both the blood and lungs. Some of these molecular interactions are novel and may serve as potential biomarkers of TB and LTBI, facilitating the development for a more sensitive, efficient, and cost-effective diagnostic assay for TB and LTBI for the Taiwanese population.

Differentially expressed microRNA in multiple sclerosis: A window into pathogenesis?

DEFF Research Database (Denmark)

Martin, Nellie Anne; Illés, Zsolt

2014-01-01

MicroRNA are small non-coding RNA that mediate mRNA translation repression or mRNA degradation, and thereby refine protein expression levels. More than 30–60% of all genes are regulated by microRNA. Exploring disease-related microRNA signatures is an emerging tool in biomarker discovery, and sile......MicroRNA are small non-coding RNA that mediate mRNA translation repression or mRNA degradation, and thereby refine protein expression levels. More than 30–60% of all genes are regulated by microRNA. Exploring disease-related microRNA signatures is an emerging tool in biomarker discovery......RNA related to multiple sclerosis has increased significantly in recent years. Differentially expressed microRNA have been identified in the whole blood, serum, plasma, cerebrospinal fluid, peripheral blood mononuclear cells, blood-derived cell subsets and brain lesions of patients with multiple sclerosis....... Most studies applied a non-candidate approach of screening by microarray and validation by quantitative polymerase chain reaction or next generation sequencing; others used a candidate-driven approach. Despite a relatively high number of multiple sclerosis-associated microRNA, just a few could...

MicroRNA Profiling Reveals Marker of Motor Neuron Disease in ALS Models.

Science.gov (United States)

Hoye, Mariah L; Koval, Erica D; Wegener, Amy J; Hyman, Theodore S; Yang, Chengran; O'Brien, David R; Miller, Rebecca L; Cole, Tracy; Schoch, Kathleen M; Shen, Tao; Kunikata, Tomonori; Richard, Jean-Philippe; Gutmann, David H; Maragakis, Nicholas J; Kordasiewicz, Holly B; Dougherty, Joseph D; Miller, Timothy M

2017-05-31

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder marked by the loss of motor neurons (MNs) in the brain and spinal cord, leading to fatally debilitating weakness. Because this disease predominantly affects MNs, we aimed to characterize the distinct expression profile of that cell type to elucidate underlying disease mechanisms and to identify novel targets that inform on MN health during ALS disease time course. microRNAs (miRNAs) are short, noncoding RNAs that can shape the expression profile of a cell and thus often exhibit cell-type-enriched expression. To determine MN-enriched miRNA expression, we used Cre recombinase-dependent miRNA tagging and affinity purification in mice. By defining the in vivo miRNA expression of MNs, all neurons, astrocytes, and microglia, we then focused on MN-enriched miRNAs via a comparative analysis and found that they may functionally distinguish MNs postnatally from other spinal neurons. Characterizing the levels of the MN-enriched miRNAs in CSF harvested from ALS models of MN disease demonstrated that one miRNA (miR-218) tracked with MN loss and was responsive to an ALS therapy in rodent models. Therefore, we have used cellular expression profiling tools to define the distinct miRNA expression of MNs, which is likely to enrich future studies of MN disease. This approach enabled the development of a novel, drug-responsive marker of MN disease in ALS rodents. SIGNIFICANCE STATEMENT Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease in which motor neurons (MNs) in the brain and spinal cord are selectively lost. To develop tools to aid in our understanding of the distinct expression profiles of MNs and, ultimately, to monitor MN disease progression, we identified small regulatory microRNAs (miRNAs) that were highly enriched or exclusive in MNs. The signal for one of these MN-enriched miRNAs is detectable in spinal tap biofluid from an ALS rat model, where its levels change as disease

microRNA expression profiles in human peripheral blood lymphocytes cultured in modeled microgravity

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National Aeronautics and Space Administration — In the present study we analyzed miRNA and mRNA expression profiles in human peripheral blood lymphocytes (PBLs) incubated in microgravity condition simulated by a...

Intrinsic noise of microRNA-regulated genes and the ceRNA hypothesis.

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Javad Noorbakhsh

Full Text Available MicroRNAs are small noncoding RNAs that regulate genes post-transciptionally by binding and degrading target eukaryotic mRNAs. We use a quantitative model to study gene regulation by inhibitory microRNAs and compare it to gene regulation by prokaryotic small non-coding RNAs (sRNAs. Our model uses a combination of analytic techniques as well as computational simulations to calculate the mean-expression and noise profiles of genes regulated by both microRNAs and sRNAs. We find that despite very different molecular machinery and modes of action (catalytic vs stoichiometric, the mean expression levels and noise profiles of microRNA-regulated genes are almost identical to genes regulated by prokaryotic sRNAs. This behavior is extremely robust and persists across a wide range of biologically relevant parameters. We extend our model to study crosstalk between multiple mRNAs that are regulated by a single microRNA and show that noise is a sensitive measure of microRNA-mediated interaction between mRNAs. We conclude by discussing possible experimental strategies for uncovering the microRNA-mRNA interactions and testing the competing endogenous RNA (ceRNA hypothesis.

Circulating MicroRNA Expression Levels Associated With Internet Gaming Disorder

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Minho Lee

2018-03-01

Full Text Available BackgroundAddictive use of the Internet and online games is a potential psychiatric disorder termed Internet gaming disorder (IGD. Altered microRNA (miRNA expression profiles have been reported in blood and brain tissue of patients with certain psychiatric disorders and suggested as biomarkers. However, there have been no reports on blood miRNA profiles in IGD.MethodsTo discover IGD-associated miRNAs, we analyzed the miRNA expression profiles of 51 samples (25 IGD and 26 controls using the TaqMan Low Density miRNA Array. For validation, we performed quantitative reverse transcription PCR with 36 independent samples (20 IGD and 16 controls.ResultsThrough discovery and independent validation, we identified three miRNAs (hsa-miR-200c-3p, hsa-miR-26b-5p, hsa-miR-652-3p that were significantly downregulated in the IGD group. Individuals with all three miRNA alterations had a much higher risk of IGD than those with no alteration [odds ratio (OR 22, 95% CI 2.29–211.11], and the ORs increased dose dependently with number of altered miRNAs. The predicted target genes of the three miRNAs were associated with neural pathways. We explored the protein expression of the three downstream target genes by western blot and confirmed that expression of GABRB2 and DPYSL2 was significantly higher in the IGD group.ConclusionWe observed that expressions of hsa-miR-200c-3p, hsa-miR-26b-5p, and hsa-miR-652-3p were downregulated in the IGD patients. Our results will be helpful to understand the pathophysiology of IGD.

Serum microRNA expression profile distinguishes enterovirus 71 and coxsackievirus 16 infections in patients with hand-foot-and-mouth disease.

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Lunbiao Cui

Full Text Available Altered circulating microRNA (miRNA profiles have been noted in patients with microbial infections. We compared host serum miRNA levels in patients with hand-foot-and-mouth disease (HFMD caused by enterovirus 71 (EV71 and coxsackievirus 16 (CVA16 as well as in other microbial infections and in healthy individuals. Among 664 different miRNAs analyzed using a miRNA array, 102 were up-regulated and 26 were down-regulated in sera of patients with enteroviral infections. Expression levels of ten candidate miRNAs were further evaluated by quantitative real-time PCR assays. A receiver operating characteristic (ROC curve analysis revealed that six miRNAs (miR-148a, miR-143, miR-324-3p, miR-628-3p, miR-140-5p, and miR-362-3p were able to discriminate patients with enterovirus infections from healthy controls with area under curve (AUC values ranged from 0.828 to 0.934. The combined six miRNA using multiple logistic regression analysis provided not only a sensitivity of 97.1% and a specificity of 92.7% but also a unique profile that differentiated enterovirial infections from other microbial infections. Expression levels of five miRNAs (miR-148a, miR-143, miR-324-3p, miR-545, and miR-140-5p were significantly increased in patients with CVA16 versus those with EV71 (p<0.05. Combination of miR-545, miR-324-3p, and miR-143 possessed a moderate ability to discrimination between CVA16 and EV71 with an AUC value of 0.761. These data indicate that sera from patients with different subtypes of enteroviral infection express unique miRNA profiles. Serum miRNA expression profiles may provide supplemental biomarkers for diagnosing and subtyping enteroviral HFMD infections.

MicroRNA expression in multiple myeloma is associated with genetic subtype, isotype and survival

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Pezzella Francesco

2011-05-01

Full Text Available Abstract Background MicroRNAs are small RNA species that regulate gene expression post-transcriptionally and are aberrantly expressed in many cancers including hematological malignancies. However, the role of microRNAs in the pathogenesis of multiple myeloma (MM is only poorly understood. We therefore used microarray analysis to elucidate the complete miRNome (miRBase version 13.0 of purified tumor (CD138+ cells from 33 patients with MM, 5 patients with monoclonal gammopathy of undetermined significance (MGUS and 9 controls. Results Unsupervised cluster analysis revealed that MM and MGUS samples have a distinct microRNA expression profile from control CD138+ cells. The majority of microRNAs aberrantly expressed in MM (109/129 were up-regulated. A comparison of these microRNAs with those aberrantly expressed in other B-cell and T-cell malignancies revealed a surprising degree of similarity (~40% suggesting the existence of a common lymphoma microRNA signature. We identified 39 microRNAs associated with the pre-malignant condition MGUS. Twenty-three (59% of these were also aberrantly expressed in MM suggesting common microRNA expression events in MM progression. MM is characterized by multiple chromosomal abnormalities of varying prognostic significance. We identified specific microRNA signatures associated with the most common IgH translocations (t(4;14 and t(11;14 and del(13q. Expression levels of these microRNAs were distinct between the genetic subtypes (by cluster analysis and correctly predicted these abnormalities in > 85% of cases using the support vector machine algorithm. Additionally, we identified microRNAs associated with light chain only myeloma, as well as IgG and IgA-type MM. Finally, we identified 32 microRNAs associated with event-free survival (EFS in MM, ten of which were significant by univariate (logrank survival analysis. Conclusions In summary, this work has identified aberrantly expressed microRNAs associated with the

miRNA and mRNA Expression Profiles Reveal Insight into Chitosan-Mediated Regulation of Plant Growth.

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Zhang, Xiaoqian; Li, Kecheng; Xing, Ronge; Liu, Song; Chen, Xiaolin; Yang, Haoyue; Li, Pengcheng

2018-04-18

Chitosan has been numerously studied as a plant growth regulator and stress tolerance inducer. To investigate the roles of chitosan as bioregulator on plant and unravel its possible metabolic responses mechanisms, we simultaneously investigated mRNAs and microRNAs (miRNAs) expression profiles of wheat seedlings in response to chitosan heptamer. We found 400 chitosan-responsive differentially expressed genes, including 268 up-regulated and 132 down-regulated mRNAs, many of which were related to photosynthesis, primary carbon and nitrogen metabolism, defense responses, and transcription factors. Moreover, miRNAs also participate in chitosan-mediated regulation on plant growth. We identified 87 known and 21 novel miRNAs, among which 56 miRNAs were induced or repressed by chitosan heptamer, such as miRNA156, miRNA159a, miRNA164, miRNA171a, miRNA319, and miRNA1127. The integrative analysis of miRNA and mRNA expression profiles in this case provides fundamental information for further investigation of regulation mechanisms of chitosan on plant growth and will facilitate its application in agriculture.

Aging-associated changes in microRNA expression profile of internal anal sphincter smooth muscle: Role of microRNA-133a

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Singh, Jagmohan; Boopathi, Ettickan; Addya, Sankar; Phillips, Benjamin; Rigoutsos, Isidore; Penn, Raymond B.

2016-01-01

A comprehensive genomic and proteomic, computational, and physiological approach was employed to examine the (previously unexplored) role of microRNAs (miRNAs) as regulators of internal anal sphincter (IAS) smooth muscle contractile phenotype and basal tone. miRNA profiling, genome-wide expression, validation, and network analyses were employed to assess changes in mRNA and miRNA expression in IAS smooth muscles from young vs. aging rats. Multiple miRNAs, including rno-miR-1, rno-miR-340-5p, rno-miR-185, rno-miR-199a-3p, rno-miR-200c, rno-miR-200b, rno-miR-31, rno-miR-133a, and rno-miR-206, were found to be upregulated in aging IAS. qPCR confirmed the upregulated expression of these miRNAs and downregulation of multiple, predicted targets (Eln, Col3a1, Col1a1, Zeb2, Myocd, Srf, Smad1, Smad2, Rhoa/Rock2, Fn1, Tagln v2, Klf4, and Acta2) involved in regulation of smooth muscle contractility. Subsequent studies demonstrated an aging-associated increase in the expression of miR-133a, corresponding decreases in RhoA, ROCK2, MYOCD, SRF, and SM22α protein expression, RhoA-signaling, and a decrease in basal and agonist [U-46619 (thromboxane A2 analog)]-induced increase in the IAS tone. Moreover, in vitro transfection of miR-133a caused a dose-dependent increase of IAS tone in strips, which was reversed by anti-miR-133a. Last, in vivo perianal injection of anti-miR-133a reversed the loss of IAS tone associated with age. This work establishes the important regulatory effect of miRNA-133a on basal and agonist-stimulated IAS tone. Moreover, reversal of age-associated loss of tone via anti-miR delivery strongly implicates miR dysregulation as a causal factor in the aging-associated decrease in IAS tone and suggests that miR-133a is a feasible therapeutic target in aging-associated rectoanal incontinence. PMID:27634012

Cigarette smoking decreases global microRNA expression in human alveolar macrophages.

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Joel W Graff

Full Text Available Human alveolar macrophages are critical components of the innate immune system. Cigarette smoking-induced changes in alveolar macrophage gene expression are linked to reduced resistance to pulmonary infections and to the development of emphysema/COPD. We hypothesized that microRNAs (miRNAs could control, in part, the unique messenger RNA (mRNA expression profiles found in alveolar macrophages of cigarette smokers. Activation of macrophages with different stimuli in vitro leads to a diverse range of M1 (inflammatory and M2 (anti-inflammatory polarized phenotypes that are thought to mimic activated macrophages in distinct tissue environments. Microarray mRNA data indicated that smoking promoted an "inverse" M1 mRNA expression program, defined by decreased expression of M1-induced transcripts and increased expression of M1-repressed transcripts with few changes in M2-regulated transcripts. RT-PCR arrays identified altered expression of many miRNAs in alveolar macrophages of smokers and a decrease in global miRNA abundance. Stratification of human subjects suggested that the magnitude of the global decrease in miRNA abundance was associated with smoking history. We found that many of the miRNAs with reduced expression in alveolar macrophages of smokers were predicted to target mRNAs upregulated in alveolar macrophages of smokers. For example, miR-452 is predicted to target the transcript encoding MMP12, an important effector of smoking-related diseases. Experimental antagonism of miR-452 in differentiated monocytic cells resulted in increased expression of MMP12. The comprehensive mRNA and miRNA expression profiles described here provide insight into gene expression regulation that may underlie the adverse effects cigarette smoking has on alveolar macrophages.

Integrating microRNA and mRNA expression profiles of neuronal progenitors to identify regulatory networks underlying the onset of cortical neurogenesis

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Barker Jeffery L

2009-08-01

Full Text Available Abstract Background Cortical development is a complex process that includes sequential generation of neuronal progenitors, which proliferate and migrate to form the stratified layers of the developing cortex. To identify the individual microRNAs (miRNAs and mRNAs that may regulate the genetic network guiding the earliest phase of cortical development, the expression profiles of rat neuronal progenitors obtained at embryonic day 11 (E11, E12 and E13 were analyzed. Results Neuronal progenitors were purified from telencephalic dissociates by a positive-selection strategy featuring surface labeling with tetanus-toxin and cholera-toxin followed by fluorescence-activated cell sorting. Microarray analyses revealed the fractions of miRNAs and mRNAs that were up-regulated or down-regulated in these neuronal progenitors at the beginning of cortical development. Nearly half of the dynamically expressed miRNAs were negatively correlated with the expression of their predicted target mRNAs. Conclusion These data support a regulatory role for miRNAs during the transition from neuronal progenitors into the earliest differentiating cortical neurons. In addition, by supplying a robust data set in which miRNA and mRNA profiles originate from the same purified cell type, this empirical study may facilitate the development of new algorithms to integrate various "-omics" data sets.

MicroRNA expression in benign breast tissue and risk of subsequent invasive breast cancer.

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Rohan, Thomas; Ye, Kenny; Wang, Yihong; Glass, Andrew G; Ginsberg, Mindy; Loudig, Olivier

2018-01-01

MicroRNAs are endogenous, small non-coding RNAs that control gene expression by directing their target mRNAs for degradation and/or posttranscriptional repression. Abnormal expression of microRNAs is thought to contribute to the development and progression of cancer. A history of benign breast disease (BBD) is associated with increased risk of subsequent breast cancer. However, no large-scale study has examined the association between microRNA expression in BBD tissue and risk of subsequent invasive breast cancer (IBC). We conducted discovery and validation case-control studies nested in a cohort of 15,395 women diagnosed with BBD in a large health plan between 1971 and 2006 and followed to mid-2015. Cases were women with BBD who developed subsequent IBC; controls were matched 1:1 to cases on age, age at diagnosis of BBD, and duration of plan membership. The discovery stage (316 case-control pairs) entailed use of the Illumina MicroRNA Expression Profiling Assay (in duplicate) to identify breast cancer-associated microRNAs. MicroRNAs identified at this stage were ranked by the strength of the correlation between Illumina array and quantitative PCR results for 15 case-control pairs. The top ranked 14 microRNAs entered the validation stage (165 case-control pairs) which was conducted using quantitative PCR (in triplicate). In both stages, linear regression was used to evaluate the association between the mean expression level of each microRNA (response variable) and case-control status (independent variable); paired t-tests were also used in the validation stage. None of the 14 validation stage microRNAs was associated with breast cancer risk. The results of this study suggest that microRNA expression in benign breast tissue does not influence the risk of subsequent IBC.

MicroRNA and mesial temporal lobe epilepsy with hippocampal sclerosis: Whole miRNome profiling of human hippocampus.

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Bencurova, Petra; Baloun, Jiri; Musilova, Katerina; Radova, Lenka; Tichy, Boris; Pail, Martin; Zeman, Martin; Brichtova, Eva; Hermanova, Marketa; Pospisilova, Sarka; Mraz, Marek; Brazdil, Milan

2017-10-01

Mesial temporal lobe epilepsy (mTLE) is a severe neurological disorder characterized by recurrent seizures. mTLE is frequently accompanied by neurodegeneration in the hippocampus resulting in hippocampal sclerosis (HS), the most common morphological correlate of drug resistance in mTLE patients. Incomplete knowledge of pathological changes in mTLE+HS complicates its therapy. The pathological mechanism underlying mTLE+HS may involve abnormal gene expression regulation, including posttranscriptional networks involving microRNAs (miRNAs). miRNA expression deregulation has been reported in various disorders, including epilepsy. However, the miRNA profile of mTLE+HS is not completely known and needs to be addressed. Here, we have focused on hippocampal miRNA profiling in 33 mTLE+HS patients and nine postmortem controls to reveal abnormally expressed miRNAs. In this study, we significantly reduced technology-related bias (the most common source of false positivity in miRNA profiling data) by combining two different miRNA profiling methods, namely next generation sequencing and miRNA-specific quantitative real-time polymerase chain reaction. These methods combined have identified and validated 20 miRNAs with altered expression in the human epileptic hippocampus; 19 miRNAs were up-regulated and one down-regulated in mTLE+HS patients. Nine of these miRNAs have not been previously associated with epilepsy, and 19 aberrantly expressed miRNAs potentially regulate the targets and pathways linked with epilepsy (such as potassium channels, γ-aminobutyric acid, neurotrophin signaling, and axon guidance). This study extends current knowledge of miRNA-mediated gene expression regulation in mTLE+HS by identifying miRNAs with altered expression in mTLE+HS, including nine novel abnormally expressed miRNAs and their putative targets. These observations further encourage the potential of microRNA-based biomarkers or therapies. Wiley Periodicals, Inc. © 2017 International League Against

Pulmonary microRNA profiling: implications in upper lobe predominant lung disease.

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Armstrong, David A; Nymon, Amanda B; Ringelberg, Carol S; Lesseur, Corina; Hazlett, Haley F; Howard, Louisa; Marsit, Carmen J; Ashare, Alix

2017-01-01

Numerous pulmonary diseases manifest with upper lobe predominance including cystic fibrosis, smoking-related chronic obstructive pulmonary disease, and tuberculosis. Zonal hypoxia, characteristic of these pulmonary maladies, and oxygen stress in general is known to exert profound effects on various important aspects of cell biology. Lung macrophages are major participants in the pulmonary innate immune response and regional differences in macrophage responsiveness to hypoxia may contribute in the development of lung disease. MicroRNAs are ubiquitous regulators of human biology and emerging evidence indicates altered microRNA expression modulates respiratory disease processes. The objective of this study is to gain insight into the epigenetic and cellular mechanisms influencing regional differences in lung disease by investigating effect of hypoxia on regional microRNA expression in the lung. All studies were performed using primary alveolar macrophages ( n  = 10) or bronchoalveolar lavage fluid ( n  = 16) isolated from human subjects. MicroRNA was assayed via the NanoString nCounter microRNA assay. Divergent molecular patterns of microRNA expression were observed in alternate lung lobes, specifically noted was disparate expression of miR-93 and miR-4454 in alveolar macrophages along with altered expression of miR-451a and miR-663a in bronchoalveolar lavage fluid. Gene ontology was used to identify potential downstream targets of divergent microRNAs. Targets include cytokines and matrix metalloproteinases, molecules that could have a significant impact on pulmonary inflammation and fibrosis. Our findings show variant regional microRNA expression associated with hypoxia in alveolar macrophages and BAL fluid in the lung-upper vs lower lobe. Future studies should address whether these specific microRNAs may act intracellularly, in a paracrine/endocrine manner to direct the innate immune response or may ultimately be involved in pulmonary host-to-pathogen trans

Expression profiling of microRNAs in human bone tissue from postmenopausal women.

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De-Ugarte, Laura; Serra-Vinardell, Jenny; Nonell, Lara; Balcells, Susana; Arnal, Magdalena; Nogues, Xavier; Mellibovsky, Leonardo; Grinberg, Daniel; Diez-Perez, Adolfo; Garcia-Giralt, Natalia

2018-01-01

Bone tissue is composed of several cell types, which express their own microRNAs (miRNAs) that will play a role in cell function. The set of total miRNAs expressed in all cell types configures the specific signature of the bone tissue in one physiological condition. The aim of this study was to explore the miRNA expression profile of bone tissue from postmenopausal women. Tissue was obtained from trabecular bone and was analyzed in fresh conditions (n = 6). Primary osteoblasts were also obtained from trabecular bone (n = 4) and human osteoclasts were obtained from monocyte precursors after in vitro differentiation (n = 5). MicroRNA expression profiling was obtained for each sample by microarray and a global miRNA analysis was performed combining the data acquired in all the microarray experiments. From the 641 miRNAs detected in bone tissue samples, 346 (54%) were present in osteoblasts and/or osteoclasts. The other 46% were not identified in any of the bone cells analyzed. Intersection of osteoblast and osteoclast arrays identified 101 miRNAs shared by both cell types, which accounts for 30-40% of miRNAs detected in these cells. In osteoblasts, 266 miRNAs were detected, of which 243 (91%) were also present in the total bone array, representing 38% of all bone miRNAs. In osteoclasts, 340 miRNAs were detected, of which 196 (58%) were also present in the bone tissue array, representing 31% of all miRNAs detected in total bone. These analyses provide an overview of miRNAs expressed in bone tissue, broadening our knowledge in the microRNA field.

Analysis of MicroRNA Expression Profiles in Weaned Pig Skeletal Muscle after Lipopolysaccharide Challenge

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Jing Zhang

2015-09-01

Full Text Available MicroRNAs (miRNAs constitute a class of non-coding RNAs that play a crucial regulatory role in skeletal muscle development and disease. Several acute inflammation conditions including sepsis and cancer are characterized by a loss of skeletal muscle due primarily to excessive muscle catabolism. As a well-known inducer of acute inflammation, a lipopolysaccharide (LPS challenge can cause serious skeletal muscle wasting. However, knowledge of the role of miRNAs in the course of inflammatory muscle catabolism is still very limited. In this study, RNA extracted from the skeletal muscle of pigs injected with LPS or saline was subjected to small RNA deep sequencing. We identified 304 conserved and 114 novel candidate miRNAs in the pig. Of these, four were significantly increased in the LPS-challenged samples and five were decreased. The expression of five miRNAs (ssc-miR-146a-5p, ssc-miR-221-5p, ssc-miR-148b-3p, ssc-miR-215 and ssc-miR-192 were selected for validation by quantitative polymerase chain reaction (qPCR, which found that ssc-miR-146a-5p and ssc-miR-221-5p were significantly upregulated in LPS-challenged pig skeletal muscle. Moreover, we treated mouse C2C12 myotubes with 1000 ng/mL LPS as an acute inflammation cell model. Expression of TNF-α, IL-6, muscle atrophy F-box (MAFbx and muscle RING finger 1 (MuRF1 mRNA was strongly induced by LPS. Importantly, miR-146a-5p and miR-221-5p also showed markedly increased expression in LPS-treated C2C12 myotubes, suggesting the two miRNAs may be involved in muscle catabolism systems in response to acute inflammation caused by a LPS challenge. To our knowledge, this study is the first to examine miRNA expression profiles in weaned pig skeletal muscle challenged with LPS, and furthers our understanding of miRNA function in the regulation of inflammatory muscle catabolism.

MicroRNA expression profiles in chronic epilepsy rats and neuroprotection from seizures by targeting miR-344a

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Liu XX

2017-07-01

Full Text Available Xixia Liu,1,2 Yuhan Liao,1 Xiuxiu Wang,1 Donghua Zou,1 Chun Luo,1 Chongdong Jian,1 Yuan Wu1 1Department of Neurology, First Affiliated Hospital of Guangxi Medical University, 2Department of Rehabilitation, People’s Hospital of Guangxi Zhuang Autonomous Region, Nanning, China Abstract: MicroRNA (miRNA is believed to play a crucial role in the cause and treatment of epilepsy by controlling gene expression. However, it is still unclear how miRNA profiles change after multiple prolonged seizures and aggravation of brain injury in chronic epilepsy (CE. To investigate the role of miRNA in epilepsy, we utilized the CE rat models with pentylenetetrazol (PTZ and miRNA profiles in the hippocampus. miRNA profiles were characterized using miRNA microarray analysis and were compared with the rats in the sham group, which received 0.9% physiological saline treatment at the same dose. Four up-regulated miRNAs (miR-139–3p, -770–5p, -127–5p, -331–3p and 5 down-regulated miRNAs (miR-802–5p, -380–5p, -183–5p, -547–5p, -344a/-344a–5p were found in the CE rats (fold change >1.5, P<0.05. Three of the dysregulated miRNAs were validated by quantitative real-time polymerase chain reaction, which revealed an outcome consistent with the initial results of the miRNA microarray analyses. Then, miR-344a agomir was intracerebroventricularly injected and followed by PTZ induction of CE models to investigate the effect of miR-344a in chronic neocortical epileptogenesis. After miRNA-344a agomir and scramble treatment, results showed a restoration of seizure behavior and a reduction in neuron damage in the cortex in miRNA-334a agomir treated rats. These data suggest that miRNA-344a might have a small modulatory effect on seizure-induced apoptosis signaling pathways in the cortex. Keywords: microRNA, chronic epilepsy, miR-344a, epigenetics, apoptosis

Antagonism pattern detection between microRNA and target expression in Ewing's sarcoma.

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Loredana Martignetti

Full Text Available MicroRNAs (miRNAs have emerged as fundamental regulators that silence gene expression at the post-transcriptional and translational levels. The identification of their targets is a major challenge to elucidate the regulated biological processes. The overall effect of miRNA is reflected on target mRNA expression, suggesting the design of new investigative methods based on high-throughput experimental data such as miRNA and transcriptome profiles. We propose a novel statistical measure of non-linear dependence between miRNA and mRNA expression, in order to infer miRNA-target interactions. This approach, which we name antagonism pattern detection, is based on the statistical recognition of a triangular-shaped pattern in miRNA-target expression profiles. This pattern is observed in miRNA-target expression measurements since their simultaneously elevated expression is statistically under-represented in the case of miRNA silencing effect. The proposed method enables miRNA target prediction to strongly rely on cellular context and physiological conditions reflected by expression data. The procedure has been assessed on synthetic datasets and tested on a set of real positive controls. Then it has been applied to analyze expression data from Ewing's sarcoma patients. The antagonism relationship is evaluated as a good indicator of real miRNA-target biological interaction. The predicted targets are consistently enriched for miRNA binding site motifs in their 3'UTR. Moreover, we reveal sets of predicted targets for each miRNA sharing important biological function. The procedure allows us to infer crucial miRNA regulators and their potential targets in Ewing's sarcoma disease. It can be considered as a valid statistical approach to discover new insights in the miRNA regulatory mechanisms.

Intra-tumor heterogeneity of microRNA-92a, microRNA-375 and microRNA-424 in colorectal cancer

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Jepsen, Rikke Karlin; Novotny, Guy Wayne; Klarskov, Louise Laurberg

2016-01-01

Various microRNAs (miRNAs) have been investigated in order to improve diagnostics and risk assessment in colorectal cancer (CRC). To clarify the potential of miRNA profiling in CRC, knowledge of intra-tumor heterogeneity in expression levels is crucial. The study aim was to estimate the intra...

Changes of microRNA profile and microRNA-mRNA regulatory network in bones of ovariectomized mice.

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An, Jee Hyun; Ohn, Jung Hun; Song, Jung Ah; Yang, Jae-Yeon; Park, Hyojung; Choi, Hyung Jin; Kim, Sang Wan; Kim, Seong Yeon; Park, Woog-Yang; Shin, Chan Soo

2014-03-01

Growing evidence shows the possibility of a role of microRNAs (miRNA) in regulating bone mass. We investigated the change of miRNAs and mRNA expression profiles in bone tissue in an ovariectomized mice model and evaluated the regulatory mechanism of bone mass mediated by miRNAs in an estrogen-deficiency state. Eight-week-old female C3H/HeJ mice underwent ovariectomy (OVX) or sham operation (Sham-op), and their femur and tibia were harvested to extract total bone RNAs after 4 weeks for microarray analysis. Eight miRNAs (miR-127, -133a, -133a*, -133b, -136, -206, -378, -378*) were identified to be upregulated after OVX, whereas one miRNA (miR-204) was downregulated. Concomitant analysis of mRNA microarray revealed that 658 genes were differentially expressed between OVX and Sham-op mice. Target prediction of differentially expressed miRNAs identified potential targets, and integrative analysis using the mRNA microarray results showed that PPARγ and CREB pathways are activated in skeletal tissues after ovariectomy. Among the potential candidates of miRNA, we further studied the role of miR-127 in vitro, which exhibited the greatest changes after OVX. We also studied the effects of miR-136, which has not been studied in the context of bone mass regulation. Transfection of miR-127 inhibitor has enhanced osteoblastic differentiation in UAMS-32 cells as measured by alkaline phosphatase activities and mRNA expression of osteoblast-specific genes, whereas miR-136 precursor has inhibited osteoblastic differentiation. Furthermore, transfection of both miR-127 and miR-136 inhibitors enhanced the osteocyte-like morphological changes and survival in MLO-Y4 cells, whereas precursors of miR-127 and -136 have aggravated dexamethasone-induced cell death. Both of the precursors enhanced osteoclastic differentiation in bone marrow macrophages, indicating that both miR-127 and -136 are negatively regulating bone mass. Taken together, these results suggest a novel insight into the

Large-scale expression analysis reveals distinct microRNA profiles at different stages of human neurodevelopment.

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Brandon Smith

Full Text Available BACKGROUND: MicroRNAs (miRNAs are short non-coding RNAs predicted to regulate one third of protein coding genes via mRNA targeting. In conjunction with key transcription factors, such as the repressor REST (RE1 silencing transcription factor, miRNAs play crucial roles in neurogenesis, which requires a highly orchestrated program of gene expression to ensure the appropriate development and function of diverse neural cell types. Whilst previous studies have highlighted select groups of miRNAs during neural development, there remains a need for amenable models in which miRNA expression and function can be analyzed over the duration of neurogenesis. PRINCIPAL FINDINGS: We performed large-scale expression profiling of miRNAs in human NTera2/D1 (NT2 cells during retinoic acid (RA-induced transition from progenitors to fully differentiated neural phenotypes. Our results revealed dynamic changes of miRNA patterns, resulting in distinct miRNA subsets that could be linked to specific neurodevelopmental stages. Moreover, the cell-type specific miRNA subsets were very similar in NT2-derived differentiated cells and human primary neurons and astrocytes. Further analysis identified miRNAs as putative regulators of REST, as well as candidate miRNAs targeted by REST. Finally, we confirmed the existence of two predicted miRNAs; pred-MIR191 and pred-MIR222 associated with SLAIN1 and FOXP2, respectively, and provided some evidence of their potential co-regulation. CONCLUSIONS: In the present study, we demonstrate that regulation of miRNAs occurs in precise patterns indicative of their roles in cell fate commitment, progenitor expansion and differentiation into neurons and glia. Furthermore, the similarity between our NT2 system and primary human cells suggests their roles in molecular pathways critical for human in vivo neurogenesis.

Integrating microRNA and mRNA expression profiling in Symbiodinium microadriaticum, a dinoflagellate symbiont of reef-building corals.

KAUST Repository

Baumgarten, Sebastian

2013-10-12

Animal and plant genomes produce numerous small RNAs (smRNAs) that regulate gene expression post-transcriptionally affecting metabolism, development, and epigenetic inheritance. In order to characterize the repertoire of endogenous smRNAs and potential gene targets in dinoflagellates, we conducted smRNA and mRNA expression profiling over 9 experimental treatments of cultures from Symbiodinium microadriaticum, a photosynthetic symbiont of scleractinian corals.

Integrating microRNA and mRNA expression profiling in Symbiodinium microadriaticum, a dinoflagellate symbiont of reef-building corals.

KAUST Repository

Baumgarten, Sebastian; Bayer, Till; Aranda, Manuel; Liew, Yi Jin; Carr, Adrian; Micklem, Gos; Voolstra, Christian R.

2013-01-01

Animal and plant genomes produce numerous small RNAs (smRNAs) that regulate gene expression post-transcriptionally affecting metabolism, development, and epigenetic inheritance. In order to characterize the repertoire of endogenous smRNAs and potential gene targets in dinoflagellates, we conducted smRNA and mRNA expression profiling over 9 experimental treatments of cultures from Symbiodinium microadriaticum, a photosynthetic symbiont of scleractinian corals.

Chronological changes in microRNA expression in the developing human brain.

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Michael P Moreau

Full Text Available MicroRNAs (miRNAs are endogenously expressed noncoding RNA molecules that are believed to regulate multiple neurobiological processes. Expression studies have revealed distinct temporal expression patterns in the developing rodent and porcine brain, but comprehensive profiling in the developing human brain has not been previously reported.We performed microarray and TaqMan-based expression analysis of all annotated mature miRNAs (miRBase 10.0 as well as 373 novel, predicted miRNAs. Expression levels were measured in 48 post-mortem brain tissue samples, representing gestational ages 14-24 weeks, as well as early postnatal and adult time points.Expression levels of 312 miRNAs changed significantly between at least two of the broad age categories, defined as fetal, young, and adult.We have constructed a miRNA expression atlas of the developing human brain, and we propose a classification scheme to guide future studies of neurobiological function.

In Situ Detection of MicroRNA Expression with RNAscope Probes.

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Yin, Viravuth P

2018-01-01

Elucidating the spatial resolution of gene transcripts provides important insight into potential gene function. MicroRNAs are short, singled-stranded noncoding RNAs that control gene expression through base-pair complementarity with target mRNAs in the 3' untranslated region (UTR) and inhibiting protein expression. However, given their small size of ~22- to 24-nt and low expression levels, standard in situ hybridization detection methods are not amendable for microRNA spatial resolution. Here, I describe a technique that employs RNAscope probe design and propriety amplification technology that provides simultaneous single molecule detection of individual microRNA and its target gene. This method allows for rapid and sensitive detection of noncoding RNA transcripts in frozen tissue sections.

MicroRNA expression profiling to identify and validate reference genes for relative quantification in colorectal cancer.

LENUS (Irish Health Repository)

Chang, Kah Hoong

2010-01-01

BACKGROUND: Advances in high-throughput technologies and bioinformatics have transformed gene expression profiling methodologies. The results of microarray experiments are often validated using reverse transcription quantitative PCR (RT-qPCR), which is the most sensitive and reproducible method to quantify gene expression. Appropriate normalisation of RT-qPCR data using stably expressed reference genes is critical to ensure accurate and reliable results. Mi(cro)RNA expression profiles have been shown to be more accurate in disease classification than mRNA expression profiles. However, few reports detailed a robust identification and validation strategy for suitable reference genes for normalisation in miRNA RT-qPCR studies. METHODS: We adopt and report a systematic approach to identify the most stable reference genes for miRNA expression studies by RT-qPCR in colorectal cancer (CRC). High-throughput miRNA profiling was performed on ten pairs of CRC and normal tissues. By using the mean expression value of all expressed miRNAs, we identified the most stable candidate reference genes for subsequent validation. As such the stability of a panel of miRNAs was examined on 35 tumour and 39 normal tissues. The effects of normalisers on the relative quantity of established oncogenic (miR-21 and miR-31) and tumour suppressor (miR-143 and miR-145) target miRNAs were assessed. RESULTS: In the array experiment, miR-26a, miR-345, miR-425 and miR-454 were identified as having expression profiles closest to the global mean. From a panel of six miRNAs (let-7a, miR-16, miR-26a, miR-345, miR-425 and miR-454) and two small nucleolar RNA genes (RNU48 and Z30), miR-16 and miR-345 were identified as the most stably expressed reference genes. The combined use of miR-16 and miR-345 to normalise expression data enabled detection of a significant dysregulation of all four target miRNAs between tumour and normal colorectal tissue. CONCLUSIONS: Our study demonstrates that the top six most

MicroRNA expression profiling to identify and validate reference genes for relative quantification in colorectal cancer

LENUS (Irish Health Repository)

Chang, Kah Hoong

2010-04-29

Abstract Background Advances in high-throughput technologies and bioinformatics have transformed gene expression profiling methodologies. The results of microarray experiments are often validated using reverse transcription quantitative PCR (RT-qPCR), which is the most sensitive and reproducible method to quantify gene expression. Appropriate normalisation of RT-qPCR data using stably expressed reference genes is critical to ensure accurate and reliable results. Mi(cro)RNA expression profiles have been shown to be more accurate in disease classification than mRNA expression profiles. However, few reports detailed a robust identification and validation strategy for suitable reference genes for normalisation in miRNA RT-qPCR studies. Methods We adopt and report a systematic approach to identify the most stable reference genes for miRNA expression studies by RT-qPCR in colorectal cancer (CRC). High-throughput miRNA profiling was performed on ten pairs of CRC and normal tissues. By using the mean expression value of all expressed miRNAs, we identified the most stable candidate reference genes for subsequent validation. As such the stability of a panel of miRNAs was examined on 35 tumour and 39 normal tissues. The effects of normalisers on the relative quantity of established oncogenic (miR-21 and miR-31) and tumour suppressor (miR-143 and miR-145) target miRNAs were assessed. Results In the array experiment, miR-26a, miR-345, miR-425 and miR-454 were identified as having expression profiles closest to the global mean. From a panel of six miRNAs (let-7a, miR-16, miR-26a, miR-345, miR-425 and miR-454) and two small nucleolar RNA genes (RNU48 and Z30), miR-16 and miR-345 were identified as the most stably expressed reference genes. The combined use of miR-16 and miR-345 to normalise expression data enabled detection of a significant dysregulation of all four target miRNAs between tumour and normal colorectal tissue. Conclusions Our study demonstrates that the top six most

Adenoid cystic carcinomas of the salivary gland, lacrimal gland, and breast are morphologically and genetically similar but have distinct microRNA expression profiles.

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Andreasen, Simon; Tan, Qihua; Agander, Tina Klitmøller; Steiner, Petr; Bjørndal, Kristine; Høgdall, Estrid; Larsen, Stine Rosenkilde; Erentaite, Daiva; Olsen, Caroline Holkmann; Ulhøi, Benedicte Parm; von Holstein, Sarah Linéa; Wessel, Irene; Heegaard, Steffen; Homøe, Preben

2018-02-21

Adenoid cystic carcinoma is among the most frequent malignancies in the salivary and lacrimal glands and has a grave prognosis characterized by frequent local recurrences, distant metastases, and tumor-related mortality. Conversely, adenoid cystic carcinoma of the breast is a rare type of triple-negative (estrogen and progesterone receptor, HER2) and basal-like carcinoma, which in contrast to other triple-negative and basal-like breast carcinomas has a very favorable prognosis. Irrespective of site, adenoid cystic carcinoma is characterized by gene fusions involving MYB, MYBL1, and NFIB, and the reason for the different clinical outcomes is unknown. In order to identify the molecular mechanisms underlying the discrepancy in clinical outcome, we characterized the phenotypic profiles, pattern of gene rearrangements, and global microRNA expression profiles of 64 salivary gland, 9 lacrimal gland, and 11 breast adenoid cystic carcinomas. All breast and lacrimal gland adenoid cystic carcinomas had triple-negative and basal-like phenotypes, while salivary gland tumors were indeterminate in 13% of cases. Aberrations in MYB and/or NFIB were found in the majority of cases in all three locations, whereas MYBL1 involvement was restricted to tumors in the salivary gland. Global microRNA expression profiling separated salivary and lacrimal gland adenoid cystic carcinoma from their respective normal glands but could not distinguish normal breast adenoid cystic carcinoma from normal breast tissue. Hierarchical clustering separated adenoid cystic carcinomas of salivary gland origin from those of the breast and placed lacrimal gland carcinomas in between these. Functional annotation of the microRNAs differentially expressed between salivary gland and breast adenoid cystic carcinoma showed these as regulating genes involved in metabolism, signal transduction, and genes involved in other cancers. In conclusion, microRNA dysregulation is the first class of molecules separating adenoid

miRNA Expression Profiles in Cerebrospinal Fluid and Blood of Patients with Acute Ischemic Stroke

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Sørensen, Sofie Sølvsten; Nygaard, Ann-Britt; Nielsen, Ming-Yuan

2014-01-01

in the cell-free fractions of CSF and blood were analyzed by a microarray technique (miRCURY LNA™ microRNA Array, Exiqon A/S, Denmark) using a quantitative PCR (qPCR) platform containing 378 miRNA primers. In total, 183 different miRNAs were detected in the CSF, of which two miRNAs (let-7c and miR-221-3p......The aims of the study were (1) to determine whether miRNAs (microRNAs) can be detected in the cerebrospinal fluid (CSF) and blood of patients with ischemic stroke and (2) to compare these miRNA profiles with corresponding profiles from other neurological patients to address whether the mi......RNA profiles of CSF or blood have potential usefulness as diagnostic biomarkers of ischemic stroke. CSF from patients with acute ischemic stroke (n = 10) and patients with other neurological diseases (n = 10) was collected by lumbar puncture. Blood samples were taken immediately after. Expression profiles...

MicroRNA Expression Profiling by Bead Array Technology in Human Tumor Cell Lines Treated with Interferon-Alpha-2a

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Siegrist Fredy

2009-01-01

Full Text Available Abstract MicroRNAs are positive and negative regulators of eukaryotic gene expression that modulate transcript abundance by specific binding to sequence motifs located prevalently in the 3' untranslated regions of target messenger RNAs (mRNA. Interferon-alpha-2a (IFNα induces a large set of protein coding genes mediating antiproliferative and antiviral responses. Here we use a global microarray-based microRNA detection platform to identify genes that are induced by IFNα in hepatoma- or melanoma-derived human tumor cell lines. Despite the enormous differences in expression levels between these models, we were able to identify microRNAs that are upregulated by IFNα in both lines suggesting the possibility that interferon-regulated microRNAs are involved in the transcriptional repression of mRNA relevant to cytokine responses.

MicroRNA expression profiles of multiple system atrophy from formalin-fixed paraffin-embedded samples.

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Wakabayashi, Koichi; Mori, Fumiaki; Kakita, Akiyoshi; Takahashi, Hitoshi; Tanaka, Shinya; Utsumi, Jun; Sasaki, Hidenao

2016-12-02

MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression. Recently, we have shown that informative miRNA data can be derived from archived formalin-fixed paraffin-embedded (FFPE) samples from postmortem cases of amyotrophic lateral sclerosis and normal controls. miRNA analysis has now been performed on FFPE samples from affected brain regions in patients with multiple system atrophy (MSA) and the same areas in neurologically normal controls. We evaluated 50 samples from patients with MSA (n=13) and controls (n=13). Twenty-six samples were selected for miRNA analysis on the basis of the criteria reported previously: (i) a formalin fixation time of less than 4 weeks, (ii) a total RNA yield per sample of more than 500ng, and (iii) sufficient quality of the RNA electrophoresis pattern. These included 11 cases of MSA and 5 controls. Thus, the success rate for analysis of RNA from FFPE samples was 52% (26 of 50). For MSA, a total of 395 and 383 miRNAs were identified in the pons and cerebellum, respectively; 5 were up-regulated and 33 were down-regulated in the pons and 5 were up-regulated and 18 were down-regulated in the cerebellum. Several miRNAs down-regulated in the pons (miR-129-2-3p and miR-129-5p) and cerebellum (miR-129-2-3p, miR-129-5p and miR-132-3p) had already been identified in frozen cerebellum from MSA patients. These findings suggest that archived FFPE postmortem samples can be a valuable source for miRNA profiling in MSA. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Differential MicroRNA Expression Profile in Myxomatous Mitral Valve Prolapse and Fibroelastic Deficiency Valves

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Yei-Tsung Chen

2016-05-01

Full Text Available Myxomatous mitral valve prolapse (MMVP and fibroelastic deficiency (FED are two common variants of degenerative mitral valve disease (DMVD, which is a leading cause of mitral regurgitation worldwide. While pathohistological studies have revealed differences in extracellular matrix content in MMVP and FED, the molecular mechanisms underlying these two disease entities remain to be elucidated. By using surgically removed valvular specimens from MMVP and FED patients that were categorized on the basis of echocardiographic, clinical and operative findings, a cluster of microRNAs that expressed differentially were identified. The expressions of has-miR-500, -3174, -17, -1193, -646, -1273e, -4298, -203, -505, and -939 showed significant differences between MMVP and FED after applying Bonferroni correction (p < 0.002174. The possible involvement of microRNAs in the pathogenesis of DMVD were further suggested by the presences of in silico predicted target sites on a number of genes reported to be involved in extracellular matrix homeostasis and marker genes for cellular composition of mitral valves, including decorin (DCN, aggrecan (ACAN, fibromodulin (FMOD, α actin 2 (ACTA2, extracellular matrix protein 2 (ECM2, desmin (DES, endothelial cell specific molecule 1 (ESM1, and platelet/ endothelial cell adhesion molecule 1 (PECAM1, as well as inverse correlations of selected microRNA and mRNA expression in MMVP and FED groups. Our results provide evidence that distinct molecular mechanisms underlie MMVP and FED. Moreover, the microRNAs identified may be targets for the future development of diagnostic biomarkers and therapeutics.

Differential expression of microRNA-675, microRNA-139-3p and microRNA-335 in benign and malignant adrenocortical tumours

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Helwig, J; Bertram, S; Sheu, S Y; Suttorp, A C; Seggewiß, J; Willscher, E; Walz, M K; Worm, K; Schmid, K W

2011-01-01

Background For the clinical management of adrenocortical neoplasms it is crucial to correctly distinguish between benign and malignant tumours. Even histomorphologically based scoring systems do not allow precise separation in single lesions, thus novel parameters are desired which offer a more accurate differentiation. The tremendous potential of microRNAs (miRNAs) as diagnostic biomarkers in surgical pathology has recently been shown in a broad variety of tumours. Methods In order to elucidate the diagnostic impact of miRNA expression in adrenocortical neoplasms, a cohort of 20 adrenocortical specimens including normal adrenal tissue (n=4), adrenocortical adenomas (ACAs) (n=9), adrenocortical carcinomas (ACCs) (n=4) and metastases (n=3) was analysed using TaqMan low density arrays to identify specific miRNA profiles in order to distinguish between benign and malignant adrenocortical lesions. Results were validated in a validation cohort (n=16). Results Concerning the differential diagnosis of ACAs and ACCs, 159 out of 667 miRNAs were up- and 89 were down-regulated in ACAs. Using real-time PCR analysis of three of the most significantly expressed single key miRNAs allowed separation of ACAs from ACCs. ACCs exhibited significantly lower levels of miR-139-3p (up to 8.49-fold, p<0.001), miR-675 (up to 23.25-fold, p<0.001) and miR-335 (up to 5.25-fold, p<0.001). A validation cohort of 16 specimen with known Weiss score showed up-regulation of miR-335 and miR-675 in the majority of cases with probable malignant course, although overlapping values exist. Conclusion miRNA profiling of miR-675 and miR-335 helps in discriminating ACCs from ACAs. miRNA analysis may indicate malignant behaviour in cases with indeterminate malignant potential. PMID:21471143

Analysis of serum microRNA expression in male workers with occupational noise-induced hearing loss

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Y.H. Li

2018-01-01

Full Text Available Occupational noise-induced hearing loss (ONIHL is a prevalent occupational disorder that impairs auditory function in workers exposed to prolonged noise. However, serum microRNA expression in ONIHL subjects has not yet been studied. We aimed to compare the serum microRNA expression profiles in male workers of ONIHL subjects and controls. MicroRNA microarray analysis revealed that four serum microRNAs were differentially expressed between controls (n=3 and ONIHL subjects (n=3. Among these microRNAs, three were upregulated (hsa-miR-3162-5p, hsa-miR-4484, hsa-miR-1229-5p and one was downregulated (hsa-miR-4652-3p in the ONIHL group (fold change >1.5 and Pbon value <0.05. Real time quantitative PCR was conducted for validation of the microRNA expression. Significantly increased serum levels of miR-1229-5p were found in ONIHL subjects compared to controls (n=10 for each group; P<0.05. A total of 659 (27.0% genes were predicted as the target genes of miR-1229-5p. These genes were involved in various pathways, such as mitogen-activated protein kinase (MAPK signaling pathway. Overexpression of miR-1229-5p dramatically inhibited the luciferase activity of 3′ UTR segment of MAPK1 (P<0.01. Compared to the negative control, HEK293T cells expressing miR-1229-5p mimics showed a significant decline in mRNA levels of MAPK1 (P<0.05. This preliminary study indicated that serum miR-1229-5p was significantly elevated in ONIHL subjects. Increased miR-1229-5p may participate in the pathogenesis of ONIHL through repressing MAPK1 signaling.

Analysis of serum microRNA expression in male workers with occupational noise-induced hearing loss.

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Li, Y H; Yang, Y; Yan, Y T; Xu, L W; Ma, H Y; Shao, Y X; Cao, C J; Wu, X; Qi, M J; Wu, Y Y; Chen, R; Hong, Y; Tan, X H; Yang, L

2018-01-11

Occupational noise-induced hearing loss (ONIHL) is a prevalent occupational disorder that impairs auditory function in workers exposed to prolonged noise. However, serum microRNA expression in ONIHL subjects has not yet been studied. We aimed to compare the serum microRNA expression profiles in male workers of ONIHL subjects and controls. MicroRNA microarray analysis revealed that four serum microRNAs were differentially expressed between controls (n=3) and ONIHL subjects (n=3). Among these microRNAs, three were upregulated (hsa-miR-3162-5p, hsa-miR-4484, hsa-miR-1229-5p) and one was downregulated (hsa-miR-4652-3p) in the ONIHL group (fold change >1.5 and Pbon value <0.05). Real time quantitative PCR was conducted for validation of the microRNA expression. Significantly increased serum levels of miR-1229-5p were found in ONIHL subjects compared to controls (n=10 for each group; P<0.05). A total of 659 (27.0%) genes were predicted as the target genes of miR-1229-5p. These genes were involved in various pathways, such as mitogen-activated protein kinase (MAPK) signaling pathway. Overexpression of miR-1229-5p dramatically inhibited the luciferase activity of 3' UTR segment of MAPK1 (P<0.01). Compared to the negative control, HEK293T cells expressing miR-1229-5p mimics showed a significant decline in mRNA levels of MAPK1 (P<0.05). This preliminary study indicated that serum miR-1229-5p was significantly elevated in ONIHL subjects. Increased miR-1229-5p may participate in the pathogenesis of ONIHL through repressing MAPK1 signaling.

Identification of several circulating microRNAs from a genome-wide circulating microRNA expression profile as potential biomarkers for impaired glucose metabolism in polycystic ovarian syndrome.

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Jiang, Linlin; Huang, Jia; Chen, Yaxiao; Yang, Yabo; Li, Ruiqi; Li, Yu; Chen, Xiaoli; Yang, Dongzi

2016-07-01

This study aimed to detect serum microRNAs (miRNAs) differentially expressed between polycystic ovary syndrome (PCOS) patients with impaired glucose metabolism (IGM), PCOS patients with normal glucose tolerance (NGT), and healthy controls. A TaqMan miRNA array explored serum miRNA profiles as a pilot study, then selected miRNAs were analyzed in a validation cohort consisting of 65 PCOS women with IGM, 65 PCOS women with NGT, and 45 healthy women The relative expression of miR-122, miR-193b, and miR-194 was up-regulated in PCOS patients compared with controls, whereas that of miR-199b-5p was down-regulated. Furthermore, miR-122, miR-193b, and miR-194 were increased in the PCOS-IGM group compared with the PCOS-NGT group. Multiple linear regression analyses revealed that miR-193b and body mass index contributed independently to explain 43.7 % (P ovarian follicle development pathways, including the insulin signaling pathway, the neurotrophin signaling pathway, the PI3K-AKT signaling pathway, and regulation of actin cytoskeleton. This study expands our knowledge of the serum miRNA expression profiles of PCOS patients with IGM and the predicted target signal pathways involved in disease pathophysiology.

MicroRNA-223 Expression Is Upregulated in Insulin Resistant Human Adipose Tissue

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Tung-Yueh Chuang

2015-01-01

Full Text Available MicroRNAs (miRNAs are short noncoding RNAs involved in posttranscriptional regulation of gene expression and influence many cellular functions including glucose and lipid metabolism. We previously reported that adipose tissue (AT from women with polycystic ovary syndrome (PCOS or controls with insulin resistance (IR revealed a differentially expressed microRNA (miRNA profile, including upregulated miR-93 in PCOS patients and in non-PCOS women with IR. Overexpressed miR-93 directly inhibited glucose transporter isoform 4 (GLUT4 expression, thereby influencing glucose metabolism. We have now studied the role of miR-223, which is also abnormally expressed in the AT of IR subjects. Our data indicates that miR-223 is significantly overexpressed in the AT of IR women, regardless of whether they had PCOS or not. miR-223 expression in AT was positively correlated with HOMA-IR. Unlike what is reported in cardiomyocytes, overexpression of miR-223 in human differentiated adipocytes was associated with a reduction in GLUT4 protein content and insulin-stimulated glucose uptake. In addition, our data suggests miR-223 regulates GLUT4 expression by direct binding to its 3′ untranslated region (3′UTR. In conclusion, in AT miR-223 is an IR-related miRNA that may serve as a potential therapeutic target for the treatment of IR-related disorders.

MicroRNA profile changes in human immunodeficiency virus type 1 (HIV-1 seropositive individuals

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Smith Stephen M

2008-12-01

Full Text Available Abstract MicroRNAs (miRNAs play diverse roles in regulating cellular and developmental functions. We have profiled the miRNA expression in peripheral blood mononuclear cells from 36 HIV-1 seropositive individuals and 12 normal controls. The HIV-1-positive individuals were categorized operationally into four classes based on their CD4+ T-cell counts and their viral loads. We report that specific miRNA signatures can be observed for each of the four classes.

Optimal consistency in microRNA expression analysis using reference-gene-based normalization.

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Wang, Xi; Gardiner, Erin J; Cairns, Murray J

2015-05-01

Normalization of high-throughput molecular expression profiles secures differential expression analysis between samples of different phenotypes or biological conditions, and facilitates comparison between experimental batches. While the same general principles apply to microRNA (miRNA) normalization, there is mounting evidence that global shifts in their expression patterns occur in specific circumstances, which pose a challenge for normalizing miRNA expression data. As an alternative to global normalization, which has the propensity to flatten large trends, normalization against constitutively expressed reference genes presents an advantage through their relative independence. Here we investigated the performance of reference-gene-based (RGB) normalization for differential miRNA expression analysis of microarray expression data, and compared the results with other normalization methods, including: quantile, variance stabilization, robust spline, simple scaling, rank invariant, and Loess regression. The comparative analyses were executed using miRNA expression in tissue samples derived from subjects with schizophrenia and non-psychiatric controls. We proposed a consistency criterion for evaluating methods by examining the overlapping of differentially expressed miRNAs detected using different partitions of the whole data. Based on this criterion, we found that RGB normalization generally outperformed global normalization methods. Thus we recommend the application of RGB normalization for miRNA expression data sets, and believe that this will yield a more consistent and useful readout of differentially expressed miRNAs, particularly in biological conditions characterized by large shifts in miRNA expression.

Reproducible pattern of microRNA in normal human skin

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Holst, Line; Kaczkowski, Bogumil; Gniadecki, Robert

2010-01-01

RNA expression pattern in normal human skin. Here we investigated miRNA expression profiles from skin biopsies of 8 healthy volunteers taken from sun protected and mildly photo damaged skin using the modified protocol for miRNA extraction. We were able to show a constant pattern of miRNA expression between......MicroRNAs (miRNAs) regulate cell growth, differentiation and apoptosis via specific targeting of messenger RNA (mRNA). Aberrant mRNA expression contributes to pathological processes such as carcinogenesis. To take advantage of miRNA profiling in skin disease it is essential to investigate mi...... different individuals. We did not find any significant differences in miRNA expression between sun protected and mildly photodamaged skin. These results may be valuable for future design of studies on miRNA expression in skin disease....

Reproducible pattern of microRNA in normal human skin

DEFF Research Database (Denmark)

Holst, Line; Kaczkowski, Bogumil; Gniadecki, Robert

2010-01-01

RNA expression pattern in normal human skin. Here we investigated miRNA expression profiles from skin biopsies of 8 healthy volunteers taken from sun protected and mildly photo damaged skin using the modified protocol for miRNA extraction. We were able to show a constant pattern of miRNA expression between...... different individuals. We did not find any significant differences in miRNA expression between sun protected and mildly photodamaged skin. These results may be valuable for future design of studies on miRNA expression in skin disease.......MicroRNAs (miRNAs) regulate cell growth, differentiation and apoptosis via specific targeting of messenger RNA (mRNA). Aberrant mRNA expression contributes to pathological processes such as carcinogenesis. To take advantage of miRNA profiling in skin disease it is essential to investigate mi...

Long-term Neuroglial Cocultures as a Brain Aging Model: Hallmarks of Senescence, MicroRNA Expression Profiles, and Comparison With In Vivo Models.

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Bigagli, Elisabetta; Luceri, Cristina; Scartabelli, Tania; Dolara, Piero; Casamenti, Fiorella; Pellegrini-Giampietro, Domenico E; Giovannelli, Lisa

2016-01-01

Our purpose was to evaluate long-term neuroglial cocultures as a model for investigating senescence in the nervous system and to assess its similarities with in vivo models. To this aim, we maintained the cultures from 15 days in vitro (mature cultures) up to 27 days in vitro (senescent cultures), measuring senescence-associated, neuronal, dendritic, and astrocytic markers. Whole microRNA expression profiles were compared with those measured in the cortex of 18- and 24-month-old C57Bl/6J aged mice and of transgenic TgCRND8 mice, a model of amyloid-β deposition. Neuroglial cocultures displayed features of cellular senescence (increased senescence-associated-β-galactosidase activity, oxidative stress, γ-H2AX expression, IL-6 production, astrogliosis) that were concentration dependently counteracted by the antiaging compound resveratrol (1-5 µM). Among the 1,080 microRNAs analyzed, 335 were downregulated or absent in 27 compared with 15 days in vitro and resveratrol reversed this effect. A substantial overlapping was found between age-associated changes in microRNA expression profiles in vitro and in TgCRND8 mice but not in physiologically aged mice, indicating that this culture model displays more similarities with pathological than physiological brain aging. Our results demonstrate that neuroglial cocultures aged in vitro can be useful for investigating the cellular and molecular mechanisms of brain aging and for preliminary testing of protective compounds. © The Author 2015. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Performance comparison of digital microRNA profiling technologies applied on human breast cancer cell lines.

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Erik Knutsen

Full Text Available MicroRNA profiling represents an important first-step in deducting individual RNA-based regulatory function in a cell, tissue, or at a specific developmental stage. Currently there are several different platforms to choose from in order to make the initial miRNA profiles. In this study we investigate recently developed digital microRNA high-throughput technologies. Four different platforms were compared including next generation SOLiD ligation sequencing and Illumina HiSeq sequencing, hybridization-based NanoString nCounter, and miRCURY locked nucleic acid RT-qPCR. For all four technologies, full microRNA profiles were generated from human cell lines that represent noninvasive and invasive tumorigenic breast cancer. This study reports the correlation between platforms, as well as a more extensive analysis of the accuracy and sensitivity of data generated when using different platforms and important consideration when verifying results by the use of additional technologies. We found all the platforms to be highly capable for microRNA analysis. Furthermore, the two NGS platforms and RT-qPCR all have equally high sensitivity, and the fold change accuracy is independent of individual miRNA concentration for NGS and RT-qPCR. Based on these findings we propose new guidelines and considerations when performing microRNA profiling.

Analysis of miRNA and mRNA Expression Profiles Highlights Alterations in Ionizing Radiation Response of Human Lymphocytes under Modeled Microgravity

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Casara, Silvia; Sales, Gabriele; Lanfranchi, Gerolamo; Celotti, Lucia; Mognato, Maddalena

2012-01-01

Background Ionizing radiation (IR) can be extremely harmful for human cells since an improper DNA-damage response (DDR) to IR can contribute to carcinogenesis initiation. Perturbations in DDR pathway can originate from alteration in the functionality of the microRNA-mediated gene regulation, being microRNAs (miRNAs) small noncoding RNA that act as post-transcriptional regulators of gene expression. In this study we gained insight into the role of miRNAs in the regulation of DDR to IR under microgravity, a condition of weightlessness experienced by astronauts during space missions, which could have a synergistic action on cells, increasing the risk of radiation exposure. Methodology/Principal Findings We analyzed miRNA expression profile of human peripheral blood lymphocytes (PBL) incubated for 4 and 24 h in normal gravity (1 g) and in modeled microgravity (MMG) during the repair time after irradiation with 0.2 and 2Gy of γ-rays. Our results show that MMG alters miRNA expression signature of irradiated PBL by decreasing the number of radio-responsive miRNAs. Moreover, let-7i*, miR-7, miR-7-1*, miR-27a, miR-144, miR-200a, miR-598, miR-650 are deregulated by the combined action of radiation and MMG. Integrated analyses of miRNA and mRNA expression profiles, carried out on PBL of the same donors, identified significant miRNA-mRNA anti-correlations of DDR pathway. Gene Ontology analysis reports that the biological category of “Response to DNA damage” is enriched when PBL are incubated in 1 g but not in MMG. Moreover, some anti-correlated genes of p53-pathway show a different expression level between 1 g and MMG. Functional validation assays using luciferase reporter constructs confirmed miRNA-mRNA interactions derived from target prediction analyses. Conclusions/Significance On the whole, by integrating the transcriptome and microRNome, we provide evidence that modeled microgravity can affects the DNA-damage response to IR in human PBL. PMID:22347458

MicroRNA Expression Profile in Penile Cancer Revealed by Next-Generation Small RNA Sequencing.

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Li Zhang

Full Text Available Penile cancer (PeCa is a relatively rare tumor entity but possesses higher morbidity and mortality rates especially in developing countries. To date, the concrete pathogenic signaling pathways and core machineries involved in tumorigenesis and progression of PeCa remain to be elucidated. Several studies suggested miRNAs, which modulate gene expression at posttranscriptional level, were frequently mis-regulated and aberrantly expressed in human cancers. However, the miRNA profile in human PeCa has not been reported before. In this present study, the miRNA profile was obtained from 10 fresh penile cancerous tissues and matched adjacent non-cancerous tissues via next-generation sequencing. As a result, a total of 751 and 806 annotated miRNAs were identified in normal and cancerous penile tissues, respectively. Among which, 56 miRNAs with significantly different expression levels between paired tissues were identified. Subsequently, several annotated miRNAs were selected randomly and validated using quantitative real-time PCR. Compared with the previous publications regarding to the altered miRNAs expression in various cancers and especially genitourinary (prostate, bladder, kidney, testis cancers, the most majority of deregulated miRNAs showed the similar expression pattern in penile cancer. Moreover, the bioinformatics analyses suggested that the putative target genes of differentially expressed miRNAs between cancerous and matched normal penile tissues were tightly associated with cell junction, proliferation, growth as well as genomic instability and so on, by modulating Wnt, MAPK, p53, PI3K-Akt, Notch and TGF-β signaling pathways, which were all well-established to participate in cancer initiation and progression. Our work presents a global view of the differentially expressed miRNAs and potentially regulatory networks of their target genes for clarifying the pathogenic transformation of normal penis to PeCa, which research resource also

MicroRNA expression in melanocytic nevi: the usefulness of formalin-fixed, paraffin-embedded material for miRNA microarray profiling

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Glud, M.; Klausen, M.; Gniadecki, R.

2009-01-01

surgical specimens are formalin fixed and paraffin embedded (FFPE). To explore whether FFPE material would be suitable for miRNA profiling in melanocytic lesions, we compared miRNA expression patterns in FFPE versus fresh frozen samples, obtained from 15 human melanocytic nevi. Out of microarray data, we...

Effects of short-term exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin on microRNA expression in zebrafish embryos

International Nuclear Information System (INIS)

Jenny, Matthew J.; Aluru, Neelakanteswar; Hahn, Mark E.

2012-01-01

Although many drugs and environmental chemicals are teratogenic, the mechanisms by which most toxicants disrupt embryonic development are not well understood. MicroRNAs, single-stranded RNA molecules of ∼ 22 nt that regulate protein expression by inhibiting mRNA translation and promoting mRNA sequestration or degradation, are important regulators of a variety of cellular processes including embryonic development and cellular differentiation. Recent studies have demonstrated that exposure to xenobiotics can alter microRNA expression and contribute to the mechanisms by which environmental chemicals disrupt embryonic development. In this study we tested the hypothesis that developmental exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a well-known teratogen, alters microRNA expression during zebrafish development. We exposed zebrafish embryos to DMSO (0.1%) or TCDD (5 nM) for 1 h at 30 hours post fertilization (hpf) and measured microRNA expression using several methods at 36 and 60 hpf. TCDD caused strong induction of CYP1A at 36 hpf (62-fold) and 60 hpf (135-fold) as determined by real-time RT-PCR, verifying the effectiveness of the exposure. MicroRNA expression profiles were determined using microarrays (Agilent and Exiqon), next-generation sequencing (SOLiD), and real-time RT-PCR. The two microarray platforms yielded results that were similar but not identical; both showed significant changes in expression of miR-451, 23a, 23b, 24 and 27e at 60 hpf. Multiple analyses were performed on the SOLiD sequences yielding a total of 16 microRNAs as differentially expressed by TCDD in zebrafish embryos. However, miR-27e was the only microRNA to be identified as differentially expressed by all three methods (both microarrays, SOLiD sequencing, and real-time RT-PCR). These results suggest that TCDD exposure causes modest changes in expression of microRNAs, including some (miR-451, 23a, 23b, 24 and 27e) that are critical for hematopoiesis and cardiovascular

Statistical Use of Argonaute Expression and RISC Assembly in microRNA Target Identification

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Stanhope, Stephen A.; Sengupta, Srikumar; den Boon, Johan; Ahlquist, Paul; Newton, Michael A.

2009-01-01

MicroRNAs (miRNAs) posttranscriptionally regulate targeted messenger RNAs (mRNAs) by inducing cleavage or otherwise repressing their translation. We address the problem of detecting m/miRNA targeting relationships in homo sapiens from microarray data by developing statistical models that are motivated by the biological mechanisms used by miRNAs. The focus of our modeling is the construction, activity, and mediation of RNA-induced silencing complexes (RISCs) competent for targeted mRNA cleavage. We demonstrate that regression models accommodating RISC abundance and controlling for other mediating factors fit the expression profiles of known target pairs substantially better than models based on m/miRNA expressions alone, and lead to verifications of computational target pair predictions that are more sensitive than those based on marginal expression levels. Because our models are fully independent of exogenous results from sequence-based computational methods, they are appropriate for use as either a primary or secondary source of information regarding m/miRNA target pair relationships, especially in conjunction with high-throughput expression studies. PMID:19779550

Cigarette smoking substantially alters plasma microRNA profiles in healthy subjects

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Takahashi, Kei; Yokota, Shin-ichi; Tatsumi, Naoyuki; Fukami, Tatsuki; Yokoi, Tsuyoshi; Nakajima, Miki, E-mail: nmiki@p.kanazawa-u.ac.jp

2013-10-01

Circulating microRNAs (miRNAs) are receiving attention as potential biomarkers of various diseases, including cancers, chronic obstructive pulmonary disease, and cardiovascular disease. However, it is unknown whether the levels of circulating miRNAs in a healthy subject might vary with external factors in daily life. In this study, we investigated whether cigarette smoking, a habit that has spread throughout the world and is a risk factor for various diseases, affects plasma miRNA profiles. We determined the profiles of 11 smokers and 7 non-smokers by TaqMan MicroRNA array analysis. A larger number of miRNAs were detected in smokers than in non-smokers, and the plasma levels of two-thirds of the detected miRNAs (43 miRNAs) were significantly higher in smokers than in non-smokers. A principal component analysis of the plasma miRNA profiles clearly separated smokers and non-smokers. Twenty-four of the miRNAs were previously reported to be potential biomarkers of disease, suggesting the possibility that smoking status might interfere with the diagnosis of disease. Interestingly, we found that quitting smoking altered the plasma miRNA profiles to resemble those of non-smokers. These results suggested that the differences in the plasma miRNA profiles between smokers and non-smokers could be attributed to cigarette smoking. In addition, we found that an acute exposure of ex-smokers to cigarette smoke (smoking one cigarette) did not cause a dramatic change in the plasma miRNA profile. In conclusion, we found that repeated cigarette smoking substantially alters the plasma miRNA profile, interfering with the diagnosis of disease or signaling potential smoking-related diseases. - Highlights: • Plasma miRNA profiles were unambiguously different between smokers and non-smokers. • Smoking status might interfere with the diagnosis of disease using plasma miRNAs. • Changes of plasma miRNA profiles may be a signal of smoking-related diseases.

Cigarette smoking substantially alters plasma microRNA profiles in healthy subjects

International Nuclear Information System (INIS)

Takahashi, Kei; Yokota, Shin-ichi; Tatsumi, Naoyuki; Fukami, Tatsuki; Yokoi, Tsuyoshi; Nakajima, Miki

2013-01-01

Circulating microRNAs (miRNAs) are receiving attention as potential biomarkers of various diseases, including cancers, chronic obstructive pulmonary disease, and cardiovascular disease. However, it is unknown whether the levels of circulating miRNAs in a healthy subject might vary with external factors in daily life. In this study, we investigated whether cigarette smoking, a habit that has spread throughout the world and is a risk factor for various diseases, affects plasma miRNA profiles. We determined the profiles of 11 smokers and 7 non-smokers by TaqMan MicroRNA array analysis. A larger number of miRNAs were detected in smokers than in non-smokers, and the plasma levels of two-thirds of the detected miRNAs (43 miRNAs) were significantly higher in smokers than in non-smokers. A principal component analysis of the plasma miRNA profiles clearly separated smokers and non-smokers. Twenty-four of the miRNAs were previously reported to be potential biomarkers of disease, suggesting the possibility that smoking status might interfere with the diagnosis of disease. Interestingly, we found that quitting smoking altered the plasma miRNA profiles to resemble those of non-smokers. These results suggested that the differences in the plasma miRNA profiles between smokers and non-smokers could be attributed to cigarette smoking. In addition, we found that an acute exposure of ex-smokers to cigarette smoke (smoking one cigarette) did not cause a dramatic change in the plasma miRNA profile. In conclusion, we found that repeated cigarette smoking substantially alters the plasma miRNA profile, interfering with the diagnosis of disease or signaling potential smoking-related diseases. - Highlights: • Plasma miRNA profiles were unambiguously different between smokers and non-smokers. • Smoking status might interfere with the diagnosis of disease using plasma miRNAs. • Changes of plasma miRNA profiles may be a signal of smoking-related diseases

MicroRNA Expression Varies according to Glucose Tolerance, Measurement Platform, and Biological Source

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S. Dias

2017-01-01

Full Text Available Dysregulated microRNA (miRNA expression is observed during type 2 diabetes (T2D, although the consistency of miRNA expression across measurement platform and biological source is uncertain. Here we report miRNA profiling in the whole blood and serum of South African women with different levels of glucose tolerance, using next generation sequencing (NGS and quantitative real time PCR (qRT-PCR. Whole blood-derived miRNAs from women with newly diagnosed T2D (n=4, impaired glucose tolerance (IGT (n=4, and normal glucose tolerance (NGT (n=4 were subjected to NGS, whereafter transcript levels of selected miRNAs were quantified in the whole blood and serum of these women using qRT-PCR. Of the five significantly differentially expressed miRNAs identified by NGS, only the directional increase of miR-27b in women with IGT compared to NGT was confirmed in whole blood and serum, using qRT-PCR. Functional enrichment of miR-27b gene targets identified biological pathways associated with glucose transport and insulin regulation. In conclusion, this study showed poor correlation in miRNA expression profiled using NGS and qRT-PCR and in whole blood and serum. The consistent increased expression of miR-27b in women with IGT compared to NGT across measurement platform and biological source holds potential as a biomarker for risk stratification in our population.

BioVLAB-MMIA: a cloud environment for microRNA and mRNA integrated analysis (MMIA) on Amazon EC2.

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Lee, Hyungro; Yang, Youngik; Chae, Heejoon; Nam, Seungyoon; Choi, Donghoon; Tangchaisin, Patanachai; Herath, Chathura; Marru, Suresh; Nephew, Kenneth P; Kim, Sun

2012-09-01

MicroRNAs, by regulating the expression of hundreds of target genes, play critical roles in developmental biology and the etiology of numerous diseases, including cancer. As a vast amount of microRNA expression profile data are now publicly available, the integration of microRNA expression data sets with gene expression profiles is a key research problem in life science research. However, the ability to conduct genome-wide microRNA-mRNA (gene) integration currently requires sophisticated, high-end informatics tools, significant expertise in bioinformatics and computer science to carry out the complex integration analysis. In addition, increased computing infrastructure capabilities are essential in order to accommodate large data sets. In this study, we have extended the BioVLAB cloud workbench to develop an environment for the integrated analysis of microRNA and mRNA expression data, named BioVLAB-MMIA. The workbench facilitates computations on the Amazon EC2 and S3 resources orchestrated by the XBaya Workflow Suite. The advantages of BioVLAB-MMIA over the web-based MMIA system include: 1) readily expanded as new computational tools become available; 2) easily modifiable by re-configuring graphic icons in the workflow; 3) on-demand cloud computing resources can be used on an "as needed" basis; 4) distributed orchestration supports complex and long running workflows asynchronously. We believe that BioVLAB-MMIA will be an easy-to-use computing environment for researchers who plan to perform genome-wide microRNA-mRNA (gene) integrated analysis tasks.

Dose-responsiveness and persistence of microRNA expression alterations induced by cigarette smoke in mouse lung

International Nuclear Information System (INIS)

Izzotti, Alberto; Larghero, Patrizia; Longobardi, Mariagrazia; Cartiglia, Cristina; Camoirano, Anna; Steele, Vernon E.; De Flora, Silvio

2011-01-01

Our previous studies demonstrated that exposure to cigarette smoke (CS), either mainstream or environmental, results in a remarkable downregulation of microRNA expression in the lung of both mice and rats. The goals of the present study were to evaluate the dose responsiveness to CS and the persistence of microRNA alterations after smoking cessation. ICR (CD-1) neonatal mice were exposed whole-body to mainstream CS, at the doses of 119, 292, 438, and 631 mg/m 3 of total particulate matter. Exposure started within 12 h after birth and continued daily for 4 weeks. The levels of bulky DNA adducts and 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodGuo) were measured by 32 P postlabeling procedures, and the expression of 697 mouse microRNAs was analyzed by microarray. The highest CS dose was lethal. Exposure to CS caused a dose-dependent increase of DNA alterations. DNA adducts and, even more sharply, 8-oxodGuo were reverted 1 and 4 weeks after smoking cessation. Exposure to CS resulted in an evident dysregulation of microRNA expression profiles, mainly in the sense of downregulation. The two lowest doses were not particularly effective, while the highest nonlethal dose produced extensive microRNA alterations. The expression of most downregulated microRNAs, including among others 7 members of the let-7 family, was restored one week after smoking cessation. However, the recovery was incomplete for a limited array of microRNAs, including mir-34b, mir-345, mir-421, mir-450b, mir-466, and mir-469. Thus, it appears that microRNAs mainly behave as biomarkers of effect and that exposure to high-dose, lasting for an adequate period of time, is needed to trigger the CS-related carcinogenesis process in the experimental animal model used.

Correlation analyses revealed global microRNA-mRNA expression associations in human peripheral blood mononuclear cells.

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Wang, Lan; Zhu, Jiang; Deng, Fei-Yan; Wu, Long-Fei; Mo, Xing-Bo; Zhu, Xiao-Wei; Xia, Wei; Xie, Fang-Fei; He, Pei; Bing, Peng-Fei; Qiu, Ying-Hua; Lin, Xiang; Lu, Xin; Zhang, Lei; Yi, Neng-Jun; Zhang, Yong-Hong; Lei, Shu-Feng

2018-02-01

MicroRNAs (miRNAs) can regulate gene expression through binding to complementary sites in the 3'-untranslated regions of target mRNAs, which will lead to existence of correlation in expression between miRNA and mRNA. However, the miRNA-mRNA correlation patterns are complex and remain largely unclear yet. To establish the global correlation patterns in human peripheral blood mononuclear cells (PBMCs), multiple miRNA-mRNA correlation analyses and expression quantitative trait locus (eQTL) analysis were conducted in this study. We predicted and achieved 861 miRNA-mRNA pairs (65 miRNAs, 412 mRNAs) using multiple bioinformatics programs, and found global negative miRNA-mRNA correlations in PBMC from all 46 study subjects. Among the 861 pairs of correlations, 19.5% were significant (P correlation network was complex and highlighted key miRNAs/genes in PBMC. Some miRNAs, such as hsa-miR-29a, hsa-miR-148a, regulate a cluster of target genes. Some genes, e.g., TNRC6A, are regulated by multiple miRNAs. The identified genes tend to be enriched in molecular functions of DNA and RNA binding, and biological processes such as protein transport, regulation of translation and chromatin modification. The results provided a global view of the miRNA-mRNA expression correlation profile in human PBMCs, which would facilitate in-depth investigation of biological functions of key miRNAs/mRNAs and better understanding of the pathogenesis underlying PBMC-related diseases.

MicroRNA profiling reveals distinct signatures in B cell chronic lymphocytic leukemias

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Calin, George Adrian; Liu, Chang-Gong; Sevignani, Cinzia; Ferracin, Manuela; Felli, Nadia; Dumitru, Calin Dan; Shimizu, Masayoshi; Cimmino, Amelia; Zupo, Simona; Dono, Mariella; Dell'Aquila, Marie L.; Alder, Hansjuerg; Rassenti, Laura; Kipps, Thomas J.; Bullrich, Florencia; Negrini, Massimo; Croce, Carlo M.

2004-01-01

Little is known about the expression levels or function of micro-RNAs (miRNAs) in normal and neoplastic cells, although it is becoming clear that miRNAs play important roles in the regulation of gene expression during development [Ambros, V. (2003) Cell 113, 673–676; McManus, M. T. (2003) Semin. Cancer Biol. 13, 253–258]. We now report the genomewide expression profiling of miRNAs in human B cell chronic lymphocytic leukemia (CLL) by using a microarray containing hundreds of human precursor and mature miRNA oligonucleotide probes. This approach allowed us to identify significant differences in miRNome expression between CLL samples and normal CD5+ B cells; data were confirmed by Northern blot analyses and real-time RT-PCR. At least two distinct clusters of CLL samples can be identified that were associated with the presence or absence of Zap-70 expression, a predictor of early disease progression. Two miRNA signatures were associated with the presence or absence of mutations in the expressed Ig variableregion genes or with deletions at 13q14, respectively. These data suggest that miRNA expression patterns have relevance to the biological and clinical behavior of this leukemia. PMID:15284443

Phenotypic and microRNA transcriptomic profiling of the MDA-MB-231 spheroid-enriched CSCs with comparison of MCF-7 microRNA profiling dataset

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Lily Boo

2017-07-01

Full Text Available Breast cancer spheroids have been widely used as in vitro models of cancer stem cells (CSCs, yet little is known about their phenotypic characteristics and microRNAs (miRNAs expression profiles. The objectives of this research were to evaluate the phenotypic characteristics of MDA-MB-231 spheroid-enriched cells for their CSCs properties and also to determine their miRNAs expression profile. Similar to our previously published MCF-7 spheroid, MDA-MB-231 spheroid also showed typical CSCs characteristics namely self-renewability, expression of putative CSCs-related surface markers and enhancement of drug resistance. From the miRNA profile, miR-15b, miR-34a, miR-148a, miR-628 and miR-196b were shown to be involved in CSCs-associated signalling pathways in both models of spheroids, which highlights the involvement of these miRNAs in maintaining the CSCs features. In addition, unique clusters of miRNAs namely miR-205, miR-181a and miR-204 were found in basal-like spheroid whereas miR-125, miR-760, miR-30c and miR-136 were identified in luminal-like spheroid. Our results highlight the roles of miRNAs as well as novel perspectives of the relevant pathways underlying spheroid-enriched CSCs in breast cancer.

MicroRNA expression in a phosphaturic mesenchymal tumour

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Darrell Green

2017-12-01

Full Text Available Phosphaturic mesenchymal tumours are a heterogeneous set of bone and soft tissue neoplasms that can cause a number of paraneoplastic syndromes such as tumour induced osteomalacia. The term phosphaturic comes from the common finding that these tumours secrete high levels of fibroblast growth factor 23 which causes renal phosphate wasting leading to hypophosphatemia. Phosphaturic mesenchymal tumours are rare and diagnosis is difficult. A very active 68 year old male presented with bone pain and muscle weakness. He was hypophosphataemic and total alkaline phosphatase was markedly elevated. The patient was placed on vitamin D supplementation but his condition progressed. In the fifth year of presentation the patient required the use of a wheelchair and described “explosive” bone pain on physical contact. Serum 1,25 dihydroxyvitamin D was low and serum fibroblast growth factor 23 was significantly elevated, raising suspicion of a phosphaturic mesenchymal tumour. A lesion was detected in his left femoral head and the patient underwent a total hip replacement. The patient displayed a rapid improvement to his condition and during a three year follow up period he returned to an active lifestyle. As molecular testing may help provide a robust diagnosis and is particularly useful in rare diseases we took a next generation sequencing approach to identify a differential expression of small RNAs in the resected tumour. Small RNAs are non-coding RNA molecules that play a key role in regulation of gene expression and can be used as specific biomarkers. We found an upregulation of miR-197. We also found a downregulation of miR-20b, miR-144 and miR-335 which is a small RNA profile typical of osteosarcoma. MiR-21, the most frequently upregulated microRNA in cancer, was downregulated. We conclude that the specific small RNA profile is typical of osteosarcoma except for the downregulation of oncogenic miR-21. Transcriptional plasticity of miR-197, which is

Analysis of plasma microRNA expression profiles revealed different cancer susceptibility in healthy young adult smokers and middle-aged smokers.

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Shi, Bing; Gao, Hongmin; Zhang, Tianyang; Cui, Qinghua

2016-04-19

Cigarette smoking is a world-wide habit and an important risk factor for cancer. It was known that cigarette smoking can change the expression of circulating microRNAs (miRNAs) in healthy middle-aged adults. However, it remains unclear whether cigarette smoking can change the levels of circulating miRNAs in young healthy smokers and whether there are differences in cancer susceptibility for the two cases. In this study, the miRNA expression profiles of 28 smokers and 12 non-smokers were determined by Agilent human MicroRNA array. We further performed bioinformatics analysis for the differentially expressed miRNAs. The result showed that 35 miRNAs were differentially expressed. Among them, 24 miRNAs were up-regulated and 11 miRNAs were down-regulated in smokers. Functional enrichment analysis showed that the deregulated miRNAs are related to immune system and hormones regulation. Strikingly, the up-regulated miRNAs are mostly associated with hematologic cancers, such as lymphoma, leukemia. As a comparison, the up-regulated plasma miRNAs in middle-aged smokers are mostly associated with solid cancers, such as hepatocellular carcinoma and lung cancer, suggesting that smoking could have different influences on young adults and middle-aged adults. In a conclusion, we identified the circulating miRNAs deregulated by cigarette smoking and revealed that the age-dependent deregulated miRNAs tend to be mainly involved in different types of human cancers.

Integrated analysis of microRNA and gene expression profiles reveals a functional regulatory module associated with liver fibrosis.

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Chen, Wei; Zhao, Wenshan; Yang, Aiting; Xu, Anjian; Wang, Huan; Cong, Min; Liu, Tianhui; Wang, Ping; You, Hong

2017-12-15

Liver fibrosis, characterized with the excessive accumulation of extracellular matrix (ECM) proteins, represents the final common pathway of chronic liver inflammation. Ever-increasing evidence indicates microRNAs (miRNAs) dysregulation has important implications in the different stages of liver fibrosis. However, our knowledge of miRNA-gene regulation details pertaining to such disease remains unclear. The publicly available Gene Expression Omnibus (GEO) datasets of patients suffered from cirrhosis were extracted for integrated analysis. Differentially expressed miRNAs (DEMs) and genes (DEGs) were identified using GEO2R web tool. Putative target gene prediction of DEMs was carried out using the intersection of five major algorithms: DIANA-microT, TargetScan, miRanda, PICTAR5 and miRWalk. Functional miRNA-gene regulatory network (FMGRN) was constructed based on the computational target predictions at the sequence level and the inverse expression relationships between DEMs and DEGs. DAVID web server was selected to perform KEGG pathway enrichment analysis. Functional miRNA-gene regulatory module was generated based on the biological interpretation. Internal connections among genes in liver fibrosis-related module were determined using String database. MiRNA-gene regulatory modules related to liver fibrosis were experimentally verified in recombinant human TGFβ1 stimulated and specific miRNA inhibitor treated LX-2 cells. We totally identified 85 and 923 dysregulated miRNAs and genes in liver cirrhosis biopsy samples compared to their normal controls. All evident miRNA-gene pairs were identified and assembled into FMGRN which consisted of 990 regulations between 51 miRNAs and 275 genes, forming two big sub-networks that were defined as down-network and up-network, respectively. KEGG pathway enrichment analysis revealed that up-network was prominently involved in several KEGG pathways, in which "Focal adhesion", "PI3K-Akt signaling pathway" and "ECM

Identification of serum microRNA biomarkers for tuberculosis using RNA-seq.

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Zhang, Hongtai; Sun, Zhaogang; Wei, Wenjing; Liu, Zhonghui; Fleming, Joy; Zhang, Shuai; Lin, Nan; Wang, Ming; Chen, Maoshan; Xu, Yuhui; Zhou, Jie; Li, Chuanyou; Bi, Lijun; Zhou, Guangming

2014-01-01

Tuberculosis (TB) remains a significant human health issue. More effective biomarkers for use in tuberculosis prevention, diagnosis, and treatment, including markers that can discriminate between healthy individuals and those with latent infection, are urgently needed. To identify a set of such markers, we used Solexa sequencing to examine microRNA expression in the serum of patients with active disease, healthy individuals with latent TB, and those with or without prior BCG inoculation. We identified 24 microRNAs that are up-regulated (2.85-1285.93 fold) and 6 microRNAs that are down-regulated (0.003-0.11 fold) (PmicroRNAs were up-regulated (2.05-2454.58 fold) and 11 were down-regulated (0.001-0.42 fold) (PmicroRNAs were differentially-expressed in BCG-inoculated relative to un-inoculated individuals (18 up-regulated 2.9-499.29 fold, 116 down-regulated 0.0002-0.5 fold), providing insights into the effects of BCG inoculation at the microRNA level. Target prediction of differentially-expressed microRNAs by microRNA-Gene Network analysis and analysis of pathways affected suggest that regulation of the host immune system by microRNAs is likely to be one of the main factors in the pathogenesis of tuberculosis. qRT-PCR validation indicated that hsa-miR-196b and hsa-miR-376c have potential as markers for active TB disease. The microRNA differential-expression profiles generated in this study provide a good foundation for the development of markers for TB diagnosis, and for investigations on the role of microRNAs in BCG-inoculated and latent-infected individuals.

Arsenic exposure triggers a shift in microRNA expression.

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Sturchio, Elena; Colombo, Teresa; Boccia, Priscilla; Carucci, Nicoletta; Meconi, Claudia; Minoia, Claudio; Macino, Giuseppe

2014-02-15

Exposure to inorganic Arsenic (iAs) through drinking water is a major public health problem affecting most countries. iAs has been classified by the International Agency for Research on Cancer as Group 1: "Carcinogenic to humans". Although numerous studies have shown the related adverse effects of iAs, sensitive appropriate biomarkers for studies of environmental epidemiology are still required. The present work aims at investigate the role of microRNAs (miRNAs), powerful negative regulators of gene expression, playing a key role in many physiological and pathological cellular processes, in iAs exposure. To this end, we analyzed miRNA changes in expression profile triggered by iAs exposure in Jurkat cell line. We used microarray technology to profile the expression of miRNAs following 2 μmol/L sodium arsenite treatment at different time points. Moreover, we performed phenotypic analysis of iAs treated cells. Real Time Polymerase Chain Reaction (RT-PCR) was used to validate miRNA microarray data and to assay expression modulation of selected relevant mRNAs. Finally, bioinformatics techniques were applied to reconstruct iAs-relevant molecular pathways and miRNA regulatory networks from the expression data. We report miRNAs modulated after iAs treatment in Jurkat cells. In particular, we highlight 36 miRNAs exhibiting consistent dysregulation and particularly a panel of 8 miRNAs which we also validated by RT-PCR analysis. Computational analysis of lists of putative target genes for these 8 miRNAs points to an involvement in arsenic-response pathways, for a subset of them, that were analyzed by RT-PCR. Furthermore, iAs exposure reveals induction of cell cycle progression and the failure of apoptosis, supporting the idea of iAs carcinogenic activity. Our study provides a list of miRNAs whose expression levels are affected by iAs treatment, corroborating the importance of proceeding with the hunt for specific subset of miRNAs, which can serve as potential biomarkers of

Comparison of the Functional microRNA Expression in Immune Cell Subsets of Neonates and Adults

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Yu, Hong-Ren; Hsu, Te-Yao; Huang, Hsin-Chun; Kuo, Ho-Chang; Li, Sung-Chou; Yang, Kuender D.; Hsieh, Kai-Sheng

2016-01-01

Diversity of biological molecules in newborn and adult immune cells contributes to differences in cell function and atopic properties. Micro RNAs (miRNAs) are reported to involve in the regulation of immune system. Therefore, determining the miRNA expression profile of leukocyte subpopulations is important for understanding immune system regulation. In order to explore the unique miRNA profiling that contribute to altered immune in neonates, we comprehensively analyzed the functional miRNA signatures of eight leukocyte subsets (polymorphonuclear cells, monocytes, CD4+ T cells, CD8+ T cells, natural killer cells, B cells, plasmacytoid dendritic cells, and myeloid dendritic cells) from both neonatal and adult umbilical cord and peripheral blood samples, respectively. We observed distinct miRNA profiles between adult and neonatal blood leukocyte subsets, including unique miRNA signatures for each cell lineage. Leukocyte miRNA signatures were altered after stimulation. Adult peripheral leukocytes had higher let-7b-5p expression levels compared to neonatal cord leukocytes across multiple subsets, irrespective of stimulation. Transfecting neonatal monocytes with a let-7b-5p mimic resulted in a reduction of LPS-induced interleukin (IL)-6 and TNF-α production, while transfection of a let-7b-5p inhibitor into adult monocytes enhanced IL-6 and TNF-α production. With this functional approach, we provide intact differential miRNA expression profiling of specific immune cell subsets between neonates and adults. These studies serve as a basis to further understand the altered immune response observed in neonates and advance the development of therapeutic strategies. PMID:28066425

Identification of serum microRNA biomarkers for tuberculosis using RNA-seq.

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Hongtai Zhang

Full Text Available Tuberculosis (TB remains a significant human health issue. More effective biomarkers for use in tuberculosis prevention, diagnosis, and treatment, including markers that can discriminate between healthy individuals and those with latent infection, are urgently needed. To identify a set of such markers, we used Solexa sequencing to examine microRNA expression in the serum of patients with active disease, healthy individuals with latent TB, and those with or without prior BCG inoculation. We identified 24 microRNAs that are up-regulated (2.85-1285.93 fold and 6 microRNAs that are down-regulated (0.003-0.11 fold (P<0.05 in patients with active TB relative to the three groups of healthy controls. In addition, 75 microRNAs were up-regulated (2.05-2454.58 fold and 11 were down-regulated (0.001-0.42 fold (P<0.05 in latent-TB infected individuals relative to BCG- inoculated individuals. Of interest, 134 microRNAs were differentially-expressed in BCG-inoculated relative to un-inoculated individuals (18 up-regulated 2.9-499.29 fold, 116 down-regulated 0.0002-0.5 fold, providing insights into the effects of BCG inoculation at the microRNA level. Target prediction of differentially-expressed microRNAs by microRNA-Gene Network analysis and analysis of pathways affected suggest that regulation of the host immune system by microRNAs is likely to be one of the main factors in the pathogenesis of tuberculosis. qRT-PCR validation indicated that hsa-miR-196b and hsa-miR-376c have potential as markers for active TB disease. The microRNA differential-expression profiles generated in this study provide a good foundation for the development of markers for TB diagnosis, and for investigations on the role of microRNAs in BCG-inoculated and latent-infected individuals.

Aberrant microRNA expression in multiple myeloma

DEFF Research Database (Denmark)

Dimopoulos, Konstantinos; Gimsing, Peter; Grønbæk, Kirsten

2013-01-01

Multiple myeloma (MM) is a devastating disease with a complex biology, and in spite of improved survivability by novel treatment strategies over the last decade, MM is still incurable by current therapy. MicroRNAs (miRNAs) are small, non-coding RNA molecules that regulate gene expression at a post...

Analysis of microRNA Expression Profiles Induced by Yiqifumai Injection in Rats with Chronic Heart Failure

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Yu Zhao

2018-02-01

Full Text Available Background: Yiqifumai Injection (YQFM is clinically used to treat various cardiovascular diseases including chronic heart failure (CHF. The efficacy of YQFM for treating heart failure has been suggested, but the mechanism of action for pharmacological effects of YQFM is unclear.Methods: Echocardiography detection, left ventricular intubation evaluation, histopathology and immunohistochemical examination were performed in CHF rats to evaluate the cardioprotective effect of YQFM. Rat miRNA microarray and bioinformatics analysis were employed to investigate the differentially expressed microRNAs. In vitro models of AngII-induced hypertrophy and t-BHP induced oxidative stress in H9c2 myocardial cells were used to validate the anti-hypertrophy and anti-apoptosis effects of YQFM. Measurement of cell surface area, ATP content and cell viability, Real-time PCR and Western blot were performed.Results: YQFM significantly improved the cardiac function of CHF rats by increasing left ventricular ejection fraction and fractional shortening, decreasing left ventricular internal diameter and enhancing cardiac output. Seven microRNAs which have a reversible regulation by YQFM treatment were found. Among them, miR-21-3p and miR-542-3p are related to myocardial hypertrophy and cell proliferation, respectively and were further verified by RT-PCR. Target gene network was established and potential related signaling pathways were predicted. YQFM could significantly alleviate AngII induced hypertrophy in cellular model. It also significantly increased cell viabilities and ATP content in t-BHP induced apoptotic cell model. Western blot analysis showed that YQFM could increase the phosphorylation of Akt.Conclusion: Our findings provided scientific evidence to uncover the mechanism of action of YQFM on miRNAs regulation against CHF by miRNA expression profile technology. The results indicated that YQFM has a potential effect on alleviate cardiac hypertrophy and apoptosis

MicroRNA transcriptome profiles during swine skeletal muscle development

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Sonstegard Tad S

2009-02-01

Full Text Available Abstract Background MicroRNA (miR are a class of small RNAs that regulate gene expression by inhibiting translation of protein encoding transcripts. To evaluate the role of miR in skeletal muscle of swine, global microRNA abundance was measured at specific developmental stages including proliferating satellite cells, three stages of fetal growth, day-old neonate, and the adult. Results Twelve potential novel miR were detected that did not match previously reported sequences. In addition, a number of miR previously reported to be expressed in mammalian muscle were detected, having a variety of abundance patterns through muscle development. Muscle-specific miR-206 was nearly absent in proliferating satellite cells in culture, but was the highest abundant miR at other time points evaluated. In addition, miR-1 was moderately abundant throughout developmental stages with highest abundance in the adult. In contrast, miR-133 was moderately abundant in adult muscle and either not detectable or lowly abundant throughout fetal and neonate development. Changes in abundance of ubiquitously expressed miR were also observed. MiR-432 abundance was highest at the earliest stage of fetal development tested (60 day-old fetus and decreased throughout development to the adult. Conversely, miR-24 and miR-27 exhibited greatest abundance in proliferating satellite cells and the adult, while abundance of miR-368, miR-376, and miR-423-5p was greatest in the neonate. Conclusion These data present a complete set of transcriptome profiles to evaluate miR abundance at specific stages of skeletal muscle growth in swine. Identification of these miR provides an initial group of miR that may play a vital role in muscle development and growth.

Evaluation of microRNA Expression in Patients with Herpes Zoster

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Xihan Li

2016-12-01

Full Text Available Reactivated varicella-zoster virus (VZV, which lies latent in the dorsal root ganglions and cranial nerves before its reactivation, is capable of causing herpes zoster (HZ, but the specific mechanism of virus reactivation and latency remains unknown. It was proposed that circulating microRNAs (miRNAs in body fluids could potentially indicate infection. However, the connection between herpes zoster and circulating miRNAs has not been demonstrated. In this study, 41 HZ patients without superinfection were selected. The serum miRNA levels were analyzed by TaqMan low density array (TLDA and confirmed individually by quantitative reverse transcription PCR (RT-qPCR analysis. Thirty-five age-matched subjects without any infectious diseases or inflammation were selected as controls. The results showed that the serum miRNA expression profiles in 41 HZ patients were different from those of control subjects. Specifically, 18 miRNAs were up-regulated and 126 were down-regulated more than two-fold in HZ patients compared with controls. The subsequent confirmation of these results by qRT-PCR, as well as receiver operating characteristic (ROC curve analysis, revealed that six kinds of miRNAs, including miR-190b, miR-571, miR-1276, miR-1303, miR-943, and miR-661, exhibited statistically significant enhanced expression levels (more than four-fold in HZ patients, compared with those of healthy controls and herpes simplex virus (HSV patients. Subsequently, it is proposed that these circulating miRNAs are capable of regulating numerous pathways and some may even participate in the inflammatory response or nervous system activity. This study has initially demonstrated that the serum miRNA expression profiles in HZ patients were different from those of uninfected individuals. Additionally, these findings also suggest that six of the altered miRNA could be potentially used as biomarkers to test for latent HZ infection.

A comparison of microRNA expression profiles from splenic hemangiosarcoma, splenic nodular hyperplasia, and normal spleens of dogs.

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Grimes, Janet A; Prasad, Nripesh; Levy, Shawn; Cattley, Russell; Lindley, Stephanie; Boothe, Harry W; Henderson, Ralph A; Smith, Bruce F

2016-12-03

Splenic masses are common in older dogs; yet diagnosis preceding splenectomy and histopathology remains elusive. MicroRNAs (miRNAs) are short, non-coding RNAs that play a role in post-transcriptional regulation, and differential expression of miRNAs between normal and tumor tissue has been used to diagnose neoplastic diseases. The objective of this study was to determine differential expression of miRNAs by use of RNA-sequencing in canine spleens that were histologically confirmed as hemangiosarcoma, nodular hyperplasia, or normal. Twenty-two miRNAs were found to be differentially expressed in hemangiosarcoma samples (4 between hemangiosarcoma and both nodular hyperplasia and normal spleen and 18 between hemangiosarcoma and normal spleen only). In particular, mir-26a, mir-126, mir-139, mir-140, mir-150, mir-203, mir-424, mir-503, mir-505, mir-542, mir-30e, mir-33b, mir-365, mir-758, mir-22, and mir-452 are of interest in the pathogenesis of hemangiosarcoma. Findings of this study confirm the hypothesis that miRNA expression profiles are different between canine splenic hemangiosarcoma, nodular hyperplasia, and normal spleens. A large portion of the differentially expressed miRNAs have roles in angiogenesis, with an additional group of miRNAs being dysregulated in vascular disease processes. Two other miRNAs have been implicated in cancer pathways such as PTEN and cell cycle checkpoints. The finding of multiple miRNAs with roles in angiogenesis and vascular disease is important, as hemangiosarcoma is a tumor of endothelial cells, which are driven by angiogenic stimuli. This study shows that miRNA dysregulation is a potential player in the pathogenesis of canine splenic hemangiosarcoma.

Detecting microRNA activity from gene expression data

LENUS (Irish Health Repository)

Madden, Stephen F

2010-05-18

Abstract Background MicroRNAs (miRNAs) are non-coding RNAs that regulate gene expression by binding to the messenger RNA (mRNA) of protein coding genes. They control gene expression by either inhibiting translation or inducing mRNA degradation. A number of computational techniques have been developed to identify the targets of miRNAs. In this study we used predicted miRNA-gene interactions to analyse mRNA gene expression microarray data to predict miRNAs associated with particular diseases or conditions. Results Here we combine correspondence analysis, between group analysis and co-inertia analysis (CIA) to determine which miRNAs are associated with differences in gene expression levels in microarray data sets. Using a database of miRNA target predictions from TargetScan, TargetScanS, PicTar4way PicTar5way, and miRanda and combining these data with gene expression levels from sets of microarrays, this method produces a ranked list of miRNAs associated with a specified split in samples. We applied this to three different microarray datasets, a papillary thyroid carcinoma dataset, an in-house dataset of lipopolysaccharide treated mouse macrophages, and a multi-tissue dataset. In each case we were able to identified miRNAs of biological importance. Conclusions We describe a technique to integrate gene expression data and miRNA target predictions from multiple sources.

Detecting microRNA activity from gene expression data.

LENUS (Irish Health Repository)

Madden, Stephen F

2010-01-01

BACKGROUND: MicroRNAs (miRNAs) are non-coding RNAs that regulate gene expression by binding to the messenger RNA (mRNA) of protein coding genes. They control gene expression by either inhibiting translation or inducing mRNA degradation. A number of computational techniques have been developed to identify the targets of miRNAs. In this study we used predicted miRNA-gene interactions to analyse mRNA gene expression microarray data to predict miRNAs associated with particular diseases or conditions. RESULTS: Here we combine correspondence analysis, between group analysis and co-inertia analysis (CIA) to determine which miRNAs are associated with differences in gene expression levels in microarray data sets. Using a database of miRNA target predictions from TargetScan, TargetScanS, PicTar4way PicTar5way, and miRanda and combining these data with gene expression levels from sets of microarrays, this method produces a ranked list of miRNAs associated with a specified split in samples. We applied this to three different microarray datasets, a papillary thyroid carcinoma dataset, an in-house dataset of lipopolysaccharide treated mouse macrophages, and a multi-tissue dataset. In each case we were able to identified miRNAs of biological importance. CONCLUSIONS: We describe a technique to integrate gene expression data and miRNA target predictions from multiple sources.

Genome Wide Expression Profiling of Cancer Cell Lines Cultured in Microgravity Reveals Significant Dysregulation of Cell Cycle and MicroRNA Gene Networks.

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Prasanna Vidyasekar

Full Text Available Zero gravity causes several changes in metabolic and functional aspects of the human body and experiments in space flight have demonstrated alterations in cancer growth and progression. This study reports the genome wide expression profiling of a colorectal cancer cell line-DLD-1, and a lymphoblast leukemic cell line-MOLT-4, under simulated microgravity in an effort to understand central processes and cellular functions that are dysregulated among both cell lines. Altered cell morphology, reduced cell viability and an aberrant cell cycle profile in comparison to their static controls were observed in both cell lines under microgravity. The process of cell cycle in DLD-1 cells was markedly affected with reduced viability, reduced colony forming ability, an apoptotic population and dysregulation of cell cycle genes, oncogenes, and cancer progression and prognostic markers. DNA microarray analysis revealed 1801 (upregulated and 2542 (downregulated genes (>2 fold in DLD-1 cultures under microgravity while MOLT-4 cultures differentially expressed 349 (upregulated and 444 (downregulated genes (>2 fold under microgravity. The loss in cell proliferative capacity was corroborated with the downregulation of the cell cycle process as demonstrated by functional clustering of DNA microarray data using gene ontology terms. The genome wide expression profile also showed significant dysregulation of post transcriptional gene silencing machinery and multiple microRNA host genes that are potential tumor suppressors and proto-oncogenes including MIR22HG, MIR17HG and MIR21HG. The MIR22HG, a tumor-suppressor gene was one of the highest upregulated genes in the microarray data showing a 4.4 log fold upregulation under microgravity. Real time PCR validated the dysregulation in the host gene by demonstrating a 4.18 log fold upregulation of the miR-22 microRNA. Microarray data also showed dysregulation of direct targets of miR-22, SP1, CDK6 and CCNA2.

Plasma microRNA profiles in rat models of hepatocellular injury, cholestasis, and steatosis.

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Yu Yamaura

Full Text Available MicroRNAs (miRNAs are small RNA molecules that function to modulate the expression of target genes, playing important roles in a wide range of physiological and pathological processes. The miRNAs in body fluids have received considerable attention as potential biomarkers of various diseases. In this study, we compared the changes of the plasma miRNA expressions by acute liver injury (hepatocellular injury or cholestasis and chronic liver injury (steatosis, steatohepatitis and fibrosis using rat models made by the administration of chemicals or special diets. Using miRNA array analysis, we found that the levels of a large number of miRNAs (121-317 miRNAs were increased over 2-fold and the levels of a small number of miRNAs (6-35 miRNAs were decreased below 0.5-fold in all models except in a model of cholestasis caused by bile duct ligation. Interestingly, the expression profiles were different between the models, and the hierarchical clustering analysis discriminated between the acute and chronic liver injuries. In addition, miRNAs whose expressions were typically changed in each type of liver injury could be specified. It is notable that, in acute liver injury models, the plasma level of miR-122, the most abundant miRNA in the liver, was more quickly and dramatically increased than the plasma aminotransferase level, reflecting the extent of hepatocellular injury. This study demonstrated that the plasma miRNA profiles could reflect the types of liver injury (e.g. acute/chronic liver injury or hepatocellular injury/cholestasis/steatosis/steatohepatitis/fibrosis and identified the miRNAs that could be specific and sensitive biomarkers of liver injury.

Differential MicroRNA Expression in Human Macrophages with Mycobacterium tuberculosis Infection of Beijing/W and Non-Beijing/W Strain Types.

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Zheng, Lin; Leung, Eric; Lee, Nelson; Lui, Grace; To, Ka-Fai; Chan, Raphael C Y; Ip, Margaret

2015-01-01

The role of microRNAs in association with Mycobacterium tuberculosis (MTB) infection and the immunology regulated by microRNAs upon MTB infection have not been fully unravelled. We examined the microRNA profiles of THP-1 macrophages upon the MTB infection of Beijing/W and non-Beijing/W clinical strains. We also studied the microRNA profiles of the host macrophages by microarray in a small cohort with active MTB disease, latent infection (LTBI), and from healthy controls. The results revealed that 14 microRNAs differentiated infections of Beijing/W from non-Beijing/W strains (PmicroRNAs in human macrophages was identified to differentiate active MTB disease from LTBI and healthy controls. Pathway analyses of these differentially expressed miRNAs suggest that the immune-regulatory interactions involving TGF-β signalling pathway take part in the dysregulation of critical TB processes in the macrophages, resulting in active expression of both cell communication and signalling transduction systems. We showed for the first time that the Beijing/W TB strains repressed a number of miRNAs expressions which may reflect their virulence characteristics in altering the host response. The unique signatures of 11 microRNAs may deserve further evaluation as candidates for biomarkers in the diagnosis of MTB and Beijing/W infections.

MicroRNA-31 functions as an oncogenic microRNA in mouse and human lung cancer cells by repressing specific tumor suppressors

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Liu, Xi; Sempere, Lorenzo F; Ouyang, Haoxu

2010-01-01

MicroRNAs (miRNAs) regulate gene expression. It has been suggested that obtaining miRNA expression profiles can improve classification, diagnostic, and prognostic information in oncology. Here, we sought to comprehensively identify the miRNAs that are overexpressed in lung cancer by conducting mi...

MicroRNA expression in nodal and extranodal Diffuse Large B-cell Lymphoma

DEFF Research Database (Denmark)

Mandrup, Charlotte; Petersen, Anders; Højfeldt, Anne Dirks

MicroRNA expression in nodal and extranodal Diffuse Large B-cell Lymphoma   C. Mandrup1, A. Petersen1, A. D. Hoejfeldt1, H. F. Thomsen1, J. Madsen1, J. Dahlgaard1, P. Johansen2, A. Bukh1, K. Dybkaer1 and H. E Johnsen1. 1Department of Hematology, 2Pathological Institute, Aalborg Hospital, Aarhus...... University Hospital, Aalborg, Denmark Introduction: The aim of this project was to analyse microRNA (miRNA) expression in nodal and extranodal diffuse large B-cell lymphoma (DLBCL). Manifestation at diagnosis may be nodal and/or extranodal. At present, there are no known determinants for none...... of the manifestations, and no way to predict the potential progression from nodal to extranodal disease. miRNA are small regulatory RNA molecules with core function to repress/cleave sequence complementary mRNA targets. Abnormalities in miRNA genetics and expression are known to affect initiation and development...

Exosomes Derived From Pancreatic Stellate Cells: MicroRNA Signature and Effects on Pancreatic Cancer Cells.

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Takikawa, Tetsuya; Masamune, Atsushi; Yoshida, Naoki; Hamada, Shin; Kogure, Takayuki; Shimosegawa, Tooru

2017-01-01

Pancreatic stellate cells (PSCs) interact with pancreatic cancer cells in the tumor microenvironment. Cell constituents including microRNAs may be exported from cells within membranous nanovesicles termed exosomes. Exosomes might play a pivotal role in intercellular communication. This study aimed to clarify the microRNA signature of PSC-derived exosomes and their effects on pancreatic cancer cells. Exosomes were prepared from the conditioned medium of immortalized human PSCs. MicroRNAs were prepared from the exosomes and their source PSCs, and the microRNA expression profiles were compared by microarray. The effects of PSC-derived exosomes on proliferation, migration, and the mRNA expression profiles were examined in pancreatic cancer cells. Pancreatic stellate cell-derived exosomes contained a variety of microRNAs including miR-21-5p. Several microRNAs such as miR-451a were enriched in exosomes compared to their source PSCs. Pancreatic stellate cell-derived exosomes stimulated the proliferation, migration and expression of mRNAs for chemokine (C - X - C motif) ligands 1 and 2 in pancreatic cancer cells. The stimulation of proliferation, migration, and chemokine gene expression by the conditioned medium of PSCs was suppressed by GW4869, an exosome inhibitor. We clarified the microRNA expression profile in PSC-derived exosomes. Pancreatic stellate cell-derived exosomes might play a role in the interactions between PSCs and pancreatic cancer cells.

Aging-induced dysregulation of dicer1-dependent microRNA expression impairs angiogenic capacity of rat cerebromicrovascular endothelial cells.

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Ungvari, Zoltan; Tucsek, Zsuzsanna; Sosnowska, Danuta; Toth, Peter; Gautam, Tripti; Podlutsky, Andrej; Csiszar, Agnes; Losonczy, Gyorgy; Valcarcel-Ares, M Noa; Sonntag, William E; Csiszar, Anna

2013-08-01

Age-related impairment of angiogenesis is likely to play a central role in cerebromicrovascular rarefaction and development of vascular cognitive impairment, but the underlying mechanisms remain elusive. To test the hypothesis that dysregulation of Dicer1 (ribonuclease III, a key enzyme of the microRNA [miRNA] machinery) impairs endothelial angiogenic capacity in aging, primary cerebromicrovascular endothelial cells (CMVECs) were isolated from young (3 months old) and aged (24 months old) Fischer 344 × Brown Norway rats. We found an age-related downregulation of Dicer1 expression both in CMVECs and in small cerebral vessels isolated from aged rats. In aged CMVECs, Dicer1 expression was increased by treatment with polyethylene glycol-catalase. Compared with young cells, aged CMVECs exhibited altered miRNA expression profile, which was associated with impaired proliferation, adhesion to vitronectin, collagen and fibronectin, cellular migration (measured by a wound-healing assay using electric cell-substrate impedance sensing technology), and impaired ability to form capillary-like structures. Overexpression of Dicer1 in aged CMVECs partially restored miRNA expression profile and significantly improved angiogenic processes. In young CMVECs, downregulation of Dicer1 (siRNA) resulted in altered miRNA expression profile associated with impaired proliferation, adhesion, migration, and tube formation, mimicking the aging phenotype. Collectively, we found that Dicer1 is essential for normal endothelial angiogenic processes, suggesting that age-related dysregulation of Dicer1-dependent miRNA expression may be a potential mechanism underlying impaired angiogenesis and cerebromicrovascular rarefaction in aging.

The microRNA Expression Profile in Donation after Cardiac Death (DCD Livers and Its Ability to Identify Primary Non Function.

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Shirin Elizabeth Khorsandi

Full Text Available Donation after cardiac death (DCD livers are marginal organs for transplant and their use is associated with a higher risk of primary non function (PNF or early graft dysfunction (EGD. The aim was to determine if microRNA (miRNA was able to discriminate between DCD livers of varying clinical outcome. DCD groups were categorized as PNF retransplanted within a week (n=7, good functional outcome (n=7 peak aspartate transaminase (AST ≤ 1000 IU/L and EGD (n=9 peak AST ≥ 2500 IU/L. miRNA was extracted from archival formalin fixed post-perfusion tru-cut liver biopsies. High throughput expression analysis was performed using miRNA arrays. Bioinformatics for expression data analysis was performed and validated with real time quantitative PCR (RT-qPCR. The function of miRNA of interest was investigated using computational biology prediction algorithms. From the array analysis 16 miRNAs were identified as significantly different (p<0.05. On RT-qPCR miR-155 and miR-940 had the highest expression across all three DCD clinical groups. Only one miRNA, miR-22, was validated with marginal significance, to have differential expression between the three groups (p=0.049. From computational biology miR-22 was predicted to affect signalling pathways that impact protein turnover, metabolism and apoptosis/cell cycle. In conclusion, microRNA expression patterns have a low diagnostic potential clinically in discriminating DCD liver quality and outcome.

Differential MicroRNA Expression in Human Macrophages with Mycobacterium tuberculosis Infection of Beijing/W and Non-Beijing/W Strain Types.

Directory of Open Access Journals (Sweden)

Lin Zheng

Full Text Available The role of microRNAs in association with Mycobacterium tuberculosis (MTB infection and the immunology regulated by microRNAs upon MTB infection have not been fully unravelled. We examined the microRNA profiles of THP-1 macrophages upon the MTB infection of Beijing/W and non-Beijing/W clinical strains. We also studied the microRNA profiles of the host macrophages by microarray in a small cohort with active MTB disease, latent infection (LTBI, and from healthy controls.The results revealed that 14 microRNAs differentiated infections of Beijing/W from non-Beijing/W strains (P<0.05. A unique signature of 11 microRNAs in human macrophages was identified to differentiate active MTB disease from LTBI and healthy controls. Pathway analyses of these differentially expressed miRNAs suggest that the immune-regulatory interactions involving TGF-β signalling pathway take part in the dysregulation of critical TB processes in the macrophages, resulting in active expression of both cell communication and signalling transduction systems.We showed for the first time that the Beijing/W TB strains repressed a number of miRNAs expressions which may reflect their virulence characteristics in altering the host response. The unique signatures of 11 microRNAs may deserve further evaluation as candidates for biomarkers in the diagnosis of MTB and Beijing/W infections.

Characterization and Profiling of Liver microRNAs by RNA-sequencing in Cattle Divergently Selected for Residual Feed Intake

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Wijdan Al-Husseini

2016-10-01

Full Text Available MicroRNAs (miRNAs are short non-coding RNAs that post-transcriptionally regulate expression of mRNAs in many biological pathways. Liver plays an important role in the feed efficiency of animals and high and low efficient cattle demonstrated different gene expression profiles by microarray. Here we report comprehensive miRNAs profiles by next-gen deep sequencing in Angus cattle divergently selected for residual feed intake (RFI and identify miRNAs related to feed efficiency in beef cattle. Two microRNA libraries were constructed from pooled RNA extracted from livers of low and high RFI cattle, and sequenced by Illumina genome analyser. In total, 23,628,103 high quality short sequence reads were obtained and more than half of these reads were matched to the bovine genome (UMD 3.1. We identified 305 known bovine miRNAs. Bta-miR-143, bta-miR-30, bta-miR-122, bta-miR-378, and bta-let-7 were the top five most abundant miRNAs families expressed in liver, representing more than 63% of expressed miRNAs. We also identified 52 homologous miRNAs and 10 novel putative bovine-specific miRNAs, based on precursor sequence and the secondary structure and utilizing the miRBase (v. 21. We compared the miRNAs profile between high and low RFI animals and ranked the most differentially expressed bovine known miRNAs. Bovine miR-143 was the most abundant miRNA in the bovine liver and comprised 20% of total expressed mapped miRNAs. The most highly expressed miRNA in liver of mice and humans, miR-122, was the third most abundant in our cattle liver samples. We also identified 10 putative novel bovine-specific miRNA candidates. Differentially expressed miRNAs between high and low RFI cattle were identified with 18 miRNAs being up-regulated and 7 other miRNAs down-regulated in low RFI cattle. Our study has identified comprehensive miRNAs expressed in bovine liver. Some of the expressed miRNAs are novel in cattle. The differentially expressed miRNAs between high and low RFI

Identifying microRNA/mRNA dysregulations in ovarian cancer.

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Miles, Gregory D; Seiler, Michael; Rodriguez, Lorna; Rajagopal, Gunaretnam; Bhanot, Gyan

2012-03-27

MicroRNAs are a class of noncoding RNA molecules that co-regulate the expression of multiple genes via mRNA transcript degradation or translation inhibition. Since they often target entire pathways, they may be better drug targets than genes or proteins. MicroRNAs are known to be dysregulated in many tumours and associated with aggressive or poor prognosis phenotypes. Since they regulate mRNA in a tissue specific manner, their functional mRNA targets are poorly understood. In previous work, we developed a method to identify direct mRNA targets of microRNA using patient matched microRNA/mRNA expression data using an anti-correlation signature. This method, applied to clear cell Renal Cell Carcinoma (ccRCC), revealed many new regulatory pathways compromised in ccRCC. In the present paper, we apply this method to identify dysregulated microRNA/mRNA mechanisms in ovarian cancer using data from The Cancer Genome Atlas (TCGA). TCGA Microarray data was normalized and samples whose class labels (tumour or normal) were ambiguous with respect to consensus ensemble K-Means clustering were removed. Significantly anti-correlated and correlated genes/microRNA differentially expressed between tumour and normal samples were identified. TargetScan was used to identify gene targets of microRNA. We identified novel microRNA/mRNA mechanisms in ovarian cancer. For example, the expression level of RAD51AP1 was found to be strongly anti-correlated with the expression of hsa-miR-140-3p, which was significantly down-regulated in the tumour samples. The anti-correlation signature was present separately in the tumour and normal samples, suggesting a direct causal dysregulation of RAD51AP1 by hsa-miR-140-3p in the ovary. Other pairs of potentially biological relevance include: hsa-miR-145/E2F3, hsa-miR-139-5p/TOP2A, and hsa-miR-133a/GCLC. We also identified sets of positively correlated microRNA/mRNA pairs that are most likely result from indirect regulatory mechanisms. Our findings identify

MicroRNA profiling reveals dysregulated microRNAs and their target gene regulatory networks in cemento-ossifying fibroma.

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Pereira, Thaís Dos Santos Fontes; Brito, João Artur Ricieri; Guimarães, André Luiz Sena; Gomes, Carolina Cavaliéri; de Lacerda, Júlio Cesar Tanos; de Castro, Wagner Henriques; Coimbra, Roney Santos; Diniz, Marina Gonçalves; Gomez, Ricardo Santiago

2018-01-01

Cemento-ossifying fibroma (COF) is a benign fibro-osseous neoplasm of uncertain pathogenesis, and its treatment results in morbidity. MicroRNAs (miRNA) are small non-coding RNAs that regulate gene expression and may represent therapeutic targets. The purpose of the study was to generate a comprehensive miRNA profile of COF compared to normal bone. Additionally, the most relevant pathways and target genes of differentially expressed miRNA were investigated by in silico analysis. Nine COF and ten normal bone samples were included in the study. miRNA profiling was carried out by using TaqMan® OpenArray® Human microRNA panel containing 754 validated human miRNAs. We identified the most relevant miRNAs target genes through the leader gene approach, using STRING and Cytoscape software. Pathways enrichment analysis was performed using DIANA-miRPath. Eleven miRNAs were downregulated (hsa-miR-95-3p, hsa-miR-141-3p, hsa-miR-205-5p, hsa-miR-223-3p, hsa-miR-31-5p, hsa-miR-944, hsa-miR-200b-3p, hsa-miR-135b-5p, hsa-miR-31-3p, hsa-miR-223-5p and hsa-miR-200c-3p), and five were upregulated (hsa-miR-181a-5p, hsa-miR-181c-5p, hsa-miR-149-5p, hsa-miR-138-5p and hsa-miR-199a-3p) in COF compared to normal bone. Eighteen common target genes were predicted, and the leader genes approach identified the following genes involved in human COF: EZH2, XIAP, MET and TGFBR1. According to the biology of bone and COF, the most relevant KEGG pathways revealed by enrichment analysis were proteoglycans in cancer, miRNAs in cancer, pathways in cancer, p53-, PI3K-Akt-, FoxO- and TGF-beta signalling pathways, which were previously found to be differentially regulated in bone neoplasms, odontogenic tumours and osteogenesis. miRNA dysregulation occurs in COF, and EZH2, XIAP, MET and TGFBR1 are potential targets for functional analysis validation. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Genome-wide annotation of porcine microRNA genes and transcriptome profiling during Actinobacillus infection

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Nielsen, Mathilde

MicroRNAs are small single stranded non-coding RNA molecules which contributes to the regulation of gene expression by primarily binding to the 3´end of protein coding mRNA, hereby inhibiting the translation process or promting degradation of the mRNA. The main focus of this PhD project was to ex......MicroRNAs are small single stranded non-coding RNA molecules which contributes to the regulation of gene expression by primarily binding to the 3´end of protein coding mRNA, hereby inhibiting the translation process or promting degradation of the mRNA. The main focus of this PhD project...

Early and late effects of prenatal corticosteroid treatment on the microRNA profiles of lung tissue in rats

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YU, HONG-REN; LI, SUNG-CHOU; TSENG, WAN-NING; TAIN, YOU-LIN; CHEN, CHIH-CHENG; SHEEN, JIUNN-MING; TIAO, MAO-MENG; KUO, HO-CHANG; HUANG, CHAO-CHENG; HSIEH, KAI-SHENG; HUANG, LI-TUNG

2016-01-01

Glucocorticoids have been administered to mothers at risk of premature delivery to induce maturation of preterm fetal lungs and prevent the development of respiratory distress syndrome. Micro (mi)RNAs serve various crucial functions in cell proliferation, differentiation and organ development; however, few studies have demonstrated an association between miRNAs and lung development. The aim of the present study was to investigate alterations in the miRNA profiles of rat lung tissue following prenatal glucocorticoid therapy for fetal lung development. The differences in miRNA expression profiles were compared between postnatal days 7 (D7) and 120 (D120) rat lung tissues, followed by validation using reverse transcription-quantitative polymerase chain reaction. The miRNA profiles of rat lung tissues following prenatal dexamethasone (DEX) therapy were also investigated. miRNAs with 2-fold changes were selected for further analysis. At D120, 6 upregulated and 6 downregulated miRNAs were detected, compared with D7. Among these differentially expressed miRNAs, miR-101-3p and miR-99b-5p were associated with the lowest and highest expressions of miRNA at D7, respectively. A limited impact on the miRNA profiles of rat lung tissues was observed following prenatal DEX treatment, which may help to further clarify the mechanisms underlying normal lung development. However, the results of the present study cannot entirely elucidate the effects of prenatal DEX treatment on the lung development of premature infants, and further studies investigating the impact of prenatal corticosteroids on fetal lung miRNA profiles are required. PMID:26997989

MirZ: an integrated microRNA expression atlas and target prediction resource.

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Hausser, Jean; Berninger, Philipp; Rodak, Christoph; Jantscher, Yvonne; Wirth, Stefan; Zavolan, Mihaela

2009-07-01

MicroRNAs (miRNAs) are short RNAs that act as guides for the degradation and translational repression of protein-coding mRNAs. A large body of work showed that miRNAs are involved in the regulation of a broad range of biological functions, from development to cardiac and immune system function, to metabolism, to cancer. For most of the over 500 miRNAs that are encoded in the human genome the functions still remain to be uncovered. Identifying miRNAs whose expression changes between cell types or between normal and pathological conditions is an important step towards characterizing their function as is the prediction of mRNAs that could be targeted by these miRNAs. To provide the community the possibility of exploring interactively miRNA expression patterns and the candidate targets of miRNAs in an integrated environment, we developed the MirZ web server, which is accessible at www.mirz.unibas.ch. The server provides experimental and computational biologists with statistical analysis and data mining tools operating on up-to-date databases of sequencing-based miRNA expression profiles and of predicted miRNA target sites in species ranging from Caenorhabditis elegans to Homo sapiens.

Environmental contaminants and microRNA regulation: Transcription factors as regulators of toxicant-altered microRNA expression

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Sollome, James; Martin, Elizabeth [Department of Environmental Science & Engineering, Gillings School of Global Public Health, University of North Carolina, Chapel Hill (United States); Sethupathy, Praveen [Department of Genetics, School of Medicine, University of North Carolina, Chapel Hill, NC (United States); Fry, Rebecca C., E-mail: rfry@unc.edu [Department of Environmental Science & Engineering, Gillings School of Global Public Health, University of North Carolina, Chapel Hill (United States); Curriculum in Toxicology, School of Medicine, University of North Carolina, Chapel Hill, NC (United States)

2016-12-01

MicroRNAs (miRNAs) regulate gene expression by binding mRNA and inhibiting translation and/or inducing degradation of the associated transcripts. Expression levels of miRNAs have been shown to be altered in response to environmental toxicants, thus impacting cellular function and influencing disease risk. Transcription factors (TFs) are known to be altered in response to environmental toxicants and play a critical role in the regulation of miRNA expression. To date, environmentally-responsive TFs that are important for regulating miRNAs remain understudied. In a state-of-the-art analysis, we utilized an in silico bioinformatic approach to characterize potential transcriptional regulators of environmentally-responsive miRNAs. Using the miRStart database, genomic sequences of promoter regions for all available human miRNAs (n = 847) were identified and promoter regions were defined as − 1000/+500 base pairs from the transcription start site. Subsequently, the promoter region sequences of environmentally-responsive miRNAs (n = 128) were analyzed using enrichment analysis to determine overrepresented TF binding sites (TFBS). While most (56/73) TFs differed across environmental contaminants, a set of 17 TFs was enriched for promoter binding among miRNAs responsive to numerous environmental contaminants. Of these, one TF was common to miRNAs altered by the majority of environmental contaminants, namely SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily A, member 3 (SMARCA3). These identified TFs represent candidate common transcriptional regulators of miRNAs perturbed by environmental toxicants. - Highlights: • Transcription factors that regulate environmentally-modulated miRNA expression are understudied • Transcription factor binding sites (TFBS) located within DNA promoter regions of miRNAs were identified. • Specific transcription factors may serve as master regulators of environmentally-mediated microRNA expression.

Environmental contaminants and microRNA regulation: Transcription factors as regulators of toxicant-altered microRNA expression

International Nuclear Information System (INIS)

Sollome, James; Martin, Elizabeth; Sethupathy, Praveen; Fry, Rebecca C.

2016-01-01

MicroRNAs (miRNAs) regulate gene expression by binding mRNA and inhibiting translation and/or inducing degradation of the associated transcripts. Expression levels of miRNAs have been shown to be altered in response to environmental toxicants, thus impacting cellular function and influencing disease risk. Transcription factors (TFs) are known to be altered in response to environmental toxicants and play a critical role in the regulation of miRNA expression. To date, environmentally-responsive TFs that are important for regulating miRNAs remain understudied. In a state-of-the-art analysis, we utilized an in silico bioinformatic approach to characterize potential transcriptional regulators of environmentally-responsive miRNAs. Using the miRStart database, genomic sequences of promoter regions for all available human miRNAs (n = 847) were identified and promoter regions were defined as − 1000/+500 base pairs from the transcription start site. Subsequently, the promoter region sequences of environmentally-responsive miRNAs (n = 128) were analyzed using enrichment analysis to determine overrepresented TF binding sites (TFBS). While most (56/73) TFs differed across environmental contaminants, a set of 17 TFs was enriched for promoter binding among miRNAs responsive to numerous environmental contaminants. Of these, one TF was common to miRNAs altered by the majority of environmental contaminants, namely SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily A, member 3 (SMARCA3). These identified TFs represent candidate common transcriptional regulators of miRNAs perturbed by environmental toxicants. - Highlights: • Transcription factors that regulate environmentally-modulated miRNA expression are understudied • Transcription factor binding sites (TFBS) located within DNA promoter regions of miRNAs were identified. • Specific transcription factors may serve as master regulators of environmentally-mediated microRNA expression

Zika virus alters the microRNA expression profile and elicits an RNAi response in Aedes aegypti mosquitoes.

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Miguel A Saldaña

2017-07-01

Full Text Available Zika virus (ZIKV, a flavivirus transmitted primarily by Aedes aegypti, has recently spread globally in an unprecedented fashion, yet we have a poor understanding of host-microbe interactions in this system. To gain insights into the interplay between ZIKV and the mosquito, we sequenced the small RNA profiles in ZIKV-infected and non-infected Ae. aegypti mosquitoes at 2, 7 and 14 days post-infection. ZIKA induced an RNAi response in the mosquito with virus-derived short interfering RNAs and PIWI-interacting RNAs dramatically increased in abundance post-infection. Further, we found 17 host microRNAs (miRNAs that were modulated by ZIKV infection at all time points. Strikingly, many of these regulated miRNAs have been reported to have their expression altered by dengue and West Nile viruses, while the response was divergent from that induced by the alphavirus Chikungunya virus in mosquitoes. This suggests that conserved miRNA responses occur within mosquitoes in response to flavivirus infection. This study expands our understanding of ZIKV-vector interactions and provides potential avenues to be further investigated to target ZIKV in the mosquito host.

MicroRNA and piRNA profiles in normal human testis detected by next generation sequencing.

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Qingling Yang

Full Text Available BACKGROUND: MicroRNAs (miRNAs are the class of small endogenous RNAs that play an important regulatory role in cells by negatively affecting gene expression at transcriptional and post-transcriptional levels. There have been extensive studies aiming to discover miRNAs and to analyze their functions in the cells from a variety of species. However, there are no published studies of miRNA profiles in human testis using next generation sequencing (NGS technology. RESULTS: We employed Solexa sequencing technology to profile miRNAs in normal human testis. Total 770 known and 5 novel human miRNAs, and 20121 piRNAs were detected, indicating that the human testis has a complex population of small RNAs. The expression of 15 known and 5 novel detected miRNAs was validated by qRT-PCR. We have also predicted the potential target genes of the abundant known and novel miRNAs, and subjected them to GO and pathway analysis, revealing the involvement of miRNAs in many important biological phenomenon including meiosis and p53-related pathways that are implicated in the regulation of spermatogenesis. CONCLUSIONS: This study reports the first genome-wide miRNA profiles in human testis using a NGS approach. The presence of large number of miRNAs and the nature of their target genes suggested that miRNAs play important roles in spermatogenesis. Here we provide a useful resource for further elucidation of the regulatory role of miRNAs and piRNAs in the spermatogenesis. It may also facilitate the development of prophylactic strategies for male infertility.

Circulating Extracellular microRNA in Systemic Autoimmunity

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Heegaard, Niels H. H.; Carlsen, Anting Liu; Skovgaard, Kerstin

2015-01-01

killer cells, neutrophil granulocytes, and monocyte-macrophages. Exploratory studies (only validated in a few cases) also show that specific profiles of circulating miRNAs are associated with different systemic autoimmune diseases including systemic lupus erythematosus (SLE), systemic sclerosis......, extracellular miRNA is protected against degradation by complexation with carrier proteins and/or by being enclosed in subcellular membrane vesicles. This, together with their tissue- and disease-specific expression, has fuelled the interest in using circulating microRNA profiles as harbingers of disease, i.......e., as diagnostic analytes and as clues to dysregulated pathways in disease. Many studies show that inflammation and immune dysregulation, e.g., in autoimmune diseases, are associated with distinct miRNA expression changes in targeted tissues and in innate and adaptive immunity cells such as lymphocytes, natural...

Microarray Analysis of microRNA Expression during Axolotl Limb Regeneration

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Holman, Edna C.; Campbell, Leah J.; Hines, John; Crews, Craig M.

2012-01-01

Among vertebrates, salamanders stand out for their remarkable capacity to quickly regrow a myriad of tissues and organs after injury or amputation. The limb regeneration process in axolotls (Ambystoma mexicanum) has been well studied for decades at the cell-tissue level. While several developmental genes are known to be reactivated during this epimorphic process, less is known about the role of microRNAs in urodele amphibian limb regeneration. Given the compelling evidence that many microRNAs tightly regulate cell fate and morphogenetic processes through development and adulthood by modulating the expression (or re-expression) of developmental genes, we investigated the possibility that microRNA levels change during limb regeneration. Using two different microarray platforms to compare the axolotl microRNA expression between mid-bud limb regenerating blastemas and non-regenerating stump tissues, we found that miR-21 was overexpressed in mid-bud blastemas compared to stump tissue. Mature A. mexicanum (“Amex”) miR-21 was detected in axolotl RNA by Northern blot and differential expression of Amex-miR-21 in blastema versus stump was confirmed by quantitative RT-PCR. We identified the Amex Jagged1 as a putative target gene for miR-21 during salamander limb regeneration. We cloned the full length 3′UTR of Amex-Jag1, and our in vitro assays demonstrated that its single miR-21 target recognition site is functional and essential for the response of the Jagged1 gene to miR-21 levels. Our findings pave the road for advanced in vivo functional assays aimed to clarify how microRNAs such as miR-21, often linked to pathogenic cell growth, might be modulating the redeployment of developmental genes such as Jagged1 during regenerative processes. PMID:23028429

Microarray analysis of microRNA expression during axolotl limb regeneration.

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Edna C Holman

Full Text Available Among vertebrates, salamanders stand out for their remarkable capacity to quickly regrow a myriad of tissues and organs after injury or amputation. The limb regeneration process in axolotls (Ambystoma mexicanum has been well studied for decades at the cell-tissue level. While several developmental genes are known to be reactivated during this epimorphic process, less is known about the role of microRNAs in urodele amphibian limb regeneration. Given the compelling evidence that many microRNAs tightly regulate cell fate and morphogenetic processes through development and adulthood by modulating the expression (or re-expression of developmental genes, we investigated the possibility that microRNA levels change during limb regeneration. Using two different microarray platforms to compare the axolotl microRNA expression between mid-bud limb regenerating blastemas and non-regenerating stump tissues, we found that miR-21 was overexpressed in mid-bud blastemas compared to stump tissue. Mature A. mexicanum ("Amex" miR-21 was detected in axolotl RNA by Northern blot and differential expression of Amex-miR-21 in blastema versus stump was confirmed by quantitative RT-PCR. We identified the Amex Jagged1 as a putative target gene for miR-21 during salamander limb regeneration. We cloned the full length 3'UTR of Amex-Jag1, and our in vitro assays demonstrated that its single miR-21 target recognition site is functional and essential for the response of the Jagged1 gene to miR-21 levels. Our findings pave the road for advanced in vivo functional assays aimed to clarify how microRNAs such as miR-21, often linked to pathogenic cell growth, might be modulating the redeployment of developmental genes such as Jagged1 during regenerative processes.

Profile of microRNA in Giant Panda Blood: A Resource for Immune-Related and Novel microRNAs.

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Mingyu Yang

Full Text Available The giant panda (Ailuropoda melanoleuca is one of the world's most beloved endangered mammals. Although the draft genome of this species had been assembled, little was known about the composition of its microRNAs (miRNAs or their functional profiles. Recent studies demonstrated that changes in the expression of miRNAs are associated with immunity. In this study, miRNAs were extracted from the blood of four healthy giant pandas and sequenced by Illumina next generation sequencing technology. As determined by miRNA screening, a total of 276 conserved miRNAs and 51 novel putative miRNAs candidates were detected. After differential expression analysis, we noticed that the expressions of 7 miRNAs were significantly up-regulated in young giant pandas compared with that of adults. Moreover, 2 miRNAs were up-regulated in female giant pandas and 1 in the male individuals. Target gene prediction suggested that the miRNAs of giant panda might be relevant to the expressions of 4,602 downstream genes. Subseuqently, the predicted target genes were conducted to KEGG (Kyoto Encyclopedia of Genes and Genomes pathway analysis and we found that these genes were mainly involved in host immunity, including the Ras signaling pathway, the PI3K-Akt signaling pathway, and the MAPK signaling pathway. In conclusion, our results provide the first miRNA profiles of giant panda blood, and the predicted functional analyses may open an avenue for further study of giant panda immunity.

Profile of microRNA in Giant Panda Blood: A Resource for Immune-Related and Novel microRNAs.

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Yang, Mingyu; Du, Lianming; Li, Wujiao; Shen, Fujun; Fan, Zhenxin; Jian, Zuoyi; Hou, Rong; Shen, Yongmei; Yue, Bisong; Zhang, Xiuyue

2015-01-01

The giant panda (Ailuropoda melanoleuca) is one of the world's most beloved endangered mammals. Although the draft genome of this species had been assembled, little was known about the composition of its microRNAs (miRNAs) or their functional profiles. Recent studies demonstrated that changes in the expression of miRNAs are associated with immunity. In this study, miRNAs were extracted from the blood of four healthy giant pandas and sequenced by Illumina next generation sequencing technology. As determined by miRNA screening, a total of 276 conserved miRNAs and 51 novel putative miRNAs candidates were detected. After differential expression analysis, we noticed that the expressions of 7 miRNAs were significantly up-regulated in young giant pandas compared with that of adults. Moreover, 2 miRNAs were up-regulated in female giant pandas and 1 in the male individuals. Target gene prediction suggested that the miRNAs of giant panda might be relevant to the expressions of 4,602 downstream genes. Subseuqently, the predicted target genes were conducted to KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis and we found that these genes were mainly involved in host immunity, including the Ras signaling pathway, the PI3K-Akt signaling pathway, and the MAPK signaling pathway. In conclusion, our results provide the first miRNA profiles of giant panda blood, and the predicted functional analyses may open an avenue for further study of giant panda immunity.

A Customized Quantitative PCR MicroRNA Panel Provides a Technically Robust Context for Studying Neurodegenerative Disease Biomarkers and Indicates a High Correlation Between Cerebrospinal Fluid and Choroid Plexus MicroRNA Expression.

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Wang, Wang-Xia; Fardo, David W; Jicha, Gregory A; Nelson, Peter T

2017-12-01

MicroRNA (miRNA) expression varies in association with different tissue types and in diseases. Having been found in body fluids including blood and cerebrospinal fluid (CSF), miRNAs constitute potential biomarkers. CSF miRNAs have been proposed as biomarkers for neurodegenerative diseases; however, there is a lack of consensus about the best candidate miRNA biomarkers and there has been variability in results from different research centers, perhaps due to technical factors. Here, we sought to optimize technical parameters for CSF miRNA studies. We examined different RNA isolation methods and performed miRNA expression profiling with TaqMan® miRNA Arrays. More specifically, we developed a customized CSF-miRNA low-density array (TLDA) panel that contains 47 targets: miRNAs shown previously to be relevant to neurodegenerative disease, miRNAs that are abundant in CSF, data normalizers, and controls for potential blood and tissue contamination. The advantages of using this CSF-miRNA TLDA panel include specificity, sensitivity, fast processing and data analysis, and cost effectiveness. We optimized technical parameters for this assay. Further, the TLDA panel can be tailored to other specific purposes. We tested whether the profile of miRNAs in the CSF resembled miRNAs isolated from brain tissue (hippocampus or cerebellum), blood, or the choroid plexus. We found that the CSF miRNA expression profile most closely resembles that of choroid plexus tissue, underscoring the potential importance of choroid plexus-derived signaling through CSF miRNAs. In summary, the TLDA miRNA array panel will enable evaluation and discovery of CSF miRNA biomarkers and can potentially be utilized in clinical diagnosis and disease stage monitoring.

Dehydration triggers differential microRNA expression in Xenopus laevis brain.

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Luu, Bryan E; Storey, Kenneth B

2015-11-15

African clawed frogs, Xenopus laevis, although primarily aquatic, have a high tolerance for dehydration, being capable of withstanding the loss of up to 32-35% of total water body water. Recent studies have shown that microRNAs play a role in the response to dehydration by the liver, kidney and ventral skin of X. laevis. MicroRNAs act by modulating the expression of mRNA transcripts, thereby affecting diverse biochemical pathways. In this study, 43 microRNAs were assessed in frog brains comparing control and dehydrated (31.2±0.83% of total body water lost) conditions. MicroRNAs of interest were measured using a modified protocol which employs polyadenylation of microRNAs prior to reverse transcription and qPCR. Twelve microRNAs that showed a significant decrease in expression (to 41-77% of control levels) in brains from dehydrated frogs (xla-miR-15a, -150, -181a, -191, -211, -218, -219b, -30c, -30e, -31, -34a, and -34b) were identified. Genomic analysis showed that the sequences of these dehydration-responsive microRNAs were highly conserved as compared with the comparable microRNAs of mice (91-100%). Suppression of these microRNAs implies that translation of the mRNA transcripts under their control could be enhanced in response to dehydration. Bioinformatic analysis using the DIANA miRPath program (v.2.0) predicted the top two KEGG pathways that these microRNAs collectively regulate: 1. Axon guidance, and 2. Long-term potentiation. Previous studies indicated that suppression of these microRNAs promotes neuroprotective pathways by increasing the expression of brain-derived neurotrophic factor and activating anti-apoptotic pathways. This suggests that similar actions may be triggered in X. laevis brains as a protective response to dehydration. Crown Copyright © 2015. Published by Elsevier B.V. All rights reserved.

Time-Dependent Expression Profiles of microRNAs and mRNAs in Rat Milk Whey

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Izumi, Hirohisa; Kosaka, Nobuyoshi; Shimizu, Takashi; Sekine, Kazunori; Ochiya, Takahiro; Takase, Mitsunori

2014-01-01

Functional RNAs, such as microRNA (miRNA) and mRNA, are present in milk, but their roles are unknown. To clarify the roles of milk RNAs, further studies using experimental animals such as rats are needed. However, it is unclear whether rat milk also contains functional RNAs and what their time dependent expression profiles are. Thus, we prepared total RNA from whey isolated from rat milk collected on days 2, 9, and 16 postpartum and analyzed using microarrays and quantitative PCR. The concentration of RNA in colostrum whey (day 2) was markedly higher than that in mature milk whey (days 9 and 16). Microarray analysis detected 161 miRNAs and 10,948 mRNA transcripts. Most of the miRNAs and mRNA transcripts were common to all tested milks. Finally, we selected some immune- and development-related miRNAs and mRNAs, and analysed them by quantitative PCR (in equal sample volumes) to determine their time-dependent changes in expression in detail. Some were significantly more highly expressed in colostrum whey than in mature milk whey, but some were expressed equally. And mRNA expression levels of some cytokines and hormones did not reflect the protein levels. It is still unknown whether RNAs in milk play biological roles in neonates. However, our data will help guide future in vivo studies using experimental animals such as rats. PMID:24533154

High Throughput qPCR Expression Profiling of Circulating MicroRNAs Reveals Minimal Sex- and Sample Timing-Related Variation in Plasma of Healthy Volunteers.

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Catherine Mooney

Full Text Available MicroRNAs are a class of small non-coding RNA that regulate gene expression at a post-transcriptional level. MicroRNAs have been identified in various body fluids under normal conditions and their stability as well as their dysregulation in disease opens up a new field for biomarker study. However, diurnal and day-to-day variation in plasma microRNA levels, and differential regulation between males and females, may affect biomarker stability. A QuantStudio 12K Flex Real-Time PCR System was used to profile plasma microRNA levels using OpenArray in male and female healthy volunteers, in the morning and afternoon, and at four time points over a one month period. Using this system we were able to run four OpenArray plates in a single run, the equivalent of 32 traditional 384-well qPCR plates or 12,000 data points. Up to 754 microRNAs can be identified in a single plasma sample in under two hours. 108 individual microRNAs were identified in at least 80% of all our samples which compares favourably with other reports of microRNA profiles in serum or plasma in healthy adults. Many of these microRNAs, including miR-16-5p, miR-17-5p, miR-19a-3p, miR-24-3p, miR-30c-5p, miR-191-5p, miR-223-3p and miR-451a are highly expressed and consistent with previous studies using other platforms. Overall, microRNA levels were very consistent between individuals, males and females, and time points and we did not detect significant differences in levels of microRNAs. These results suggest the suitability of this platform for microRNA profiling and biomarker discovery and suggest minimal confounding influence of sex or sample timing. However, the platform has not been subjected to rigorous validation which must be demonstrated in future biomarker studies where large differences may exist between disease and control samples.

Comparison of microarray platforms for measuring differential microRNA expression in paired normal/cancer colon tissues.

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Maurizio Callari

Full Text Available BACKGROUND: Microarray technology applied to microRNA (miRNA profiling is a promising tool in many research fields; nevertheless, independent studies characterizing the same pathology have often reported poorly overlapping results. miRNA analysis methods have only recently been systematically compared but only in few cases using clinical samples. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the inter-platform reproducibility of four miRNA microarray platforms (Agilent, Exiqon, Illumina, and Miltenyi, comparing nine paired tumor/normal colon tissues. The most concordant and selected discordant miRNAs were further studied by quantitative RT-PCR. Globally, a poor overlap among differentially expressed miRNAs identified by each platform was found. Nevertheless, for eight miRNAs high agreement in differential expression among the four platforms and comparability to qRT-PCR was observed. Furthermore, most of the miRNA sets identified by each platform are coherently enriched in data from the other platforms and the great majority of colon cancer associated miRNA sets derived from the literature were validated in our data, independently from the platform. Computational integration of miRNA and gene expression profiles suggested that anti-correlated predicted target genes of differentially expressed miRNAs are commonly enriched in cancer-related pathways and in genes involved in glycolysis and nutrient transport. CONCLUSIONS: Technical and analytical challenges in measuring miRNAs still remain and further research is required in order to increase consistency between different microarray-based methodologies. However, a better inter-platform agreement was found by looking at miRNA sets instead of single miRNAs and through a miRNAs - gene expression integration approach.

Research resources: comparative microRNA profiles in human corona radiata cells and cumulus oophorus cells detected by next-generation small RNA sequencing.

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Xian-Hong Tong

Full Text Available During folliculogenesis, cumulus cells surrounding the oocyte differentiate into corona radiata cells (CRCs and cumulus oophorus cells (COCs, which are involved in gonadal steroidogenesis and the development of germ cells. Several studies suggested that microRNAs (miRNAs play an important regulatory role at the post-transcriptional level in cumulus cells. However, comparative miRNA profiles and associated processes in human CRCs and COCs have not been reported before. In this study, miRNA profiles were obtained from CRCs and COCs using next generation sequencing in women undergoing controlled ovarian stimulation for IVF. A total of 785 and 799 annotated miRNAs were identified in CRCs and COCs, while high expression levels of six novel miRNAs were detected both in CRCs and in COCs. In addition, different expression patterns in CRCs and COCs were detected in 72 annotated miRNAs. To confirm the miRNA profile in COCs and CRCs, quantitative real-time PCR was used to validate the expression of annotated miRNAs, differentially expressed miRNAs, and novel miRNAs. The miRNAs in the let-7 family were found to be involved in the regulation of a broad range of biological processes in both cumulus cell populations, which was accompanied by a large amount of miRNA editing. Bioinformatics analysis showed that amino acid and energy metabolism were targeted significantly by miRNAs that were differentially expressed between CRCs and COCs. Our work extends the current knowledge of the regulatory role of miRNAs and their targeted pathways in folliculogenesis, and provides novel candidates for molecular biomarkers in the research of female infertility.

Chimira: analysis of small RNA sequencing data and microRNA modifications.

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Vitsios, Dimitrios M; Enright, Anton J

2015-10-15

Chimira is a web-based system for microRNA (miRNA) analysis from small RNA-Seq data. Sequences are automatically cleaned, trimmed, size selected and mapped directly to miRNA hairpin sequences. This generates count-based miRNA expression data for subsequent statistical analysis. Moreover, it is capable of identifying epi-transcriptomic modifications in the input sequences. Supported modification types include multiple types of 3'-modifications (e.g. uridylation, adenylation), 5'-modifications and also internal modifications or variation (ADAR editing or single nucleotide polymorphisms). Besides cleaning and mapping of input sequences to miRNAs, Chimira provides a simple and intuitive set of tools for the analysis and interpretation of the results (see also Supplementary Material). These allow the visual study of the differential expression between two specific samples or sets of samples, the identification of the most highly expressed miRNAs within sample pairs (or sets of samples) and also the projection of the modification profile for specific miRNAs across all samples. Other tools have already been published in the past for various types of small RNA-Seq analysis, such as UEA workbench, seqBuster, MAGI, OASIS and CAP-miRSeq, CPSS for modifications identification. A comprehensive comparison of Chimira with each of these tools is provided in the Supplementary Material. Chimira outperforms all of these tools in total execution speed and aims to facilitate simple, fast and reliable analysis of small RNA-Seq data allowing also, for the first time, identification of global microRNA modification profiles in a simple intuitive interface. Chimira has been developed as a web application and it is accessible here: http://www.ebi.ac.uk/research/enright/software/chimira. aje@ebi.ac.uk Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press.

Large-scale identification of microRNA targets in murine Dgcr8-deficient embryonic stem cell lines.

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Matthew P A Davis

Full Text Available Small RNAs such as microRNAs play important roles in embryonic stem cell maintenance and differentiation. A broad range of microRNAs is expressed in embryonic stem cells while only a fraction of their targets have been identified. We have performed large-scale identification of embryonic stem cell microRNA targets using a murine embryonic stem cell line deficient in the expression of Dgcr8. These cells are heavily depleted for microRNAs, allowing us to reintroduce specific microRNA duplexes and identify refined target sets. We used deep sequencing of small RNAs, mRNA expression profiling and bioinformatics analysis of microRNA seed matches in 3' UTRs to identify target transcripts. Consequently, we have identified a network of microRNAs that converge on the regulation of several important cellular pathways. Additionally, our experiments have revealed a novel candidate for Dgcr8-independent microRNA genesis and highlighted the challenges currently facing miRNA annotation.

Role of Melt Curve Analysis in Interpretation of Nutrigenomics' MicroRNA Expression Data.

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Ahmed, Farid E; Gouda, Mostafa M; Hussein, Laila A; Ahmed, Nancy C; Vos, Paul W; Mohammad, Mahmoud A

2017-01-01

This article illustrates the importance of melt curve analysis (MCA) in interpretation of mild nutrogenomic micro(mi)RNA expression data, by measuring the magnitude of the expression of key miRNA molecules in stool of healthy human adults as molecular markers, following the intake of Pomegranate juice (PGJ), functional fermented sobya (FS), rich in potential probiotic lactobacilli, or their combination. Total small RNA was isolated from stool of 25 volunteers before and following a three-week dietary intervention trial. Expression of 88 miRNA genes was evaluated using Qiagen's 96 well plate RT 2 miRNA qPCR arrays. Employing parallel coordinates plots, there was no observed significant separation for the gene expression (Cq) values, using Roche 480® PCR LightCycler instrument used in this study, and none of the miRNAs showed significant statistical expression after controlling for the false discovery rate. On the other hand, melting temperature profiles produced during PCR amplification run, found seven significant genes (miR-184, miR-203, miR-373, miR-124, miR-96, miR-373 and miR-301a), which separated candidate miRNAs that could function as novel molecular markers of relevance to oxidative stress and immunoglobulin function, for the intake of polyphenol (PP)-rich, functional fermented foods rich in lactobacilli (FS), or their combination. We elaborate on these data, and present a detailed review on use of melt curves for analyzing nutigenomic miRNA expression data, which initially appear to show no significant expressions, but are actually more subtle than this simplistic view, necessitating the understanding of the role of MCA for a comprehensive understanding of what the collective expression and MCA data collectively imply. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

RBiomirGS: an all-in-one miRNA gene set analysis solution featuring target mRNA mapping and expression profile integration

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Jing Zhang

2018-01-01

Full Text Available Background With the continuous discovery of microRNA’s (miRNA association with a wide range of biological and cellular processes, expression profile-based functional characterization of such post-transcriptional regulation is crucial for revealing its significance behind particular phenotypes. Profound advancement in bioinformatics has been made to enable in depth investigation of miRNA’s role in regulating cellular and molecular events, resulting in a huge quantity of software packages covering different aspects of miRNA functional analysis. Therefore, an all-in-one software solution is in demand for a comprehensive yet highly efficient workflow. Here we present RBiomirGS, an R package for a miRNA gene set (GS analysis. Methods The package utilizes multiple databases for target mRNA mapping, estimates miRNA effect on the target mRNAs through miRNA expression profile and conducts a logistic regression-based GS enrichment. Additionally, human ortholog Entrez ID conversion functionality is included for target mRNAs. Results By incorporating all the core steps into one package, RBiomirGS eliminates the need for switching between different software packages. The modular structure of RBiomirGS enables various access points to the analysis, with which users can choose the most relevant functionalities for their workflow. Conclusions With RBiomirGS, users are able to assess the functional significance of the miRNA expression profile under the corresponding experimental condition by minimal input and intervention. Accordingly, RBiomirGS encompasses an all-in-one solution for miRNA GS analysis. RBiomirGS is available on GitHub (http://github.com/jzhangc/RBiomirGS. More information including instruction and examples can be found on website (http://kenstoreylab.com/?page_id=2865.

Plasma and White Blood Cells Show Different miRNA Expression Profiles in Parkinson's Disease.

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Schwienbacher, Christine; Foco, Luisa; Picard, Anne; Corradi, Eloina; Serafin, Alice; Panzer, Jörg; Zanigni, Stefano; Blankenburg, Hagen; Facheris, Maurizio F; Giannini, Giulia; Falla, Marika; Cortelli, Pietro; Pramstaller, Peter P; Hicks, Andrew A

2017-06-01

Parkinson's disease (PD) diagnosis is based on the assessment of motor symptoms, which manifest when more than 50% of dopaminergic neurons are degenerated. To date, no validated biomarkers are available for the diagnosis of PD. The aims of the present study are to evaluate whether plasma and white blood cells (WBCs) are interchangeable biomarker sources and to identify circulating plasma-based microRNA (miRNA) biomarkers for an early detection of PD. We profiled plasma miRNA levels in 99 L-dopa-treated PD patients from two independent data collections, in ten drug-naïve PD patients, and in unaffected controls matched by sex and age. We evaluated expression levels by reverse transcription and quantitative real-time PCR (RT-qPCR) and combined the results from treated PD patients using a fixed effect inverse-variance weighted meta-analysis. We revealed different expression profiles comparing plasma and WBCs and drug-naïve and L-dopa-treated PD patients. We observed an upregulation trend for miR-30a-5p in L-dopa-treated PD patients and investigated candidate target genes by integrated in silico analyses. We could not analyse miR-29b-3p, normally expressed in WBCs, due to the very low expression in plasma. We observed different expression profiles in WBCs and plasma, suggesting that they are both suitable but not interchangeable peripheral sources for biomarkers. We revealed miR-30a-5p as a potential biomarker for PD in plasma. In silico analyses suggest that miR-30a-5p might have a regulatory role in mitochondrial dynamics and autophagy. Further investigations are needed to confirm miR-30a-5p deregulation and targets and to investigate the influence of L-dopa treatment on miRNA expression levels.

Integrated microRNA and mRNA signatures associated with survival in triple negative breast cancer.

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Cascione, Luciano; Gasparini, Pierluigi; Lovat, Francesca; Carasi, Stefania; Pulvirenti, Alfredo; Ferro, Alfredo; Alder, Hansjuerg; He, Gang; Vecchione, Andrea; Croce, Carlo M; Shapiro, Charles L; Huebner, Kay

2013-01-01

Triple negative breast cancer (TNBC) is a heterogeneous disease at the molecular, pathologic and clinical levels. To stratify TNBCs, we determined microRNA (miRNA) expression profiles, as well as expression profiles of a cancer-focused mRNA panel, in tumor, adjacent non-tumor (normal) and lymph node metastatic lesion (mets) tissues, from 173 women with TNBCs; we linked specific miRNA signatures to patient survival and used miRNA/mRNA anti-correlations to identify clinically and genetically different TNBC subclasses. We also assessed miRNA signatures as potential regulators of TNBC subclass-specific gene expression networks defined by expression of canonical signal pathways.Tissue specific miRNAs and mRNAs were identified for normal vs tumor vs mets comparisons. miRNA signatures correlated with prognosis were identified and predicted anti-correlated targets within the mRNA profile were defined. Two miRNA signatures (miR-16, 155, 125b, 374a and miR-16, 125b, 374a, 374b, 421, 655, 497) predictive of overall survival (P = 0.05) and distant-disease free survival (P = 0.009), respectively, were identified for patients 50 yrs of age or younger. By multivariate analysis the risk signatures were independent predictors for overall survival and distant-disease free survival. mRNA expression profiling, using the cancer-focused mRNA panel, resulted in clustering of TNBCs into 4 molecular subclasses with different expression signatures anti-correlated with the prognostic miRNAs. Our findings suggest that miRNAs play a key role in triple negative breast cancer through their ability to regulate fundamental pathways such as: cellular growth and proliferation, cellular movement and migration, Extra Cellular Matrix degradation. The results define miRNA expression signatures that characterize and contribute to the phenotypic diversity of TNBC and its metastasis.

Characterization of MicroRNA Expression Profiles and Identification of Potential Biomarkers in Leprosy.

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Jorge, Karina T O S; Souza, Renan P; Assis, Marieta T A; Araújo, Marcelo G; Locati, Massimo; Jesus, Amélia M R; Dias Baptista, Ida M F; Lima, Cristiano X; Teixeira, Antônio L; Teixeira, Mauro M; Soriani, Frederico M

2017-05-01

Leprosy is an important cause of disability in the developing world. Early diagnosis is essential to allow for cure and to interrupt transmission of this infection. MicroRNAs (miRNAs) are important factors for host-pathogen interaction and they have been identified as biomarkers for various infectious diseases. The expression profile of 377 microRNAs were analyzed by TaqMan low-density array (TLDA) in skin lesions of tuberculoid and lepromatous leprosy patients as well as skin specimens from healthy controls. In a second step, 16 microRNAs were selected for validation experiments with reverse transcription-quantitative PCR (qRT-PCR) in skin samples from new individuals. Principal-component analysis followed by logistic regression model and receiver operating characteristic (ROC) curve analyses were performed to evaluate the diagnostic potential of selected miRNAs. Four patterns of differential expression were identified in the TLDA experiment, suggesting a diagnostic potential of miRNAs in leprosy. After validation experiments, a combination of four miRNAs (miR-101, miR-196b, miR-27b, and miR-29c) was revealed as able to discriminate between healthy control and leprosy patients with 80% sensitivity and 91% specificity. This set of miRNAs was also able to discriminate between lepromatous and tuberculoid patients with a sensitivity of 83% and 80% specificity. In this work, it was possible to identify a set of miRNAs with good diagnostic potential for leprosy. Copyright © 2017 American Society for Microbiology.

RNA-Seq reveals MicroRNA expression signature and genetic polymorphism associated with growth and muscle quality traits in rainbow trout

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The role of microRNA expression and genetic variation in microRNA-binding sites of target genes on growth and muscle quality traits is poorly characterized. We used RNA-Seq approach to investigate their importance on 5 growth and muscle quality traits: whole body weight (WBW), muscle yield, muscle c...

Identification of a Polyomavirus microRNA Highly Expressed in Tumors

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Chen, Chun Jung; Cox, Jennifer E.; Azarm, Kristopher; Wylie, Karen N.; Woolard, Kevin D.; Pesavento, Patricia A.; Sullivan, Christopher S.

2014-01-01

Polyomaviruses (PyVs) are associated with tumors including Merkel cell carcinoma (MCC). Several PyVs encode microRNAs (miRNAs) but to date no abundant PyV miRNAs have been reported in tumors. To better understand the function of the Merkel cell PyV (MCPyV) miRNA, we examined phylogenetically-related viruses for miRNA expression. We show that two primate PyVs and the more distantly-related raccoon PyV (RacPyV) encode miRNAs that share genomic position and partial sequence identity with MCPyV miRNAs. Unlike MCPyV miRNA in MCC, RacPyV miRNA is highly abundant in raccoon tumors. RacPyV miRNA negatively regulates reporters of early viral (T antigen) transcripts, yet robust viral miRNA expression is tolerated in tumors. We also identify raccoon miRNAs expressed in RacPyV-associated neuroglial brain tumors, including several likely oncogenic miRNAs (oncomiRs). This work describes the first PyV miRNA abundantly expressed in tumors and is consistent with a possible role for both host and viral miRNAs in RacPyV-associated tumors. PMID:25514573

MicroRNA and histopathological characterization of pure mucinous breast carcinoma

International Nuclear Information System (INIS)

Zhou, Feng; Li, Shuai; Meng, Hui-Min; Qi, Li-Qiang; Gu, Lin

2013-01-01

Pure mucinous breast carcinoma (PMBC) is an uncommon histological type of breast cancer characterized by a large amount of mucin production. MicroRNA (miRNA) is a large class of small noncoding RNA of about 22 nt involved in the regulation of various biological processes. This study aims to identify the miRNA expression profile in PMBC. MiRNA expression profiles in 11 PMBCs were analyzed by miRNA-microarray and real-time polymerase chain reaction (PCR). Thirty-one PMBCs and 27 invasive ductal carcinoma of no special types (IDC-NSTs) were assessed by immunohistochemistry using antibodies against ER, PR-progesterone receptor, HER2, Ki-67, Bcl-2, p53, PCNA, and CK5 and 6. We analyzed the miRNA expression in 11 PMBCs and corresponding normal tissues using miRNA-microarray and real-time PCR, and found that miR-143 and miR-224-5p were significantly downregulated in mucinous carcinoma tissue. Compared with IDC-NSTs, PMBC showed a significantly higher ER positive rate, lower HER-2 positive rate, and lower cell proliferation rates. To our knowledge, this is the first study to demonstrate the miRNA expression profile of PMBC, and our findings may lead to further understanding of this type of breast cancer

Impact of gastro-oesophageal reflux on microRNA expression, location and function.

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Smith, Cameron M; Michael, Michael Z; Watson, David I; Tan, Grace; Astill, David St J; Hummel, Richard; Hussey, Damian J

2013-01-08

Ulceration of the oesophageal squamous mucosa (ulcerative oesophagitis) is a pathological manifestation of gastro-oesophageal reflux disease, and is a major risk factor for the development of Barrett's oesophagus. Barrett's oesophagus is characterised by replacement of reflux-damaged oesophageal squamous epithelium with a columnar intestinal-like epithelium. We previously reported discovery of microRNAs that are differentially expressed between oesophageal squamous mucosa and Barrett's oesophagus mucosa. Now, to better understand early steps in the initiation of Barrett's oesophagus, we assessed the expression, location and function of these microRNAs in oesophageal squamous mucosa from individuals with ulcerative oesophagitis. Quantitative real-time PCR was used to compare miR-21, 143, 145, 194, 203, 205 and 215 expression levels in oesophageal mucosa from individuals without pathological gastro-oesophageal reflux to individuals with ulcerative oesophagitis. Correlations between microRNA expression and messenger RNA differentiation markers BMP-4, CK8 and CK14 were analyzed. The cellular localisation of microRNAs within the oesophageal mucosa was determined using in-situ hybridisation. microRNA involvement in proliferation and apoptosis was assessed following transfection of a human squamous oesophageal mucosal cell line (Het-1A). miR-143, miR-145 and miR-205 levels were significantly higher in gastro-oesophageal reflux compared with controls. Elevated miR-143 expression correlated with BMP-4 and CK8 expression, and elevated miR-205 expression correlated negatively with CK14 expression. Endogenous miR-143, miR-145 and miR-205 expression was localised to the basal layer of the oesophageal epithelium. Transfection of miR-143, 145 and 205 mimics into Het-1A cells resulted in increased apoptosis and decreased proliferation. Elevated miR-143, miR-145 and miR-205 expression was observed in oesophageal squamous mucosa of individuals with ulcerative oesophagitis. These mi

How the RNA isolation method can affect microRNA microarray results

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Podolska, Agnieszka; Kaczkowski, Bogumil; Litman, Thomas

2011-01-01

RNA microarray analysis on porcine brain tissue. One method is a phenol-guanidine isothiocyanate-based procedure that permits isolation of total RNA. The second method, miRVana™ microRNA isolation, is column based and recovers the small RNA fraction alone. We found that microarray analyses give different results...... that depend on the RNA fraction used, in particular because some microRNAs appear very sensitive to the RNA isolation method. We conclude that precautions need to be taken when comparing microarray studies based on RNA isolated with different methods.......The quality of RNA is crucial in gene expression experiments. RNA degradation interferes in the measurement of gene expression, and in this context, microRNA quantification can lead to an incorrect estimation. In the present study, two different RNA isolation methods were used to perform micro...

Comparison of commercial exosome isolation kits for circulating exosomal microRNA profiling.

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Ding, Meng; Wang, Cheng; Lu, Xiaolan; Zhang, Cuiping; Zhou, Zhen; Chen, Xi; Zhang, Chen-Yu; Zen, Ke; Zhang, Chunni

2018-06-01

Circulating exosomal microRNAs (miRNAs) are valuable biomarker candidates; however, information on the characterization and mutual agreement of commercial kits for circulating exosomal miRNA profiling is scarce. Here, we analyzed the advantages and weaknesses of four commonly used commercial kits for exosomal miRNA profiling and their application to the sample of serum and/or plasma, respectively. NanoSight and Western blotting were conducted to evaluate the efficiency and purity of the isolated exosomes. In our conditions, the size distribution of the isolated particles was appropriate (40-150 nm), and ExoQuick™ Exosome Precipitation Solution (EXQ) generated a relatively high yield of exosomes. Nevertheless, albumin impurity was ubiquitous for all the four kits, and Total Exosome Isolation for serum or plasma (TEI) yielded a relatively pure isolation. We further performed Illumina sequencing combined with RT-qPCR to determine the ability of these kits for miRNA profiling. There was significant correlation of the exosomal miRNA profile and specific miRNAs between kits, but with differences depending on methods. exoRNeasy Serum/Plasma Midi Kit (EXR) and EXQ performed better in the specific exosomal miRNAs recovery. Intraassay CVs for specific miRNA measurement were 0.88-3.82, 1.19-3.77, 0-2.70, and 1.23-9.11% for EXR, TEI, EXQ, and RIBO™ Exosome Isolation Reagent (REI), respectively. In each kit, serum yielded a higher abundance of exosomes and exosomal miRNAs than plasma, yet with more albumin impurity. In conclusion, our data provide some valuable guidance for the methodology of disease biomarker identification of circulation exosomal miRNAs. Graphical abstract Circulating exosomal microRNAs (miRNAs) are valuable biomarker candidates; however, information on the characterization and mutual agreement of commercial kits for circulating exosomal miRNA profiling is scarce. In this study, we compared four commonly used commercially available kits for exosomal mi

Differential expression profiles of microRNAs in liver of 60Co γ-ray irradiated mice

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Sun Xiujin; Cui Fengmei; Huang Chengcheng; Hu Mingjiang; Wang Daojin; Tu Yu

2011-01-01

Objective: To investigate the differential expression profiles of microRNAs in the liver of 60 Co γ-ray irradiated mice using microRNA microarray and to explore their main functions by bioinformatic analysis. Methods: After SPF C57BL/6J mice expose to 4 Gy-single whole body radiation,total number of peripheral WBC and the fMNPCE were measured at 3 d.The differentially expressed miRNAs in mouse liver were detected with miRNA microarray, miRNA-124 and miR-34a were confirmed by real time RT-PCR assay. Bioinformatic analysis was applied to explore target genes and the main functions of the differential expressed miRNAs. Results: Compared with control group, the total number of peripheral WBC decreased (t=2.87, P<0.05), while the fMNPCE in bone marrow increased (t=-2.91, P<0.05) after 4 Gy γ-ray irradiation.miRNA microarray revealed that 17 miRNAs were differentially expressed, in which 9 up-regulated, 8 down-regulated. The expression levels of miR-124 and miR-34a were coincident with the result of real time RT-PCR. GO analysis showed that some pathways including adherens junction and cell cycle were suppressed, while some immune-related pathways were activated. Conclusions: miR-34a and miR-194 were involved in the regulation of acute radiation damage, some other miRNAs including miR-124, miR-382 and miR-92a * also played important roles in radiation process. (authors)

The MicroRNA Repertoire of Symbiodinium, the Dinoflagellate Symbiont of Reef-Building Corals

KAUST Repository

Baumgarten, Sebastian

2013-07-01

Animal and plant genomes produce numerous small RNAs (smRNAs) that regulate gene expression post-transcriptionally affecting metabolism, development, and epigenetic inheritance. In order to characterize the repertoire of endogenous microRNAs and potential gene targets, we conducted smRNA and mRNA expression profiling over nine experimental treatments of cultures from the dinoflagellate Symbiodinium sp. A1, a photosynthetic symbiont of scleractinian corals. We identified a total of 75 novel smRNAs in Symbiodinum sp. A1 that share stringent key features with functional microRNAs from other model organisms. A subset of 38 smRNAs was predicted independently over all nine treatments and their putative gene targets were identified. We found 3,187 animal-like target sites in the 3’UTRs of 12,858 mRNAs and 53 plantlike target sites in 51,917 genes. Furthermore, we identified the core RNAi protein machinery in Symbiodinium. Integration of smRNA and mRNA expression profiling identified a variety of processes that could be under microRNA control, e.g. regulation of translation, DNA modification, and chromatin silencing. Given that Symbiodinium seems to have a paucity of transcription factors and differentially expressed genes, identification and characterization of its smRNA repertoire establishes the possibility of a range of gene regulatory mechanisms in dinoflagellates acting post-transcriptionally.

Expression of microRNA related to bone remodeling regulation in plasma in patients with acromegaly

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Tatiana A. Grebennikova

2017-11-01

Full Text Available Backgraund. MiсroRNA are small regulatory factors that regulate gene expression by post-transcriptional regulation of mRNA, playing an important role in numerous cellular processes including organogenesis, apoptosis, cell proliferation and differentiation. Acromegaly causes bone fragility, but the pathogenetic mechanism is generally unknown. Aim. To evaluate levels of microRNA related to bone remodeling regulation in plasma samples from patients with acromegaly Materials and methods. Fasting plasma samples were taken and stored in aliquot at ≤ -80°C from consecutive subjects with clinically evident and biochemically confirmed active acromegaly and healthy volunteers matched by age, sex and body mass index (BMI. miRNeasy Serum/Plasma Kit, TaqMan Advanced miRNA cDNA Synthesis Kit, TaqMan Advanced miRNA Assays were used to assay plasma miRNA expression. Insulin-like growth factor 1 (IGF1 was measured by immunochemiluminescence assay (Liaison. Results. We enrolled 40 subjects 22 patients suffered from acromegaly and 18 matched healthy controls matched by sex, age and BMI. The median age of patients with acromegaly was 42 years (Q25;Q75 – 37;43 with no difference among the groups, p=0.205; BMI – 28 (24;32 kg/m2, p=0.253. The median IGF1 in subjects with acromegaly – 622 (514;1000 ng/ml was significantly higher as compared to the control group (p<0.001. Patients with acromegaly had significantly higher expression of microRNA-100-5р (p=0.051, microRNA-550а-5р (p=0.048, microRNA-7b-5р (p=0.005 and microRNA-96-5р (p=0.042 among 27 bone-specific microRNA tested in plasma Conclusions. This study reveals that several microRNAs, known to regulate bone remodeling can be detected in plasma samples of patients with acromegaly and may be suggested as biomarkers for skeletal involvement in patients with acromegaly.

Profiling of REST-dependent microRNAs reveals dynamic modes of expression

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Zhengliang eGao

2012-05-01

Full Text Available Multipotent neural stem cells (NSCs possess the ability to self-renew and differentiate into both neurons and glia. However, the detailed mechanisms underlying NSC fate decisions are not well understood. Recent work suggest that the interaction between cell-type specific transcription factors and microRNAs (miRNAs is important as resident neural stem/progenitor cells give rise to functionally mature neurons. Recently, we demonstrated that the transcriptional repressor REST (RE1-silencing transcription factor is essential to prevent precocious neuronal differentiation and maintain NSC self-renewal in the adult hippocampus. Here we show that REST is required for orchestrating the expression of distinct subsets of miRNAs in primary mouse NSC cultures, a physiologically relevant cell type. Using miRNA array profiling, we identified known REST-regulated miRNA genes, as well as previously uncharacterized REST-dependent miRNAs. Interestingly, REST-regulated miRNAs undergo dynamic expression changes under differentiation conditions over time, but not under proliferation conditions. These results suggest that REST functions in a context-dependent manner through its target miRNAs for mediating neuronal production.

MicroRNA-122 is involved in oxidative stress in isoniazid-induced liver injury in mice.

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Song, L; Zhang, Z R; Zhang, J L; Zhu, X B; He, L; Shi, Z; Gao, L; Li, Y; Hu, B; Feng, F M

2015-10-27

Many studies have shown that the pathogenesis of liver injury includes oxidative stress. MicroRNA-122 may be a marker for the early diagnosis of drug-induced liver injury. However, the relationship between microRNA-122 and oxidative stress in anti-tuberculosis drug-induced liver injury remains unknown. We measured changes in tissue microRNA-122 levels and indices of oxidative stress during liver injury in mice after administration of isoniazid, a first-line anti-tuberculosis drug. We quantified microRNA-122 expression and indices of oxidative stress at 7 time points, including 1, 3, and 5 days and 1, 2, 3, and 4 weeks. The tissue microRNA-122 levels and oxidative stress significantly changed at 3 and 5 days, suggesting that isoniazid-induced liver injury reduces oxidative stress and microRNA-122 expression compared to in the control group (P microRNA-122, began to change at 5 days (P microRNA-122 profile may affect oxidative stress by regulating mitochondrial ribosome protein S11 gene during isoniazid-induced liver injury, which may contribute to the response mechanisms of microRNA-122 and oxidative stress.

MicroRNA-gene expression network in murine liver during Schistosoma japonicum infection.

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Pengfei Cai

Full Text Available BACKGROUND: Schistosomiasis japonica remains a significant public health problem in China and Southeast Asian countries. The most typical and serious outcome of the chronic oriental schistosomiasis is the progressive granuloma and fibrosis in the host liver, which has been a major medical challenge. However, the molecular mechanism underling the hepatic pathogenesis is still not clear. METHODOLOGY AND PRINCIPAL FINDINGS: Using microarrays, we quantified the temporal gene expression profiles in the liver of Schistosoma japonicum-infected BALB/c mice at 15, 30, and 45 day post infection (dpi with that from uninfected mice as controls. Gene expression alternation associated with liver damage was observed in the initial phase of infection (dpi 15, which became more magnificent with the onset of egg-laying. Up-regulated genes were dominantly associated with inflammatory infiltration, whereas down-regulated genes primarily led to the hepatic functional disorders. Simultaneously, microRNA profiles from the same samples were decoded by Solexa sequencing. More than 130 miRNAs were differentially expressed in murine liver during S. japonicum infection. MiRNAs significantly dysregulated in the mid-phase of infection (dpi 30, such as mmu-miR-146b and mmu-miR-155, may relate to the regulation of hepatic inflammatory responses, whereas miRNAs exhibiting a peak expression in the late phase of infection (dpi 45, such as mmu-miR-223, mmu-miR-146a/b, mmu-miR-155, mmu-miR-34c, mmu-miR-199, and mmu-miR-134, may represent a molecular signature of the development of schistosomal hepatopathy. Further, a dynamic miRNA-gene co-expression network in the progression of infection was constructed. CONCLUSIONS AND SIGNIFICANCE: This study presents a global view of dynamic expression of both mRNA and miRNA transcripts in murine liver during S. japonicum infection, and highlights that miRNAs may play a variety of regulatory roles in balancing the immune responses during the

Biomarker MicroRNAs for Diagnosis of Oral Squamous Cell Carcinoma Identified Based on Gene Expression Data and MicroRNA-mRNA Network Analysis

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Zhang, Hui; Li, Tangxin; Zheng, Linqing

2017-01-01

Oral squamous cell carcinoma is one of the most malignant tumors with high mortality rate worldwide. Biomarker discovery is critical for early diagnosis and precision treatment of this disease. MicroRNAs are small noncoding RNA molecules which often regulate essential biological processes and are good candidates for biomarkers. By integrative analysis of both the cancer-associated gene expression data and microRNA-mRNA network, miR-148b-3p, miR-629-3p, miR-27a-3p, and miR-142-3p were screened as novel diagnostic biomarkers for oral squamous cell carcinoma based on their unique regulatory abilities in the network structure of the conditional microRNA-mRNA network and their important functions. These findings were confirmed by literature verification and functional enrichment analysis. Future experimental validation is expected for the further investigation of their molecular mechanisms. PMID:29098014

MicroRNA involvement in glioblastoma pathogenesis

International Nuclear Information System (INIS)

Novakova, Jana; Slaby, Ondrej; Vyzula, Rostislav; Michalek, Jaroslav

2009-01-01

MicroRNAs are endogenously expressed regulatory noncoding RNAs. Altered expression levels of several microRNAs have been observed in glioblastomas. Functions and direct mRNA targets for these microRNAs have been relatively well studied over the last years. According to these data, it is now evident, that impairment of microRNA regulatory network is one of the key mechanisms in glioblastoma pathogenesis. MicroRNA deregulation is involved in processes such as cell proliferation, apoptosis, cell cycle regulation, invasion, glioma stem cell behavior, and angiogenesis. In this review, we summarize the current knowledge of miRNA functions in glioblastoma with an emphasis on its significance in glioblastoma oncogenic signaling and its potential to serve as a disease biomarker and a novel therapeutic target in oncology.

MicroRNA-146a expression as a potential biomarker for rheumatoid ...

African Journals Online (AJOL)

MicroRNA-146a expression as a potential biomarker for rheumatoid arthritis in Egypt. Heba Mohamed Abdelkader Elsayed, Walaa Shawky Khater, Ayman Asaad Ibrahim, Maha Salah El-din Hamdy, Nashwa Aly Morshedy ...

Analysis of plasma microRNA expression profiles in male textile workers with noise-induced hearing loss.

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Ding, Lu; Liu, Jing; Shen, Huan-Xi; Pan, Li-Ping; Liu, Qing-Dong; Zhang, Heng-Dong; Han, Lei; Shuai, Li-Guo; Ding, En-Min; Zhao, Qiu-Ni; Wang, Bo-Shen; Zhu, Bao-Li

2016-03-01

Circulating microRNAs (miRNAs) have attracted interests as non-invasive biomarkers of physiological and pathological conditions, which may be applied in noise-induced hearing loss (NIHL). However, no epidemiology studies have yet examined the potential effects of NIHL or noise exposure on miRNA expression profiles. We sought to identify permanent NIHL-related miRNAs and to predict the biological functions of the putative genes encoding the indicated miRNAs. In the discovery stage, we used a microarray assay to detect the miRNA expression profiles between pooled plasma samples from 10 noise-exposed individuals with normal hearing and 10 NIHL patients. In addition, we conducted a preliminary validation of six candidate miRNAs in the same 20 workers. Subsequently, three miRNAs were selected for expanded validation in 23 non-exposed individuals with normal hearing and 46 noise-exposed textile workers which including 23 noise-exposed workers with normal hearing and 23 NIHL patients. Moreover, we predicted the biological functions of the putative target genes using a Gene Ontology (GO) function enrichment analysis. In the discovery stage, compared with the noise exposures with normal hearing, 73 miRNAs demonstrated at least a 1.5-fold differential expression in the NIHL patients. In the preliminary validation, compared with the noise exposures, the plasma levels of miR-16-5p, miR-24-3p, miR-185-5p and miR-451a were all upregulated (P < 0.001) in the NIHL patients. In the expanded validation stage, compared with the non-exposures, the plasma levels of miR-24, miR-185-5p and miR-451a were all significantly downregulated (P < 0.001) in the exposures. And compared with the noise exposures, the plasma levels of miR-185-5p and miR-451a were slightly elevated (P < 0.001) in the NIHL patients, which were consistent with the results of preliminary validation and microarray analysis. The two indicated plasma miRNAs may be biomarkers of indicating responses to noise exposure

MicroRNA Expression during Viral Infection or PolyI:C Stimulation in a Fish Model

DEFF Research Database (Denmark)

Kristensen, Lasse Bøgelund Juel; Schyth, Brian Dall; Lorenzen, Niels

Fish are important as small vertebrate models for studying various aspects of development and disease. MicroRNA regulation in fish has so far received attention especially in studies of their expression and function during embryonic development. In the studies carried out at the National Veterinary...... Institute in Århus we aim at using fish models for studying microRNA regulation during viral infection. In the studies presented here we make use of a qPCR method to detect miRNAs in fish cells. We present results regarding the expression of the immunologically relevant microRNAs, miR-155, miR-146a and mi......R-146b in fish cells during infection with the fish pathogenic virus viral hemorrhagic septicemia virus (VHSV) and during immune stimulation with double stranded RNA (polyI:C). We highlight the need of finding stable normalization genes for microRNA detection....

RNA Helicase DDX5 Regulates MicroRNA Expression and Contributes to Cytoskeletal Reorganization in Basal Breast Cancer Cells

Energy Technology Data Exchange (ETDEWEB)

Wang, Daojing; Huang, Jing; Hu, Zhi

2011-11-15

RNA helicase DDX5 (also p68) is involved in all aspects of RNA metabolism and serves as a transcriptional co-regulator, but its functional role in breast cancer remains elusive. Here, we report an integrative biology study of DDX5 in breast cancer, encompassing quantitative proteomics, global MicroRNA profiling, and detailed biochemical characterization of cell lines and human tissues. We showed that protein expression of DDX5 increased progressively from the luminal to basal breast cancer cell lines, and correlated positively with that of CD44 in the basal subtypes. Through immunohistochemistry analyses of tissue microarrays containing over 200 invasive human ductal carcinomas, we observed that DDX5 was upregulated in the majority of malignant tissues, and its expression correlated strongly with those of Ki67 and EGFR in the triple-negative tumors. We demonstrated that DDX5 regulated a subset of MicroRNAs including miR-21 and miR-182 in basal breast cancer cells. Knockdown of DDX5 resulted in reorganization of actin cytoskeleton and reduction of cellular proliferation. The effects were accompanied by upregulation of tumor suppressor PDCD4 (a known miR-21 target); as well as upregulation of cofilin and profilin, two key proteins involved in actin polymerization and cytoskeleton maintenance, as a consequence of miR-182 downregulation. Treatment with miR-182 inhibitors resulted in morphologic phenotypes resembling those induced by DDX5 knockdown. Using bioinformatics tools for pathway and network analyses, we confirmed that the network for regulation of actin cytoskeleton was predominantly enriched for the predicted downstream targets of miR-182. Our results reveal a new functional role of DDX5 in breast cancer via the DDX5→miR-182→actin cytoskeleton pathway, and suggest the potential clinical utility of DDX5 and its downstream MicroRNAs in the theranostics of breast cancer.

OP17MICRORNA PROFILING USING SMALL RNA-SEQ IN PAEDIATRIC LOW GRADE GLIOMAS

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Jeyapalan, Jennie N.; Jones, Tania A.; Tatevossian, Ruth G.; Qaddoumi, Ibrahim; Ellison, David W.; Sheer, Denise

2014-01-01

INTRODUCTION: MicroRNAs regulate gene expression by targeting mRNAs for translational repression or degradation at the post-transcriptional level. In paediatric low-grade gliomas a few key genetic mutations have been identified, including BRAF fusions, FGFR1 duplications and MYB rearrangements. Our aim in the current study is to profile aberrant microRNA expression in paediatric low-grade gliomas and determine the role of epigenetic changes in the aetiology and behaviour of these tumours. METHOD: MicroRNA profiling of tumour samples (6 pilocytic, 2 diffuse, 2 pilomyxoid astrocytomas) and normal brain controls (4 adult normal brain samples and a primary glial progenitor cell-line) was performed using small RNA sequencing. Bioinformatic analysis included sequence alignment, analysis of the number of reads (CPM, counts per million) and differential expression. RESULTS: Sequence alignment identified 695 microRNAs, whose expression was compared in tumours v. normal brain. PCA and hierarchical clustering showed separate groups for tumours and normal brain. Computational analysis identified approximately 400 differentially expressed microRNAs in the tumours compared to matched location controls. Our findings will then be validated and integrated with extensive genetic and epigenetic information we have previously obtained for the full tumour cohort. CONCLUSION: We have identified microRNAs that are differentially expressed in paediatric low-grade gliomas. As microRNAs are known to target genes involved in the initiation and progression of cancer, they provide critical information on tumour pathogenesis and are an important class of biomarkers.

Differentially Expressed microRNAs and Target Genes Associated with Plastic Internode Elongation in Alternanthera philoxeroides in Contrasting Hydrological Habitats

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Gengyun Li

2017-12-01

Full Text Available Phenotypic plasticity is crucial for plants to survive in changing environments. Discovering microRNAs, identifying their targets and further inferring microRNA functions in mediating plastic developmental responses to environmental changes have been a critical strategy for understanding the underlying molecular mechanisms of phenotypic plasticity. In this study, the dynamic expression patterns of microRNAs under contrasting hydrological habitats in the amphibious species Alternanthera philoxeroides were identified by time course expression profiling using high-throughput sequencing technology. A total of 128 known and 18 novel microRNAs were found to be differentially expressed under contrasting hydrological habitats. The microRNA:mRNA pairs potentially associated with plastic internode elongation were identified by integrative analysis of microRNA and mRNA expression profiles, and were validated by qRT-PCR and 5′ RLM-RACE. The results showed that both the universal microRNAs conserved across different plants and the unique microRNAs novelly identified in A. philoxeroides were involved in the responses to varied water regimes. The results also showed that most of the differentially expressed microRNAs were transiently up-/down-regulated at certain time points during the treatments. The fine-scale temporal changes in microRNA expression highlighted the importance of time-series sampling in identifying stress-responsive microRNAs and analyzing their role in stress response/tolerance.

A Downmodulated MicroRNA Profiling in Patients with Gastric Cancer

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Tao Zhang

2017-01-01

Full Text Available Objective. Here, we aim to investigate the microRNA (miR profiling in human gastric cancer (GC. Methods. Tumoral and matched peritumoral gastric specimens were collected from 12 GC patients who underwent routine surgery. A high-throughput miR sequencing method was applied to detect the aberrantly expressed miRs in a subset of 6 paired samples. The stem-loop quantitative real-time polymerase chain reaction (qRT-PCR assay was subsequently performed to confirm the sequencing results in the remaining 6 paired samples. The profiling results were also validated in vitro in three human GC cell lines (BGC-823, MGC-803, and GTL-16 and a normal gastric epithelial cell line (GES-1. Results. The miR sequencing approach detected 5 differentially expressed miRs, hsa-miR-132-3p, hsa-miR-155-5p, hsa-miR-19b-3p, hsa-miR-204-5p, and hsa-miR-30a-3p, which were significantly downmodulated between the tumoral and peritumoral GC tissues. Most of the results were further confirmed by qRT-PCR, while no change was observed for hsa-miR-30a-3p. The in vitro finding also agreed with the results of both miR sequencing and qRT-PCR for hsa-miR-204-5p, hsa-miR-155-5p, and hsa-miR-132-3p. Conclusion. Together, our findings may serve to identify new molecular alterations as well as to enrich the miR profiling in human GC.

Comparative Profiling of microRNA Expression in Soybean Seeds from Genetically Modified Plants and their Near-Isogenic Parental Lines

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Wang, Yong; Lan, Qingkuo; Zhao, Xin; Xu, Wentao; Li, Feiwu; Wang, Qinying; Chen, Rui

2016-01-01

MicroRNAs (miRNAs) have been widely demonstrated to play fundamental roles in gene regulation in most eukaryotes. To date, there has been no study describing the miRNA composition in genetically modified organisms (GMOs). In this study, small RNAs from dry seeds of two GM soybean lines and their parental cultivars were investigated using deep sequencing technology and bioinformatic approaches. As a result, several differentially expressed gma-miRNAs were found between the GM and non-GM soybeans. Meanwhile, more differentially expressed gma-miRNAs were identified between distantly relatednon-GM soybeans, indicating that the miRNA components of soybean seeds varied among different soybean lines, including the GM and non-GM soybeans, and the extent of difference might be related to their genetic relationship. Additionally, fourteen novel gma-miRNA candidates were predicted in soybean seeds including a potential bidirectionally transcribed miRNA family with two genomic loci (gma-miR-N1). Our findings firstly provided useful data for miRNA composition in edible GM crops and also provided valuable information for soybean miRNA research. PMID:27214227

Comparative Profiling of microRNA Expression in Soybean Seeds from Genetically Modified Plants and their Near-Isogenic Parental Lines.

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Yong Wang

Full Text Available MicroRNAs (miRNAs have been widely demonstrated to play fundamental roles in gene regulation in most eukaryotes. To date, there has been no study describing the miRNA composition in genetically modified organisms (GMOs. In this study, small RNAs from dry seeds of two GM soybean lines and their parental cultivars were investigated using deep sequencing technology and bioinformatic approaches. As a result, several differentially expressed gma-miRNAs were found between the GM and non-GM soybeans. Meanwhile, more differentially expressed gma-miRNAs were identified between distantly relatednon-GM soybeans, indicating that the miRNA components of soybean seeds varied among different soybean lines, including the GM and non-GM soybeans, and the extent of difference might be related to their genetic relationship. Additionally, fourteen novel gma-miRNA candidates were predicted in soybean seeds including a potential bidirectionally transcribed miRNA family with two genomic loci (gma-miR-N1. Our findings firstly provided useful data for miRNA composition in edible GM crops and also provided valuable information for soybean miRNA research.

Comparative Profiling of microRNA Expression in Soybean Seeds from Genetically Modified Plants and their Near-Isogenic Parental Lines.

Science.gov (United States)

Wang, Yong; Lan, Qingkuo; Zhao, Xin; Xu, Wentao; Li, Feiwu; Wang, Qinying; Chen, Rui

2016-01-01

MicroRNAs (miRNAs) have been widely demonstrated to play fundamental roles in gene regulation in most eukaryotes. To date, there has been no study describing the miRNA composition in genetically modified organisms (GMOs). In this study, small RNAs from dry seeds of two GM soybean lines and their parental cultivars were investigated using deep sequencing technology and bioinformatic approaches. As a result, several differentially expressed gma-miRNAs were found between the GM and non-GM soybeans. Meanwhile, more differentially expressed gma-miRNAs were identified between distantly relatednon-GM soybeans, indicating that the miRNA components of soybean seeds varied among different soybean lines, including the GM and non-GM soybeans, and the extent of difference might be related to their genetic relationship. Additionally, fourteen novel gma-miRNA candidates were predicted in soybean seeds including a potential bidirectionally transcribed miRNA family with two genomic loci (gma-miR-N1). Our findings firstly provided useful data for miRNA composition in edible GM crops and also provided valuable information for soybean miRNA research.

Impact of gastro-oesophageal reflux on microRNA expression, location and function

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Smith Cameron M

2013-01-01

Full Text Available Abstract Background Ulceration of the oesophageal squamous mucosa (ulcerative oesophagitis is a pathological manifestation of gastro-oesophageal reflux disease, and is a major risk factor for the development of Barrett’s oesophagus. Barrett’s oesophagus is characterised by replacement of reflux-damaged oesophageal squamous epithelium with a columnar intestinal-like epithelium. We previously reported discovery of microRNAs that are differentially expressed between oesophageal squamous mucosa and Barrett’s oesophagus mucosa. Now, to better understand early steps in the initiation of Barrett’s oesophagus, we assessed the expression, location and function of these microRNAs in oesophageal squamous mucosa from individuals with ulcerative oesophagitis. Methods Quantitative real-time PCR was used to compare miR-21, 143, 145, 194, 203, 205 and 215 expression levels in oesophageal mucosa from individuals without pathological gastro-oesophageal reflux to individuals with ulcerative oesophagitis. Correlations between microRNA expression and messenger RNA differentiation markers BMP-4, CK8 and CK14 were analyzed. The cellular localisation of microRNAs within the oesophageal mucosa was determined using in-situ hybridisation. microRNA involvement in proliferation and apoptosis was assessed following transfection of a human squamous oesophageal mucosal cell line (Het-1A. Results miR-143, miR-145 and miR-205 levels were significantly higher in gastro-oesophageal reflux compared with controls. Elevated miR-143 expression correlated with BMP-4 and CK8 expression, and elevated miR-205 expression correlated negatively with CK14 expression. Endogenous miR-143, miR-145 and miR-205 expression was localised to the basal layer of the oesophageal epithelium. Transfection of miR-143, 145 and 205 mimics into Het-1A cells resulted in increased apoptosis and decreased proliferation. Conclusions Elevated miR-143, miR-145 and miR-205 expression was observed in

Adenoid cystic carcinomas of the salivary gland, lacrimal gland, and breast are morphologically and genetically similar but have distinct microRNA expression profiles

DEFF Research Database (Denmark)

Andreasen, Simon; Tan, Qihua; Agander, Tina Klitmøller

2018-01-01

Adenoid cystic carcinoma is among the most frequent malignancies in the salivary and lacrimal glands and has a grave prognosis characterized by frequent local recurrences, distant metastases, and tumor-related mortality. Conversely, adenoid cystic carcinoma of the breast is a rare type of triple......-negative (estrogen and progesterone receptor, HER2) and basal-like carcinoma, which in contrast to other triple-negative and basal-like breast carcinomas has a very favorable prognosis. Irrespective of site, adenoid cystic carcinoma is characterized by gene fusions involving MYB, MYBL1, and NFIB, and the reason...... for the different clinical outcomes is unknown. In order to identify the molecular mechanisms underlying the discrepancy in clinical outcome, we characterized the phenotypic profiles, pattern of gene rearrangements, and global microRNA expression profiles of 64 salivary gland, 9 lacrimal gland, and 11 breast...

Design, Construction, and Validation of Artificial MicroRNA Vectors Using Agrobacterium-Mediated Transient Expression System.

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Bhagwat, Basdeo; Chi, Ming; Han, Dianwei; Tang, Haifeng; Tang, Guiliang; Xiang, Yu

2016-01-01

Artificial microRNA (amiRNA) technology utilizes microRNA (miRNA) biogenesis pathway to produce artificially selected small RNAs using miRNA gene backbone. It provides a feasible strategy for inducing loss of gene function, and has been applied in functional genomics study, improvement of crop quality and plant virus disease resistance. A big challenge in amiRNA applications is the unpredictability of silencing efficacy of the designed amiRNAs and not all constructed amiRNA candidates would be expressed effectively in plant cells. We and others found that high efficiency and specificity in RNA silencing can be achieved by designing amiRNAs with perfect or almost perfect sequence complementarity to their targets. In addition, we recently demonstrated that Agrobacterium-mediated transient expression system can be used to validate amiRNA constructs, which provides a simple, rapid and effective method to select highly expressible amiRNA candidates for stable genetic transformation. Here, we describe the methods for design of amiRNA candidates with perfect or almost perfect base-pairing to the target gene or gene groups, incorporation of amiRNA candidates in miR168a gene backbone by one step inverse PCR amplification, construction of plant amiRNA expression vectors, and assay of transient expression of amiRNAs in Nicotiana benthamiana through agro-infiltration, small RNA extraction, and amiRNA Northern blot.

MicroRNA Biomarkers in Neurodegenerative Diseases and Emerging Nano-Sensors Technology

DEFF Research Database (Denmark)

Shah, Pratik; Cho, Seok Keun; Thulstrup, Peter Waaben

2017-01-01

MicroRNAs (miRNAs) are essential small RNA molecules (20–24 nt) that negatively regulate the expression of target genes at the post-transcriptional level. Due to their roles in a variety of biological processes, the aberrant expression profiles of miRNAs have been identified as biomarkers for man...

Expression of genes for microRNA-processing enzymes is altered in advanced non-alcoholic fatty liver disease.

Science.gov (United States)

Sharma, Haveesh; Estep, Michael; Birerdinc, Aybike; Afendy, Arian; Moazzez, Amir; Elariny, Hazem; Goodman, Zachary; Chandhoke, Vikas; Baranova, Ancha; Younossi, Zobair M

2013-08-01

Recently, microRNAs (miRNA) have been linked to the pathogenesis of non-alcoholic fatty liver disease (NAFLD) and its progression to non-alcoholic steatohepatitis (NASH). First transcribed as pri-miRNA, these molecules are further processed by a complex of endonuclear and cytosolic RNA binding molecules to form mature miRNAs. The aim of this study is to investigate mechanisms of miRNA regulation in the visceral adipose of obese NAFLD patients via measuring expression of miRNA processing enzymes and pri-miRNA. Total RNAs were extracted from visceral adipose tissue (VAT) samples collected from patients undergoing bariatric surgery. All patients had biopsy-proven NAFLD (NASH patients [n = 12] and non-NASH NAFLD [n = 12]). For each patient, we profiled mRNA levels for three miRNA processing elements (Drosha, DGCR8, and Dicer1) and seven pri-miRNAs (pri-miR-125b-2, pri-miR-16-2, pri-miR-26a-1, pri-miR-26a-2, pri-miR-7-1, pri-miR-7-2, and pri-miR-7-3). Expression of Dicer1, Drosha and DGCR8 was significantly increased within the NASH cohort along with expression of pri-miR-7-1. The presence of focal necrosis on the liver biopsy correlated significantly with levels of Dicer1 and DGRC8. Both NASH and ballooning degeneration of hepatocytes correlated negatively with the expression levels of hsa-miR-125b. Histologic NASH correlated positively with the expression levels of pri-miR-16-2 and pri-miR-7-1. The presence of the hepatocyte's ballooning degeneration in the liver biopsy correlated positively with pri-miR-26a-1 and pri-miR-7-1. The expression profile of pri-miR-125b-2 also correlated positively with body mass index. Our findings support the hypothesis that VAT-derived miRNA may contribute to the pathogenesis of NASH in obese patients. © 2013 Journal of Gastroenterology and Hepatology Foundation and Wiley Publishing Asia Pty Ltd.

Integrated Analysis of Dysregulated ncRNA and mRNA Expression Profiles in Humans Exposed to Carbon Nanotubes.

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Anna A Shvedova

Full Text Available As the application of carbon nanotubes (CNT in consumer products continues to rise, studies have expanded to determine the associated risks of exposure on human and environmental health. In particular, several lines of evidence indicate that exposure to multi-walled carbon nanotubes (MWCNT could pose a carcinogenic risk similar to asbestos fibers. However, to date the potential markers of MWCNT exposure are not yet explored in humans.In the present study, global mRNA and ncRNA expression profiles in the blood of exposed workers, having direct contact with MWCNT aerosol for at least 6 months (n = 8, were compared with expression profiles of non-exposed (n = 7 workers (e.g., professional and/or technical staff from the same manufacturing facility.Significant changes in the ncRNA and mRNA expression profiles were observed between exposed and non-exposed worker groups. An integrative analysis of ncRNA-mRNA correlations was performed to identify target genes, functional relationships, and regulatory networks in MWCNT-exposed workers. The coordinated changes in ncRNA and mRNA expression profiles revealed a set of miRNAs and their target genes with roles in cell cycle regulation/progression/control, apoptosis and proliferation. Further, the identified pathways and signaling networks also revealed MWCNT potential to trigger pulmonary and cardiovascular effects as well as carcinogenic outcomes in humans, similar to those previously described in rodents exposed to MWCNTs.This study is the first to investigate aberrant changes in mRNA and ncRNA expression profiles in the blood of humans exposed to MWCNT. The significant changes in several miRNAs and mRNAs expression as well as their regulatory networks are important for getting molecular insights into the MWCNT-induced toxicity and pathogenesis in humans. Further large-scale prospective studies are necessary to validate the potential applicability of such changes in mRNAs and miRNAs as prognostic markers

Comparison of microRNA expression using different preservation methods of matched psoriatic skin samples

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Løvendorf, Marianne B; Zibert, John R; Hagedorn, Peter H

2012-01-01

MicroRNAs are non-coding RNA molecules modulating gene expression post-transcriptionally. Formalin-fixed, paraffin-embedding (FFPE) is a standard preservation method often used in clinical practices, but induces RNA degradation. Extracting high-quality RNA from human skin can be challenging as skin...

mirEX: a platform for comparative exploration of plant pri-miRNA expression data.

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Bielewicz, Dawid; Dolata, Jakub; Zielezinski, Andrzej; Alaba, Sylwia; Szarzynska, Bogna; Szczesniak, Michal W; Jarmolowski, Artur; Szweykowska-Kulinska, Zofia; Karlowski, Wojciech M

2012-01-01

mirEX is a comprehensive platform for comparative analysis of primary microRNA expression data. RT-qPCR-based gene expression profiles are stored in a universal and expandable database scheme and wrapped by an intuitive user-friendly interface. A new way of accessing gene expression data in mirEX includes a simple mouse operated querying system and dynamic graphs for data mining analyses. In contrast to other publicly available databases, the mirEX interface allows a simultaneous comparison of expression levels between various microRNA genes in diverse organs and developmental stages. Currently, mirEX integrates information about the expression profile of 190 Arabidopsis thaliana pri-miRNAs in seven different developmental stages: seeds, seedlings and various organs of mature plants. Additionally, by providing RNA structural models, publicly available deep sequencing results, experimental procedure details and careful selection of auxiliary data in the form of web links, mirEX can function as a one-stop solution for Arabidopsis microRNA information. A web-based mirEX interface can be accessed at http://bioinfo.amu.edu.pl/mirex.

Detection and comparison of microRNA expression in the serum of Doberman Pinschers with dilated cardiomyopathy and healthy controls.

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Steudemann, Carola; Bauersachs, Stefan; Weber, Karin; Wess, Gerhard

2013-01-17

Dilated cardiomyopathy (DCM) is the most common heart disease in Doberman Pinschers. MicroRNAs (miRNAs) are short non-coding RNAs playing important roles in gene regulation. Different miRNA expression patterns have been described for DCM in humans and might represent potential diagnostic markers. There are no studies investigating miRNA expression profiles in canine DCM. The aims of this study were to screen the miRNA expression profile of canine serum using miRNA microarray and to compare expression patterns of a group of Doberman Pinschers with DCM and healthy controls. Eight Doberman Pinschers were examined by echocardiography and 24-hour-ECG and classified as healthy (n=4) or suffering from DCM (n=4). Total RNA was extracted from serum and hybridized on a custom-designed 8x60k miRNA microarray (Agilent) containing probes for 1368 individual miRNAs. Although total RNA concentrations were very low in serum samples, 404 different miRNAs were detectable with sufficient signal intensity on miRNA microarray. 22 miRNAs were differentially expressed in the two groups (p1.5), but did not reach statistical significance after multiple testing correction (false discovery rate adjusted p>0.05). Five miRNAs were selected for further analysis using quantitative Real-Time RT-PCR (qPCR) assays. No significant differences were found using specific miRNA qPCR assays (p>0.05). Numerous miRNAs can be detected in canine serum. Between healthy and DCM dogs, miRNA expression changes could be detected, but the results did not reach statistical significance most probably due to the small group size. miRNAs are potential new circulating biomarkers in veterinary medicine and should be investigated in larger patient groups and additional canine diseases.

Natural RNA circles function as efficient microRNA sponges

DEFF Research Database (Denmark)

Hansen, Thomas Birkballe; Jensen, Trine I; Clausen, Bettina Hjelm

2013-01-01

MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression that act by direct base pairing to target sites within untranslated regions of messenger RNAs. Recently, miRNA activity has been shown to be affected by the presence of miRNA sponge transcripts, the so-called comp......MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression that act by direct base pairing to target sites within untranslated regions of messenger RNAs. Recently, miRNA activity has been shown to be affected by the presence of miRNA sponge transcripts, the so......-called competing endogenous RNA in humans and target mimicry in plants. We previously identified a highly expressed circular RNA (circRNA) in human and mouse brain. Here we show that this circRNA acts as a miR-7 sponge; we term this circular transcript ciRS-7 (circular RNA sponge for miR-7). ciRS-7 contains more...... sponge, suggesting that miRNA sponge effects achieved by circRNA formation are a general phenomenon. This study serves as the first, to our knowledge, functional analysis of a naturally expressed circRNA....

Analysis of microRNA expression profiling identifies miR-155 and miR-155* as potential diagnostic markers for active tuberculosis: a preliminary study.

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Wu, Jing; Lu, Chanyi; Diao, Ni; Zhang, Shu; Wang, Sen; Wang, Feifei; Gao, Yan; Chen, Jiazhen; Shao, Lingyun; Lu, Jingning; Zhang, Xuelian; Weng, Xinhua; Wang, Honghai; Zhang, Wenhong; Huang, Yuxian

2012-01-01

To explore biologic behaviors and disease relevance of microRNAs (miRNAs) in the development of active tuberculosis (ATB), we investigated the expression profile of Mycobacterium tuberculosis (MTB) purified protein derivative (PPD)-induced miRNAs to determine the specific miRNAs involved in the pathogenesis of ATB. The expression profile of miRNA under PPD challenge was first measured using microarray analysis in peripheral blood mononuclear cells isolated from ATB patients and healthy controls (HC). The remarkably reactive miRNAs were then validated in a larger cohort by quantitative real-time polymerase chain reaction (qRT-PCR). The receiver operating characteristic (ROC) curve was plotted to evaluate the diagnostic value of the determined PPD-responsive miRNAs. The potential targets for those miRNAs were also predicted by computational programs. Fourteen of 866 human miRNAs exhibited at least 1.8-fold difference in the ratio of expression level before and after stimulation with PPD between the ATB and HC groups. The qRT-PCR study validated the findings from microarray-based screening, in which miR-155 exhibited a fold change of 1.4 in the HC group and 3.7 in the ATB group upon PPD stimulation (p microRNAs exhibited no differences between the ATB and HC groups. miR-155 and miR-155* exhibited characteristic expression by TB-specific antigen, suggesting that they can be potential diagnostic markers under the challenge of specific MTB antigens. Copyright © 2012 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

Isolation of microRNA targets using biotinylated synthetic microRNAs

DEFF Research Database (Denmark)

Ørom, Ulf Andersson; Lund, Anders H

2007-01-01

MicroRNAs are small regulatory RNAs found in multicellular organisms where they post-transcriptionally regulate gene expression. In animals, microRNAs bind mRNAs via incomplete base pairings making the identification of microRNA targets inherently difficult. Here, we present a detailed method...... for experimental identification of microRNA targets based on affinity purification of tagged microRNAs associated with their targets. Udgivelsesdato: 2007-Oct...

Expression profile of microRNA-146a along HPV-induced multistep carcinogenesis: a study in HPV16 transgenic mice.

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Araújo, Rita; Santos, Joana M O; Fernandes, Mara; Dias, Francisca; Sousa, Hugo; Ribeiro, Joana; Bastos, Margarida M S M; Oliveira, Paula A; Carmo, Diogo; Casaca, Fátima; Silva, Sandra; Medeiros, Rui; Gil da Costa, Rui M

2018-02-01

Persistent human papillomavirus (HPV) infection is associated with the development of certain types of cancer and the dysregulation of microRNAs has been implicated in HPV-associated carcinogenesis. This is the case of microRNA-146a (miR-146a), which is thought to regulate tumor-associated inflammation. We sought to investigate the expression levels of miR-146a during HPV16-mediated carcinogenesis using skin samples from K14-HPV16 transgenic mice which develop the consecutive phases of the carcinogenesis process. Female transgenic (HPV +/- ) and wild-type (HPV -/- ) mice were sacrificed at 24-26 weeks-old or 28-30 weeks-old. Chest and ear skin samples from HPV +/- and HPV -/- mice were histologically classified and used for microRNA extraction and quantification by qPCR. Chest skin samples from 24 to 26 weeks-old HPV +/- mice presented diffuse epidermal hyperplasia and only 22.5% showed multifocal dysplasia, while at 28-30 weeks-old all (100.0%) HPV +/- animals showed epidermal dysplasia. All HPV +/- ear skin samples showed carcinoma in situ (CIS). MiR-146a expression levels were higher in HPV +/- compared to HPV -/- mice (p = 0.006). There was also an increase in miR-146a expression in dysplastic skin lesions compared with hyperplasic lesions (p = 0.011). Samples showing CIS had a significant decrease in miR-146a expression when compared to samples showing epidermal hyperplasia (p = 0.018) and epidermal dysplasia (p = 0.009). These results suggest that HPV16 induces the overexpression of miR-146a in the initial stages of carcinogenesis (hyperplasia and dysplasia), whereas decreases its expression at later stages (CIS). Taken together, these data implicate and suggest different roles of miR-146a in HPV-mediated carcinogenesis.

Characterization of circulating microRNA expression in patients with a ventricular septal defect.

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Dong Li

Full Text Available Ventricular septal defect (VSD, one of the most common types of congenital heart disease (CHD, results from a combination of environmental and genetic factors. Recent studies demonstrated that microRNAs (miRNAs are involved in development of CHD. This study was to characterize the expression of miRNAs that might be involved in the development or reflect the consequences of VSD.MiRNA microarray analysis and reverse transcription-polymerase chain reaction (RT-PCR were employed to determine the miRNA expression profile from 3 patients with VSD and 3 VSD-free controls. 3 target gene databases were employed to predict the target genes of differentially expressed miRNAs. miRNAs that were generally consensus across the three databases were selected and then independently validated using real time PCR in plasma samples from 20 VSD patients and 15 VSD-free controls. Target genes of validated 8 miRNAs were predicted using bioinformatic methods.36 differentially expressed miRNAs were found in the patients with VSD and the VSD-free controls. Compared with VSD-free controls, expression of 15 miRNAs were up-regulated and 21 miRNAs were downregulated in the VSD group. 15 miRNAs were selected based on database analysis results and expression levels of 8 miRNAs were validated. The results of the real time PCR were consistent with those of the microarray analysis. Gene ontology analysis indicated that the top target genes were mainly related to cardiac right ventricle morphogenesis. NOTCH1, HAND1, ZFPM2, and GATA3 were predicted as targets of hsa-let-7e-5p, hsa-miR-222-3p and hsa-miR-433.We report for the first time the circulating miRNA profile for patients with VSD and showed that 7 miRNAs were downregulated and 1 upregulated when matched to VSD-free controls. Analysis revealed target genes involved in cardiac development were probably regulated by these miRNAs.

Characterization of circulating microRNA expression in patients with a ventricular septal defect.

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Li, Dong; Ji, Long; Liu, Lianbo; Liu, Yizhi; Hou, Haifeng; Yu, Kunkun; Sun, Qiang; Zhao, Zhongtang

2014-01-01

Ventricular septal defect (VSD), one of the most common types of congenital heart disease (CHD), results from a combination of environmental and genetic factors. Recent studies demonstrated that microRNAs (miRNAs) are involved in development of CHD. This study was to characterize the expression of miRNAs that might be involved in the development or reflect the consequences of VSD. MiRNA microarray analysis and reverse transcription-polymerase chain reaction (RT-PCR) were employed to determine the miRNA expression profile from 3 patients with VSD and 3 VSD-free controls. 3 target gene databases were employed to predict the target genes of differentially expressed miRNAs. miRNAs that were generally consensus across the three databases were selected and then independently validated using real time PCR in plasma samples from 20 VSD patients and 15 VSD-free controls. Target genes of validated 8 miRNAs were predicted using bioinformatic methods. 36 differentially expressed miRNAs were found in the patients with VSD and the VSD-free controls. Compared with VSD-free controls, expression of 15 miRNAs were up-regulated and 21 miRNAs were downregulated in the VSD group. 15 miRNAs were selected based on database analysis results and expression levels of 8 miRNAs were validated. The results of the real time PCR were consistent with those of the microarray analysis. Gene ontology analysis indicated that the top target genes were mainly related to cardiac right ventricle morphogenesis. NOTCH1, HAND1, ZFPM2, and GATA3 were predicted as targets of hsa-let-7e-5p, hsa-miR-222-3p and hsa-miR-433. We report for the first time the circulating miRNA profile for patients with VSD and showed that 7 miRNAs were downregulated and 1 upregulated when matched to VSD-free controls. Analysis revealed target genes involved in cardiac development were probably regulated by these miRNAs.

MicroRNA expression profiles of drug-resistance breast cancer cells and their exosomes.

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Zhong, Shanliang; Chen, Xiu; Wang, Dandan; Zhang, Xiaohui; Shen, Hongyu; Yang, Sujin; Lv, Mengmeng; Tang, Jinhai; Zhao, Jianhua

2016-04-12

Exosomes have been shown to transmit drug resistance through delivering miRNAs. We aimed to explore their roles in breast cancer. Three resistant sublines were established by exposing parental MDA-MB-231 cell line to docetaxel, epirubicin and vinorelbine, respectively. Preneoadjuvant chemotherapy biopsies and paired surgically-resected specimens embedded in paraffin from 23 breast cancer patients were collected. MiRNA expression profiles of the cell lines and their exosomes were evaluated using microarray. The result showed that most miRNAs in exosomes had a lower expression level than that in cells, however, some miRNAs expressed higher in exosomes than in cells, suggesting a number of miRNAs is concentrated in exosomes. Among the dysregulated miRNAs, 22 miRNAs were consistently up-regulated in exosomes and their cells of origin. We further found that 12 of the 22 miRNAs were significantly up-regulated after preneoadjuvant chemotherapy. Further study of the role of these 12 miRNAs in acquisition of drug resistance is needed to clarify their contribution to chemoresistance.

MicroRNA expression profile associated with response to neoadjuvant chemoradiotherapy in locally advanced rectal cancer patients

International Nuclear Information System (INIS)

Svoboda, Marek; Sana, Jiri; Fabian, Pavel; Kocakova, Ilona; Gombosova, Jana; Nekvindova, Jana; Radova, Lenka; Vyzula, Rostislav; Slaby, Ondrej

2012-01-01

Rectal cancer accounts for approximately one third of all colorectal cancers (CRC), which belong among leading causes of cancer deaths worldwide. Standard treatment for locally advanced rectal cancer (cT3/4 and/or cN+) includes neoadjuvant chemoradiotherapy with fluoropyrimidines (capecitabine or 5-fluorouracil) followed by radical surgical resection. Unfortunately, a significant proportion of tumors do not respond enough to the neoadjuvant treatment and these patients are at risk of relapse. MicroRNAs (miRNAs) are small non-coding RNAs playing significant roles in the pathogenesis of many cancers including rectal cancer. MiRNAs could present the new predictive biomarkers for rectal cancer patients. We selected 20 patients who underwent neoadjuvant chemoradiotherapy for advanced rectal cancer and whose tumors were classified as most sensitive or resistant to the treatment. These two groups were compared using large-scale miRNA expression profiling. Expression levels of 8 miRNAs significantly differed between two groups. MiR-215, miR-190b and miR-29b-2* have been overexpressed in non-responders, and let-7e, miR-196b, miR-450a, miR-450b-5p and miR-99a* have shown higher expression levels in responders. Using these miRNAs 9 of 10 responders and 9 of 10 non-responders (p < 0.05) have been correctly classified. Our pilot study suggests that miRNAs are part of the mechanisms that are involved in response of rectal cancer to the chemoradiotherapy and that miRNAs may be promising predictive biomarkers for such patients. In most miRNAs we identified (miR-215, miR-99a*, miR-196b, miR-450b-5p and let-7e), the connection between their expression and radioresistance or chemoresistance to inhibitors of thymidylate synthetase was already established

Human colon cancer profiles show differential microRNA expression depending on mismatch repair status and are characteristic of undifferentiated proliferative states

International Nuclear Information System (INIS)

Sarver, Aaron L; Cunningham, Julie M; Subramanian, Subbaya; Wang, Liang; Smyrk, Tom C; Rodrigues, Cecilia MP; Thibodeau, Stephen N; Steer, Clifford J; French, Amy J; Borralho, Pedro M; Thayanithy, Venugopal; Oberg, Ann L; Silverstein, Kevin AT; Morlan, Bruce W; Riska, Shaun M; Boardman, Lisa A

2009-01-01

Colon cancer arises from the accumulation of multiple genetic and epigenetic alterations to normal colonic tissue. microRNAs (miRNAs) are small, non-coding regulatory RNAs that post-transcriptionally regulate gene expression. Differential miRNA expression in cancer versus normal tissue is a common event and may be pivotal for tumor onset and progression. To identify miRNAs that are differentially expressed in tumors and tumor subtypes, we carried out highly sensitive expression profiling of 735 miRNAs on samples obtained from a statistically powerful set of tumors (n = 80) and normal colon tissue (n = 28) and validated a subset of this data by qRT-PCR. Tumor specimens showed highly significant and large fold change differential expression of the levels of 39 miRNAs including miR-135b, miR-96, miR-182, miR-183, miR-1, and miR-133a, relative to normal colon tissue. Significant differences were also seen in 6 miRNAs including miR-31 and miR-592, in the direct comparison of tumors that were deficient or proficient for mismatch repair. Examination of the genomic regions containing differentially expressed miRNAs revealed that they were also differentially methylated in colon cancer at a far greater rate than would be expected by chance. A network of interactions between these miRNAs and genes associated with colon cancer provided evidence for the role of these miRNAs as oncogenes by attenuation of tumor suppressor genes. Colon tumors show differential expression of miRNAs depending on mismatch repair status. miRNA expression in colon tumors has an epigenetic component and altered expression that may reflect a reversion to regulatory programs characteristic of undifferentiated proliferative developmental states

Plasma microRNA profiles distinguish lethal injury in acetaminophen toxicity: A research study

Institute of Scientific and Technical Information of China (English)

Jeanine Ward; Shashi Bala; Jan Petrasek; Gyongyi Szabo

2012-01-01

AIM:To investigate plasma microRNA (miRNA) profiles indicative of hepatotoxicity in the setting of lethal acetaminophen (APAP) toxicity in mice.METHODS:Using plasma from APAP poisoned mice,either lethally (500 mg/kg) or sublethally (150 mg/kg) dosed,we screened commercially available murine microRNA libraries (SABiosciences,Qiagen Sciences,MD) to evaluate for unique miRNA profiles between these two dosing parameters.RESULTS:We distinguished numerous,unique plasma miRNAs both up- and downregulated in lethally compared to sublethally dosed mice.Of note,many of the greatest up- and downregulated miRNAs,namely 574-5p,466g,466f-3p,375,29c,and 148a,have been shown to be associated with asthma in prior studies.Interestingly,a relationship between APAP and asthma has been previously well described in the literature,with an as yet unknown mechanism of pathology.There was a statistically significant increase in alanine aminotransferase levels in the lethal compared to sublethal APAP dosing groups at the 12 h time point (P ＜0.001).There was 90％ mortality in the lethally compared to sublethally dosed mice at the 48 h time point (P =0.011).CONCLUSION:We identified unique plasma miRNAs both up- and downregulated in APAP poisoning which are correlated to asthma development.

A genomic portrait of the genetic architecture and regulatory impact of microRNA expression in response to infection.

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Siddle, Katherine J; Deschamps, Matthieu; Tailleux, Ludovic; Nédélec, Yohann; Pothlichet, Julien; Lugo-Villarino, Geanncarlo; Libri, Valentina; Gicquel, Brigitte; Neyrolles, Olivier; Laval, Guillaume; Patin, Etienne; Barreiro, Luis B; Quintana-Murci, Lluís

2014-05-01

MicroRNAs (miRNAs) are critical regulators of gene expression, and their role in a wide variety of biological processes, including host antimicrobial defense, is increasingly well described. Consistent with their diverse functional effects, miRNA expression is highly context dependent and shows marked changes upon cellular activation. However, the genetic control of miRNA expression in response to external stimuli and the impact of such perturbations on miRNA-mediated regulatory networks at the population level remain to be determined. Here we assessed changes in miRNA expression upon Mycobacterium tuberculosis infection and mapped expression quantitative trait loci (eQTL) in dendritic cells from a panel of healthy individuals. Genome-wide expression profiling revealed that ∼40% of miRNAs are differentially expressed upon infection. We find that the expression of 3% of miRNAs is controlled by proximate genetic factors, which are enriched in a promoter-specific histone modification associated with active transcription. Notably, we identify two infection-specific response eQTLs, for miR-326 and miR-1260, providing an initial assessment of the impact of genotype-environment interactions on miRNA molecular phenotypes. Furthermore, we show that infection coincides with a marked remodeling of the genome-wide relationships between miRNA and mRNA expression levels. This observation, supplemented by experimental data using the model of miR-29a, sheds light on the role of a set of miRNAs in cellular responses to infection. Collectively, this study increases our understanding of the genetic architecture of miRNA expression in response to infection, and highlights the wide-reaching impact of altering miRNA expression on the transcriptional landscape of a cell.

Heart structure-specific transcriptomic atlas reveals conserved microRNA-mRNA interactions.

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Vacchi-Suzzi, Caterina; Hahne, Florian; Scheubel, Philippe; Marcellin, Magali; Dubost, Valerie; Westphal, Magdalena; Boeglen, Catherine; Büchmann-Møller, Stine; Cheung, Ming Sin; Cordier, André; De Benedetto, Christopher; Deurinck, Mark; Frei, Moritz; Moulin, Pierre; Oakeley, Edward; Grenet, Olivier; Grevot, Armelle; Stull, Robert; Theil, Diethilde; Moggs, Jonathan G; Marrer, Estelle; Couttet, Philippe

2013-01-01

MicroRNAs are short non-coding RNAs that regulate gene expression at the post-transcriptional level and play key roles in heart development and cardiovascular diseases. Here, we have characterized the expression and distribution of microRNAs across eight cardiac structures (left and right ventricles, apex, papillary muscle, septum, left and right atrium and valves) in rat, Beagle dog and cynomolgus monkey using microRNA sequencing. Conserved microRNA signatures enriched in specific heart structures across these species were identified for cardiac valve (miR-let-7c, miR-125b, miR-127, miR-199a-3p, miR-204, miR-320, miR-99b, miR-328 and miR-744) and myocardium (miR-1, miR-133b, miR-133a, miR-208b, miR-30e, miR-499-5p, miR-30e*). The relative abundance of myocardium-enriched (miR-1) and valve-enriched (miR-125b-5p and miR-204) microRNAs was confirmed using in situ hybridization. MicroRNA-mRNA interactions potentially relevant for cardiac functions were explored using anti-correlation expression analysis and microRNA target prediction algorithms. Interactions between miR-1/Timp3, miR-125b/Rbm24, miR-204/Tgfbr2 and miR-208b/Csnk2a2 were identified and experimentally investigated in human pulmonary smooth muscle cells and luciferase reporter assays. In conclusion, we have generated a high-resolution heart structure-specific mRNA/microRNA expression atlas for three mammalian species that provides a novel resource for investigating novel microRNA regulatory circuits involved in cardiac molecular physiopathology.

Heart structure-specific transcriptomic atlas reveals conserved microRNA-mRNA interactions.

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Caterina Vacchi-Suzzi

Full Text Available MicroRNAs are short non-coding RNAs that regulate gene expression at the post-transcriptional level and play key roles in heart development and cardiovascular diseases. Here, we have characterized the expression and distribution of microRNAs across eight cardiac structures (left and right ventricles, apex, papillary muscle, septum, left and right atrium and valves in rat, Beagle dog and cynomolgus monkey using microRNA sequencing. Conserved microRNA signatures enriched in specific heart structures across these species were identified for cardiac valve (miR-let-7c, miR-125b, miR-127, miR-199a-3p, miR-204, miR-320, miR-99b, miR-328 and miR-744 and myocardium (miR-1, miR-133b, miR-133a, miR-208b, miR-30e, miR-499-5p, miR-30e*. The relative abundance of myocardium-enriched (miR-1 and valve-enriched (miR-125b-5p and miR-204 microRNAs was confirmed using in situ hybridization. MicroRNA-mRNA interactions potentially relevant for cardiac functions were explored using anti-correlation expression analysis and microRNA target prediction algorithms. Interactions between miR-1/Timp3, miR-125b/Rbm24, miR-204/Tgfbr2 and miR-208b/Csnk2a2 were identified and experimentally investigated in human pulmonary smooth muscle cells and luciferase reporter assays. In conclusion, we have generated a high-resolution heart structure-specific mRNA/microRNA expression atlas for three mammalian species that provides a novel resource for investigating novel microRNA regulatory circuits involved in cardiac molecular physiopathology.

The microRNA and messengerRNA profile of the RNA-induced silencing complex in human primary astrocyte and astrocytoma cells.

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Moser, Joanna J; Fritzler, Marvin J

2010-10-18

GW/P bodies are cytoplasmic ribonucleoprotein-rich foci involved in microRNA (miRNA)-mediated messenger RNA (mRNA) silencing and degradation. The mRNA regulatory functions within GW/P bodies are mediated by GW182 and its binding partner hAgo2 that bind miRNA in the RNA-induced silencing complex (RISC). To date there are no published reports of the profile of miRNA and mRNA targeted to the RISC or a comparison of the RISC-specific miRNA/mRNA profile differences in malignant and non-malignant cells. RISC mRNA and miRNA components were profiled by microarray analysis of malignant human U-87 astrocytoma cells and its non-malignant counterpart, primary human astrocytes. Total cell RNA as well as RNA from immunoprecipitated RISC was analyzed. The novel findings were fourfold: (1) miRNAs were highly enriched in astrocyte RISC compared to U-87 astrocytoma RISC, (2) astrocytoma and primary astrocyte cells each contained unique RISC miRNA profiles as compared to their respective cellular miRNA profiles, (3) miR-195, 10b, 29b, 19b, 34a and 455-3p levels were increased and the miR-181b level was decreased in U-87 astrocytoma RISC as compared to astrocyte RISC, and (4) the RISC contained decreased levels of mRNAs in primary astrocyte and U-87 astrocytoma cells. The observation that miR-34a and miR-195 levels were increased in the RISC of U-87 astrocytoma cells suggests an oncogenic role for these miRNAs. Differential regulation of mRNAs by specific miRNAs is evidenced by the observation that three miR34a-targeted mRNAs and two miR-195-targeted mRNAs were downregulated while one miR-195-targeted mRNA was upregulated. Biological pathway analysis of RISC mRNA components suggests that the RISC plays a pivotal role in malignancy and other conditions. This study points to the importance of the RISC and ultimately GW/P body composition and function in miRNA and mRNA deregulation in astrocytoma cells and possibly in other malignancies.

The microRNA and messengerRNA profile of the RNA-induced silencing complex in human primary astrocyte and astrocytoma cells.

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Joanna J Moser

2010-10-01

Full Text Available GW/P bodies are cytoplasmic ribonucleoprotein-rich foci involved in microRNA (miRNA-mediated messenger RNA (mRNA silencing and degradation. The mRNA regulatory functions within GW/P bodies are mediated by GW182 and its binding partner hAgo2 that bind miRNA in the RNA-induced silencing complex (RISC. To date there are no published reports of the profile of miRNA and mRNA targeted to the RISC or a comparison of the RISC-specific miRNA/mRNA profile differences in malignant and non-malignant cells.RISC mRNA and miRNA components were profiled by microarray analysis of malignant human U-87 astrocytoma cells and its non-malignant counterpart, primary human astrocytes. Total cell RNA as well as RNA from immunoprecipitated RISC was analyzed. The novel findings were fourfold: (1 miRNAs were highly enriched in astrocyte RISC compared to U-87 astrocytoma RISC, (2 astrocytoma and primary astrocyte cells each contained unique RISC miRNA profiles as compared to their respective cellular miRNA profiles, (3 miR-195, 10b, 29b, 19b, 34a and 455-3p levels were increased and the miR-181b level was decreased in U-87 astrocytoma RISC as compared to astrocyte RISC, and (4 the RISC contained decreased levels of mRNAs in primary astrocyte and U-87 astrocytoma cells.The observation that miR-34a and miR-195 levels were increased in the RISC of U-87 astrocytoma cells suggests an oncogenic role for these miRNAs. Differential regulation of mRNAs by specific miRNAs is evidenced by the observation that three miR34a-targeted mRNAs and two miR-195-targeted mRNAs were downregulated while one miR-195-targeted mRNA was upregulated. Biological pathway analysis of RISC mRNA components suggests that the RISC plays a pivotal role in malignancy and other conditions. This study points to the importance of the RISC and ultimately GW/P body composition and function in miRNA and mRNA deregulation in astrocytoma cells and possibly in other malignancies.

MicroRNA-196a & microRNA-101 expression in Barrett's oesophagus in patients with medically and surgically treated gastro-oesophageal reflux

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Bright Tim

2011-02-01

Full Text Available Abstract Background Proton pump inhibitor (PPI medication and surgical fundoplication are used for the control of gastro-oesophageal reflux in patients with Barrett's oesophagus, but differ in their effectiveness for both acid and bile reflux. This might impact on the inflammatory processes that are associated with progression of Barrett's oesophagus to cancer, and this may be evident in the gene expression profile and microRNA expression pattern in Barrett's oesophagus mucosa. We hypothesised that two miRNAs with inflammatory and oncogenic roles, miR-101 and miR-196a, are differentially expressed in Barrett's oesophagus epithelium in patients with reflux treated medically vs. surgically. Findings Mucosal tissue was obtained at endoscopy from patients with Barrett's oesophagus whose reflux was controlled by proton pump inhibitor (PPI therapy (n = 20 or by fundoplication (n = 19. RNA was extracted and the expression of miR-101 and miR-196a was measured using real-time reverse transcription - polymerase chain reaction. There were no significant differences in miR-101 and miR-196a expression in Barrett's oesophagus epithelium in patients treated by PPI vs. fundoplication (p = 0.768 and 0.211 respectively. Secondary analysis showed a correlation between miR-196a expression and Barrett's oesophagus segment length (p = 0.014. Conclusion The method of reflux treatment did not influence the expression of miR-101 and miR-196a in Barrett's oesophagus. This data does not provide support to the hypothesis that surgical treatment of reflux better prevents cancer development in Barrett's oesophagus. The association between miR-196a expression and Barrett's oesophagus length is consistent with a tumour promoting role for miR-196a in Barrett's oesophagus.

MicroRNA-196a & microRNA-101 expression in Barrett's oesophagus in patients with medically and surgically treated gastro-oesophageal reflux

Science.gov (United States)

2011-01-01

Background Proton pump inhibitor (PPI) medication and surgical fundoplication are used for the control of gastro-oesophageal reflux in patients with Barrett's oesophagus, but differ in their effectiveness for both acid and bile reflux. This might impact on the inflammatory processes that are associated with progression of Barrett's oesophagus to cancer, and this may be evident in the gene expression profile and microRNA expression pattern in Barrett's oesophagus mucosa. We hypothesised that two miRNAs with inflammatory and oncogenic roles, miR-101 and miR-196a, are differentially expressed in Barrett's oesophagus epithelium in patients with reflux treated medically vs. surgically. Findings Mucosal tissue was obtained at endoscopy from patients with Barrett's oesophagus whose reflux was controlled by proton pump inhibitor (PPI) therapy (n = 20) or by fundoplication (n = 19). RNA was extracted and the expression of miR-101 and miR-196a was measured using real-time reverse transcription - polymerase chain reaction. There were no significant differences in miR-101 and miR-196a expression in Barrett's oesophagus epithelium in patients treated by PPI vs. fundoplication (p = 0.768 and 0.211 respectively). Secondary analysis showed a correlation between miR-196a expression and Barrett's oesophagus segment length (p = 0.014). Conclusion The method of reflux treatment did not influence the expression of miR-101 and miR-196a in Barrett's oesophagus. This data does not provide support to the hypothesis that surgical treatment of reflux better prevents cancer development in Barrett's oesophagus. The association between miR-196a expression and Barrett's oesophagus length is consistent with a tumour promoting role for miR-196a in Barrett's oesophagus. PMID:21352563

Evaluation of microRNA stability in plasma and serum from healthy dogs

DEFF Research Database (Denmark)

Enelund, Lars; Nielsen, Lise Nikolic; Cirera, Susanna

2017-01-01

. METHODS: The levels of four microRNAs (cfa-let-7a, cfa-miR-16, cfa-miR-23a and cfa-miR-26a) known to be stably expressed from other canine studies, have been measured by real-time quantitative PCR. RESULTS: MicroRNA levels were found sufficiently stable for gene profiling in serum- and plasma stored...

Small RNA sequencing reveals metastasis-related microRNAs in lung adenocarcinoma

DEFF Research Database (Denmark)

Daugaard, Iben; Venø, Morten T.; Yan, Yan

2017-01-01

The majority of lung cancer deaths are caused by metastatic disease. MicroRNAs (miRNAs) are posttranscriptional regulators of gene expression and miRNA dysregulation can contribute to metastatic progression. Here, small RNA sequencing was used to profile the miRNA and piwi-interacting RNA (piRNA......) transcriptomes in relation to lung cancer metastasis. RNA-seq was performed using RNA extracted from formalin-fixed paraffin embedded (FFPE) lung adenocarcinomas (LAC) and brain metastases from 8 patients, and LACs from 8 patients without detectable metastatic disease. Impact on miRNA and piRNA transcriptomes...... was subtle with 9 miRNAs and 8 piRNAs demonstrating differential expression between metastasizing and non-metastasizing LACs. For piRNAs, decreased expression of piR-57125 was the most significantly associated with distant metastasis. Validation by RT-qPCR in a LAC cohort comprising 52 patients confirmed...

Cesium Toxicity Alters MicroRNA Processing and AGO1 Expressions in Arabidopsis thaliana.

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Il Lae Jung

Full Text Available MicroRNAs (miRNAs are short RNA fragments that play important roles in controlled gene silencing, thus regulating many biological processes in plants. Recent studies have indicated that plants modulate miRNAs to sustain their survival in response to a variety of environmental stimuli, such as biotic stresses, cold, drought, nutritional starvation, and toxic heavy metals. Cesium and radio-cesium contaminations have arisen as serious problems that both impede plant growth and enter the food chain through contaminated plants. Many studies have been performed to define plant responses against cesium intoxication. However, the complete profile of miRNAs in plants during cesium intoxication has not been established. Here we show the differential expression of the miRNAs that are mostly down-regulated during cesium intoxication. Furthermore, we found that cesium toxicity disrupts both the processing of pri-miRNAs and AGONOUTE 1 (AGO1-mediated gene silencing. AGO 1 seems to be especially destabilized by cesium toxicity, possibly through a proteolytic regulatory pathway. Our study presents a comprehensive profile of cesium-responsive miRNAs, which is distinct from that of potassium, and suggests two possible mechanisms underlying the cesium toxicity on miRNA metabolism.

MicroRNA expression profiles from eggs of different qualities associated with post-ovulatory ageing in rainbow trout (Oncorhynchus mykiss)

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Background: Egg quality is an important aspect in rainbow trout farming. Post-ovulatory aging is one of the most important factors affecting egg quality. MicroRNAs (miRNAs) are the major regulators in various biological processes and their expression profiles could serve as reliable biomarkers for v...

Evaluation of a new high-dimensional miRNA profiling platform

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Lamblin Anne-Francoise

2009-08-01

Full Text Available Abstract Background MicroRNAs (miRNAs are a class of approximately 22 nucleotide long, widely expressed RNA molecules that play important regulatory roles in eukaryotes. To investigate miRNA function, it is essential that methods to quantify their expression levels be available. Methods We evaluated a new miRNA profiling platform that utilizes Illumina's existing robust DASL chemistry as the basis for the assay. Using total RNA from five colon cancer patients and four cell lines, we evaluated the reproducibility of miRNA expression levels across replicates and with varying amounts of input RNA. The beta test version was comprised of 735 miRNA targets of Illumina's miRNA profiling application. Results Reproducibility between sample replicates within a plate was good (Spearman's correlation 0.91 to 0.98 as was the plate-to-plate reproducibility replicates run on different days (Spearman's correlation 0.84 to 0.98. To determine whether quality data could be obtained from a broad range of input RNA, data obtained from amounts ranging from 25 ng to 800 ng were compared to those obtained at 200 ng. No effect across the range of RNA input was observed. Conclusion These results indicate that very small amounts of starting material are sufficient to allow sensitive miRNA profiling using the Illumina miRNA high-dimensional platform. Nonlinear biases were observed between replicates, indicating the need for abundance-dependent normalization. Overall, the performance characteristics of the Illumina miRNA profiling system were excellent.

MicroRNA Expression in Laser Micro-dissected Breast Cancer Tissue Samples - a Pilot Study.

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Seclaman, Edward; Narita, Diana; Anghel, Andrei; Cireap, Natalia; Ilina, Razvan; Sirbu, Ioan Ovidiu; Marian, Catalin

2017-10-28

Breast cancer continues to represent a significant public health burden despite outstanding research advances regarding the molecular mechanisms of cancer biology, biomarkers for diagnostics and prognostic and therapeutic management of this disease. The studies of micro RNAs in breast cancer have underlined their potential as biomarkers and therapeutic targets; however most of these studies are still done on largely heterogeneous whole breast tissue samples. In this pilot study we have investigated the expression of four micro RNAs (miR-21, 145, 155, 92) known to be involved in breast cancer, in homogenous cell populations collected by laser capture microdissection from breast tissue section slides. Micro RNA expression was assessed by real time PCR, and associations with clinical and pathological characteristics were also explored. Our results have confirmed previous associations of miR-21 expression with poor prognosis characteristics of breast cancers such as high stage, large and highly proliferative tumors. No statistically significant associations were found with the other micro RNAs investigated, possibly due to the small sample size of our study. Our results also suggest that miR-484 could be a suitable endogenous control for data normalization in breast tissues, these results needing further confirmation by future studies. In summary, our pilot study showed the feasibility of detecting micro RNAs expression in homogenous laser captured microdissected invasive breast cancer samples, and confirmed some of the previously reported associations with poor prognostic characteristics of breast tumors.

Regulatory roles and therapeutic potential of microRNA in sarcoma.

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Lim, Hui Jun; Yang, Jia-Lin

2016-01-01

MicroRNAs (miRNAs) are single-stranded noncoding RNAs involved in various biological processes, including cell differentiation and development. They play multiple key roles as tumour suppressors, oncogenes or both in particular cases. This review aims to summarise current findings of the expression of miRNAs and their role in clinical oncology. Current knowledge regarding the involvement of miRNAs in different sarcoma subtypes will be assessed, in conjunction with their potential application as therapeutic targets. Relevant articles in scientific databases were identified using a combination of search terms, including "microRNA," "deregulation," "sarcoma," and "targeted therapy". These databases included Medline, Embase, Cochrane Review, Pubmed and Scopus. Aberrant miRNA expression patterns have been identified in a range of sarcoma subtypes, and differences in miRNA expression profiles between malignant cells and their normal counterparts suggests that miRNAs play key roles in sarcoma development. The identification of unique miRNA patterns in individual tumour types could possibly be used as a diagnostic tool in sarcoma. Moreover, identification of these miRNAs provides novel targets for the development of therapeutic strategies in distinct sarcoma subtypes. miRNAs hold significant potential as diagnostic biomarkers, as well as therapeutic targets in sarcoma. Possible future clinical applications include the use of miRNA pathways as therapeutic targets or miRNA expression profiling as a means of patient selection. The involvement miRNAs will undoubtedly contribute to the advancement of future targeted therapeutic interventions in sarcoma, and further establishment of appropriate delivery systems is vital for their use in clinical settings. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Modulation of microRNA activity by semi-microRNAs (smiRNAs

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Isabelle ePlante

2012-06-01

Full Text Available The ribonuclease Dicer plays a central role in the microRNA pathway by catalyzing the formation of 19 to 24-nucleotide (nt long microRNAs. Subsequently incorporated into Ago2 effector complexes, microRNAs are known to regulate messenger RNA (mRNA translation. Whether shorter RNA species derived from microRNAs exist and play a role in mRNA regulation remains unknown. Here, we report the serendipitous discovery of a 12-nt long RNA species corresponding to the 5’ region of the microRNA let-7, and tentatively termed semi-microRNA, or smiRNA. Using a smiRNA derived from the precursor of miR-223 as a model, we show that 12-nt long smiRNA species are devoid of any direct mRNA regulatory activity, as assessed in a reporter gene activity assay in transfected cultured human cells. However, smiR-223 was found to modulate the ability of the microRNA from which it derives to mediate translational repression or cleavage of reporter mRNAs. Our findings suggest that smiRNAs may be generated along the microRNA pathway and participate to the control of gene expression by regulating the activity of the related full-length mature microRNA in vivo.

Comparison of microRNA profiles between benign and malignant salivary gland tumors in tissue, blood and saliva samples: a prospective, case-control study

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Cinpolat, Ovgu; Unal, Zeynep Nil; Ismi, Onur; Gorur, Aysegul; Unal, Murat

2017-01-01

Abstract Introduction: Salivary gland tumors (SGTs) are rare head and neck malignancies consisting of a spectrum of tumors with different biological behaviors. Objective: In this study we aimed to find out differential expression of microRNA profiles between benign and malignant SGTs. Methods: We investigated the possible role of 95 microRNAs in the 20 patients with salivary gland tumors with comparison of 17 patients without malignancy or salivary gland diseases. Sixteen of the tumors wer...

Genomic analysis of microRNA time-course expression in liver of mice treated with genotoxic carcinogen N-ethyl-N-nitrosourea

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Guo Lei

2010-10-01

Full Text Available Abstract Background Dysregulated expression of microRNAs (miRNAs has been previously observed in human cancer tissues and shown promise in defining tumor status. However, there is little information as to if or when expression changes of miRNAs occur in normal tissues after carcinogen exposure. Results To explore the possible time-course changes of miRNA expression induced by a carcinogen, we treated mice with one dose of 120 mg/kg N-ethyl-N-nitrosourea (ENU, a model genotoxic carcinogen, and vehicle control. The miRNA expression profiles were assessed in the mouse livers in a time-course design. miRNAs were isolated from the livers at days 1, 3, 7, 15, 30 and 120 after the treatment and their expression was determined using a miRNA PCR Array. Principal component analysis of the miRNA expression profiles showed that miRNA expression at post-treatment days (PTDs 7 and 15 were different from those at the other time points and the control. The number of differentially expressed miRNAs (DEMs changed over time (3, 5, 14, 32, 5 and 5 at PTDs 1, 3, 7, 15, 30 and 120, respectively. The magnitude of the expression change varied with time with the highest changes at PTDs 7 or 15 for most of the DEMs. In silico functional analysis of the DEMs at PTDs 7 and 15 indicated that the major functions of these ENU-induced DEMs were associated with DNA damage, DNA repair, apoptosis and other processes related to carcinogenesis. Conclusion Our results showed that many miRNAs changed their expression to respond the exposure of the genotoxic carcinogen ENU and the number and magnitude of the changes were highest at PTDs 7 to 15. Thus, one to two weeks after the exposure is the best time for miRNA expression sampling.

MicroRNA-21 Increases Proliferation and Cisplatin Sensitivity of Osteosarcoma-Derived Cells.

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Vanita Vanas

Full Text Available Osteosarcoma is the most common primary bone tumor and poor prognosis for osteosarcoma patients is mainly due to chemotherapy resistance. MicroRNAs are important to maintain pathophysiological mechanisms of cancer and influence cell sensitivity to chemotherapy. In this study, we tested the functions of microRNA-21 for malignant features as well as for drug resistance of osteosarcoma. We used Northern blot to measure microRNA-21 levels in osteosarcoma-derived cell lines. MicroRNA-21 activity was modulated by either expressing a sponge to decrease its activity in an osteosarcoma-derived cell line expressing high levels of microRNA-21 or by introducing pri-microRNA-21 in a cell line with low endogenous levels. Cell migration was determined in a scratch assay and cell proliferation was measured by performing growth curve analysis. Sensitivity of the cells towards chemotherapeutics was investigated by performing cell viability assays and calculating the IC50 values. While cell migration was unaffected by modulated microRNA-21 levels, microRNA-21 inhibition slowed proliferation and exogenously expressed microRNA-21 promoted this process. Modulated microRNA-21 activity failed to effect sensitivity of osteosarcoma-derived cell lines to doxorubicin or methotrexate. Contrarily, reduction of microRNA-21 activity resulted in enhanced resistance towards cisplatin while ectopic expression of microRNA-21 showed the opposite effect. Increased microRNA-21 levels repressed the expression of Sprouty2 and ectopic expression of Sprouty2 was able to largely rescue the observed effects of microRNA-21 in osteosarcoma. In summary, our data indicate that in osteosarcoma microRNA-21 expression is an important component for regulation of cell proliferation and for determining sensitivity to cisplatin.

[Expression profiles and bioinformatic analysis of miRNA in human dental pulp cells during endothelial differentiation].

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Gong, Qimei; Jiang, Hongwei; Wang, Jinming; Ling, Junqi

2014-05-01

To investigate the differential expression profile and bioinformatic analysis of microRNA (miRNA) in human dental pulp cells (DPC) during endothelial differentiation. DPC were cultured in endothelial induction medium (50 µg/L vascular endothelial growth factor, 10 µg/L basic fibroblast growth factor and 2% fetal calf serum) for 7 days. Meanwhile non-induced DPC were used as control.Quantitative real-time PCR (qRT-PCR) was applied to detect vascular endothelial marker genes [CD31, von Willebrand factor (vWF) and vascular endothelial-cadherin (VE-cadherin)] and in vitro tube formation on matrigel was used to analyze the angiogenic ability of differentiated cells. And then miRNA expression profiles of DPC were examined using miRNA microarray and then the differentially expressed miRNA were validated by qRT-PCR. Furthermore, bioinformatic analysis was employed to predict the target genes of miRNA and to analyze the possible biological functions and signaling pathways that were involved in DPC after induction. The relative mRNA level of CD31, vWF and VE-cadherin in the control group were (3.48 ± 0.22) ×10(-4), (3.13 ± 0.31) ×10(-4) and (39.60 ± 2.36) ×10(-4), and (19.57 ± 2.20) ×10(-4), (48.13 ± 0.54) ×10(-4) and (228.00 ± 8.89) ×10(-4) in the induced group. The expressions of CD31, vWF and VE-cadherin were increased significantly in endothelial induced DPC compared to the control group (P functions, such as the regulation of transcription, cell motion, blood vessel morphogenesis, angiogenesis and cytoskeletal protein, and signaling pathways including the mitogen-activated protein kinase (MAPK) and the Wnt signaling pathway. The differential miRNA expression identified in this study may be involved in governing DPC endothelial differentiation, thus contributing to the future research on regulatory mechanisms in dental pulp angiogenesis.

An Update on Cellular MicroRNA Expression in Human Papillomavirus-Associated Head and Neck Squamous Cell Carcinoma.

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Emmett, Sarah; Whiteman, David C; Panizza, Benedict J; Antonsson, Annika

2018-06-19

Squamous cell carcinoma of mucosal sites in the head and neck (HNSCC) is the sixth most common cause of cancer worldwide, and despite advances in conventional management, it still has significant morbidity and mortality associated with both diagnosis and treatment. Advances in our understanding of the biological mechanisms underlying this disease have demonstrated a significant difference between human papillomavirus (HPV)-associated, HPV and tobacco associated, and HPV-negative disease. It remains important to further elucidate the biologic and genetic differences between HPV-associated and tobacco-associated disease, with the aim of earlier diagnosis through screening, and advances in management including the development of novel therapeutic agents. MicroRNAs (miRNAs) are small, non-coding RNAs that function as post-transcriptional regulators of gene expression, and have effects on almost every cellular function, and have potentially important applications to diagnosis, management and prognosis in HNSCC. Establishing a cellular miRNA expression profile for HPV-associated disease may therefore have important implications for the screening and treatment of this disease. This review summarises the current findings regarding miRNA expression in mucosal HNSCC, and focuses particularly on miRNA expression in HPV-associated tumours. © 2018 S. Karger AG, Basel.

Effects of active acromegaly on bone mRNA and microRNA expression patterns.

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Belaya, Zhanna; Grebennikova, Tatiana; Melnichenko, Galina; Nikitin, Alexey; Solodovnikov, Alexander; Brovkina, Olga; Grigoriev, Andrey; Rozhinskaya, Liudmila; Lutsenko, Alexander; Dedov, Ivan

2018-04-01

To evaluate the response of bone to chronic long-term growth hormone (GH) and insulin-like growth factor-1 (IGF1) excess by measuring the expression of selected mRNA and microRNA (miR) in bone tissue samples of patients with active acromegaly. Case-control study. Bone tissue samples were obtained during transsphenoidal adenomectomy from the sphenoid bone (sella turcica) from 14 patients with clinically and biochemically confirmed acromegaly and 10 patients with clinically non-functioning pituitary adenoma (NFPA) matched by sex and age. Expression of genes involved in the regulation of bone remodeling was studied using quantitative polymerase chain reaction (qPCR). Of the genes involved in osteoblast and osteoclast activity, only alkaline phosphatase (ALP) mRNA was 50% downregulated in patients with acromegaly. GH excess caused increased expression of the Wnt signaling antagonists ( DKK1) and agonists ( WNT10B) and changes in the levels of miR involved in mesenchymal stem cell commitment to chondrocytes (miR-199a-5p) or adipocytes (miR-27-5p, miR-125b-5p, miR-34a-5p, miR-188-3p) P  Acromegaly had minimal effects on tested mRNAs specific to osteoblast or osteoclast function except for downregulated ALP expression. The expressions of miR known to be involved in mesenchymal stem cell commitment and downregulated TWIST1 expression suggest acromegaly has a negative effect on osteoblastogenesis. © 2018 European Society of Endocrinology.

Changes in miRNA expression profile of space-flown Caenorhabditis elegans during Shenzhou-8 mission

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Xu, Dan; Gao, Ying; Huang, Lei; Sun, Yeqing

2014-04-01

Recent advances in the field of molecular biology have demonstrated that small non-coding microRNAs (miRNAs) have a broad effect on gene expression networks and play a key role in biological responses to environmental stressors. However, little is known about how space radiation exposure and altered gravity affect miRNA expression. The "International Space Biological Experiments" project was carried out in November 2011 by an international collaboration between China and Germany during the Shenzhou-8 (SZ-8) mission. To study the effects of spaceflight on Caenorhabditis elegans (C. elegans), we explored the expression profile miRNA changes in space-flown C. elegans. Dauer C. elegans larvae were taken by SZ-8 spacecraft and experienced the 16.5-day shuttle spaceflight. We performed miRNA microarray analysis, and the results showed that 23 miRNAs were altered in a complex space environment and different expression patterns were observed in the space synthetic and radiation environments. Most putative target genes of the altered miRNAs in the space synthetic environment were predicted to be involved in developmental processes instead of in the regulation of transcription, and the enrichment of these genes was due to space radiation. Furthermore, integration analysis of the miRNA and mRNA expression profiles confirmed that twelve genes were differently regulated by seven miRNAs. These genes may be involved in embryonic development, reproduction, transcription factor activity, oviposition in a space synthetic environment, positive regulation of growth and body morphogenesis in a space radiation environment. Specifically, we found that cel-miR-52, -55, and -56 of the miR-51 family were sensitive to space environmental stressors and could regulate biological behavioural responses and neprilysin activity through the different isoforms of T01C4.1 and F18A12.8. These findings suggest that C. elegans responded to spaceflight by altering the expression of miRNAs and some target

MicroRNA 10a marks regulatory T cells

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Jeker, Lukas T; Zhou, Xuyu; Gershberg, Kseniya

2012-01-01

MicroRNAs (miRNAs) are crucial for regulatory T cell (Treg) stability and function. We report that microRNA-10a (miR-10a) is expressed in Tregs but not in other T cells including individual thymocyte subsets. Expression profiling in inbred mouse strains demonstrated that non-obese diabetic (NOD......) mice with a genetic susceptibility for autoimmune diabetes have lower Treg-specific miR-10a expression than C57BL/6J autoimmune resistant mice. Inhibition of miR-10a expression in vitro leads to reduced FoxP3 expression levels and miR-10a expression is lower in unstable "exFoxP3" T cells. Unstable...... and phenotype of natural Treg nor the capacity of conventional T cells to induce FoxP3 in response to TGFβ, RA, or a combination of the two. Thus, miR-10a is selectively expressed in Treg but inhibition by antagomiRs or genetic ablation resulted in discordant effects on FoxP3....

miRBase: integrating microRNA annotation and deep-sequencing data.

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Kozomara, Ana; Griffiths-Jones, Sam

2011-01-01

miRBase is the primary online repository for all microRNA sequences and annotation. The current release (miRBase 16) contains over 15,000 microRNA gene loci in over 140 species, and over 17,000 distinct mature microRNA sequences. Deep-sequencing technologies have delivered a sharp rise in the rate of novel microRNA discovery. We have mapped reads from short RNA deep-sequencing experiments to microRNAs in miRBase and developed web interfaces to view these mappings. The user can view all read data associated with a given microRNA annotation, filter reads by experiment and count, and search for microRNAs by tissue- and stage-specific expression. These data can be used as a proxy for relative expression levels of microRNA sequences, provide detailed evidence for microRNA annotations and alternative isoforms of mature microRNAs, and allow us to revisit previous annotations. miRBase is available online at: http://www.mirbase.org/.

Transient Gene and miRNA Expression Profile Changes of Confluent Human Fibroblast Cells in Space

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Zhang, Ye; Lu, Tao; Wong, Michael; Feiveson, Alan; Stodieck, Louis; Karouia, Fathi; Wang, Xiaoyu; Wu, Honglu

2015-01-01

Microgravity or an altered gravity environment from the static 1 gravitational constant has been shown to influence global gene expression patterns and protein levels in cultured cells. However, most of the reported studies conducted in space or using simulated microgravity on the ground have focused on the growth or differentiation of the cells. Whether non-dividing cultured cells will sense the presence of microgravity in space has not been specifically addressed. In an experiment conducted on the International Space Station, confluent human fibroblast cells were fixed after being cultured in space for 3 and 14 days for investigations of gene and miRNA (microRNA) expression profile changes in these cells. A fibroblast is a type of cell that synthesizes the extracellular matrix and collagen, the structural framework for tissues, and plays a critical role in wound healing and other functions. Results of the experiment showed that on Day 3, both the flown and ground cells were still proliferating slowly even though they were confluent, as measured by the expression of the protein Ki-67 positive cells, and the cells in space grew slightly faster. Gene and miRNA expression data indicated activation of NF(sub kappa)B (nuclear factor kappa-light-chain-enhancer of activated B cells) and other growth related pathways involving HGF and VEGF in the flown cells. On Day 14 when the cells were mostly non-dividing, the gene and miRNA expression profiles between the flight and ground samples were indistinguishable. Comparison of gene and miRNA expressions in the Day 3 samples in respect to Day 14 revealed that most of the changes observed on Day 3 were related to cell growth for both the flown and ground cells. Analysis of cytoskeleton changes by immunohistochemistry staining of the cells with antibodies for alpha-tubulin showed no difference between the flight and ground samples. Results of our study suggest that in true non-dividing human fibroblast cells, microgravity in

MicroRNA-99 family members suppress Homeobox A1 expression in epithelial cells.

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Chen, Dan; Chen, Zujian; Jin, Yi; Dragas, Dragan; Zhang, Leitao; Adjei, Barima S; Wang, Anxun; Dai, Yang; Zhou, Xiaofeng

2013-01-01

The miR-99 family is one of the evolutionarily most ancient microRNA families, and it plays a critical role in developmental timing and the maintenance of tissue identity. Recent studies, including reports from our group, suggested that the miR-99 family regulates various physiological processes in adult tissues, such as dermal wound healing, and a number of disease processes, including cancer. By combining 5 independent genome-wide expression profiling experiments, we identified a panel of 266 unique transcripts that were down-regulated in epithelial cells transfected with miR-99 family members. A comprehensive bioinformatics analysis using 12 different sequence-based microRNA target prediction algorithms revealed that 81 out of these 266 down-regulated transcripts are potential direct targets for the miR-99 family. Confirmation experiments and functional analyses were performed to further assess 6 selected miR-99 target genes, including mammalian Target of rapamycin (mTOR), Homeobox A1 (HOXA1), CTD small phosphatase-like (CTDSPL), N-myristoyltransferase 1 (NMT1), Transmembrane protein 30A (TMEM30A), and SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5 (SMARCA5). HOXA1 is a known proto-oncogene, and it also plays an important role in embryonic development. The direct targeting of the miR-99 family to two candidate binding sequences located in the HOXA1 mRNA was confirmed using a luciferase reporter gene assay and a ribonucleoprotein-immunoprecipitation (RIP-IP) assay. Ectopic transfection of miR-99 family reduced the expression of HOXA1, which, in consequence, down-regulated the expression of its downstream gene (i.e., Bcl-2) and led to reduced proliferation and cell migration, as well as enhanced apoptosis. In summary, we identified a number of high-confidence miR-99 family target genes, including proto-oncogene HOXA1, which may play an important role in regulating epithelial cell proliferation and migration during

MicroRNA-99 family members suppress Homeobox A1 expression in epithelial cells.

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Dan Chen

Full Text Available The miR-99 family is one of the evolutionarily most ancient microRNA families, and it plays a critical role in developmental timing and the maintenance of tissue identity. Recent studies, including reports from our group, suggested that the miR-99 family regulates various physiological processes in adult tissues, such as dermal wound healing, and a number of disease processes, including cancer. By combining 5 independent genome-wide expression profiling experiments, we identified a panel of 266 unique transcripts that were down-regulated in epithelial cells transfected with miR-99 family members. A comprehensive bioinformatics analysis using 12 different sequence-based microRNA target prediction algorithms revealed that 81 out of these 266 down-regulated transcripts are potential direct targets for the miR-99 family. Confirmation experiments and functional analyses were performed to further assess 6 selected miR-99 target genes, including mammalian Target of rapamycin (mTOR, Homeobox A1 (HOXA1, CTD small phosphatase-like (CTDSPL, N-myristoyltransferase 1 (NMT1, Transmembrane protein 30A (TMEM30A, and SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5 (SMARCA5. HOXA1 is a known proto-oncogene, and it also plays an important role in embryonic development. The direct targeting of the miR-99 family to two candidate binding sequences located in the HOXA1 mRNA was confirmed using a luciferase reporter gene assay and a ribonucleoprotein-immunoprecipitation (RIP-IP assay. Ectopic transfection of miR-99 family reduced the expression of HOXA1, which, in consequence, down-regulated the expression of its downstream gene (i.e., Bcl-2 and led to reduced proliferation and cell migration, as well as enhanced apoptosis. In summary, we identified a number of high-confidence miR-99 family target genes, including proto-oncogene HOXA1, which may play an important role in regulating epithelial cell proliferation and

Common features of microRNA target prediction tools

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Sarah M. Peterson

2014-02-01

Full Text Available The human genome encodes for over 1800 microRNAs, which are short noncoding RNA molecules that function to regulate gene expression post-transcriptionally. Due to the potential for one microRNA to target multiple gene transcripts, microRNAs are recognized as a major mechanism to regulate gene expression and mRNA translation. Computational prediction of microRNA targets is a critical initial step in identifying microRNA:mRNA target interactions for experimental validation. The available tools for microRNA target prediction encompass a range of different computational approaches, from the modeling of physical interactions to the incorporation of machine learning. This review provides an overview of the major computational approaches to microRNA target prediction. Our discussion highlights three tools for their ease of use, reliance on relatively updated versions of miRBase, and range of capabilities, and these are DIANA-microT-CDS, miRanda-mirSVR, and TargetScan. In comparison across all microRNA target prediction tools, four main aspects of the microRNA:mRNA target interaction emerge as common features on which most target prediction is based: seed match, conservation, free energy, and site accessibility. This review explains these features and identifies how they are incorporated into currently available target prediction tools. MicroRNA target prediction is a dynamic field with increasing attention on development of new analysis tools. This review attempts to provide a comprehensive assessment of these tools in a manner that is accessible across disciplines. Understanding the basis of these prediction methodologies will aid in user selection of the appropriate tools and interpretation of the tool output.

Human Milk MicroRNA and Total RNA Differ Depending on Milk Fractionation.

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Alsaweed, Mohammed; Hepworth, Anna R; Lefèvre, Christophe; Hartmann, Peter E; Geddes, Donna T; Hassiotou, Foteini

2015-10-01

MicroRNA have been recently discovered in human milk signifying potentially important functions for both the lactating breast and the infant. Whilst human milk microRNA have started to be explored, little data exist on the evaluation of sample processing, and analysis to ensure that a full spectrum of microRNA can be obtained. Human milk comprises three main fractions: cells, skim milk, and lipids. Typically, the skim milk fraction has been measured in isolation despite evidence that the lipid fraction may contain more microRNA. This study aimed to standardize isolation of microRNA and total RNA from all three fractions of human milk to determine the most appropriate sampling and analysis procedure for future studies. Three different methods from eight commercially available kits were tested for their efficacy in extracting total RNA and microRNA from the lipid, skim, and cell fractions of human milk. Each fraction yielded different concentrations of RNA and microRNA, with the highest quantities found in the cell and lipid fractions, and the lowest in skim milk. The column-based phenol-free method was the most efficient extraction method for all three milk fractions. Two microRNAs were expressed and validated in the three milk fractions by qPCR using the three recommended extraction kits for each fraction. High expression levels were identified in the skim and lipid milk factions for these microRNAs. These results suggest that careful consideration of both the human milk sample preparation and extraction protocols should be made prior to embarking upon research in this area. © 2015 The Authors. Journal of Cellular Biochemistry Published by Wiley Periodicals, Inc.

microRNA expression in the neural retina: Focus on Müller glia.

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Quintero, Heberto; Lamas, Mónica

2018-03-01

The neural retina hosts a unique specialized type of macroglial cell that not only preserves retinal homeostasis, function, and integrity but also may serve as a source of new neurons during regenerative processes: the Müller cell. Precise microRNA-driven mechanisms of gene regulation impel and direct the processes of Müller glia lineage acquisition from retinal progenitors during development, the triggering of their response to retinal degeneration and, in some cases, Müller cell reprogramming and regenerative events. In this review we survey the recent reports describing, through functional assays, the regulatory role of microRNAs in Müller cell physiology, differentiation potential, and retinal pathology. We discuss also the evidence based on expression analysis that points out the relevance of a Müller glia-specific microRNA signature that would orchestrate these processes. © 2017 Wiley Periodicals, Inc.

Analysis of microRNA profile of Anopheles sinensis by deep sequencing and bioinformatic approaches.

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Feng, Xinyu; Zhou, Xiaojian; Zhou, Shuisen; Wang, Jingwen; Hu, Wei

2018-03-12

microRNAs (miRNAs) are small non-coding RNAs widely identified in many mosquitoes. They are reported to play important roles in development, differentiation and innate immunity. However, miRNAs in Anopheles sinensis, one of the Chinese malaria mosquitoes, remain largely unknown. We investigated the global miRNA expression profile of An. sinensis using Illumina Hiseq 2000 sequencing. Meanwhile, we applied a bioinformatic approach to identify potential miRNAs in An. sinensis. The identified miRNA profiles were compared and analyzed by two approaches. The selected miRNAs from the sequencing result and the bioinformatic approach were confirmed with qRT-PCR. Moreover, target prediction, GO annotation and pathway analysis were carried out to understand the role of miRNAs in An. sinensis. We identified 49 conserved miRNAs and 12 novel miRNAs by next-generation high-throughput sequencing technology. In contrast, 43 miRNAs were predicted by the bioinformatic approach, of which two were assigned as novel. Comparative analysis of miRNA profiles by two approaches showed that 21 miRNAs were shared between them. Twelve novel miRNAs did not match any known miRNAs of any organism, indicating that they are possibly species-specific. Forty miRNAs were found in many mosquito species, indicating that these miRNAs are evolutionally conserved and may have critical roles in the process of life. Both the selected known and novel miRNAs (asi-miR-281, asi-miR-184, asi-miR-14, asi-miR-nov5, asi-miR-nov4, asi-miR-9383, and asi-miR-2a) could be detected by quantitative real-time PCR (qRT-PCR) in the sequenced sample, and the expression patterns of these miRNAs measured by qRT-PCR were in concordance with the original miRNA sequencing data. The predicted targets for the known and the novel miRNAs covered many important biological roles and pathways indicating the diversity of miRNA functions. We also found 21 conserved miRNAs and eight counterparts of target immune pathway genes in An. sinensis

MicroRNA genes preferentially expressed in dendritic cells contain sites for conserved transcription factor binding motifs in their promoters

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Huynen Martijn A

2011-06-01

Full Text Available Abstract Background MicroRNAs (miRNAs play a fundamental role in the regulation of gene expression by translational repression or target mRNA degradation. Regulatory elements in miRNA promoters are less well studied, but may reveal a link between their expression and a specific cell type. Results To explore this link in myeloid cells, miRNA expression profiles were generated from monocytes and dendritic cells (DCs. Differences in miRNA expression among monocytes, DCs and their stimulated progeny were observed. Furthermore, putative promoter regions of miRNAs that are significantly up-regulated in DCs were screened for Transcription Factor Binding Sites (TFBSs based on TFBS motif matching score, the degree to which those TFBSs are over-represented in the promoters of the up-regulated miRNAs, and the extent of conservation of the TFBSs in mammals. Conclusions Analysis of evolutionarily conserved TFBSs in DC promoters revealed preferential clustering of sites within 500 bp upstream of the precursor miRNAs and that many mRNAs of cognate TFs of the conserved TFBSs were indeed expressed in the DCs. Taken together, our data provide evidence that selected miRNAs expressed in DCs have evolutionarily conserved TFBSs relevant to DC biology in their promoters.

Liposomal curcumin alters chemosensitivity of breast cancer cells to Adriamycin via regulating microRNA expression.

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Zhou, Siying; Li, Jian; Xu, Hanzi; Zhang, Sijie; Chen, Xiu; Chen, Wei; Yang, Sujin; Zhong, Shanliang; Zhao, Jianhua; Tang, Jinhai

2017-07-30

Emerging evidence suggests that curcumin can overcome drug resistance to classical chemotherapies, but poor bioavailability and low absorption have limited its clinical use and the mechanisms remain unclear. Also, Adriamycin (Adr) is one of the most active cytotoxic agents in breast cancer; however, the high resistant rate of Adr leads to a poor prognosis. We utilized encapsulation in liposomes as a strategy to improve the bioavailability of curcumin and demonstrated that liposomal curcumin altered chemosensitivity of Adr-resistant MCF-7 human breast cancer (MCF-7/Adr) by MTT assay. The miRNA and mRNA expression profiles of MCF-7/S, MCF-7/Adr and curcumin-treated MCF-7/Adr cells were analyzed by microarray and further confirmed by real-time PCR. We focused on differentially expressed miR-29b-1-5p to explore the involvement of miR-29b-1-5p in the resistance of Adr. Candidate genes of dysregulated miRNAs were identified by prediction algorithms based on gene expression profiles. Networks of KEGG pathways were organized by the selected dysregulated miRNAs. Moreover, protein-protein interaction (PPI) was utilized to map protein interaction networks of curcumin regulated proteins. We first demonstrated liposomal curcumin could rescue part of Adriamycin resistance in breast cancer and further identified 67 differentially expressed microRNAs among MCF-7/S, MCF-7/Adr and curcumin-treated MCF-7/Adr. The results showed that lower expressed miR-29b-1-5p decreased the IC50 of MCF-7/Adr cells and higher expressed miR-29b-1-5p, weaken the effects of liposomal curcumin to Adr-resistance. Besides, we found that 20 target genes (mRNAs) of each dysregulated miRNA were not only predicted by prediction algorithms, but also differentially expressed in the microarray. The results showed that MAPK, mTOR, PI3K-Akt, AMPK, TNF, Ras signaling pathways and several target genes such as PPARG, RRM2, SRSF1and EPAS1, may associate with drug resistance of breast cancer cells to Adr. We determined

Plasma processing conditions substantially influence circulating microRNA biomarker levels.

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Cheng, Heather H; Yi, Hye Son; Kim, Yeonju; Kroh, Evan M; Chien, Jason W; Eaton, Keith D; Goodman, Marc T; Tait, Jonathan F; Tewari, Muneesh; Pritchard, Colin C

2013-01-01

Circulating, cell-free microRNAs (miRNAs) are promising candidate biomarkers, but optimal conditions for processing blood specimens for miRNA measurement remain to be established. Our previous work showed that the majority of plasma miRNAs are likely blood cell-derived. In the course of profiling lung cancer cases versus healthy controls, we observed a broad increase in circulating miRNA levels in cases compared to controls and that higher miRNA expression correlated with higher platelet and particle counts. We therefore hypothesized that the quantity of residual platelets and microparticles remaining after plasma processing might impact miRNA measurements. To systematically investigate this, we subjected matched plasma from healthy individuals to stepwise processing with differential centrifugation and 0.22 µm filtration and performed miRNA profiling. We found a major effect on circulating miRNAs, with the majority (72%) of detectable miRNAs substantially affected by processing alone. Specifically, 10% of miRNAs showed 4-30x variation, 46% showed 30-1,000x variation, and 15% showed >1,000x variation in expression solely from processing. This was predominantly due to platelet contamination, which persisted despite using standard laboratory protocols. Importantly, we show that platelet contamination in archived samples could largely be eliminated by additional centrifugation, even in frozen samples stored for six years. To minimize confounding effects in microRNA biomarker studies, additional steps to limit platelet contamination for circulating miRNA biomarker studies are necessary. We provide specific practical recommendations to help minimize confounding variation attributable to plasma processing and platelet contamination.

Microparticle conferred microRNA profiles - implications in the transfer and dominance of cancer traits

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Jaiswal Ritu

2012-06-01

Full Text Available Abstract Background Microparticles (MPs are membrane vesicles which are released from normal and malignant cells following a process of budding and detachment from donor cells. MPs contain surface antigens, proteins and genetic material and serve as vectors of intercellular communication. MPs comprise the major source of systemic RNA including microRNA (miRNA, the aberrant expression of which appears to be associated with stage, progression and spread of many cancers. Our previous study showed that MPs carry both transcripts and miRNAs associated with the acquisition of multidrug resistance in cancer. Results Herein, we expand on our previous finding and demonstrate that MPs carry the transcripts of the membrane vesiculation machinery (floppase and scramblase as well as nucleic acids encoding the enzymes essential for microRNA biogenesis (Drosha, Dicer and Argonaute. We also demonstrate using microarray miRNA profiling analysis, the selective packaging of miRNAs (miR-1228*, miR-1246, miR-1308, miR-149*, miR-455-3p, miR-638 and miR-923 within the MP cargo upon release from the donor cells. Conclusions These miRNAs are present in both haematological and non-haematological cancer cells and are involved in pathways implicated in cancer pathogenesis, membrane vesiculation and cascades regulated by ABC transporters. Our recent findings reinforce our earlier reports that MP transfer ‘re-templates’ recipient cells so as to reflect donor cell traits. We now demonstrate that this process is likely to occur via a process of selective packaging of nucleic acid species, including regulatory nucleic acids upon MP vesiculation. These findings have significant implications in understanding the cellular basis governing the intercellular acquisition and dominance of deleterious traits in cancers.

Diagnostic and Prognostic MicroRNA Biomarkers for Prostate Cancer in Cell-free Urine

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Fredsøe, Jacob Christian; Rasmussen, Anne Karin; Thomsen, Anni Rønfeldt

2017-01-01

Background: Widespread use of prostate-specific antigen (PSA) testing for prostate cancer (PC) detection has led to extensive overdiagnosis and overtreatment. Urine-based microRNA (miRNA) biomarkers could be useful in PC diagnosis and prognosis. Objective: To train and validate urine-based micro......RNA (miRNA) biomarkers that may assist in PC diagnosis and prognosis. Design, setting, and participants: We profiled the expression levels of 92 miRNAs via reverse transcriptase–poymerase chain reaction in cell-free urine samples from 29 patients with benign prostatic hyperplasia (BPH) and 215 patients...... could help in primary diagnosis of PC and guide treatment decisions. Further validation studies are warranted. Patient summary: Using two large patient cohorts, we searched for novel prostate cancer biomarkers in urine. We found two new sets of microRNA biomarkers in urine that could accurately predict...

Expression of human ARGONAUTE 2 inhibits endogenous microRNA activity in Arabidopsis

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Ira eDeveson

2013-04-01

Full Text Available Plant and animal microRNA (miRNA pathways share many analogous components, the ARGONAUTE (AGO proteins being foremost among them. We sought to ascertain the degree of functional conservation shared by Homo sapiens ARGONAUTE 2 (HsAGO2 and Arabidopsis thaliana ARGONAUTE 1 (AtAGO1, which are the predominant AGO family members involved with miRNA activity in their respective species. Transgenic Arabidopsis plants expressing HsAGO2 were indistinguishable from counterparts over-expressing AtAGO1, each group exhibiting the morphological and molecular hallmarks of miRNA-pathway loss-of-function alleles. However, unlike AtAGO1, HsAGO2 was unable to rescue the ago1-27 allele. We conclude that, despite the evolutionary gulf between them, HsAGO2 is likely capable of interacting with some component/s of the Arabidopsis miRNA pathway, thereby perturbing its operation, although differences have arisen such that HsAGO2 alone is insufficient to confer efficient silencing of miRNA targets in planta.

MicroRNA MiR-17 retards tissue growth and represses fibronectin expression.

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Shan, Sze Wan; Lee, Daniel Y; Deng, Zhaoqun; Shatseva, Tatiana; Jeyapalan, Zina; Du, William W; Zhang, Yaou; Xuan, Jim W; Yee, Siu-Pok; Siragam, Vinayakumar; Yang, Burton B

2009-08-01

MicroRNAs (miRNAs) are single-stranded regulatory RNAs, frequently expressed as clusters. Previous studies have demonstrated that the six-miRNA cluster miR-17~92 has important roles in tissue development and cancers. However, the precise role of each miRNA in the cluster is unknown. Here we show that overexpression of miR-17 results in decreased cell adhesion, migration and proliferation. Transgenic mice overexpressing miR-17 showed overall growth retardation, smaller organs and greatly reduced haematopoietic cell lineages. We found that fibronectin and the fibronectin type-III domain containing 3A (FNDC3A) are two targets that have their expression repressed by miR-17, both in vitro and in transgenic mice. Several lines of evidence support the notion that miR-17 causes cellular defects through its repression of fibronectin expression. Our single miRNA expression assay may be evolved to allow the manipulation of individual miRNA functions in vitro and in vivo. We anticipate that this could serve as a model for studying gene regulation by miRNAs in the development of gene therapy.

MicroRNA and protein profiling of brain metastasis competent cell-derived exosomes.

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Laura Camacho

Full Text Available Exosomes are small membrane vesicles released by most cell types including tumor cells. The intercellular exchange of proteins and genetic material via exosomes is a potentially effective approach for cell-to-cell communication and it may perform multiple functions aiding to tumor survival and metastasis. We investigated microRNA and protein profiles of brain metastatic (BM versus non-brain metastatic (non-BM cell-derived exosomes. We studied the cargo of exosomes isolated from brain-tropic 70W, MDA-MB-231BR, and circulating tumor cell brain metastasis-selected markers (CTC1BMSM variants, and compared them with parental non-BM MeWo, MDA-MB-231P and CTC1P cells, respectively. By performing microRNA PCR array we identified one up-regulated (miR-210 and two down-regulated miRNAs (miR-19a and miR-29c in BM versus non-BM exosomes. Second, we analyzed the proteomic content of cells and exosomes isolated from these six cell lines, and detected high expression of proteins implicated in cell communication, cell cycle, and in key cancer invasion and metastasis pathways. Third, we show that BM cell-derived exosomes can be internalized by non-BM cells and that they effectively transport their cargo into cells, resulting in increased cell adhesive and invasive potencies. These results provide a strong rationale for additional investigations of exosomal proteins and miRNAs towards more profound understandings of exosome roles in brain metastasis biogenesis, and for the discovery and application of non-invasive biomarkers for new therapies combating brain metastasis.

The effects of environmental chemical carcinogens on the microRNA machinery.

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Izzotti, A; Pulliero, A

2014-07-01

The first evidence that microRNA expression is early altered by exposure to environmental chemical carcinogens in still healthy organisms was obtained for cigarette smoke. To date, the cumulative experimental data indicate that similar effects are caused by a variety of environmental carcinogens, including polycyclic aromatic hydrocarbons, nitropyrenes, endocrine disruptors, airborne mixtures, carcinogens in food and water, and carcinogenic drugs. Accordingly, the alteration of miRNA expression is a general mechanism that plays an important pathogenic role in linking exposure to environmental toxic agents with their pathological consequences, mainly including cancer development. This review summarizes the existing experimental evidence concerning the effects of chemical carcinogens on the microRNA machinery. For each carcinogen, the specific microRNA alteration signature, as detected in experimental studies, is reported. These data are useful for applying microRNA alterations as early biomarkers of biological effects in healthy organisms exposed to environmental carcinogens. However, microRNA alteration results in carcinogenesis only if accompanied by other molecular damages. As an example, microRNAs altered by chemical carcinogens often inhibits the expression of mutated oncogenes. The long-term exposure to chemical carcinogens causes irreversible suppression of microRNA expression thus allowing the transduction into proteins of mutated oncogenes. This review also analyzes the existing knowledge regarding the mechanisms by which environmental carcinogens alter microRNA expression. The underlying molecular mechanism involves p53-microRNA interconnection, microRNA adduct formation, and alterations of Dicer function. On the whole, reported findings provide evidence that microRNA analysis is a molecular toxicology tool that can elucidate the pathogenic mechanisms activated by environmental carcinogens. Copyright © 2014 Elsevier GmbH. All rights reserved.

Distinct Expression Profiles and Novel Targets of MicroRNAs in Human Spermatogonia, Pachytene Spermatocytes, and Round Spermatids between OA Patients and NOA Patients

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Chencheng Yao

2017-12-01

Full Text Available Human spermatogenesis includes three main stages, namely, the mitosis of spermatogonia, meiosis of spermatocytes, and spermiogenesis of spermatids, which are precisely regulated by epigenetic and genetic factors. Abnormality of epigenetic and genetic factors can result in aberrant spermatogenesis and eventual male infertility. However, epigenetic regulators in controlling each stage of normal and abnormal human spermatogenesis remain unknown. Here, we have revealed for the first time the distinct microRNA profiles in human spermatogonia, pachytene spermatocytes, and round spermatids between obstructive azoospermia (OA patients and non-obstructive azoospermia (NOA patients. Human spermatogonia, pachytene spermatocytes, and round spermatids from OA patients and NOA patients were isolated using STA-PUT velocity sedimentation and identified by numerous hallmarks for these cells. RNA deep sequencing showed that 396 microRNAs were differentially expressed in human spermatogonia between OA patients and NOA patients and 395 differentially expressed microRNAs were found in human pachytene spermatocytes between OA patients and NOA patients. Moreover, 378 microRNAs were differentially expressed in human round spermatids between OA patients and NOA patients. The differential expression of numerous microRNAs identified by RNA deep sequencing was verified by real-time PCR. Moreover, a number of novel targeting genes for microRNAs were predicted using various kinds of software and further verified by real-time PCR. This study thus sheds novel insights into epigenetic regulation of human normal spermatogenesis and the etiology of azoospermia, and it could offer new targets for molecular therapy to treat male infertility.

MicroRNA-Mediated Dynamic Bidirectional Shift between the Subclasses of Glioblastoma Stem-like Cells

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Arun K. Rooj

2017-06-01

Full Text Available Large-scale transcriptomic profiling of glioblastoma (GBM into subtypes has provided remarkable insight into the pathobiology and heterogeneous nature of this disease. The mechanisms of speciation and inter-subtype transitions of these molecular subtypes require better characterization to facilitate the development of subtype-specific targeting strategies. The deregulation of microRNA expression among GBM subtypes and their subtype-specific targeting mechanisms are poorly understood. To reveal the underlying basis of microRNA-driven complex subpopulation dynamics within the heterogeneous intra-tumoral ecosystem, we characterized the expression of the subtype-enriched microRNA-128 (miR-128 in transcriptionally and phenotypically diverse subpopulations of patient-derived glioblastoma stem-like cells. Because microRNAs are capable of re-arranging the molecular landscape in a cell-type-specific manner, we argue that alterations in miR-128 levels are a potent mechanism of bidirectional transitions between GBM subpopulations, resulting in intermediate hybrid stages and emphasizing highly intricate intra-tumoral networking.

A cytometry microparticle platform approach for screening tobacco microRNA changes after agrobacterium delivery

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Powell, Joshua D.; Chen, Qiang; Mason, Hugh S.

2016-08-01

Abstract Key message nta-miR-398 is significantly up-regulated while nta-miR-428d is significantly down-regulated in tobacco after agroinfiltration AbstractMicroRNAs are a class of non-coding regulatory RNAs that can modulate development as well as alter innate antiviral defenses in plants. In this study we explored host changes at the microRNA level within tobacco (Nicotiana benthamiana) after expression of a recombinant anti-Ebola GP1 antibody through Agrobacterium tumefaciens agroinfiltration delivery. A multiplex nanoparticle-based cytometry assay tracked the host expression changes of 53 tobacco microRNAs. Our results revealed that the most abundant microRNAs in actively growing leaves corresponded to nanoparticle probes specific to nta-mir-6149 and nta-miR-168b. After agroinfiltration, probes targeting nta-mir-398 and nta-mir-482d were significantly altered in their respective expression levels and were further verified through RT-qPCR analysis. To our knowledge this study is the first to profile microRNA expression in tobacco after agroinfiltration using a multiplex nanoparticle approach.

MicroRNA Implications across Neurodevelopment and Neuropathology

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Sabata Martino

2009-01-01

Full Text Available MicroRNAs (miRNAs have rapidly emerged as biologically important mediators of posttranscriptional and epigenetic regulation in both plants and animals. miRNAs function through a variety of mechanisms including mRNA degradation and translational repression; additionally, miRNAs may guide gene expression by serving as transcription factors. miRNAs are highly expressed in human brain. Tissue and cell type-specific enrichments of certain miRNAs within the nervous system argue for a biological significance during neurodevelopmental stages. On the other hand, a large number of studies have reported links between alterations of miRNA homeostasis and pathologic conditions such as cancer, heart diseases, and neurodegeneration. Thus, profiles of distinct or aberrant miRNA signatures have most recently surged as one of the most fascinating interests in current biology. Here, the most recent insights into the involvement of miRNAs in the biology of the nervous system and the occurrence of neuropathological disorders are reviewed and discussed.

Impact of MicroRNA Levels, Target-Site Complementarity, and Cooperativity on Competing Endogenous RNA-Regulated Gene Expression.

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Denzler, Rémy; McGeary, Sean E; Title, Alexandra C; Agarwal, Vikram; Bartel, David P; Stoffel, Markus

2016-11-03

Expression changes of competing endogenous RNAs (ceRNAs) have been proposed to influence microRNA (miRNA) activity and thereby regulate other transcripts containing miRNA-binding sites. Here, we find that although miRNA levels define the extent of repression, they have little effect on the magnitude of the ceRNA expression change required to observe derepression. Canonical 6-nt sites, which typically mediate modest repression, can nonetheless compete for miRNA binding, with potency ∼20% of that observed for canonical 8-nt sites. In aggregate, low-affinity/background sites also contribute to competition. Sites with extensive additional complementarity can appear as more potent, but only because they induce miRNA degradation. Cooperative binding of proximal sites for the same or different miRNAs does increase potency. These results provide quantitative insights into the stoichiometric relationship between miRNAs and target abundance, target-site spacing, and affinity requirements for ceRNA-mediated gene regulation, and the unusual circumstances in which ceRNA-mediated gene regulation might be observed. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Gene expression profiles in BCL11B-siRNA treated malignant T cells

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Grabarczyk Piotr

2011-05-01

Full Text Available Abstract Background Downregulation of the B-cell chronic lymphocytic leukemia (CLL/lymphoma11B (BCL11B gene by small interfering RNA (siRNA leads to growth inhibition and apoptosis of the human T-cell acute lymphoblastic leukemia (T-ALL cell line Molt-4. To further characterize the molecular mechanism, a global gene expression profile of BCL11B-siRNA -treated Molt-4 cells was established. The expression profiles of several genes were further validated in the BCL11B-siRNA -treated Molt-4 cells and primary T-ALL cells. Results 142 genes were found to be upregulated and 109 genes downregulated in the BCL11B-siRNA -treated Molt-4 cells by microarray analysis. Among apoptosis-related genes, three pro-apoptotic genes, TNFSF10, BIK, BNIP3, were upregulated and one anti-apoptotic gene, BCL2L1 was downregulated. Moreover, the expression of SPP1 and CREBBP genes involved in the transforming growth factor (TGF-β pathway was down 16-fold. Expression levels of TNFSF10, BCL2L1, SPP1, and CREBBP were also examined by real-time PCR. A similar expression pattern of TNFSF10, BCL2L1, and SPP1 was identified. However, CREBBP was not downregulated in the BLC11B-siRNA -treated Molt-4 cells. Conclusion BCL11B-siRNA treatment altered expression profiles of TNFSF10, BCL2L1, and SPP1 in both Molt-4 T cell line and primary T-ALL cells.

microRNA profiling in the zoonotic parasite Echinococcus canadensis using a high-throughput approach.

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Macchiaroli, Natalia; Cucher, Marcela; Zarowiecki, Magdalena; Maldonado, Lucas; Kamenetzky, Laura; Rosenzvit, Mara Cecilia

2015-02-06

microRNAs (miRNAs), a class of small non-coding RNAs, are key regulators of gene expression at post-transcriptional level and play essential roles in fundamental biological processes such as development and metabolism. The particular developmental and metabolic characteristics of cestode parasites highlight the importance of studying miRNA gene regulation in these organisms. Here, we perform a comprehensive analysis of miRNAs in the parasitic cestode Echinococcus canadensis G7, one of the causative agents of the neglected zoonotic disease cystic echinococcosis. Small RNA libraries from protoscoleces and cyst walls of E. canadensis G7 and protoscoleces of E. granulosus sensu stricto G1 were sequenced using Illumina technology. For miRNA prediction, miRDeep2 core algorithm was used. The output list of candidate precursors was manually curated to generate a high confidence set of miRNAs. Differential expression analysis of miRNAs between stages or species was estimated with DESeq. Expression levels of selected miRNAs were validated using poly-A RT-qPCR. In this study we used a high-throughput approach and found transcriptional evidence of 37 miRNAs thus expanding the miRNA repertoire of E. canadensis G7. Differential expression analysis showed highly regulated miRNAs between life cycle stages, suggesting a role in maintaining the features of each developmental stage or in the regulation of developmental timing. In this work we characterize conserved and novel Echinococcus miRNAs which represent 30 unique miRNA families. Here we confirmed the remarkable loss of conserved miRNA families in E. canadensis, reflecting their low morphological complexity and high adaptation to parasitism. We performed the first in-depth study profiling of small RNAs in the zoonotic parasite E. canadensis G7. We found that miRNAs are the preponderant small RNA silencing molecules, suggesting that these small RNAs could be an essential mechanism of gene regulation in this species. We also

microRNA-320/RUNX2 axis regulates adipocytic differentiation of human mesenchymal (skeletal) stem cells

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Hamam, D; Ali, D; Vishnubalaji, R

2014-01-01

The molecular mechanisms promoting lineage-specific commitment of human mesenchymal (skeletal or stromal) stem cells (hMSCs) into adipocytes (ADs) are not fully understood. Thus, we performed global microRNA (miRNA) and gene expression profiling during adipocytic differentiation of h...... differentiation and accelerated formation of mature ADs in ex vivo cultures. Integrated analysis of bioinformatics and global gene expression profiling in miR-320c overexpressing cells and during adipocytic differentiation of hMSC identified several biologically relevant gene targets for miR-320c including RUNX2...

Serum microRNA profiles in athyroid patients on and off levothyroxine therapy.

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Massolt, Elske T; Chaker, Layal; Visser, Theo J; Gillis, Ad J M; Dorssers, Lambert C J; Beukhof, Carolien M; Kam, Boen L R; Franssen, Gaston J; Brigante, Giulia; van Ginhoven, Tessa M; Visser, W Edward; Looijenga, Leendert H J; Peeters, Robin P

2018-01-01

Levothyroxine replacement treatment in hypothyroidism is unable to restore physiological thyroxine and triiodothyronine concentrations in serum and tissues completely. Normal serum thyroid stimulating hormone (TSH) concentrations reflect only pituitary euthyroidism and, therefore, novel biomarkers representing tissue-specific thyroid state are needed. MicroRNAs (miRNAs), small non-coding regulatory RNAs, exhibit tissue-specific expression patterns and can be detectable in serum. Previous studies have demonstrated differential expression of (precursors of) miRNAs in tissues under the influence of thyroid hormone. To study if serum miRNA profiles are changed in different thyroid states. We studied 13 athyroid patients (6 males) during TSH suppressive therapy and after 4 weeks of thyroid hormone withdrawal. A magnetic bead capture system was used to isolate 384 defined miRNAs from serum. Subsequently, the TaqMan Array Card 3.0 platform was used for profiling after individual target amplification. Mean age of the subjects was 44.0 years (range 20-61 years). Median TSH levels were 88.9 mU/l during levothyroxine withdrawal and 0.006 mU/l during LT4 treatment with a median dosage of 2.1 μg/kg. After normalization to allow inter-sample analysis, a paired analysis did not demonstrate a significant difference in expression of any of the 384 miRNAs analyzed on and off LT4 treatment. Although we previously showed an up-regulation of pri-miRNAs 133b and 206 in hypothyroid state in skeletal muscle, the present study does not supply evidence that thyroid state also affects serum miRNAs in humans.

Shielding the messenger (RNA): microRNA-based anticancer therapies

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Sotillo, Elena; Thomas-Tikhonenko, Andrei

2011-01-01

It has been a decade since scientists realized that microRNAs (miRNAs) are not an oddity invented by worms to regulate gene expression at post-transcriptional levels. Rather, many of these 21–22-nucleotide-short RNAs exist in invertebrates and vertebrates alike and some of them are in fact highly conserved. miRNAs are now recognized as an important class of non-coding small RNAs that inhibit gene expression by targeting mRNA stability and translation. In the last ten years, our knowledge of the miRNAs world was expanding at vertiginous speed, propelled by the development of computational engines for miRNA identification and target prediction, biochemical tools and techniques to modulate miRNA activity, and last but not least, the emergence of miRNA-centric animal models. One important conclusion that has emerged from this effort is that many microRNAs and their cognate targets are strongly implicated in cancer, either as oncogenes or tumor and metastasis suppressors. In this review we will discuss the diverse role that miRNAs play in cancer initiation and progression and also the tools with which miRNA expression could be corrected in vivo. While the idea of targeting microRNAs towards therapeutic ends is getting considerable traction, basic, translational, and clinical research done in the next few years will tell whether this promise is well-founded. PMID:21514318

P2X7 mRNA expression in non-small cell lung cancer: MicroRNA regulation and prognostic value

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BOLDRINI, LAURA; GIORDANO, MIRELLA; ALÌ, GRETA; MELFI, FRANCA; ROMANO, GAETANO; LUCCHI, MARCO; FONTANINI, GABRIELLA

2014-01-01

The human P2X7 receptor is significant and exhibits several functions in neoplasia. At present, little is known with regard to its regulation. P2X7 expression may be regulated post-transcriptionally and putative microRNA (miRNA) binding sites are considered to be involved. The aim of this study was to determine whether miRNAs (miR-21, let-7 g and miR-205) regulate P2X7 mRNA stability. In addition, the impact of P2X7 expression in patients with non-small cell lung cancer (NSCLC) was investigat...

Identification of Novel and Conserved microRNAs in Homalodisca vitripennis, the Glassy-Winged Sharpshooter by Expression Profiling.

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Raja Sekhar Nandety

Full Text Available The glassy-winged sharpshooter (GWSS Homalodisca vitripennis (Hemiptera: Cicadellidae, is a xylem-feeding leafhopper and an important vector of the bacterium Xylella fastidiosa; the causal agent of Pierce's disease of grapevines. MicroRNAs are a class of small RNAs that play an important role in the functional development of various organisms including insects. In H. vitripennis, we identified microRNAs using high-throughput deep sequencing of adults followed by computational and manual annotation. A total of 14 novel microRNAs that are not found in the miRBase were identified from adult H. vitripennis. Conserved microRNAs were also found in our datasets. By comparison to our previously determined transcriptome sequence of H. vitripennis, we identified the potential targets of the microRNAs in the transcriptome. This microRNA profile information not only provides a more nuanced understanding of the biological and physiological mechanisms that govern gene expression in H. vitripennis, but may also lead to the identification of novel mechanisms for biorationally designed management strategies through the use of microRNAs.

Unravelling the complexity of microRNA-mediated gene regulation in black pepper (Piper nigrum L.) using high-throughput small RNA profiling.

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Asha, Srinivasan; Sreekumar, Sweda; Soniya, E V

2016-01-01

Analysis of high-throughput small RNA deep sequencing data, in combination with black pepper transcriptome sequences revealed microRNA-mediated gene regulation in black pepper ( Piper nigrum L.). Black pepper is an important spice crop and its berries are used worldwide as a natural food additive that contributes unique flavour to foods. In the present study to characterize microRNAs from black pepper, we generated a small RNA library from black pepper leaf and sequenced it by Illumina high-throughput sequencing technology. MicroRNAs belonging to a total of 303 conserved miRNA families were identified from the sRNAome data. Subsequent analysis from recently sequenced black pepper transcriptome confirmed precursor sequences of 50 conserved miRNAs and four potential novel miRNA candidates. Stem-loop qRT-PCR experiments demonstrated differential expression of eight conserved miRNAs in black pepper. Computational analysis of targets of the miRNAs showed 223 potential black pepper unigene targets that encode diverse transcription factors and enzymes involved in plant development, disease resistance, metabolic and signalling pathways. RLM-RACE experiments further mapped miRNA-mediated cleavage at five of the mRNA targets. In addition, miRNA isoforms corresponding to 18 miRNA families were also identified from black pepper. This study presents the first large-scale identification of microRNAs from black pepper and provides the foundation for the future studies of miRNA-mediated gene regulation of stress responses and diverse metabolic processes in black pepper.

Accurate microRNA target prediction correlates with protein repression levels

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Simossis Victor A

2009-09-01

Full Text Available Abstract Background MicroRNAs are small endogenously expressed non-coding RNA molecules that regulate target gene expression through translation repression or messenger RNA degradation. MicroRNA regulation is performed through pairing of the microRNA to sites in the messenger RNA of protein coding genes. Since experimental identification of miRNA target genes poses difficulties, computational microRNA target prediction is one of the key means in deciphering the role of microRNAs in development and disease. Results DIANA-microT 3.0 is an algorithm for microRNA target prediction which is based on several parameters calculated individually for each microRNA and combines conserved and non-conserved microRNA recognition elements into a final prediction score, which correlates with protein production fold change. Specifically, for each predicted interaction the program reports a signal to noise ratio and a precision score which can be used as an indication of the false positive rate of the prediction. Conclusion Recently, several computational target prediction programs were benchmarked based on a set of microRNA target genes identified by the pSILAC method. In this assessment DIANA-microT 3.0 was found to achieve the highest precision among the most widely used microRNA target prediction programs reaching approximately 66%. The DIANA-microT 3.0 prediction results are available online in a user friendly web server at http://www.microrna.gr/microT

Integrated analysis of miRNA and mRNA expression profiles in tilapia gonads at an early stage of sex differentiation.

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Tao, Wenjing; Sun, Lina; Shi, Hongjuan; Cheng, Yunying; Jiang, Dongneng; Fu, Beide; Conte, Matthew A; Gammerdinger, William J; Kocher, Thomas D; Wang, Deshou

2016-05-04

MicroRNAs (miRNAs) represent a second regulatory network that has important effects on gene expression and protein translation during biological process. However, the possible role of miRNAs in the early stages of fish sex differentiation is not well understood. In this study, we carried an integrated analysis of miRNA and mRNA expression profiles to explore their possibly regulatory patterns at the critical stage of sex differentiation in tilapia. We identified 279 pre-miRNA genes in tilapia genome, which were highly conserved in other fish species. Based on small RNA library sequencing, we identified 635 mature miRNAs in tilapia gonads, in which 62 and 49 miRNAs showed higher expression in XX and XY gonads, respectively. The predicted targets of these sex-biased miRNAs (e.g., miR-9, miR-21, miR-30a, miR-96, miR-200b, miR-212 and miR-7977) included genes encoding key enzymes in steroidogenic pathways (Cyp11a1, Hsd3b, Cyp19a1a, Hsd11b) and key molecules involved in vertebrate sex differentiation (Foxl2, Amh, Star1, Sf1, Dmrt1, and Gsdf). These genes also showed sex-biased expression in tilapia gonads at 5 dah. Some miRNAs (e.g., miR-96 and miR-737) targeted multiple genes involved in steroid synthesis, suggesting a complex miRNA regulatory network during early sex differentiation in this fish. The sequence and expression patterns of most miRNAs in tilapia are conserved in fishes, indicating the basic functions of vertebrate miRNAs might share a common evolutionary origin. This comprehensive analysis of miRNA and mRNA at the early stage of molecular sex differentiation in tilapia XX and XY gonads lead to the discovery of differentially expressed miRNAs and their putative targets, which will facilitate studies of the regulatory network of molecular sex determination and differentiation in fishes.

Hypoxia regulates microRNA expression in the human carotid body

International Nuclear Information System (INIS)

Mkrtchian, Souren; Lee, Kian Leong; Kåhlin, Jessica; Ebberyd, Anette; Poellinger, Lorenz; Fagerlund, Malin Jonsson; Eriksson, Lars I.

2017-01-01

The carotid body (CB) is the key sensing organ for physiological oxygen levels in the body. Under conditions of low oxygen (hypoxia), the CB plays crucial roles in signaling to the cardiorespiratory center in the medulla oblongata for the restoration of oxygen homeostasis. How hypoxia regulates gene expression in the human CB remains poorly understood. While limited information on transcriptional regulation in animal CBs is available, the identity and impact of important post-transcriptional regulators such as non-coding RNAs, and in particular miRNAs are not known. Here we show using ex vivo experiments that indeed a number of miRNAs are differentially regulated in surgically removed human CB slices when acute hypoxic conditions were applied. Analysis of the hypoxia-regulated miRNAs shows that they target biological pathways with upregulation of functions related to cell proliferation and immune response and downregulation of cell differentiation and cell death functions. Comparative analysis of the human CB miRNAome with the global miRNA expression patterns of a large number of different human tissues showed that the CB miRNAome had a unique profile which reflects its highly specialized functional status. Nevertheless, the human CB miRNAome is most closely related to the miRNA expression pattern of brain tissues indicating that they may have the most similar developmental origins. - Highlights: • Hypoxia triggers differential expression of many miRNAs in the human carotid body. • This can lead to the upregulation of proliferation and immune response functions. • CB expression profile in the carotid body resembles the miRNA expression pattern in the brain. • miRNAs are involved in the regulation of carotid body functions including oxygen sensing.

Hypoxia regulates microRNA expression in the human carotid body

Energy Technology Data Exchange (ETDEWEB)

Mkrtchian, Souren, E-mail: souren.mkrtchian@ki.se [Section for Anesthesiology and Intensive Care Medicine, Department of Physiology and Pharmacology, Karolinska Institute, SE-171 77 Stockholm (Sweden); Lee, Kian Leong, E-mail: csilkl@nus.edu.sg [Cancer Science Institute of Singapore, National University of Singapore, 117599 Singapore (Singapore); Kåhlin, Jessica [Section for Anesthesiology and Intensive Care Medicine, Department of Physiology and Pharmacology, Karolinska Institute, SE-171 77 Stockholm (Sweden); Function Perioperative Medicine and Intensive Care, Karolinska University Hospital, SE-171 76 Stockholm (Sweden); Ebberyd, Anette [Section for Anesthesiology and Intensive Care Medicine, Department of Physiology and Pharmacology, Karolinska Institute, SE-171 77 Stockholm (Sweden); Poellinger, Lorenz [Cancer Science Institute of Singapore, National University of Singapore, 117599 Singapore (Singapore); Department of Cell and Molecular Biology, Karolinska Institute, SE-171 77 Stockholm (Sweden); Fagerlund, Malin Jonsson; Eriksson, Lars I. [Section for Anesthesiology and Intensive Care Medicine, Department of Physiology and Pharmacology, Karolinska Institute, SE-171 77 Stockholm (Sweden); Function Perioperative Medicine and Intensive Care, Karolinska University Hospital, SE-171 76 Stockholm (Sweden)

2017-03-15

The carotid body (CB) is the key sensing organ for physiological oxygen levels in the body. Under conditions of low oxygen (hypoxia), the CB plays crucial roles in signaling to the cardiorespiratory center in the medulla oblongata for the restoration of oxygen homeostasis. How hypoxia regulates gene expression in the human CB remains poorly understood. While limited information on transcriptional regulation in animal CBs is available, the identity and impact of important post-transcriptional regulators such as non-coding RNAs, and in particular miRNAs are not known. Here we show using ex vivo experiments that indeed a number of miRNAs are differentially regulated in surgically removed human CB slices when acute hypoxic conditions were applied. Analysis of the hypoxia-regulated miRNAs shows that they target biological pathways with upregulation of functions related to cell proliferation and immune response and downregulation of cell differentiation and cell death functions. Comparative analysis of the human CB miRNAome with the global miRNA expression patterns of a large number of different human tissues showed that the CB miRNAome had a unique profile which reflects its highly specialized functional status. Nevertheless, the human CB miRNAome is most closely related to the miRNA expression pattern of brain tissues indicating that they may have the most similar developmental origins. - Highlights: • Hypoxia triggers differential expression of many miRNAs in the human carotid body. • This can lead to the upregulation of proliferation and immune response functions. • CB expression profile in the carotid body resembles the miRNA expression pattern in the brain. • miRNAs are involved in the regulation of carotid body functions including oxygen sensing.

MicroRNA Expression in Formalin-fixed Paraffin-embedded Cancer Tissue

DEFF Research Database (Denmark)

Boisen, Mogens Karsbøl; Dehlendorff, Christian; Linnemann, Dorte

2015-01-01

of miRNA expression in FFPE tissue samples from patients with colorectal (CRC) and pancreatic (PC) cancer and to quantify the variability associated with sample age and fixation. METHODS: High-throughput miRNA profiling results from 203 CRC and 256 PC FFPE samples as well as from 37 paired frozen....../FFPE samples from nine other CRC tumors (methodological samples) were used. Candidate reference miRNAs were identified by their correlation with global mean expression. The stability of reference genes was analyzed according to published methods. The association between sample age and global mean mi...... to global mean expression in each cancer type. Nine of these miRNAs were present in both lists, and miR-103a-3p was the most stable reference miRNA for both CRC and PC FFPE tissue. The optimal number of reference miRNAs was 4 in CRC and 10 in PC. Sample age had a significant effect on global mi...

Extracellular microRNA signature in chronic kidney disease.

Science.gov (United States)

Muralidharan, Jagdeesan; Ramezani, Ali; Hubal, Monica; Knoblach, Susan; Shrivastav, Shashi; Karandish, Sara; Scott, Richard; Maxwell, Nirmal; Ozturk, Savas; Beddhu, Srinivasan; Kopp, Jeffrey B; Raj, Dominic S

2017-06-01

MicroRNAs (miRNAs) are noncoding RNAs that regulate posttranscriptional gene expression. In this study we characterized the circulating and urinary miRNA pattern associated with reduced glomerular filtration rate, using Affymetrix GeneChip miR 4.0 in 28 patients with chronic kidney disease (CKD). Top miRNA discoveries from the human studies were validated in an Alb/TGFβ mouse model of CKD, and in rat renal proximal tubular cells (NRK52E) exposed to TGFβ1. Plasma and urinary levels of procollagen III N-terminal propeptide and collagen IV were elevated in patients with decreased estimated glomerular filtration rate (eGFR). Expression of 384 urinary and 266 circulatory miRNAs were significantly different between CKD patients with eGFR ≥30 vs. kidney fibrosis, and specific urinary and plasma miRNA profile may have diagnostic and prognostic utility in CKD. Copyright © 2017 the American Physiological Society.

Microarray profiling of microRNAs expressed in testis tissues of developing primates

DEFF Research Database (Denmark)

Yan, Naihong; Lu, Yilu; Sun, Huaqin

2009-01-01

MicroRNAs (miRNAs) are small non-coding RNA molecules that have been identified as potent regulators of gene expression. Recent studies indicate that miRNAs are involved in mammalian spermatogenesis but the mechanism of regulation is largely unknown....

MicroRNA Profiling in Aqueous Humor of Individual Human Eyes by Next-Generation Sequencing.

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Wecker, Thomas; Hoffmeier, Klaus; Plötner, Anne; Grüning, Björn Andreas; Horres, Ralf; Backofen, Rolf; Reinhard, Thomas; Schlunck, Günther

2016-04-01

Extracellular microRNAs (miRNAs) in aqueous humor were suggested to have a role in transcellular signaling and may serve as disease biomarkers. The authors adopted next-generation sequencing (NGS) techniques to further characterize the miRNA profile in single samples of 60 to 80 μL human aqueous humor. Samples were obtained at the outset of cataract surgery in nine independent, otherwise healthy eyes. Four samples were used to extract RNA and generate sequencing libraries, followed by an adapter-driven amplification step, electrophoretic size selection, sequencing, and data analysis. Five samples were used for quantitative PCR (qPCR) validation of NGS results. Published NGS data on circulating miRNAs in blood were analyzed in comparison. One hundred fifty-eight miRNAs were consistently detected by NGS in all four samples; an additional 59 miRNAs were present in at least three samples. The aqueous humor miRNA profile shows some overlap with published NGS-derived inventories of circulating miRNAs in blood plasma with high prevalence of human miR-451a, -21, and -16. In contrast to blood, miR-184, -4448, -30a, -29a, -29c, -19a, -30d, -205, -24, -22, and -3074 were detected among the 20 most prevalent miRNAs in aqueous humor. Relative expression patterns of miR-451a, -202, and -144 suggested by NGS were confirmed by qPCR. Our data illustrate the feasibility of miRNA analysis by NGS in small individual aqueous humor samples. Intraocular cells as well as blood plasma contribute to the extracellular aqueous humor miRNome. The data suggest possible roles of miRNA in intraocular cell adhesion and signaling by TGF-β and Wnt, which are important in intraocular pressure regulation and glaucoma.

Inflammation-related microRNA expression level in the bovine milk is affected by mastitis.

Science.gov (United States)

Lai, Yu-Chang; Fujikawa, Takuro; Maemura, Tadashi; Ando, Takaaki; Kitahara, Go; Endo, Yasuyuki; Yamato, Osamu; Koiwa, Masateru; Kubota, Chikara; Miura, Naoki

2017-01-01

MicroRNA (miRNA) in tissue and liquid samples have been shown to be associated with many diseases including inflammation. We aimed to identify inflammation-related miRNA expression level in the bovine mastitis milk. Expression level of inflammation-related miRNA in milk from mastitis-affected and normal cows was analyzed using qPCR. We found that expression level of miR-21, miR-146a, miR-155, miR-222, and miR-383 was significantly upregulated in California mastitis test positive (CMT+) milk. We further analyzed these miRNA using a chip-based QuantStudio Digital PCR System. The digital PCR results correlated with those of qPCR, demonstrating upregulation of miR-21, miR-146a, miR-155, miR-222, and miR-383 in CMT+ milk. In conclusion, we identified miRNA that are upregulated in CMT+ milk. These miRNA exhibited sensitivity and specificity greater than 80% for differentiating between CMT+ milk and normal milk. Our findings suggest that inflammation-related miRNA expression level in the bovine milk was affected by mastitis, and miRNA in milk have potential for use as biomarkers of bovine mastitis.

A microRNA feedback loop regulates global microRNA abundance during aging.

Science.gov (United States)

Inukai, Sachi; Pincus, Zachary; de Lencastre, Alexandre; Slack, Frank J

2018-02-01

Expression levels of many microRNAs (miRNAs) change during aging, notably declining globally in a number of organisms and tissues across taxa. However, little is known about the mechanisms or the biological relevance for this change. We investigated the network of genes that controls miRNA transcription and processing during C. elegans aging. We found that miRNA biogenesis genes are highly networked with transcription factors and aging-associated miRNAs. In particular, miR-71, known to influence life span and itself up-regulated during aging, represses alg-1 /Argonaute expression post-transcriptionally during aging. Increased ALG-1 abundance in mir-71 loss-of-function mutants led to globally increased miRNA expression. Interestingly, these mutants demonstrated widespread mRNA expression dysregulation and diminished levels of variability both in gene expression and in overall life span. Thus, the progressive molecular decline often thought to be the result of accumulated damage over an organism's life may be partially explained by a miRNA-directed mechanism of age-associated decline. © 2018 Inukai et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

Role of microRNA-7 and selenoprotein P in hepatocellular carcinoma.

Science.gov (United States)

Tarek, Marwa; Louka, Manal Louis; Khairy, Eman; Ali-Labib, Randa; Zakaria Zaky, Doaa; Montasser, Iman F

2017-05-01

There is an obvious need to diagnose hepatocellular carcinoma using novel non-invasive and sensitive biomarkers. In this regard, the aim of this study was to evaluate and correlate both relative quantification of microRNA-7 using quantitative real time polymerase chain reaction and quantitative analysis of selenoprotein P using enzyme-linked immunosorbent assay in sera of hepatocellular carcinoma patients, chronic liver disease patients, as well as normal healthy subjects in order to establish a new diagnostic biomarker with a valid non-invasive technique. In addition, this study aimed to investigate whether changes in selenium supply affect microRNA-7 expression and selenoprotein P levels in human hepatocarcinoma cell line (HepG2). The results showed a highly significant decrease in serum microRNA-7 relative quantification values and selenoprotein P levels in malignant group in comparison with benign and control groups. The best cutoff for serum microRNA-7 and selenoprotein P to discriminate hepatocellular carcinoma group from benign and control groups was 0.06 and 4.30 mg/L, respectively. Furthermore, this study showed that changes in selenium supply to HepG2 cell line can alter the microRNA-7 profile and are paralleled by changes in the concentration of its target protein (selenoprotein P). Hence, serum microRNA-7 and selenoprotein P appear to be potential non-invasive diagnostic markers for hepatocellular carcinoma. Moreover, the results suggest that selenium could be used as an anticancer therapy for hepatocellular carcinoma by affecting both microRNA-7 and selenoprotein P.

Expression Profiling of Circulating MicroRNAs in Canine Myxomatous Mitral Valve Disease

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Qinghong Li

2015-06-01

Full Text Available MicroRNAs (miRNAs are small non-coding RNAs that have shown promise as noninvasive biomarkers in cardiac disease. This study was undertaken to investigate the miRNA expression profile in dogs with myxomatous mitral valve disease (MMVD. 277 miRNAs were quantified using RT-qPCR from six normal dogs (American College of Veterinary Internal Medicine Stage A, six dogs with MMVD mild to moderate cardiac enlargement (ACVIM Stage B1/B2 and six dogs with MMVD and congestive heart failure (ACVIM Stage C/D. Eleven miRNAs were differentially expressed (False Discovery Rate < 0.05. Dogs in Stage B1/B2 or C/D had four upregulated miRNAs, including three cfa-let-7/cfa-miR-98 family members, while seven others were downregulated, compared to Stage A. Expression of six of the 11 miRNAs also were significantly different between dogs in Stage C/D and those in Stage B1/B2. The expression changes were greater as disease severity increased. These miRNAs may be candidates for novel biomarkers and may provide insights into genetic regulatory pathways in canine MMVD.

Characterization of global microRNA expression reveals oncogenic potential of miR-145 in metastatic colorectal cancer

International Nuclear Information System (INIS)

Arndt, Greg M; Retzlaff, Kathy; Bittner, Anton; Raponi, Mitch; Dossey, Lesley; Cullen, Lara M; Lai, Angela; Druker, Riki; Eisbacher, Michael; Zhang, Chunyan; Tran, Nham; Fan, Hongtao

2009-01-01

MicroRNAs (MiRNAs) are short non-coding RNAs that control protein expression through various mechanisms. Their altered expression has been shown to be associated with various cancers. The aim of this study was to profile miRNA expression in colorectal cancer (CRC) and to analyze the function of specific miRNAs in CRC cells. MirVana miRNA Bioarrays were used to determine the miRNA expression profile in eight CRC cell line models, 45 human CRC samples of different stages, and four matched normal colon tissue samples. SW620 CRC cells were stably transduced with miR-143 or miR-145 expression vectors and analyzed in vitro for cell proliferation, cell differentiation and anchorage-independent growth. Signalling pathways associated with differentially expressed miRNAs were identified using a gene set enrichment analysis. The expression analysis of clinical CRC samples identified 37 miRNAs that were differentially expressed between CRC and normal tissue. Furthermore, several of these miRNAs were associated with CRC tumor progression including loss of miR-133a and gain of miR-224. We identified 11 common miRNAs that were differentially expressed between normal colon and CRC in both the cell line models and clinical samples. In vitro functional studies indicated that miR-143 and miR-145 appear to function in opposing manners to either inhibit or augment cell proliferation in a metastatic CRC model. The pathways targeted by miR-143 and miR-145 showed no significant overlap. Furthermore, gene expression analysis of metastatic versus non-metastatic isogenic cell lines indicated that miR-145 targets involved in cell cycle and neuregulin pathways were significantly down-regulated in the metastatic context. MiRNAs showing altered expression at different stages of CRC could be targets for CRC therapies and be further developed as potential diagnostic and prognostic analytes. The identified biological processes and signalling pathways collectively targeted by co-expressed miRNAs in

Human amniotic epithelial cell feeder layers maintain human iPS cell pluripotency via inhibited endogenous microRNA-145 and increased Sox2 expression

International Nuclear Information System (INIS)

Liu, Te; Cheng, Weiwei; Huang, Yongyi; Huang, Qin; Jiang, Lizhen; Guo, Lihe

2012-01-01

Currently, human induced pluripotent stem (iPS) cells were generated from patient or disease-specific sources and share the same key properties as embryonic stem cells. This makes them attractive for personalized medicine, drug screens or cellular therapy. Long-term cultivation and maintenance of normal iPS cells in an undifferentiated self-renewing state are a major challenge. Our previous studies have shown that human amniotic epithelial cells (HuAECs) could provide a good source of feeder cells for mouse and human embryonic stem cells, or spermatogonial stem cells, but the mechanism for this is unknown. Here, we examined the effect of endogenous microRNA-145 regulation on Sox2 expression in human iPS cells by HuAECs feeder cells regulation, and in turn on human iPS cells pluripotency. We found that human IPS cells transfected with a microRNA-145 mutant expressed Sox2 at high levels, allowing iPS to maintain a high level of AP activity in long-term culture and form teratomas in SCID mice. Expression of stem cell markers was increased in iPS transfected with the microRNA-145 mutant, compared with iPS was transfected with microRNA-145. Besides, the expression of Drosha proteins of the microRNA-processor complex, required for the generation of precursor pre-miRNA, was significantly increased in human iPS cells cultured on MEF but not on HuAECs. Taken together, these results suggest that endogenous Sox2 expression may be regulated by microRNA-145 in human iPS cells with HuAECs feeder cells, and Sox2 is a crucial component required for maintenance of them in an undifferentiated, proliferative state capable of self-renewal. Highlights: ► microRNA-145 inhibits Sox2 expression in human iPS cells. ► microRNA-145 suppresses the self-renewal and pluripotency of human iPS cells. ► HuAECs regulate expression of microRNA-145 and Sox2 in human iPS cells. ► HuAECs feeder layers maintain human iPS cells pluripotency. ► HuAECs negatively regulates the synthesis of

Human amniotic epithelial cell feeder layers maintain human iPS cell pluripotency via inhibited endogenous microRNA-145 and increased Sox2 expression

Energy Technology Data Exchange (ETDEWEB)

Liu, Te, E-mail: liute79@yahoo.com [School of Environmental Science and Engineering, Donghua University, Shanghai 201620 (China); Shanghai Geriatric Institute of Chinese Medicine, Shanghai 200031 (China); Cheng, Weiwei [International Peace Maternity and Child Health Hospital, Shanghai Jiaotong University, Shanghai 200030 (China); Huang, Yongyi [Laboratoire PROTEE, Batiment R, Universite du Sud Toulon-Var, 83957 LA GARDE Cedex (France); Huang, Qin; Jiang, Lizhen [Institute of Biochemistry and Cell Biology, Shanghai Institute for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Guo, Lihe, E-mail: liute79@yahoo.com [Institute of Biochemistry and Cell Biology, Shanghai Institute for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China)

2012-02-15

Currently, human induced pluripotent stem (iPS) cells were generated from patient or disease-specific sources and share the same key properties as embryonic stem cells. This makes them attractive for personalized medicine, drug screens or cellular therapy. Long-term cultivation and maintenance of normal iPS cells in an undifferentiated self-renewing state are a major challenge. Our previous studies have shown that human amniotic epithelial cells (HuAECs) could provide a good source of feeder cells for mouse and human embryonic stem cells, or spermatogonial stem cells, but the mechanism for this is unknown. Here, we examined the effect of endogenous microRNA-145 regulation on Sox2 expression in human iPS cells by HuAECs feeder cells regulation, and in turn on human iPS cells pluripotency. We found that human IPS cells transfected with a microRNA-145 mutant expressed Sox2 at high levels, allowing iPS to maintain a high level of AP activity in long-term culture and form teratomas in SCID mice. Expression of stem cell markers was increased in iPS transfected with the microRNA-145 mutant, compared with iPS was transfected with microRNA-145. Besides, the expression of Drosha proteins of the microRNA-processor complex, required for the generation of precursor pre-miRNA, was significantly increased in human iPS cells cultured on MEF but not on HuAECs. Taken together, these results suggest that endogenous Sox2 expression may be regulated by microRNA-145 in human iPS cells with HuAECs feeder cells, and Sox2 is a crucial component required for maintenance of them in an undifferentiated, proliferative state capable of self-renewal. Highlights: Black-Right-Pointing-Pointer microRNA-145 inhibits Sox2 expression in human iPS cells. Black-Right-Pointing-Pointer microRNA-145 suppresses the self-renewal and pluripotency of human iPS cells. Black-Right-Pointing-Pointer HuAECs regulate expression of microRNA-145 and Sox2 in human iPS cells. Black-Right-Pointing-Pointer HuAECs feeder

Deep sequencing of cardiac microRNA-mRNA interactomes in clinical and experimental cardiomyopathy.

Science.gov (United States)

Matkovich, Scot J; Dorn, Gerald W

2015-01-01

MicroRNAs are a family of short (~21 nucleotide) noncoding RNAs that serve key roles in cellular growth and differentiation and the response of the heart to stress stimuli. As the sequence-specific recognition element of RNA-induced silencing complexes (RISCs), microRNAs bind mRNAs and prevent their translation via mechanisms that may include transcript degradation and/or prevention of ribosome binding. Short microRNA sequences and the ability of microRNAs to bind to mRNA sites having only partial/imperfect sequence complementarity complicate purely computational analyses of microRNA-mRNA interactomes. Furthermore, computational microRNA target prediction programs typically ignore biological context, and therefore the principal determinants of microRNA-mRNA binding: the presence and quantity of each. To address these deficiencies we describe an empirical method, developed via studies of stressed and failing hearts, to determine disease-induced changes in microRNAs, mRNAs, and the mRNAs targeted to the RISC, without cross-linking mRNAs to RISC proteins. Deep sequencing methods are used to determine RNA abundances, delivering unbiased, quantitative RNA data limited only by their annotation in the genome of interest. We describe the laboratory bench steps required to perform these experiments, experimental design strategies to achieve an appropriate number of sequencing reads per biological replicate, and computer-based processing tools and procedures to convert large raw sequencing data files into gene expression measures useful for differential expression analyses.

The MicroRNA-200 Family Is Upregulated in Endometrial Carcinoma

Science.gov (United States)

Snowdon, Jaime; Zhang, Xiao; Childs, Tim; Tron, Victor A.; Feilotter, Harriet

2011-01-01

Background MicroRNAs (miRNAs, miRs) are small non-coding RNAs that negatively regulate gene expression at the post-transcriptional level. MicroRNAs are dysregulated in cancer and may play essential roles in tumorigenesis. Additionally, miRNAs have been shown to have prognostic and diagnostic value in certain types of cancer. The objective of this study was to identify dysregulated miRNAs in endometrioid endometrial adenocarcinoma (EEC) and the precursor lesion, complex atypical hyperplasia (CAH). Methodology We compared the expression profiles of 723 human miRNAs from 14 cases of EEC, 10 cases of CAH, and 10 normal proliferative endometria controls using Agilent Human miRNA arrays following RNA extraction from formalin-fixed paraffin-embedded (FFPE) tissues. The expression of 4 dysregulated miRNAs was validated using real time reverse transcription-PCR. Results Forty-three miRNAs were dysregulated in EEC and CAH compared to normal controls (p<0.05). The entire miR-200 family (miR-200a/b/c, miR-141, and miR-429) was up-regulated in cases of EEC. Conclusions This information contributes to the candidate miRNA expression profile that has been generated for EEC and shows that certain miRNAs are dysregulated in the precursor lesion, CAH. These miRNAs in particular may play important roles in tumorigenesis. Examination of miRNAs that are consistently dysregulated in various studies of EEC, like the miR-200 family, will aid in the understanding of the role that miRNAs play in tumorigenesis in this tumour type. PMID:21897839

The microRNA-200 family is upregulated in endometrial carcinoma.

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Jaime Snowdon

Full Text Available BACKGROUND: MicroRNAs (miRNAs, miRs are small non-coding RNAs that negatively regulate gene expression at the post-transcriptional level. MicroRNAs are dysregulated in cancer and may play essential roles in tumorigenesis. Additionally, miRNAs have been shown to have prognostic and diagnostic value in certain types of cancer. The objective of this study was to identify dysregulated miRNAs in endometrioid endometrial adenocarcinoma (EEC and the precursor lesion, complex atypical hyperplasia (CAH. METHODOLOGY: We compared the expression profiles of 723 human miRNAs from 14 cases of EEC, 10 cases of CAH, and 10 normal proliferative endometria controls using Agilent Human miRNA arrays following RNA extraction from formalin-fixed paraffin-embedded (FFPE tissues. The expression of 4 dysregulated miRNAs was validated using real time reverse transcription-PCR. RESULTS: Forty-three miRNAs were dysregulated in EEC and CAH compared to normal controls (p<0.05. The entire miR-200 family (miR-200a/b/c, miR-141, and miR-429 was up-regulated in cases of EEC. CONCLUSIONS: This information contributes to the candidate miRNA expression profile that has been generated for EEC and shows that certain miRNAs are dysregulated in the precursor lesion, CAH. These miRNAs in particular may play important roles in tumorigenesis. Examination of miRNAs that are consistently dysregulated in various studies of EEC, like the miR-200 family, will aid in the understanding of the role that miRNAs play in tumorigenesis in this tumour type.

RNA deep sequencing reveals differential microRNA expression during development of sea urchin and sea star.

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Sabah Kadri

Full Text Available microRNAs (miRNAs are small (20-23 nt, non-coding single stranded RNA molecules that act as post-transcriptional regulators of mRNA gene expression. They have been implicated in regulation of developmental processes in diverse organisms. The echinoderms, Strongylocentrotus purpuratus (sea urchin and Patiria miniata (sea star are excellent model organisms for studying development with well-characterized transcriptional networks. However, to date, nothing is known about the role of miRNAs during development in these organisms, except that the genes that are involved in the miRNA biogenesis pathway are expressed during their developmental stages. In this paper, we used Illumina Genome Analyzer (Illumina, Inc. to sequence small RNA libraries in mixed stage population of embryos from one to three days after fertilization of sea urchin and sea star (total of 22,670,000 reads. Analysis of these data revealed the miRNA populations in these two species. We found that 47 and 38 known miRNAs are expressed in sea urchin and sea star, respectively, during early development (32 in common. We also found 13 potentially novel miRNAs in the sea urchin embryonic library. miRNA expression is generally conserved between the two species during development, but 7 miRNAs are highly expressed in only one species. We expect that our two datasets will be a valuable resource for everyone working in the field of developmental biology and the regulatory networks that affect it. The computational pipeline to analyze Illumina reads is available at http://www.benoslab.pitt.edu/services.html.

RNA Deep Sequencing Reveals Differential MicroRNA Expression during Development of Sea Urchin and Sea Star

Science.gov (United States)

Kadri, Sabah; Hinman, Veronica F.; Benos, Panayiotis V.

2011-01-01

microRNAs (miRNAs) are small (20–23 nt), non-coding single stranded RNA molecules that act as post-transcriptional regulators of mRNA gene expression. They have been implicated in regulation of developmental processes in diverse organisms. The echinoderms, Strongylocentrotus purpuratus (sea urchin) and Patiria miniata (sea star) are excellent model organisms for studying development with well-characterized transcriptional networks. However, to date, nothing is known about the role of miRNAs during development in these organisms, except that the genes that are involved in the miRNA biogenesis pathway are expressed during their developmental stages. In this paper, we used Illumina Genome Analyzer (Illumina, Inc.) to sequence small RNA libraries in mixed stage population of embryos from one to three days after fertilization of sea urchin and sea star (total of 22,670,000 reads). Analysis of these data revealed the miRNA populations in these two species. We found that 47 and 38 known miRNAs are expressed in sea urchin and sea star, respectively, during early development (32 in common). We also found 13 potentially novel miRNAs in the sea urchin embryonic library. miRNA expression is generally conserved between the two species during development, but 7 miRNAs are highly expressed in only one species. We expect that our two datasets will be a valuable resource for everyone working in the field of developmental biology and the regulatory networks that affect it. The computational pipeline to analyze Illumina reads is available at http://www.benoslab.pitt.edu/services.html. PMID:22216218

Proanthocyanidins modulate microRNA expression in human HepG2 cells.

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Anna Arola-Arnal

Full Text Available Mi(croRNAs are small non-coding RNAs of 18-25 nucleotides in length that modulate gene expression at the post-transcriptional level. These RNAs have been shown to be involved in a several biological processes, human diseases and metabolic disorders. Proanthocyanidins, which are the most abundant polyphenol class in the human diet, have positive health effects on a variety of metabolic disorders such as inflammation, obesity, diabetes and insulin resistance. The present study aimed to evaluate whether proanthocyanidin-rich natural extracts modulate miRNA expression. Using microarray analysis and Q-PCR, we investigated miRNA expression in HepG2 cells treated with proanthocyanidins. Our results showed that when HepG2 cells were treated with grape seed proanthocyanidin extract (GSPE, cocoa proanthocyanidin extract (CPE or pure epigallocatechin gallate isolated from green tea (EGCG, fifteen, six and five differentially expressed miRNAs, respectively, were identified out of 904 mRNAs. Specifically, miR-30b* was downregulated by the three treatments, and treatment with GSPE or CPE upregulated miR-1224-3p, miR-197 and miR-532-3p. Therefore, these results provide evidence of the capacity of dietary proanthocyanidins to influence microRNA expression, suggesting a new mechanism of action of proanthocyanidins.

MicroRNA miR-328 regulates zonation morphogenesis by targeting CD44 expression.

Science.gov (United States)

Wang, Chia-Hui; Lee, Daniel Y; Deng, Zhaoqun; Jeyapalan, Zina; Lee, Shao-Chen; Kahai, Shireen; Lu, Wei-Yang; Zhang, Yaou; Yang, Burton B

2008-06-18

Morphogenesis is crucial to initiate physiological development and tumor invasion. Here we show that a microRNA controls zonation morphogenesis by targeting hyaluronan receptor CD44. We have developed a novel system to study microRNA functions by generating constructs expressing pre-miRNAs and mature miRNAs. Using this system, we have demonstrated that expression of miR-328 reduced cell adhesion, aggregation, and migration, and regulated formation of capillary structure. Protein analysis indicated that miR-328 repressed CD44 expression. Activities of luciferase constructs harboring the target site in CD44, but not the one containing mutation, were repressed by miR-328. Zonation morphogenesis appeared in cells transfected by miR-328: miR-328-transfected cells were present on the surface of zonating structures while the control cells stayed in the middle. MiR-328-mediated CD44 actions was validated by anti-CD44 antibody, hyaluronidase, CD44 siRNA, and CD44 expression constructs. In vivo experiments showed that CD44-silencing cells appeared as layers on the surfaces of nodules or zonating structures. Immuno-histochemistry also exhibited CD44-negative cells on the surface layers of normal rat livers and the internal zones of Portal veins. Our results demonstrate that miR-328 targets CD44, which is essential in regulating zonation morphogenesis: silencing of CD44 expression is essential in sealing the zonation structures to facilitate their extension and to inhibit complex expansion.

MicroRNA miR-328 regulates zonation morphogenesis by targeting CD44 expression.

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Chia-Hui Wang

Full Text Available Morphogenesis is crucial to initiate physiological development and tumor invasion. Here we show that a microRNA controls zonation morphogenesis by targeting hyaluronan receptor CD44. We have developed a novel system to study microRNA functions by generating constructs expressing pre-miRNAs and mature miRNAs. Using this system, we have demonstrated that expression of miR-328 reduced cell adhesion, aggregation, and migration, and regulated formation of capillary structure. Protein analysis indicated that miR-328 repressed CD44 expression. Activities of luciferase constructs harboring the target site in CD44, but not the one containing mutation, were repressed by miR-328. Zonation morphogenesis appeared in cells transfected by miR-328: miR-328-transfected cells were present on the surface of zonating structures while the control cells stayed in the middle. MiR-328-mediated CD44 actions was validated by anti-CD44 antibody, hyaluronidase, CD44 siRNA, and CD44 expression constructs. In vivo experiments showed that CD44-silencing cells appeared as layers on the surfaces of nodules or zonating structures. Immuno-histochemistry also exhibited CD44-negative cells on the surface layers of normal rat livers and the internal zones of Portal veins. Our results demonstrate that miR-328 targets CD44, which is essential in regulating zonation morphogenesis: silencing of CD44 expression is essential in sealing the zonation structures to facilitate their extension and to inhibit complex expansion.

MicroRNA-26a-mediated regulation of interleukin-2 expression in transformed avian lymphocyte lines

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Smith Lorraine P

2010-05-01

Full Text Available Abstract Background Micro(miRNAs are a class of small non-coding RNAs that play critical roles in the induction of various cancers, including lymphomas induced by oncogenic viruses. While some of the miRNAs are oncogenic, miRNAs such as miR-26a are consistently downregulated in a number of cancers, demonstrating their potential tumor suppressor functions. Global miRNA expression profiles of a number of virus-transformed avian lymphoma cell lines have shown downregulation of gga-miR-26a expression, irrespective of molecular mechanisms of transformation or the viral aetiology. The neoplastic transformation of lymphocytes by many viruses accompanies high levels of proliferative responses, mostly mediated through cytokines such as IL-2. Chicken IL-2 can modulate T-cell proliferation and cytotoxicity in vitro and in vivo and dysregulation of IL-2 expression is observed in diseases such as leukaemia. Results The expression levels of gga-miR-26a in chicken lymphoma cells transformed by 3 distinct avian oncogenic viruses, viz Marek's disease virus (MDV, avian leukosis virus (ALV and Reticuloendotheliosis virus (REV were consistently downregulated compared to the levels in the normal lymphocytes. This downregulation of miR-26a regardless of the viral etiology and molecular mechanisms of transformation was consistent with the tumor suppressor role of this miRNA. Notwithstanding this well-established role in cancer, we demonstrate the additional role of this miRNA in directly targeting chicken IL-2 through reporter and biochemical assays. The downregulation of miR-26a can relieve the suppressive effect of this miRNA on IL-2 expression. Conclusions We show that miR-26a is globally downregulated in a number of avian lymphoma cells irrespective of the mechanisms of transformation, reiterating the highly conserved tumor suppressor function of this miRNA. However, with the potential for directly targeting chicken IL-2, the downregulation of miR-26a in these

Tissue-specific mRNA expression profiling in grape berry tissues

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Grimplet, Jerome; Deluc, Laurent G; Tillett, Richard L; Wheatley, Matthew D; Schlauch, Karen A; Cramer, Grant R; Cushman, John C

2007-01-01

Background Berries of grape (Vitis vinifera) contain three major tissue types (skin, pulp and seed) all of which contribute to the aroma, color, and flavor characters of wine. The pericarp, which is composed of the exocarp (skin) and mesocarp (pulp), not only functions to protect and feed the developing seed, but also to assist in the dispersal of the mature seed by avian and mammalian vectors. The skin provides volatile and nonvolatile aroma and color compounds, the pulp contributes organic acids and sugars, and the seeds provide condensed tannins, all of which are important to the formation of organoleptic characteristics of wine. In order to understand the transcriptional network responsible for controlling tissue-specific mRNA expression patterns, mRNA expression profiling was conducted on each tissue of mature berries of V. vinifera Cabernet Sauvignon using the Affymetrix GeneChip® Vitis oligonucleotide microarray ver. 1.0. In order to monitor the influence of water-deficit stress on tissue-specific expression patterns, mRNA expression profiles were also compared from mature berries harvested from vines subjected to well-watered or water-deficit conditions. Results Overall, berry tissues were found to express approximately 76% of genes represented on the Vitis microarray. Approximately 60% of these genes exhibited significant differential expression in one or more of the three major tissue types with more than 28% of genes showing pronounced (2-fold or greater) differences in mRNA expression. The largest difference in tissue-specific expression was observed between the seed and pulp/skin. Exocarp tissue, which is involved in pathogen defense and pigment production, showed higher mRNA abundance relative to other berry tissues for genes involved with flavonoid biosynthesis, pathogen resistance, and cell wall modification. Mesocarp tissue, which is considered a nutritive tissue, exhibited a higher mRNA abundance of genes involved in cell wall function and

Tissue-specific mRNA expression profiling in grape berry tissues

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Cramer Grant R

2007-06-01

Full Text Available Abstract Background Berries of grape (Vitis vinifera contain three major tissue types (skin, pulp and seed all of which contribute to the aroma, color, and flavor characters of wine. The pericarp, which is composed of the exocarp (skin and mesocarp (pulp, not only functions to protect and feed the developing seed, but also to assist in the dispersal of the mature seed by avian and mammalian vectors. The skin provides volatile and nonvolatile aroma and color compounds, the pulp contributes organic acids and sugars, and the seeds provide condensed tannins, all of which are important to the formation of organoleptic characteristics of wine. In order to understand the transcriptional network responsible for controlling tissue-specific mRNA expression patterns, mRNA expression profiling was conducted on each tissue of mature berries of V. vinifera Cabernet Sauvignon using the Affymetrix GeneChip® Vitis oligonucleotide microarray ver. 1.0. In order to monitor the influence of water-deficit stress on tissue-specific expression patterns, mRNA expression profiles were also compared from mature berries harvested from vines subjected to well-watered or water-deficit conditions. Results Overall, berry tissues were found to express approximately 76% of genes represented on the Vitis microarray. Approximately 60% of these genes exhibited significant differential expression in one or more of the three major tissue types with more than 28% of genes showing pronounced (2-fold or greater differences in mRNA expression. The largest difference in tissue-specific expression was observed between the seed and pulp/skin. Exocarp tissue, which is involved in pathogen defense and pigment production, showed higher mRNA abundance relative to other berry tissues for genes involved with flavonoid biosynthesis, pathogen resistance, and cell wall modification. Mesocarp tissue, which is considered a nutritive tissue, exhibited a higher mRNA abundance of genes involved in cell

MicroRNA modulation in complex regional pain syndrome

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Orlova Irina A

2011-11-01

Full Text Available Abstract Background Aberrant expression of small noncoding RNAs called microRNAs (miRNAs is a common feature of several human diseases. The objective of the study was to identify miRNA modulation in patients with complex regional pain syndrome (CRPS a chronic pain condition resulting from dysfunction in the central and/or peripheral nervous systems. Due to a multitude of inciting pathologies, symptoms and treatment conditions, the CRPS patient population is very heterogeneous. Our goal was to identify differentially expressed miRNAs in blood and explore their utility in patient stratification. Methods We profiled miRNAs in whole blood from 41 patients with CRPS and 20 controls using TaqMan low density array cards. Since neurogenic inflammation is known to play a significant role in CRPS we measured inflammatory markers including chemokines, cytokines, and their soluble receptors in blood from the same individuals. Correlation analyses were performed for miRNAs, inflammatory markers and other parameters including disease symptoms, medication, and comorbid conditions. Results Three different groups emerged from miRNA profiling. One group was comprised of 60% of CRPS patients and contained no control subjects. miRNA profiles from the remaining patients were interspersed among control samples in the other two groups. We identified differential expression of 18 miRNAs in CRPS patients. Analysis of inflammatory markers showed that vascular endothelial growth factor (VEGF, interleukin1 receptor antagonist (IL1Ra and monocyte chemotactic protein-1 (MCP1 were significantly elevated in CRPS patients. VEGF and IL1Ra showed significant correlation with the patients reported pain levels. Analysis of the patients who were clustered according to their miRNA profile revealed correlations that were not significant in the total patient population. Correlation analysis of miRNAs detected in blood with additional parameters identified miRNAs associated with

Elevated levels of circulating microRNA-200 family members correlate with serous epithelial ovarian cancer

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Kan Casina WS

2012-12-01

Full Text Available Abstract Background There is a critical need for improved diagnostic markers for high grade serous epithelial ovarian cancer (SEOC. MicroRNAs are stable in the circulation and may have utility as biomarkers of malignancy. We investigated whether levels of serum microRNA could discriminate women with high-grade SEOC from age matched healthy volunteers. Methods To identify microRNA of interest, microRNA expression profiling was performed on 4 SEOC cell lines and normal human ovarian surface epithelial cells. Total RNA was extracted from 500 μL aliquots of serum collected from patients with SEOC (n = 28 and age-matched healthy donors (n = 28. Serum microRNA levels were assessed by quantitative RT-PCR following preamplification. Results microRNA (miR-182, miR-200a, miR-200b and miR-200c were highly overexpressed in the SEOC cell lines relative to normal human ovarian surface epithelial cells and were assessed in RNA extracted from serum as candidate biomarkers. miR-103, miR-92a and miR -638 had relatively invariant expression across all ovarian cell lines, and with small-nucleolar C/D box 48 (RNU48 were assessed in RNA extracted from serum as candidate endogenous normalizers. No correlation between serum levels and age were observed (age range 30-79 years for any of these microRNA or RNU48. Individually, miR-200a, miR-200b and miR-200c normalized to serum volume and miR-103 were significantly higher in serum of the SEOC cohort (P  Conclusions We identified serum microRNAs able to discriminate patients with high grade SEOC from age-matched healthy controls. The addition of these microRNAs to current testing regimes may improve diagnosis for women with SEOC.

UVA and UVB Irradiation Differentially Regulate microRNA Expression in Human Primary Keratinocytes

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Kraemer, Anne; Chen, I-Peng; Henning, Stefan; Faust, Alexandra; Volkmer, Beate; Atkinson, Michael J.; Moertl, Simone; Greinert, Ruediger

2013-01-01

MicroRNA (miRNA)-mediated regulation of the cellular transcriptome is an important epigenetic mechanism for fine-tuning regulatory pathways. These include processes related to skin cancer development, progression and metastasis. However, little is known about the role of microRNA as an intermediary in the carcinogenic processes following exposure to UV-radiation. We now show that UV irradiation of human primary keratinocytes modulates the expression of several cellular miRNAs. A common set of miRNAs was influenced by exposure to both UVA and UVB. However, each wavelength band also activated a distinct subset of miRNAs. Common sets of UVA- and UVB-regulated miRNAs harbor the regulatory elements GLYCA-nTRE, GATA-1-undefined-site-13 or Hox-2.3-undefined-site-2 in their promoters. In silico analysis indicates that the differentially expressed miRNAs responding to UV have potential functions in the cellular pathways of cell growth and proliferation. Interestingly, the expression of miR-23b, which is a differentiation marker of human keratinocytes, is remarkably up-regulated after UVA irradiation. Studying the interaction between miR-23b and its putative skin-relevant targets using a Luciferase reporter assay revealed that RRAS2 (related RAS viral oncogene homolog 2), which is strongly expressed in highly aggressive malignant skin cancer, to be a direct target of miR-23b. This study demonstrates for the first time a differential miRNA response to UVA and UVB in human primary keratinocytes. This suggests that selective regulation of signaling pathways occurs in response to different UV energies. This may shed new light on miRNA-regulated carcinogenic processes involved in UV-induced skin carcinogenesis. PMID:24391759

UVA and UVB irradiation differentially regulate microRNA expression in human primary keratinocytes.

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Anne Kraemer

Full Text Available MicroRNA (miRNA-mediated regulation of the cellular transcriptome is an important epigenetic mechanism for fine-tuning regulatory pathways. These include processes related to skin cancer development, progression and metastasis. However, little is known about the role of microRNA as an intermediary in the carcinogenic processes following exposure to UV-radiation. We now show that UV irradiation of human primary keratinocytes modulates the expression of several cellular miRNAs. A common set of miRNAs was influenced by exposure to both UVA and UVB. However, each wavelength band also activated a distinct subset of miRNAs. Common sets of UVA- and UVB-regulated miRNAs harbor the regulatory elements GLYCA-nTRE, GATA-1-undefined-site-13 or Hox-2.3-undefined-site-2 in their promoters. In silico analysis indicates that the differentially expressed miRNAs responding to UV have potential functions in the cellular pathways of cell growth and proliferation. Interestingly, the expression of miR-23b, which is a differentiation marker of human keratinocytes, is remarkably up-regulated after UVA irradiation. Studying the interaction between miR-23b and its putative skin-relevant targets using a Luciferase reporter assay revealed that RRAS2 (related RAS viral oncogene homolog 2, which is strongly expressed in highly aggressive malignant skin cancer, to be a direct target of miR-23b. This study demonstrates for the first time a differential miRNA response to UVA and UVB in human primary keratinocytes. This suggests that selective regulation of signaling pathways occurs in response to different UV energies. This may shed new light on miRNA-regulated carcinogenic processes involved in UV-induced skin carcinogenesis.

Long non-coding RNA expression profiling of mouse testis during postnatal development.

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Jin Sun

Full Text Available Mammalian testis development and spermatogenesis play critical roles in male fertility and continuation of a species. Previous research into the molecular mechanisms of testis development and spermatogenesis has largely focused on the role of protein-coding genes and small non-coding RNAs, such as microRNAs and piRNAs. Recently, it has become apparent that large numbers of long (>200 nt non-coding RNAs (lncRNAs are transcribed from mammalian genomes and that lncRNAs perform important regulatory functions in various developmental processes. However, the expression of lncRNAs and their biological functions in post-natal testis development remain unknown. In this study, we employed microarray technology to examine lncRNA expression profiles of neonatal (6-day-old and adult (8-week-old mouse testes. We found that 8,265 lncRNAs were expressed above background levels during post-natal testis development, of which 3,025 were differentially expressed. Candidate lncRNAs were identified for further characterization by an integrated examination of genomic context, gene ontology (GO enrichment of their associated protein-coding genes, promoter analysis for epigenetic modification, and evolutionary conservation of elements. Many lncRNAs overlapped or were adjacent to key transcription factors and other genes involved in spermatogenesis, such as Ovol1, Ovol2, Lhx1, Sox3, Sox9, Plzf, c-Kit, Wt1, Sycp2, Prm1 and Prm2. Most differentially expressed lncRNAs exhibited epigenetic modification marks similar to protein-coding genes and tend to be expressed in a tissue-specific manner. In addition, the majority of differentially expressed lncRNAs harbored evolutionary conserved elements. Taken together, our findings represent the first systematic investigation of lncRNA expression in the mammalian testis and provide a solid foundation for further research into the molecular mechanisms of lncRNAs function in mammalian testis development and spermatogenesis.

Optimization and analysis of a quantitative real-time PCR-based technique to determine microRNA expression in formalin-fixed paraffin-embedded samples

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Reis Patricia P

2010-06-01

Full Text Available Abstract Background MicroRNAs (miRs are non-coding RNA molecules involved in post-transcriptional regulation, with diverse functions in tissue development, differentiation, cell proliferation and apoptosis. miRs may be less prone to degradation during formalin fixation, facilitating miR expression studies in formalin-fixed paraffin-embedded (FFPE tissue. Results Our study demonstrates that the TaqMan Human MicroRNA Array v1.0 (Early Access platform is suitable for miR expression analysis in FFPE tissue with a high reproducibility (correlation coefficients of 0.95 between duplicates, p 35, we show that reproducibility between technical replicates, equivalent dilutions, and FFPE vs. frozen samples is best in the high abundance stratum. We also demonstrate that the miR expression profiles of FFPE samples are comparable to those of fresh-frozen samples, with a correlation of up to 0.87 (p Conclusion Our study thus demonstrates the utility, reproducibility, and optimization steps needed in miR expression studies using FFPE samples on a high-throughput quantitative PCR-based miR platform, opening up a realm of research possibilities for retrospective studies.

The rectal cancer microRNAome - microRNA expression in rectal cancer and matched normal mucosa

DEFF Research Database (Denmark)

Gaedcke, Jochen; Grade, Marian; Camps, Jordi

2012-01-01

PURPOSE: miRNAs play a prominent role in a variety of physiologic and pathologic biologic processes, including cancer. For rectal cancers, only limited data are available on miRNA expression profiles, whereas the underlying genomic and transcriptomic aberrations have been firmly established. We t...

Roles of microRNA-15 family in normal and pathological late lung development

OpenAIRE

Sakkas, Elpidoforos

2016-01-01

MicroRNAs are key regulators of organogenesis and during the last years many studies focused on microRNA expression during embryonic development. To date, there is no study to report possible roles of microRNAs in late lung development and especially during the alveolarization process. The objective of this study was to identify microRNAs that are deregulated under hyperoxic conditions and to assess whether microRNA expression can be modulated in vivo. Lung microRNA expression screening wa...

Thiol-linked alkylation of RNA to assess expression dynamics.

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Herzog, Veronika A; Reichholf, Brian; Neumann, Tobias; Rescheneder, Philipp; Bhat, Pooja; Burkard, Thomas R; Wlotzka, Wiebke; von Haeseler, Arndt; Zuber, Johannes; Ameres, Stefan L

2017-12-01

Gene expression profiling by high-throughput sequencing reveals qualitative and quantitative changes in RNA species at steady state but obscures the intracellular dynamics of RNA transcription, processing and decay. We developed thiol(SH)-linked alkylation for the metabolic sequencing of RNA (SLAM seq), an orthogonal-chemistry-based RNA sequencing technology that detects 4-thiouridine (s 4 U) incorporation in RNA species at single-nucleotide resolution. In combination with well-established metabolic RNA labeling protocols and coupled to standard, low-input, high-throughput RNA sequencing methods, SLAM seq enabled rapid access to RNA-polymerase-II-dependent gene expression dynamics in the context of total RNA. We validated the method in mouse embryonic stem cells by showing that the RNA-polymerase-II-dependent transcriptional output scaled with Oct4/Sox2/Nanog-defined enhancer activity, and we provide quantitative and mechanistic evidence for transcript-specific RNA turnover mediated by post-transcriptional gene regulatory pathways initiated by microRNAs and N 6 -methyladenosine. SLAM seq facilitates the dissection of fundamental mechanisms that control gene expression in an accessible, cost-effective and scalable manner.

NMD and microRNA expression profiling of the HPCX1 locus reveal MAGEC1 as a candidate prostate cancer predisposition gene

International Nuclear Information System (INIS)

Mattila, Henna; Schindler, Martin; Isotalo, Jarkko; Ikonen, Tarja; Vihinen, Mauno; Oja, Hannu; Tammela, Teuvo LJ; Wahlfors, Tiina; Schleutker, Johanna

2011-01-01

Several predisposition loci for hereditary prostate cancer (HPC) have been suggested, including HPCX1 at Xq27-q28, but due to the complex structure of the region, the susceptibility gene has not yet been identified. In this study, nonsense-mediated mRNA decay (NMD) inhibition was used for the discovery of truncating mutations. Six prostate cancer (PC) patients and their healthy brothers were selected from a group of HPCX1-linked families. Expression analyses were done using Agilent 44 K oligoarrays, and selected genes were screened for mutations by direct sequencing. In addition, microRNA expression levels in the lymphoblastic cells were analyzed to trace variants that might alter miRNA expression and explain partly an inherited genetic predisposion to PC. Seventeen genes were selected for resequencing based on the NMD array, but no truncating mutations were found. The most interesting variant was MAGEC1 p.Met1?. An association was seen between the variant and unselected PC (OR = 2.35, 95% CI = 1.10-5.02) and HPC (OR = 3.38, 95% CI = 1.10-10.40). miRNA analysis revealed altogether 29 miRNAs with altered expression between the PC cases and controls. miRNA target analysis revealed that 12 of them also had possible target sites in the MAGEC1 gene. These miRNAs were selected for validation process including four miRNAs located in the X chromosome. The expressions of 14 miRNAs were validated in families that contributed to the significant signal differences in Agilent arrays. Further functional studies are needed to fully understand the possible contribution of these miRNAs and MAGEC1 start codon variant to PC

Expression levels of the microRNA maturing microprocessor complex component DGCR8 and the RNA-induced silencing complex (RISC) components argonaute-1, argonaute-2, PACT, TARBP1, and TARBP2 in epithelial skin cancer.

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Sand, Michael; Skrygan, Marina; Georgas, Dimitrios; Arenz, Christoph; Gambichler, Thilo; Sand, Daniel; Altmeyer, Peter; Bechara, Falk G

2012-11-01

The microprocessor complex mediates intranuclear biogenesis of precursor microRNAs from the primary microRNA transcript. Extranuclear, mature microRNAs are incorporated into the RNA-induced silencing complex (RISC) before interaction with complementary target mRNA leads to transcriptional repression or cleavage. In this study, we investigated the expression profiles of the microprocessor complex subunit DiGeorge syndrome critical region gene 8 (DGCR8) and the RISC components argonaute-1 (AGO1), argonaute-2 (AGO2), as well as double-stranded RNA-binding proteins PACT, TARBP1, and TARBP2 in epithelial skin cancer and its premalignant stage. Patients with premalignant actinic keratoses (AK, n = 6), basal cell carcinomas (BCC, n = 15), and squamous cell carcinomas (SCC, n = 7) were included in the study. Punch biopsies were harvested from the center of the tumors (lesional), from healthy skin sites (intraindividual controls), and from healthy skin sites in a healthy control group (n = 16; interindividual control). The DGCR8, AGO1, AGO2, PACT, TARBP1, and TARBP2 mRNA expression levels were detected by quantitative real-time reverse transcriptase polymerase chain reaction. The DGCR8, AGO1, AGO2, PACT, and TARBP1 expression levels were significantly higher in the AK, BCC, and SCC groups than the healthy controls (P  0.05). This study indicates that major components of the miRNA pathway, such as the microprocessor complex and RISC, are dysregulated in epithelial skin cancer. Copyright © 2011 Wiley Periodicals, Inc.

A conformation-induced fluorescence method for microRNA detection

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Aw, Sherry S; Tang, Melissa Xm; Teo, Yin Nah

2016-01-01

and quantify microRNAs may aid research into novel aspects of microRNA biology and contribute to the development of diagnostics. By introducing an additional stem loop into the fluorescent RNA Spinach and altering its 3' and 5' ends, we have generated a new RNA, Pandan, that functions as the basis for a micro......MicroRNAs play important roles in a large variety of biological systems and processes through their regulation of target mRNA expression, and show promise as clinical biomarkers. However, their small size presents challenges for tagging or direct detection. Innovation in techniques to sense......RNA sensor. Pandan contains two sequence-variable stem loops that encode complementary sequence for a target microRNA of interest. In its sensor form, it requires the binding of a target microRNA in order to reconstitute the RNA scaffold for fluorophore binding and fluorescence. Binding of the target micro...

Customization of Artificial MicroRNA Design.

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Van Vu, Tien; Do, Vinh Nang

2017-01-01

RNAi approaches, including microRNA (miRNA) regulatory pathway, offer great tools for functional characterization of unknown genes. Moreover, the applications of artificial microRNA (amiRNA) in the field of plant transgenesis have also been advanced to engineer pathogen-resistant or trait-improved transgenic plants. Until now, despite the high potency of amiRNA approach, no commercial plant cultivar expressing amiRNAs with improved traits has been released yet. Beside the issues of biosafety policies, the specificity and efficacy of amiRNAs are of major concerns. Sufficient cares should be taken for the specificity and efficacy of amiRNAs due to their potential off-target effects and other issues relating to in vivo expression of pre-amiRNAs. For these reasons, the proper design of amiRNAs with the lowest off-target possibility is very important for successful applications of the approach in plant. Therefore, there are many studies with the aim to improve the amiRNA design and amiRNA expressing backbones for obtaining better specificity and efficacy. However, the requirement for an efficient reference for the design is still needed. In the present chapter, we attempt to summarize and discuss all the major concerns relating to amiRNA design with the hope to provide a significant guideline for this approach.

microRNA Response to Listeria monocytogenes Infection in Epithelial Cells

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Izar, Benjamin; Mannala, Gopala Krishna; Mraheil, Mobarak Abu; Chakraborty, Trinad; Hain, Torsten

2012-01-01

microRNAs represent a family of very small non-coding RNAs that control several physiologic and pathologic processes, including host immune response and cancer by antagonizing a number of target mRNAs. There is limited knowledge about cell expression and the regulatory role of microRNAs following bacterial infections. We investigated whether infection with a Gram-positive bacterium leads to altered expression of microRNAs involved in the host cell response in epithelial cells. Caco-2 cells were infected with Listeria monocytogenes EGD-e, a mutant strain (ΔinlAB or Δhly) or incubated with purified listeriolysin (LLO). Total RNA was isolated and microRNA and target gene expression was compared to the expression in non-infected cells using microRNA microarrays and qRT-PCR. We identified and validated five microRNAs (miR- 146b, miR-16, let-7a1, miR-145 and miR-155) that were significantly deregulated following listerial infection. We show that expression patterns of particular microRNAs strongly depend on pathogen localization and the presence of bacterial effector proteins. Strikingly, miR-155 which was shown to have an important role in inflammatory responses during infection was induced by wild-type bacteria, by LLO-deficient bacteria and following incubation with purified LLO. It was downregulated following ΔinlAB infection indicating a new potent role for internalins in listerial pathogenicity and miRNA regulation. Concurrently, we observed differences in target transcript expression of the investigated miRNAs. We provide first evidence that L. monocytogenes infection leads to deregulation of a set of microRNAs with important roles in host response. Distinct microRNA expression depends on both LLO and pathogen localization. PMID:22312311

Extract of Stellerachamaejasme L(ESC) inhibits growth and metastasis of human hepatocellular carcinoma via regulating microRNA expression.

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Liu, Xiaoni; Wang, Shuang; Xu, Jianji; Kou, Buxin; Chen, Dexi; Wang, Yajie; Zhu, Xiaoxin

2018-03-20

MicroRNAs(miRNAs)are involved in the initiation and progression of hepatocellular carcinoma. ESC, an extract of Stellerachamaejasme L, had been confirmed as a potential anti-tumor extract of Traditional Chinese Medicine. In light of the important role of miRNAs in hepatocellular carcinoma, we questioned whether the inhibitory effects of ESC on hepatocellular carcinoma (HCC) were associated with miRNAs. The proliferation inhibition of ESC on HCC cells was measured with MTT assay. The migration inhibition of ESC on HCC cells was measured with transwell assay. The influences of ESC on growth and metastasis inhibition were evaluated with xenograft tumor model of HCC. Protein expressions were measured with western blot and immunofluorescence methods and miRNA profiles were detected with miRNA array. Differential miRNA and target mRNAs were verified with real-time PCR. The results showed that ESC could inhibit proliferation and epithelial mesenchymal transition (EMT) in HCC cells in vitro and tumor growth and metastasis in xenograft models in vivo. miRNA array results showed that 69 differential miRNAs in total of 429 ones were obtained in MHCC97H cells treated by ESC. hsa-miR-107, hsa-miR-638, hsa-miR-106b-5p were selected to be validated with real-time PCR method in HepG2 and MHCC97H cells. Expressions of hsa-miR-107 and hsa-miR-638 increased obviously in HCC cells treated by ESC. Target genes of three miRNAs were also validated with real-time PCR. Interestingly, only target genes of hsa-miR-107 changed greatly. ESC downregulated the MCL1, SALL4 and BCL2 gene expressions significantly but did not influence the expression of CACNA2D1. The findings suggested ESC regressed growth and metastasis of human hepatocellular carcinoma via regulating microRNAs expression and their corresponding target genes.

MicroRNA-33 promotes the replicative senescence of mouse embryonic fibroblasts by suppressing CDK6

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Xu, Shun; Huang, Haijiao; Li, Nanhong; Zhang, Bing; Jia, Yubin; Yang, Yukun; Yuan, Yuan; Xiong, Xing-dong; Wang, Dengchuan; Zheng, Hui-ling [Institute of Aging Research, Guangdong Medical University, Dongguan (China); Guangdong Provincial Key Laboratory of Medical Molecular Diagnostics, Dongguan (China); Institute of Biochemistry & Molecular Biology, Guangdong Medical University, Zhanjiang (China); Liu, Xinguang, E-mail: xgliu64@126.com [Institute of Aging Research, Guangdong Medical University, Dongguan (China); Guangdong Provincial Key Laboratory of Medical Molecular Diagnostics, Dongguan (China); Institute of Biochemistry & Molecular Biology, Guangdong Medical University, Zhanjiang (China)

2016-05-13

MicroRNAs are a large class of tiny noncoding RNAs, which have emerged as critical regulators of gene expression, and thus are involved in multiple cellular processes, including cellular senescence. MicroRNA-33 has previously been established to exert crucial effect on cell proliferation, lipid metabolism and cholesterol metabolism. Nonetheless, the association between microRNA-33 and cellular senescence and its underlying molecular mechanism are far to be elucidated. The present study has attempted to probe into the effect of microRNA-33 on MEFs senescence. Our data unveiled that microRNA-33 was dramatically down-regulated in senescent MEFs compared to the young MEFs, and ectopic expression of microRNA-33 promoted MEFs senescence, while knock-down of microRNA-33 exhibited a protective effect against senescence phenotype. Moreover, we verified CDK6 as a direct target of microRNA-33 in mouse. Silencing of CDK6 induced the premature senescence phenotype of MEFs similarly as microRNA-33, while enforced expression of CDK6 significantly reverse the senescence-induction effect of microRNA-33. Taken together, our results suggested that microRNA-33 enhanced the replicative senescence of MEFs potentially by suppressing CDK6 expression. -- Highlights: •MicroRNA-33 was dramatically down-regulated in senescent MEF cells. •Altered expression of microRNA-33 exerted a critical role in MEFs senescence. •MicroRNA-33 promoted the replicative senescence of MEFs via targeting of CDK6.

MicroRNA-33 promotes the replicative senescence of mouse embryonic fibroblasts by suppressing CDK6

International Nuclear Information System (INIS)

Xu, Shun; Huang, Haijiao; Li, Nanhong; Zhang, Bing; Jia, Yubin; Yang, Yukun; Yuan, Yuan; Xiong, Xing-dong; Wang, Dengchuan; Zheng, Hui-ling; Liu, Xinguang

2016-01-01

MicroRNAs are a large class of tiny noncoding RNAs, which have emerged as critical regulators of gene expression, and thus are involved in multiple cellular processes, including cellular senescence. MicroRNA-33 has previously been established to exert crucial effect on cell proliferation, lipid metabolism and cholesterol metabolism. Nonetheless, the association between microRNA-33 and cellular senescence and its underlying molecular mechanism are far to be elucidated. The present study has attempted to probe into the effect of microRNA-33 on MEFs senescence. Our data unveiled that microRNA-33 was dramatically down-regulated in senescent MEFs compared to the young MEFs, and ectopic expression of microRNA-33 promoted MEFs senescence, while knock-down of microRNA-33 exhibited a protective effect against senescence phenotype. Moreover, we verified CDK6 as a direct target of microRNA-33 in mouse. Silencing of CDK6 induced the premature senescence phenotype of MEFs similarly as microRNA-33, while enforced expression of CDK6 significantly reverse the senescence-induction effect of microRNA-33. Taken together, our results suggested that microRNA-33 enhanced the replicative senescence of MEFs potentially by suppressing CDK6 expression. -- Highlights: •MicroRNA-33 was dramatically down-regulated in senescent MEF cells. •Altered expression of microRNA-33 exerted a critical role in MEFs senescence. •MicroRNA-33 promoted the replicative senescence of MEFs via targeting of CDK6.

MicroRNA-10b expression in breast cancer and its clinical association.

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Jianhui Zhang

Full Text Available MicroRNAs (miRNAs are short non-coding RNA molecules that play a significant role in many types of cancers including breast cancer. In the current study, we evaluated the expression levels of microR-10b (miR-10b in 115 breast cancer patients from Sichuan Cancer Center. Real time reverse transcription-PCR was used to assess miR-10b expression. Clinical data including disease stage, survival status, age, ER/PR/HER2 status, molecular subtypes, tumor size, lymph node status and Ki-67 expression levels were correlated with miR-10b expression levels. Our data showed that the miR-10b expression is correlated with disease stage, living status and tumor sizes. We also found that miR-10b expression levels are higher in the lymph node positive group and the Ki-67 higher scoring group (score > 20. No statistically significant differences were observed based on age or molecular sub-type grouping. In conclusion, miR-10b may be a biomarker for breast cancer and is a potential treatment target.

Cross disease analysis of co-functional microRNA pairs on a reconstructed network of disease-gene-microRNA tripartite.

Science.gov (United States)

Peng, Hui; Lan, Chaowang; Zheng, Yi; Hutvagner, Gyorgy; Tao, Dacheng; Li, Jinyan

2017-03-24

MicroRNAs always function cooperatively in their regulation of gene expression. Dysfunctions of these co-functional microRNAs can play significant roles in disease development. We are interested in those multi-disease associated co-functional microRNAs that regulate their common dysfunctional target genes cooperatively in the development of multiple diseases. The research is potentially useful for human disease studies at the transcriptional level and for the study of multi-purpose microRNA therapeutics. We designed a computational method to detect multi-disease associated co-functional microRNA pairs and conducted cross disease analysis on a reconstructed disease-gene-microRNA (DGR) tripartite network. The construction of the DGR tripartite network is by the integration of newly predicted disease-microRNA associations with those relationships of diseases, microRNAs and genes maintained by existing databases. The prediction method uses a set of reliable negative samples of disease-microRNA association and a pre-computed kernel matrix instead of kernel functions. From this reconstructed DGR tripartite network, multi-disease associated co-functional microRNA pairs are detected together with their common dysfunctional target genes and ranked by a novel scoring method. We also conducted proof-of-concept case studies on cancer-related co-functional microRNA pairs as well as on non-cancer disease-related microRNA pairs. With the prioritization of the co-functional microRNAs that relate to a series of diseases, we found that the co-function phenomenon is not unusual. We also confirmed that the regulation of the microRNAs for the development of cancers is more complex and have more unique properties than those of non-cancer diseases.

Integrated Analysis of Long Noncoding RNA and mRNA Expression Profile in Advanced Laryngeal Squamous Cell Carcinoma.

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Ling Feng

Full Text Available Long non-coding RNA (lncRNA plays an important role in tumorigenesis. However, the expression pattern and function of lncRNAs in laryngeal squamous cell carcinoma (LSCC are still unclear. To investigate the aberrantly expressed lncRNAs and mRNAs in advanced LSCC, we screened lncRNA and mRNA expression profiles in 9 pairs of primary Stage IVA LSCC tissues and adjacent non-neoplastic tissues by lncRNA and mRNA integrated microarrays. Gene Ontology and pathway analysis were performed to find out the significant function and pathway of the differentially expressed mRNAs, gene-gene functional interaction network and ceRNA network were constructed to select core mRNAs, and lncRNA-mRNA expression correlation network was built to identify the interactions between lncRNA and mRNA. qRT-PCR was performed to further validate the expressions of selected lncRNAs and mRNAs in advanced LSCC. We found 1459 differentially expressed lncRNAs and 2381 differentially expressed mRNAs, including 846 up-regulated lncRNAs and 613 down-regulated lncRNAs, 1542 up-regulated mRNAs and 839 down-regulated mRNAs. The mRNAs ITGB1, HIF1A, and DDIT4 were selected as core mRNAs, which are mainly involved in biological processes, such as matrix organization, cell cycle, adhesion, and metabolic pathway. LncRNA-mRNA expression correlation network showed LncRNA NR_027340, MIR31HG were positively correlated with ITGB1, HIF1A respectively. LncRNA SOX2-OT was negatively correlated with DDIT4. qRT-PCR further validated the expression of these lncRNAs and mRNAs. The work provides convincing evidence that the identified lncRNAs and mRNAs are potential biomarkers in advanced LSCC for further future studies.

Changes in miRNA Expression Profiling during Neuronal Differentiation and Methyl Mercury-Induced Toxicity in Human in Vitro Models

Directory of Open Access Journals (Sweden)

Giorgia Pallocca

2014-08-01

Full Text Available MicroRNAs (miRNAs are implicated in the epigenetic regulation of several brain developmental processes, such as neurogenesis, neuronal differentiation, neurite outgrowth, and synaptic plasticity. The main aim of this study was to evaluate whether miRNA expression profiling could be a useful approach to detect in vitro developmental neurotoxicity. For this purpose, we assessed the changes in miRNA expression caused by methyl mercury chloride (MeHgCl, a well-known developmental neurotoxicant, comparing carcinoma pluripotent stem cells (NT-2 with human embryonic stem cells (H9, both analyzed during the early stage of neural progenitor commitment into neuronal lineage. The data indicate the activation of two distinct miRNA signatures, one activated upon neuronal differentiation and another upon MeHgCl-induced toxicity. Particularly, exposure to MeHgCl elicited, in both neural models, the down-regulation of the same six out of the ten most up-regulated neuronal pathways, as shown by the up-regulation of the corresponding miRNAs and further assessment of gene ontology (GO term and pathway enrichment analysis. Importantly, some of these common miRNA-targeted pathways defined in both cell lines are known to play a role in critical developmental processes, specific for neuronal differentiation, such as axon guidance and neurotrophin-regulated signaling. The obtained results indicate that miRNAs expression profiling could be a promising tool to assess developmental neurotoxicity pathway perturbation, contributing towards improved predictive human toxicity testing.

Comprehensive microRNA profiling reveals potential augmentation of the IL1 pathway in rheumatic heart valve disease.

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Lu, Qiyu; Sun, Yi; Duan, Yuyin; Li, Bin; Xia, Jianming; Yu, Songhua; Zhang, Guimin

2018-03-16

Valvular heart disease is a leading cause of cardiovascular mortality, especially in China. More than a half of valvular heart diseases are caused by acute rheumatic fever. microRNA is involved in many physiological and pathological processes. However, the miRNA profile of the rheumatic valvular heart disease is unknown. This research is to discuss microRNAs and their target gene pathways involved in rheumatic heart valve disease. Serum miRNA from one healthy individual and four rheumatic heart disease patients were sequenced. Specific differentially expressed miRNAs were quantified by Q-PCR in 40 patients, with 20 low-to-moderate rheumatic mitral valve stenosis patients and 20 severe mitral valve stenosis patients. The target relationship between certain miRNA and predicted target genes were analysis by Luciferase reporter assay. The IL-1β and IL1R1 expression levels were analyzed by immunohistochemistry and western blot in the mitral valve from surgery of mitral valve replacement. The results showed that 13 and 91 miRNAs were commonly upregulated or downregulated in all four patients. Nine miRNAs, 1 upregulated and 8 downregulated, that had a similar fold change in all 4 patients were selected for quantitative PCR verification. The results showed similar results from miRNA sequencing. Within these 9 tested miRNAs, hsa-miR-205-3p and hsa-miR-3909 showed a low degree of dispersion between the members of each group. Hsa miR-205-3p and hsa-miR-3909 were predicted to target the 3'UTR of IL-1β and IL1R1 respectively. This was verified by luciferase reporter assays. Immunohistochemistry and Western blot results showed that the mitral valve from rheumatic valve heart disease showed higher levels of IL- 1β and IL1R1 expression compared with congenital heart valve disease. This suggested a difference between rheumatic heart valve disease and other types of heart valve diseases, with more inflammatory responses in the former. In the present study, by next generation

Small Molecule, Big Prospects: MicroRNA in Pregnancy and Its Complications

Directory of Open Access Journals (Sweden)

Meng Cai

2017-01-01

Full Text Available MicroRNAs are small, noncoding RNA molecules that regulate target gene expression in the posttranscriptional level. Unlike siRNA, microRNAs are “fine-tuners” rather than “switches” in the regulation of gene expression; thus they play key roles in maintaining tissue homeostasis. The aberrant microRNA expression is implicated in the disease process. To date, numerous studies have demonstrated the regulatory roles of microRNAs in various pathophysiological conditions. In contrast, the study of microRNA in pregnancy and its associated complications, such as preeclampsia (PE, fetal growth restriction (FGR, and preterm labor, is a young field. Over the last decade, the knowledge of pregnancy-related microRNAs has increased and the molecular mechanisms by which microRNAs regulate pregnancy or its associated complications are emerging. In this review, we focus on the recent advances in the research of pregnancy-related microRNAs, especially their function in pregnancy-associated complications and the potential clinical applications. Here microRNAs that associate with pregnancy are classified as placenta-specific, placenta-associated, placenta-derived circulating, and uterine microRNA according to their localization and origin. MicroRNAs offer a great potential for developing diagnostic and therapeutic targets in pregnancy-related disorders.

The ROS-sensitive microRNA-9/9* controls the expression of mitochondrial tRNA-modifying enzymes and is involved in the molecular mechanism of MELAS syndrome.

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Meseguer, Salvador; Martínez-Zamora, Ana; García-Arumí, Elena; Andreu, Antonio L; Armengod, M-Eugenia

2015-01-01

Mitochondrial dysfunction activates mitochondria-to-nucleus signaling pathways whose components are mostly unknown. Identification of these components is important to understand the molecular mechanisms underlying mitochondrial diseases and to discover putative therapeutic targets. MELAS syndrome is a rare neurodegenerative disease caused by mutations in mitochondrial (mt) DNA affecting mt-tRNA(Leu(UUR)). Patient and cybrid cells exhibit elevated oxidative stress. Moreover, mutant mt-tRNAs(Leu(UUR)) lack the taurine-containing modification normally present at the wobble uridine (U34) of wild-type mt-tRNA(Leu(UUR)), which is considered an etiology of MELAS. However, the molecular mechanism is still unclear. We found that MELAS cybrids exhibit a significant decrease in the steady-state levels of several mt-tRNA-modification enzymes, which is not due to transcriptional regulation. We demonstrated that oxidative stress mediates an NFkB-dependent induction of microRNA-9/9*, which acts as a post-transcriptional negative regulator of the mt-tRNA-modification enzymes GTPBP3, MTO1 and TRMU. Down-regulation of these enzymes by microRNA-9/9* affects the U34 modification status of non-mutant tRNAs and contributes to the MELAS phenotype. Anti-microRNA-9 treatments of MELAS cybrids reverse the phenotype, whereas miR-9 transfection of wild-type cells mimics the effects of siRNA-mediated down-regulation of GTPBP3, MTO1 and TRMU. Our data represent the first evidence that an mt-DNA disease can directly affect microRNA expression. Moreover, we demonstrate that the modification status of mt-tRNAs is dynamic and that cells respond to stress by modulating the expression of mt-tRNA-modifying enzymes. microRNA-9/9* is a crucial player in mitochondria-to-nucleus signaling as it regulates expression of nuclear genes in response to changes in the functional state of mitochondria. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email