Decreased Expression of miR-21, miR-26a, miR-29a, and miR-142-3p in CD4+ T Cells and Peripheral Blood from Tuberculosis Patients

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Schattling, Stefanie; Kohns, Malte; Sander-Jülch, Claudia; Walzl, Gerhard; Hesseling, Anneke; Mayatepek, Ertan; Fleischer, Bernhard; Marx, Florian M.; Jacobsen, Marc

2013-01-01

The vast majority of Mycobacterium tuberculosis (M. tuberculosis) infected individuals are protected from developing tuberculosis and T cells are centrally involved in this process. MicroRNAs (miRNA) regulate T-cell functions and are biomarker candidates of disease susceptibility and treatment efficacy in M. tuberculosis infection. We determined the expression profile of 29 selected miRNAs in CD4+ T cells from tuberculosis patients and contacts with latent M. tuberculosis infection (LTBI). These analyses showed lower expression of miR-21, miR-26a, miR-29a, and miR-142-3p in CD4+ T cells from tuberculosis patients. Whole blood miRNA candidate analyses verified decreased expression of miR-26a, miR-29a, and miR-142-3p in children with tuberculosis as compared to healthy children with LTBI. Despite marked variances between individual donor samples, trends of increased miRNA candidate expression during treatment and recovery were observed. Functional in vitro analysis identified increased miR-21 and decreased miR-26a expression after re-stimulation of T cells. In vitro polarized Interleukin-17 positive T-cell clones showed activation-dependent miR-29a up-regulation. In order to characterize the role of miR-29a (a described suppressor of Interferon-γ in tuberculosis), we analyzed M. tuberculosis specific Interferon-γ expressing T cells in children with tuberculosis and healthy contacts but detected no correlation between miR-29a and Interferon-γ expression. Suppression of miR-29a in primary human T cells by antagomirs indicated no effect on Interferon-γ expression after in vitro activation. Finally, classification of miRNA targets revealed only a moderate overlap between the candidates. This may reflect differential roles of miR-21, miR-26a, miR-29a, and miR-142-3p in T-cell immunity against M. tuberculosis infection and disease. PMID:23613882

Differential expression of miR-139, miR-486 and miR-21 in breast cancer patients sub-classified according to lymph node status

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Rask, Lene; Balslev, Eva; Søkilde, Rolf

2014-01-01

PURPOSE: Therapeutic decisions in breast cancer are increasingly guided by prognostic and predictive biomarkers. Non-protein-coding microRNAs (miRNAs) have recently been found to be deregulated in breast cancers and, in addition, to be correlated with several clinico-pathological features. One...... of the most consistently up-regulated miRNAs is miR-21. Here, we specifically searched for differentially expressed miRNAs in high-risk breast cancer patients as compared to low-risk breast cancer patients. In the same patients, we also compared miR-21 expression with the expression of its presumed target...... PTEN. METHODS: Both microarray and RT-qPCR techniques were used to assess miRNA expression levels in lymph node-positive and -negative human invasive ductal carcinoma tissues. Simultaneously, PTEN protein expression levels were assessed using immunohistochemistry. RESULTS: miR-486-5p and miR-139-5p...

Targeting of the tumor suppressor GRHL3 by a miR-21-dependent proto-oncogenic network results in PTEN loss and tumorigenesis.

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Darido, Charbel; Georgy, Smitha R; Wilanowski, Tomasz; Dworkin, Sebastian; Auden, Alana; Zhao, Quan; Rank, Gerhard; Srivastava, Seema; Finlay, Moira J; Papenfuss, Anthony T; Pandolfi, Pier Paolo; Pearson, Richard B; Jane, Stephen M

2011-11-15

Despite its prevalence, the molecular basis of squamous cell carcinoma (SCC) remains poorly understood. Here, we identify the developmental transcription factor Grhl3 as a potent tumor suppressor of SCC in mice, and demonstrate that targeting of Grhl3 by a miR-21-dependent proto-oncogenic network underpins SCC in humans. Deletion of Grhl3 in adult epidermis evokes loss of expression of PTEN, a direct GRHL3 target, resulting in aggressive SCC induced by activation of PI3K/AKT/mTOR signaling. Restoration of Pten expression completely abrogates SCC formation. Reduced levels of GRHL3 and PTEN are evident in human skin, and head and neck SCC, associated with increased expression of miR-21, which targets both tumor suppressors. Our data define the GRHL3-PTEN axis as a critical tumor suppressor pathway in SCC. 2011 Elsevier Inc. All rights reserved.

Establishment of Lipofection Protocol for Efficient miR-21 Transfection into Cortical Neurons In Vitro.

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Han, Zhaoli; Ge, Xintong; Tan, Jin; Chen, Fanglian; Gao, Huabin; Lei, Ping; Zhang, Jianning

2015-12-01

Dysregulated microRNAs in neurons could cause many nervous system diseases. The therapeutic manipulation of these pathogenic microRNAs necessitates novel, efficient delivery systems to facilitate microRNA modulators targeting neurons with minimal off-target effects. The study aimed to establish a lipofection protocol to upregulate expression levels of miR-21 in neurons under different conditions, including different serum-free medium, transfection conditions, and reagent concentration, by evaluating the expression levels of miR-21 and neuron injury. The expression levels of miR-21 were higher in neurons transfected by Neurobasal-A than by DMEM. Expression levels of miR-21 were already the highest at the ratio RNAiMAX:miR-21 = 3:5, but the increase of RNAiMAX's concentration had not caused the further upregulation of expression level of miR-21. Neuron injury was condition dependent and dose dependent after transfection. Compared to S-Neurobasal groups, neurons have a smaller injury in N-Neurobasal groups, and compared to ratios RNAiMAX:miR-21 = 4:5, 5:5, neuron injury was smaller at ratios of RNAiMAX:miR-21 = 1:5, 2:5, 3:5. Without the pretreatment of starvation in vitro, the lipofection protocol was that RNAiMAX/miR-21 agomir complexes were diluted in Neurobasal-A at the ratio of RNAiMAX:miR-21 = 3:5.

Decreased expression of miR-21, miR-26a, miR-29a, and miR-142-3p in CD4⁺ T cells and peripheral blood from tuberculosis patients.

Science.gov (United States)

Kleinsteuber, Katja; Heesch, Kerrin; Schattling, Stefanie; Kohns, Malte; Sander-Jülch, Claudia; Walzl, Gerhard; Hesseling, Anneke; Mayatepek, Ertan; Fleischer, Bernhard; Marx, Florian M; Jacobsen, Marc

2013-01-01

The vast majority of Mycobacterium tuberculosis (M. tuberculosis) infected individuals are protected from developing tuberculosis and T cells are centrally involved in this process. MicroRNAs (miRNA) regulate T-cell functions and are biomarker candidates of disease susceptibility and treatment efficacy in M. tuberculosis infection. We determined the expression profile of 29 selected miRNAs in CD4(+) T cells from tuberculosis patients and contacts with latent M. tuberculosis infection (LTBI). These analyses showed lower expression of miR-21, miR-26a, miR-29a, and miR-142-3p in CD4(+) T cells from tuberculosis patients. Whole blood miRNA candidate analyses verified decreased expression of miR-26a, miR-29a, and miR-142-3p in children with tuberculosis as compared to healthy children with LTBI. Despite marked variances between individual donor samples, trends of increased miRNA candidate expression during treatment and recovery were observed. Functional in vitro analysis identified increased miR-21 and decreased miR-26a expression after re-stimulation of T cells. In vitro polarized Interleukin-17 positive T-cell clones showed activation-dependent miR-29a up-regulation. In order to characterize the role of miR-29a (a described suppressor of Interferon-γ in tuberculosis), we analyzed M. tuberculosis specific Interferon-γ expressing T cells in children with tuberculosis and healthy contacts but detected no correlation between miR-29a and Interferon-γ expression. Suppression of miR-29a in primary human T cells by antagomirs indicated no effect on Interferon-γ expression after in vitro activation. Finally, classification of miRNA targets revealed only a moderate overlap between the candidates. This may reflect differential roles of miR-21, miR-26a, miR-29a, and miR-142-3p in T-cell immunity against M. tuberculosis infection and disease.

Declining Physical Performance Associates with Serum FasL, miR-21, and miR-146a in Aging Sprinters

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Reeta Kangas

2017-01-01

Full Text Available Aging is associated with systemic inflammation and cellular apoptosis accelerating physiological dysfunctions. Whether physically active way of life affects these associations is unclear. This study measured the levels of serum inflammatory and apoptotic molecules, their change over 10 years, and their associations with physical performance in sprint-trained male athletes. HsCRP, cell counts, HGB, FasL, miR-21, and miR-146a were measured cross-sectionally (n=67, 18–90 yrs and serum FasL, miR-21, and miR-146a and their aging-related associations with physical performance were assessed over a 10-year follow-up (n=49, 50–90 yrs. The cross-sectional study showed positive age correlations for neutrophils and negative for lymphocytes, red blood cells, HGB, FasL, and miR-146a. During the 10-year follow-up, FasL decreased (P=0.017 and miR-21 (P<0.001 and miR-146a (P=0.005 levels increased. When combining the molecule levels, aging, and physical performance, FasL associated with countermovement jump and bench press (P<0.001, miR-21 and miR-146a with knee flexion (P=0.023; P<0.001, and bench press (P=0.004; P<0.001 and miR-146a with sprint performance (P<0.001. The studied serum molecules changed in an age-dependent manner and were associated with declining physical performance. They have potential as biomarkers of aging-related processes influencing the development of physiological dysfunctions. Further research is needed focusing on the origins and targets of circulating microRNAs to clarify their function in various tissues with aging.

Macrophage deficiency of miR-21 promotes apoptosis, plaque necrosis, and vascular inflammation during atherogenesis.

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Canfrán-Duque, Alberto; Rotllan, Noemi; Zhang, Xinbo; Fernández-Fuertes, Marta; Ramírez-Hidalgo, Cristina; Araldi, Elisa; Daimiel, Lidia; Busto, Rebeca; Fernández-Hernando, Carlos; Suárez, Yajaira

2017-09-01

Atherosclerosis, the major cause of cardiovascular disease, is a chronic inflammatory disease characterized by the accumulation of lipids and inflammatory cells in the artery wall. Aberrant expression of microRNAs has been implicated in the pathophysiological processes underlying the progression of atherosclerosis. Here, we define the contribution of miR-21 in hematopoietic cells during atherogenesis. Interestingly, we found that miR-21 is the most abundant miRNA in macrophages and its absence results in accelerated atherosclerosis, plaque necrosis, and vascular inflammation. miR-21 expression influences foam cell formation, sensitivity to ER-stress-induced apoptosis, and phagocytic clearance capacity. Mechanistically, we discovered that the absence of miR-21 in macrophages increases the expression of the miR-21 target gene, MKK3, promoting the induction of p38-CHOP and JNK signaling. Both pathways enhance macrophage apoptosis and promote the post-translational degradation of ABCG1, a transporter that regulates cholesterol efflux in macrophages. Altogether, these findings reveal a major role for hematopoietic miR-21 in atherogenesis. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

miR-21 promotes fibrogenic epithelial-to-mesenchymal transition of epicardial mesothelial cells involving Programmed Cell Death 4 and Sprouty-1

DEFF Research Database (Denmark)

Brønnum, Hasse; Andersen, Ditte C; Schneider, Mikael

2013-01-01

and miR-21-dependent targeting of Programmed Cell Death 4 (PDCD4) and Sprouty Homolog 1 (SPRY1) significantly contributed to the development of a fibroblastoid phenotype. However, PDCD4- and SPRY1-targeting was not entirely ascribable to all phenotypic effects from miR-21, underscoring the pleiotropic......, especially TGF-β, promoted EMT progression in EMC cultures, which resulted in differential expression of numerous miRNAs, especially the pleiotropic miR-21. Accordingly, ectopic expression of miR-21 substantially promoted the fibroblast-like phenotype arising from fibrogenic EMT, whereas an antagonist...... that targeted miR-21 blocked this effect, as assessed on the E-cadherin/α-smooth muscle actin balance, cell viability, matrix activity, and cell motility, thus making miR-21 a relevant target of EMC-derived fibrosis. Several mRNA targets of miR-21 was differentially regulated during fibrogenic EMT of EMCs...

A miR-21 inhibitor enhances apoptosis and reduces G2-M accumulation induced by ionizing radiation in human glioblastoma U251 cells

International Nuclear Information System (INIS)

Li, Yi; Li, Qiang; Asai, Akio; Kawamoto, Keiji; Zhao Shiguang; Zhen Yunbo; Teng Lei

2011-01-01

MicroRNAs (miRNAs) are small noncoding RNAs that take part in diverse biological processes by suppressing target gene expression. Elevated expression of miR-21 has been reported in many types of human cancers. Radiotherapy is a standard adjuvant treatment for patients with glioblastoma. However, the resistance of glioblastoma cells to radiation limits the success of this treatment. In this study, we found that miR-21 expression was upregulated in response to ionizing radiation (IR) in U251 cells, which suggested that miR-21 could be involved in the response of U251 cells to radiation. We showed that a miR-21 inhibitor enhanced IR-induced glioblastoma cell growth arrest and increased the level of apoptosis, which was probably caused by abrogation of the G 2 -M arrest induced by IR. Further research demonstrated that the miR-21 inhibitor induced the upregulation of Cdc25A. Taken together, these findings suggest that miR-21 inhibitor can increase IR-induced growth arrest and apoptosis in U251 glioblastoma cells, at least in part by abrogating G 2 -M arrest, and that Cdc25A is a potential target of miR-21. (author)

Evaluation of miR-21 and miR-375 as prognostic biomarkers in esophageal cancer

DEFF Research Database (Denmark)

Winther, Mette; Alsner, Jan; Tramm, Trine

2015-01-01

analyses identified miR-21 as an independent prognostic marker for DSS in EAC [HR 3.52 (95% CI 1.06-11.69)]. High miR-375 was not correlated with improved prognosis in either histology. However, Forest plots demonstrated that both miR-21 and miR-375 were of prognostic impact in ESCC. CONCLUSION...... chemotherapy were analyzed. Expression levels of miR-21 and miR-375 were quantified using Affymetrix GeneChip miRNA 1.0 Array. The Cox proportional hazards model was used to assess the correlation of miR-21 and miR-375 with disease-specific survival (DSS) and overall survival (OS). Forest plots were performed...... to evaluate the prognostic impact of miR-21 and miR-375 in the present study and previously published reports. RESULTS: In ESCC, patients with miR-21 expression levels above median showed a trend towards poorer DSS and OS. When dividing miR-21 expression by tertiles, high levels of miR-21 significantly...

MiR-21/PTEN Axis Promotes Skin Wound Healing by Dendritic Cells Enhancement.

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Han, Zhaofeng; Chen, Ya; Zhang, Yile; Wei, Aizhou; Zhou, Jian; Li, Qian; Guo, Lili

2017-10-01

A number of miRNAs associated with wound repair have been identified and characterized, but the mechanism has not been fully clarified. MiR-21 is one of wound-related lncRNAs, and the study aimed to explore the functional involvement of miR-21 and its concrete mechanism in wound healing. In this study, the rat model of skin wounds was established. The expression of miR-21, PTEN and related molecules of wound tissues or cells was determined by quantitative real-time PCR and Western blot, respectively. The regulatory role of miR-21 on PTEN was examined by luciferase reporter gene assay. Flow cytometry assay was applied to measure cell number changes. MiR-21 was upregulated at 6, 24, 48, 72 h after model establishment, and the increase reached a maximum at 24 h in wound tissues. MMP-9 expression presented the same tread as miR-21 and was significantly enhanced within 6 h of wound formation, and then remained to be increased to the maximum at 24 h. The increase of miR-21 was accompanied by the increase of cell total number and DCs ratio in wound fluids. MiR-21 overexpression significantly improved the healing of skin wounds and increased the ratio of DCs in rats. The results of using FL confirmed that miR-21 overexpression obviously promoted DCs differentiation. Additionally, miR-21 could activate AKT/PI3K signaling pathway via inhibition of PTEN. MiR-21 contributes to wound healing via inhibition of PTEN that activated AKT/PI3K signaling pathway to increase DCs. J. Cell. Biochem. 118: 3511-3519, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Intense light-elicited upregulation of miR-21 facilitates glycolysis and cardioprotection through Per2-dependent mechanisms.

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Colleen Marie Bartman

Full Text Available A wide search for ischemic preconditioning (IPC mechanisms of cardioprotection identified the light elicited circadian rhythm protein Period 2 (Per2 to be cardioprotective. Studies on cardiac metabolism found a key role for light elicited Per2 in mediating metabolic dependence on carbohydrate metabolism. To profile Per2 mediated pathways following IPC of the mouse heart, we performed a genome array and identified 352 abundantly expressed and well-characterized Per2 dependent micro RNAs. One prominent result of our in silico analysis for cardiac Per2 dependent micro RNAs revealed a selective role for miR-21 in the regulation of hypoxia and metabolic pathways. Based on this Per2 dependency, we subsequently found a diurnal expression pattern for miR-21 with higher miR-21 expression levels at Zeitgeber time (ZT 15 compared to ZT3. Gain or loss of function studies for miR-21 using miRNA mimics or miRNA inhibitors and a Seahorse Bioanalyzer uncovered a critical role of miR-21 for cellular glycolysis, glycolytic capacity, and glycolytic reserve. Exposing mice to intense light, a strategy to induce Per2, led to a robust induction of cardiac miR-21 tissue levels and decreased infarct sizes, which was abolished in miR-21-/- mice. Similarly, first translational studies in humans using intense blue light exposure for 5 days in healthy volunteers resulted in increased plasma miR-21 levels which was associated with increased phosphofructokinase activity, the rate-limiting enzyme in glycolysis. Together, we identified miR-21 as cardioprotective downstream target of Per2 and suggest intense light therapy as a potential strategy to enhance miR-21 activity and subsequent carbohydrate metabolism in humans.

Clinical relevance of microRNA miR-21, miR-31, miR-92a, miR-101, miR-106a and miR-145 in colorectal cancer

International Nuclear Information System (INIS)

Schee, Kristina; Boye, Kjetil; Abrahamsen, Torveig Weum; Fodstad, Øystein; Flatmark, Kjersti

2012-01-01

MicroRNAs (miRNAs) regulate gene expression by binding to mRNA, and can function as oncogenes or tumor suppressors depending on the target. In this study, using qRT-PCR, we examined the expression of six miRNAs (miR-21, miR-31, miR-92a, miR-101, miR-106a and miR-145) in tumors from 193 prospectively recruited patients with colorectal cancer, and associations with clinicopathological parameters and patient outcome were analyzed. The miRNAs were chosen based on previous studies for their biomarker potential and suggested biological relevance in colorectal cancer. The miRNA expression was examined by qRT-PCR. Associations between miRNA expression and clinicopathological variables were explored using Mann–Whitney U and Kruskal-Wallis test while survival was estimated using the Kaplan-Meier method and compared using the log-rank test. MiR-101 was hardly expressed in the tumor samples, while for the other miRNAs, variable expression levels and expression ranges were observed, with miR-21 being most abundantly expressed relative to the reference (RNU44). In our study cohort, major clinical significance was demonstrated only for miR-31, as high expression was associated with advanced tumor stage and poor differentiation. No significant associations were found between expression of the investigated miRNAs and metastasis-free or overall survival. Investigating the expression of six miRNAs previously identified as candidate biomarkers in colorectal cancer, few clinically relevant associations were detected in our patient cohort. Our results emphasize the importance of validating potential tumor markers in independent patient cohorts, and indicate that the role of miRNAs as colorectal cancer biomarkers is still undetermined

MiR-146b-5p overexpression attenuates stemness and radioresistance of glioma stem cells by targeting HuR/lincRNA-p21/β-catenin pathway

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Yang, Wei; Yu, Hongquan; Shen, Yueming; Liu, Yingying; Yang, Zhanshan; Sun, Ting

2016-01-01

A stem-like subpopulation existed in GBM cells, called glioma stem cells (GSCs), might contribute to cancer invasion, angiogenesis, immune evasion, and therapeutic resistance, providing a rationale to eliminate GSCs population and their supporting niche for successful GBM treatment. LincRNA-p21, a novel regulator of cell proliferation, apoptosis and DNA damage response, is found to be downregulated in several types of tumor. However, little is known about the role of lincRNA-p21 in stemness and radioresistance of GSCs and its regulating mechanisms. In this study, we found that lincRNA-p21 negatively regulated the expression and activity of β-catenin in GSCs. Downregulation of lincRNA-p21 in GSCs was resulted from upregulation of Hu antigen R (HuR) expression caused by miR-146b-5p downregulation. MiR-146b-5p overexpression increased apoptosis and radiosensitivity, decreased cell viability, neurosphere formation capacity and stem cell marker expression, and induced differentiation in GSCs. Moreover, knock-down lincRNA-p21 or HuR and β-catenin overexpression could rescue the phenotypic changes resulted from miR-146b-5p overexpression in GSCs. These findings suggest that targeting the miR-146b-5p/HuR/lincRNA-p21/β-catenin signaling pathway may be valuable therapeutic strategies against glioma. PMID:27166258

MiR-21 is involved in radiation-induced bystander effects

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Xu, Shuai; Ding, Nan; Pei, Hailong; Hu, Wentao; Wei, Wenjun; Zhang, Xurui; Zhou, Guangming; Wang, Jufang

2014-01-01

Radiation-induced bystander effects are well-established phenomena, in which DNA damage responses are induced not only in the directly irradiated cells but also in the non-irradiated bystander cells through intercellular signal transmission. Recent studies hint that bystander effects are possibly mediated via small non-coding RNAs, especially microRNAs. Thus, more details about the roles of microRNA in bystander effects are urgently needed to be elucidated. Here we demonstrated that bystander effects were induced in human fetal lung MRC-5 fibroblasts through medium-mediated way by different types of radiation. We identified a set of differentially expressed microRNAs in the cell culture medium after irradiation, among which the up-regulation of miR-21 was further verified with qRT-PCR. In addition, we found significant upregulation of miR-21 in both directly irradiated cells and bystander cells, which was confirmed by the expression of miR-21 precursor and its target genes. Transfection of miR-21 mimics into non-irradiated MRC-5 cells caused bystander-like effects. Taken together, our data reveals that miR-21 is involved in radiation-induced bystander effects. Elucidation of such a miRNA-mediated bystander effect is of utmost importance in understanding the biological processes related to ionizing radiation and cell-to-cell communication. PMID:25483031

Multiple Myeloma-Derived Exosomes Regulate the Functions of Mesenchymal Stem Cells Partially via Modulating miR-21 and miR-146a

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Qian Cheng

2017-01-01

Full Text Available Exosomes derived from cancer cells can affect various functions of mesenchymal stem cells (MSCs via conveying microRNAs (miRs. miR-21 and miR-146a have been demonstrated to regulate MSC proliferation and transformation. Interleukin-6 (IL-6 secreted from transformed MSCs in turn favors the survival of multiple myeloma (MM cells. However, the effects of MM exosomes on MSC functions remain largely unclear. In this study, we investigated the effects of OPM2 (a MM cell line exosomes (OPM2-exo on regulating the proliferation, cancer-associated fibroblast (CAF transformation, and IL-6 secretion of MSCs and determined the role of miR-21 and miR-146a in these effects. We found that OPM2-exo harbored high levels of miR-21 and miR-146a and that OPM2-exo coculture significantly increased MSC proliferation with upregulation of miR-21 and miR-146a. Moreover, OPM2-exo induced CAF transformation of MSCs, which was evidenced by increased fibroblast-activated protein (FAP, α-smooth muscle actin (α-SMA, and stromal-derived factor 1 (SDF-1 expressions and IL-6 secretion. Inhibition of miR-21 or miR-146a reduced these effects of OPM2-exo on MSCs. In conclusion, MM could promote the proliferation, CAF transformation, and IL-6 secretion of MSCs partially through regulating miR21 and miR146a.

STAT5 induces miR-21 expression in cutaneous T cell lymphoma

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Lindahl, Lise M; Fredholm, Simon; Joseph, Claudine

2016-01-01

was inhibited by Tofacitinib (CP-690550), a clinical-grade JAK3 inhibitor. Chromatin immunoprecipitation (ChIP) analysis showed direct binding of STAT5 to the miR-21 promoter. Cytokine starvation ex vivo triggered a decrease in miR-21 expression, whereas IL-2 induced an increased miR-21 expression in primary SS...

MiR-144 Increases Intestinal Permeability in IBS-D Rats by Targeting OCLN and ZO1

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Qiuke Hou

2017-12-01

Full Text Available Background/Aims: Irritable bowel syndrome with diarrhoea (IBS-D is a chronic, functional bowel disorder characterized by abdominal pain or diarrhoea and altered bowel habits, which correlate with intestinal hyperpermeability. MicroRNAs (miRNAs are involved in regulating intestinal permeability in IBS-D. However, the role of miRNAs in regulating intestinal permeability and protecting the epithelial barrier remains unclear. Our goals were to (i identify differential expression of miRNAs and their targets in the distal colon of IBS-D rats; (ii verify in vitro whether occludin (OCLN and zonula occludens 1 (ZO1/TJP1 were direct targets of miR-144 and were down-regulated in IBS-D rats; and (iii determine whether down-regulation of miR-144 in vitro could reverse the pathological hallmarks of intestinal hyperpermeability via targeting OCLN and ZO1. Methods: The IBS-D rat model was established using 4% acetic acid and evaluated by haematoxylin-eosin (HE staining. The distal colon was obtained in order to perform miRNA microarray analysis and to isolate and culture colonic epithelial cells. When differential expression of miRNA was found, the results were verified by qRT-PCR, and the target genes were further explored by bioinformatics analysis. Correlation analyses were carried out to compare the expression of miRNA and target genes. Then, mutants, miRNA mimics and inhibitors of the target genes were constructed and transfected to colonic epithelial cells. qRT-PCR, western blotting, enzyme-linked immunosorbent assays (ELISAs and dual-luciferase assays were used to investigate the expression of miR-144 and OCLN, ZO1 in IBS-D rats. Results: There were 8 up-regulated and 18 down-regulated miRNAs identified in the IBS-D rat model. Of these, miR-144 was markedly up-regulated and resulted in the down-regulation of OCLN and ZO1 expression. Overexpression of miR-144 by transfection of miR-144 precursor markedly inhibited the expression of OCLN and ZO1. Further

Resveratrol Inhibits ROS-Promoted Activation and Glycolysis of Pancreatic Stellate Cells via Suppression of miR-21

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Bin Yan

2018-01-01

Full Text Available Activation of pancreatic stellate cells (PSCs initiates pancreatic fibrosis in chronic pancreatitis and furnishes a niche that enhances the malignancy of pancreatic cancer cells (PCCs in pancreatic ductal adenocarcinoma (PDAC. Resveratrol (RSV, a natural polyphenol, exhibits potent antioxidant and anticancer effects. However, whether and how RSV influences the biological properties of activated PSCs and the effects of these changes on tumor remain unknown. In the present study, we found that RSV impeded hydrogen peroxide-driven reactive oxygen species- (ROS- induced activation, invasion, migration, and glycolysis of PSCs. In addition, miR-21 expression in activated PSCs was downregulated after RSV treatment, whereas the PTEN protein level increased. miR-21 silencing attenuated ROS-induced activation, invasion, migration, and glycolysis of PSCs, whereas the overexpression of miR-21 rescued the responses of PSCs treated with RSV. Moreover, RSV or N-acetyl-L-cysteine (NAC administration or miR-21 knockdown in PSCs reduced the invasion and migration of PCCs in coculture, and the effects of RSV were partly reversed by miR-21 upregulation. Collectively, RSV inhibits PCC invasion and migration through suppression of ROS/miR-21-mediated activation and glycolysis in PSCs. Therefore, targeting miR-21-mediated glycolysis by RSV in tumor stroma may serve as a new strategy for clinical PDAC prevention or treatment.

Altering β-cell number through stable alteration of miR-21 and miR-34a expression

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Backe, Marie Balslev; Novotny, Guy Wayne; Christensen, Dan Ploug

2014-01-01

RNAs, miR-21 and miR-34a, may be involved in mediating cytokine-induced β-cell dysfunction. Therefore, manipulation of miR-21 and miR-34a levels may potentially be beneficial to β cells. To study the effect of long-term alterations of miR-21 or miR-34a levels upon net β-cell number, we stably overexpressed...

Folate status, folate-related genes and serum miR-21 expression: Implications for miR-21 as a biomarker

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Emma Louise Beckett

2015-12-01

General significance: This study demonstrates that serum miR-21 expression correlates with folate status and related genetic status. This may have consequences for the proposed use of miR-21 as a colorectal cancer biomarker.

Upregulation of miR21 and repression of Grhl3 by leptin mediates sinusoidal endothelial injury in experimental nonalcoholic steatohepatitis.

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Sahar Pourhoseini

Full Text Available Sinusoidal endothelial dysfunction (SED has been found to be an early event in nonalcoholic steatohepatitis (NASH progression but the molecular mechanisms underlying its causation remains elusive. We hypothesized that adipokine leptin worsens sinusoidal injury by decreasing functionally active nitric oxide synthase 3 (NOS3 via miR21. Using rodent models of NASH, and transgenic mice lacking leptin and leptin receptor, results showed that hyperleptinemia caused a 4-5 fold upregulation of hepatic miR21 as assessed by qRTPCR. The upregulation of miR21 led to a time-dependent repression of its target protein Grhl3 levels as shown by western blot analyses. NOS3-p/NOS3 ratio which is controlled by Grhl3 was significantly decreased in NASH models. SED markers ICAM-1, VEGFR-2, and E-selectin as assessed by immunofluorescence microscopy were significantly up regulated in the progressive phases of NASH. Lack of leptin or its receptor in vivo, reversed the upregulation of miR21 and restored the levels of Grhl3 and NOS3-p/NOS3 ratio coupled with decreased SED dysfunction markers. Interestingly, leptin supplementation in mice lacking leptin, significantly enhanced miR21 levels, decreased Grhl3 repression and NOS3 phosphorylation. Leptin supplementation in isolated primary endothelial cells, Kupffer cells and stellate cells showed increased mir21 expression in stellate cells while sinusoidal injury was significantly higher in all cell types. Finally miR21 KO mice showed increased NOS3-p/NOS3 ratio and reversed SED markers in the rodent models of NASH. The experimental results described here show a close association of leptin-induced miR21 in aiding sinusoidal injury in NASH.

High expression of miR-21 in tumor stroma correlates with increased cancer cell proliferation in human breast cancer

DEFF Research Database (Denmark)

Rask, Lene; Balslev, Eva; Jørgensen, Stine

2011-01-01

features, we performed in situ hybridization and semi-quantitative assessment of the miR-21 signal on 12 LN negative grade I (assumed low risk), and 12 LN positive grade II (high risk) breast cancers. miR-21 was predominantly seen in cancer associated fibroblast-like cells, with no difference in expression......Low-risk and high-risk breast cancer patients are stratified primarily according to their lymph node (LN) status and grading. However, some low-risk patients relapse, and some high-risk patients have a favorable clinical outcome, implying a need for better prognostic and predictive tests. Micro...... RNAs are often aberrantly expressed in cancer and microRNA-21 is upregulated in a variety of cancers, including breast cancer. High miR-21 levels have been associated with poor prognosis. To determine the cellular localization of miR-21 and to compare its expression levels with histopathological...

Myostatin Promotes Interleukin-1β Expression in Rheumatoid Arthritis Synovial Fibroblasts through Inhibition of miR-21-5p

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Sung-Lin Hu

2017-12-01

Full Text Available Rheumatoid arthritis (RA is characterized by the infiltration of a number of pro-inflammatory cytokines into synovial fluid and patients with RA often develop joint destruction and deficits in muscle mass. The growth factor myostatin is a key regulator linking muscle mass and bone structure. We sought to determine whether myostatin regulates rheumatoid synovial fibroblast activity and inflammation in RA. We found that levels of myostatin and interleukin (IL-1β (a key pro-inflammatory cytokine in RA in synovial fluid from RA patients were overexpressed and positively correlated. In in vitro investigations, we found that myostatin dose-dependently regulated IL-1β expression through the ERK, JNK, and AP-1 signal-transduction pathways. Computational analysis confirmed that miR-21-5p directly targets the expression of the 3′ untranslated region (3′ UTR of IL-1β. Treatment of cells with myostatin inhibited miR-21-5p expression and miR-21-5p mimic prevented myostatin-induced enhancement of IL-1β expression, showing an inverse correlation between miR-21-5p and IL-1β expression during myostatin treatment. We also found significantly increased paw swelling in an animal model of collagen-induced arthritis (CIA, compared with controls; immunohistochemistry staining revealed substantially higher levels of myostatin and IL-1β expression in CIA tissue. Our evidence indicates that myostatin regulates IL-1β production. Thus, targeting myostatin may represent a potential therapeutic target for RA.

Effect of microRNA-21 on the proliferation of human degenerated nucleus pulposus by targeting programmed cell death 4

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B. Chen

2016-01-01

Full Text Available This study aims to explore the effect of microRNA-21 (miR-21 on the proliferation of human degenerated nucleus pulposus (NP by targeting programmed cell death 4 (PDCD4 tumor suppressor. NP tissues were collected from 20 intervertebral disc degeneration (IDD patients, and from 5 patients with traumatic spine fracture. MiR-21 expressions were tested. NP cells from IDD patients were collected and divided into blank control group, negative control group (transfected with miR-21 negative sequences, miR-21 inhibitor group (transfected with miR-21 inhibitors, miR-21 mimics group (transfected with miR-21 mimics and PDCD4 siRNA group (transfected with PDCD4 siRNAs. Cell growth was estimated by Cell Counting Kit-8; PDCD4, MMP-2,MMP-9 mRNA expressions were evaluated by qRT-PCR; PDCD4, c-Jun and p-c-Jun expressions were tested using western blot. In IDD patients, the expressions of miR-21 and PDCD4 mRNA were respectively elevated and decreased (both P<0.05. The miR-21 expressions were positively correlated with Pfirrmann grades, but negatively correlated with PDCD4 mRNA (both P<0.001. In miR-21 inhibitor group, cell growth, MMP-2 and MMP-9 mRNA expressions, and p-c-Jun protein expressions were significantly lower, while PDCD4 mRNA and protein expressions were higher than the other groups (all P<0.05. These expressions in the PDCD4 siRNA and miR-21 mimics groups was inverted compared to that in the miR-21 inhibitor group (all P<0.05. MiR-21 could promote the proliferation of human degenerated NP cells by targeting PDCD4, increasing phosphorylation of c-Jun protein, and activating AP-1-dependent transcription of MMPs, indicating that miR-21 may be a crucial biomarker in the pathogenesis of IDD.

Adrenaline inhibits osteogenesis via repressing miR-21 expression.

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Chen, Danying; Wang, Zuolin

2017-01-01

Sympathetic signaling is involved in bone homeostasis; however, the cellular and molecular mechanisms remain unknown. In this study, we found that the psychological stress mediator adrenaline inhibited osteogenic differentiation of human bone marrow-derived stem cells (hMSC) by reducing microRNA-21 (miR-21) expression. Briefly, adrenaline significantly inhibited the osteogenic differentiation of hMSCs, as observed with both Alizarin red staining and maker gene expression (RUNX2, OSX, OCN, and OPN). During this process, miR-21 was suppressed by adrenaline via inhibition of histone acetylation, as verified by H3K9Ac chromatin immunoprecipitation (ChIP) assay. MiR-21 was confirmed to promote hMSC osteogenic differentiation, and overexpression of miR-21 reversed the impeditive effect of adrenaline on hMSC osteogenic differentiation. Our results demonstrate that down-regulation of miR-21 is responsible for the adrenaline-mediated inhibition of hMSC osteogenic differentiation. These findings indicate a regulation of bone metabolism by psychological stress and also provide a molecular basis for psychological stress-associated bone diseases. © 2016 International Federation for Cell Biology.

Lentiviral CRISPR/Cas9 vector mediated miR-21 gene editing inhibits the epithelial to mesenchymal transition in ovarian cancer cells.

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Huo, Wenying; Zhao, Guannan; Yin, Jinggang; Ouyang, Xuan; Wang, Yinan; Yang, Chuanhe; Wang, Baojing; Dong, Peixin; Wang, Zhixiang; Watari, Hidemichi; Chaum, Edward; Pfeffer, Lawrence M; Yue, Junming

2017-01-01

CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats) mediated genome editing is a powerful approach for loss of function studies. Here we report that lentiviral CRISPR/Cas9 vectors are highly efficient in introducing mutations in the precursor miRNA sequence, thus leading to the loss of miRNA expression and function. We constructed four different lentiviral CRISPR/Cas9 vectors that target different regions of the precursor miR-21 sequence and found that these lentiviral CRISPR/Cas9 miR-21 gRNA vectors induced mutations in the precursor sequences as shown by DNA surveyor mutation assay and Sanger sequencing. Two miR-21 lentiviral CRISPR/Cas9 gRNA vectors were selected to probe miR-21 function in ovarian cancer SKOV3 and OVCAR3 cell lines. Our data demonstrate that disruption of pre-miR-21 sequences leads to reduced cell proliferation, migration and invasion. Moreover, CRISPR/Cas9-mediated miR-21 gene editing sensitizes both SKOV3 and OVCAR3 cells to chemotherapeutic drug treatment. Disruption of miR-21 leads to the inhibition of epithelial to mesenchymal transition (EMT) in both SKOV3 and OVCAR3 cells as evidenced by the upregulation of epithelial cell marker E-cadherin and downregulation of mesenchymal marker genes, vimentin and Snai2. The miR-21 target genes PDCD4 and SPRY2 were upregulated in cells transduced with miR-21gRNAs compared to controls. Our study indicates that lentiviral CRISPR/Cas9-mediated miRNA gene editing is an effective approach to address miRNA function, and disruption of miR-21 inhibits EMT in ovarian cancer cells.

MiR-21 is induced in endothelial cells by shear stress and modulates apoptosis and eNOS activity

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Weber, Martina; Baker, Meredith B.; Moore, Jeffrey P. [Division of Cardiology, Emory University, 1639 Pierce Drive, WMB 319, Atlanta, GA 30322 (United States); Searles, Charles D., E-mail: csearle@emory.edu [Division of Cardiology, Emory University, 1639 Pierce Drive, WMB 319, Atlanta, GA 30322 (United States); Atlanta Veterans Administration Medical Center, 1670 Clarimont Road, Decatur, GA 30033 (United States)

2010-03-19

Mechanical forces associated with blood flow play an important role in regulating vascular signaling and gene expression in endothelial cells (ECs). MicroRNAs (miRNAs) are a class of noncoding RNAs that posttranscriptionally regulate the expression of genes involved in diverse cell functions, including differentiation, growth, proliferation, and apoptosis. miRNAs are known to have an important role in modulating EC biology, but their expression and functions in cells subjected to shear stress conditions are unknown. We sought to determine the miRNA expression profile in human ECs subjected to unidirectional shear stress and define the role of miR-21 in shear stress-induced changes in EC function. TLDA array and qRT-PCR analysis performed on HUVECs exposed to prolonged unidirectional shear stress (USS, 24 h, 15 dynes/cm{sup 2}) identified 13 miRNAs whose expression was significantly upregulated (p < 0.05). The miRNA with the greatest change was miR-21; it was increased 5.2-fold (p = 0.002) in USS-treated versus control cells. Western analysis demonstrated that PTEN, a known target of miR-21, was downregulated in HUVECs exposed to USS or transfected with pre-miR-21. Importantly, HUVECs overexpressing miR-21 had decreased apoptosis and increased eNOS phosphorylation and nitric oxide (NO{sup {center_dot}}) production. These data demonstrate that shear stress forces regulate the expression of miRNAs in ECs, and that miR-21 influences endothelial biology by decreasing apoptosis and activating the NO{sup {center_dot}} pathway. These studies advance our understanding of the mechanisms by which shear stress forces modulate vascular homeostasis.

Diagnostic and prognostic potential of serum miR-7, miR-16, miR-25, miR-93, miR-182, miR-376a and miR-429 in ovarian cancer patients.

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Meng, Xiaodan; Joosse, Simon A; Müller, Volkmar; Trillsch, Fabian; Milde-Langosch, Karin; Mahner, Sven; Geffken, Maria; Pantel, Klaus; Schwarzenbach, Heidi

2015-11-03

Owing to late diagnosis in advanced disease stages, prognosis of patients with epithelial ovarian cancer (EOC) is poor. The quantification of deregulated levels of microRNAs could facilitate earlier diagnosis and improve prognosis of EOC. Seven microRNAs (miR-7, miR-16, miR-25, miR-93, miR-182, miR-376a and miR-429) were quantified in the serum of 180 EOC patients and 66 healthy women by TaqMan PCR microRNA assays. Median follow-up time was 21 months. The effects of miR-7 and miR-429 on apoptosis, cell proliferation, migration and invasion were investigated in two (EOC) cell lines. Serum levels of miR-25 (P=0.0001) and miR-93 (P=0.0001) were downregulated, whereas those of miR-7 (P=0.001) and miR-429 (P=0.0001) were upregulated in EOC patients compared with healthy women. The four microRNAs discriminated EOC patients from healthy women with a sensitivity of 93% and a specificity of 92%. The levels of miR-429 positively correlated with CA125 values (P=0.0001) and differed between FIGO I-II and III-IV stages (P=0.001). MiR-429 was an independent predictor of overall survival (P=0.011). Overexpressed miR-429 in SKOV3 cells led to suppression of cell migration (P=0.037) and invasion (P=0.011). Increased levels of miR-7 were associated with lymph node metastases (P=0.0001) and FIGO stages III-IV (P=0.0001). Overexpressed miR-7 in SKOV3 cells resulted in increased cell migration (P=0.001) and invasion (P=0.011). Additionally, the increased levels of miR-376a correlated with FIGO stages III-IV (P=0.02). Our data indicate the diagnostic potential of miR-7, miR-25, miR-93 and miR-429 in EOC and the prognostic potential of miR-429. This microRNA panel may be promising molecules to be targeted in the treatment of EOC.

LincRNA-p21 inhibits invasion and metastasis of hepatocellular carcinoma through miR-9/E-cadherin cascade signaling pathway molecular mechanism

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Ding G

2017-06-01

Full Text Available Gangqiang Ding, Zhen Peng, Jia Shang, Yi Kang, Huibin Ning, Chongshan Mao Department of Infectious Diseases, People’s Hospital of Zhengzhou University, Henan Provincial People’s Hospital, Zhengzhou, China Abstract: In the previous study, it was found that long intergenic noncoding RNA-p21 (lincRNA-p21 was downregulated in hepatocellular carcinoma (HCC and lincRNA-p21 overexpression inhibited tumor invasion through inducing epithelial–mesenchymal transition. However, the underlying mechanism was not fully elaborated. In this study, lincRNA-p21 expression was measured in 12 paired HCC and nontumor adjacent normal tissues by ­quantitative real-time polymerase chain reaction. The effects of lincRNA-p21 on HCC cells were studied using lentivirus expressing lincRNA-p21 vector in vitro. The association between lincRNA-p21 level and miR-9 level was tested with the Spearman rank correlation. The effects of miR-9 on HCC cells were studied by using miR-9 inhibitor in vitro. Luciferase assay was used to validate the target of miR-9. The results showed that lincRNA-p21 was downregulated in human HCC tissues and cell lines. LincRNA-p21 overexpression significantly inhibited HCC cell migration and invasion in vitro. Besides, lincRNA-p21 negatively regulated miR-9 expression level, and miR-9 was upregulated in human HCC tissues and cells. MiR-9 knockdown inhibited HCC cell migration and invasion in vitro. Finally, the luciferase assay results showed that E-cadherin was a direct target of miR-9. The expression level of E-cadherin was found to be regulated by lincRNA-p21 and miR-9. Altogether, the results suggested that lincRNA-p21 inhibits migration and invasion of HCC cells through regulating miR-9-mediated E-cadherin cascade signaling pathway. Keywords: hepatocellular carcinoma, lincRNA-p21, miR-9, E-cadherin, epithelial–mesenchymal transition

MicroRNA-21 directly targets MARCKS and promotes apoptosis resistance and invasion in prostate cancer cells

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Li, Tao; Li, Dong; Sha, Jianjun; Sun, Peng; Huang, Yiran

2009-01-01

Prostate cancer is one of the most common malignant cancers in men. Recent studies have shown that microRNA-21 (miR-21) is overexpressed in various types of cancers including prostate cancer. Studies on glioma, colon cancer cells, hepatocellular cancer cells and breast cancer cells have indicated that miR-21 is involved in tumor growth, invasion and metastasis. However, the roles of miR-21 in prostate cancer are poorly understood. In this study, the effects of miR-21 on prostate cancer cell proliferation, apoptosis, and invasion were examined. In addition, the targets of miR-21 were identified by a reported RISC-coimmunoprecipitation-based biochemical method. Inactivation of miR-21 by antisense oligonucleotides in androgen-independent prostate cancer cell lines DU145 and PC-3 resulted in sensitivity to apoptosis and inhibition of cell motility and invasion, whereas cell proliferation were not affected. We identified myristoylated alanine-rich protein kinase c substrate (MARCKS), which plays key roles in cell motility, as a new target in prostate cancer cells. Our data suggested that miR-21 could promote apoptosis resistance, motility, and invasion in prostate cancer cells and these effects of miR-21 may be partly due to its regulation of PDCD4, TPM1, and MARCKS. Gene therapy using miR-21 inhibition strategy may therefore be useful as a prostate cancer therapy.

miR-543 promotes gastric cancer cell proliferation by targeting SIRT1

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Li, Juan; Dong, Guoying; Wang, Bo; Gao, Wei; Yang, Qing

2016-01-01

SIRT1, a class III histone deacetylase, exerts inhibitory effects on tumorigenesis and is downregulated in gastric cancer. However, the role of microRNAs in the regulation of SIRT1 in gastric cancer is still largely unknown. Here, we identified miR-543 as a predicted upstream regulator of SIRT1 using 3 different bioinformatics databases. Mimics of miR-543 significantly inhibited the expression of SIRT1, whereas an inhibitor of miR-543 increased SIRT1 expression. MiR-543 directly targeted the 3′-UTR of SIRT1, and both of the two binding sites contributed to the inhibitory effects. In gastric epithelium-derived cell lines, miR-543 promoted cell proliferation and cell cycle progression, and overexpression of SIRT1 rescued the above effects of miR-543. The inhibitory effects of miR-543 on SIRT1 were also validated using clinical gastric cancer samples. Moreover, we found that miR-543 expression was positively associated with tumor size, clinical grade, TNM stage and lymph node metastasis in gastric cancer patients. Our results identify a new regulatory mechanism of miR-543 on SIRT1 expression in gastric cancer, and raise the possibility that the miR-543/SIRT1 pathway may serve as a potential target for the treatment of gastric cancer. - Highlights: • SIRT1 is a novel target of miR-543. • miR-543 promotes gastric cancer cell proliferation and cell cycle progression by targeting SIRT1. • miR-543 is upregulated in GC and positively associated with tumor size, clinical grade, TNM stage and lymph node metastasis. • miR-543 is negatively correlated with SIRT1 expression in gastric cancer tissues.

Hypomethylation of miR-142 promoter and upregulation of microRNAs that target the oxytocin receptor gene in the autism prefrontal cortex.

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Mor, Michal; Nardone, Stefano; Sams, Dev Sharan; Elliott, Evan

2015-01-01

MicroRNAs are small RNA molecules that regulate the translation of protein from gene transcripts and are a powerful mechanism to regulate gene networks. Next-generation sequencing technologies have produced important insights into gene transcription changes that occur in the brain of individuals diagnosed with autism spectrum disorder (asd). However, these technologies have not yet been employed to uncover changes in microRNAs in the brain of individuals diagnosed with asd. Small RNA next-generation sequencing was performed on RNA extracted from 12 human autism brain samples and 12 controls. Real-time PCR was used to validate a sample of the differentially expressed microRNAs, and bioinformatic analysis determined common pathways of gene targets. MicroRNA expression data was correlated to genome-wide DNA methylation data to determine if there is epigenetic regulation of dysregulated microRNAs in the autism brain. Luciferase assays, real-time PCR, and Western blot analysis were used to determine how dysregulated microRNAs may regulate the expression and translation of an autism-related gene transcript. We determined that miR-142-5p, miR-142-3p, miR-451a, miR-144-3p, and miR-21-5p are overexpressed in the asd brain. Furthermore, the promoter region of the miR-142 gene is hypomethylated in the same brain samples, suggesting that epigenetics plays a role in dysregulation of microRNAs in the brain. Bioinformatic analysis revealed that these microRNAs target genes that are involved in synaptic function. Further bioinformatic analysis, coupled with in vitro luciferase assays, determined that miR-451a and miR-21-5p can target the oxytocin receptor (OXTR) gene. OXTR gene expression is increased in these same brain samples, and there is a positive correlation between miR-21-5p and OXTR expression. However, miR-21-5p expression negatively correlates to production of OXTR protein from the OXTR transcript. Therefore, we suggest that miR-21-5p may attenuate OXTR expression in

Regulation of miR-21 expression in human melanoma via UV-ray-induced melanin pigmentation.

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Lin, Kuan-Yu; Chen, Chien-Min; Lu, Cheng-You; Cheng, Chun-Yuan; Wu, Yu-Hsin

2017-08-01

Excessive environmental ultraviolet (UV) radiation produces genetic mutations that can lead to skin cancer. This study was designed to assess the potential inhibitory activity of microRNA-21 (miR-21) on the UV irradiation-stimulated melanogenesis signal pathway in melanoma cells. The molecular mechanism of miR-21-induced inhibitory activity on UV-ray-stimulated melanogenesis-regulating proteins was examined in A375.S2 human melanoma and B16F10 mouse melanoma cells. UV irradiation for 30 min induced melanogenesis signal pathway by increasing melanin production and the number of A375.S2 cells. Similarly, UV radiation increased the expression of α-melanocyte-stimulating hormone (α-MSH) protein and decreased the melanogenesis-regulating signal, such as EGFR and Akt phosphorylation. Notably, miR-21 overexpression in UV-ray-stimulated A375.S2 cells decreased α-MSH expression and increased EGFR and Akt phosphorylation levels. Furthermore, miR-21 on UV-ray- induced melanogenesis was down-regulated by the Akt inhibitor and the EGFR inhibitor (Gefitinib). Results suggest that the suppressive activity of miR-21 on UV-ray-stimulated melanogenesis may involve the down-regulation of α-MSH and the activation in both of EGFR and Akt. © 2017 Wiley Periodicals, Inc.

miR-21 modulates tumor outgrowth induced by human adipose tissue-derived mesenchymal stem cells in vivo

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Shin, Keun Koo; Lee, Ae Lim; Kim, Jee Young [Department of Physiology, School of Medicine, Pusan National University, Yangsan, Gyeongnam 626-870 (Korea, Republic of); Medical Research Center for Ischemic Tissue Engineering, Pusan National University, Yangsan, Gyeongnam 626-870 (Korea, Republic of); BK21 Medical Science Education Center, School of Medicine, Pusan National University, Yangsan, Gyeongnam 626-870 (Korea, Republic of); Lee, Sun Young [Department of Physiology, School of Medicine, Pusan National University, Yangsan, Gyeongnam 626-870 (Korea, Republic of); Medical Research Center for Ischemic Tissue Engineering, Pusan National University, Yangsan, Gyeongnam 626-870 (Korea, Republic of); Bae, Yong Chan [Department of Plastic Surgery, School of Medicine, Pusan National University, Pusan 602-739 (Korea, Republic of); Jung, Jin Sup, E-mail: jsjung@pusan.ac.kr [Department of Physiology, School of Medicine, Pusan National University, Yangsan, Gyeongnam 626-870 (Korea, Republic of); Medical Research Center for Ischemic Tissue Engineering, Pusan National University, Yangsan, Gyeongnam 626-870 (Korea, Republic of); BK21 Medical Science Education Center, School of Medicine, Pusan National University, Yangsan, Gyeongnam 626-870 (Korea, Republic of); Medical Research Institute, Pusan National University, Pusan 602-739 (Korea, Republic of)

2012-06-15

Highlights: Black-Right-Pointing-Pointer miR-21 modulates hADSC-induced increase of tumor growth. Black-Right-Pointing-Pointer The action is mostly mediated by the modulation of TGF-{beta} signaling. Black-Right-Pointing-Pointer Inhibition of miR-21 enhances the blood flow recovery in hindlimb ischemia. -- Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in clinical situations, due principally to their potential use in regenerative medicine and tissue engineering applications. However, the therapeutic application of MSCs remains limited, unless the favorable effects of MSCs on tumor growth in vivo, and the long-term safety of the clinical applications of MSCs, can be more thoroughly understood. In this study, we determined whether microRNAs can modulate MSC-induced tumor outgrowth in BALB/c nude mice. Overexpression of miR-21 in human adipose-derived stem cells (hADSCs) inhibited hADSC-induced tumor growth, and inhibition of miR-21 increased it. Downregulation of transforming growth factor beta receptor II (TGFBR2), but not of signal transducer and activator of transcription 3, in hADSCs showed effects similar to those of miR-21 overexpression. Downregulation of TGFBR2 and overexpression of miR21 decreased tumor vascularity. Inhibition of miR-21 and the addition of TGF-{beta} increased the levels of vascular endothelial growth factor and interleukin-6 in hADSCs. Transplantation of miR-21 inhibitor-transfected hADSCs increased blood flow recovery in a hind limb ischemia model of nude mice, compared with transplantation of control oligo-transfected cells. These findings indicate that MSCs might favor tumor growth in vivo. Thus, it is necessary to study the long-term safety of this technique before MSCs can be used as therapeutic tools in regenerative medicine and tissue engineering.

Role of Kallistatin Treatment in Aging and Cancer by Modulating miR-34a and miR-21 Expression

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Julie Chao

2017-01-01

Full Text Available Kallistatin is an endogenous protein that regulates differential signaling pathways and a wide spectrum of biological activities via its two structural elements: an active site and a heparin-binding domain. Kallistatin via its heparin-binding site inhibits vascular inflammation and oxidative stress by antagonizing TNF-α-induced NADPH oxidase activity, NF-κB activation, and inflammatory gene expression in endothelial cells. Moreover, kallistatin via its active site inhibits microRNA-34a (miR-34a synthesis and stimulates eNOS and SIRT1 expression in endothelial progenitor cells, whereas its heparin-binding site is crucial for blocking TNF-α-induced miR-21 expression and oxidative stress, thus reducing cellular senescence. By downregulating miR-34a and miR-21 expression, kallistatin treatment attenuates oxidative damage and aortic senescence in streptozotocin-induced diabetic mice and extends Caenorhabditis elegans lifespan under stress conditions. Likewise, kallistatin through the heparin-binding site inhibits TGF-β-induced miR-21 synthesis and oxidative stress in endothelial cells, resulting in inhibition of endothelial-mesenchymal transition, a process contributing to fibrosis and cancer. Furthermore, kallistatin’s active site is essential for stimulating miR-34a and p53 expression and inhibiting the miR-21-Akt-Bcl-2 signaling pathway, thus inducing apoptosis in breast cancer cells. These findings reveal novel mechanisms of kallistatin in protection against senescence, aging, and cancer development by modulating miR-34a and miR-21 levels and inhibiting oxidative stress.

miR-11 regulates pupal size of Drosophila melanogaster via directly targeting Ras85D.

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Li, Yao; Li, Shengjie; Jin, Ping; Chen, Liming; Ma, Fei

2017-01-01

MicroRNAs play diverse roles in various physiological processes during Drosophila development. In the present study, we reported that miR-11 regulates pupal size during Drosophila metamorphosis via targeting Ras85D with the following evidences: pupal size was increased in the miR-11 deletion mutant; restoration of miR-11 in the miR-11 deletion mutant rescued the increased pupal size phenotype observed in the miR-11 deletion mutant; ectopic expression of miR-11 in brain insulin-producing cells (IPCs) and whole body shows consistent alteration of pupal size; Dilps and Ras85D expressions were negatively regulated by miR-11 in vivo; miR-11 targets Ras85D through directly binding to Ras85D 3'-untranslated region in vitro; removal of one copy of Ras85D in the miR-11 deletion mutant rescued the increased pupal size phenotype observed in the miR-11 deletion mutant. Thus, our current work provides a novel mechanism of pupal size determination by microRNAs during Drosophila melanogaster metamorphosis. Copyright © 2017 the American Physiological Society.

Uncovering Direct Targets of MiR-19a Involved in Lung Cancer Progression.

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Kumiko Yamamoto

Full Text Available Micro RNAs (miRNAs regulate the expression of target genes posttranscriptionally by pairing incompletely with mRNA in a sequence-specific manner. About 30% of human genes are regulated by miRNAs, and a single miRNA is capable of reducing the production of hundreds of proteins by means of incomplete pairing upon miRNA-mRNA binding. Lately, evidence implicating miRNAs in the development of lung cancers has been emerging. In particular, miR-19a, which is highly expressed in malignant lung cancer cells, is considered the key miRNA for tumorigenesis. However, its direct targets remain underreported. In the present study, we focused on six potential miR-19a target genes selected by miRNA target prediction software. To evaluate these genes as direct miR-19a target genes, we performed luciferase, pull-down, and western blot assays. The luciferase activity of plasmids with each miR-19a-binding site was observed to decrease, while increased luciferase activity was observed in the presence of anti-miR-19a locked nucleic acid (LNA. The pull-down assay showed biotinylated miR-19a to bind to AGO2 protein and to four of six potential target mRNAs. Western blot analysis showed that the expression levels of the four genes changed depending on treatment with miR-19a mimic or anti-miR-19a-LNA. Finally, FOXP1, TP53INP1, TNFAIP3, and TUSC2 were identified as miR-19a targets. To examine the function of these four target genes in lung cancer cells, LK79 (which has high miR-19a expression and A549 (which has low miR-19a expression were used. The expression of the four target proteins was higher in A549 than in LK79 cells. The four miR-19a target cDNA expression vectors suppressed cell viability, colony formation, migration, and invasion of A549 and LK79 cells, but LK79 cells transfected with FOXP1 and TP53INP1 cDNAs showed no difference compared to the control cells in the invasion assay.

MiR-223 suppresses cell proliferation by targeting IGF-1R.

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Cheng You Jia

Full Text Available To study the roles of microRNA-223 (miR-223 in regulation of cell growth, we established a miR-223 over-expression model in HeLa cells infected with miR-223 by Lentivirus pLL3.7 system. We observed in this model that miR-223 significantly suppressed the proliferation, growth rate, colony formation of HeLa cells in vitro, and in vivo tumorigenicity or tumor formation in nude mice. To investigate the mechanisms involved, we scanned and examined the potential and putative target molecules of miR-223 by informatics, quantitative PCR and Western blot, and found that insulin-like growth factor-1 receptor (IGF-1R was the functional target of miR-223 inhibition of cell proliferation. Targeting IGF-1R by miR-223 was not only seen in HeLa cells, but also in leukemia and hepatoma cells. The downstream pathway, Akt/mTOR/p70S6K, to which the signal was mediated by IGF-1R, was inhibited as well. The relative luciferase activity of the reporter containing wild-type 3'UTR(3'untranslated region of IGF-1R was significantly suppressed, but the mutant not. Silence of IGF-1R expression by vector-based short hairpin RNA resulted in the similar inhibition with miR-223. Contrarily, rescued IGF-1R expression in the cells that over-expressed miR-223, reversed the inhibition caused by miR-223 via introducing IGF-1R cDNA that didn't contain the 3'UTR. Meanwhile, we also noted that miR-223 targeted Rasa1, but the downstream molecules mediated by Rasa1 was neither targeted nor regulated. Therefore we believed that IGF-1R was the functional target for miR-223 suppression of cell proliferation and its downstream PI3K/Akt/mTOR/p70S6K pathway suppressed by miR-223 was by targeting IGF-1R.

MiR-145 functions as a tumor suppressor targeting NUAK1 in human intrahepatic cholangiocarcinoma

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Xiong, Xinkui; Sun, Daoyi; Chai, Hao; Shan, Wengang [Liver Transplantation Center, First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu Province (China); Key Laboratory of Living Donor Liver Transplantation, Ministry of Public Health, Nanjing, Jiangsu Province (China); Yu, Yue [Key Laboratory of Living Donor Liver Transplantation, Ministry of Public Health, Nanjing, Jiangsu Province (China); Pu, Liyong [Liver Transplantation Center, First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu Province (China); Key Laboratory of Living Donor Liver Transplantation, Ministry of Public Health, Nanjing, Jiangsu Province (China); Cheng, Feng, E-mail: docchengfeng@njmu.edu.cn [Liver Transplantation Center, First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu Province (China); Key Laboratory of Living Donor Liver Transplantation, Ministry of Public Health, Nanjing, Jiangsu Province (China)

2015-09-18

The dysregulation of micro (mi)RNAs is associated with cancer development. The miRNA miR-145 is downregulated in intrahepatic cholangiocarcinoma (ICC); however, its precise role in tumor progression has not yet been elucidated. Novel (nua) kinase family (NUAK)1 functions as an oncogene in various cancers and is a putative target of miR-145 regulation. In this study, we investigated the regulation of NUAK1 by miR-145 in ICC. We found that miR-145 level was significantly decreased in ICC tissue and cell lines, which corresponded with an increase in NUAK1 expression. NUAK1 was found to be a direct target of miR-145 regulation. The overexpression of miR-145 in ICC cell lines inhibited proliferation, growth, and invasion by suppressing NUAK1 expression, which was associated with a decrease in Akt signaling and matrix metalloproteinase protein expression. Similar results were observed by inhibiting NUAK1 expression. These results demonstrate that miR-145 can prevent ICC progression by targeting NUAK1 and its downstream effectors, and can therefore be useful for clinical diagnosis and targeted therapy of ICC. - Highlights: • MiR-145 suppresses ICC proliferation and invasion abilities. • We demonstrated that miR-145 directly targets NUAK1 in ICC. • MiR-145 expression in ICC was associated with Akt signaling and MMPs expression.

Dual role of miR-21 in CD4+ T-cells: activation-induced miR-21 supports survival of memory T-cells and regulates CCR7 expression in naive T-cells.

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Katarzyna Smigielska-Czepiel

Full Text Available Immune cell-type specific miRNA expression patterns have been described but the detailed role of single miRNAs in the function of T-cells remains largely unknown. We investigated the role of miR-21 in the function of primary human CD4+ T-cells. MiR-21 is substantially expressed in T-cells with a memory phenotype, and is robustly upregulated upon αCD3/CD28 activation of both naive and memory T-cells. By inhibiting the endogenous miR-21 function in activated naive and memory T-cells, we showed that miR-21 regulates fundamentally different aspects of T-cell biology, depending on the differentiation status of the T-cell. Stable inhibition of miR-21 function in activated memory T-cells led to growth disadvantage and apoptosis, indicating that the survival of memory T-cells depends on miR-21 function. In contrast, stable inhibition of miR-21 function in activated naive T-cells did not result in growth disadvantage, but led to a significant induction of CCR7 protein expression. Direct interaction between CCR7 and miR-21 was confirmed in a dual luciferase reporter assay. Our data provide evidence for a dual role of miR-21 in CD4+ T cells; Regulation of T-cell survival is confined to activated memory T-cells, while modulation of potential homing properties, through downregulation of CCR7 protein expression, is observed in activated naive T-cells.

Targeting miR-155 to Treat Experimental Scleroderma.

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Yan, Qingran; Chen, Jie; Li, Wei; Bao, Chunde; Fu, Qiong

2016-02-01

Scleroderma is a refractory autoimmune skin fibrotic disorder. Alterations of microRNAs in lesional skin could be a new approach to treating the disease. Here, we found that expression of miR-155 was up regulated in lesional skin tissue from patients with either systemic or localized scleroderma, and correlated with fibrosis area. Then we demonstrated the potential of miR-155 as a therapeutic target in pre-clinical scleroderma models. MiR-155(-/-) mice were resistant to bleomycin induced skin fibrosis. Moreover, topical antagomiR-155 could effectively treat mice primed with subcutaneous bleomycin. In primary skin fibroblast, miR-155 silencing could inhibit collagen synthesis function, as well as signaling intensity of two pro-fibrotic pathways, Wnt/β-catenin and Akt, simultaneously. We further showed that miR-155 could regulate the two pathways via directly targeting casein kinase 1α (CK1α) and Src homology 2-containing inositol phosphatase-1 (SHIP-1), as previous reports. Mice with miR-155 knockout or topical antagomir-155 treatment showed inhibited Wnt/β-catenin and Akt signaling in skin upon bleomycin challenge. Together, our data suggest the potential of miR-155 silencing as a promising treatment for dermal fibrosis, especially in topical applications.

Functional specialization of the plant miR396 regulatory network through distinct microRNA-target interactions.

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Juan M Debernardi

2012-01-01

Full Text Available MicroRNAs (miRNAs are ∼21 nt small RNAs that regulate gene expression in animals and plants. They can be grouped into families comprising different genes encoding similar or identical mature miRNAs. Several miRNA families are deeply conserved in plant lineages and regulate key aspects of plant development, hormone signaling, and stress response. The ancient miRNA miR396 regulates conserved targets belonging to the GROWTH-REGULATING FACTOR (GRF family of transcription factors, which are known to control cell proliferation in Arabidopsis leaves. In this work, we characterized the regulation of an additional target for miR396, the transcription factor bHLH74, that is necessary for Arabidopsis normal development. bHLH74 homologs with a miR396 target site could only be detected in the sister families Brassicaceae and Cleomaceae. Still, bHLH74 repression by miR396 is required for margin and vein pattern formation of Arabidopsis leaves. MiR396 contributes to the spatio-temporal regulation of GRF and bHLH74 expression during leaf development. Furthermore, a survey of miR396 sequences in different species showed variations in the 5' portion of the miRNA, a region known to be important for miRNA activity. Analysis of different miR396 variants in Arabidopsis thaliana revealed that they have an enhanced activity toward GRF transcription factors. The interaction between the GRF target site and miR396 has a bulge between positions 7 and 8 of the miRNA. Our data indicate that such bulge modulates the strength of the miR396-mediated repression and that this modulation is essential to shape the precise spatio-temporal pattern of GRF2 expression. The results show that ancient miRNAs can regulate conserved targets with varied efficiency in different species, and we further propose that they could acquire new targets whose control might also be biologically relevant.

Xenosensor CAR mediates down-regulation of miR-122 and up-regulation of miR-122 targets in the liver

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Kazantseva, Yuliya A.; Yarushkin, Andrei A.; Mostovich, Lyudmila A. [The Institute of Molecular Biology and Biophysics, Timakova str., 2/12, Novosibirsk 630117 (Russian Federation); Pustylnyak, Yuliya A. [Novosibirsk State University, Pirogova str., 2, Novosibirsk 630090 (Russian Federation); Pustylnyak, Vladimir O., E-mail: pustylnyak@ngs.ru [The Institute of Molecular Biology and Biophysics, Timakova str., 2/12, Novosibirsk 630117 (Russian Federation); Novosibirsk State University, Pirogova str., 2, Novosibirsk 630090 (Russian Federation); The Institute International Tomography Center of the Russian Academy of Sciences, Institutskaya str. 3-A, Novosibirsk 630090 (Russian Federation)

2015-10-01

MiR-122 is a major hepatic microRNA, accounting for more than 70% of the total liver miRNA population. It has been shown that miR-122 is associated with liver diseases, including hepatocellular carcinoma. Mir-122 is an intergenic miRNA with its own promoter. Pri-miR-122 expression is regulated by liver-enriched transcription factors, mainly by HNF4α, which mediates the expression via the interaction with a specific DR1 site. It has been shown that phenobarbital-mediated activation of constitutive androstane receptor (CAR), xenobiotic nuclear receptor, is associated with a decrease in miR-122 in the liver. In the present study, we investigated HNF4α–CAR cross-talk in the regulation of miR-122 levels and promitogenic signalling in mouse livers. The level of miR-122 was significantly repressed by treatment with 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP), which is an agonist of mouse CAR. ChIP assays demonstrated that TCPOBOP-activated CAR inhibited HNF4α transactivation by competing with HNF4α for binding to the DR1 site in the pri-miR-122 promoter. Such transcription factor replacement was strongly correlated with miR-122 down-regulation. Additionally, the decrease in miR-122 levels produced by CAR activation is accompanied by an increase in mRNA and cellular protein levels of E2f1 and its accumulation on the target cMyc gene promoter. The increase in accumulation of E2f1 on the target cMyc gene promoter is accompanied by an increase in cMyc levels and transcriptional activity. Thus, our results provide evidence to support the conclusion that CAR activation decreases miR-122 levels through suppression of HNF4α transcriptional activity and indirectly regulates the promitogenic protein cMyc. HNF4α–CAR cross-talk may provide new opportunities for understanding liver diseases and developing more effective therapeutic approaches to better drug treatments. - Highlights: • CAR activation decreased the level of miR-122 in mouse livers. • CAR decreases

Xenosensor CAR mediates down-regulation of miR-122 and up-regulation of miR-122 targets in the liver

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Kazantseva, Yuliya A.; Yarushkin, Andrei A.; Mostovich, Lyudmila A.; Pustylnyak, Yuliya A.; Pustylnyak, Vladimir O.

2015-01-01

MiR-122 is a major hepatic microRNA, accounting for more than 70% of the total liver miRNA population. It has been shown that miR-122 is associated with liver diseases, including hepatocellular carcinoma. Mir-122 is an intergenic miRNA with its own promoter. Pri-miR-122 expression is regulated by liver-enriched transcription factors, mainly by HNF4α, which mediates the expression via the interaction with a specific DR1 site. It has been shown that phenobarbital-mediated activation of constitutive androstane receptor (CAR), xenobiotic nuclear receptor, is associated with a decrease in miR-122 in the liver. In the present study, we investigated HNF4α–CAR cross-talk in the regulation of miR-122 levels and promitogenic signalling in mouse livers. The level of miR-122 was significantly repressed by treatment with 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP), which is an agonist of mouse CAR. ChIP assays demonstrated that TCPOBOP-activated CAR inhibited HNF4α transactivation by competing with HNF4α for binding to the DR1 site in the pri-miR-122 promoter. Such transcription factor replacement was strongly correlated with miR-122 down-regulation. Additionally, the decrease in miR-122 levels produced by CAR activation is accompanied by an increase in mRNA and cellular protein levels of E2f1 and its accumulation on the target cMyc gene promoter. The increase in accumulation of E2f1 on the target cMyc gene promoter is accompanied by an increase in cMyc levels and transcriptional activity. Thus, our results provide evidence to support the conclusion that CAR activation decreases miR-122 levels through suppression of HNF4α transcriptional activity and indirectly regulates the promitogenic protein cMyc. HNF4α–CAR cross-talk may provide new opportunities for understanding liver diseases and developing more effective therapeutic approaches to better drug treatments. - Highlights: • CAR activation decreased the level of miR-122 in mouse livers. • CAR decreases

MiR-29c regulates the expression of miR-34c and miR-449a by targeting DNA methyltransferase 3a and 3b in nasopharyngeal carcinoma

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Niu, Man; Gao, Dan; Wen, Qiuyuan; Wei, Pingpin; Pan, Suming; Shuai, Cijun; Ma, Huiling; Xiang, Juanjuan; Li, Zheng; Fan, Songqing; Li, Guiyuan; Peng, Shuping

2016-01-01

Nasopharyngeal carcinoma (NPC) is prevalent in South East Asia and Southern China particularly, despite the reported 5-year survival ratio is relative higher than other deadly cancers such as liver, renal, pancreas cancer, the lethality is characterized by high metastatic potential in the early stage and high recurrence rate after radiation treatment. MicroRNA-29c was found to be down-regulated in the serum as well as in the tissue of nasopharyngeal carcinoma tissue. In this study, we found accidentally that the transfection of pre-miR-29c or miR-29c mimics significantly increases the expression level of miR-34c and miR-449a but doesn’t affect that of miR-222 using real-time quantitative PCR in nasopharyngeal carcinoma cell lines. To explore the molecular mechanism of the regulatory role, the cells are treated with 5-Aza-2-deoxycytidine (5-Aza-CdR) treatment and the level of miR-34c and miR-449a but not miR-222 accumulated by the treatment. DNA methyltransferase 3a, 3b were down-regulated by the 5-Aza-CdR treatment with western blot and real-time quantitative PCR. We found that pre-miR-29c or miR-29c mimics significantly increases the expression level of miR-34c and miR-449a. We further found DNA methyltransferase 3a and 3b are the target gene of miR-29c. Restoration of miR-29c in NPC cells down-regulated DNA methyltransferase 3a, 3b, but not DNA methyltransferase T1. The regulation of miR-29c/DNMTs/miR-34c/449a is an important molecular axis of NPC development and targeting DNMTs or restoring of miR-29c might be a promising therapy strategy for the prevention of NPC

MiR-181b regulates steatosis in nonalcoholic fatty liver disease via targeting SIRT1.

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Wang, Yunxia; Zhu, Kongxi; Yu, Weihua; Wang, Hongjuan; Liu, Lan; Wu, Qiong; Li, Shuai; Guo, Jianqiang

2017-11-04

Non-alcoholic fatty liver diseases (NAFLD) is one of the leading cause of chronic liver diseases in the world. However, the pathogenesis of NAFLD is still unclear. Emerging studies have demonstrated that microRNAs (miRs) are profoundly involved in NAFLD and related metabolic diseases. Here, we investigated the mechanisms by which miR-181b influences NAFLD via direct targeting SIRT1. The expression of miR181b was up-regulated while SIRT1 was down-regulated in both human NAFLD patients and high fat diet (HFD) induced NAFDL mice model. And palmitic acid (PA) treatment increased the miR-181b expression while decreased SIRT1 expression in HepG2 cells. Further, we identified that SIRT1 is a direct downstream target of miR-181b. Ectopic expression of miR-181b significantly repressed the 3'-UTR reporter activities of SIRT1 in a dose-dependent manner, while the effect of miR-181b was interrupted when the binding site of miR-181b within the SIRT1 3'-UTR was mutated. And overexpression of miR-181b reduced both the mRNA and protein levels of SIRT1 in HepG2 cells. We also found that inhibition of miR-181b expression alleviates hepatic steatosis both in vitro and in vivo. And the effect of miR-181b on steatosis was blocked by SIRT1 overexpression. Taken together, our data indicated that increased expression of miR-181b potentially contributes to altered lipid metabolism in NAFLD. Downregulation of miR-34a may be a therapeutic strategy against NAFLD by regulating its target SIRT1. Copyright © 2017 Elsevier Inc. All rights reserved.

Deregulation of miR-100, miR-99a and miR-199b in tissues and plasma coexists with increased expression of mTOR kinase in endometrioid endometrial carcinoma

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Torres, Anna; Torres, Kamil; Pesci, Anna; Ceccaroni, Marcello; Paszkowski, Tomasz; Cassandrini, Paola; Zamboni, Giuseppe; Maciejewski, Ryszard

2012-01-01

Alterations of mTOR gene expression have been implicated in the pathogenesis of endometrioid endometrial cancer however only few studies explored the cause of increased mTOR activation in this malignancy. miRNAs are small, noncoding RNAs, which were proven to regulated gene expression at the posttranscriptional level. The study aimed to explore deregulation of miRNAs targeting mTOR kinase (miR-99a, miR-100 and miR-199b) as a possible cause of its altered expression in EEC tissues. In addition expression of the three miRNAs was investigated in plasma of EEC patients and was assessed in terms of diagnostic and prognostic utility. We investigated expression of mTOR kinase transcripts in 46 fresh tissue samples. Expression of miR-99a, miR-100 and miR-199b was investigated in the same group of fresh samples, and in additional 58 FFPE sections as well as in 48 plasma samples using qPCR. Relative quantification was performed using experimentally validated endogenous controls. mTOR kinase expression was increased in EEC tissues and was accompanied by decreased expression of all three miRNAs. Down-regulation of the investigated miRNAs was discovered in plasma of EEC patients and miRNA signatures classified EEC tissues (miR-99a/miR-100/miR-199b) and plasma (miR-99a/miR-199b) samples with higher accuracy in comparison to single miRNAs. We also revealed that miR-100 was an independent prognostic marker of overall survival. We conclude that increased expression of mTOR kinase coexists with down-regulation of its targeting miRNAs, which could suggest a new mechanism of mTOR pathway alterations in EEC. In addition, our findings implicate that miRNA signatures can be considered promising biomarkers for early detection and prognosis of endometrioid endometrial carcinoma

miR-122 targets pyruvate kinase M2 and affects metabolism of hepatocellular carcinoma.

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Angela M Liu

Full Text Available In contrast to normal differentiated cells that depend on mitochondrial oxidative phosphorylation for energy production, cancer cells have evolved to utilize aerobic glycolysis (Warburg's effect, with benefit of providing intermediates for biomass production. MicroRNA-122 (miR-122 is highly expressed in normal liver tissue regulating a wide variety of biological processes including cellular metabolism, but is reduced in hepatocellular carcinoma (HCC. Overexpression of miR-122 was shown to inhibit cancer cell proliferation, metastasis, and increase chemosensitivity, but its functions in cancer metabolism remains unknown. The present study aims to identify the miR-122 targeted genes and to investigate the associated regulatory mechanisms in HCC metabolism. We found the ectopic overexpression of miR-122 affected metabolic activities of HCC cells, evidenced by the reduced lactate production and increased oxygen consumption. Integrated gene expression analysis in a cohort of 94 HCC tissues revealed miR-122 level tightly associated with a battery of glycolytic genes, in which pyruvate kinase (PK gene showed the strongest anti-correlation coefficient (Pearson r = -0.6938, p = <0.0001. In addition, reduced PK level was significantly associated with poor clinical outcomes of HCC patients. We found isoform M2 (PKM2 is the dominant form highly expressed in HCC and is a direct target of miR-122, as overexpression of miR-122 reduced both the mRNA and protein levels of PKM2, whereas PKM2 re-expression abrogated the miR-122-mediated glycolytic activities. The present study demonstrated the regulatory role of miR-122 on PKM2 in HCC, having an implication of therapeutic intervention targeting cancer metabolic pathways.

Coevolution Pattern and Functional Conservation or Divergence of miR167s and their targets across Diverse Plant Species.

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Barik, Suvakanta; Kumar, Ashutosh; Sarkar Das, Shabari; Yadav, Sandeep; Gautam, Vibhav; Singh, Archita; Singh, Sharmila; Sarkar, Ananda K

2015-10-13

microRNAs (miRNAs), a class of endogenously produced small non-coding RNAs of 20-21 nt length, processed from precursor miRNAs, regulate many developmental processes by negatively regulating the target genes in both animals and plants. The coevolutionary pattern of a miRNA family and their targets underscores its functional conservation or diversification. The miR167 regulates various aspects of plant development in Arabidopsis by targeting ARF6 and ARF8. The evolutionary conservation or divergence of miR167s and their target genes are poorly understood till now. Here we show the evolutionary relationship among 153 MIR167 genes obtained from 33 diverse plant species. We found that out of the 153 of miR167 sequences retrieved from the "miRBase", 27 have been annotated to be processed from the 3' end, and have diverged distinctively from the other miR167s produced from 5' end. Our analysis reveals that gma-miR167h/i and mdm-miR167a are processed from 3' end and have evolved separately, diverged most resulting in novel targets other than their known ones, and thus led to functional diversification, especially in apple and soybean. We also show that mostly conserved miR167 sequences and their target AUXIN RESPONSE FACTORS (ARFs) have gone through parallel evolution leading to functional diversification among diverse plant species.

MiR-155 modulates the progression of neuropathic pain through targeting SGK3.

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Liu, Shaoxing; Zhu, Bo; Sun, Yan; Xie, Xianfeng

2015-01-01

This study aimed to illustrate the potential effects of miR-155 in neuropathic pain and its potential mechanism. Spragure-Dawley (SD) rats were used for neuropathic pain model of bilateral chronic constriction injury (bCCI) construction. Effects of miR-155 expression on pain threshold of mechanical stimuli (MWT), paw withdrawal threshold latency (PMTL) and cold threshold were analyzed. Target for miR-155 was analyzed using bioinformatics methods. Moreover, effects of miR-155 target gene expression on pain thresholds were also assessed. Compared with the controls and sham group, miR-155 was overexpressed in neuropathic pain rats (P<0.05), but miR-155 slicing could significantly decreased the pain thresholds (P<0.05). Serum and glucocorticoid regulated protein kinase 3 (SGK3) was predicted as the target gene for miR-155, and miR-155 expression was negatively correlated to SGK3 expression. Furthermore, SGK3 overexpression could significantly decreased the pain thresholds which was the same as miR-155 (P<0.05). Moreover, miR-155 slicing and SGK3 overexpression could significantly decrease the painthreshold. The data presented in this study suggested that miR-155 slicing could excellently alleviate neuropathic pain in rats through targeting SGK3 expression. miR-155 may be a potential therapeutic target for neuropathic pain treatment.

Potential impact of miR-137 and its targets in schizophrenia

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Carrie eWright

2013-04-01

Full Text Available The significant impact of microRNAs (miRNAs on disease pathology is becoming increasingly evident. These small non-coding RNAs have the ability to post-transcriptionally silence the expression of thousands of genes. Therefore, dysregulation of even a single miRNA could confer a large polygenic effect. Schizophrenia is a genetically complex illness thought to involve multiple genes each contributing a small risk. Large genome-wide association studies identified miR-137, a miRNA shown to be involved in neuronal maturation, as one of the top risk genes. To assess the potential mechanism of impact of miR-137 in this disorder and identify its targets, we used a combination of literature searches, Ingenuity Pathway Analysis (IPA, and freely accessible bioinformatics resources. Using TargetScan and the Schizophrenia Gene Resource (SZGR database, we found that in addition to CSMD1, C10orf26, CACNA1C, TCF4, and ZNF804A, five schizophrenia risk genes whose transcripts are also validated miR-137 targets, there are other schizophrenia-associated genes that may be targets of miR-137, including ERBB4, GABRA1, GRIN2A, GRM5, GSK3B, NRG2 and HTR2C. IPA analyses of all the potential targets identified several nervous system functions as the top canonical pathways including synaptic long-term potentiation, a process implicated in learning and memory mechanisms and recently shown to be altered in patients with schizophrenia. Among the subset of targets involved in nervous system development and function, the top scoring pathways were ephrin receptor signaling and axonal guidance, processes that are critical for proper circuitry formation and were shown to be disrupted in schizophrenia. These results suggest that miR-137 may indeed play a substantial role in the genetic etiology of schizophrenia by regulating networks involved in neural development and brain function.

Upregulation of miR-150* and miR-630 induces apoptosis in pancreatic cancer cells by targeting IGF-1R.

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Lulu Farhana

Full Text Available MicroRNAs have been implicated in many critical cellular processes including apoptosis. We have previously found that apoptosis in pancreatic cancer cells was induced by adamantyl retinoid-related (ARR molecule 3-Cl-AHPC. Here we report that 3-Cl-AHPC-dependent apoptosis involves regulating a number of microRNAs including miR-150* and miR-630. 3-Cl-AHPC stimulated miR-150* expression and caused decreased expression of c-Myb and IGF-1R in the pancreatic cancer cells. 3-Cl-AHPC-mediated reduction of c-Myb resulted in diminished binding of c-Myb with IGF-1R and Bcl-2 promoters, thereby causing repression of their transcription and protein expression. Over-expression of miR-150* also resulted in diminished levels of c-Myb and Bcl-2 proteins. Furthermore, the addition of the miRNA inhibitor 2'-O-methylated miR-150 blocked 3-Cl-AHPC-mediated increase in miR-150* levels and abrogated loss of c-Myb protein. Knockdown of c-Myb in PANC-1 cells resulted in enhanced apoptosis both in the presence or absence of 3-Cl-AHPC confirming the anti-apoptotic property of c-Myb. Overexpression of miR-630 also induced apoptosis in the pancreatic cancer cells and inhibited target protein IGF-1R mRNA and protein expression. Together these results implicate key roles for miR-150* and miR-630 and their targeting of IGF-1R to promote apoptosis in pancreatic cancer cells.

miR-181a Targets RGS16 to Promote Chondrosarcoma Growth, Angiogenesis, and Metastasis.

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Sun, Xiaojuan; Charbonneau, Cherie; Wei, Lei; Chen, Qian; Terek, Richard M

2015-09-01

Chondrosarcoma is the most common primary malignant bone tumor in adults, has no effective systemic treatment, and patients with this disease have poor survival. Altered expression of microRNA (miR) is involved in tumorigenesis; however, its role in chondrosarcoma is undetermined. miR-181a is overexpressed in high-grade chondrosarcoma, is upregulated by hypoxia, and increases VEGF expression. Here, the purpose was to determine the mechanism of miR-181a regulation of VEGF, determine whether miR-181a overexpression promotes tumor progression, and to evaluate an antagomir-based approach for chondrosarcoma treatment. Therapeutic inhibition of miR-181a decreased expression of VEGF and MMP1 in vitro, and angiogenesis, MMP1 activity, tumor growth, and lung metastasis, all by more than 50%, in a xenograft mouse model. A target of miR-181a is a regulator of G-protein signaling 16 (RGS16), a negative regulator of CXC chemokine receptor 4 (CXCR4) signaling. CXCR4 signaling is increased in chondrosarcoma, its expression is also increased by hypoxia, and is associated with angiogenesis and metastasis; however, receptor blockade is only partially effective. RGS16 expression is restored after miR-181a inhibition and partially accounts for the antiangiogenic and antimetastatic effects of miR-181a inhibition. These data establish miR-181a as an oncomiR that promotes chondrosarcoma progression through a new mechanism involving enhancement of CXCR4 signaling by inhibition of RGS16. Targeting miR-181a can inhibit tumor angiogenesis, growth, and metastasis, thus suggesting the possibility of antagomir-based therapy in chondrosarcoma. ©2015 American Association for Cancer Research.

miR-27 regulates mitochondrial networks by directly targeting the mitochondrial fission factor.

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Tak, Hyosun; Kim, Jihye; Jayabalan, Aravinth Kumar; Lee, Heejin; Kang, Hoin; Cho, Dong-Hyung; Ohn, Takbum; Nam, Suk Woo; Kim, Wook; Lee, Eun Kyung

2014-11-28

Mitochondrial morphology is dynamically regulated by forming small, fragmented units or interconnected networks, and this is a pivotal process that is used to maintain mitochondrial homeostasis. Although dysregulation of mitochondrial dynamics is related to the pathogenesis of several human diseases, its molecular mechanism is not fully elucidated. In this study, we demonstrate the potential role of miR-27 in the regulation of mitochondrial dynamics. Mitochondrial fission factor (MFF) mRNA is a direct target of miR-27, whose ectopic expression decreases MFF expression through binding to its 3'-untranslated region. Expression of miR-27 results in the elongation of mitochondria as well as an increased mitochondrial membrane potential and mitochondrial ATP level. Our results suggest that miR-27 is a novel regulator affecting morphological mitochondrial changes by targeting MFF.

Novel Triazole linked 2-phenyl benzoxazole derivatives induce apoptosis by inhibiting miR-2, miR-13 and miR-14 function in Drosophila melanogaster.

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Mondal, Tanmoy; Lavanya, A V S; Mallick, Akash; Dadmala, Tulshiram L; Kumbhare, Ravindra M; Bhadra, Utpal; Bhadra, Manika Pal

2017-06-01

Apoptosis is an important phenomenon in multi cellular organisms for maintaining tissue homeostasis and embryonic development. Defect in apoptosis leads to a number of disorders like- autoimmune disorder, immunodeficiency and cancer. 21-22 nucleotides containing micro RNAs (miRNAs/miRs) function as a crucial regulator of apoptosis alike other cellular pathways. Recently, small molecules have been identified as a potent inducer of apoptosis. In this study, we have identified novel Triazole linked 2-phenyl benzoxazole derivatives (13j and 13h) as a negative regulator of apoptosis inhibiting micro RNAs (miR-2, miR-13 and miR-14) in a well established in vivo model Drosophila melanogaster where the process of apoptosis is very similar to human apoptosis. These compounds inhibit miR-2, miR-13 and miR-14 activity at their target sites, which induce an increased caspase activity, and in turn influence the caspase dependent apoptotic pathway. These two compounds also increase the mitochondrial reactive oxygen species (ROS) level to trigger apoptotic cell death.

miR-137 suppresses tumor growth of malignant melanoma by targeting aurora kinase A

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Chang, Xiao; Zhang, Haiping [Department of Dermatology and Venereal Disease, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China); Lian, Shi [Department of Dermatology and Venereal Disease, Capital Medical University, Beijing 100069 (China); Zhu, Wei, E-mail: zhuwei_2020@163.com [Department of Dermatology and Venereal Disease, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China)

2016-07-01

As an oncogene, aurora kinase A (AURKA) is overexpressed in various types of human cancers. However, the expression and roles of AURKA in malignant melanoma are largely unknown. In this study, a miR-137-AURKA axis was revealed to regulate melanoma growth. We found a significant increase in levels of AURKA in melanoma. Both genetic knockdown and pharmacologic inhibition of AURKA decreased tumor cell growth in vitro and in vivo. Further found that miR-137 reduced AURKA expression through interaction with its 3′ untranslated region (3′UTR) and that miR-137 was negatively correlated with AURKA expression in melanoma specimens. Overexpression of miR-137 decreased cell proliferation and colony formation in vitro. Notably, re-expression of AURKA significantly rescued miR-137-mediated suppression of cell growth and clonality. In summary, these results reveal that miR-137 functions as a tumor suppressor by targeting AURKA, providing new insights into investigation of therapeutic strategies against malignant melanoma. -- Highlights: •First reported overexpression of AURKA in melanoma. •Targeting AURKA inhibits melanoma growth in vitro and in vivo. •Further found miR-137 suppressed cell growth by binding to AURKA 3′UTR. •Re-expression of AURKA rescued miR-137-mediated suppression. •miR-137-AURKA axis may be potential therapeutic targets of melanoma.

Hypoxia-induced aggressiveness of pancreatic cancer cells is due to increased expression of VEGF, IL-6 and miR-21, which can be attenuated by CDF treatment.

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Bin Bao

Full Text Available Hypoxia is known to play critical roles in cell survival, angiogenesis, tumor invasion, and metastasis. Hypoxia mediated over-expression of hypoxia-inducible factor (HIF has been shown to be associated with therapeutic resistance, and contributes to poor prognosis of cancer patients. Emerging evidence suggest that hypoxia and HIF pathways contributes to the acquisition of epithelial-to-mesenchymal transition (EMT, maintenance of cancer stem cell (CSC functions, and also maintains the vicious cycle of inflammation-all which lead to therapeutic resistance. However, the precise molecular mechanism(s by which hypoxia/HIF drives these events are not fully understood. Here, we show, for the first time, that hypoxia leads to increased expression of VEGF, IL-6, and CSC signature genes Nanog, Oct4 and EZH2 consistent with increased cell migration/invasion and angiogenesis, and the formation of pancreatospheres, concomitant with increased expression of miR-21 and miR-210 in human pancreatic cancer (PC cells. The treatment of PC cells with CDF, a novel synthetic compound inhibited the production of VEGF and IL-6, and down-regulated the expression of Nanog, Oct4, EZH2 mRNAs, as well as miR-21 and miR-210 under hypoxia. CDF also led to decreased cell migration/invasion, angiogenesis, and formation of pancreatospheres under hypoxia. Moreover, CDF decreased gene expression of miR-21, miR-210, IL-6, HIF-1α, VEGF, and CSC signatures in vivo in a mouse orthotopic model of human PC. Collectively, these results suggest that the anti-tumor activity of CDF is in part mediated through deregulation of tumor hypoxic pathways, and thus CDF could become a novel, and effective anti-tumor agent for PC therapy.

ADAR1 attenuates allogeneic graft rejection by suppressing miR-21 biogenesis in macrophages and promoting M2 polarization.

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Li, Junjie; Xie, Jiangang; Liu, Shanshou; Li, Xiao; Zhang, Dongliang; Wang, Xianqi; Jiang, Jinquan; Hu, Wei; Zhang, Yuan; Jin, Boquan; Zhuang, Ran; Yin, Wen

2018-04-25

ADAR1 (adenosine deaminase acting on double-stranded RNA 1) is an RNA-editing enzyme that mediates adenosine-to-inosine RNA editing events, an important post-transcriptional modification mechanism that can alter the coding properties of mRNA or regulate microRNA biogenesis. ADAR1 also regulates the innate immune response. Here, we have demonstrated that ADAR1 expression increased in LPS-stimulated macrophages. Silencing ADAR1 by using small interfering RNA in macrophages resulted in the pronounced polarization of macrophages to M1, whereas ADAR1 overexpression promoted M2 polarization, which indicated that ADAR1 can inhibit macrophage hyperpolarization and prevent immune hyperactivity. The RNA-RNP immunoprecipitation binding assay demonstrated a direct interaction between ADAR1 and miR-21 precursor. Significant up-regulation in IL-10 and down-regulation in miR-21 were observed in ADAR1-overexpressing macrophages. We evaluated miR-21 target mRNAs and macrophage polarization signaling pathways and found that forkhead box protein O1 (Foxo1) was up-regulated in cells that overexpressed ADAR1. In a mouse allogeneic skin transplantation model, grafts in the ADAR1-overexpressed group survived longer and suffered less immune cell infiltration. In ADAR1-overexpressed recipients, splenic macrophages were significantly polarized to M2, and levels of sera IL-10 were markedly higher than those in the control group. In summary, ADAR1 modulates macrophage M2 polarization via the ADAR1-miR-21-Foxo1-IL-10 axis, thereby suppressing allogeneic graft rejection.-Li, J., Xie, J., Liu, S., Li, X., Zhang, D., Wang, X., Jiang, J., Hu, W., Zhang, Y., Jin, B., Zhuang, R., Yin, W. ADAR1 attenuates allogeneic graft rejection by suppressing miR-21 biogenesis in macrophages and promoting M2 polarization.

Functionalized graphene oxide mediated adriamycin delivery and miR-21 gene silencing to overcome tumor multidrug resistance in vitro.

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Feng Zhi

Full Text Available Multidrug resistance (MDR is a major impediment to successful cancer chemotherapy. Co-delivery of novel MDR-reversing agents and anticancer drugs to cancer cells holds great promise for cancer treatment. MicroRNA-21 (miR-21 overexpression is associated with the development and progression of MDR in breast cancer, and it is emerging as a novel and promising MDR-reversing target. In this study, a multifunctional nanocomplex, composed of polyethylenimine (PEI/poly(sodium 4-styrenesulfonates (PSS/graphene oxide (GO and termed PPG, was prepared using the layer-by-layer assembly method to evaluate the reversal effects of PPG as a carrier for adriamycin (ADR along with miR-21 targeted siRNA (anti-miR-21 in cancer drug resistance. ADR was firstly loaded onto the PPG surface (PPGADR by physical mixing and anti-miR-21 was sequentially loaded onto PPGADR through electric absorption to form (anti-miR-21PPGADR. Cell experiments showed that PPG significantly enhanced the accumulation of ADR in MCF-7/ADR cells (an ADR resistant breast cancer cell line and exhibited much higher cytotoxicity than free ADR, suggesting that PPG could effectively reverse ADR resistance of MCF-7/ADR. Furthermore, the enhanced therapeutic efficacy of PPG could be correlated with effective silencing of miR-21 and with increased accumulation of ADR in drug-resistant tumor cells. The endocytosis study confirmed that PPG could effectively carry drug molecules into cells via the caveolae and clathrin-mediated endocytosis pathways. These results suggest that this PPG could be a potential and efficient non-viral vector for reversing MDR, and the strategy of combining anticancer drugs with miRNA therapy to overcome MDR could be an attractive approach in cancer treatment.

Comprehensive RNA dataset of AGO2 associated RNAs in Jurkat cells following miR-21 over-expression

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Claudia Carissimi

2016-06-01

Full Text Available We set out to identify miR-21 targets in Jurkat cells using a high-throughput biochemical approach (10.1016/j.biochi.2014.09.021 [1]. Using a specific monoclonal antibody raised against AGO2, RISC complexes were immunopurified in Jurkat cells over-expressing miR-21 following lentiviral trasduction as well as in Jurkat control cells lines. A parallel immunoprecipitation using isotype-matched rat IgG was performed as a control. AGO2 associated mRNAs were profiled by microarray (GEO: GSE37212. AGO2 bound miRNAs were profiled by RNA-seq.

Uvrag targeting by Mir125a and Mir351 modulates autophagy associated with Ewsr1 deficiency.

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Kim, Yunha; Kang, Young-Sook; Lee, Na-Young; Kim, Ki Yoon; Hwang, Yu Jin; Kim, Hyun-Wook; Rhyu, Im Joo; Her, Song; Jung, Min-Kyung; Kim, Sun; Lee, Chai-Jin; Ko, Seyoon; Kowall, Neil W; Lee, Sean Bong; Lee, Junghee; Ryu, Hoon

2015-01-01

The EWSR1 (EWS RNA-binding protein 1/Ewing Sarcoma Break Point Region 1) gene encodes a RNA/DNA binding protein that is ubiquitously expressed and involved in various cellular processes. EWSR1 deficiency leads to impairment of development and accelerated senescence but the mechanism is not known. Herein, we found that EWSR1 modulates the Uvrag (UV radiation resistance associated) gene at the post-transcription level. Interestingly, EWSR1 deficiency led to the activation of the DROSHA-mediated microprocessor complex and increased the level of Mir125a and Mir351, which directly target Uvrag. Moreover, the Mir125a- and Mir351-mediated reduction of Uvrag was associated with the inhibition of autophagy that was confirmed in ewsr1 knockout (KO) MEFs and ewsr1 KO mice. Taken together, our data indicate that EWSR1 is involved in the post-transcriptional regulation of Uvrag via a miRNA-dependent pathway, resulting in the deregulation of autophagy inhibition. The mechanism of Uvrag and autophagy regulation by EWSR1 provides new insights into the role of EWSR1 deficiency-related cellular dysfunction.

MicroRNA and cellular targets profiling reveal miR-217 and miR-576-3p as proviral factors during Oropouche infection.

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Victor Emmanuel Viana Geddes

2018-05-01

Full Text Available Oropouche Virus is the etiological agent of an arbovirus febrile disease that affects thousands of people and is widespread throughout Central and South American countries. Although isolated in 1950's, still there is scarce information regarding the virus biology and its prevalence is likely underestimated. In order to identify and elucidate interactions with host cells factors and increase the understanding about the Oropouche Virus biology, we performed microRNA (miRNA and target genes screening in human hepatocarcinoma cell line HuH-7. Cellular miRNAs are short non-coding RNAs that regulates gene expression post-transcriptionally and play key roles in several steps of viral infections. The large scale RT-qPCR based screening found 13 differentially expressed miRNAs in Oropouche infected cells. Further validation confirmed that miR-217 and miR-576-3p were 5.5 fold up-regulated at early stages of virus infection (6 hours post-infection. Using bioinformatics and pathway enrichment analysis, we predicted the cellular targets genes for miR-217 and miR-576-3p. Differential expression analysis of RNA from 95 selected targets revealed genes involved in innate immunity modulation, viral release and neurological disorder outcomes. Further analysis revealed the gene of decapping protein 2 (DCP2, a previous known restriction factor for bunyaviruses transcription, as a miR-217 candidate target that is progressively down-regulated during Oropouche infection. Our analysis also showed that activators genes involved in innate immune response through IFN-β pathway, as STING (Stimulator of Interferon Genes and TRAF3 (TNF-Receptor Associated Factor 3, were down-regulated as the infection progress. Inhibition of miR-217 or miR-576-3p restricts OROV replication, decreasing viral RNA (up to 8.3 fold and virus titer (3 fold. Finally, we showed that virus escape IFN-β mediated immune response increasing the levels of cellular miR-576-3p resulting in a decreasing of

Data of expression status of miR- 29a and its putative target mitochondrial apoptosis regulatory gene DRP1 upon miR-15a and miR-214 inhibition

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Muhammad Ishtiaq Jan

2018-02-01

Full Text Available Data is about the mitochondrial apoptosis regulatory framework genes PUMA, DRP1 (apoptotic, and ARC (anti-apoptotic analysis after the employment of their controlling miRNAs inhibitors. The data represents putative conserved targeting of seed regions of miR-15a, miR-29a, and miR-214 with respective target genes PUMA, DRP1, and ARC. Data is of cross interference in expression levels of one miRNA family, miR-29a and its putative target DRP1 upon the inhibitory treatment of other miRNAs 15a and 214. Keywords: DRP1, miR-15a, Apoptosis, miRNAs inhibition

miR-21-3p is a positive regulator of L1CAM in several human carcinomas.

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Doberstein, Kai; Bretz, Niko P; Schirmer, Uwe; Fiegl, Heidi; Blaheta, Roman; Breunig, Christian; Müller-Holzner, Elisabeth; Reimer, Dan; Zeimet, Alain G; Altevogt, Peter

2014-11-28

Expression of L1 cell adhesion molecule (L1CAM) occurs frequently in human cancers and is associated with poor prognosis in cancers such as ovarian, endometrial, breast, renal cell carcinoma and pancreatic ductal adenocarcinoma. L1CAM promotes cell motility, invasion, chemoresistance and metastasis formation. Elucidating genetic processes involved in the expression of L1CAM in cancers is of considerable importance. Transcription factors such as SLUG, β-catenin/TCF-LEF, PAX8 and VHL have been implicated in the re-activation of L1CAM in various types of cancers. There is increasing evidence that micro-RNAs can also have strong effects on gene expression. Here we have identified miR-21-3p as a positive regulator of L1CAM expression. Over-expression of miR-21-3p (miR-21*) but not the complementary sequence miR-21-5p (miR-21) could strongly augment L1CAM expression in renal, endometrial and ovarian carcinoma derived cell lines by an unknown mechanism involving transcriptional activation of the L1CAM gene. In patient cohorts from renal, endometrial and ovarian cancers we observed a strong positive correlation of L1CAM and miR-21-3p expressions. Although L1CAM alone was a reliable marker for overall and disease free survival, the combination of L1CAM and miR-21-3p expressions strongly enhanced the predictive power. Our findings shed new light on the complex regulation of L1CAM in cancers and advocate the use of L1CAM/miR-21-3p for diagnostic application. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

miR-99 inhibits cervical carcinoma cell proliferation by targeting TRIB2.

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Xin, Jia-Xuan; Yue, Zhen; Zhang, Shuai; Jiang, Zhong-Hua; Wang, Ping-Yu; Li, You-Jie; Pang, Min; Xie, Shu-Yang

2013-10-01

MicroRNAs (miRNAs) have significant roles in cell processes, including proliferation, apoptosis and stress responses. To investigate the involvement of miR-99 in the inhibition of HeLa cell proliferation, an miR-99 gene expression vector (pU6.1/miR-99), which overexpressed miR-99 in HeLa cells after transient transfection, was constructed. The expression of miR-99 was detected by qPCR. Cell proliferation and apoptosis were analyzed by cell viability, proliferation and apoptosis assays, as well as by electron microscopy. The results showed that overexpression of miR-99 in HeLa cells increased the HeLa cell mortality rate. Moreover, miR-99 overexpression was able to markedly inhibit HeLa cell proliferation according to the 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The cell apoptosis rate was significantly higher in pU6.1/miR-99-treated cells compared with that in the control cultures. Increases in intracellular electron density, as well as the proportion of nuclear plasma, blebbing phenomena and apoptotic bodies were observed in pU6.1/miR-99-treated cells compared with control cultures according to electron microscopy analysis. The Tribbles 2 (TRIB2) 3'-untranslated region was also observed to be targeted by miR-99 and the results further demonstrated that miR-99 was able to negatively regulate TRIB2 expression in HeLa cells The results indicate that miR-99 acts as a tumor suppressor gene in HeLa cells, establishing a theoretical basis for its application in cancer therapeutics.

Conservation and diversification of the miR166 family in soybean and potential roles of newly identified miR166s.

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Li, Xuyan; Xie, Xin; Li, Ji; Cui, Yuhai; Hou, Yanming; Zhai, Lulu; Wang, Xiao; Fu, Yanli; Liu, Ranran; Bian, Shaomin

2017-02-01

microRNA166 (miR166) is a highly conserved family of miRNAs implicated in a wide range of cellular and physiological processes in plants. miR166 family generally comprises multiple miR166 members in plants, which might exhibit functional redundancy and specificity. The soybean miR166 family consists of 21 members according to the miRBase database. However, the evolutionary conservation and functional diversification of miR166 family members in soybean remain poorly understood. We identified five novel miR166s in soybean by data mining approach, thus enlarging the size of miR166 family from 21 to 26 members. Phylogenetic analyses of the 26 miR166s and their precursors indicated that soybean miR166 family exhibited both evolutionary conservation and diversification, and ten pairs of miR166 precursors with high sequence identity were individually grouped into a discrete clade in the phylogenetic tree. The analysis of genomic organization and evolution of MIR166 gene family revealed that eight segmental duplications and four tandem duplications might occur during evolution of the miR166 family in soybean. The cis-elements in promoters of MIR166 family genes and their putative targets pointed to their possible contributions to the functional conservation and diversification. The targets of soybean miR166s were predicted, and the cleavage of ATHB14-LIKE transcript was experimentally validated by RACE PCR. Further, the expression patterns of the five newly identified MIR166s and 12 target genes were examined during seed development and in response to abiotic stresses, which provided important clues for dissecting their functions and isoform specificity. This study enlarged the size of soybean miR166 family from 21 to 26 members, and the 26 soybean miR166s exhibited evolutionary conservation and diversification. These findings have laid a foundation for elucidating functional conservation and diversification of miR166 family members, especially during seed development or

Irradiation-Induced Cardiac Connexin-43 and miR-21 Responses Are Hampered by Treatment with Atorvastatin and Aspirin

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Csilla Viczenczova

2018-04-01

Full Text Available Radiation of the chest during cancer therapy is deleterious to the heart, mostly due to oxidative stress and inflammation related injury. A single sub-lethal dose of irradiation has been shown to result in compensatory up-regulation of the myocardial connexin-43 (Cx43, activation of the protein kinase C (PKC signaling along with the decline of microRNA (miR-1 and an increase of miR-21 levels in the left ventricle (LV. We investigated whether drugs with antioxidant, anti-inflammatory or vasodilating properties, such as aspirin, atorvastatin, and sildenafil, may affect myocardial response in the LV and right ventricle (RV following chest irradiation. Adult, male Wistar rats were subjected to a single sub-lethal dose of chest radiation at 25 Gy and treated with aspirin (3 mg/day, atorvastatin (0.25 mg/day, and sildenafil (0.3 mg/day for six weeks. Cx43, PKCε and PKCδ proteins expression and levels of miR-1 as well as miR-21 were determined in the LV and RV. Results showed that the suppression of miR-1 was associated with an increase of total and phosphorylated forms of Cx43 as well as PKCε expression in the LV while having no effect in the RV post-irradiation as compared to the non-irradiated rats. Treatment with aspirin and atorvastatin prevented an increase in the expression of Cx43 and PKCε without change in the miR-1 levels. Furthermore, treatment with aspirin, atorvastatin, and sildenafil completely prevented an increase of miR-21 in the LV while having partial effect in the RV post irradiation. The increase in pro-apoptotic PKCδ was not affected by any of the used treatment. In conclusion, irradiation and drug-induced changes were less pronounced in the RV as compared to the LV. Treatment with aspirin and atorvastatin interfered with irradiation-induced compensatory changes in myocardial Cx43 protein and miR-21 by preventing their elevation, possibly via amelioration of oxidative stress and inflammation.

Gender-dependent expression of leading and passenger strand of miR-21 and miR-16 in human colorectal cancer and adjacent colonic tissues.

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Hasáková, K; Bezakova, J; Vician, M; Reis, R; Zeman, M; Herichova, I

2017-12-30

miRNAs are small regulatory RNA molecules involved in posttranscriptional gene silencing. Their biosynthesis results in the formation of duplex consisting of a leading and a passenger strand of mature miRNA. The leading strand exhibits the main activity but recent findings indicate a certain role of the passenger strand as well. Deregulated levels of miRNA were found in many types of cancers including colorectal cancer. miR-21 and miR-16 were indicated as possible markers of colorectal cancer, however, small attention to gender differences in their expression was paid so far. Therefore, the aim of our study was to investigate the expression of miR-21-5p, miR-21-3p, miR-16-5p and miR-16-3p in human colorectal cancer tissue and compare it to the adjacent tissues taken during surgery in men and women separately. Our results showed an up-regulation of all measured miRNAs in tumor tissue compared to adjacent tissues. As expected, tumors and adjacent tissues exhibited a significantly higher expression of leading miRNAs compared to passenger strand of miR-21 and miR-16. The expression of leading and passenger strand of miR-21 and miR-16 positively correlated exhibiting the highest correlation coefficient in the distal tissue. The expression pattern showed gender-dependent differences, with higher levels of miRNA in men than in women. Our findings indicate a gender-related expression pattern of miRNA, which should be considered as an important factor in generating new prognostic or diagnostic biomarkers.

miR-340 alleviates chemoresistance of osteosarcoma cells by targeting ZEB1.

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Yan, Haibin; Zhang, Bingyun; Fang, Chongbin; Chen, Liqiu

2018-06-01

Chemoresistance during treatment of osteosarcoma (OS) is attracting more and more attention as the main clinical obstacle. The purpose of this study was to elucidate the role of miR-340 in chemoresistance of OS. Plasmid construction and transfection, miRNA arrays, PCR analyses, and western blot analysis, as well as MTT, apoptosis, and luciferase assays were carried out in MG-63 cells and MG-63/cisplatin (DDP)-resistant cells. The results showed that miR-340 was downregulated in OS tissues and drug-resistant OS cells. Moreover, a negative correlation was observed between miR-340 and ZEB1 expression in OS tissues. Forced expression of miR-340 in drug-resistant OS cells significantly reduced multidrug resistance-1 and P-gp expression. Overexpression of miR-340 enhanced sensitivity to DDP by inhibiting viability and promoting apoptosis. The luciferase assay and western blot analysis identified ZEB1 as a direct target of miR-340, and miR-340 negatively regulated ZEB1 expression. Ectopic expression of ZEB1 reversed the effects of miR-340 on P-gp expression, cell viability, and apoptosis. miR-340 alleviated chemoresistance of OS cells by targeting ZEB1. Our results indicate that targeting miR-340 may be a potential therapeutic approach to treat drug-resistant OS.

miR398 and miR395 are involved in response to SO2 stress in Arabidopsis thaliana.

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Li, Lihong; Yi, Huilan; Xue, Meizhao; Yi, Min

2017-11-01

Sulfur dioxide (SO 2 ) is a common air pollutant that has adverse effects on plants. MicroRNAs (miRNAs) are small noncoding RNA that play critical roles in plant development and stress response. In this study, we found that two miRNAs, miR398 and miR395, were differentially expressed in Arabidopsis shoots under SO 2 stress. The expression of miR398 was down-regulated, and the transcript levels of its target genes, Cu/Zn superoxide dismutases (CSD1 and CSD2), were increased during SO 2 exposure. The activity of superoxide dismutase (SOD), one of the major antioxidant enzymes, was enhanced with the increase in the CSD transcript level, suggesting an important role of miR398 in response to SO 2 -induced oxidative stress. Meanwhile, the expression of miR395 was increased, and the transcript levels of its target genes, ATP sulfurylases (APS3 and APS4) and a low-affinity sulfate transporter (SULTR2;1), were decreased in Arabidopsis shoots, showing that miR395 played important roles in the regulation of sulfate assimilation and translocation during SO 2 exposure. The content of glutathione (GSH), an important sulfur-containing antioxidant, was enhanced with the changes in sulfur metabolism in Arabidopsis shoots under SO 2 stress. These results showed that both miR398 and miR395 were involved in protecting plants from oxidative damage during SO 2 exposure. Many stress-responsive cis-elements were found in the promoter regions of MIR398 and MIR395, suggesting that these miRNAs might respond to various environmental conditions, including SO 2 stress. Overall, our study provides an insight into the regulatory roles of miRNAs in response to SO 2 stress in plants, and highlights the molecular mechanisms of plant adaptation to environmental stress.

miR-486-3p, miR-139-5p, and miR-21 as Biomarkers for the Detection of Oral Tongue Squamous Cell Carcinoma

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Zujian Chen

2017-01-01

Full Text Available Oral tongue squamous cell carcinoma (TSCC is a complex disease with extensive genetic and epigenetic defects, including microRNA deregulation. The aims of the present study were to test the feasibility of performing the microRNA profiling analysis on archived TSCC specimens and to assess the potential diagnostic utility of the identified microRNA biomarkers for the detection of TSCC. TaqMan array-based microRNA profiling analysis was performed on 10 archived TSCC samples and their matching normal tissues. A panel of 12 differentially expressed microRNAs was identified. Eight of these differentially expressed microRNAs were validated in an independent sample set. A random forest (RF classification model was built with miR-486-3p, miR-139-5p, and miR-21, and it was able to detect TSCC with a sensitivity of 100% and a specificity of 86.7% (overall error rate = 6.7%. As such, this study demonstrated the utility of the archived clinical specimens for microRNA biomarker discovery. The feasibility of using microRNA biomarkers (miR-486-3p, miR-139-5p, and miR-21 for the detection of TSCC was confirmed.

MiR-32 promotes gastric carcinoma tumorigenesis by targeting Kruppel-like factor 4

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Yan, Chao [Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, Beijing, 100730 (China); Yu, Jianchun, E-mail: yu_jchpumch@163.com [Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, Beijing, 100730 (China); Liu, Yuqin [Cell Culture Center, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100005 (China); Kang, Weiming; Ma, Zhiqiang; Zhou, Li [Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, Beijing, 100730 (China)

2015-11-27

Gastric cancer (GC) is a prevalent malignant cancer worldwide and is highly lethal because of its fast growth. Currently, the clinical therapy options for GC remain limited. MiR-32 has been reported as an oncogenic microRNA in many cancers, but its role in GC is unclear. Here, we found that miR-32 was overexpressed in GC tissues compared with adjacent normal tissue, and miR-32 was higher in GC patients' plasma compared with healthy individuals. Furthermore, we have identified miR-32 to be oncogenic, by promoting gastric cell proliferation, migration and invasion. We also identified Kruppel-like factor 4 (KLF4) as a direct target of miR-32. Knockdown of KLF4 promoted proliferation, migration and invasion of GC cells. We conclude that miR-32 promotes GC cell proliferation, migration and invasion by targeting KLF4, suggesting that the miR-32-KLF4 pathway may be useful in clinical diagnosis and therapeutics. - Highlights: • miR-32 was overexpression in GC tissues than adjacent normal tissue. • miR-32 was higher in GC patients' plasma compared with healthy people. • miR-32 promotes GC cell proliferation, migration and invasion by targeting KLF4.

miR-184 and miR-150 promote renal glomerular mesangial cell aging by targeting Rab1a and Rab31.

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Liu, Xiujuan; Fu, Bo; Chen, Dapeng; Hong, Quan; Cui, Jing; Li, Jin; Bai, Xueyuan; Chen, Xiangmei

2015-08-15

The molecular mechanism of kidney aging is not well understood, but the abnormal expression of miRNAs with aging is considered to be an important contributor. miR-184 and miR-150 were screened using a miRNA microarray and qRT-PCR and found to be significantly upregulated in 24-month-old rats. Rat renal primary glomerular mesangial cells (GMCs) were isolated from 3-month and 24-month-old rats for the in vitro analysis of the roles of miR-184 and miR-150 in kidney aging. Bioinformatics analyses suggested that Rab1a and Rab31, which are associated with cell autophagy, were targeted by both miR-184 and miR-150. miR-184 and miR-150 were increased significantly in aging GMCs versus young cells, while Rab1a and Rab31 were significantly lower in aging cells. Furthermore, dual luciferase reporter assays revealed that miR-184 and miR-150 bound to the 3'-UTR of Rab1a and Rab31 mRNAs. Transfection of miR-184 and miR-150 mimics into young GMCs suppressed the expression of Rab1a and Rab31. Transfected cells showed lower autophagy activities and higher levels of cellular oxidative products, leading to the aging of young GMCs. However, miR-184 and miR-150 inhibitors promoted autophagy and reduced oxidative damage by upregulating Rab1a and Rab31 in old GMCs. In conclusion, miR-184 and miR-150 inhibited autophagy, promoting GMC aging. Copyright © 2015 Elsevier Inc. All rights reserved.

MiR-217 is down-regulated in psoriasis and promotes keratinocyte differentiation via targeting GRHL2

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Zhu, Haigang; Hou, Liyue; Liu, Jingjing; Li, Zhiming

2016-01-01

MiR-217 is a well-known tumor suppressor, and its down-regulation has been shown in a wide range of solid and leukaemic cancers. However, the biological role of miR-217 in psoriasis pathogenesis, especially in keratinocyte hyperproliferation and differentiation, is not clearly understood. In this study, we found the expression of miR-217 was markedly down-regulated in psoriasis keratinocytes of psoriatic patients. In addition, overexpression of miR-217 inhibited the proliferation and promoted the differentiation of primary human keratinocytes. On the contrary, inhibition of endogenous miR-217 increased cell proliferation and delayed differentiation. Furthermore, Grainyhead-like 2 (GRHL2) was identified as a direct target of miR-217 by luciferase reporter assay. The expression of miR-217 and GRHL2 was inversely correlated in both transfected keratinocytes and in psoriasis lesional skin. Moreover, knocking down GRHL2 expression by siRNA enhanced keratinocyte differentiation. Taken together, our results demonstrate a role for miR-217 in the regulation of keratinocyte differentiation, partially through the regulation of GRHL2. - Highlights: • miR-217 is down-regulated in psoriasis skin lesions. • miR-217 inhibits the proliferation and promotes differentiation of keratinocytes. • GRHL2 is a novel target of miR-217 in keratinocytes. • GRHL2 is up-regulated and inversely correlated with miR-217 in psoriasis skin lesions.

MiR-217 is down-regulated in psoriasis and promotes keratinocyte differentiation via targeting GRHL2

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Zhu, Haigang; Hou, Liyue; Liu, Jingjing; Li, Zhiming, E-mail: lizm_1001@sina.com

2016-02-26

MiR-217 is a well-known tumor suppressor, and its down-regulation has been shown in a wide range of solid and leukaemic cancers. However, the biological role of miR-217 in psoriasis pathogenesis, especially in keratinocyte hyperproliferation and differentiation, is not clearly understood. In this study, we found the expression of miR-217 was markedly down-regulated in psoriasis keratinocytes of psoriatic patients. In addition, overexpression of miR-217 inhibited the proliferation and promoted the differentiation of primary human keratinocytes. On the contrary, inhibition of endogenous miR-217 increased cell proliferation and delayed differentiation. Furthermore, Grainyhead-like 2 (GRHL2) was identified as a direct target of miR-217 by luciferase reporter assay. The expression of miR-217 and GRHL2 was inversely correlated in both transfected keratinocytes and in psoriasis lesional skin. Moreover, knocking down GRHL2 expression by siRNA enhanced keratinocyte differentiation. Taken together, our results demonstrate a role for miR-217 in the regulation of keratinocyte differentiation, partially through the regulation of GRHL2. - Highlights: • miR-217 is down-regulated in psoriasis skin lesions. • miR-217 inhibits the proliferation and promotes differentiation of keratinocytes. • GRHL2 is a novel target of miR-217 in keratinocytes. • GRHL2 is up-regulated and inversely correlated with miR-217 in psoriasis skin lesions.

Epigenetically altered miR-193b targets cyclin D1 in prostate cancer

International Nuclear Information System (INIS)

Kaukoniemi, Kirsi M; Rauhala, Hanna E; Scaravilli, Mauro; Latonen, Leena; Annala, Matti; Vessella, Robert L; Nykter, Matti; Tammela, Teuvo L J; Visakorpi, Tapio

2015-01-01

Micro-RNAs (miRNA) are important regulators of gene expression and often differentially expressed in cancer and other diseases. We have previously shown that miR-193b is hypermethylated in prostate cancer (PC) and suppresses cell growth. It has been suggested that miR-193b targets cyclin D1 in several malignancies. Here, our aim was to determine if miR-193b targets cyclin D1 in prostate cancer. Our data show that miR-193b is commonly methylated in PC samples compared to benign prostate hyperplasia. We found reduced miR-193b expression (P < 0.05) in stage pT3 tumors compared to pT2 tumors in a cohort of prostatectomy specimens. In 22Rv1 PC cells with low endogenous miR-193b expression, the overexpression of miR-193b reduced CCND1mRNA levels and cyclin D1 protein levels. In addition, the exogenous expression of miR-193b decreased the phosphorylation level of RB, a target of the cyclin D1-CDK4/6 pathway. Moreover, according to a reporter assay, miR-193b targeted the 3’UTR of CCND1 in PC cells and the CCND1 activity was rescued by expressing CCND1 lacking its 3’UTR. Immunohistochemical analysis of cyclin D1 showed that castration-resistant prostate cancers have significantly (P = 0.0237) higher expression of cyclin D1 compared to hormone-naïve cases. Furthermore, the PC cell lines 22Rv1 and VCaP, which express low levels of miR-193b and high levels of CCND1, showed significant growth retardation when treated with a CDK4/6 inhibitor. In contrast, the inhibitor had no effect on the growth of PC-3 and DU145 cells with high miR-193b and low CCND1 expression. Taken together, our data demonstrate that miR-193b targets cyclin D1 in prostate cancer

STAT3-regulated exosomal miR-21 promotes angiogenesis and is involved in neoplastic processes of transformed human bronchial epithelial cells.

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Liu, Yi; Luo, Fei; Wang, Bairu; Li, Huiqiao; Xu, Yuan; Liu, Xinlu; Shi, Le; Lu, Xiaolin; Xu, Wenchao; Lu, Lu; Qin, Yu; Xiang, Quanyong; Liu, Qizhan

2016-01-01

Although microRNA (miRNA) enclosed in exosomes can mediate intercellular communication, the roles of exosomal miRNA and angiogenesis in lung cancer remain unclear. We investigated functions of STAT3-regulated exosomal miR-21 derived from cigarette smoke extract (CSE)-transformed human bronchial epithelial (HBE) cells in the angiogenesis of CSE-induced carcinogenesis. miR-21 levels in serum were higher in smokers than those in non-smokers. The medium from transformed HBE cells promoted miR-21 levels in normal HBE cells and angiogenesis of human umbilical vein endothelial cells (HUVEC). Transformed cells transferred miR-21 into normal HBE cells via exosomes. Knockdown of STAT3 reduced miR-21 levels in exosomes derived from transformed HBE cells, which blocked the angiogenesis. Exosomes derived from transformed HBE cells elevated levels of vascular endothelial growth factor (VEGF) in HBE cells and thereby promoted angiogenesis in HUVEC cells. Inhibition of exosomal miR-21, however, decreased VEGF levels in recipient cells, which blocked exosome-induced angiogenesis. Thus, miR-21 in exosomes leads to STAT3 activation, which increases VEGF levels in recipient cells, a process involved in angiogenesis and malignant transformation of HBE cells. These results, demonstrating the function of exosomal miR-21 from transformed HBE cells, provide a new perspective for intervention strategies to prevent carcinogenesis of lung cancer. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

miR-181b functions as an oncomiR in colorectal cancer by targeting PDCD4

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Yanqing Liu

2016-09-01

Full Text Available Abstract Programmed cell death 4 (PDCD4 is a RNA-binding protein that acts as a tumor suppressor in many cancer types, including colorectal cancer (CRC. During CRC carcinogenesis, PDCD4 protein levels remarkably decrease, but the underlying molecular mechanism for decreased PDCD4 expression is not fully understood. In this study, we performed bioinformatics analysis to identify miRNAs that potentially target PDCD4. We demonstrated miR-181b as a direct regulator of PDCD4. We further showed that activation of IL6/STAT3 signaling pathway increased miR-181b expression and consequently resulted in downregulation of PDCD4 in CRC cells. In addition, we investigated the biological effects of PDCD4 inhibition by miR-181b both in vitro and in vivo and found that miR-181b could promote cell proliferation and migration and suppress apoptosis in CRC cells and accelerate tumor growth in xenograft mice, potentially through targeting PDCD4. Taken together, this study highlights an oncomiR role for miR-181b in regulating PDCD4 in CRC and suggests that miR-181b may be a novel molecular therapeutic target for CRC.

PAI-1–regulated miR-21 defines a novel age-associated fibrogenic pathway in muscular dystrophy

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Ardite, Esther; Perdiguero, Eusebio; Vidal, Berta; Gutarra, Susana; Serrano, Antonio L.

2012-01-01

Disruption of skeletal muscle homeostasis by substitution with fibrotic tissue constitutes the principal cause of death in Duchenne muscular dystrophy (DMD) patients, yet the implicated fibrogenic mechanisms remain poorly understood. This study identifies the extracellular PAI-1/urokinase-type plasminogen activator (uPA) balance as an important regulator of microribonucleic acid (miR)–21 biogenesis, controlling age-associated muscle fibrosis and dystrophy progression. Genetic loss of PAI-1 in mdx dystrophic mice anticipated muscle fibrosis through these sequential mechanisms: the alteration of collagen metabolism by uPA-mediated proteolytic processing of transforming growth factor (TGF)–β in muscle fibroblasts and the activation of miR-21 expression, which inhibited phosphatase and tensin homologue and enhanced AKT signaling, thus endowing TGF-β with a remarkable cell proliferation–promoting potential. Age-associated fibrogenesis and muscle deterioration in mdx mice, as well as exacerbated dystrophy in young PAI-1−/− mdx mice, could be reversed by miR-21 or uPA-selective interference, whereas forced miR-21 overexpression aggravated disease severity. The PAI-1–miR-21 fibrogenic axis also appeared dysregulated in muscle of DMD patients, providing a basis for effectively targeting fibrosis and muscular dystrophies in currently untreatable individuals. PMID:22213800

MiR-200a is involved in rat epididymal development by targeting β-catenin mRNA

Institute of Scientific and Technical Information of China (English)

Xiaojiang Wu; Botao Zhao; Wei Li; Yue Chen; Ruqiang Liang; Lin Li; Youxin Jin; Kangcheng Ruan

2012-01-01

The expression of 350 microRNAs (miRNAs) in epididymis of rat from postnatal development to adult (from postnatal days 7-70) was profiled with home-made miRNA microarray.Among them,48 miRNAs changed significantly, in which the expression of miR-200a increased obviously with time,in a good agreement with that obtained from northern blot analysis.The real-time quantitative-polymerase chain reaction result indicated that temporal expression of rat β-catenin was exactly inversed to that of miR-200a during rat epididymal development,implying that miR-200a might also target β-catenin mRNA in rat epididymis as reported by Saydam et al.in humans.The bioinformatic analysis indicated that 3' untranslated region of rat β-catenin mRNA did contain a putative binding site for miR-200a.Meanwhile,it was found that the sequence of this binding site was different from that of human β-catenin mRNA with a deletion of two adjacent nucleotides (U and C).But the results of luciferase targeting assay in HEK 293T cells and the overexpression of miR-200a in rat NRK cells demonstrated that miR-200a did target rat β-catenin mRNA and cause the suppression of its expression.All these results show that miR-200a should be involved in rat epididymal development by targeting β-catenin mRNA of rat and suppressing its expression.

miR-625 suppresses cell proliferation and migration by targeting HMGA1 in breast cancer

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Zhou, Wen-bin; Zhong, Cai-neng; Luo, Xun-peng; Zhang, Ya-yuan; Zhang, Gui-ying [Department of Breast Surgery, Second Clinical Medical College of Jinan University, Shenzhen People' s Hospital, Shenzhen, Guangdong Province (China); Zhou, Dong-xian, E-mail: 1072241978@qq.com [Department of Breast Surgery, Second Clinical Medical College of Jinan University, Shenzhen People' s Hospital, Shenzhen, Guangdong Province (China); Liu, Li-ping, E-mail: leoliping@aliyun.com [Department of Hepatobiliary and Pancreas Surgery, Second Clinical Medical College of Jinan University, Shenzhen People' s Hospital, Shenzhen, Guangdong Province (China)

2016-02-19

Dysregulation of microRNA contributes to the high incidence and mortality of breast cancer. Here, we show that miR-625 was frequently down-regulated in breast cancer. Decrease of miR-625 was closely associated with estrogen receptor (P = 0.004), human epidermal growth factor receptor 2 (P = 0.003) and clinical stage (P = 0.001). Kaplan–Meier and multivariate analyses indicated miR-625 as an independent factor for unfavorable prognosis (hazard ratio = 2.654, 95% confident interval: 1.300–5.382, P = 0.007). Re-expression of miR-625 impeded, whereas knockdown of miR-625 enhanced cell viabilities and migration abilities in breast cancer cells. HMGA1 was confirmed as a direct target of miR-625. The expressions of HMGA1 mRNA and protein were induced by miR-625 mimics, but reduced by miR-625 inhibitor. Re-introduction of HMGA1 in cells expressing miR-625 distinctly abrogated miR-625-mediated inhibition of cell growth. Taken together, our data demonstrate that miR-625 suppresses cell proliferation and migration by targeting HMGA1 and suggest miR-625 as a promising prognostic biomarker and a potential therapeutic target for breast cancer. - Highlights: • miR-625 expression was significantly decreased in breast cancer. • Decrease of miR-625 was associated with poor clinical outcomes and unfavorable overall survival. • miR-625 overexpression inhibits cell proliferation and migration in vitro. • miR-625 directly targets and suppresses the expression of HMGA1.

MiR-19a targets suppressor of cytokine signaling 1 to modulate the progression of neuropathic pain.

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Wang, Conghui; Jiang, Qi; Wang, Min; Li, Dong

2015-01-01

We aimed to investigate whether miR-19a is associated with neuropathic pain and elucidate the underlying regulatory mechanism. We established a neuropathic pain model of bilateral chronic constriction injury (bCCI). Then bCCI rats were injected with mo-miR-19a, siR-SOCS1 or blank expression vector through a microinjection syringe via an intrathecal catheter on 3 day before surgery and after surgery. Behavioral tests, such as mechanical allodynia, thermal hyperalgesia and acetone induced cold allodynia, were performed to evaluate the pain threshold. Besides, quantitative real-time polymerase chain reaction (qRT-PCR) was performed to determine the expression of miR-19a and western blotting was carried out to measure the expression of SOCS1. miR-19a expression levels were markedly increased in neuropathic pain models. Moreover, miR-19a significantly attenuated mechanical allodynia and thermal hyperalgesia, and similar results were obtained after knockdown of SOCS1 expression. However, miR-19a markedly increased the times that the rats appeared a sign of cold allodynia, and knockdown of SOCS1 expression had similar effects. Besides, the results of bioinformatics analysis and western blotting analysis were all confirmed that SOCS1 was a direct target of miR-19a in neuropathic pain models. Our finding indicate that SOCS1 is a direct target of miR-19a in neuropathic pain rats and miR-19a may play a critical role in regulating of neuropathic pain via targeting SOCS1.

Effects of microRNA-21 on Nerve Cell Regeneration and Neural Function Recovery in Diabetes Mellitus Combined with Cerebral Infarction Rats by Targeting PDCD4.

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Guo, Yun-Bao; Ji, Tie-Feng; Zhou, Hong-Wei; Yu, Jin-Lu

2018-03-01

We aimed to determine the effect and mechanism of microRNA-21 (miR-21) on nerve cell regeneration and nerve functional recovery in diabetes mellitus combined with cerebral infarction (DM + CI) rats by targeting PDCD4. A total of 125 male Wistar rats were selected for DM + CI rat model construction and assigned into the blank, miR-21 mimics, mimics control, miR-21 inhibitor, inhibitor control, miR-21 inhibitor + si-PDCD4 and si-PDCD4 groups. And, 20 healthy rats were selected for the normal group. Triphenylterazolium chloride (TTC) staining and HE staining were used for determination of the area of CI and pathological changes, respectively. Behaviors of rats in the eight groups were determined by forelimb placement test and balance beam walking test. Immunohistochemical staining, double immunofluorescence staining assay, Western blotting, and qRT-PCR were used to detect expressions of miR-21, PDCD4, HNA, Nestin, NeuN, β-III-Tub, PTEN, FasL, and GFAP. DNA laddering and TUNEL staining was used for cell apoptosis. TTC and HE staining confirmed that 87.5% rats were induced into CI + DM models successfully. Results of forelimb placement test and balance beam walking test showed that miR-21 mimics, and si-PCDC4 improved the nerve defect of model rats. Comparing with the blank group at the same time, rats in the miR-21 inhibitor group displayed significant decrease in the forelimb placement test score, significant increase in the balance beam walking test score, and exacerbation of nerve defect, while rats in the miR-21 mimics and si-PCDC4 groups displayed significant increase in forelimb placement test score and significant decrease in the balance beam walking test score and improvement of nerve defect situation. The HNA, Nestin, and PDCD4 expressions were decreased and the NeuN, β-III-Tub, and GFAP expressions were increased in the miR-21 mimics and si-PDCD4 groups comparing with the blank group. The results of miR-21 inhibitor group were on the contrary. In

miR-598 acts as a tumor suppressor in human gastric cancer by targeting IGF-1R.

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Liu, Na; Yang, Hua; Wang, Hong

2018-01-01

In recent years, the aberrant expression of miR-598 in tumorigenesis has been demonstrated, as well as the fact that the IGF-1R pathway is also involved in the development of human gastric cancer (GC). The present study aimed to investigate the molecular mechanisms underlying miR-598-regulated IGF-1R expression in human GC. We analyzed the expression of miR-598 and IGF-1R in GC samples and cells, and evaluated the clinical significance of miR-598 and IGF-1R in GC patients. Furthermore, in vitro and in vivo assays were used to investigate the molecular mechanisms of miR-598 and IGF-1R. miR-598 expression was frequently downregulated in GC tissues and cells, and significantly correlated with poor prognosis, vascular invasion, TNM stage, and lymph node metastases as well as IGF-1R expression. The overexpression of miR-598 obviously inhibited cell proliferation, migration, invasion, and induced cell cycle arrest in the G1/S phase, and increased the apoptosis of GC cells. The overexpression of miR-598 also significantly inhibited ERK1/2 and Akt phosphorylation level. In vivo assay validated the inhibitory effect of miR-598 on tumor growth. Further studies showed that miR-598 inhibited IGF-1R protein expression by directly targeting its 3'-UTR. Besides, over-expression of IGF-1R reversed the inhibitory effects of miR-598, while suppression of IGF-1R expression showed inverse effects. miR-598 suppresses GC cell proliferation, migration and invasion by directly targeting IGF-1R expression. Thus, miR-598 may be a useful target for GC patients.

DNA damage responsive miR-33b-3p promoted lung cancer cells survival and cisplatin resistance by targeting p21WAF1/CIP1.

Science.gov (United States)

Xu, Shun; Huang, Haijiao; Chen, Yu-Ning; Deng, Yun-Ting; Zhang, Bing; Xiong, Xing-Dong; Yuan, Yuan; Zhu, Yanmei; Huang, Haiyong; Xie, Luoyijun; Liu, Xinguang

2016-11-01

Cisplatin is the most potent and widespread used chemotherapy drug for lung cancer treatment. However, the development of resistance to cisplatin is a major obstacle in clinical therapy. The principal mechanism of cisplatin is the induction of DNA damage, thus the capability of DNA damage response (DDR) is a key factor that influences the cisplatin sensitivity of cancer cells. Recent advances have demonstrated that miRNAs (microRNAs) exerted critical roles in DNA damage response; nonetheless, the association between DNA damage responsive miRNAs and cisplatin resistance and its underlying molecular mechanism still require further investigation. The present study has attempted to identify differentially expressed miRNAs in cisplatin induced DNA damage response in lung cancer cells, and probe into the effects of the misexpressed miRNAs on cisplatin sensitivity. Deep sequencing showed that miR-33b-3p was dramatically down-regulated in cisplatin-induced DNA damage response in A549 cells; and ectopic expression of miR-33b-3p endowed the lung cancer cells with enhanced survival and decreased γH2A.X expression level under cisplatin treatment. Consistently, silencing of miR-33b-3p in the cisplatin-resistant A549/DDP cells evidently sensitized the cells to cisplatin. Furthermore, we identified CDKN1A (p21) as a functional target of miR-33b-3p, a critical regulator of G1/S checkpoint, which potentially mediated the protection effects of miR-33b-3p against cisplatin. In aggregate, our results suggested that miR-33b-3p modulated the cisplatin sensitivity of cancer cells might probably through impairing the DNA damage response. And the knowledge of the drug resistance conferred by miR-33b-3p has great clinical implications for improving the efficacy of chemotherapies for treating lung cancers.

MDRL lncRNA regulates the processing of miR-484 primary transcript by targeting miR-361.

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Kun Wang

2014-07-01

Full Text Available Long noncoding RNAs (lncRNAs are emerging as new players in gene regulation, but whether lncRNAs operate in the processing of miRNA primary transcript is unclear. Also, whether lncRNAs are involved in the regulation of the mitochondrial network remains to be elucidated. Here, we report that a long noncoding RNA, named mitochondrial dynamic related lncRNA (MDRL, affects the processing of miR-484 primary transcript in nucleus and regulates the mitochondrial network by targeting miR-361 and miR-484. The results showed that miR-361 that predominantly located in nucleus can directly bind to primary transcript of miR-484 (pri-miR-484 and prevent its processing by Drosha into pre-miR-484. miR-361 is able to regulate mitochondrial fission and apoptosis by regulating miR-484 levels. In exploring the underlying molecular mechanism by which miR-361 is regulated, we identified MDRL and demonstrated that it could directly bind to miR-361 and downregulate its expression levels, which promotes the processing of pri-miR-484. MDRL inhibits mitochondrial fission and apoptosis by downregulating miR-361, which in turn relieves inhibition of miR-484 processing by miR-361. Our present study reveals a novel regulating model of mitochondrial fission program which is composed of MDRL, miR-361 and miR-484. Our work not only expands the function of the lncRNA pathway in gene regulation but also establishes a new mechanism for controlling miRNA expression.

Mir-452-3p: A Potential Tumor Promoter That Targets the CPEB3/EGFR Axis in Human Hepatocellular Carcinoma

Science.gov (United States)

Tang, Hui; Zhang, Jianwen; Yu, Zhenyu; Ye, Linsen; Li, Kun; Ding, Fan; Feng, Xiao

2017-01-01

Purpose: We proposed to investigate the effects of miR-452-3p on the proliferation and mobility of hepatocellular carcinoma (HCC) cells by targeting cytoplasmic polyadenylation element binding protein 3/estimated glomerular filtration rate (CPEB3/EGFR) axis. Methods: Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect miR-452-3p expression in 84 pairs of HCC tissues and adjacent tissues. Luciferase reporter assay was employed to examine the relationship between miR-452-3p and CPEB3. Microculture tetrazolium (MTT) assay, colony formation assay, flow cytometry detection, wound healing assay, and transwell assay were used to detect cell proliferation, cycle arrest, apoptosis, and mobility, respectively, in HCC, HepG2, and Huh-7. Western blot was used to detect protein expression levels in EGFR signaling pathway. Kaplan-Meier survival analysis was conducted to analyze the correlation between the miR-452-3p and CPEB3 expression levels and the survival of patients with HCC. Results: MiRNA-452-3p was found significantly upregulated in 84 human HCC sample tissues and cells in comparison with adjacent tissues and normal liver epithelial cells (P < .01). Luciferase reporter assay demonstrated that CPEB3 was a direct target of miR-452-3p. Overexpression of miR-452-3p promoted cell proliferation and mobility and suppressed apoptosis. MiR-452-3p enhanced EGFR and phosphorylated AKT (pAKT) expression but inhibited p21 expression level. Conclusion: MiR-452-3p promoted HCC cell proliferation and mobility by directly targeting the CPEB3/EGFR axis. PMID:29332449

Epigenetic modification of miR-10a regulates renal damage by targeting CREB1 in type 2 diabetes mellitus

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Shan, Qun, E-mail: shanp@jsnu.edu.cn; Zheng, Guihong, E-mail: ghzhengsd@jsnu.edu.cn; Zhu, Aihua, E-mail: ahzhu@jsnu.edu.cn; Cao, Li, E-mail: 948113717@qq.com; Lu, Jun, E-mail: lu-jun75@163.com; Wu, Dongmei, E-mail: wdm8610@jsnu.edu.cn; Zhang, ZiFeng, E-mail: zhangzifengsuper@jsnu.edu.cn; Fan, Shaohua, E-mail: fshfly@126.com; Sun, Chunhui, E-mail: 306484866@qq.com; Hu, Bin, E-mail: hubin@jsnu.edu.cn; Zheng, Yuanlin, E-mail: ylzheng@jsnu.edu.cn

2016-09-01

Emerging evidence has shown that microRNA-mediated gene expression modulation plays a crucial role in the pathogenesis of type 2 diabetes mellitus, but the novel miRNAs involved in type 2 diabetes and its functional regulatory mechanisms still need to be determined. In this study, we assessed the role of miR-10a in extracellular matrix accumulation in the kidney of diabetic mellitus induced by combining administration of chronic high fat diet (HFD) and low dosage of streptozotocin (STZ, 35 mg/kg). Here, we found that HFD/STZ administration decreased the level of microRNA (miR-10a) expression in ICR strain mice. Overexpression of miR-10a alleviated the increased ratio of urine albumin-to-creatinine (ACR) ratio of HFD/STZ mice. In contrast, knockdown of miR-10a increased the ratio of kidney ACR in naïve mice. Furthermore, cAMP response element binding protein 1 (CREB1) was validated as a target of miR-10a in vitro and in vivo. CREB1 and its downstream fibronectin (FN, extracellular matrix) were increased in HFD/STZ-treated mice, which was reversed by kidney miR-10a overexpression. The content of CREB1 and FN was increased by miR-10a knockdown in kidney of naïve mice. Furthermore, histone deacetylase 3 (HDAC3) was revealed to be increased in kidney of HFD/STZ mice, accompanied with the augmentation of ACR ratio and FN level. Knockdown of HDAC3 with siRNA significantly caused the increase of miR-10a, resulting in the decrease in CREB1 and FN expression in kidney of HFD/STZ mice. Contrarily, HDAC3 overexpression mediated by lentivirus decreased miR-10a content, and enhanced ACR value, CREB1 and FN formation in naïve mice. Collectively, these results elucidate that HDAC3/miR-10a/CREB1 serves as a new mechanism underlying kidney injury, providing potential therapeutic targets in type 2 diabetes. - Highlights: • Diabetes induces the decrease of miR-10a level in the kidney. • MiR-10a overexpression improves kidney damage of diabetes. • MiR-10a targeting CREB1/FN

Epigenetic modification of miR-10a regulates renal damage by targeting CREB1 in type 2 diabetes mellitus

International Nuclear Information System (INIS)

Shan, Qun; Zheng, Guihong; Zhu, Aihua; Cao, Li; Lu, Jun; Wu, Dongmei; Zhang, ZiFeng; Fan, Shaohua; Sun, Chunhui; Hu, Bin; Zheng, Yuanlin

2016-01-01

Emerging evidence has shown that microRNA-mediated gene expression modulation plays a crucial role in the pathogenesis of type 2 diabetes mellitus, but the novel miRNAs involved in type 2 diabetes and its functional regulatory mechanisms still need to be determined. In this study, we assessed the role of miR-10a in extracellular matrix accumulation in the kidney of diabetic mellitus induced by combining administration of chronic high fat diet (HFD) and low dosage of streptozotocin (STZ, 35 mg/kg). Here, we found that HFD/STZ administration decreased the level of microRNA (miR-10a) expression in ICR strain mice. Overexpression of miR-10a alleviated the increased ratio of urine albumin-to-creatinine (ACR) ratio of HFD/STZ mice. In contrast, knockdown of miR-10a increased the ratio of kidney ACR in naïve mice. Furthermore, cAMP response element binding protein 1 (CREB1) was validated as a target of miR-10a in vitro and in vivo. CREB1 and its downstream fibronectin (FN, extracellular matrix) were increased in HFD/STZ-treated mice, which was reversed by kidney miR-10a overexpression. The content of CREB1 and FN was increased by miR-10a knockdown in kidney of naïve mice. Furthermore, histone deacetylase 3 (HDAC3) was revealed to be increased in kidney of HFD/STZ mice, accompanied with the augmentation of ACR ratio and FN level. Knockdown of HDAC3 with siRNA significantly caused the increase of miR-10a, resulting in the decrease in CREB1 and FN expression in kidney of HFD/STZ mice. Contrarily, HDAC3 overexpression mediated by lentivirus decreased miR-10a content, and enhanced ACR value, CREB1 and FN formation in naïve mice. Collectively, these results elucidate that HDAC3/miR-10a/CREB1 serves as a new mechanism underlying kidney injury, providing potential therapeutic targets in type 2 diabetes. - Highlights: • Diabetes induces the decrease of miR-10a level in the kidney. • MiR-10a overexpression improves kidney damage of diabetes. • MiR-10a targeting CREB1/FN

NR2F2 inhibits Smad7 expression and promotes TGF-β-dependent epithelial-mesenchymal transition of CRC via transactivation of miR-21.

Science.gov (United States)

Wang, Hao; Nie, Lei; Wu, Lei; Liu, Qiufang; Guo, Xueyan

2017-03-25

Metastasis is one of the most decisive factors influencing CRC patient prognosis and current studies suggest that a molecular mechanism known as EMT broadly regulates cancer metastasis. NR2F2 is a key molecule in the development of CRC, but the roles and underlying mechanisms of NR2F2 in TGF-β induced EMT in CRC remain largely unknown. In the current study, we were interested to examine the role of NR2F2 in the TGF-β-induced EMT in CRC. Here, we found NR2F2 was upregulated in CRC cells and promotes TGF-β-induced EMT in CRC. Using comparative miRNA profiling TGF-β pre-treated CRC cells in which NR2F2 had been knocked down with that of control cells, we identified miR-21 as a commonly downregulated miRNA in HT29 cells treated with TGF-β and NR2F2 siRNA, and its downregulation inhibiting migration and invasion of CRC cells. Moreover, we found NR2F2 could transcriptional activated miR-21 expression by binding to miR-21 promoter in HT29 by ChIP and luciferase assay. In the last, our data demonstrated that Smad7 was the direct target of miR-21 in CRC cells. Thus, NR2F2 could promote TGF-β-induced EMT and inhibit Smad7 expression via transactivation of miR-21, and NR2F2 may be a new common therapeutic target for CRC. Copyright © 2017 Elsevier Inc. All rights reserved.

MiR-18a regulates the proliferation, migration and invasion of human glioblastoma cell by targeting neogenin

International Nuclear Information System (INIS)

Song, Yichen; Wang, Ping; Zhao, Wei; Yao, Yilong; Liu, Xiaobai; Ma, Jun; Xue, Yixue; Liu, Yunhui

2014-01-01

MiR-17-92 cluster has recently been reported as an oncogene in some tumors. However, the association of miR-18a, an important member of this cluster, with glioblastoma remains unknown. Therefore, this study aims to investigate the expression of miR-18a in glioblastoma and its role in biological behavior of U87 and U251 human glioblastoma cell lines. Quantitative RT-PCR results showed that miR-18a was highly expressed in glioblastoma tissues and U87 and U251 cell lines compared with that in human brain tissues and primary normal human astrocytes, and the expression levels were increased along with the rising pathological grades of glioblastoma. Neogenin was identified as the target gene of miR-18a by dual-luciferase reporter assays. RT-PCR and western blot results showed that its expression levels were decreased along with the rising pathological grades of glioblastoma. Inhibition of miR-18a expression was established by transfecting exogenous miR-18a inhibitor into U87 and U251 cells, and its effects on the biological behavior of glioblastoma cells were studied using CCK-8 assay, transwell assay and flow cytometry. Inhibition of miR-18a expression in U87 and U251 cells significantly up-regulated neogenin, and dramatically suppressed the abilities of cell proliferation, migration and invasion, induced cell cycle arrest and promoted cellular apoptosis. Collectively, these results suggest that miR-18a may regulate biological behavior of human glioblastoma cells by targeting neogenin, and miR-18a can serve as a potential target in the treatment of glioblastoma. - Highlights: • MiR-18a was highly expressed in glioblastoma tissues and U87 and U251 cell lines. • Neogenin was identified as the target gene of miR-18a. • Neogenin expressions were decreased along with the rising pathological grades of glioblastoma. • Inhibition of miR-18a suppressed biological behavior of glioma cells by up-regulating neogenin

MiR-18a regulates the proliferation, migration and invasion of human glioblastoma cell by targeting neogenin

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Song, Yichen, E-mail: jeff200064017@163.com [Department of Neurosurgery, Shengjing Hospital of China Medical University, Shenyang 110004 (China); Wang, Ping, E-mail: pingwang8000@163.com [Department of Neurobiology, College of Basic Medicine, China Medical University, Shenyang 110001 (China); Institute of Pathology and Pathophysiology, China Medical University, Shenyang 110001 (China); Zhao, Wei, E-mail: 15669746@qq.com [Department of Neurobiology, College of Basic Medicine, China Medical University, Shenyang 110001 (China); Institute of Pathology and Pathophysiology, China Medical University, Shenyang 110001 (China); Yao, Yilong, E-mail: yaoyilong_322@163.com [Department of Neurosurgery, Shengjing Hospital of China Medical University, Shenyang 110004 (China); Liu, Xiaobai, E-mail: paganizonda1991@qq.com [The 96th Class, 7-year Program, China Medical University, Shenyang, Liaoning Province 110001 (China); Ma, Jun, E-mail: majun_724@163.com [Department of Neurobiology, College of Basic Medicine, China Medical University, Shenyang 110001 (China); Institute of Pathology and Pathophysiology, China Medical University, Shenyang 110001 (China); Xue, Yixue, E-mail: xueyixue888@163.com [Department of Neurobiology, College of Basic Medicine, China Medical University, Shenyang 110001 (China); Institute of Pathology and Pathophysiology, China Medical University, Shenyang 110001 (China); Liu, Yunhui, E-mail: liuyh@sj-hospital.org [Department of Neurosurgery, Shengjing Hospital of China Medical University, Shenyang 110004 (China)

2014-05-15

MiR-17-92 cluster has recently been reported as an oncogene in some tumors. However, the association of miR-18a, an important member of this cluster, with glioblastoma remains unknown. Therefore, this study aims to investigate the expression of miR-18a in glioblastoma and its role in biological behavior of U87 and U251 human glioblastoma cell lines. Quantitative RT-PCR results showed that miR-18a was highly expressed in glioblastoma tissues and U87 and U251 cell lines compared with that in human brain tissues and primary normal human astrocytes, and the expression levels were increased along with the rising pathological grades of glioblastoma. Neogenin was identified as the target gene of miR-18a by dual-luciferase reporter assays. RT-PCR and western blot results showed that its expression levels were decreased along with the rising pathological grades of glioblastoma. Inhibition of miR-18a expression was established by transfecting exogenous miR-18a inhibitor into U87 and U251 cells, and its effects on the biological behavior of glioblastoma cells were studied using CCK-8 assay, transwell assay and flow cytometry. Inhibition of miR-18a expression in U87 and U251 cells significantly up-regulated neogenin, and dramatically suppressed the abilities of cell proliferation, migration and invasion, induced cell cycle arrest and promoted cellular apoptosis. Collectively, these results suggest that miR-18a may regulate biological behavior of human glioblastoma cells by targeting neogenin, and miR-18a can serve as a potential target in the treatment of glioblastoma. - Highlights: • MiR-18a was highly expressed in glioblastoma tissues and U87 and U251 cell lines. • Neogenin was identified as the target gene of miR-18a. • Neogenin expressions were decreased along with the rising pathological grades of glioblastoma. • Inhibition of miR-18a suppressed biological behavior of glioma cells by up-regulating neogenin.

MiR-142-3p Functions as a Potential Tumor Suppressor in Human Osteosarcoma by Targeting HMGA1

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Guoxing Xu

2014-04-01

Full Text Available Background/Aims: Mounting evidence has shown that aberrant expression of miRNAs correlates with human cancers, and that miRNAs can function as tumor suppressors or oncogenes. Here, we investigated the role and mechanism of miR-142-3p in human osteosarcoma. Methods: We used quantitative real-time RT-PCR to measure the expression of miR-142-3p in human osteosarcoma cell lines and tissues. The roles of miR-142-3p in osteosarcoma development were studied using cultured HOS, MG63 and Saos-2 cells and tumor xenograft analyses in nude mice; their target genes were also investigated. Results: We found that miR-142-3p was significantly downregulated in osteosarcoma cell lines and clinical specimens. Overexpression of miR-142-3p suppressed osteosarcoma cell proliferation, migration and invasion, whereas miR-142-3p knockdown increased these parameters. The xenograft mouse model also revealed the suppressive effect of miR-142-3p on tumor growth. High mobility group AT-hook 1 (HMGA1 was identified as a target of miR-142-3p. Downregulation of HMGA1 induced effects on osteosarcoma cell lines similar to those induced by miR-142-3p. In contrast, restoration of HMGA1 abrogated the effects induced by miR-142-3p up-regulation. Conclusion: These results indicated that miR-142-3p may function as a tumor suppressor by targeting HMGA1 in osteosarcoma.

Urtica dioica extract suppresses miR-21 and metastasis-related genes in breast cancer.

Science.gov (United States)

Mansoori, Behzad; Mohammadi, Ali; Hashemzadeh, Shahriar; Shirjang, Solmaz; Baradaran, Ali; Asadi, Milad; Doustvandi, Mohammad Amin; Baradaran, Behzad

2017-09-01

Breast cancer has a high prevalence among women worldwide. Tumor invasion and metastasis still remains an open issue that causes most of the therapeutic failures and remains the prime cause of patient mortality. Hence, there is an unmet need to develop the most effective therapeutic approach with the lowest side effects and highest cytotoxicity that will effectively arrest or eradicate metastasis. An MTT assay and scratch test were used to assess the cytotoxicity and migration effects of Urtica dioica on the breast cancer cells. The QRT-PCR was used to study the expression levels of miR-21, MMP1, MMP9, MMP13, CXCR4, vimentin, and E-cadherin. The results of gene expression in tumoral groups confirmed the overexpression of miR-21, MMP1, MMP9, MMP13, vimentin, and CXCR4, and the lower expression of E-cadherin compared to control groups (PUrtica dioica significantly inhibited breast cancer cell proliferation. Moreover, findings from the scratch assay exhibited the inhibitory effects of Urtica dioica on the migration of breast cancer cell lines. Urtica dioica extract could inhibit cancer cell migration by regulating miR-21, MMP1, MMP9, MMP13, vimentin, CXCR4, and E-Cadherin. Moreover, our findings demonstrated that the extract could decrease miR-21 expression, which substantially lessens the overexpressed MMP1, MMP9, MMP13, vimentin, and CXCR4 and increases E-cadherin in the tumoral group. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

The miRNA Pull Out Assay as a Method to Validate the miR-28-5p Targets Identified in Other Tumor Contexts in Prostate Cancer

DEFF Research Database (Denmark)

Rizzo, Milena; Berti, Gabriele; Russo, Francesco

2017-01-01

targets in the pull out sample. We showed that E2F6, TEX-261, MAPK1, MPL, N4BP1, and RAP1B but not BAG1, OTUB1, MAD2L1, and p21 were significantly enriched, suggesting that not all the miR-28-5p targets are regulated by this miRNA in PCa. We then verified whether the miR-28-5p-interacting targets were...

Prognostic significance of miR-205 in endometrial cancer.

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Mihriban Karaayvaz

Full Text Available microRNAs have emerged as key regulators of gene expression, and their altered expression has been associated with tumorigenesis and tumor progression. Thus, microRNAs have potential as both cancer biomarkers and/or potential novel therapeutic targets. Although accumulating evidence suggests the role of aberrant microRNA expression in endometrial carcinogenesis, there are still limited data available about the prognostic significance of microRNAs in endometrial cancer. The goal of this study is to investigate the prognostic value of selected key microRNAs in endometrial cancer by the analysis of archival formalin-fixed paraffin-embedded tissues.Total RNAs were extracted from 48 paired normal and endometrial tumor specimens using Trizol based approach. The expression of miR-26a, let-7g, miR-21, miR-181b, miR-200c, miR-192, miR-215, miR-200c, and miR-205 were quantified by real time qRT-PCR expression analysis. Targets of the differentially expressed miRNAs were quantified using immunohistochemistry. Statistical analysis was performed by GraphPad Prism 5.0.The expression levels of miR-200c (P<0.0001 and miR-205 (P<0.0001 were significantly increased in endometrial tumors compared to normal tissues. Kaplan-Meier survival analysis revealed that high levels of miR-205 expression were associated with poor patient overall survival (hazard ratio, 0.377; Logrank test, P = 0.028. Furthermore, decreased expression of a miR-205 target PTEN was detected in endometrial cancer tissues compared to normal tissues.miR-205 holds a unique potential as a prognostic biomarker in endometrial cancer.

Inhibition of miR-155, a therapeutic target for breast cancer, prevented in cancer stem cell formation.

Science.gov (United States)

Zuo, Jiangcheng; Yu, Yalan; Zhu, Man; Jing, Wei; Yu, Mingxia; Chai, Hongyan; Liang, Chunzi; Tu, Jiancheng

2018-02-06

Breast cancer is a common cancer in women of worldwide. Cancer cells with stem-like properties played important roles in breast cancer, such as relapse, metastasis and treatment resistance. Micro-RNA-155 (miR-155) is a well-known oncogenic miRNA overexpressed in many human cancers. The expression levels of miR-155 in 38 pairs of cancer tissues and adjacent normal tissues from breast cancer patients were detected using quantitative real-time PCR. The invasive cell line MDA-MB-231 was used to quantify the expression of miR-155 by tumor-sphere forming experiment. Soft agar colony formation assay and tumor xenografts was used to explore whether the inhibition of miR-155 could reduce proliferation of cancer cells in vivo and vitro. In the study, we found miR-155 was upregulated in BC. Soft agar colony formation assay and tumor xenografts showed inhibition of miR-155 could significantly reduce proliferation of cancer cells in vivo and vitro, which confirmed that miR-155 is an effective therapeutic target of breast cancer. Sphere-forming experiment showed that overexpression of miR-155 significantly correlated with stem-like properties. Expressions of ABCG2, CD44 and CD90 were repressed by inhibition of miR-155, but CD24 was promoted. Interestingly, inhibition of miR-155 rendered MDA-MB-231 cells more sensitive to Doxorubicinol, which resulted in an increase of inhibition rate from 20.23% to 68.72%. Expression of miR-155 not only was a therapeutic target but also was associated with cancer stem cell formation and Doxorubicinol sensitivity. Our results underscore the importance of miR-155 as a therapeutic target and combination of Doxorubicinol and miR-155-silencing would be a potential way to cure breast cancer.

miR-203a is involved in HBx-induced inflammation by targeting Rap1a

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Wu, AiRong [Department of gastroenterology, The First affiliated Hospital of Soochow University, Suzhou 215006 (China); Chen, Huo [Institutes of Biology and Medical Sciences, Soochow University, Suzhou 215123 (China); Xu, ChunFang [Department of gastroenterology, The First affiliated Hospital of Soochow University, Suzhou 215006 (China); Zhou, Ji; Chen, Si [Institutes of Biology and Medical Sciences, Soochow University, Suzhou 215123 (China); Shi, YuQi [Department of gastroenterology, The First affiliated Hospital of Soochow University, Suzhou 215006 (China); Xu, Jie [Institutes of Biology and Medical Sciences, Soochow University, Suzhou 215123 (China); Gan, JianHe, E-mail: j_pzhang@suda.edu.cn [Department of gastroenterology, The First affiliated Hospital of Soochow University, Suzhou 215006 (China); Zhang, JinPing, E-mail: ganjianhe@aliyun.com [Institutes of Biology and Medical Sciences, Soochow University, Suzhou 215123 (China)

2016-11-15

Hepatitis B virus (HBV) causes acute and chronic hepatitis, and is one of the major causes of cirrhosis and hepatocellular carcinoma. Accumulating evidence suggests that inflammation is the key factor for liver cirrhosis and hepatocellular carcinoma. MicroRNAs play important roles in many biological processes. Here, we aim to explore the function of microRNAs in the HBX-induced inflammation. First, microarray experiment showed that HBV{sup +} liver samples expressed higher level of miR-203a compared to HBV{sup -} liver samples. To verify these alterations, HBx-coding plasmid was transfected into HepG2 cells to overexpress HBx protein. The real-time PCR results suggested that over-expression of HBx could induce up-regulation of miR-203a. To define how up-regulation of miR-203a can induce liver cells inflammation, we over-expressed miR-203a in HepG2 cells. Annexin V staining and BrdU staining suggested that overexpression of miR-203a significantly increased the cell apoptosis and proliferation, meanwhile, over-expression of miR-203a could lead to a decrease in G0/G1 phase cells and an increase in G2/M phase cells. Some cytokines production including IL-6 and IL-8 were significantly increased, but TGFβ and IFNγ were decreased in miR-203a over-expressed HepG2 cells. Luciferase reporter assay experiments, protein mass-spectrum assay and real-time PCR all together demonstrated that Rap1a was the target gene of miR-203a. Further experiments showed that these alterations were modulated through PI3K/ERK/p38/NFκB pathways. These data suggested that HBV-infection could up-regulate the expression of miR-203a, thus down regulated the expression of Rap1a and affected the PI3K/ERK/p38/NFκB pathways, finally induced the hepatitis inflammation. - Highlights: • HBX induces the over-expression of miR-203a in HepG2 cells. • miR-203a targets Rap1a to induce the inflammation in HepG2 cells. • miR-203a regulates the apoptosis and cell cycles of HepG2 cells. • miR-203a alters

miR-221 and miR-222 expression affects the proliferation potential of human prostate carcinoma cell lines by targeting p27Kip1.

Science.gov (United States)

Galardi, Silvia; Mercatelli, Neri; Giorda, Ezio; Massalini, Simone; Frajese, Giovanni Vanni; Ciafrè, Silvia Anna; Farace, Maria Giulia

2007-08-10

MicroRNAs are short regulatory RNAs that negatively modulate protein expression at a post-transcriptional level and are deeply involved in the pathogenesis of several types of cancers. Here we show that miR-221 and miR-222, encoded in tandem on chromosome X, are overexpressed in the PC3 cellular model of aggressive prostate carcinoma, as compared with LNCaP and 22Rv1 cell line models of slowly growing carcinomas. In all cell lines tested, we show an inverse relationship between the expression of miR-221 and miR-222 and the cell cycle inhibitor p27(Kip1). We recognize two target sites for the microRNAs in the 3' untranslated region of p27 mRNA, and we show that miR-221/222 ectopic overexpression directly results in p27 down-regulation in LNCaP cells. In those cells, we demonstrate that the ectopic overexpression of miR-221/222 strongly affects their growth potential by inducing a G(1) to S shift in the cell cycle and is sufficient to induce a powerful enhancement of their colony-forming potential in soft agar. Consistently, miR-221 and miR-222 knock-down through antisense LNA oligonucleotides increases p27(Kip1) in PC3 cells and strongly reduces their clonogenicity in vitro. Our results suggest that miR-221/222 can be regarded as a new family of oncogenes, directly targeting the tumor suppressor p27(Kip1), and that their overexpression might be one of the factors contributing to the oncogenesis and progression of prostate carcinoma through p27(Kip1) down-regulation.

miR-106a-5p inhibits the proliferation and migration of astrocytoma cells and promotes apoptosis by targeting FASTK.

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Feng Zhi

Full Text Available Astrocytomas are common malignant intracranial tumors that comprise the majority of adult primary central nervous system tumors. MicroRNAs (miRNAs are small, non-coding RNAs (20-24 nucleotides that post-transcriptionally modulate gene expression by negatively regulating the stability or translational efficiency of their target mRNAs. In our previous studies, we found that the downregulation of miR-106a-5p in astrocytomas is associated with poor prognosis. However, its specific gene target(s and underlying functional mechanism(s in astrocytomas remain unclear. In this study, we used mRNA microarray experiments to measure global mRNA expression in the presence of increased or decreased miR-106a-5p levels. We then performed bioinformatics analysis based on multiple target prediction algorithms to obtain candidate target genes that were further validated by computational predictions, western blot analysis, quantitative real-time PCR, and the luciferase reporter assay. Fas-activated serine/threonine kinase (FASTK was identified as a direct target of miR-106a-5p. In human astrocytomas, miR-106a-5p is downregulated and negatively associated with clinical staging, whereas FASTK is upregulated and positively associated with advanced clinical stages, at both the protein and mRNA levels. Furthermore, Kaplan-Meier analysis revealed that the reduced expression of miR-106a-5p or the increased expression of FASTK is significantly associated with poor survival outcome. These results further supported the finding that FASTK is a direct target gene of miR-106a-5p. Next, we explored the function of miR-106a-5p and FASTK during astrocytoma progression. Through gain-of-function and loss-of-function studies, we demonstrated that miR-106a-5p can significantly inhibit cell proliferation and migration and can promote cell apoptosis in vitro. The knockdown of FASTK induced similar effects on astrocytoma cells as those induced by the overexpression of miR-106a-5p. These

The miRNA Pull Out Assay as a Method to Validate the miR-28-5p Targets Identified in Other Tumor Contexts in Prostate Cancer

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Milena Rizzo

2017-01-01

Full Text Available miR-28-5p is an intragenic miRNA which is underexpressed in several tumor types showing a tumor suppressor (TS activity. Routinely, the known miR-28-5p targets are validated in specific tumor contexts but it is unclear whether these targets are also being regulated in other tumor types. To this end, we adopted the miRNA pull out assay to capture the miR-28-5p targets in DU-145 prostate cancer (PCa cells. Firstly, we demonstrated that miR-28-5p acts as a TS-miRNA in PCa, affecting cell proliferation, survival, and apoptosis. Secondly, we evaluated the enrichment of the 10 validated miR-28-5p targets in the pull out sample. We showed that E2F6, TEX-261, MAPK1, MPL, N4BP1, and RAP1B but not BAG1, OTUB1, MAD2L1, and p21 were significantly enriched, suggesting that not all the miR-28-5p targets are regulated by this miRNA in PCa. We then verified whether the miR-28-5p-interacting targets were regulated by this miRNA. We selected E2F6, the most enriched target in the pull out sample, and demonstrated that miR-28-5p downregulated E2F6 at the protein level suggesting that our approach was effective. In general terms, these findings support the miRNA pull out assay as a useful method to identify context-specific miRNA targets.

MiR-637 maintains the balance between adipocytes and osteoblasts by directly targeting Osterix.

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Zhang, Jin-fang; Fu, Wei-ming; He, Ming-liang; Wang, Hua; Wang, Wei-mao; Yu, Shi-cang; Bian, Xiu-Wu; Zhou, Jin; Lin, Marie C M; Lu, Gang; Poon, Wai-sang; Kung, Hsiang-fu

2011-11-01

Bone development is dynamically regulated by homeostasis, in which a balance between adipocytes and osteoblasts is maintained. Disruption of this differentiation balance leads to various bone-related metabolic diseases, including osteoporosis. In the present study, a primate-specific microRNA (miR-637) was found to be involved in the differentiation of human mesenchymal stem cells (hMSCs). Our preliminary data indicated that miR-637 suppressed the growth of hMSCs and induced S-phase arrest. Expression of miR-637 was increased during adipocyte differentiation (AD), whereas it was decreased during osteoblast differentiation (OS), which suggests miR-637 could act as a mediator of adipoosteogenic differentiation. Osterix (Osx), a significant transcription factor of osteoblasts, was shown to be a direct target of miR-637, which significantly enhanced AD and suppressed OS in hMSCs through direct suppression of Osx expression. Furthermore, miR-637 also significantly enhanced de novo adipogenesis in nude mice. In conclusion, our data indicated that the expression of miR-637 was indispensable for maintaining the balance of adipocytes and osteoblasts. Disruption of miR-637 expression patterns leads to irreversible damage to the balance of differentiation in bone marrow.

Thermo-chemotherapy Induced miR-218 upregulation inhibits the invasion of gastric cancer via targeting Gli2 and E-cadherin.

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Ruan, Qiang; Fang, Zhi-Yuan; Cui, Shu-Zhong; Zhang, Xiang-Liang; Wu, Yin-Bing; Tang, Hong-Sheng; Tu, Yi-Nuo; Ding, Yan

2015-08-01

Thermo-chemotherapy has been proven to reduce the invasion capability of cancer cells. However, the molecular mechanism underlying this anti-invasion effect is still unclear. In this study, the role of thermo-chemotherapy in the inhibition of tumor invasion was studied. The results demonstrated that expression of miR-218 was downregulated in gastric cancer tissues, which had a positive correlation with tumor invasion and metastasis. In vitro thermo-chemotherapy increased miR-218 expression in SGC7901 cells and inhibited both proliferation and invasion of cancer cells. Gli2 was identified as a downstream target of miR-218, and its expression was negatively regulated by miR-218. The thermo-chemotherapy induced miR-218 upregulation was also accompanied by increasing of E-cadherin expression. In conclusion, the present study indicates that thermo-chemotherapy can effectively decrease the invasion capability of cancer cells and increase cell-cell adhesion. miR-218 and its downstream target Gli2, as well as E-cadherin, participate in the anti-invasion process.

Dual Role of miR-21 in CD4+T-Cells : Activation-Induced miR-21 Supports Survival of Memory T-Cells and Regulates CCR7 Expression in Naive T-Cells

NARCIS (Netherlands)

Smigielska-Czepiel, Katarzyna; van den Berg, Anke; Jellema, Pytrick; Slezak-Prochazka, Izabella; Maat, Henny; van den Bos, Hilda; van der Lei, Roelof Jan; Kluiver, Joost; Brouwer, Elisabeth; Boots, Anne Mieke H.; Kroesen, Bart-Jan

2013-01-01

Immune cell-type specific miRNA expression patterns have been described but the detailed role of single miRNAs in the function of T-cells remains largely unknown. We investigated the role of miR-21 in the function of primary human CD4+ T-cells. MiR-21 is substantially expressed in T-cells with a

MiR-124 suppresses cell proliferation in hepatocellular carcinoma by targeting PIK3CA

International Nuclear Information System (INIS)

Lang, Qingbo; Ling, Changquan

2012-01-01

Highlights: ► PIK3CA is a novel target of miR-124 in HepG2 cells. ► MiR-124 suppresses cell proliferation by downregulating PIK3CA expression. ► MiR-124 regulates the PI3K/Akt pathway in HepG2 cells. ► MiR-124 overexpression inhibits the tumorigenesis in nude mice. -- Abstract: MicroRNAs (miRNAs) have crucial roles in the development and progression of human cancers, including hepatocellular carcinoma (HCC). Recent studies have shown that microRNA-124 (miR-124) was downregulated in HCC; however, the underlying mechanisms by which miR-124 suppresses tumorigenesis in HCC are largely unknown. In this study, we report that phosphoinositide 3-kinase catalytic subunit alpha (PIK3CA) is a novel target of miR-124 in HepG2 cells. Overexpression of miR-124 resulted in decreased expression of PIK3CA at both mRNA and protein levels. We found that miR-124 overexpression markedly suppressed cell proliferation by inducing G1-phase cell-cycle arrest in vitro. Consistent with the restoring miR-124 expression, PIK3CA knockdown suppressed cell proliferation, whereas overexpression of PIK3CA abolished the suppressive effect of miR-124. Mechanistic studies showed that miR-124-mediated reduction of PIK3CA resulted in suppression of PI3K/Akt pathway. The expressions of Akt and mTOR, key components of the PI3K/Akt pathway, were all downregulated. Moreover, we found overexpressed miR-124 effectively repressed tumor growth in xenograft animal experiments. Taken together, our results demonstrate that miR-124 functions as a growth-suppressive miRNA and plays an important role in inhibiting the tumorigenesis through targeting PIK3CA.

MiR-124 suppresses cell proliferation in hepatocellular carcinoma by targeting PIK3CA

Energy Technology Data Exchange (ETDEWEB)

Lang, Qingbo [Department of Traditional Chinese Medicine, Changhai Hospital, Second Military Medical University, Shanghai 200433 (China); Ling, Changquan, E-mail: lingchangquan@hotmail.com [Department of Traditional Chinese Medicine, Changhai Hospital, Second Military Medical University, Shanghai 200433 (China)

2012-09-21

Highlights: Black-Right-Pointing-Pointer PIK3CA is a novel target of miR-124 in HepG2 cells. Black-Right-Pointing-Pointer MiR-124 suppresses cell proliferation by downregulating PIK3CA expression. Black-Right-Pointing-Pointer MiR-124 regulates the PI3K/Akt pathway in HepG2 cells. Black-Right-Pointing-Pointer MiR-124 overexpression inhibits the tumorigenesis in nude mice. -- Abstract: MicroRNAs (miRNAs) have crucial roles in the development and progression of human cancers, including hepatocellular carcinoma (HCC). Recent studies have shown that microRNA-124 (miR-124) was downregulated in HCC; however, the underlying mechanisms by which miR-124 suppresses tumorigenesis in HCC are largely unknown. In this study, we report that phosphoinositide 3-kinase catalytic subunit alpha (PIK3CA) is a novel target of miR-124 in HepG2 cells. Overexpression of miR-124 resulted in decreased expression of PIK3CA at both mRNA and protein levels. We found that miR-124 overexpression markedly suppressed cell proliferation by inducing G1-phase cell-cycle arrest in vitro. Consistent with the restoring miR-124 expression, PIK3CA knockdown suppressed cell proliferation, whereas overexpression of PIK3CA abolished the suppressive effect of miR-124. Mechanistic studies showed that miR-124-mediated reduction of PIK3CA resulted in suppression of PI3K/Akt pathway. The expressions of Akt and mTOR, key components of the PI3K/Akt pathway, were all downregulated. Moreover, we found overexpressed miR-124 effectively repressed tumor growth in xenograft animal experiments. Taken together, our results demonstrate that miR-124 functions as a growth-suppressive miRNA and plays an important role in inhibiting the tumorigenesis through targeting PIK3CA.

miR-758-5p regulates cholesterol uptake via targeting the CD36 3'UTR.

Science.gov (United States)

Li, Bi-Rong; Xia, Lin-Qin; Liu, Jing; Liao, Lin-Ling; Zhang, Yang; Deng, Min; Zhong, Hui-Juan; Feng, Ting-Ting; He, Ping-Ping; Ouyang, Xin-Ping

2017-12-09

miR-758-3p plays an important role via regulting ABCA1-mediated cholesterol efflux in atherosclerosis. However, the mechanism of miR-758-5p in cholesterol metabolism is still unclear. Here, we revealed that miR-758-5p decreased total cholesterol accumulation in THP-1 macrophage derived foam cells through markedly reducing cholesterol uptake, and no effect on the cholesterol efflux. Interestingly, computational analysis suggests that CD36 may be a target gene of miR-758-5p. Our study further demonstrated that miR-758-5p decreased CD36 expression at both protein and mRNA levels via targeting the CD36 3'UTR in THP-1 macrophage derived foam cells. The present present study concluded that miR-758-5p decreases lipid accumulation of foam cell via regulating CD36-mediated the cholesterol uptake. Therefore, targeting miR-758-5p may offer a promising strategy to treat atherosclerotic vascular disease. Copyright © 2017. Published by Elsevier Inc.

miR-26a suppresses autophagy in swine Sertoli cells by targeting ULK2.

Science.gov (United States)

Ran, M; Li, Z; Cao, R; Weng, B; Peng, F; He, C; Chen, B

2018-05-14

A large number of microRNAs (miRNAs) have been detected from porcine testicular tissues thanks to the development of high-throughput sequencing technology. However, the regulatory roles of most identified miRNAs in swine testicular development or spermatogenesis are poorly understood. In our previous study, ULK2 (uncoordinated-51-like kinase 2) was predicted as a target gene of miR-26a. In this study, we aimed to investigate the role of miR-26a in swine Sertoli cell autophagy. The relative expression of miR-26a and ULK2 levels has a significant negative correlation (R 2  = .5964, p ≤ .01) in nine developmental stages of swine testicular tissue. Dual-luciferase reporter assay results show that miR-26a directly targets the 3'UTR of the ULK2 gene (position 618-624). In addition, both the mRNA and protein expression of ULK2 were downregulated by miR-26a in swine Sertoli cells. These results indicate that miR-26a targets the ULK2 gene and downregulates its expression in swine Sertoli cells. Based on the expression of marker genes (LC3, p62 and Beclin-1), overexpression of miR-26a or knock-down of ULK2 inhibits swine Sertoli cell autophagy. Taken together, these findings demonstrate that miR-26a suppresses autophagy in swine Sertoli cells by targeting ULK2. © 2018 Blackwell Verlag GmbH.

miR-31 Regulates Spermatogonial Stem Cells Meiosis via Targeting Stra8.

Science.gov (United States)

Wang, Yingjie; Zuo, Qisheng; Bi, Yulin; Zhang, Wenhui; Jin, Jing; Zhang, Liangliang; Zhang, Ya-Ni; Li, Bichun

2017-12-01

Stra8 (stimulated by retinoic acid gene 8) is a specific gene that is expressed in mammalian germ cells during transition from mitosis to meiosis and plays a key role in the initiation of meiosis in mammals and birds. So, the evaluation of the Stra8 pathway in cSSCs may provide a deeper insight into mammalian spermatogenesis. miRNA was also an important regulating factor for meiosis of SSCs. However, there is currently no data indicating that miRNA regulate the meiosis of SSCs via Stra8. Here, we predicted the prospective miRNA targeting to Stra8 using the online Bioinformatics database-Targetscan, and performed an analysis of the dual-luciferase recombinant vector, pGL3-CMV-LUC-MCS-Stra8-3'UTR. miR-31 mimics (miR-31m), miR-31 inhibitors (miR-31i), Control (NC, scrambled oligonucleotides transfection) were transfected into cSSCs; Stra8 and miRNA were analyzed by RT-qPCR, immunofluorescence, and Western blot. The detection of haploid was conducted by flow cytometry. The results showed that miR-31 regulates meiosis of cSSCs via targeting Stra8 in vitro and in vivo. Our study identifies a new regulatory pathway that miR-31 targets Stra8 and inhibits spermatogenesis. J. Cell. Biochem. 118: 4844-4853, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

NFkappaB activation is essential for miR-21 induction by TGFβ1 in high glucose conditions

International Nuclear Information System (INIS)

Madhyastha, Radha; Madhyastha, HarishKumar; Pengjam, Yutthana; Nakajima, Yuichi; Omura, Sayuri; Maruyama, Masugi

2014-01-01

Highlights: • Transforming growth factor beta 1 (TGFβ1) induces miR-21 in high glucose conditions. • NFkappaB activation and subsequent ROS generation are necessary for TGFβ1’s effect. • TGFβ1 facilitates binding of NFkB p65 to miR-21 promoter. • SMAD proteins bind to R-SBE sites on primary miR-21, in NFkB dependent manner. - Abstract: Transforming growth factor beta1 (TGFβ1) is a pleiotropic growth factor with a very broad spectrum of effects on wound healing. Chronic non-healing wounds such as diabetic foot ulcers express reduced levels of TGFβ1. On the other hand, our previous studies have shown that the microRNA miR-21 is differentially regulated in diabetic wounds and that it promotes migration of fibroblast cells. Although interplay between TGFβ1 and miR-21 are studied in relation to cancer, their interaction in the context of chronic wounds has not yet been investigated. In this study, we examined if TGFβ1 could stimulate miR-21 in fibroblasts that are subjected to high glucose environment. MiR-21 was, in fact, induced by TGFβ1 in high glucose conditions. The induction by TGFβ1 was dependent on NFκB activation and subsequent ROS generation. TGFβ1 was instrumental in degrading the NFκB inhibitor IκBα and facilitating the nuclear translocation of NFκB p65 subunit. EMSA studies showed enhanced DNA binding activity of NFκB in the presence of TGFβ1. ChIP assay revealed binding of p65 to miR-21 promoter. NFκB activation was also required for the nuclear translocation of Smad 4 protein and subsequent direct interaction of Smad proteins with primary miR-21 as revealed by RNA-IP studies. Our results show that manipulation of TGFβ1–NFκB–miR-21 pathway could serve as an innovative approach towards therapeutics to heal diabetic ulcers

NFkappaB activation is essential for miR-21 induction by TGFβ1 in high glucose conditions

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Madhyastha, Radha, E-mail: radharao@med.miyazaki-u.ac.jp; Madhyastha, HarishKumar; Pengjam, Yutthana; Nakajima, Yuichi; Omura, Sayuri; Maruyama, Masugi

2014-09-05

Highlights: • Transforming growth factor beta 1 (TGFβ1) induces miR-21 in high glucose conditions. • NFkappaB activation and subsequent ROS generation are necessary for TGFβ1’s effect. • TGFβ1 facilitates binding of NFkB p65 to miR-21 promoter. • SMAD proteins bind to R-SBE sites on primary miR-21, in NFkB dependent manner. - Abstract: Transforming growth factor beta1 (TGFβ1) is a pleiotropic growth factor with a very broad spectrum of effects on wound healing. Chronic non-healing wounds such as diabetic foot ulcers express reduced levels of TGFβ1. On the other hand, our previous studies have shown that the microRNA miR-21 is differentially regulated in diabetic wounds and that it promotes migration of fibroblast cells. Although interplay between TGFβ1 and miR-21 are studied in relation to cancer, their interaction in the context of chronic wounds has not yet been investigated. In this study, we examined if TGFβ1 could stimulate miR-21 in fibroblasts that are subjected to high glucose environment. MiR-21 was, in fact, induced by TGFβ1 in high glucose conditions. The induction by TGFβ1 was dependent on NFκB activation and subsequent ROS generation. TGFβ1 was instrumental in degrading the NFκB inhibitor IκBα and facilitating the nuclear translocation of NFκB p65 subunit. EMSA studies showed enhanced DNA binding activity of NFκB in the presence of TGFβ1. ChIP assay revealed binding of p65 to miR-21 promoter. NFκB activation was also required for the nuclear translocation of Smad 4 protein and subsequent direct interaction of Smad proteins with primary miR-21 as revealed by RNA-IP studies. Our results show that manipulation of TGFβ1–NFκB–miR-21 pathway could serve as an innovative approach towards therapeutics to heal diabetic ulcers.

miR-151-3p Targets TWIST1 to Repress Migration of Human Breast Cancer Cells.

Directory of Open Access Journals (Sweden)

Ting-Chih Yeh

Full Text Available TWIST1 is a highly conserved basic helix-loop-helix transcription factor that contributes to cancer metastasis by promoting an epithelial-mesenchymal transition and repressing E-cadherin gene expression in breast cancer. In this study, we explored the potential role of miR-151 in TWIST1 expression and cancer properties in human breast cancer cells. We found that the human TWIST1 3'UTR contains a potential binging site for miR-151-3p at the putative target sequence 5'-CAGUCUAG-3'. Using a TWIST1-3'UTR luciferase reporter assay, we demonstrated that the target sequence within the TWIST1 3'UTR is required for miR-151-3p regulation of TWIST1 expression. Moreover, we found that ectopic expression of miR-151-3p by infection with adenoviruses expressing miR-151 significantly decreased TWIST1 expression, migration and invasion, but did not affect cell growth and tumorsphere formation of human breast cancer cells. In addition, overexpression of the protein coding region without the 3'UTR of TWIST1 reversed the repression of cell migration by miR-151-3p. Furthermore, knockdown of miR-151-3p increased TWIST1 expression, reduced E-cadherin expression, and enhanced cell migration. In conclusion, these results suggest that miR-151-3p directly regulates TWIST1 expression by targeting the TWIST1 3'UTR and thus repressing the migration and invasion of human breast cancer cells by enhancing E-cadherin expression. Our findings add to accumulating evidence that microRNAs are involved in breast cancer progression by modulating TWIST1 expression.

A Double Negative Loop Comprising ETV6/RUNX1 and MIR181A1 Contributes to Differentiation Block in t(12;21)-Positive Acute Lymphoblastic Leukemia.

Science.gov (United States)

Yang, Yung-Li; Yen, Ching-Tzu; Pai, Chen-Hsueh; Chen, Hsuan-Yu; Yu, Sung-Liang; Lin, Chien-Yu; Hu, Chung-Yi; Jou, Shiann-Tarng; Lin, Dong-Tsamn; Lin, Shu-Rung; Lin, Shu-Wha

2015-01-01

Childhood acute lymphoblastic leukemia (ALL) with t(12;21), which results in expression of the ETV6/RUNX1 fusion gene, is the most common chromosomal lesion in precursor-B (pre-B) ALL. We identified 17 microRNAs that were downregulated in ETV6/RUNX1+ compared with ETV6/RUNX1- clinical samples. Among these microRNAs, miR-181a-1 was the most significantly reduced (by ~75%; P regulatory region of MIR181A1, and knockdown of ETV6/RUNX1 increased miR-181a-1 level. We further showed that miR-181a (functional counterpart of miR-181a-1) could target ETV6/RUNX1 and cause a reduction in the level of the oncoprotein ETV6/RUNX1, cell growth arrest, an increase in apoptosis, and induction of cell differentiation in ETV6/RUNX1+ cell line. Moreover, ectopic expression of miR-181a also resulted in decreased CD10 hyperexpression in ETV6/RUNX1+ primary patient samples. Taken together, our results demonstrate that MIR181A1 and ETV6/RUNX1 regulate each other, and we propose that a double negative loop involving MIR181A1 and ETV6/RUNX1 may contribute to ETV6/RUNX1-driven arrest of differentiation in pre-B ALL.

DeepMirTar: a deep-learning approach for predicting human miRNA targets.

Science.gov (United States)

Wen, Ming; Cong, Peisheng; Zhang, Zhimin; Lu, Hongmei; Li, Tonghua

2018-06-01

MicroRNAs (miRNAs) are small noncoding RNAs that function in RNA silencing and post-transcriptional regulation of gene expression by targeting messenger RNAs (mRNAs). Because the underlying mechanisms associated with miRNA binding to mRNA are not fully understood, a major challenge of miRNA studies involves the identification of miRNA-target sites on mRNA. In silico prediction of miRNA-target sites can expedite costly and time-consuming experimental work by providing the most promising miRNA-target-site candidates. In this study, we reported the design and implementation of DeepMirTar, a deep-learning-based approach for accurately predicting human miRNA targets at the site level. The predicted miRNA-target sites are those having canonical or non-canonical seed, and features, including high-level expert-designed, low-level expert-designed, and raw-data-level, were used to represent the miRNA-target site. Comparison with other state-of-the-art machine-learning methods and existing miRNA-target-prediction tools indicated that DeepMirTar improved overall predictive performance. DeepMirTar is freely available at https://github.com/Bjoux2/DeepMirTar_SdA. lith@tongji.edu.cn, hongmeilu@csu.edu.cn. Supplementary data are available at Bioinformatics online.

The Involvement of miR-29b-3p in Arterial Calcification by Targeting Matrix Metalloproteinase-2

Directory of Open Access Journals (Sweden)

Wenhong Jiang

2017-01-01

Full Text Available Vascular calcification is a risk predictor and common pathological change in cardiovascular diseases that are associated with elastin degradation and phenotypic transformation of vascular smooth muscle cells via gelatinase matrix metalloproteinase-2 (MMP2. However, the mechanisms involved in this process remain unclear. In this study, we investigated the relationships between miR-29b-3p and MMP2, to confirm miR-29b-3p-mediated MMP2 expression at the posttranscriptional level in arterial calcification. In male Sprague Dawley rats, arterial calcification was induced by subcutaneous injection of a toxic dose of cholecalciferol. In vivo, the quantitative real-time polymerase chain reaction (qRT-PCR showed that MMP2 expression was upregulated in calcified arterial tissues, and miR-29b-3p expression was downregulated. There was a negative correlation between MMP2 mRNA expression and miR-29b-3p levels (P=0.0014, R2=0.481. Western blotting showed that MMP2 expression was significantly increased in rats treated with cholecalciferol. In vitro, overexpression of miR-29b-3p led to decreased MMP2 expression in rat vascular smooth muscle cells, while downregulation of miR-29b-3p expression led to increased MMP2 expression. Moreover, the luciferase reporter assay confirmed that MMP2 is the direct target of miR-29b-3p. Together, our results demonstrated that a role of miR-29b-3p in vascular calcification involves targeting MMP2.

miR-137 inhibits the invasion of melanoma cells through downregulation of multiple oncogenic target genes.

Science.gov (United States)

Luo, Chonglin; Tetteh, Paul W; Merz, Patrick R; Dickes, Elke; Abukiwan, Alia; Hotz-Wagenblatt, Agnes; Holland-Cunz, Stefan; Sinnberg, Tobias; Schittek, Birgit; Schadendorf, Dirk; Diederichs, Sven; Eichmüller, Stefan B

2013-03-01

MicroRNAs are small noncoding RNAs that regulate gene expression and have important roles in various types of cancer. Previously, miR-137 was reported to act as a tumor suppressor in different cancers, including malignant melanoma. In this study, we show that low miR-137 expression is correlated with poor survival in stage IV melanoma patients. We identified and validated two genes (c-Met and YB1) as direct targets of miR-137 and confirmed two previously known targets, namely enhancer of zeste homolog 2 (EZH2) and microphthalmia-associated transcription factor (MITF). Functional studies showed that miR-137 suppressed melanoma cell invasion through the downregulation of multiple target genes. The decreased invasion caused by miR-137 overexpression could be phenocopied by small interfering RNA knockdown of EZH2, c-Met, or Y box-binding protein 1 (YB1). Furthermore, miR-137 inhibited melanoma cell migration and proliferation. Finally, miR-137 induced apoptosis in melanoma cell lines and decreased BCL2 levels. In summary, our study confirms that miR-137 acts as a tumor suppressor in malignant melanoma and reveals that miR-137 regulates multiple targets including c-Met, YB1, EZH2, and MITF.

miR-1271 inhibits migration, invasion and epithelial-mesenchymal transition by targeting ZEB1 and TWIST1 in pancreatic cancer cells

Energy Technology Data Exchange (ETDEWEB)

Liu, Huaize [Department of Developmental Genetics, Nanjing Medical University, Nanjing 210029 (China); Wang, Han [The First Clinical Medical College of Nanjing Medical University, Nanjing 210029 (China); Liu, Xiaoxiao [Department of Biotechnology, Nanjing Medical University, Nanjing 210029 (China); Yu, Tingting, E-mail: tingting@njmu.edu.cn [Department of Developmental Genetics, Nanjing Medical University, Nanjing 210029 (China)

2016-04-01

Pancreatic cancer (PC) remains one of the most lethal types of cancer in adults. The purpose of this study was to determine the role of miR-1271 in regulation of epithelial mesenchymal transition (EMT) and metastasis of pancreatic cancer cells. miR-1271 was identified to be significantly down-regulated in PC tissues by miRNA array. Also, an increase of EMT-regulators ZEB1 and TWIST1 expression level is accompanied by a decrease of miR-1271. We showed that expression of miR-1271 was significantly down-regulated in PC tissues as compared with that in normal tissues. In addition, our results showed that miR-1271 expression levels were decreased while ZEB1 and TWIST1 expression levels were increased in detected PC cell lines. Moreover, ectopic expression of miR-1271 suppressed and antagomiR-1271 promoted proliferation, migration, and invasion in SW1990 and PANC-1 cells. Bioinformatics coupled with luciferase and Western blot assays also revealed that miR-1271 inhibited expression of ZEB1 and TWIST1, which are master regulators of tumor metastasis. Our study first indicates that miR-1271 functions as a suppressor in regulating of pancreatic cancer EMT by targeting ZEB1 and TWIST1, and it promise as a therapeutic target and prognostic marker for metastatic pancreatic cancer. - Highlights: • miR-1271 is downregulated in pancreatic cancer tissues and cell lines. • miR-1271 regulates cell metastasis ability and EMT marker expression. . • miR-1271 directly targets ZEB1 and TWIST1. • ZEB1 and TWIST1 are functionally related to the effects of miR-1271.

miR-1271 inhibits migration, invasion and epithelial-mesenchymal transition by targeting ZEB1 and TWIST1 in pancreatic cancer cells

International Nuclear Information System (INIS)

Liu, Huaize; Wang, Han; Liu, Xiaoxiao; Yu, Tingting

2016-01-01

Pancreatic cancer (PC) remains one of the most lethal types of cancer in adults. The purpose of this study was to determine the role of miR-1271 in regulation of epithelial mesenchymal transition (EMT) and metastasis of pancreatic cancer cells. miR-1271 was identified to be significantly down-regulated in PC tissues by miRNA array. Also, an increase of EMT-regulators ZEB1 and TWIST1 expression level is accompanied by a decrease of miR-1271. We showed that expression of miR-1271 was significantly down-regulated in PC tissues as compared with that in normal tissues. In addition, our results showed that miR-1271 expression levels were decreased while ZEB1 and TWIST1 expression levels were increased in detected PC cell lines. Moreover, ectopic expression of miR-1271 suppressed and antagomiR-1271 promoted proliferation, migration, and invasion in SW1990 and PANC-1 cells. Bioinformatics coupled with luciferase and Western blot assays also revealed that miR-1271 inhibited expression of ZEB1 and TWIST1, which are master regulators of tumor metastasis. Our study first indicates that miR-1271 functions as a suppressor in regulating of pancreatic cancer EMT by targeting ZEB1 and TWIST1, and it promise as a therapeutic target and prognostic marker for metastatic pancreatic cancer. - Highlights: • miR-1271 is downregulated in pancreatic cancer tissues and cell lines. • miR-1271 regulates cell metastasis ability and EMT marker expression. . • miR-1271 directly targets ZEB1 and TWIST1. • ZEB1 and TWIST1 are functionally related to the effects of miR-1271.

miR-34 increases in vitro PANC-1 cell sensitivity to gemcitabine via targeting Slug/PUMA.

Science.gov (United States)

Zhang, Qing-An; Yang, Xu-Hai; Chen, Dong; Yan, Xiang; Jing, Fu-Chun; Liu, Hong-Qian; Zhang, Ronghua

2018-01-01

miR-34 was deregulated in tumor tissues compared with corresponding noncancerous tissue samples. Furthermore, miR-34 may contribute to cancer-stromal interaction associated with cancer progression. However, whether miR-34 could decrease chemoresistance of cancer cells to chemotherapeutic agent remains unclear. In our study, we examined whether overexpression of miR-34 could sensitize gemcitabine -mediated apoptosis in human pancreatic cancer PANC-1 cells. We found that miR-34 markedly induced gemcitabine -mediated apoptosis in PANC-1 cells. miR-34 induced down-regulation of Slug expression and upregulation of p53 up-regulated modulator of apoptosis (PUMA) expression. The over-expression of Slug or downregulation of PUMA by Slug cDNA or PUMA siRNA transfection markedly blocked miR-34-induced gemcitabine sensitization. Furthermore, miR-34 induced PUMA expression by downregulation of Slug. Taken together, our study demonstrates that miR-34 enhances sensitization against gemcitabine-mediated apoptosis through the down-regulation of Slug expression, and up-regulation of Slug-dependent PUMA expression.

MiR-328 suppresses the survival of esophageal cancer cells by targeting PLCE1

International Nuclear Information System (INIS)

Han, Na; Zhao, Wenchao; Zhang, Zhongmian; Zheng, Pengyuan

2016-01-01

Esophageal cancer (EC) is the sixth leading cause of death worldwide. Recent studies have highlighted the vital role of microRNAs (miRNAs) in EC development and diagnosis. In our study, qPCR analysis showed that miRNA-328 was expressed at significantly low levels in EC109 and EC9706 cells. The results also showed that overexpression of miR-328 by lentivirus-mediated gene transfer markedly inhibited cell proliferation and invasion, and enhanced apoptosis; whereas, inhibition of miR-328 significantly promoted cell proliferation and invasion, and suppressed apoptosis in EC109 and EC9706 cells. Dual-luciferase reporter assay confirmed that miR-328 directly targeted phospholipase C epsilon 1 (PLCE1) by binding to target sequences in the 3′-UTR. qPCR and Western blot analysis showed that the PLCE1 was overexpressed in EC109 and EC9706 cells. Additionally, we found that miR-328 overexpression decreased PLCE1 mRNA and protein levels, while miR-328 inhibition enhanced the PLCE1 expression. Further analysis showed that PLCE1 overexpression rescued the inhibitory effect of miR-328 on cell proliferation and invasion, and repressed the promotive effect of miR-328 on cell apoptosis. In conclusion, our results suggest that miR-328 suppresses the survival of EC cells by regulating PLCE1 expression, which might be a potential therapeutic method for EC. - Highlights: • PLCE1 was a target gene of miR-328. • MiR-328 overexpression decreased PLCE1 expression. • PLCE1 overexpression rescued the inhibitory effect of miR-328 on the survival of EC cells.

MiR-328 suppresses the survival of esophageal cancer cells by targeting PLCE1

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Han, Na [Department of Oncology, The Second Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, 450014 (China); Zhao, Wenchao [Department of Physiology and Neurobiology, College of Basic Medical Sciences, Zhengzhou University, Zhengzhou, Henan, 450001 (China); Zhang, Zhongmian [Department of Oncology, The Second Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, 450014 (China); Zheng, Pengyuan, E-mail: pengyuanzhengcn@163.com [No.3, Kangfuqian Street, Department of Gastroenterology, The Fifth Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, 450052 (China); No.3, Kangfuqian Street, Medical Microecology and Clinical Nutrition Research Institute of Zhengzhou University, Zhengzhou, Henan, 450052 (China)

2016-01-29

Esophageal cancer (EC) is the sixth leading cause of death worldwide. Recent studies have highlighted the vital role of microRNAs (miRNAs) in EC development and diagnosis. In our study, qPCR analysis showed that miRNA-328 was expressed at significantly low levels in EC109 and EC9706 cells. The results also showed that overexpression of miR-328 by lentivirus-mediated gene transfer markedly inhibited cell proliferation and invasion, and enhanced apoptosis; whereas, inhibition of miR-328 significantly promoted cell proliferation and invasion, and suppressed apoptosis in EC109 and EC9706 cells. Dual-luciferase reporter assay confirmed that miR-328 directly targeted phospholipase C epsilon 1 (PLCE1) by binding to target sequences in the 3′-UTR. qPCR and Western blot analysis showed that the PLCE1 was overexpressed in EC109 and EC9706 cells. Additionally, we found that miR-328 overexpression decreased PLCE1 mRNA and protein levels, while miR-328 inhibition enhanced the PLCE1 expression. Further analysis showed that PLCE1 overexpression rescued the inhibitory effect of miR-328 on cell proliferation and invasion, and repressed the promotive effect of miR-328 on cell apoptosis. In conclusion, our results suggest that miR-328 suppresses the survival of EC cells by regulating PLCE1 expression, which might be a potential therapeutic method for EC. - Highlights: • PLCE1 was a target gene of miR-328. • MiR-328 overexpression decreased PLCE1 expression. • PLCE1 overexpression rescued the inhibitory effect of miR-328 on the survival of EC cells.

miR-129 suppresses tumor cell growth and invasion by targeting PAK5 in hepatocellular carcinoma

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Zhai, Jian [Department II of Interventional Radiology, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438 (China); Qu, Shuping [Department II of Special Medical Care, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438 (China); Li, Xiaowei; Zhong, Jiaming; Chen, Xiaoxia [Department II of Interventional Radiology, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438 (China); Qu, Zengqiang, E-mail: drquzengqiang@163.com [Department II of Interventional Radiology, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438 (China); Wu, Dong, E-mail: wudongstc@126.com [Department II of Special Medical Care, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438 (China)

2015-08-14

Emerging evidence suggests that microRNAs (miRNAs) play important roles in regulating HCC development and progression; however, the mechanisms by which their specific functions and mechanisms remained to be further explored. miR-129 has been reported in gastric cancers, lung cancer and colon cancer. In this study, we disclosed a new tumor suppresser function of miR-129 in HCC. We also found the downregulation of miR-129 occurred in nearly 3/4 of the tumors examined (56/76) compared with adjacent nontumorous tissues, which was more importantly, correlated to the advanced stage and vascular invasion. We then demonstrated that miR-129 overexpression attenuated HCC cells proliferation and invasion, inducing apoptosis in vitro. Moreover, we used miR-129 antagonist and found that anti-miR-129 promoted HCC cells malignant phenotypes. Mechanistically, our further investigations revealed that miR-129 suppressed cell proliferation and invasion by targeting the 3’-untranslated region of PAK5, as well as miR-129 silencing up-regulated PAK5 expression. Moreover, miR-129 expression was inversely correlated with PAK5 expression in 76 cases of HCC samples. RNA interference of PAK5 attenuated anti-miR-129 mediated cell proliferation and invasion in HCC cells. Taken together, these results demonstrated that miR-129 suppressed tumorigenesis and progression by directly targeting PAK5, defining miR-129 as a potential treatment target for HCC. - Highlights: • Decreased of miR-129 is found in HCC and associated with advanced stage and metastasis. • miR-129 suppresses proliferation and invasion of HCC cells. • miR-129 directly targets the 3′ UTR of PAK5 and diminishes PAK5 expression. • PAK5 is involved in miR-129 mediated suppression functions.

Epigenetic modification of miR-10a regulates renal damage by targeting CREB1 in type 2 diabetes mellitus.

Science.gov (United States)

Shan, Qun; Zheng, Guihong; Zhu, Aihua; Cao, Li; Lu, Jun; Wu, Dongmei; Zhang, ZiFeng; Fan, Shaohua; Sun, Chunhui; Hu, Bin; Zheng, Yuanlin

2016-09-01

Emerging evidence has shown that microRNA-mediated gene expression modulation plays a crucial role in the pathogenesis of type 2 diabetes mellitus, but the novel miRNAs involved in type 2 diabetes and its functional regulatory mechanisms still need to be determined. In this study, we assessed the role of miR-10a in extracellular matrix accumulation in the kidney of diabetic mellitus induced by combining administration of chronic high fat diet (HFD) and low dosage of streptozotocin (STZ, 35mg/kg). Here, we found that HFD/STZ administration decreased the level of microRNA (miR-10a) expression in ICR strain mice. Overexpression of miR-10a alleviated the increased ratio of urine albumin-to-creatinine (ACR) ratio of HFD/STZ mice. In contrast, knockdown of miR-10a increased the ratio of kidney ACR in naïve mice. Furthermore, cAMP response element binding protein 1 (CREB1) was validated as a target of miR-10a in vitro and in vivo. CREB1 and its downstream fibronectin (FN, extracellular matrix) were increased in HFD/STZ-treated mice, which was reversed by kidney miR-10a overexpression. The content of CREB1 and FN was increased by miR-10a knockdown in kidney of naïve mice. Furthermore, histone deacetylase 3 (HDAC3) was revealed to be increased in kidney of HFD/STZ mice, accompanied with the augmentation of ACR ratio and FN level. Knockdown of HDAC3 with siRNA significantly caused the increase of miR-10a, resulting in the decrease in CREB1 and FN expression in kidney of HFD/STZ mice. Contrarily, HDAC3 overexpression mediated by lentivirus decreased miR-10a content, and enhanced ACR value, CREB1 and FN formation in naïve mice. Collectively, these results elucidate that HDAC3/miR-10a/CREB1 serves as a new mechanism underlying kidney injury, providing potential therapeutic targets in type 2 diabetes. Copyright © 2016. Published by Elsevier Inc.

Upregulation of Mir-21 Levels in the Vitreous Humor Is Associated with Development of Proliferative Vitreoretinal Disease.

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Ayumi Usui-Ouchi

Full Text Available MicroRNAs (miRNAs are small noncoding RNAs that regulate gene expression by post-transcriptional inhibition of mRNA translation. Dysregulation of miRNAs, including circulating miRNAs, has been reported to play an important role in the development of various diseases, including fibrotic diseases. Aberrant expression of miRNAs in the vitreous humor of vitreoretinal diseased eyes has been reported. However, the expression pattern of miRNAs present in the vitreous humor of proliferative vitreoretinal disease (PVD patients, including proliferative diabetic retinopathy (PDR, and proliferative vitreoretinopathy (PVR, remains unknown. To investigate the factors important for the development of PVD, we characterized the miRNAs present in the vitreous humor of PVD patients and analyzed the expression profiles of 377 miRNAs using quantitative polymerase chain reaction-based miRNA arrays. The expression of a specific subset of miRNAs, previously reported to be associated with the development of angiogenesis and fibrosis, was significantly altered in the vitreous of PVD patients. Among these miRNAs, we identified miR-21 as a candidate fibrotic miRNA with an important role in the pathogenesis of PVD. Increased miR-21 levels in the vitreous were associated with retinal fibrosis, including PVR and PDR. Because epithelial-mesenchymal transition (EMT of retinal pigment epithelial cells (RPECs plays a critical role in retinal fibrosis, the expression of miR-21 in human RPECs was determined. Its expression in RPECs was induced by transforming growth factor-β, a key growth factor involved in fibrogenesis, and was enhanced by high glucose culture conditions, suggesting that miR-21 expression positively correlates with disease progression. Gain- and loss-of-function studies revealed that miR-21 promoted cell proliferation and migration of ARPE-19 cells without affecting EMT-related gene expression. Together, our studies have identified miR-21 as a potential disease

miR-326 targets antiapoptotic Bcl-xL and mediates apoptosis in human platelets.

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Shifang Yu

Full Text Available Platelets play crucial roles in hemostasis, thrombosis, wound healing, inflammation, angiogenesis, and tumor metastases. Because they are anucleated blood cells, platelets lack nuclear DNA, but they do contain mitochondrial DNA, which plays a key role in regulating apoptosis. Recent evidence has suggested that miRNAs are also involved in regulating gene expression and apoptosis in platelets. Our previous study showed that the expression of miR-326 increased visibly when apheresis platelets were stored in vitro. The antiapoptotic Bcl-2 family regulator Bcl-xL has been identified as a putative target of miR-326. In the present study, dual reporter luciferase assays were used to characterize the function of miR-326 in the regulation of the apoptosis of platelet cells. These assays demonstrated that miR-326 bound to the 3'-translated region of Bcl-xL. To directly assess the functional effects of miR-326 expression, levels of Bcl-xL and the apoptotic status of stored apheresis platelets were measured after transfection of miR-326 mimic or inhibitor. Results indicated that miR-326 inhibited Bcl-xL expression and induced apoptosis in stored platelets. Additionally, miR-326 inhibited Bcl-2 protein expression and enhanced Bak expression, possibly through an indirect mechanism, though there was no effect on the expression of Bax. The effect of miR-326 appeared to be limited to apoptosis, with no significant effect on platelet activation. These results provide new insight into the molecular mechanisms affecting differential platelet gene regulation, which may increase understanding of the role of platelet apoptosis in multiple diseases.

miR-92a family and their target genes in tumorigenesis and metastasis

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Li, Molin, E-mail: molin_li@hotmail.com [Department of Pathophysiology, Basic Medical Science of Dalian Medical University, Dalian 116044 (China); Institute of Cancer Stem Cell, Dalian Medical University Cancer Center, Dalian 116044 (China); Guan, Xingfang; Sun, Yuqiang [Department of Pathophysiology, Basic Medical Science of Dalian Medical University, Dalian 116044 (China); Mi, Jun [Institute of Cancer Stem Cell, Dalian Medical University Cancer Center, Dalian 116044 (China); Shu, Xiaohong [College of Pharmacy, Dalian Medical University Cancer Center, Dalian 116044 (China); Liu, Fang [Department of Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian 116027 (China); Li, Chuangang, E-mail: li_chuangang@sina.com [Department of Surgery, The Second Affiliated Hospital of Dalian Medical University, Dalian 116027 (China)

2014-04-15

The miR-92a family, including miR-25, miR-92a-1, miR-92a-2 and miR-363, arises from three different paralog clusters miR-17-92, miR-106a-363, and miR-106b-25 that are highly conservative in the process of evolution, and it was thought as a group of microRNAs (miRNAs) correlated with endothelial cells. Aberrant expression of miR-92a family was detected in multiple cancers, and the disturbance of miR-92a family was related with tumorigenesis and tumor development. In this review, the progress on the relationship between miR-92a family and their target genes and malignant tumors will be summarized. - Highlights: • Aberrant expression of miR-92a, miR-25 and miR-363 can be observed in many kinds of malignant tumors. • The expression of miR-92a family is regulated by LOH, epigenetic alteration, transcriptional factors such as SP1, MYC, E2F, wild-type p53 etc. • Roles of miR-92a family in tumorigenesis and development: promoting cell proliferation, invasion and metastasis, inhibiting cell apoptosis.

miR-92a family and their target genes in tumorigenesis and metastasis

International Nuclear Information System (INIS)

Li, Molin; Guan, Xingfang; Sun, Yuqiang; Mi, Jun; Shu, Xiaohong; Liu, Fang; Li, Chuangang

2014-01-01

The miR-92a family, including miR-25, miR-92a-1, miR-92a-2 and miR-363, arises from three different paralog clusters miR-17-92, miR-106a-363, and miR-106b-25 that are highly conservative in the process of evolution, and it was thought as a group of microRNAs (miRNAs) correlated with endothelial cells. Aberrant expression of miR-92a family was detected in multiple cancers, and the disturbance of miR-92a family was related with tumorigenesis and tumor development. In this review, the progress on the relationship between miR-92a family and their target genes and malignant tumors will be summarized. - Highlights: • Aberrant expression of miR-92a, miR-25 and miR-363 can be observed in many kinds of malignant tumors. • The expression of miR-92a family is regulated by LOH, epigenetic alteration, transcriptional factors such as SP1, MYC, E2F, wild-type p53 etc. • Roles of miR-92a family in tumorigenesis and development: promoting cell proliferation, invasion and metastasis, inhibiting cell apoptosis

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.

Directory of Open Access Journals (Sweden)

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.

In Silico Prediction and Validation of Gfap as an miR-3099 Target in Mouse Brain.

Science.gov (United States)

Abidin, Shahidee Zainal; Leong, Jia-Wen; Mahmoudi, Marzieh; Nordin, Norshariza; Abdullah, Syahril; Cheah, Pike-See; Ling, King-Hwa

2017-08-01

MicroRNAs are small non-coding RNAs that play crucial roles in the regulation of gene expression and protein synthesis during brain development. MiR-3099 is highly expressed throughout embryogenesis, especially in the developing central nervous system. Moreover, miR-3099 is also expressed at a higher level in differentiating neurons in vitro, suggesting that it is a potential regulator during neuronal cell development. This study aimed to predict the target genes of miR-3099 via in-silico analysis using four independent prediction algorithms (miRDB, miRanda, TargetScan, and DIANA-micro-T-CDS) with emphasis on target genes related to brain development and function. Based on the analysis, a total of 3,174 miR-3099 target genes were predicted. Those predicted by at least three algorithms (324 genes) were subjected to DAVID bioinformatics analysis to understand their overall functional themes and representation. The analysis revealed that nearly 70% of the target genes were expressed in the nervous system and a significant proportion were associated with transcriptional regulation and protein ubiquitination mechanisms. Comparison of in situ hybridization (ISH) expression patterns of miR-3099 in both published and in-house-generated ISH sections with the ISH sections of target genes from the Allen Brain Atlas identified 7 target genes (Dnmt3a, Gabpa, Gfap, Itga4, Lxn, Smad7, and Tbx18) having expression patterns complementary to miR-3099 in the developing and adult mouse brain samples. Of these, we validated Gfap as a direct downstream target of miR-3099 using the luciferase reporter gene system. In conclusion, we report the successful prediction and validation of Gfap as an miR-3099 target gene using a combination of bioinformatics resources with enrichment of annotations based on functional ontologies and a spatio-temporal expression dataset.

Mechanical and IL-1β Responsive miR-365 Contributes to Osteoarthritis Development by Targeting Histone Deacetylase 4

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Xu Yang

2016-03-01

Full Text Available Mechanical stress plays an important role in the initiation and progression of osteoarthritis. Studies show that excessive mechanical stress can directly damage the cartilage extracellular matrix and shift the balance in chondrocytes to favor catabolic activity over anabolism. However, the underlying mechanism remains unknown. MicroRNAs (miRNAs are emerging as important regulators in osteoarthritis pathogenesis. We have found that mechanical loading up-regulated microRNA miR-365 in growth plate chondrocytes, which promotes chondrocyte differentiation. Here, we explored the role of the mechanical responsive microRNA miR-365 in pathogenesis of osteoarthritis (OA. We found that miR-365 was up-regulated by cyclic loading and IL-1β stimulation in articular chondrocytes through a mechanism that involved the transcription factor NF-κB. miR-365 expressed significant higher level in rat anterior cruciate ligament (ACL surgery induced OA cartilage as well as human OA cartilage from primary OA patients and traumatic OA Patients. Overexpression of miR-365 in chondrocytes increases gene expression of matrix degrading enzyme matrix metallopeptidase 13 (MMP13 and collagen type X (Col X. The increase in miR-365 expression in OA cartilage and in response to IL-1 may contribute to the abnormal gene expression pattern characteristic of OA. Inhibition of miR-365 down-regulated IL-1β induced MMP13 and Col X gene expression. We further showed histone deacetylase 4 (HDAC4 is a direct target of miR-365, which mediates mechanical stress and inflammation in OA pathogenesis. Thus, miR-365 is a critical regulator of mechanical stress and pro-inflammatory responses, which contributes cartilage catabolism. Manipulation of the expression of miR-365 in articular chondrocytes by miR-365 inhibitor may be a potent therapeutic target for the prevention and treatment of osteoarthritis.

Valsartan ameliorates KIR2.1 in rats with myocardial infarction via the NF-κB-miR-16 pathway.

Science.gov (United States)

Li, Xinran; Hu, Hesheng; Wang, Ye; Xue, Mei; Li, Xiaolu; Cheng, Wenjuan; Xuan, Yongli; Yin, Jie; Yang, Na; Yan, Suhua

2016-09-30

MicroRNAs have an important role in regulating arrhythmogenesis. MicroRNA-16 (miR-16) is predicted to target KCNJ2. The regulation of miR-16 is primarily due to NF-κB. Whether valsartan could downregulate miR-16 via the inhibition of NF-κB after MI and whether miR-16 targets KCNJ2 remain unclear. MI rats received valsartan or saline for 7days. The protein levels of NF-κB p65, inhibitor κBα (IκBα), and Kir2.1 were detected by Western blot analysis. The mRNA levels of Kir2.1 and miR-16 were examined by quantitative real-time PCR. Whole cell patch-clamp techniques were applied to record IK1. MiR-16 expression was higher in the infarct border, and was accompanied by a depressed IK1/KIR2.1 level. Additionally, miR-16 overexpression suppressed KCNJ2/KIR2.1 expression. In contrast, miR-16 inhibition or binding-site mutation enhanced KCNJ2/KIR2.1 expression, establishing KCNJ2 as a miR-16 target. In the MI rats, compared to saline treatment, valsartan reduced NF-κB p65 and miR-16 expression and increased IκBα and Kir2.1 expression. In vitro, angiotensin II increased miR-16 expression and valsartan inhibited it. Overexpressing miR-16 in cells treated with valsartan abrogated its beneficial effect on KCNJ2/Kir2.1. NF-κB activation directly upregulates miR-16 expression. miR-16 controls KCNJ2 expression, and valsartan ameliorates Kir2.1 after MI partly depending on the NF-κB-miR-16 pathway. Copyright © 2015 Elsevier B.V. All rights reserved.

Thrombin-induced, TNFR-dependent miR-181c downregulation promotes MLL1 and NF-κB target gene expression in human microglia.

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Yin, Min; Chen, Zhiying; Ouyang, Yetong; Zhang, Huiyan; Wan, Zhigang; Wang, Han; Wu, Wei; Yin, Xiaoping

2017-06-29

Controlling thrombin-driven microglial activation may serve as a therapeutic target for intracerebral hemorrhage (ICH). Here, we investigated microRNA (miRNA)-based regulation of thrombin-driven microglial activation using an in vitro thrombin toxicity model applied to primary human microglia. A miRNA array identified 22 differential miRNA candidates. Quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) identified miR-181c as the most significantly downregulated miRNA. TargetScan analysis identified mixed lineage leukemia-1 (MLL1) as a putative gene target for miR-181c. qRT-PCR was applied to assess tumor necrosis factor-alpha (TNF-α), miR-181c, and MLL1 levels following thrombin or proteinase-activated receptor-4-specific activating peptide (PAR4AP) exposure. Anti-TNF-α antibodies and tumor necrosis factor receptor (TNFR) silencing were employed to test TNF-α/TNFR dependence. A dual-luciferase reporter system and miR-181c mimic transfection assessed whether mir-181c directly binds to and negatively regulates MLL1. Nuclear factor kappa-B (NF-κB)-dependent luciferase reporter assays and NF-κB target gene expression were assessed in wild-type (MLL1+) and MLL1-silenced cells. Thrombin or PAR4AP-induced miR-181c downregulation (p < 0.05) and MLL1 upregulation (p < 0.05) that were dependent upon TNF-α/TNFR. miR-181c decreased wild-type MLL1 3'-UTR luciferase reporter activity (p < 0.05), and a miR-181c mimic suppressed MLL1 expression (p < 0.05). Thrombin treatment increased, while miR-181c reduced, NF-κB activity and NF-κB target gene expression in both wild-type (MLL1+) and MLL1-silenced cells (p < 0.05). Thrombin-induced, TNF-α/TNFR-dependent miR-181c downregulation promotes MLL1 expression, increases NF-κB activity, and upregulates NF-κB target gene expression. As miR-181c opposes thrombin's stimulation of pro-inflammatory NF-κB activity, miR-181c mimic therapy may show promise in controlling thrombin

The key role of miR-21-regulated SOD2 in the medium-mediated bystander responses in human fibroblasts induced by α-irradiated keratinocytes

International Nuclear Information System (INIS)

Tian, Wenqian; Yin, Xiaoming; Wang, Longxiao; Wang, Jingdong; Zhu, Wei; Cao, Jianping; Yang, Hongying

2015-01-01

Highlights: • After co-culture with α-irradiated HaCaT cells, WS1 cells displayed oxidative stress and DNA damage. • Increased miR-21 expression in bystander cells was critical to the occurrence of RIBEs. • SOD2 of bystander cells played an important role in bystander responses. • miR-21 mediated bystander effects through its regulation on SOD2. - Abstract: Radiation-induced bystander effect (RIBE) is well accepted in the radiation research field by now, but the underlying molecular mechanisms for better understanding this phenomenon caused by intercellular communication and intracellular signal transduction are still incomplete. Although our previous study has demonstrated an important role of miR-21 of unirradiated bystander cells in RIBEs, the direct evidence for the hypothesis that RIBE is epigenetically regulated is still limited and how miR-21 mediates RIBEs is unknown. Reactive oxygen species (ROS) have been demonstrated to be involved in RIBEs, however, the roles of anti-oxidative stress system of cells in RIBEs are unclear. Using transwell insert co-culture system, we investigated medium-mediated bystander responses in WS1 human fibroblasts after co-culture with HaCaT keratinocytes traversed by α-particles. Results showed that the ROS levels in unirradiated bystander WS1 cells were significantly elevated after 30 min of co-culture, and 53BP1 foci, a surrogate marker of DNA damage, were obviously induced after 3 h of co-culture. This indicates the occurrence of oxidative stress and DNA damage in bystander WS1 cells after co-culture with irradiated keratinocytes. Furthermore, the expression of miR-21 was increased in bystander WS1 cells, downregulation of miR-21 eliminated the bystander responses, overexpression of miR-21 alone could induce bystander-like oxidative stress and DNA damage in WS1 cells. These data indicate an important mediating role of miR-21 in RIBEs. In addition, MnSOD or SOD2 in WS1 cells was involved in the bystander effects

The key role of miR-21-regulated SOD2 in the medium-mediated bystander responses in human fibroblasts induced by α-irradiated keratinocytes

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Tian, Wenqian; Yin, Xiaoming; Wang, Longxiao; Wang, Jingdong; Zhu, Wei; Cao, Jianping [School of Radiation Medicine and Protection, Medical College of Soochow University/Collaborative Innovation Center of Radiation Medicine of Jiangsu Higher Education Institutions, 199 Renai Road, Suzhou Industrial Park, Suzhou, Jiangsu Province 215123 (China); Yang, Hongying, E-mail: yanghongying@suda.edu.cn [School of Radiation Medicine and Protection, Medical College of Soochow University/Collaborative Innovation Center of Radiation Medicine of Jiangsu Higher Education Institutions, 199 Renai Road, Suzhou Industrial Park, Suzhou, Jiangsu Province 215123 (China); Institute of Radiotherapy & Oncology, Soochow University (China)

2015-10-15

Highlights: • After co-culture with α-irradiated HaCaT cells, WS1 cells displayed oxidative stress and DNA damage. • Increased miR-21 expression in bystander cells was critical to the occurrence of RIBEs. • SOD2 of bystander cells played an important role in bystander responses. • miR-21 mediated bystander effects through its regulation on SOD2. - Abstract: Radiation-induced bystander effect (RIBE) is well accepted in the radiation research field by now, but the underlying molecular mechanisms for better understanding this phenomenon caused by intercellular communication and intracellular signal transduction are still incomplete. Although our previous study has demonstrated an important role of miR-21 of unirradiated bystander cells in RIBEs, the direct evidence for the hypothesis that RIBE is epigenetically regulated is still limited and how miR-21 mediates RIBEs is unknown. Reactive oxygen species (ROS) have been demonstrated to be involved in RIBEs, however, the roles of anti-oxidative stress system of cells in RIBEs are unclear. Using transwell insert co-culture system, we investigated medium-mediated bystander responses in WS1 human fibroblasts after co-culture with HaCaT keratinocytes traversed by α-particles. Results showed that the ROS levels in unirradiated bystander WS1 cells were significantly elevated after 30 min of co-culture, and 53BP1 foci, a surrogate marker of DNA damage, were obviously induced after 3 h of co-culture. This indicates the occurrence of oxidative stress and DNA damage in bystander WS1 cells after co-culture with irradiated keratinocytes. Furthermore, the expression of miR-21 was increased in bystander WS1 cells, downregulation of miR-21 eliminated the bystander responses, overexpression of miR-21 alone could induce bystander-like oxidative stress and DNA damage in WS1 cells. These data indicate an important mediating role of miR-21 in RIBEs. In addition, MnSOD or SOD2 in WS1 cells was involved in the bystander effects

MiR-519d-3p suppresses invasion and migration of trophoblast cells via targeting MMP-2.

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Jie Ding

Full Text Available Our study was approved by the Medical Ethics Committee of Tang Du Hospital, Fourth Military Medical University and complied strictly with national ethical guidelines. Preeclampsia (PE is a specific clinical disorder characterized by gestational hypertension and proteinuria and is a leading cause of maternal and perinatal mortality worldwide. The miR-519d-3p is upregulated in the maternal plasma of patients with PE which indicates a possible association between this microRNA and the pathogenesis of PE. No studies to date have addressed the effect of miR-519d-3p on the invasion and migration of trophoblast cells. In our study, we found that miR-519d-3p expression was elevated in placental samples from patients with PE. In vitro, overexpression of miR-519d-3p significantly inhibited trophoblast cell migration and invasion, whereas transfection of a miR-519d-3p inhibitor enhanced trophoblast cell migration and invasion. Luciferase assays confirmed that matrix metalloproteinase-2 (MMP-2 is a direct target of miR-519d-3p. Quantitative real-time PCR and western blot assays showed that overexpression of miR-519d-3p downregulated MMP-2 mRNA and protein expression. Knockdown of MMP-2 using a siRNA attenuated the increased trophoblast migration and invasion promoted by the miR-519d-3p inhibitor. In placentas from patients with PE or normal pregnancies, a negative correlation between the expression of MMP-2 and miR-519d-3p was observed using the Pearson correlation and linear regression analysis. Our present findings suggest that upregulation of miR-519d-3p may contribute to the development of PE by inhibiting trophoblast cell migration and invasion via targeting MMP-2; miR-519d-3p may represent a potential predictive and therapeutic target for PE.

miR-4458 suppresses glycolysis and lactate production by directly targeting hexokinase2 in colon cancer cells

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Qin, Yaguang; Cheng, Chuanyao; Lu, Hong, E-mail: honglu6512@163.com; Wang, Yaqiu

2016-01-01

miR-4458, a new tumor-suppressor, was reported to down-regulated in human hepatocellular carcinoma. The expression status, roles and inhibitory mechanisms of miR-4458 in other tumors still need to be clarified. The aim of this study is to investigate the effects of miR-4458 and to elucidate the potential mechanism in colon cancer cells. Using bioinformatic databases, we predicted that hexokinase2 (HK2), a rate-limiting enzyme in the glycolytic pathway, was a target of miR-4458, so the effects of miR-4458 on glycolysis and lactate production was assessed in colon cancer cells. We found that miR-4458 was down-regulated and HK2 was up-regulated in colon cancer cells. Overexpression of miR-4458 inhibited proliferation, glycolysis, and lactate production under both normoxic and hypoxic conditions. Luciferase activity assays showed that HK2 was a direct target of miR-4458. Moreover, knockdown of HK2 by specific RNAi also suppressed proliferation, glycolysis, and lactate production under both normoxic and hypoxic conditions. In conclusion, our findings suggested that miR-4458 inhibited the progression of colon cancer cells by inhibition of glycolysis and lactate production via directly targeting HK2 mRNA. - Highlights: • miR-4458 is down-regulated in colon cancer cells. • miR-4458 suppresses proliferation, glycolysis, and lactate production. • HK2 is a target of miR-4458. • HK2 knockdown inhibits proliferation, glycolysis, and lactate production.

miR-150 suppresses the proliferation and tumorigenicity of leukemia stem cells by targeting the Nanog signaling pathway

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Dan-dan Xu

2016-11-01

Full Text Available Proliferation, a key feature of cancer cells, accounts for the majority of cancer-related diseases resulting in mortality. MicroRNAs (miRNAs plays important post-transcriptional modulation roles by acting on multiple signaling pathways, but the underlying mechanism in proliferation and tumorigenicity is unclear. Here, we identified the role of miR-150 in proliferation and tumorigenicity in leukemia stem cells (LSCs (CD34+CD38- cells. miR-150 expression was significantly down-regulated in LSCs from leukemia cell lines and clinical samples. Functional assays demonstrated that increased miR-150 expression inhibited proliferation and clonal and clonogenic growth, enhanced chemosensitivity, and attenuated tumorigenic activity of LSCs in vitro. Transplantation animal studies revealed that miR-150 overexpression progressively abrogates tumour growth. Immunohistochemistry assays demonstrated that miR-150 overexpression enhanced caspase-3 level and reduced Ki-67 level. Moreover, luciferase reporter assays indicated Nanog is a direct and functional target of miR-150. Nanog silencing using small interfering RNA recapitulated anti-proliferation and tumorigenicity inhibition effects. Furthermore, miR-150 directly down-regulated the expression of other cancer stem cell factors including Notch2 and CTNNB1. These results provide insights into the specific biological behaviour of miR-150 in regulating LSC proliferation and tumorigenicity. Targeting this miR-150/Nanog axis would be a helpful therapeutic strategy to treat acute myeloid leukemia.

MIR21 drives resistance to Heat Shock Protein 90 inhibition in cholangiocarcinoma

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Lampis, Andrea; Carotenuto, Pietro; Vlachogiannis, Georgios

2018-01-01

grown as subcutaneous xenograft tumors in mice. RESULTS: Cells with IDH1 and PBRM1 mutations had the highest level of sensitivity to histone deacetylase inhibitors. HSP90 inhibitors were effective in all cell lines, irrespective of mutations. Sensitivity of cells to HSP90 inhibitors correlated inversely...... was shown to target the DnaJ heatshockprotein family (Hsp40) member B5 (DNAJB5). Transgenic expression of DNAJB5 in CCA cells that overexpressed MIR21 re-sensitized them to HSP90 inhibitors. Sensitivity of PDOs to HSP90 inhibitors, in culture and when grown as xenograft tumors in mice, depended...

A Double Negative Loop Comprising ETV6/RUNX1 and MIR181A1 Contributes to Differentiation Block in t(12;21-Positive Acute Lymphoblastic Leukemia.

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Yung-Li Yang

Full Text Available Childhood acute lymphoblastic leukemia (ALL with t(12;21, which results in expression of the ETV6/RUNX1 fusion gene, is the most common chromosomal lesion in precursor-B (pre-B ALL. We identified 17 microRNAs that were downregulated in ETV6/RUNX1+ compared with ETV6/RUNX1- clinical samples. Among these microRNAs, miR-181a-1 was the most significantly reduced (by ~75%; P < 0.001. Using chromatin immunoprecipitation, we demonstrated that ETV6/RUNX1 directly binds the regulatory region of MIR181A1, and knockdown of ETV6/RUNX1 increased miR-181a-1 level. We further showed that miR-181a (functional counterpart of miR-181a-1 could target ETV6/RUNX1 and cause a reduction in the level of the oncoprotein ETV6/RUNX1, cell growth arrest, an increase in apoptosis, and induction of cell differentiation in ETV6/RUNX1+ cell line. Moreover, ectopic expression of miR-181a also resulted in decreased CD10 hyperexpression in ETV6/RUNX1+ primary patient samples. Taken together, our results demonstrate that MIR181A1 and ETV6/RUNX1 regulate each other, and we propose that a double negative loop involving MIR181A1 and ETV6/RUNX1 may contribute to ETV6/RUNX1-driven arrest of differentiation in pre-B ALL.

Non-canonical microRNAs miR-320 and miR-702 promote proliferation in Dgcr8-deficient embryonic stem cells

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Kim, Byeong-Moo; Choi, Michael Y.

2012-01-01

Highlights: ► Embryonic stem cells (ESCs) lacking non-canonical miRNAs proliferate slower. ► miR-320 and miR-702 are two non-canonical miRNAs expressed in ESCs. ► miR-320 and miR-702 promote proliferation of Dgcr8-deficient ESCs. ► miR-320 targets p57 and helps to release Dgcr8-deficient ESCs from G1 arrest. ► miR-702 targets p21 and helps to release Dgcr8-deficient ESCs from G1 arrest. -- Abstract: MicroRNAs are known to contribute significantly to stem cell phenotype by post-transcriptionally regulating gene expression. Most of our knowledge of microRNAs comes from the study of canonical microRNAs that require two sequential cleavages by the Drosha/Dgcr8 heterodimer and Dicer to generate mature products. In contrast, non-canonical microRNAs bypass the cleavage by the Drosha/Dgcr8 heterodimer within the nucleus but still require cytoplasmic cleavage by Dicer. The function of non-canonical microRNAs in embryonic stem cells (ESCs) remains obscure. It has been hypothesized that non-canonical microRNAs have important roles in ESCs based upon the phenotypes of ESC lines that lack these specific classes of microRNAs; Dicer-deficient ESCs lacking both canonical and non-canonical microRNAs have much more severe proliferation defect than Dgcr8-deficient ESCs lacking only canonical microRNAs. Using these cell lines, we identified two non-canonical microRNAs, miR-320 and miR-702, that promote proliferation of Dgcr8-deficient ESCs by releasing them from G1 arrest. This is accomplished by targeting the 3′-untranslated regions of the cell cycle inhibitors p57 and p21 and thereby inhibiting their expression. This is the first report of the crucial role of non-canonical microRNAs in ESCs.

miR-935 suppresses gastric signet ring cell carcinoma tumorigenesis by targeting Notch1 expression

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Yan, Chao [Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, Beijing, 100730 (China); Yu, Jianchun, E-mail: yu_jchpumch@163.com [Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, Beijing, 100730 (China); Kang, Weiming [Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, Beijing, 100730 (China); Liu, Yuqin [Cell Culture Center, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100005 (China); Ma, Zhiqiang; Zhou, Li [Department of General Surgery, Peking Union Medical College Hospital, Chinese Academy of Medical Science and Peking Union Medical College, Beijing, 100730 (China)

2016-01-29

Gastric signet ring cell carcinoma (GSRCC) is a unique pathological type of gastric carcinoma that is extremely invasive and has a poor prognosis. Expression of microRNAs (miRNAs) has been closely linked to the carcinogenesis of gastric cancer and has been considered as a powerful prognostic marker. The function of miR-935 has never been reported in cancer before. We found, using microRNA array, that expression of miR-935 in GSRCC cell lines is lower than in non-GSRCC cell lines, and enhanced expression of miR-935 in GSRCC cell-lines inhibit cell proliferation, migration and invasion. We also identified Notch1 as a direct target of miR-935. Knockdown of Notch1 reduced proliferation, migration/invasion of GSRCC cells, and overexpression Notch1's activated form (Notch intracellular domain) could rescue miR-935's tumor suppressive effect on GSRCC. Expression of miR-935 was lower in gastric carcinoma tissue than in paired normal tissue samples, and lower in GSRCC than in non-GSRCC. Our results demonstrate the inverse correlation between the expression of miR-935 and Notch1 in gastric tissues. We conclude that miR-935 inhibits gastric carcinoma cell proliferation, migration and invasion by targeting Notch1, suggesting potential applications of the miR-935-Notch1 pathway in gastric cancer clinical diagnosis and therapeutics, especially in gastric signet ring cell carcinoma. - Highlights: • The expression of miR-935 is lower in GC tissue than in paired normal tissue. • The expression of miR-935 is lower in GSRCC tissue than in non-GSRCC. • Enhanced expression of miR-935 suppresses tumorigenesis of GSRCC. • Notch1 is a direct target of miR-935.

miR-181a promotes G1/S transition and cell proliferation in pediatric acute myeloid leukemia by targeting ATM.

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Liu, Xiaodan; Liao, Wang; Peng, Hongxia; Luo, Xuequn; Luo, Ziyan; Jiang, Hua; Xu, Ling

2016-01-01

Abnormal expression of miRNAs is intimately related to a variety of human cancers. The purpose of this study is to confirm the expression of miR-181a and elucidate its physiological function and mechanism in pediatric acute myeloid leukemia (AML). Pediatric AML patients and healthy controls were enrolled, and the expression of miR-181a and ataxia telangiectasia mutated (ATM) in tissues were examined using quantitative PCR. Moreover, cell proliferation and cell cycle were evaluated in several cell lines (HL60, NB4 and K562) by using flow cytometry after transfected with miR-181a mimics and inhibitors, or ATM siRNA and control siRNA. Finally, ATM as the potential target protein of miR-181a was examined. We found that miR-181a was significantly increased in pediatric AML, which showed an inverse association with ATM expression. Overexpressed miR-181a in cell lines significantly enhanced cell proliferation, as well as increased the ratio of S-phase cells by miR-181a mimics transfection in vitro. Luciferase activity of the reporter construct identified ATM as the direct molecular target of miR-181a. ATM siRNA transfection significantly enhanced cell proliferation and increased the ratio of S-phase cells in vitro. The results revealed novel mechanism through which miR-181a regulates G1/S transition and cell proliferation in pediatric AML by regulating the tumor suppressor ATM, providing insights into the molecular mechanism in pediatric AML.

Demethylation-mediated miR-129-5p up-regulation inhibits malignant phenotype of osteogenic osteosarcoma by targeting Homo sapiens valosin-containing protein (VCP).

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Long, Xin Hua; Zhou, Yun Fei; Peng, Ai Fen; Zhang, Zhi Hong; Chen, Xuan Yin; Chen, Wen Zhao; Liu, Jia Ming; Huang, Shan Hu; Liu, Zhi Li

2015-05-01

Previous studies demonstrated that increased Homo sapiens valosin-containing protein (VCP) may be involved in osteosarcoma (OS) metastasis. However, the underlying mechanism of VCP over-expression in OS remains unknown. In the present study, we found a significantly negative correlation between miR-129-5p and VCP protein expression in OS tissues with pulmonary metastasis (Spearman's rho, rs = -0.948). Bioinformatical prediction, Luciferase reporter assay, Western blot, and RT-PCR assays performed on OS cells indicated that VCP is a target of miR-129-5p. In addition, three CPG islands in the region of miR-129-5p promoter were detected by bioinformatical prediction, and significantly higher expression of miR-129-5p and lower methylation level of miR-129-2 gene in OS cells treated with 5-Aza-2'-deoxycytidine (a potent DNA demethylating agent) than in those untreated cells were observed. Furthermore, lower migratory and invasive ability was found in cells with elevated miR-129-5p than in those with decreased miR-129-5p. These findings indicated that increased miR-129-5p may be mediated by demethylation and inhibit OS cell migration and invasion by targeting VCP in OS, and targeting miR-129-5p/VCP signaling pathway may serve as a therapeutic strategy for OS management, although further studies will be necessary.

Prediction of target genes for miR-140-5p in pulmonary arterial hypertension using bioinformatics methods.

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Li, Fangwei; Shi, Wenhua; Wan, Yixin; Wang, Qingting; Feng, Wei; Yan, Xin; Wang, Jian; Chai, Limin; Zhang, Qianqian; Li, Manxiang

2017-12-01

The expression of microRNA (miR)-140-5p is known to be reduced in both pulmonary arterial hypertension (PAH) patients and monocrotaline-induced PAH models in rat. Identification of target genes for miR-140-5p with bioinformatics analysis may reveal new pathways and connections in PAH. This study aimed to explore downstream target genes and relevant signaling pathways regulated by miR-140-5p to provide theoretical evidences for further researches on role of miR-140-5p in PAH. Multiple downstream target genes and upstream transcription factors (TFs) of miR-140-5p were predicted in the analysis. Gene ontology (GO) enrichment analysis indicated that downstream target genes of miR-140-5p were enriched in many biological processes, such as biological regulation, signal transduction, response to chemical stimulus, stem cell proliferation, cell surface receptor signaling pathways. Kyoto Encyclopedia of Genes and Genome (KEGG) pathway analysis found that downstream target genes were mainly located in Notch, TGF-beta, PI3K/Akt, and Hippo signaling pathway. According to TF-miRNA-mRNA network, the important downstream target genes of miR-140-5p were PPI, TGF-betaR1, smad4, JAG1, ADAM10, FGF9, PDGFRA, VEGFA, LAMC1, TLR4, and CREB. After thoroughly reviewing published literature, we found that 23 target genes and seven signaling pathways were truly inhibited by miR-140-5p in various tissues or cells; most of these verified targets were in accordance with our present prediction. Other predicted targets still need further verification in vivo and in vitro .

MiR-30a-5p suppresses tumor growth in colon carcinoma by targeting DTL

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Baraniskin, Alexander; Birkenkamp-Demtröder, Karin; Maghnouj, Abdelouahid

2012-01-01

MicroRNAs (miRNAs) are small non-coding RNAs that are involved in different biological processes by suppressing target gene expression. Altered expression of miR-30a-5p has been reported in colon carcinoma. To elucidate its potential biological role in colon cancer, miR-30a-5p was overexpressed via...... with in silico miRNA target prediction, we identified the denticleless protein homolog (DTL) as a potential miRNA-30a-5p target. Subsequent reporter gene assays confirmed the predicted miR-30a-5p binding site in the 3'untranslated region of DTL. Importantly, overexpression of DTL in HCT116 cells partially...... is frequently overexpressed in colorectal cancer. Thus, our data suggest that restoring miR-30a-5p function may prove useful as therapeutic strategy for tumors with reduced miR-30a-5p expression....

Differential Impact of miR-21 on Pain and Associated Affective and Cognitive Behavior after Spared Nerve Injury in B7-H1 ko Mouse

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Franziska Karl

2017-07-01

Full Text Available MicroRNAs (miRNAs are increasingly recognized as regulators of immune and neuronal gene expression and are potential master switches in neuropathic pain pathophysiology. miR-21 is a promising candidate that may link the immune and the pain system. To investigate the pathophysiological role of miR-21 in neuropathic pain, we assessed mice deficient of B7 homolog 1 (B7-H1, a major inhibitor of inflammatory responses. In previous studies, an upregulation of miR-21 had been shown in mouse lymphocytes. Young (8 weeks, middle-aged (6 months, and old (12 months B7-H1 ko mice and wildtype littermates (WT received a spared nerve injury (SNI. We assessed thermal withdrawal latencies and mechanical withdrawal thresholds. Further, we performed tests for anxiety-like and cognitive behavior. Quantitative real time PCR was used to determine miR-21 relative expression in peripheral nerves, and dorsal root ganglia (DRG at distinct time points after SNI. We found mechanical hyposensitivity with increasing age of naïve B7-H1 ko mice. Young and middle-aged B7-H1 ko mice were more sensitive to mechanical stimuli compared to WT mice (young: p < 0.01, middle-aged: p < 0.05. Both genotypes developed mechanical and heat hypersensitivity (p < 0.05 after SNI, without intergroup differences. No relevant differences were found after SNI in three tests for anxiety like behavior in B7-H1 ko and WT mice. Also, SNI had no effect on cognition. B7-H1 ko and WT mice showed a higher miR-21 expression (p < 0.05 and invasion of macrophages and T cells in the injured nerve 7 days after SNI without intergroup differences. Our study reveals that increased miR-21 expression in peripheral nerves after SNI is associated with reduced mechanical and heat withdrawal thresholds. These results point to a role of miR-21 in the pathophysiology of neuropathic pain, while affective behavior and cognition seem to be spared. Contrary to expectations, B7-H1 ko mice did not show higher miR-21

miR-122 targets NOD2 to decrease intestinal epithelial cell injury in Crohn’s disease

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Chen, Yu; Wang, Chengxiao; Liu, Ying; Tang, Liwei; Zheng, Mingxia [Department of Pediatrics, Jiangwan Hospital of Shanghai, Shanghai 200434 (China); Xu, Chundi [Department of Pediatrics, Ruijin affiliated to Shanghai Jiaotong University School of Medicine, Shanghai, 200025 (China); Song, Jian, E-mail: jiansongkxy@126.com [Department of Gastroenterology, Jiangwan Hospital of Shanghai, Shanghai 200434 (China); Meng, Xiaochun [Department of Pediatrics, Jiangwan Hospital of Shanghai, Shanghai 200434 (China)

2013-08-16

Highlights: •NOD2 is a target gene of miR-122. •miR-122 inhibits LPS-induced apoptosis by suppressing NOD2 in HT-29 cells. •miR-122 reduces the expression of pro-inflammatory cytokines (TNF-α and IFN-γ). •miR-122 promotes the release of anti-inflammatory cytokines (IL-4 and IL-10). •NF-κB signaling pathway is involved in inflammatory response induced by LPS. -- Abstract: Crohn’s disease (CD) is one of the two major types of inflammatory bowel disease (IBD) thought to be caused by genetic and environmental factors. Recently, miR-122 was found to be deregulated in association with CD progression. However, the underlying molecular mechanisms remain unclear. In the present study, the gene nucleotide-binding oligomerization domain 2 (NOD2/CARD15), which is strongly associated with susceptibility to CD, was identified as a functional target of miR-122. MiR-122 inhibited LPS-induced apoptosis by suppressing NOD2 in HT-29 cells. NOD2 interaction with LPS initiates signal transduction mechanisms resulting in the activation of nuclear factor κB (NF-κB) and the stimulation of downstream pro-inflammatory events. The activation of NF-κB was inhibited in LPS-stimulated HT-29 cells pretreated with miR-122 precursor or NOD2 shRNA. The expression of the pro-inflammatory cytokines TNF-α and IFN-γ was significantly decreased, whereas therelease of the anti-inflammatory cytokines IL-4 and IL-10 was increased in LPS-stimulated HT-29 cells pretreated with miR-122 precursor, NOD2 shRNA or the NF-κB inhibitor QNZ. Taken together, these results indicate that miR-122 and its target gene NOD2 may play an important role in the injury of intestinal epithelial cells induced by LPS.

IL-4 Up-Regulates MiR-21 and the MiRNAs Hosted in the CLCN5 Gene in Chronic Lymphocytic Leukemia.

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Natalia Ruiz-Lafuente

Full Text Available Interleukin 4 (IL-4 induces B-cell differentiation and survival of chronic lymphocytic leukemia (CLL cells. MicroRNAs (miRNAs regulate mRNA and protein expression, and several miRNAs, deregulated in CLL, might play roles as oncogenes or tumor suppressors. We have studied the miRNA profile of CLL, and its response to IL-4, by oligonucleotide microarrays, resulting in the detection of a set of 129 mature miRNAs consistently expressed in CLL, which included 41 differentially expressed compared to normal B cells (NBC, and 6 significantly underexpressed in ZAP-70 positive patients. IL-4 stimulation brought about up-regulation of the 5p and 3p mature variants of the miR-21 gene, which maps immediately downstream to the VMP1 gene, and of the mature forms generated from the miR-362 (3p and 5p, miR-500a (3p, miR-502 (3p, and miR-532 (3p and 5p genes, which map within the third intron of the CLCN5 gene. Both genes are in turn regulated by IL-4, suggesting that these miRNAs were regulated by IL-4 as passengers from their carrier genes. Their levels of up-regulation by IL-4 significantly correlated with cytoprotection. MiR-21 has been reported to be leukemogenic, associated to bad prognosis in CLL, and the miRNA more frequently overexpressed in human cancer. Up-regulation by IL-4 of miR-21 and the miRNAs hosted in the CLCN5 locus may contribute to evasion of apoptosis of CLL cells. These findings indicate that the IL-4 pathway and the miRNAs induced by IL-4 are promising targets for the development of novel therapies in CLL.

Cisplatin-induced downregulation of miR-199a-5p increases drug resistance by activating autophagy in HCC cell

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Xu, Ning; Zhang, Jianjun; Shen, Conghuan; Luo, Yi; Xia, Lei; Xue, Feng; Xia, Qiang

2012-01-01

Highlights: ► miR-199a-5p levels were significantly decreased after cisplatin treatment. ► Cisplatin treatment induced autophagy activation. ► Cisplatin-induced downregulation of miR-199a-5p increases drug resistance by activating autophagy in HCC cell. -- Abstract: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Systemic chemotherapy plays an important role in the treatment of patients with advanced liver cancer. However, chemoresistance to cisplatin is a major limitation of cisplatin-based chemotherapy in the clinic, and the underlying mechanism of such resistance is not fully understood. In the study, we found that miR-199a-5p levels were significantly reduced in HCC patients treated with cisplatin-based chemotherapy. Cisplatin treatment also resulted in decreased miR-199a-5p levels in human HCC cell lines. Forced expression of miR-199a-5p promoted cisplatin-induced inhibition of cell proliferation. Cisplatin treatment activated autophagy in Huh7 and HepG2 cells, which increased cell proliferation. We further demonstrated that downregulated miR-199a-5p enhanced autophagy activation by targeting autophagy-associated gene 7 (ATG7). More important, autophagy inhibition abrogated miR-199a-5p downregulation-induced cell proliferation. These data demonstrated that miR-199a-5p/autophagy signaling represents a novel pathway regulating chemoresistance, thus offering a new target for chemotherapy of HCC.

Cisplatin-induced downregulation of miR-199a-5p increases drug resistance by activating autophagy in HCC cell

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Xu, Ning; Zhang, Jianjun; Shen, Conghuan; Luo, Yi; Xia, Lei; Xue, Feng [Department of Transplantation and Hepatic Surgery, Renji Hospital, Shanghai Jiaotong University School of Medicine, 1630 Dongfang Road, Shanghai 200127, People' s Republic of China (China); Xia, Qiang, E-mail: xiaqiang1@yahoo.com.cn [Department of Transplantation and Hepatic Surgery, Renji Hospital, Shanghai Jiaotong University School of Medicine, 1630 Dongfang Road, Shanghai 200127, People' s Republic of China (China)

2012-07-13

Highlights: Black-Right-Pointing-Pointer miR-199a-5p levels were significantly decreased after cisplatin treatment. Black-Right-Pointing-Pointer Cisplatin treatment induced autophagy activation. Black-Right-Pointing-Pointer Cisplatin-induced downregulation of miR-199a-5p increases drug resistance by activating autophagy in HCC cell. -- Abstract: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Systemic chemotherapy plays an important role in the treatment of patients with advanced liver cancer. However, chemoresistance to cisplatin is a major limitation of cisplatin-based chemotherapy in the clinic, and the underlying mechanism of such resistance is not fully understood. In the study, we found that miR-199a-5p levels were significantly reduced in HCC patients treated with cisplatin-based chemotherapy. Cisplatin treatment also resulted in decreased miR-199a-5p levels in human HCC cell lines. Forced expression of miR-199a-5p promoted cisplatin-induced inhibition of cell proliferation. Cisplatin treatment activated autophagy in Huh7 and HepG2 cells, which increased cell proliferation. We further demonstrated that downregulated miR-199a-5p enhanced autophagy activation by targeting autophagy-associated gene 7 (ATG7). More important, autophagy inhibition abrogated miR-199a-5p downregulation-induced cell proliferation. These data demonstrated that miR-199a-5p/autophagy signaling represents a novel pathway regulating chemoresistance, thus offering a new target for chemotherapy of HCC.

LncRNA MEG3 enhances cisplatin sensitivity in non-small cell lung cancer by regulating miR-21-5p/SOX7 axis

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Wang P

2017-10-01

Full Text Available Pei Wang,* Dong Chen,* Hongbing Ma, Yong LiDepartment of Cardiothoracic Surgery, Huaihe Hospital of Henan University, Kaifeng, People’s Republic of China *These authors contributed equally to this work Background: Long noncoding RNAs (lncRNAs have been revealed to play essential role in drug resistance of multiple cancers. LncRNA MEG3 was previously reported to be associated with cisplatin (DDP resistance in non-small cell lung cancer (NSCLC cells. However, the molecular mechanism of MEG3 affecting DDP resistance in NSCLC remains to be further illustrated. In this study, we attempted to discuss whether MEG3 also could function as a competing endogenous RNA to regulate DDP resistance in NSCLC.Materials and methods: The expression of MEG3, miR-21-5p, and sex-determining region Y-box 7 (SOX7 in NSCLC tissues or cells was examined by quantitative real-time polymerase chain reaction (qRT-PCR. 3-(4,5-Dimethylthazol-2-yl-2,5-diphenyltetrazolium bromide (MTT, flow cytometry, and caspase-3 activity analysis were applied to assess the DDP sensitivity of NSCLC cells. The interaction between MEG3, miR-21-5p, and SOX7 was explored by luciferase reporter assay, RNA immunoprecipitation (RIP assay, qRT-PCR, and Western blot. Mouse NSCLC transplanted tumor was established to verify the functional role of MEG3 in DDP resistance in vivo.Results: MEG3 was downregulated in DDP-resistant NSCLC cells. Overexpression of MEG3 enhanced DDP sensitivity of NSCLC cells in vitro. MEG3 directly interacted with miR-21-5p and suppressed its expression. miR-21-5p significantly abolished the effects of MEG3 on DDP resistance via modulating cell proliferation and apoptosis. SOX7 was identified as a direct target of miR-21-5p and MEG3 positively regulated SOX7 expression by suppressing miR-21-5p. Moreover, MEG3 knockdown-induced pro-proliferative and anti-apoptotic effects were reversed in DDP-resistant NSCLC cells by upregulating SOX7. Furthermore, upregulation of MEG3 induced

MiR-223 targeting MAFB suppresses proliferation and migration of nasopharyngeal carcinoma cells

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Yang, Wanyong; Lan, Xi; Li, Dongmin; Li, Tao; Lu, Shemin

2015-01-01

Mounting evidence suggests that miRNAs have major functions in tumor pathogenesis, and this study aimed to identify the candidate miRNA and investigate its role in nasopharyngeal carcinoma (NPC). MiRNA and mRNA expressions were screened by microarray assays. The cell proliferation, colony formation and migration ability were measured by MTT, soft agar and wound healing assays, respectively. The tumor growth suppression was evaluated by xenografting in nude mice. The plasma miR-223 levels in NPC patients were detected by TaqMan analysis. Real-time quantitative PCR and Western blotting were used to confirm miR-223 and MAFB expression levels. The targeting relationship between miR-223 and MAFB was verified using dual luciferase reporter assay. The miR-223 expression was decreased in CNE-1, CNE-2 cells as compared with NP69 cells, an immortalized human nasopharyngeal epithelial cell line, and its level also reduced in NPC patients’ plasma as compared with healthy controls. Exogenous expression of miR-223 in CNE-2 cells could inhibit cell proliferation both in vitro and in vivo. Extrogenous miR-223 in CNE-2 cells would decrease the ability of colony formation and migration. MAFB, a transcription factor of Maf family members, was identified as a target gene of miR-223. We found that migration and invasion abilities were inhibited by MAFB silencing. MiR-223 negatively regulates the growth and migration of NPC cells via reducing MAFB expression, and this finding provides a novel insight into understanding miR-223 regulation mechanism in nasopharyngeal carcinoma tumorigenesis

miR-21 Expression in Cancer Cells may Not Predict Resistance to Adjuvant Trastuzumab in Primary Breast Cancer

DEFF Research Database (Denmark)

Nielsen, Boye Schnack; Balslev, Eva; Poulsen, Tim Svenstrup

2014-01-01

, predominantly in cancer cells, or in both stromal and cancer cells. There was no obvious difference between the HER2-positive and HER2-negative tumors in terms of the miR-21 expression patterns and intensities. To explore the possibility that miR-21 expression levels and/or cellular localization could predict...... expression patterns and intensities revealed no association between the miR-21 scores in the cancer cell population (p = 0.69) or the stromal cells population (p = 0.13) and recurrent disease after adjuvant trastuzumab. Thus, our findings show that elevated miR-21 expression does not predict resistance......Trastuzumab is established as standard care for patients with HER2-positive breast cancer both in the adjuvant and metastatic setting. However, 50% of the patients do not respond to the trastuzumab therapy, and therefore new predictive biomarkers are highly warranted. MicroRNAs (miRs) constitute...

Ionizing Radiation–Inducible miR-27b Suppresses Leukemia Proliferation via Targeting Cyclin A2

Energy Technology Data Exchange (ETDEWEB)

Wang, Bo; Li, Dongping; Kovalchuk, Anna; Litvinov, Dmitry; Kovalchuk, Olga, E-mail: olga.kovalchuk@uleth.ca

2014-09-01

Purpose: Ionizing radiation is a common carcinogen that is important for the development of leukemia. However, the underlying epigenetic mechanisms remain largely unknown. The goal of the study was to explore microRNAome alterations induced by ionizing radiation (IR) in murine thymus, and to determine the role of IR-inducible microRNA (miRNA/miR) in the development of leukemia. Methods and Materials: We used the well-established C57BL/6 mouse model and miRNA microarray profiling to identify miRNAs that are differentially expressed in murine thymus in response to irradiation. TIB152 human leukemia cell line was used to determine the role of estrogen receptor–α (ERα) in miR-27b transcription. The biological effects of ectopic miR-27b on leukemogenesis were measured by western immunoblotting, cell viability, apoptosis, and cell cycle analyses. Results: Here, we have shown that IR triggers the differential expression of miR-27b in murine thymus tissue in a dose-, time- and sex-dependent manner. miR-27b was significantly down-regulated in leukemia cell lines CCL119 and TIB152. Interestingly, ERα was overexpressed in those 2 cell lines, and it was inversely correlated with miR-27b expression. Therefore, we used TIB152 as a model system to determine the role of ERα in miR-27b expression and the contribution of miR-27b to leukemogenesis. β-Estradiol caused a rapid and transient reduction in miR-27b expression reversed by either ERα-neutralizing antibody or ERK1/2 inhibitor. Ectopic expression of miR-27b remarkably suppressed TIB152 cell proliferation, at least in part, by inducing S-phase arrest. In addition, it attenuated the expression of cyclin A2, although it had no effect on the levels of PCNA, PPARγ, CDK2, p21, p27, p-p53, and cleaved caspase-3. Conclusion: Our data reveal that β-estradiol/ERα signaling may contribute to the down-regulation of miR-27b in acute leukemia cell lines through the ERK1/2 pathway, and that miR-27b may function as a tumor

Ionizing Radiation–Inducible miR-27b Suppresses Leukemia Proliferation via Targeting Cyclin A2

International Nuclear Information System (INIS)

Wang, Bo; Li, Dongping; Kovalchuk, Anna; Litvinov, Dmitry; Kovalchuk, Olga

2014-01-01

Purpose: Ionizing radiation is a common carcinogen that is important for the development of leukemia. However, the underlying epigenetic mechanisms remain largely unknown. The goal of the study was to explore microRNAome alterations induced by ionizing radiation (IR) in murine thymus, and to determine the role of IR-inducible microRNA (miRNA/miR) in the development of leukemia. Methods and Materials: We used the well-established C57BL/6 mouse model and miRNA microarray profiling to identify miRNAs that are differentially expressed in murine thymus in response to irradiation. TIB152 human leukemia cell line was used to determine the role of estrogen receptor–α (ERα) in miR-27b transcription. The biological effects of ectopic miR-27b on leukemogenesis were measured by western immunoblotting, cell viability, apoptosis, and cell cycle analyses. Results: Here, we have shown that IR triggers the differential expression of miR-27b in murine thymus tissue in a dose-, time- and sex-dependent manner. miR-27b was significantly down-regulated in leukemia cell lines CCL119 and TIB152. Interestingly, ERα was overexpressed in those 2 cell lines, and it was inversely correlated with miR-27b expression. Therefore, we used TIB152 as a model system to determine the role of ERα in miR-27b expression and the contribution of miR-27b to leukemogenesis. β-Estradiol caused a rapid and transient reduction in miR-27b expression reversed by either ERα-neutralizing antibody or ERK1/2 inhibitor. Ectopic expression of miR-27b remarkably suppressed TIB152 cell proliferation, at least in part, by inducing S-phase arrest. In addition, it attenuated the expression of cyclin A2, although it had no effect on the levels of PCNA, PPARγ, CDK2, p21, p27, p-p53, and cleaved caspase-3. Conclusion: Our data reveal that β-estradiol/ERα signaling may contribute to the down-regulation of miR-27b in acute leukemia cell lines through the ERK1/2 pathway, and that miR-27b may function as a tumor

The IGF-I/JAK2-STAT3/miR-21 signaling pathway may be associated with human renal cell carcinoma cell growth.

Science.gov (United States)

Su, Ying; Zhao, An; Cheng, Guoping; Xu, Jingjing; Ji, Enming; Sun, Wenyong

2017-07-04

Renal cell carcinoma (RCC) is the highest mortality rate of the genitourinary cancers, and the treatment options are very limited. Thus, identification of molecular mechanisms underlying RCC tumorigenesis, is critical for identifying biomarkers for RCC diagnosis and prognosis. To validate whether the IGF-I/JAK2-STAT3/miR-21 signaling pathway is associated with human RCC cell growth. qRT-PCR and Western blotting were used to detect the mRNA and protein expression levels, respectively. The MTT assay was performed to determine cell survival rate. The Annexin V-FITC/PI apoptosis detection kit was used to detect cell apoptosis. We employed RCC tissues and cell lines (A498; ACHN; Caki-1; Caki-2 and 786-O) in the study. IGF-I, and its inhibitor (NT-157) were administrated to detect the effects of IGF-I on the expression of miR-21 and p-JAK2. JAK2 inhibitor (AG490), and si-STAT3 were used to detect the effects of JAK2/STAT3 signaling pathway on the expression of miR-21. In our study, we firstly showed that the expression levels of IGF-I and miR-21 were up-regulated in RCC tissues and cell lines. After exogenous IGF-I treatment, the expression levels of miR-21, p-IGF-IR and p-JAK2 were significantly increased, whereas NT-157 treatment showed the reversed results. Further study indicated that JAK2 inhibitor or si-STAT3 significantly reversed the IGF-I-induced miR-21 expression level. Finally, we found that IGF-I treatment significantly prompted human RCC cell survival and inhibited cell apoptosis, and NT-157 treatment showed the reversed results. The IGF-I/JAK2-STAT3/miR-21 signaling pathway may be associated with human RCC cell growth.

miR-320a regulates cell proliferation and apoptosis in multiple myeloma by targeting pre-B-cell leukemia transcription factor 3

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Lu, Yinghao [Jiangsu Institute of Hematology, First Affiliated Hospital of Soochow University, Key Laboratory of Thrombosis and Hemostasis Under Ministry of Health, Collaborative Innovation Center of Hematology, Suzhou, 215006 (China); Department of Hematology, Affiliated Hospital of Guizhou Medical University, The Hematopoietic Stem Cell Transplant Center of Guizhou Province, Blood Diseases Diagnosis and Treatment Center of Guizhou Province, Guiyang, 550004, Guizhou Province (China); Wu, Depei, E-mail: wudepei@medmail.com.cn [Jiangsu Institute of Hematology, First Affiliated Hospital of Soochow University, Key Laboratory of Thrombosis and Hemostasis Under Ministry of Health, Collaborative Innovation Center of Hematology, Suzhou, 215006 (China); Wang, Jishi, E-mail: lgylhlyh@aliyun.com [Department of Hematology, Affiliated Hospital of Guizhou Medical University, The Hematopoietic Stem Cell Transplant Center of Guizhou Province, Blood Diseases Diagnosis and Treatment Center of Guizhou Province, Guiyang, 550004, Guizhou Province (China); Li, Yan; Chai, Xiao; Kang, Qian [Department of Hematology, Affiliated Hospital of Guizhou Medical University, The Hematopoietic Stem Cell Transplant Center of Guizhou Province, Blood Diseases Diagnosis and Treatment Center of Guizhou Province, Guiyang, 550004, Guizhou Province (China)

2016-05-13

Aberrant expression of microRNAs (miRNAs) is implicated in cancer development and progression. While miR-320a is reported to be deregulated in many malignancy types, its biological role in multiple myeloma (MM) remains unclear. Here, we observed reduced expression of miR-320a in MM samples and cell lines. Ectopic expression of miR-320a dramatically suppressed cell viability and clonogenicity and induced apoptosis in vitro. Mechanistic investigation led to the identification of Pre-B-cellleukemia transcription factor 3 (PBX3) as a novel and direct downstream target of miR-320a. Interestingly, reintroduction of PBX3 abrogated miR-320a-induced MM cell growth inhibition and apoptosis. In a mouse xenograft model, miR-320a overexpression inhibited tumorigenicity and promoted apoptosis. Our findings collectively indicate that miR-320a inhibits cell proliferation and induces apoptosis in MM cells by directly targeting PBX3, supporting its utility as a novel and potential therapeutic agent for miRNA-based MM therapy. -- Highlights: •Expression of miR-320a in MM cell induces apoptosis in vitro. •miR-320a represses PBX3 via targeting specific sequences in the 3′UTR region. •Exogenous expression of PBX3 reverses the effects of miR-320a in inhibiting MM cell growth and promoting apoptosis. •Overexpression of miR-320a inhibits tumor growth and increases apoptosis in vivo.

miR-613 inhibits the growth and invasiveness of human hepatocellular carcinoma via targeting DCLK1

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Wang, Wenyao, E-mail: wangwy117@163.com; Zhang, Hongfei; Wang, Lichao; Zhang, Shaojun; Tang, Miao

2016-05-13

microRNAs (miRNAs) play key regulatory roles in various biological processes. In this study, we aimed to determine the expression and biological roles of miR-613 in hepatocellular carcinoma (HCC). Compared with non-cancerous liver tissues, miR-613 was significantly downregulated in HCC tissues. Ectopic expression of miR-613 significantly suppressed the proliferation and invasion of Hep3B and SMMC-7721 HCC cells. Bioinformatic and luciferase reporter analysis identified doublecortin-like kinase 1 (DCLK1) as a direct target of miR-613. Overexpression of miR-613 inhibited the expression of DCLK1 in HCC cells. There was a significant inverse correlation between miR-613 and DCLK1 protein expression in HCC samples. Small interfering RNA-mediated silencing of DCLK1 phenocopied the suppressive effects of miR-613 in HCC cells. Rescue experiments demonstrated that co-transfection of DCLK1 lacking the 3′-untranslated region partially prevented miR-613-induced suppression of HCC cell proliferation and invasion. In vivo studies confirmed that miR-613 overexpression retarded the growth of Hep3B xenograft tumors in nude mice, coupled with a reduction in the percentage of Ki67-positive tumor cells and DCLK1 protein expression. In conclusion, we provide first evidence for the suppressive activity of miR-613 in HCC, which is causally linked to targeting of DCLK1. Restoration of miR-613 may provide a potential therapeutic strategy for HCC. - Highlights: • miR-613 inhibits the aggressive phenotypes of HCC cells. • DCLK1 is a direct target of miR-613 in HCC. • miR-613 impairs HCC tumorigenesis in vivo.

miR-370 suppresses HBV gene expression and replication by targeting nuclear factor IA.

Science.gov (United States)

Fan, Hongxia; Lv, Ping; Lv, Jing; Zhao, Xiaopei; Liu, Min; Zhang, Guangling; Tang, Hua

2017-05-01

Hepatitis B virus (HBV) infection is a major health problem worldwide. The roles of microRNAs in the regulation of HBV expression are being increasingly recognized. In this study, we found that overexpression of miR-370 suppressed HBV gene expression and replication in Huh7 cells, whereas antisense knockdown of endogenous miR-370 enhanced HBV gene expression and replication in Huh7 cells and HepG2.2.15 cells. Further, we identified the transcription factor nuclear factor IA (NFIA) as a new host target of miR-370. Overexpression and knockdown studies showed that NFIA stimulated HBV gene expression and replication. Importantly, overexpression of NFIA counteracted the effect of miR-370 on HBV gene expression and replication. Further mechanistic studies showed that miR-370 suppressed HBV replication and gene expression by repressing HBV Enhancer I activity, and one of the NFIA binding site in the Enhancer I element was responsible for the repressive effect of miR-370 on HBV Enhancer I activity. Altogether, our results demonstrated that miR-370 suppressed HBV gene expression and replication through repressing NFIA expression, which stimulates HBV replication via direct regulation on HBV Enhancer I activities. Our findings may provide a new antiviral strategy for HBV infection. J. Med. Virol. 89:834-844, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

IAA-Ala Resistant3, an evolutionarily conserved target of miR167, mediates Arabidopsis root architecture changes during high osmotic stress

KAUST Repository

Kinoshita, Natsuko

2012-09-01

The functions of microRNAs and their target mRNAs in Arabidopsis thaliana development have been widely documented; however, roles of stress-responsive microRNAs and their targets are not as well understood. Using small RNA deep sequencing and ATH1 microarrays to profile mRNAs, we identified IAA-Ala Resistant3 (IAR3) as a new target of miR167a. As expected, IAR3 mRNA was cleaved at the miR167a complementary site and under high osmotic stress miR167a levels decreased, whereas IAR3 mRNA levels increased. IAR3 hydrolyzes an inactive form of auxin (indole-3-acetic acid [IAA]-alanine) and releases bioactive auxin (IAA), a central phytohormone for root development. In contrast with the wild type, iar3 mutants accumulated reduced IAA levels and did not display high osmotic stress-induced root architecture changes. Transgenic plants expressing a cleavage-resistant form of IAR3 mRNA accumulated high levels of IAR3 mRNAs and showed increased lateral root development compared with transgenic plants expressing wild-type IAR3. Expression of an inducible noncoding RNA to sequester miR167a by target mimicry led to an increase in IAR3 mRNA levels, further confirming the inverse relationship between the two partners. Sequence comparison revealed the miR167 target site on IAR3 mRNA is conserved in evolutionarily distant plant species. Finally, we showed that IAR3 is required for drought tolerance. © 2012 American Society of Plant Biologists. All rights reserved.

IAA-Ala Resistant3, an evolutionarily conserved target of miR167, mediates Arabidopsis root architecture changes during high osmotic stress

KAUST Repository

Kinoshita, Natsuko; Wang, Huan; Kasahara, Hiroyuki; Liu, Jun; MacPherson, Cameron R.; Machida, Yasunori; Kamiya, Yuji; Hannah, Matthew A.; Chuaa, Nam Hai

2012-01-01

The functions of microRNAs and their target mRNAs in Arabidopsis thaliana development have been widely documented; however, roles of stress-responsive microRNAs and their targets are not as well understood. Using small RNA deep sequencing and ATH1 microarrays to profile mRNAs, we identified IAA-Ala Resistant3 (IAR3) as a new target of miR167a. As expected, IAR3 mRNA was cleaved at the miR167a complementary site and under high osmotic stress miR167a levels decreased, whereas IAR3 mRNA levels increased. IAR3 hydrolyzes an inactive form of auxin (indole-3-acetic acid [IAA]-alanine) and releases bioactive auxin (IAA), a central phytohormone for root development. In contrast with the wild type, iar3 mutants accumulated reduced IAA levels and did not display high osmotic stress-induced root architecture changes. Transgenic plants expressing a cleavage-resistant form of IAR3 mRNA accumulated high levels of IAR3 mRNAs and showed increased lateral root development compared with transgenic plants expressing wild-type IAR3. Expression of an inducible noncoding RNA to sequester miR167a by target mimicry led to an increase in IAR3 mRNA levels, further confirming the inverse relationship between the two partners. Sequence comparison revealed the miR167 target site on IAR3 mRNA is conserved in evolutionarily distant plant species. Finally, we showed that IAR3 is required for drought tolerance. © 2012 American Society of Plant Biologists. All rights reserved.

miR-630 targets IGF1R to regulate response to HER-targeting drugs and overall cancer cell progression in HER2 over-expressing breast cancer.

Science.gov (United States)

Corcoran, Claire; Rani, Sweta; Breslin, Susan; Gogarty, Martina; Ghobrial, Irene M; Crown, John; O'Driscoll, Lorraine

2014-03-24

While the treatment of HER2 over-expressing breast cancer with recent HER-targeted drugs has been highly effective for some patients, primary (also known as innate) or acquired resistance limits the success of these drugs. microRNAs have potential as diagnostic, prognostic and predictive biomarkers, as well as replacement therapies. Here we investigated the role of microRNA-630 (miR-630) in breast cancer progression and as a predictive biomarker for response to HER-targeting drugs, ultimately yielding potential as a therapeutic approach to add value to these drugs. We investigated the levels of intra- and extracellular miR-630 in cells and conditioned media from breast cancer cell lines with either innate- or acquired- resistance to HER-targeting lapatinib and neratinib, compared to their corresponding drug sensitive cell lines, using qPCR. To support the role of miR-630 in breast cancer, we examined the clinical relevance of this miRNA in breast cancer tumours versus matched peritumours. Transfection of miR-630 mimics and inhibitors was used to manipulate the expression of miR-630 to assess effects on response to HER-targeting drugs (lapatinib, neratinib and afatinib). Other phenotypic changes associated with cellular aggressiveness were evaluated by motility, invasion and anoikis assays. TargetScan prediction software, qPCR, immunoblotting and ELISAs, were used to assess miR-630's regulation of mRNA, proteins and their phosphorylated forms. We established that introducing miR-630 into cells with innate- or acquired- resistance to HER-drugs significantly restored the efficacy of lapatinib, neratinib and afatinib; through a mechanism which we have determined to, at least partly, involve miR-630's regulation of IGF1R. Conversely, we demonstrated that blocking miR-630 induced resistance/insensitivity to these drugs. Cellular motility, invasion, and anoikis were also observed as significantly altered by miR-630 manipulation, whereby introducing miR-630 into cells

MiR-285 targets P450 (CYP6N23) to regulate pyrethroid resistance in Culex pipiens pallens.

Science.gov (United States)

Tian, Mengmeng; Liu, Bingqian; Hu, Hongxia; Li, Xixi; Guo, Qin; Zou, Feifei; Liu, Xianmiao; Hu, Mengxue; Guo, Juxin; Ma, Lei; Zhou, Dan; Sun, Yan; Shen, Bo; Zhu, Changliang

2016-12-01

MicroRNAs play critical roles in post-transcriptional regulation of gene expression, which participate in the modulation of almost all of the cellular processes. Although emerging evidence indicates that microRNAs are related with antineoplastic drugs resistance, whether microRNAs are responsible for insecticide resistance in mosquitos is poorly understood. In this paper, we found that miR-285 was significantly upregulated in the deltamethrin-resistant strain of Culex pipiens pallens, and overexpression miR-285 through microinjection increased mosquito survival rate against deltamethrin treatement. Using bioinformatic software, quantitative reverse transcription PCR, luciferase reporter assay and microinjection approaches, we conformed that CYP6N23 was the target of miR-285. Lower expression of CYP6N23 was observed in the deltamethrin-resistant strain. While, mosquito mortality rate was decreased after downregulating expression of CYP6N23 by dsRNA against CYP6N23 or miR-285 mimic microinjection. These findings revealed that miR-285 could target CYP6N23 to regulate pyrethroid resistance, providing new insights into mosquito insecticide resistance surveillance and control.

miR-425 inhibits melanoma metastasis through repression of PI3K-Akt pathway by targeting IGF-1.

Science.gov (United States)

Liu, Pei; Hu, Yaotian; Ma, Ling; Du, Min; Xia, Lin; Hu, Zhensheng

2015-10-01

miR-425 is a potential tumor suppressor in cancer, but its role in melanoma is still unknown. We aim to investigate miR-425 expression in melanoma tissues and cell lines. Next, cell proliferation, cell cycle, apoptosis and metastasis will be studied using lentivirus-mediated gain-of-function studies. The predicted results are stable miR-425 inhibits cell proliferation and metastasis and induced cell apoptosis. It is predicted that IGF-1 is a potential target gene of miR-495 by bioinformatics analysis. Then luciferase assay analysis identifies IGF-1 as a new direct target gene of miR-425 and miR-425 inhibits melanoma cancer progression via IGF-1. Collectively, our findings suggested that miR-425 may function as a tumor suppressor in melanoma by targeting IGF-1. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

The prognostic importance of miR-21 in stage II colon cancer: a population-based study

DEFF Research Database (Denmark)

Kjaer-Frifeldt, S.; Hansen, T. F.; Nielsen, B. S.

2012-01-01

that increasing miR-21 expression levels were significantly correlated to decreasing RF-CSS. Further investigations of the clinical importance of miR-21 in the selection of high-risk stage II colon cancer patients are merited. British Journal of Cancer (2012) 107, 1169-1174. doi:10.1038/bjc.2012.365 www......BACKGROUND: Despite several years of research and attempts to develop prognostic models a considerable fraction of stage II colon cancer patients will experience relapse within few years from their operation. The aim of the present study was to investigate the prognostic importance of miRNA-21 (mi......-free cancer-specific survival (RF-CSS): HR = 1.26; 95% CI: 1.15-1.60; P importance and was found to be significantly related to poor RF-CSS: HR 1.41; 95% CI: 1.19-1.67; P

Identification of targets of miR-200b by a SILAC-based quantitative proteomic approach

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Arivusudar Marimuthu

2014-09-01

Full Text Available miRNAs regulate gene expression by binding to cognate mRNAs causing mRNA degradation or translational repression. Mass spectrometry-based proteomic analysis is being widely used to identify miRNA targets. The miR-200b miRNA cluster is often overexpressed in multiple cancer types, but the identity of the targets remains elusive. Using SILAC-based analysis, we examined the effects of overexpression of a miR-200b mimic or a control miRNA in fibrosarcoma cells. We identified around 300 potential targets of miR-200b based on a change in the expression of protein levels. We validated a subset of potential targets at the transcript level using quantitative PCR.

miR-206 inhibits cell proliferation, migration, and invasion by targeting BAG3 in human cervical cancer.

Science.gov (United States)

Wang, Yingying; Tian, Yongjie

2018-01-02

miR-206 and bcl2-associated athanogene 3 (BAG3) have been suggested as important regulators in various cancer types. However, the biological role of miR-206 and BAG3 in cervical cancer (CC) remains unclear. Here, we investigated the expressions and mechanisms of miR-206 and BAG3 in cervical cancer using in vitro and in vivo assays. In the present study, miR-206 expression was expressed at a lower level in CC tissues and cells than adjacent normal tissues and NEEC cells. By contrast, BAG3 mRNA and protein were expressed at higher levels in CC tissues and cells. Furthermore, miR-206 overexpression repressed cell proliferation, migration and invasion in vitro, and the 3'-untranslated region (3'-UTR) of BAG3 was a direct target of miR-206. miR-206 overexpression also inhibited EGFR, Bcl-2 and MMP2/9 protein expression, but promoted Bax protein expression. Besides, BAG3 over-expression partially abrogated miR-206-inhibited cell proliferation and invasion, while BAG3 silencing enhanced miR206-mediated inhibition. In vivo assay revealed that miR-206 repressed tumor growth in nude mice xenograft model. In conclusion, miR-206 inhibits cell proliferation, migration, and invasion by targeting BAG3 in human cervical cancer. Thus, miR-206-BAG3 can be used as a useful target for cervical cancer.

Inhibition of 14q32 MicroRNAs miR-329, miR-487b, miR-494, and miR-495 Increases Neovascularization and Blood Flow Recovery After Ischemia

DEFF Research Database (Denmark)

Welten, S. M. J.; Bastiaansen, Ajnm; de Jong, R. C. M.

2014-01-01

in mice after single femoral artery ligation. Methods and Results: Gene silencing oligonucleotides (GSOs) were used to inhibit 4 14q32 microRNAs, miR-329, miR-487b, miR-494, and miR-495, 1 day before double femoral artery ligation. Blood flow recovery was followed by laser Doppler perfusion imaging. All 4...... GSOs clearly improved blood flow recovery after ischemia. Mice treated with GSO-495 or GSO-329 showed increased perfusion already after 3 days (30% perfusion versus 15% in control), and those treated with GSO-329 showed a full recovery of perfusion after 7 days (versus 60% in control). Increased...

Preclinical Evaluation of miR-15/107 Family Members as Multifactorial Drug Targets for Alzheimer's Disease

Directory of Open Access Journals (Sweden)

Sepideh Parsi

2015-01-01

Full Text Available Alzheimer's disease (AD is a multifactorial, fatal neurodegenerative disorder characterized by the abnormal accumulation of Aβ and Tau deposits in the brain. There is no cure for AD, and failure at different clinical trials emphasizes the need for new treatments. In recent years, significant progress has been made toward the development of miRNA-based therapeutics for human disorders. This study was designed to evaluate the efficiency and potential safety of miRNA replacement therapy in AD, using miR-15/107 paralogues as candidate drug targets. We identified miR-16 as a potent inhibitor of amyloid precursor protein (APP and BACE1 expression, Aβ peptide production, and Tau phosphorylation in cells. Brain delivery of miR-16 mimics in mice resulted in a reduction of AD-related genes APP, BACE1, and Tau in a region-dependent manner. We further identified Nicastrin, a γ-secretase component involved in Aβ generation, as a target of miR-16. Proteomics analysis identified a number of additional putative miR-16 targets in vivo, including α-Synuclein and Transferrin receptor 1. Top-ranking biological networks associated with miR-16 delivery included AD and oxidative stress. Collectively, our data suggest that miR-16 is a good candidate for future drug development by targeting simultaneously endogenous regulators of AD biomarkers (i.e., Aβ and Tau, inflammation, and oxidative stress.

miR-1 is increased in pulmonary hypertension and downregulates Kv1.5 channels in rat pulmonary arteries.

Science.gov (United States)

Mondejar-Parreño, Gema; Callejo, María; Barreira, Bianca; Morales-Cano, Daniel; Esquivel-Ruiz, Sergio; Moreno, Laura; Cogolludo, Angel; Perez-Vizcaino, Francisco

2018-05-02

■The expression of miR-1 is increased in lungs from the Hyp/Su5416 PAH rat model. ■PASMC from this animal model are more depolarised and show decreased expression and activity of Kv1.5. ■miR-1 directly targets Kv1.5 channels, reduces Kv1.5 activity and induces membrane depolarization. ■Antagomir-1 prevents Kv1.5 channel downregulation and the depolarization induced by hypoxia/Su5416 exposition. Impairment of voltage-dependent potassium channel (Kv) plays a central role in the development of cardiovascular diseases, including pulmonary arterial hypertension (PAH). MicroRNAs (miRNAs) are non-coding RNAs that regulate gene expression by binding to the 3'-UTR region of specific mRNAs. The aim of this study was to analyze the effects of miR-1 on Kv channel function in pulmonary arteries (PA). Kv channel activity was studied in PA from healthy animals transfected with miR-1 or scrambled-miR. Kv currents were studied using the whole-cell configuration of patch-clamp technique. The characterization of the Kv1.5 currents was performed with the selective inhibitor DPO-1. miR-1 expression was increased and Kv1.5 channels were decreased in lungs from a rat model of PAH induced by hypoxia and Su5416. miR-1 transfection increased cell capacitance, reduced Kv1.5 currents and induced membrane depolarization in isolated pulmonary artery smooth muscle cells (PASMCs). Luciferase reporter assay indicated that KCNA5, which encodes Kv1.5 channels, is a direct target gene of miR-1. Incubation of PA with Su5416 and hypoxia (3% O 2 ) increased miR-1 and induced a decline in Kv1.5 currents, which was prevented by antagomiR-1. In conclusion, these data indicate that miR-1 induces PASMC hypertrophy and reduces the activity and expression of Kv channels, suggesting a pathophysiological role in PAH. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Involvement of miR-485-5p in hepatocellular carcinoma progression targeting EMMPRIN.

Science.gov (United States)

Sun, Xiangjun; Liu, Yonglei; Li, Ming; Wang, Mingchun; Wang, Yutong

2015-05-01

EMMPRIN plays important roles in cancer development, which includes EMMPRIN 1, 2, 3, and 4 isoforms. EMMPRIN2 is the main component in human cancers, but its regulation by miRNAs is still unclear. In this study, we will investigate the mechanism of EMMPRIN regulation in hepatocellular carcinoma (HCC) by miRNAs. Through RT-PCR, we found that EMMPRIN2 was the main isoform in HCC cells. EMMPRIN2 was down-regulated significantly by predicted miRNAs and miR-485-5p was one of the miRNA that regulated EMMPRIN in HCC cell lines. It was verified that EMMPRIN was a target gene of miR-485-5p by using luciferase analysis assay. We found that miR-485-5p was significantly downregulated in HCC tissues and that its expression was inversely correlated with the TNM stage and metastasis in HCC samples. Results of cellular functions in HCC showed that miR-485-5p could inhibit cell proliferation and metastasis. Additionally, miR-485-5p overexpression suppressed HCC growth in vivo by down-regulation of EMMPRIN. Our study for the first time demonstrated that miR-485-5p represses HCC invasive and metastatic capacities by targeting EMMPRIN expression. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Nanoparticle-antagomiR based targeting of miR-31 to induce osterix and osteocalcin expression in mesenchymal stem cells.

Directory of Open Access Journals (Sweden)

Mark McCully

Full Text Available Mesenchymal stem cells are multipotent adult stem cells capable of generating bone, cartilage and fat, and are thus currently being exploited for regenerative medicine. When considering osteogenesis, developments have been made with regards to chemical induction (e.g. differentiation media and physical induction (e.g. material stiffness, nanotopography, targeting established early transcription factors or regulators such as runx2 or bone morphogenic proteins and promoting increased numbers of cells committing to osteo-specific differentiation. Recent research highlighted the involvement of microRNAs in lineage commitment and terminal differentiation. Herein, gold nanoparticles that confer stability to short single stranded RNAs were used to deliver MiR-31 antagomiRs to both pre-osteoblastic cells and primary human MSCs in vitro. Results showed that blocking miR-31 led to an increase in osterix protein in both cell types at day 7, with an increase in osteocalcin at day 21, suggesting MSC osteogenesis. In addition, it was noted that antagomiR sequence direction was important, with the 5 prime reading direction proving more effective than the 3 prime. This study highlights the potential that miRNA antagomiR-tagged nanoparticles offer as novel therapeutics in regenerative medicine.

miR-216b suppresses breast cancer growth and metastasis by targeting SDCBP

International Nuclear Information System (INIS)

Jana, Samir; Sengupta, Suman; Biswas, Subir; Chatterjee, Annesha; Roy, Himansu; Bhattacharyya, Arindam

2017-01-01

Breast cancer is the most deadly cancer among women and the second leading cause of cancer death worldwide. Treatment effectiveness is complicated with tumor invasiveness/drug resistance. To tailor treatments more effectively to individual patients, it is important to define tumor growth and metastasis at molecular levels. SDCBP is highly overexpressed and associated with a strikingly poor prognosis in breast cancer. However the post transcriptional regulation of SDCBP overexpression remains to be an unexplored area. Our study reveals that miR-216b directly regulates SDCBP expression by binding to its 3′UTR region. miR-216b is a tumor suppressive miRNA and it is underexpressed during metastatic breast cancer. Consequently, overexpression of miR-216b resulted in decreased proliferation, migration and invasion in BC cell lines by modulating the expression of SDCBP. Inhibition of miR-216b divergent the tumor suppressive role by inducing the growth proliferation, migration and invasion in vitro. There is therefore a negative correlation between the expression of miR-216b and its target gene SDCBP in the BC tissue samples as well as cell lines. Simultaneous expression of miR-216b and SDCBP rescued the growth, migration and invasion effect suggesting that tumor suppressive action of miR-216b may be directly mediated by SDCBP. In summary, the study identifies miR-216b as a regulator of SDCBP expression in breast cancer which can potentially be targeted for developing newer therapies for the effective treatment of this killer disease.

miR-101 suppresses HBV replication and expression by targeting FOXO1 in hepatoma carcinoma cell lines.

Science.gov (United States)

Wang, Yanjing; Tian, Hui

2017-05-20

microRNAs (miRNAs) have been identified to participate in the progression of cancers and in the infection of viruses. miR-101 expression has been found to be suppressed by HBV, however, the regulatory relationship between miR-101 and HBV replication remains elusive. In this report, miR-101 was significantly downregulated in HepG2.2.15 cells with HBV expression. miR-101 overexpression dramatically suppressed HBV replication and expression. Oppositely, overexpression of FOXO1 significantly promoted HBV replication and expression. Moreover, luciferase reporter analysis, qRT-PCR analysis and western blot assay confirmed that FOXO1 was a functional target of miR-101. Furthermore, restored FOXO1 expression abolished the inhibitory effect of miR-101 overexpression on HBV replication and expression in HepG2.2.15 cells. Our data suggested that miR-101 suppressed HBV replication and expression partially by targeting FOXO1, providing new insights into the molecular mechanisms of miR-101 in HBV-host interactions and a promising therapeutic target for HBV replication. Copyright © 2017 Elsevier Inc. All rights reserved.

MiR-564 functions as a tumor suppressor in human lung cancer by targeting ZIC3

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Yang, Bin [Department of Oncology, Hubei Cancer Hospital, Wuhan, Hubei 430079 (China); Jia, Lin [Department of Nephrology, The Central Hospital of Wuhan, Wuhan, Hubei 430079 (China); Guo, Qiaojuan [Department of Radiation Oncology, Fujian Provincial Cancer Hospital, Provincial Clinical College of Fujian Medical University, Fuzhou, Fujian 350000 (China); Ren, Hui; Hu, Desheng; Zhou, Xiaoyi; Ren, Qingrong [Department of Oncology, Hubei Cancer Hospital, Wuhan, Hubei 430079 (China); Hu, Yanping, E-mail: huyp1989@163.com [Department of Oncology, Hubei Cancer Hospital, Wuhan, Hubei 430079 (China); Xie, Tao, E-mail: xietao930@hotmail.com [Department of Radiation Oncology, Hubei Cancer Hospital, Wuhan, Hubei 430079 (China)

2015-11-27

Although miR-564 was reported to be dysregulated in human malignancy, the function and mechanism of miR-564 in tumorigenesis remains unknown. In the present study, we found that miR-564 frequently downregulated in lung cancer cells and significantly inhibited cell proliferation, cell cycle progression, motility, and the tumorigenicity of lung cancer cells. Moreover, we identified zic family member 3 (ZIC3) as a direct target of miR-564. ZIC3 overexpression impaired the suppressive effects of miR-564 on the capacity of lung cancer cells for proliferation and motility. Finally, we detected the expression level of miR-564 and ZIC3 protein in tissue specimens, and found a significant negative correlation between them. Patients with low levels of miR-564 showed a poorer overall survival. Taken together, our present study revealed the tumor suppressor role of miR-564, indicating restoration of miR-564 as a potential therapeutic strategy for the treatment of lung cancer. - Highlights: • MiR-564 inhibits cancer cell proliferation, cell cycle progression, migration, and invasion. • miR-564 suppresses the tumorigenicity of lung cancer cell in vivo. • ZIC3 is a direct and functional target of miR-564. • The expression of miR-564 was negatively correlated with ZIC3 protein in tumors. • Both low miR-564 and high ZIC3 was associated with tumor stage and prognosis.

MiR-564 functions as a tumor suppressor in human lung cancer by targeting ZIC3

International Nuclear Information System (INIS)

Yang, Bin; Jia, Lin; Guo, Qiaojuan; Ren, Hui; Hu, Desheng; Zhou, Xiaoyi; Ren, Qingrong; Hu, Yanping; Xie, Tao

2015-01-01

Although miR-564 was reported to be dysregulated in human malignancy, the function and mechanism of miR-564 in tumorigenesis remains unknown. In the present study, we found that miR-564 frequently downregulated in lung cancer cells and significantly inhibited cell proliferation, cell cycle progression, motility, and the tumorigenicity of lung cancer cells. Moreover, we identified zic family member 3 (ZIC3) as a direct target of miR-564. ZIC3 overexpression impaired the suppressive effects of miR-564 on the capacity of lung cancer cells for proliferation and motility. Finally, we detected the expression level of miR-564 and ZIC3 protein in tissue specimens, and found a significant negative correlation between them. Patients with low levels of miR-564 showed a poorer overall survival. Taken together, our present study revealed the tumor suppressor role of miR-564, indicating restoration of miR-564 as a potential therapeutic strategy for the treatment of lung cancer. - Highlights: • MiR-564 inhibits cancer cell proliferation, cell cycle progression, migration, and invasion. • miR-564 suppresses the tumorigenicity of lung cancer cell in vivo. • ZIC3 is a direct and functional target of miR-564. • The expression of miR-564 was negatively correlated with ZIC3 protein in tumors. • Both low miR-564 and high ZIC3 was associated with tumor stage and prognosis.

miR-130b targets NKD2 and regulates the Wnt signaling to promote proliferation and inhibit apoptosis in osteosarcoma cells

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Li, Zhi [Department of Human Anatomy and Histoembryology, College of Basic Medical Sciences, Jilin University (China); Li, Youjun, E-mail: liyoujunn@126.com [Department of Human Anatomy and Histoembryology, College of Basic Medical Sciences, Jilin University (China); Wang, Nan; Yang, Lifeng; Zhao, Wei; Zeng, Xiandong [Central Hospital Affiliated to Shenyang Medical College (China)

2016-03-18

miR-130b was significantly up-regulated in osteosarcoma (OS) cells. Naked cuticle homolog 2 (NKD2) inhibited tumor growth and metastasis in OS by suppressing Wnt signaling. We used three miRNA target analysis tools to identify potential targets of miR-130b, and found that NKD2 is a potential target of miR-130b. Based on these findings, we hypothesize that miR-130b might target NKD2 and regulate the Wnt signaling to promote OS growth. We detected the expression of miR-130b and NKD2 mRNA and protein by quantitative Real-Time PCR (qRT-PCR) and western blot assays, respectively, and found up-regulation of miR-130b and down-regulation of NKD2 mRNA and protein exist in OS cell lines. MTT and flow cytometry assays showed that miR-130b inhibitors inhibit proliferation and promote apoptosis in OS cells. Furthermore, we showed that NKD2 is a direct target of miR-130b, and miR-130b regulated proliferation and apoptosis of OS cells by targeting NKD2. We further investigated whether miR-130b and NKD2 regulate OS cell proliferation and apoptosis by inhibiting Wnt signaling, and the results confirmed our speculation that miR-130b targets NKD2 and regulates the Wnt signaling to promote proliferation and inhibit apoptosis of OS cells. These findings will offer new clues for OS development and progression, and novel potential therapeutic targets for OS. - Highlights: • miR-130b is up-regulated and NKD2 is down-regulated in osteosarcoma cell lines. • Down-regulation of miR-130b inhibits proliferation of osteosarcoma cells. • Down-regulation of miR-130b promotes apoptosis of osteosarcoma cells. • miR-130b directly targets NKD2. • NKD2 regulates OS cell proliferation and apoptosis by inhibiting the Wnt signaling.

miR-130b targets NKD2 and regulates the Wnt signaling to promote proliferation and inhibit apoptosis in osteosarcoma cells

International Nuclear Information System (INIS)

Li, Zhi; Li, Youjun; Wang, Nan; Yang, Lifeng; Zhao, Wei; Zeng, Xiandong

2016-01-01

miR-130b was significantly up-regulated in osteosarcoma (OS) cells. Naked cuticle homolog 2 (NKD2) inhibited tumor growth and metastasis in OS by suppressing Wnt signaling. We used three miRNA target analysis tools to identify potential targets of miR-130b, and found that NKD2 is a potential target of miR-130b. Based on these findings, we hypothesize that miR-130b might target NKD2 and regulate the Wnt signaling to promote OS growth. We detected the expression of miR-130b and NKD2 mRNA and protein by quantitative Real-Time PCR (qRT-PCR) and western blot assays, respectively, and found up-regulation of miR-130b and down-regulation of NKD2 mRNA and protein exist in OS cell lines. MTT and flow cytometry assays showed that miR-130b inhibitors inhibit proliferation and promote apoptosis in OS cells. Furthermore, we showed that NKD2 is a direct target of miR-130b, and miR-130b regulated proliferation and apoptosis of OS cells by targeting NKD2. We further investigated whether miR-130b and NKD2 regulate OS cell proliferation and apoptosis by inhibiting Wnt signaling, and the results confirmed our speculation that miR-130b targets NKD2 and regulates the Wnt signaling to promote proliferation and inhibit apoptosis of OS cells. These findings will offer new clues for OS development and progression, and novel potential therapeutic targets for OS. - Highlights: • miR-130b is up-regulated and NKD2 is down-regulated in osteosarcoma cell lines. • Down-regulation of miR-130b inhibits proliferation of osteosarcoma cells. • Down-regulation of miR-130b promotes apoptosis of osteosarcoma cells. • miR-130b directly targets NKD2. • NKD2 regulates OS cell proliferation and apoptosis by inhibiting the Wnt signaling.

The Micro-RNA172c-APETALA2-1 Node as a Key Regulator of the Common Bean-Rhizobium etli Nitrogen Fixation Symbiosis1[OPEN

Science.gov (United States)

Nova-Franco, Bárbara; Íñiguez, Luis P.; Valdés-López, Oswaldo; Leija, Alfonso; Fuentes, Sara I.; Ramírez, Mario; Paul, Sujay

2015-01-01

Micro-RNAs are recognized as important posttranscriptional regulators in plants. The relevance of micro-RNAs as regulators of the legume-rhizobia nitrogen-fixing symbiosis is emerging. The objective of this work was to functionally characterize the role of micro-RNA172 (miR172) and its conserved target APETALA2 (AP2) transcription factor in the common bean (Phaseolus vulgaris)-Rhizobium etli symbiosis. Our expression analysis revealed that mature miR172c increased upon rhizobial infection and continued increasing during nodule development, reaching its maximum in mature nodules and decaying in senescent nodules. The expression of AP2-1 target showed a negative correlation with miR172c expression. A drastic decrease in miR172c and high AP2-1 mRNA levels were observed in ineffective nodules. Phenotypic analysis of composite bean plants with transgenic roots overexpressing miR172c or a mutated AP2-1 insensitive to miR172c cleavage demonstrated the pivotal regulatory role of the miR172 node in the common bean-rhizobia symbiosis. Increased miR172 resulted in improved root growth, increased rhizobial infection, increased expression of early nodulation and autoregulation of nodulation genes, and improved nodulation and nitrogen fixation. In addition, these plants showed decreased sensitivity to nitrate inhibition of nodulation. Through transcriptome analysis, we identified 114 common bean genes that coexpressed with AP2-1 and proposed these as being targets for transcriptional activation by AP2-1. Several of these genes are related to nodule senescence, and we propose that they have to be silenced, through miR172c-induced AP2-1 cleavage, in active mature nodules. Our work sets the basis for exploring the miR172-mediated improvement of symbiotic nitrogen fixation in common bean, the most important grain legume for human consumption. PMID:25739700

miR-203 inhibits melanoma invasive and proliferative abilities by targeting the polycomb group gene BMI1

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Chang, Xiao [Department of Dermatology and Venereal Disease, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China); Sun, Yong [Department of Burn and Plastic Surgery, Huai’an First People’s Hospital, Nanjing Medical University, Huai’an 223300 (China); Han, Siqi [Department of Medical Oncology, Jinling Hospital, Nanjing 210002 (China); Zhu, Wei [Department of Dermatology and Venereal Disease, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China); Zhang, Haiping, E-mail: zhanghaiping_2000@163.com [Department of Dermatology and Venereal Disease, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China); Lian, Shi, E-mail: lianshi_2020@163.com [Department of Dermatology and Venereal Disease, Capital Medical University, Beijing 100069 (China)

2015-01-02

Highlights: • First reported deregulation of miR-203 and up-regulation of BMI1 in metastatic melanoma. • miR-203 decreased BMI1 expression by directly binding to 3′UTR. • Further found miR-203 overexpression suppressed cell invasion and stemness. • Re-expression of BMI1 rescued miR-203-mediated suppression. • miR-203-BMI1 axis may be potential therapeutic targets of melanoma metastasis. - Abstract: Metastasis is the major problem in malignant melanoma, posing a therapeutic challenge to clinicians. The investigation of the underlying mechanism driving this progress remains a large unmet need. In this study, we revealed a miR-203-BMI1 axis that regulated melanoma metastasis. We found significantly deregulation of miR-203 and up-regulation of BMI1 in melanoma, particularly in metastatic melanoma. An inverse correlation between the levels of miR-203 and BMI1 was further observed in melanoma tissues and cell lines. We also identified BMI1 as a downstream target gene of miR-203, which bound to the 3′UTR of BMI1. Overexpression of miR-203 was associated with decreased BMI1 expression and impaired cell invasion and tumor sphere formation activities. Re-expression of BMI1 markedly rescued miR-203-mediated suppression of these events. Taken together, our results demonstrated that miR-203 regulated melanoma invasive and proliferative abilities in part by targeting BMI1, providing new insights into potential mechanisms of melanoma metastasis.

Non-canonical microRNAs miR-320 and miR-702 promote proliferation in Dgcr8-deficient embryonic stem cells

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Kim, Byeong-Moo [Department of Medicine/GI Unit, Massachusetts General Hospital, Boston, MA 02114 (United States); Department of Medicine, Harvard Medical School, Boston, MA 02115 (United States); Choi, Michael Y., E-mail: mchoi@partners.org [Department of Medicine/GI Unit, Massachusetts General Hospital, Boston, MA 02114 (United States); Department of Medicine, Harvard Medical School, Boston, MA 02115 (United States); Harvard Stem Cell Institute, Boston, MA 02114 (United States)

2012-09-21

Highlights: Black-Right-Pointing-Pointer Embryonic stem cells (ESCs) lacking non-canonical miRNAs proliferate slower. Black-Right-Pointing-Pointer miR-320 and miR-702 are two non-canonical miRNAs expressed in ESCs. Black-Right-Pointing-Pointer miR-320 and miR-702 promote proliferation of Dgcr8-deficient ESCs. Black-Right-Pointing-Pointer miR-320 targets p57 and helps to release Dgcr8-deficient ESCs from G1 arrest. Black-Right-Pointing-Pointer miR-702 targets p21 and helps to release Dgcr8-deficient ESCs from G1 arrest. -- Abstract: MicroRNAs are known to contribute significantly to stem cell phenotype by post-transcriptionally regulating gene expression. Most of our knowledge of microRNAs comes from the study of canonical microRNAs that require two sequential cleavages by the Drosha/Dgcr8 heterodimer and Dicer to generate mature products. In contrast, non-canonical microRNAs bypass the cleavage by the Drosha/Dgcr8 heterodimer within the nucleus but still require cytoplasmic cleavage by Dicer. The function of non-canonical microRNAs in embryonic stem cells (ESCs) remains obscure. It has been hypothesized that non-canonical microRNAs have important roles in ESCs based upon the phenotypes of ESC lines that lack these specific classes of microRNAs; Dicer-deficient ESCs lacking both canonical and non-canonical microRNAs have much more severe proliferation defect than Dgcr8-deficient ESCs lacking only canonical microRNAs. Using these cell lines, we identified two non-canonical microRNAs, miR-320 and miR-702, that promote proliferation of Dgcr8-deficient ESCs by releasing them from G1 arrest. This is accomplished by targeting the 3 Prime -untranslated regions of the cell cycle inhibitors p57 and p21 and thereby inhibiting their expression. This is the first report of the crucial role of non-canonical microRNAs in ESCs.

miR-103 Promotes Neurite Outgrowth and Suppresses Cells Apoptosis by Targeting Prostaglandin-Endoperoxide Synthase 2 in Cellular Models of Alzheimer's Disease.

Science.gov (United States)

Yang, Hui; Wang, Hongcai; Shu, Yongwei; Li, Xuling

2018-01-01

miR-103 has been reported to be decreased in brain of transgenic mouse model of Alzheimer's disease (AD) and in cerebrospinal fluid (CSF) of AD patients, while the detailed mechanism of its effect on AD is obscure, thus this study aimed to investigate the effect of miR-103 expression on neurite outgrowth and cells apoptosis as well as its targets in cellular models of AD. Blank mimic (NC1-mimic), miR-103 mimic, blank inhibitor (NC2-mimic) and miR-103 inhibitor plasmids were transferred into PC12 cellular AD model and Cellular AD model of cerebral cortex neurons which were established by Aβ1-42 insult. Rescue experiment was subsequently performed by transferring Prostaglandin-endoperoxide synthase 2 (PTGS2) and miR-103 mimic plasmid. mRNA and protein expressions were detected by qPCR and Western Blot assays. Total neurite outgrowth was detected by microscope, cells apoptosis was determined by Hoechst/PI assay, and apoptotic markers Caspase 3 and p38 expressions were determined by Western Blot assay. In both PC12 and cerebral cortex neurons cellular AD models, miR-103 mimic increases the total neurite outgrowth compared with NC1-mimic, while miR-103 inhibitor decreases the total neurite outgrowth than NC2-inhibitor. The apoptosis rate was decreased in miR-103 mimic group than NC1-mimic group while increased in miR-103 inhibitor group than NC2-inhibitor group. PTGS2, Adisintegrin and metalloproteinase 10 (ADAM10) and neprilysin (NEP) were selected as target genes of miR-103 by bioinformatics analysis. And PTGS2 was found to be conversely regulated by miR-103 expression while ADAM10 and NEP were not affected. After transfection by PTGS2 and miR-103 mimic plasmid in PC12 cellular AD model, the total neurite growth was shortened compared with miR-103 mimic group, and cells apoptosis was enhanced which indicated PTGS2 mimic attenuated the influence of miR-103 mimic on progression of AD. In conclusion, miR-103 promotes total neurite outgrowth and inhibits cells apoptosis

miR-198 Represses the Proliferation of HaCaT Cells by Targeting Cyclin D2

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Jian Wang

2015-07-01

Full Text Available Background: MiR-198 has been considered as an inhibitor of cell proliferation, invasion, migration and a promoter of apoptosis in most cancer cells, while its effect on non-cancer cells is poorly understood. Methods: The effect of miR-198 transfection on HaCaT cell proliferation was firstly detected using Cell Count Kit-8 and the cell cycle progression was analyzed by flow cytometry. Using bioinformatics analyses and luciferase assay, a new target of miR-198 was searched and identified. Then, the effect of the new target gene of miR-198 on cell proliferation and cell cycle was also detected. Results: Here we showed that miR-198 directly bound to the 3′-UTR of CCND2 mRNA, which was a key regulator in cell cycle progression. Overexpressed miR-198 repressed CCND2 expression at mRNA and protein levels and subsequently led to cell proliferation inhibition and cell cycle arrest in the G1 phase. Transfection ofSiCCND2 in HaCaT cells showed similar inhibitory effects on cell proliferation and cell cycle progression. Conclusion: In conclusion, we have identified that miR-198 inhibited HaCaT cell proliferation by directly targeting CCND2.

[miR-182 promotes cell proliferation of cervical cancer cells by targeting adenomatous polyposis coli (APC) gene].

Science.gov (United States)

Li, Pei; Hu, Jing; Zhang, Ying; Li, Jianping; Dang, Yunzhi; Zhang, Rui; Wei, Lichun; Shi, Mei

2018-02-01

Objective To investigate the role and mechanism of microRNA-182 (miR-182) in the proliferation of cervical cancer cells. Methods With liposome-mediated transient transfection method, the level of miR-182 in HeLa and SiHa cells was increased or decreased. CCK-8 assay and colony formation assay were used to observe the effect of miR-182 on the proliferation of cervical cancer cells. Using bioinformatics predictions, real-time quantitative PCR, and dual luciferase reporter assay, we clarified the role of miR-182 in posttranscriptional regulation of adenomatous polyposis coli (APC) gene and its effect on the downstream molecules (c-Myc and cyclin D1) of Wnt singling pathway. Results Up-regulation of miR-182 significantly promoted the proliferation of cervical cancer cells, while down-regulation of miR-182 significantly inhibited the proliferation of cervical cancer cells. Over-expression of miR-182 inhibited the expression of APC gene in cervical cancer cells and the regulation of miR-182 affected the expression of canonical Wnt signaling pathway downstream molecules in cervical cancer cells. Conclusion The miR-182 stimulates canonical Wnt signaling pathway by targeting APC gene and enhances the proliferation of cervical cancer cells.

miR-411-5p inhibits proliferation and metastasis of breast cancer cell via targeting GRB2

International Nuclear Information System (INIS)

Zhang, Yunda; Xu, Guoxing; Liu, Gang; Ye, Yongzhi; Zhang, Chuankai; Fan, Chuannan; Wang, Haibin; Cai, Huali; Xiao, Rui; Huang, Zhengjie; Luo, Qi

2016-01-01

miR-411-5p (previously called miR-411) is severely involved in human diseases, however, the relationship between miR-411-5p and breast cancer has not been investigated thoroughly. Here, we found that the expression of miR-411-5p was downregulated in breast cancer tissues compared with their matched adjacent non-neoplastic tissues. In addition, the expression of miR-411-5p was also lower in breast cancer cell lines in contrast with MCF-10A. Moreover, we investigated the target and mechanism of miR-411-5p in breast cancer using mimic and inhibitor, and demonstrated the involvement of GRB2 and Ras activation. Ectopic expression of miR-411-5p suppressed the breast cancer cell proliferation, migration and invasion while low expression of miR-411-5p exhibited the opposite effect. Furthermore, GRB2 was demonstrated to be significantly overexpressed in breast cancer tissues compared with normal tissues, and low expression of GRB2 had a longer overall survival compared with high expression of GRB2 in breast cancer. In general, our study shed light on the miR-411-5p related mechanism in the progression of breast cancer and, miR-411-5p/GRB2/Ras axis is potential to be molecular target for breast cancer therapy. - Highlights: • miR-411-5p is downregulated in breast cancer tissues and cell lines. • miR-411-5p inhibits breast cancer cells growth, migration and invasion in vitro. • GRB2 is a direct target of miR-411-5p in breast cancer. • GRB2 is overexpressed in breast cancer and associates with disease outcome. • miR-411-5p suppresses breast cancer progression though GRB2-SOS-Ras pathway.

miR-411-5p inhibits proliferation and metastasis of breast cancer cell via targeting GRB2

Energy Technology Data Exchange (ETDEWEB)

Zhang, Yunda [Department of Gastrointestinal Surgery, First Affiliated Hospital of Xiamen University, Xiamen 361003 (China); State Key Laboratory of Cellular Stress Biology, Innovation Center for Cell Signaling Network, School of Life Sciences, Xiamen University, Xiamen 361102 (China); Xu, Guoxing [Department of Gastrointestinal Surgery, First Affiliated Hospital of Xiamen University, Xiamen 361003 (China); Department of Gastrointestinal Surgery, First Clinical Medical College of Fujian Medical University, Fuzhou 350005 (China); Liu, Gang; Ye, Yongzhi [Department of Gastrointestinal Surgery, First Affiliated Hospital of Xiamen University, Xiamen 361003 (China); Zhang, Chuankai [Department of Gastrointestinal Surgery, First Affiliated Hospital of Xiamen University, Xiamen 361003 (China); State Key Laboratory of Cellular Stress Biology, Innovation Center for Cell Signaling Network, School of Life Sciences, Xiamen University, Xiamen 361102 (China); Fan, Chuannan [State Key Laboratory of Cellular Stress Biology, Innovation Center for Cell Signaling Network, School of Life Sciences, Xiamen University, Xiamen 361102 (China); Wang, Haibin; Cai, Huali; Xiao, Rui [Department of Gastrointestinal Surgery, First Affiliated Hospital of Xiamen University, Xiamen 361003 (China); Department of Gastrointestinal Surgery, First Clinical Medical College of Fujian Medical University, Fuzhou 350005 (China); Huang, Zhengjie, E-mail: huangzhengjie@xmu.edu.cn [Department of Gastrointestinal Surgery, First Affiliated Hospital of Xiamen University, Xiamen 361003 (China); Department of Gastrointestinal Surgery, First Clinical Medical College of Fujian Medical University, Fuzhou 350005 (China); Luo, Qi, E-mail: luoqixmzsh@126.com [Department of Gastrointestinal Surgery, First Affiliated Hospital of Xiamen University, Xiamen 361003 (China); Department of Gastrointestinal Surgery, First Clinical Medical College of Fujian Medical University, Fuzhou 350005 (China)

2016-08-05

miR-411-5p (previously called miR-411) is severely involved in human diseases, however, the relationship between miR-411-5p and breast cancer has not been investigated thoroughly. Here, we found that the expression of miR-411-5p was downregulated in breast cancer tissues compared with their matched adjacent non-neoplastic tissues. In addition, the expression of miR-411-5p was also lower in breast cancer cell lines in contrast with MCF-10A. Moreover, we investigated the target and mechanism of miR-411-5p in breast cancer using mimic and inhibitor, and demonstrated the involvement of GRB2 and Ras activation. Ectopic expression of miR-411-5p suppressed the breast cancer cell proliferation, migration and invasion while low expression of miR-411-5p exhibited the opposite effect. Furthermore, GRB2 was demonstrated to be significantly overexpressed in breast cancer tissues compared with normal tissues, and low expression of GRB2 had a longer overall survival compared with high expression of GRB2 in breast cancer. In general, our study shed light on the miR-411-5p related mechanism in the progression of breast cancer and, miR-411-5p/GRB2/Ras axis is potential to be molecular target for breast cancer therapy. - Highlights: • miR-411-5p is downregulated in breast cancer tissues and cell lines. • miR-411-5p inhibits breast cancer cells growth, migration and invasion in vitro. • GRB2 is a direct target of miR-411-5p in breast cancer. • GRB2 is overexpressed in breast cancer and associates with disease outcome. • miR-411-5p suppresses breast cancer progression though GRB2-SOS-Ras pathway.

The key role of miR-21-regulated SOD2 in the medium-mediated bystander responses in human fibroblasts induced by α-irradiated keratinocytes.

Science.gov (United States)

Tian, Wenqian; Yin, Xiaoming; Wang, Longxiao; Wang, Jingdong; Zhu, Wei; Cao, Jianping; Yang, Hongying

2015-10-01

Radiation-induced bystander effect (RIBE) is well accepted in the radiation research field by now, but the underlying molecular mechanisms for better understanding this phenomenon caused by intercellular communication and intracellular signal transduction are still incomplete. Although our previous study has demonstrated an important role of miR-21 of unirradiated bystander cells in RIBEs, the direct evidence for the hypothesis that RIBE is epigenetically regulated is still limited and how miR-21 mediates RIBEs is unknown. Reactive oxygen species (ROS) have been demonstrated to be involved in RIBEs, however, the roles of anti-oxidative stress system of cells in RIBEs are unclear. Using transwell insert co-culture system, we investigated medium-mediated bystander responses in WS1 human fibroblasts after co-culture with HaCaT keratinocytes traversed by α-particles. Results showed that the ROS levels in unirradiated bystander WS1 cells were significantly elevated after 30min of co-culture, and 53BP1 foci, a surrogate marker of DNA damage, were obviously induced after 3h of co-culture. This indicates the occurrence of oxidative stress and DNA damage in bystander WS1 cells after co-culture with irradiated keratinocytes. Furthermore, the expression of miR-21 was increased in bystander WS1 cells, downregulation of miR-21 eliminated the bystander responses, overexpression of miR-21 alone could induce bystander-like oxidative stress and DNA damage in WS1 cells. These data indicate an important mediating role of miR-21 in RIBEs. In addition, MnSOD or SOD2 in WS1 cells was involved in the bystander effects, overexpression of SOD2 abolished the bystander oxidative stress and DNA damage, indicating that SOD2 was critical to the induction of RIBEs. Moreover, we found that miR-21 regulated SOD2, suggesting that miR-21 might mediate bystander responses through its regulation on SOD2. In conclusion, this study revealed a profound role of miR-21-regulated SOD2 of unirradiated WS1

miR-503 suppresses tumor cell proliferation and metastasis by directly targeting RNF31 in prostate cancer

International Nuclear Information System (INIS)

Guo, Jia; Liu, Xiuheng; Wang, Min

2015-01-01

Microarray data analyses were performed to search for metastasis-associated oncogenes in prostate cancer (PCa). RNF31 mRNA expressions in tumor tissues and benign prostate tissues were evaluated. The RNF31 protein expression levels were also analyzed by western blot and immunohistochemistry. Luciferase reporter assays were used to identify miRNAs that can regulate RNF31. The effect of RNF31 on PCa progression was studied in vitro and in vivo. We found that RNF31 was significantly increased in PCa and its expression level was highly correlated with seminal vesicle invasion, clinical stage, prostate specific antigen (PSA) level, Gleason score, and BCR. Silence of RNF31 suppressed PCa cell proliferation and metastasis in vitro and in vivo. miR-503 can directly regulate RNF31. Enforced expression of miR-503 inhibited the expression of RNF31 significantly and the restoration of RNF31 expression reversed the inhibitory effects of miR-503 on PCa cell proliferation and metastasis. These findings collectively indicated an oncogene role of RNF31 in PCa progression which can be regulated by miR-503, suggesting that RNF31 could serve as a potential prognostic biomarker and therapeutic target for PCa. - Highlights: • RNF31 is a potential metastasis associated gene and is associated with prostate cancer progression. • Silence of RNF31 inhibits PCa cell colony formation, migration and invasion. • RNF31 as a direct target of miR-503. • miR-503 can regulate cell proliferation, invasion and migration by targeting RNF31. • RNF31 plays an important role in PCa growth and metastasis in vivo

miR-503 suppresses tumor cell proliferation and metastasis by directly targeting RNF31 in prostate cancer

Energy Technology Data Exchange (ETDEWEB)

Guo, Jia; Liu, Xiuheng, E-mail: l_xiuheng@163.com; Wang, Min

2015-09-04

Microarray data analyses were performed to search for metastasis-associated oncogenes in prostate cancer (PCa). RNF31 mRNA expressions in tumor tissues and benign prostate tissues were evaluated. The RNF31 protein expression levels were also analyzed by western blot and immunohistochemistry. Luciferase reporter assays were used to identify miRNAs that can regulate RNF31. The effect of RNF31 on PCa progression was studied in vitro and in vivo. We found that RNF31 was significantly increased in PCa and its expression level was highly correlated with seminal vesicle invasion, clinical stage, prostate specific antigen (PSA) level, Gleason score, and BCR. Silence of RNF31 suppressed PCa cell proliferation and metastasis in vitro and in vivo. miR-503 can directly regulate RNF31. Enforced expression of miR-503 inhibited the expression of RNF31 significantly and the restoration of RNF31 expression reversed the inhibitory effects of miR-503 on PCa cell proliferation and metastasis. These findings collectively indicated an oncogene role of RNF31 in PCa progression which can be regulated by miR-503, suggesting that RNF31 could serve as a potential prognostic biomarker and therapeutic target for PCa. - Highlights: • RNF31 is a potential metastasis associated gene and is associated with prostate cancer progression. • Silence of RNF31 inhibits PCa cell colony formation, migration and invasion. • RNF31 as a direct target of miR-503. • miR-503 can regulate cell proliferation, invasion and migration by targeting RNF31. • RNF31 plays an important role in PCa growth and metastasis in vivo.

miR-409-3p suppresses breast cancer cell growth and invasion by targeting Akt1

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Zhang, Guoqiang [Department of Breast Surgery, Qilu Hospital, Shandong University, Jinan 250012 (China); Department of Thyroid and Breast Surgery, Hospital Affiliated to Binzhou Medical University, 661 Second Huanghe Street, Binzhou 256603 (China); Liu, Zengyan [Department of Hematology, Hospital Affiliated to Binzhou Medical University, 661 Second Huanghe Street, Binzhou 256603 (China); Xu, Hao [Department of General Surgery, The First Affiliated Hospital of Nanjing Medical University, Nanjing 210029 (China); Yang, Qifeng, E-mail: qifengy_sdu1@163.com [Department of Breast Surgery, Qilu Hospital, Shandong University, Jinan 250012 (China)

2016-01-08

Altered levels and functions of microRNAs (miRNAs) are correlated with carcinogenesis. While miR-409-3p has been shown to play important roles in several cancer types, its function in the context of breast cancer (BC) remains unknown. In this study, miR-409-3p was significantly downregulated in BC tissues and cell lines, compared with the corresponding control counterparts. Overexpression of miR-409-3p inhibited BC cell proliferation, migration and invasion in vitro and suppressed tumor growth in vivo. Notably, miR-409-3p induced downregulation of Akt1 protein through binding to its 3′ untranslated region (UTR). Conversely, restoring Akt1 expression rescued the suppressive effects of miR-409-3p. Our data collectively indicate that miR-409-3p functions as a tumor suppressor in BC through downregulating Akt1, supporting the targeting of the novel miR-409-3p/Akt1 axis as a potentially effective therapeutic approach for BC. - Highlights: • miR-409-3p inhibits proliferation, migration and invasion of BC cells. • miR-409-3p suppresses tumor growth in nude mice. • Akt1 is a direct downstream target of miR-409-3p. • Ectopic expression of Akt1 reverses the effects of miR-409-3p on cell proliferation, migration and invasion.

Dysregulated miR-127-5p contributes to type II collagen degradation by targeting matrix metalloproteinase-13 in human intervertebral disc degeneration.

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Hua, Wen-Bin; Wu, Xing-Huo; Zhang, Yu-Kun; Song, Yu; Tu, Ji; Kang, Liang; Zhao, Kang-Cheng; Li, Shuai; Wang, Kun; Liu, Wei; Shao, Zeng-Wu; Yang, Shu-Hua; Yang, Cao

2017-08-01

Intervertebral disc degeneration (IDD) is a chronic disease associated with the degradation of extracellular matrix (ECM). Matrix metalloproteinase (MMP)-13 is a major enzyme that mediates the degradation of ECM components. MMP-13 has been predicted to be a potential target of miR-127-5p. However, the exact function of miR-127-5p in IDD is still unclear. We designed this study to evaluate the correlation between miR-127-5p level and the degeneration of human intervertebral discs and explore the potential mechanisms. miR-127-5p levels and MMP-13 mRNA levels were detected by quantitative real-time polymerase chain reaction (qPCR). To determine whether MMP-13 is a target of miR-127-5p, dual luciferase reporter assays were performed. miR-127-5p mimic and miR-127-5p inhibitor were used to overexpress or downregulate miR-127-5p expression in human NP cells, respectively. Small interfering RNA (siRNA) was used to knock down MMP-13 expression in human NP cells. Type II collagen expression in human NP cells was detected by qPCR, western blotting, and immunofluorescence staining. We confirmed that miR-127-5p was significantly downregulated in nucleus pulposus (NP) tissue of degenerative discs and its expression was inversely correlated with MMP-13 mRNA levels. We reveal that MMP-13 may act as a target of miR-127-5p. Expression of miR-127-5p was inversely correlated with type II collagen expression in human NP cells. Moreover, suppression of MMP-13 expression by siRNA blocked downstream signaling and increased type II collagen expression. Dysregulated miR-127-5p contributed to the degradation of type II collagen by targeting MMP-13 in human IDD. Our findings highlight that miR-127-5p may serve as a new therapeutic target in IDD. Copyright © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

Dysregulated Expression of the MicroRNA miR-137 and Its Target YBX1 Contribute to the Invasive Characteristics of Malignant Pleural Mesothelioma.

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Johnson, Thomas G; Schelch, Karin; Cheng, Yuen Y; Williams, Marissa; Sarun, Kadir H; Kirschner, Michaela B; Kao, Steven; Linton, Anthony; Klebe, Sonja; McCaughan, Brian C; Lin, Ruby C Y; Pirker, Christine; Berger, Walter; Lasham, Annette; van Zandwijk, Nico; Reid, Glen

2018-02-01

Malignant pleural mesothelioma (MPM) is an aggressive malignancy linked to asbestos exposure. On a genomic level, MPM is characterized by frequent chromosomal deletions of tumor suppressors, including microRNAs. MiR-137 plays a tumor suppressor role in other cancers, so the aim of this study was to characterize it and its target Y-box binding protein 1 (YBX1) in MPM. Expression, methylation, and copy number status of miR-137 and its host gene MIR137HG were assessed by polymerase chain reaction. Luciferase reporter assays confirmed a direct interaction between miR-137 and Y-box binding protein 1 gene (YBX1). Cells were transfected with a miR-137 inhibitor, miR-137 mimic, and/or YBX1 small interfering RNA, and growth, colony formation, migration and invasion assays were conducted. MiR-137 expression varied among MPM cell lines and tissue specimens, which was associated with copy number variation and promoter hypermethylation. High miR-137 expression was linked to poor patient survival. The miR-137 inhibitor did not affect target levels or growth, but interestingly, it increased miR-137 levels by means of mimic transfection suppressed growth, migration, and invasion, which was linked to direct YBX1 downregulation. YBX1 was overexpressed in MPM cell lines and inversely correlated with miR-137. RNA interference-mediated YBX1 knockdown significantly reduced cell growth, migration, and invasion. MiR-137 can exhibit a tumor-suppressive function in MPM by targeting YBX1. YBX1 knockdown significantly reduces tumor growth, migration, and invasion of MPM cells. Therefore, YBX1 represents a potential target for novel MPM treatment strategies. Copyright © 2017 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.

Chondroitin sulfate-functionalized polyamidoamine as a tumor-targeted carrier for miR-34a delivery.

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Chen, Wenqi; Liu, Yong; Liang, Xiao; Huang, Yu; Li, Quanshun

2017-07-15

Chondroitin sulfate (CS) was modified on a polyamidoamine dendrimer (PAMAM) through Michael addition to construct a tumor-targeted carrier CS-PAMAM for miR-34a delivery. The derivative CS-PAMAM was demonstrated to achieve an efficient cellular uptake of miR-34a in a CD44-dependent endocytosis way and further facilitate the endosomal escape of miR-34a after 4h. Through the miR-34a delivery, obvious inhibition of cell proliferation could be detected which was attributed to the enhancement of cell apoptosis and cell cycle arrest, and meanwhile the cell migration and invasion has been observed to be inhibited. Finally, the intravenous injection of CS-PAMAM/miR-34a formulation into mice bearing human lung adenocarcinoma cell A549 xenografts could efficiently inhibit the tumor growth and induce the tumor apoptosis owing to the enhanced accumulation of miR-34a in tumor tissue. Overall, CS-PAMAM is potential to be used as a tumor-targeted oligonucleotide carrier for achieving tumor gene therapy. The cationic dendrimer PAMAM was modified by chondroitin sulfate (CS) through Michael addition to construct a tumor-targeted carrier CS-PAMAM for miR-34a delivery. The introduction of CS could achieve an efficient cellular uptake and intracellular transfection of miR-34a in a CD44-dependent endocytosis manner. The miR-34a delivery could execute the anti-proliferation activity by simultaneously inducing cell apoptosis and cell cycle arrest, and also the anti-migration activity. The CS-PAMAM-mediated systemic delivery of miR-34a showed significant inhibition of tumor growth and induction of tumor apoptosis using a mice model of subcutaneously implanted tumors. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Methylation-mediated silencing of miR-124 facilitates chondrogenesis by targeting NFATc1 under hypoxic conditions.

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Gong, Ming; Liang, Tangzhao; Jin, Song; Dai, Xuejun; Zhou, Zhiyu; Gao, Manman; Huang, Sheng; Luo, Jiaquan; Zou, Lijin; Zou, Xuenong

2017-01-01

Chondrogenic differentiation of mesenchymal stem cells is regulated by many different pathways. Recent studies have established that hypoxia and epigenetic alterations potently affect expression of chondrogenesis marker genes. Sox9 is generally regarded as a master regulator of chondrogenesis and microRNA-124 (miRNA-124) regulates gene expression in murine bone marrow-derived mesenchymal stem cells. Therefore, in this study we investigated whether epigenetic regulation of miRNA-124 could affect the expression of Sox9 and thereby regulate chondrogenesis. A cell pellet culture model was used to induce chondrogenesis in C3H10T1/2 cells under hypoxic conditions (2% O 2 ) to determine the effects of hypoxia on miR-124 expression and DNA methylation. The expression of miR-124 was significantly downregulated under hypoxic conditions compared to normoxic conditions (21% O 2 ). The expression of chondrogenesis marker genes was significantly increased under hypoxic conditions. Bisulfite sequencing of the CpG islands in the promoter region of miR-124-3 showed that CpG methylation was significantly increased under hypoxic conditions. Treating the cells with the DNA demethylating agent 5'-AZA significantly increased miR-124 expression and decreased expression of markers of chondrogenesis. Overexpressing miR-124 under hypoxic conditions inhibited NFATc1 reporter activity. NFATc1 was shown to bind to the promoter region of Sox9. Taken together, our data provide evidence that miR-124 acts as an inhibitor of NFATc1. Under hypoxic conditions when miR-124 is downregulated by methylation of CpG islands in the promoter, NFATc1 can bind to the Sox9 promoter and induce the expression of Sox9 leading to chondrogenesis. These results support the role of epigenetic regulation in establishing and maintaining a chondrogenic phenotype.

MiR-422a targets MAPKK6 and regulates cell growth and apoptosis in colorectal cancer cells.

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Li, Peng; Li, Qingmin; Zhang, Yanqiang; Sun, Shaojun; Liu, Shuntao; Lu, Zhaoxi

2018-03-19

The important role of miR-422a in tumor has been reported in several studies. Recent research discovered that the expression of miR-422a was significantly decreased in colorectal cancer tissues, providing miR-422a as a tumor suppressor in CRC. However, the concrete mechanism of miR-422a regulating CRC cell is still unclear. In this study, we demonstrated that miR-422a could inhibit CRC cell growth and promote cell apoptosis via in vitro analyses. Moreover, computational methods were adopted to identify the targets of miR-422a. We found MAPKK6 was the direct target of miR-422a. Consequently, we further elucidated that miR-422a inhibited CRC cell growth and induced cell apoptosis by inhibiting p38/MAPK pathway. Besides that, we established the tumor xenograft model using nude mice and the inhibitory effects on tumor volumes and weights by miR-422a mimic transfection were also detected. Taken together, these findings demonstrated miR-422a exerted anti-cancer activities on CRC, which could be potentially used for CRC prognosis prediction and treatment. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

miR-497 suppresses epithelial–mesenchymal transition and metastasis in colorectal cancer cells by targeting fos-related antigen-1

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Zhang N

2016-10-01

Full Text Available Nan Zhang,1 Quan Shen,2 Pingping Zhang3 1Department of General Surgery, First Affiliated Hospital of Henan University of Chinese Medicine, 2Department of Hepatobiliary Surgery, Henan Provincial People’s Hospital, Zhengzhou, 3Department of Integrated Chinese and Western Medicine, Tianjin First Central Hospital, Tianjin, People’s Republic of China Objective: MicroRNAs have key roles in tumor metastasis. The acquisition of metastatic capability by cancer cells is associated with epithelial–mesenchymal transition (EMT. Here, we describe the role and molecular mechanism of miR-497 in colorectal cancer (CRC cell EMT, migration, and invasion.Methods: Quantitative real-time polymerase chain reaction and Western blot assays were performed to detect the expression levels of miR-497 and Fos-related antigen-1 (Fra-1 in the CRC cells. HCT116 and SW480 cells with miR-497 overexpression or Fra-1 low expression were constructed by lipofection. Target prediction and luciferase reporter assays were performed to investigate whether Fra-1 is one of the targets of miR-497. Western blot and Transwell assays were performed to detect the effects of miR-497 and Fra-1 on CRC cell EMT, migration and invasion.Results: We searched the miRanda, TargetScan, and PicTar databases and found that Fra-1, a key driver of CRC metastasis, is a potential target of miR-497. Quantitative real-time polymerase chain reaction and Western blot analysis verified downregulation of miR-497 and upregulation of Fra-1 in CRC cells. Western blot and Transwell assays showed that overexpression of miR-497 suppresses CRC cell EMT, migration, and invasion. Luciferase gene reporter assay revealed that Fra-1 is a downstream target of miR-497 as miR-497 bound directly to the 3' untranslated region of Fra-1 messenger RNA. An inverse correlation was also found between miR-497 and Fra-1 in HCT116 and SW480 cells. Furthermore, knockdown of Fra-1 recuperated the effects of miR-497 overexpression

MiR-153 targets the nuclear factor-1 family and protects against teratogenic effects of ethanol exposure in fetal neural stem cells

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Pai-Chi Tsai

2014-07-01

Full Text Available Ethanol exposure during pregnancy is an established cause of birth defects, including neurodevelopmental defects. Most adult neurons are produced during the second trimester-equivalent period. The fetal neural stem cells (NSCs that generate these neurons are an important but poorly understood target for teratogenesis. A cohort of miRNAs, including miR-153, may serve as mediators of teratogenesis. We previously showed that ethanol decreased, while nicotine increased miR-153 expression in NSCs. To understand the role of miR-153 in the etiology of teratology, we first screened fetal cortical NSCs cultured ex vivo, by microarray and quantitative RT-PCR analyses, to identify cell-signaling mRNAs and gene networks as important miR-153 targets. Moreover, miR-153 over-expression prevented neuronal differentiation without altering neuroepithelial cell survival or proliferation. Analysis of 3′UTRs and in utero over-expression of pre-miR-153 in fetal mouse brain identified Nfia (nuclear factor-1A and its paralog, Nfib, as direct targets of miR-153. In utero ethanol exposure resulted in a predicted expansion of Nfia and Nfib expression in the fetal telencephalon. In turn, miR-153 over-expression prevented, and partly reversed, the effects of ethanol exposure on miR-153 target transcripts. Varenicline, a partial nicotinic acetylcholine receptor agonist that, like nicotine, induces miR-153 expression, also prevented and reversed the effects of ethanol exposure. These data collectively provide evidence for a role for miR-153 in preventing premature NSC differentiation. Moreover, they provide the first evidence in a preclinical model that direct or pharmacological manipulation of miRNAs have the potential to prevent or even reverse effects of a teratogen like ethanol on fetal development.

miR-19, a component of the oncogenic miR-17∼92 cluster, targets the DNA-end resection factor CtIP

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Hühn, D; Kousholt, A N; Sørensen, Claus Storgaard

2014-01-01

MicroRNA-19 (miR-19) was recently identified as the key oncogenic component of the polycistronic miR-17∼92 cluster, also known as oncomiR-1, which is frequently upregulated or amplified in multiple tumor types. However, the gene targets and the pathways underlying the tumor-promoting activity of mi......R-19 still remain largely elusive. CtIP/RBBP8 promotes DNA-end resection, a critical step in the repair of DNA double-strand breaks (DSBs) by homologous recombination (HR), and is considered to function as a tumor suppressor. In this study, we show that miR-19 downregulates CtIP expression by binding...... to two highly conserved sequences located in the 3'-untranslated region of CtIP mRNA. We further demonstrate that CtIP expression is repressed by miR-19 during continuous genotoxic stress in a p53-dependent manner. Finally, we report that miR-19 impairs CtIP-mediated DNA-end resection, which results...

The long noncoding RNA TUG1 regulates blood-tumor barrier permeability by targeting miR-144.

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Cai, Heng; Xue, Yixue; Wang, Ping; Wang, Zhenhua; Li, Zhen; Hu, Yi; Li, Zhiqing; Shang, Xiuli; Liu, Yunhui

2015-08-14

Blood-tumor barrier (BTB) limits the delivery of chemotherapeutic agent to brain tumor tissues. Long non-coding RNAs (lncRNAs) have been shown to play critical regulatory roles in various biologic processes of tumors. However, the role of lncRNAs in BTB permeability is unclear. LncRNA TUG1 (taurine upregulated gene 1) was highly expressed in glioma vascular endothelial cells from glioma tissues. It also upregulated in glioma co-cultured endothelial cells (GEC) from BTB model in vitro. Knockdown of TUG1 increased BTB permeability, and meanwhile down-regulated the expression of the tight junction proteins ZO-1, occludin, and claudin-5. Both bioinformatics and luciferase reporter assays demonstrated that TUG1 influenced BTB permeability via binding to miR-144. Furthermore, Knockdown of TUG1 also down-regulated Heat shock transcription factor 2 (HSF2), a transcription factor of the heat shock transcription factor family, which was defined as a direct and functional downstream target of miR-144. HSF2 up-regulated the promoter activities and interacted with the promoters of ZO-1, occludin, and claudin-5 in GECs. In conclusion, our results indicate that knockdown of TUG1 increased BTB permeability via binding to miR-144 and then reducing EC tight junction protein expression by targeting HSF2. Thus, TUG1 may represent a useful future therapeutic target for enhancing BTB permeability.

miR-375-3p negatively regulates osteogenesis by targeting and decreasing the expression levels of LRP5 and β-catenin.

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Tianhao Sun

Full Text Available Wnt signaling pathways are essential for bone formation. Previous studies showed that Wnt signaling pathways were regulated by miR-375. Thus, we aim to explore whether miR-375 could affect osteogenesis. In the present study, we investigated the roles of miR-375 and its downstream targets. Firstly, we revealed that miR-375-3p negatively modulated osteogenesis by suppressing positive regulators of osteogenesis and promoting negative regulators of osteogenesis. In addition, the results of TUNEL cell apoptosis assay showed that miR-375-3p induced MC3T3-E1 cell apoptosis. Secondly, miR-375-3p targeted low-density lipoprotein receptor-related protein 5 (LRP5, a co-receptor of the Wnt signaling pathways, and β-catenin as determined by luciferase activity assay, and it decreased the expression levels of LRP5 and β-catenin. Thirdly, the decline of protein levels of β-catenin was determined by immunocytochemistry and immunofluorescence. Finally, silence of LRP5 in osteoblast precursor cells resulted in diminished cell viability and cell proliferation as detected by WST-1-based colorimetric assay. Additionally, all the parameters including the relative bone volume from μCT measurement suggested that LRP5 knockout in mice resulted in a looser and worse-connected trabeculae. The mRNA levels of important negative modulators relating to osteogenesis increased after the functions of LRP5 were blocked in mice. Last but not least, the expression levels of LRP5 increased during the osteogenesis of MC3T3-E1, while the levels of β-catenin decreased in bone tissues from osteoporotic patients with vertebral compression fractures. In conclusion, we revealed miR-375-3p negatively regulated osteogenesis by targeting LRP5 and β-catenin. In addition, loss of functions of LRP5 damaged bone formation in vivo. Clinically, miR-375-3p and its targets might be used as diagnostic biomarkers for osteoporosis and might be also as novel therapeutic agents in osteoporosis

miR-518b Enhances Human Trophoblast Cell Proliferation Through Targeting Rap1b and Activating Ras-MAPK Signal

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Ming Liu

2018-03-01

Full Text Available Preeclampsia is a pregnancy-specific complication defined as newly onset gestational hypertension and proteinuria. Deficiency in placental development is considered as the predominant cause of preeclampsia. Our previous study found that the expression of miR-518b increased significantly in the preeclamptic placentas, indicating the potential participation of this small RNA in the occurrence of preeclampsia. In this study, data analysis using multiple databases predicted Rap1b as a candidate target of miR-518b. An evident decrease in Rap1b expression was observed in preeclamptic placentas when compared with the control placentas, which was negatively correlated with the level of miR-518b. Based on the data of in situ hybridization and immunohistochemistry showing that Rap1b exhibited similar localization with miR-518b in villous cytotrophoblast cells and column trophoblasts, we further explored their function in regulating trophoblast cell proliferation. In HTR8/SVneo cells, exogenous transfection of miR-518b reduced the expression of Rap1b, and dual-luciferase reporter assay validated Rap1b as the direct target of miR-518b. The small RNA could increase the BrdU incorporation and the ratio of cells at S phase, and enhance the phosphorylation of Raf-1 and ERK1/2. Such growth-promoting effect could be efficiently reversed by Rap1b overexpression. The data indicate that miR-518b can promote trophoblast cell proliferation via Rap1b–Ras–MAPK pathway, and the aberrant upregulation of miR-518b in preeclamptic placenta may contribute to the excessive trophoblast proliferation. The study provides new evidence to further understand the etiology of preeclampsia.

miR-16 promotes the apoptosis of human cancer cells by targeting FEAT

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Liang, Hongwei; Fu, Zheng; Jiang, Xueyuan; Wang, Nan; Wang, Feng; Wang, Xueliang; Zhang, Suyang; Wang, Yanbo; Yan, Xin; Guan, Wen-xian; Zhang, Chen-Yu; Zen, Ke; Zhang, Yujing; Chen, Xi; Zhou, Guangxin

2015-01-01

Although human cancers have heterogeneous combinations of altered oncogenes, some crucial genes are universally dysregulated in most cancers. One such gene, FEAT (faint expression in normal tissues, aberrant overexpression in tumors), is uniformly overexpressed in a variety of human cancers and plays an important role in tumorigenesis by suppressing apoptosis. However, the precise molecular mechanism through which FEAT is upregulated during tumorigenesis remains largely unknown. In this study, we used bioinformatic analyses to search for miRNAs that potentially target FEAT. We examined the expression of FEAT protein level by western blotting and miR-16 level by qRT-PCR assay. Cancer cell lines (A549, MCF-7 and Huh-7) with miR-16 upregulation and FEAT silencing were established and the effects on apoptosis of cancer cells in vitro were assessed. Luciferase reporter assay was also performed to investigate the interaction between miR-16 and FEAT. We identified a specific target site for miR-16 in the 3′-untranslated region (3′-UTR) of FEAT. Consistent with the bioinformatic analyses, we identified an inverse correlation between the miR-16 and FEAT protein levels in lung cancer, breast cancer, and hepatocellular cancer tissues. We then experimentally validated miR-16 as a direct regulator of FEAT using cell transfection and luciferase assays. Finally, we demonstrated that the repression of FEAT by miR-16 promoted the apoptosis of cancer cells. Our findings provide the first clues regarding the role of miR-16 as a tumor suppressor in cancer cells through the inhibition of FEAT translation. The online version of this article (doi:10.1186/s12885-015-1458-8) contains supplementary material, which is available to authorized users

The micro-RNA72c-APETALA2-1 node as a key regulator of the common bean-Rhizobium etli nitrogen fixation symbiosis.

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Nova-Franco, Bárbara; Íñiguez, Luis P; Valdés-López, Oswaldo; Alvarado-Affantranger, Xochitl; Leija, Alfonso; Fuentes, Sara I; Ramírez, Mario; Paul, Sujay; Reyes, José L; Girard, Lourdes; Hernández, Georgina

2015-05-01

Micro-RNAs are recognized as important posttranscriptional regulators in plants. The relevance of micro-RNAs as regulators of the legume-rhizobia nitrogen-fixing symbiosis is emerging. The objective of this work was to functionally characterize the role of micro-RNA172 (miR172) and its conserved target APETALA2 (AP2) transcription factor in the common bean (Phaseolus vulgaris)-Rhizobium etli symbiosis. Our expression analysis revealed that mature miR172c increased upon rhizobial infection and continued increasing during nodule development, reaching its maximum in mature nodules and decaying in senescent nodules. The expression of AP2-1 target showed a negative correlation with miR172c expression. A drastic decrease in miR172c and high AP2-1 mRNA levels were observed in ineffective nodules. Phenotypic analysis of composite bean plants with transgenic roots overexpressing miR172c or a mutated AP2-1 insensitive to miR172c cleavage demonstrated the pivotal regulatory role of the miR172 node in the common bean-rhizobia symbiosis. Increased miR172 resulted in improved root growth, increased rhizobial infection, increased expression of early nodulation and autoregulation of nodulation genes, and improved nodulation and nitrogen fixation. In addition, these plants showed decreased sensitivity to nitrate inhibition of nodulation. Through transcriptome analysis, we identified 114 common bean genes that coexpressed with AP2-1 and proposed these as being targets for transcriptional activation by AP2-1. Several of these genes are related to nodule senescence, and we propose that they have to be silenced, through miR172c-induced AP2-1 cleavage, in active mature nodules. Our work sets the basis for exploring the miR172-mediated improvement of symbiotic nitrogen fixation in common bean, the most important grain legume for human consumption. © 2015 American Society of Plant Biologists. All Rights Reserved.

miR-18a promotes cell proliferation of esophageal squamous cell carcinoma cells by increasing cylin D1 via regulating PTEN-PI3K-AKT-mTOR signaling axis

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Zhang, Weiguo, E-mail: weiguozhangHU@gmail.com; Lei, Caipeng; Fan, Junli; Wang, Jing

2016-08-12

Esophageal squamous cell carcinoma (ESCC) is one of the lethal cancers with a high incidence rate in Asia. Cyclin D1 is overexpressed and plays an important role in the carcinogenesis of ESCC; however the mechanism of the deregulation of Cyclin D1 in ESCC remains to be determined. In the study, we found that miR-18a promotes the expression Cyclin D1 by targeting PTEN in eophageal squamous cell carcinoma TE13 and Eca109 cells. Transfection of miR-18a mimetics increased cyclin D1, while transfection of miR-18a antagomir decreased D1. Moreover, miR-18a-mediated upregulation of cyclin D1 was accompanied with downregulation of PTEN, which is a direct target of miR-18a, and increase of the phosphorylation of AKT and S6K1. In addition, pharmacologic inhibition of AKT or mTOR kinases abolished the increase of cyclinD1 by miR-18a, which was accompanied with decreased phosphorylation of Rb−S780 and inhibition of cell proliferation. Our results demonstrated the upregulation of miR-18a promoted cell proliferation by increasing cylin D1 via regulating PTEN-PI3K-AKT-mTOR signaling axis, suggesting that small molecule inhibitors of AKT-mTOR signaling are potential agents for the treatment of ESCC patients with upregulation of miR-17-92 cluster. - Highlights: • miR-18a promotes the proliferation of ESCC cells. • miR-18a increase cyclin D1 expression in ESCC cells. • miR-18a directly targets PTEN in ESCC cells. • Inhibition of AKT-mTOR prevents miR-18a-induced cyclin D1 in ESCC cells. • miR-18a antagomir sensitizes ESCC cells to cisplatin.

miR-29c targets TNFAIP3, inhibits cell proliferation and induces apoptosis in hepatitis B virus-related hepatocellular carcinoma

International Nuclear Information System (INIS)

Wang, Chun-Mei; Wang, Yan; Fan, Chun-Guang; Xu, Fei-Fei; Sun, Wen-Sheng; Liu, Yu-Gang; Jia, Ji-Hui

2011-01-01

Highlights: → miR-29c was significantly downregulated in HBV-related HCC. → TNFAIP3 was found to be inversely correlated with miR-29c levels and identified as a target of miR-29c. → Overexpression of miR-29c suppressed TNFAIP3. → miR-29c inhibited HBV DNA replication, cell proliferation and induced apoptosis. -- Abstract: Recent studies have revealed that microRNA-29c (miR-29c) is involved in a variety of biological processes including carcinogenesis. Here, we report that miR-29c was significantly downregulated in hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) cell lines as well as in clinical tissues compared with their corresponding controls. Tumor necrosis factor alpha-induced protein 3 (TNFAIP3), a key regulator in inflammation and immunity, was found to be inversely correlated with miR-29c levels and was identified as a target of miR-29c. Overexpression of miR-29c in HepG2.2.15 cells effectively suppressed TNFAIP3 expression and HBV DNA replication as well as inhibited cell proliferation and induced apoptosis. We conclude that miR-29c may play an important role as a tumor suppressive microRNA in the development and progression of HBV-related HCC by targeting TNFAIP3. Thus miR-29c and TNFAIP3 represent key diagnostic markers and potential therapeutic targets for the prevention and treatment of HBV infection.

miR-29c targets TNFAIP3, inhibits cell proliferation and induces apoptosis in hepatitis B virus-related hepatocellular carcinoma

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Wang, Chun-Mei [Department of Microbiology, Shandong University School of Medicine, Jinan 250012 (China); Department of Pathophysiology, Shandong University School of Medicine, Jinan 250012 (China); Wang, Yan; Fan, Chun-Guang; Xu, Fei-Fei [Department of Pathophysiology, Shandong University School of Medicine, Jinan 250012 (China); Sun, Wen-Sheng [Institute of Immunology, Shandong University School of Medicine, Jinan 250012 (China); Liu, Yu-Gang, E-mail: liu.yugang@sdu.edu.cn [Department of Pathophysiology, Shandong University School of Medicine, Jinan 250012 (China); Jia, Ji-Hui, E-mail: jiajihui@sdu.edu.cn [Department of Microbiology, Shandong University School of Medicine, Jinan 250012 (China)

2011-08-05

Highlights: {yields} miR-29c was significantly downregulated in HBV-related HCC. {yields} TNFAIP3 was found to be inversely correlated with miR-29c levels and identified as a target of miR-29c. {yields} Overexpression of miR-29c suppressed TNFAIP3. {yields} miR-29c inhibited HBV DNA replication, cell proliferation and induced apoptosis. -- Abstract: Recent studies have revealed that microRNA-29c (miR-29c) is involved in a variety of biological processes including carcinogenesis. Here, we report that miR-29c was significantly downregulated in hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) cell lines as well as in clinical tissues compared with their corresponding controls. Tumor necrosis factor alpha-induced protein 3 (TNFAIP3), a key regulator in inflammation and immunity, was found to be inversely correlated with miR-29c levels and was identified as a target of miR-29c. Overexpression of miR-29c in HepG2.2.15 cells effectively suppressed TNFAIP3 expression and HBV DNA replication as well as inhibited cell proliferation and induced apoptosis. We conclude that miR-29c may play an important role as a tumor suppressive microRNA in the development and progression of HBV-related HCC by targeting TNFAIP3. Thus miR-29c and TNFAIP3 represent key diagnostic markers and potential therapeutic targets for the prevention and treatment of HBV infection.

MiR-27a Promotes Hemin-Induced Erythroid Differentiation of K562 Cells by Targeting CDC25B

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Dongsheng Wang

2018-03-01

Full Text Available Background/Aims: MicroRNAs (miRNAs play a crucial role in erythropoiesis. MiR-23a∼27a∼24-2 clusters have been proven to take part in erythropoiesis via some proteins. CDC25B (cell division control Cdc2 phosphostase B is also the target of mir-27a; whether it regulates erythropoiesis and its mechanism are unknown. Methods: To evaluate the potential role of miR-27a during erythroid differentiation, we performed miR-27a gain- and loss-of-function experiments on hemin-induced K562 cells. We detected miR-27a expression after hemin stimulation at different time points. At the same time, the γ-globin gene also was measured via real-time PCR. According to the results of the chips, we screened the target protein of miR-27a through a dual-luciferase reporter assay and identified it via Western blot analyses. To evaluate the function of CDC25B, benzidine staining and flow cytometry were employed to detect the cell differentiation and cell cycle. Results: We found that miR-27a promotes hemin-induced erythroid differentiation of human K562 cells by targeting cell division cycle 25 B (CDC25B. Overexpression of miR-27a promotes the differentiation of hemin-induced K562 cells, as demonstrated by γ-globin overexpression. The inhibition of miR-27a expression suppresses erythroid differentiation, thus leading to a reduction in the γ-globin gene. CDC25B was identified as a new target of miR-27a during erythroid differentiation. Overexpression of miR-27a led to decreased CDC25B expression after hemin treatment, and CDC25B was up-regulated when miR-27a expression was inhibited. Moreover, the inhibition of CDC25B affected erythroid differentiation, as assessed by γ-globin expression. Conclusion: This study is the first report of the interaction between miR-27a and CDC25B, and it improves the understanding of miRNA functions during erythroid differentiation.

Monocyte to macrophage differentiation-associated (MMD) targeted by miR-140-5p regulates tumor growth in non-small cell lung cancer

International Nuclear Information System (INIS)

Li, Weina; He, Fei

2014-01-01

Highlights: • Expression of MMD is increased in lung cancer tissues. • Knockdown of MMD inhibits growth of A549 and LLC cells in vitro and in vivo. • MMD is a direct functional target of miR-140-5p. • MiR-140-5p/MMD axis regulates Erk1/2 signaling. - Abstract: Monocyte to macrophage differentiation-associated (MMD) is identified in macrophages as a gene associated with the differentiation from monocytes to macrophages. Recent microarray analysis for non-small cell lung cancer (NSCLC) suggests that MMD is an important signature associated with relapse and survival among patients with NSCLC. Therefore, we speculate that MMD likely plays a role in lung cancer. In this study, we found that the protein level of MMD was increased in lung cancer compared to benign lung tissues, and knockdown of MMD inhibited the growth of A549 and Lewis lung cancer cells (LLC) in vitro and in vivo. Integrated analysis demonstrated that MMD was a direct functional target of miR-140-5p. Furthermore, we found that miR-140-5p/MMD axis could affect the cell proliferation of lung cancer cells by regulating Erk signaling. Together, our results highlight the significance of miR-140-5p/MMD axis in lung cancer, and miR-140-5p/MMD axis could serve as new molecular targets for the therapy against lung cancer

Genetic Ablation of miR-33 Increases Food Intake, Enhances Adipose Tissue Expansion, and Promotes Obesity and Insulin Resistance

Directory of Open Access Journals (Sweden)

Nathan L. Price

2018-02-01

Full Text Available While therapeutic modulation of miRNAs provides a promising approach for numerous diseases, the promiscuous nature of miRNAs raises concern over detrimental off-target effects. miR-33 has emerged as a likely target for treatment of cardiovascular diseases. However, the deleterious effects of long-term anti-miR-33 therapies and predisposition of miR-33−/− mice to obesity and metabolic dysfunction exemplify the possible pitfalls of miRNA-based therapies. Our work provides an in-depth characterization of miR-33−/− mice and explores the mechanisms by which loss of miR-33 promotes insulin resistance in key metabolic tissues. Contrary to previous reports, our data do not support a direct role for SREBP-1-mediated lipid synthesis in promoting these effects. Alternatively, in adipose tissue of miR-33−/− mice, we observe increased pre-adipocyte proliferation, enhanced lipid uptake, and impaired lipolysis. Moreover, we demonstrate that the driving force behind these abnormalities is increased food intake, which can be prevented by pair feeding with wild-type animals.

miR-122 promotes hepatic antioxidant defense of genetically improved farmed tilapia (GIFT, Oreochromis niloticus) exposed to cadmium by directly targeting a metallothionein gene

International Nuclear Information System (INIS)

Qiang, Jun; Tao, Yi-Fan; He, Jie; Xu, Pao; Bao, Jin-Wen; Sun, Yi-Lan

2017-01-01

Highlights: • MiR-122 regulated tilapia MT by directly targeting MT 3′UTR. • MiR-122 level was negatively related to MT level under Cd stress. • MiR-122 silencing caused up-regulation of MT expression. • MiR-122 loss relieved liver stress and stimulated antioxidant enzymes. - Abastract: MicroRNAs (miRNAs) are small, non-coding RNAs that regulate target gene expression by binding to the 3′untranslated region (3′UTR) of the target mRNA. MiRNAs regulate a large variety of genes, including those involved in liver homeostasis and energy metabolism. Down-regulated levels of hepatic miR-122 were found in genetically improved farmed tilapia (GIFT, Oreochromis niloticus) exposed to cadmium (Cd) stress. Here, we report for the first time that reduction of miR-122 post-transcriptionally increased metallothionein (MT) mRNA levels by binding to its 3′UTR, as shown by a 3′ UTR luciferase reporter assay. The expression levels of miR-122 were negatively related to MT levels in GIFT under Cd stress. We performed in vivo functional analysis of miR-122 by injecting the fish with a miR-122 antagomir. Inhibition of miR-122 levels in GIFT liver caused a significant increase in MT expression, affected white blood cell and red blood cell counts, and serum alanine and aspartate aminotransferase activities, and glucose levels, all of which may help to relieve Cd stress-related liver stress. miR-122 silencing modulated oxidative stress and stimulated the activity of antioxidant enzymes. Our findings indicate that miR-122 regulated MT levels by binding to the 3′UTR of MT mRNA, and this interaction affected Cd stress induction and the resistance response in GIFT. We concluded that miR-122 plays an important role in regulating the stress response in GIFT liver. Our findings may contribute to understanding the mechanisms of miRNA-mediated gene regulation in tilapia in response to environmental stresses.

miR-122 promotes hepatic antioxidant defense of genetically improved farmed tilapia (GIFT, Oreochromis niloticus) exposed to cadmium by directly targeting a metallothionein gene

Energy Technology Data Exchange (ETDEWEB)

Qiang, Jun, E-mail: Qiangj@ffrc.cn [Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization, Ministry of Agriculture, Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences, Wuxi 214081, Jiangsu (China); Tao, Yi-Fan [Wuxi Fisheries College, Nanjing Agricultural University, Wuxi 214081 (China); He, Jie [Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization, Ministry of Agriculture, Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences, Wuxi 214081, Jiangsu (China); Xu, Pao, E-mail: Xup@ffrc.cn [Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization, Ministry of Agriculture, Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences, Wuxi 214081, Jiangsu (China); Bao, Jin-Wen [Wuxi Fisheries College, Nanjing Agricultural University, Wuxi 214081 (China); Sun, Yi-Lan [Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization, Ministry of Agriculture, Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences, Wuxi 214081, Jiangsu (China)

2017-01-15

Highlights: • MiR-122 regulated tilapia MT by directly targeting MT 3′UTR. • MiR-122 level was negatively related to MT level under Cd stress. • MiR-122 silencing caused up-regulation of MT expression. • MiR-122 loss relieved liver stress and stimulated antioxidant enzymes. - Abastract: MicroRNAs (miRNAs) are small, non-coding RNAs that regulate target gene expression by binding to the 3′untranslated region (3′UTR) of the target mRNA. MiRNAs regulate a large variety of genes, including those involved in liver homeostasis and energy metabolism. Down-regulated levels of hepatic miR-122 were found in genetically improved farmed tilapia (GIFT, Oreochromis niloticus) exposed to cadmium (Cd) stress. Here, we report for the first time that reduction of miR-122 post-transcriptionally increased metallothionein (MT) mRNA levels by binding to its 3′UTR, as shown by a 3′ UTR luciferase reporter assay. The expression levels of miR-122 were negatively related to MT levels in GIFT under Cd stress. We performed in vivo functional analysis of miR-122 by injecting the fish with a miR-122 antagomir. Inhibition of miR-122 levels in GIFT liver caused a significant increase in MT expression, affected white blood cell and red blood cell counts, and serum alanine and aspartate aminotransferase activities, and glucose levels, all of which may help to relieve Cd stress-related liver stress. miR-122 silencing modulated oxidative stress and stimulated the activity of antioxidant enzymes. Our findings indicate that miR-122 regulated MT levels by binding to the 3′UTR of MT mRNA, and this interaction affected Cd stress induction and the resistance response in GIFT. We concluded that miR-122 plays an important role in regulating the stress response in GIFT liver. Our findings may contribute to understanding the mechanisms of miRNA-mediated gene regulation in tilapia in response to environmental stresses.

MiR-34a-3p alters proliferation and apoptosis of meningioma cells in vitro and is directly targeting SMAD4, FRAT1 and BCL2

Science.gov (United States)

Werner, Tamara V.; Hart, Martin; Nickels, Ruth; Kim, Yoo-Jin; Menger, Michael D.; Bohle, Rainer M.; Keller, Andreas; Ludwig, Nicole; Meese, Eckart

2017-01-01

Micro (mi)RNAs are short, noncoding RNAs and deregulation of miRNAs and their targets are implicated in tumor generation and progression in many cancers. Meningiomas are mostly benign, slow growing tumors of the central nervous system with a small percentage showing a malignant phenotype. Following in silico prediction of potential targets of miR-34a-3p, SMAD4, FRAT1, and BCL2 have been confirmed as targets by dual luciferase assays with co-expression of miR-34a-3p and reporter gene constructs containing the respective 3'UTRs. Disruption of the miR-34a-3p binding sites in the 3'UTRs resulted in loss of responsiveness to miR-34a-3p overexpression. In meningioma cells, overexpression of miR-34a-3p resulted in decreased protein levels of SMAD4, FRAT1 and BCL2, while inhibition of miR-34a-3p led to increased levels of these proteins as confirmed by Western blotting. Furthermore, deregulation of miR-34a-3p altered cell proliferation and apoptosis of meningioma cells in vitro. We show that SMAD4, FRAT1 and BCL2 are direct targets of miR-34a-3p and that deregulation of miR-34a-3p alters proliferation and apoptosis of meningioma cells in vitro. As part of their respective signaling pathways, which are known to play a role in meningioma genesis and progression, deregulation of SMAD4, FRAT1 and BCL2 might contribute to the aberrant activation of these signaling pathways leading to increased proliferation and inhibition of apoptosis in meningiomas. PMID:28340489

MicroRNA, miR-374b, directly targets Myf6 and negatively regulates C2C12 myoblasts differentiation

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Ma, Zhiyuan; Sun, Xiaorui; Xu, Dequan; Xiong, Yuanzhu; Zuo, Bo, E-mail: zuobo@mail.hzau.edu.cn

2015-11-27

Myogenesis is a complex process including myoblast proliferation, differentiation and myotube formation and is controlled by myogenic regulatory factors (MRFs), MyoD, MyoG, Myf5 and Myf6 (also known as MRF4). MicroRNA is a kind of ∼22 nt-long non-coding small RNAs, and act as key transcriptional or post-transcriptional regulators of gene expression. Identification of miRNAs involved in the regulation of muscle genes could improve our understanding of myogenesis process. In this study, we investigated the regulation of Myf6 gene by miRNAs. We showed that miR-374b specifically bound to the 3'untranslated region (UTR) of Myf6 and down-regulated the expression of Myf6 gene at both mRNA and protein level. Furthermore, miR-374b is ubiquitously expressed in the tissues of adult C57BL6 mouse, and the mRNA abundance increases first and then decreases during C2C12 myoblasts differentiation. Over-expression of miR-374b impaired C2C12 cell differentiation, while inhibiting miR-374b expression by 2′-O-methyl antisense oligonucleotides promoted C2C12 cell differentiation. Taken together, our findings identified miR-374b directly targets Myf6 and negatively regulates myogenesis. - Highlights: • MiR-374b directly targets 3′UTR of Myf6. • MiR-374b negatively regulates Myf6 in C2C12 cells. • MiR-374b abundance significiently changes during C2C12 cells differentiation. • MiR-374b negatively regulates C2C12 cells differentiation.

Knockdown of long non-coding RNA HOTAIR increases miR-454-3p by targeting Stat3 and Atg12 to inhibit chondrosarcoma growth.

Science.gov (United States)

Bao, Xing; Ren, Tingting; Huang, Yi; Sun, Kunkun; Wang, Shidong; Liu, Kuisheng; Zheng, Bingxin; Guo, Wei

2017-02-09

Current practices for the therapy of chondrosarcoma, including wide-margin surgical resection and chemotherapy, are less than satisfactory. Recently, emerging evidence has demonstrated that long non-coding RNAs (lncRNAs) have an essential role in the initiation and progression of tumors. As a typical lncRNA, HOTAIR is significantly overexpressed in various tumors. However, the function and potential biological mechanisms of HOTAIR in human chondrosarcoma remain unknown. Quantitative RT-PCR demonstrated that HOTAIR expression was upregulated in chondrosarcoma tissues and cell lines. High HOTAIR expression is correlated with tumor stage and poor prognosis. Functional experiments reveal that HOTAIR knockdown leads to growth inhibition of human chondrosarcoma cells in vitro and in vivo. In addition to cycle arrest and apoptosis, knockdown of HOTAIR inhibits autophagy, which favors cell death. Mechanistically, we demonstrated that HOTAIR induced DNA methylation of miR-454-3p by recruiting EZH2 and DNMT1 to the miR-454-3p promoter regions, which markedly silences miR-454-3p expression. Further analysis revealed that STAT3 and ATG12 are targets of miR-454-3p, initiate HOTAIR deficiency-induced apoptosis and reduce autophagy. Collectively, our data reveal the roles and functional mechanisms of HOTAIR in human chondrosarcoma and suggest that HOTAIR may act as a prognostic biomarker and potential therapeutic target for chondrosarcoma.

miRNA Expression Profile after Status Epilepticus and Hippocampal Neuroprotection by Targeting miR-132

Science.gov (United States)

Jimenez-Mateos, Eva M.; Bray, Isabella; Sanz-Rodriguez, Amaya; Engel, Tobias; McKiernan, Ross C.; Mouri, Genshin; Tanaka, Katsuhiro; Sano, Takanori; Saugstad, Julie A.; Simon, Roger P.; Stallings, Raymond L.; Henshall, David C.

2011-01-01

When an otherwise harmful insult to the brain is preceded by a brief, noninjurious stimulus, the brain becomes tolerant, and the resulting damage is reduced. Epileptic tolerance develops when brief seizures precede an episode of prolonged seizures (status epilepticus). MicroRNAs (miRNAs) are small, noncoding RNAs that function as post-transcriptional regulators of gene expression. We investigated how prior seizure preconditioning affects the miRNA response to status epilepticus evoked by intra-amygdalar kainic acid in mice. The miRNA was extracted from the ipsilateral CA3 subfield 24 hours after focal-onset status epilepticus in animals that had previously received either seizure preconditioning (tolerance) or no preconditioning (injury), and mature miRNA levels were measured using TaqMan low-density arrays. Expression of 21 miRNAs was increased, relative to control, after status epilepticus alone, and expression of 12 miRNAs was decreased. Increased miR-132 levels were matched with increased binding to Argonaute-2, a constituent of the RNA-induced silencing complex. In tolerant animals, expression responses of >40% of the injury-group-detected miRNAs differed, being either unchanged relative to control or down-regulated, and this included miR-132. In vivo microinjection of locked nucleic acid-modified oligonucleotides (antagomirs) against miR-132 depleted hippocampal miR-132 levels and reduced seizure-induced neuronal death. Thus, our data strongly suggest that miRNAs are important regulators of seizure-induced neuronal death. PMID:21945804

miR-335 negatively regulates osteosarcoma stem cell-like properties by targeting POU5F1.

Science.gov (United States)

Guo, Xiaodong; Yu, Ling; Zhang, Zhengpei; Dai, Guo; Gao, Tian; Guo, Weichun

2017-01-01

Evidence is accumulating to link cancer stem cells to the pathogenesis and progression of osteosarcoma. The aim of this study is to investigate the role of miR-335 in osteosarcoma stem cells. Tumor spheroid culture and flow cytometry were applied to screen out osteosarcoma stem cells. Real-time quantitative PCR was used to detect the expression level of miR-335 in MG63, U2OS and 143B osteosarcoma stem cells. The relationship of miR-335 expression with osteosarcoma stem cells was then analyzed. Transwell assay and transplantation assay were performed to elucidate biological effects of miR-335 on cell invasion and vivo tumor formation. Western Blot and luciferase assays were executed to investigate the regulation of POU5F1 by miR-335. The expression of miR-335 in osteosarcoma stem cells was lower than their differentiated counterparts. Cells expressing miR-335 possessed decreased stem cell-like properties. Gain or loss of function assays were applied to find that miR-335 antagonist promoted stem cell-like properties as well as invasion. Luciferase report and transfection assay showed that POU5F1 was downregulated by miR-335. Pre-miR-335 resulted in tumor enhanced sensitivity to traditional chemotherapy, whereas anti-miR-335 promoted chemoresistance. Finally, the inhibitory effect of miR-335 on in vivo tumor formation showed that combination of pre-miR-335 with cisplatin further reduced the tumor size, and miR-335 brought down the sphere formation capacity induced by cisplatin. The current study demonstrates that miR-335 negatively regulates osteosarcoma stem cell-like properties by targeting POU5F1, and miR-335 could target CSCs to synergize with traditional chemotherapeutic agents to overcome osteosarcoma.

MiR-137 inhibited cell proliferation and migration of vascular smooth muscle cells via targeting IGFBP-5 and modulating the mTOR/STAT3 signaling.

Directory of Open Access Journals (Sweden)

Jin Pan

Full Text Available Abnormal proliferation of vascular smooth muscle cells (VSMCs contributes to the development of cardiovascular diseases. Studies have shown the great impact of microRNAs (miRNAs on the cell proliferation of VSMCs. This study examined the effects of miR-137 on the cell proliferation and migration of VSMCs and also explored the underlying molecular mechanisms. The mRNA and protein expression levels were determined by qRT-PCR and western blot assays, respectively. The CCK-8 assay, wound healing assay and transwell migration assay were performed to measure cell proliferation and migration of VSMCs. The miR-137-targeted 3'untranslated region of insulin-like growth factor-binding protein-5 (IGFBP-5 was confirmed by luciferase reporter assay. Platelet-derived growth factor-bb (PDGF-bb treatment enhanced cell proliferation and suppressed the expression of miR-137 in VSMCs. The gain-of-function and loss-of-function assays showed that overexpression of miR-137 suppressed the cell proliferation and migration, and also inhibited the expression of matrix genes of VSMCs; down-regulation of miR-137 had the opposite effects on VSMCs. Bioinformatics analysis and luciferase report assay results showed that IGFBP-5 was a direct target of miR-137, and miR-137 overexpression suppressed the IGFBP-5 expression and down-regulation of miR-137 increased the IGFBP-5 expression in VSMCs. PDGF-bb treatment also increased the IGFBP-5 mRNA expression. In addition, enforced expression of IGFBP-5 reversed the inhibitory effects of miR-137 on cell proliferation and migration of VSMCs. More importantly, overexpression of miR-137 also suppressed the activity of mTOR/STAT3 signaling in VSMCs. Taken together, the results suggest that miR-137 may suppress cell proliferation and migration of VSMCs via targeting IGFBP-5 and modulating mTOR/STAT3 signaling pathway.

miR-92a is upregulated in cervical cancer and promotes cell proliferation and invasion by targeting FBXW7

International Nuclear Information System (INIS)

Zhou, Chuanyi; Shen, Liangfang; Mao, Lei; Wang, Bing; Li, Yang; Yu, Huizhi

2015-01-01

MicroRNAs (miRNAs) are involved in the cervical carcinogenesis and progression. In this study, we investigated the role of miR-92a in progression and invasion of cervical cancer. MiR-92a was significantly upregulated in cervical cancer tissues and cell lines. Overexpression of miR-92a led to remarkably enhanced proliferation by promoting cell cycle transition from G1 to S phase and significantly enhanced invasion of cervical cancer cells, while its knockdown significantly reversed these cellular events. Bioinformatics analysis suggested F-box and WD repeat domain-containing 7 (FBXW7) as a novel target of miR-92a, and miR-92a suppressed the expression level of FBXW7 mRNA by direct binding to its 3′-untranslated region (3′UTR). Expression of miR-92a was negatively correlated with FBXW7 in cervical cancer tissues. Furthermore, Silencing of FBXW7 counteracted the effects of miR-92a suppression, while its overexpression reversed oncogenic effects of miR-92a. Together, these findings indicate that miR-92a acts as an onco-miRNA and may contribute to the progression and invasion of cervical cancer, suggesting miR-92a as a potential novel diagnostic and therapeutic target of cervical cancer. - Highlights: • miR-92a is elevated in cervical cancer tissues and cell lines. • miR-92a promotes cervical cancer cell proliferation, cell cycle transition from G1 to S phase and invasion. • FBXW7 is a direct target of miR-92a. • FBXW7 counteracts the oncogenic effects of miR-92a on cervical cancer cells

MiR-30e suppresses proliferation of hepatoma cells via targeting prolyl 4-hydroxylase subunit alpha-1 (P4HA1) mRNA

Energy Technology Data Exchange (ETDEWEB)

Feng, Guoxing [State Key Laboratory of Medicinal Chemical Biology, Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin (China); Shi, Hui [State Key Laboratory of Medicinal Chemical Biology, Department of Biochemistry, College of Life Sciences, Nankai University, Tianjin (China); Li, Jiong; Yang, Zhe [State Key Laboratory of Medicinal Chemical Biology, Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin (China); Fang, Runping; Ye, Lihong [State Key Laboratory of Medicinal Chemical Biology, Department of Biochemistry, College of Life Sciences, Nankai University, Tianjin (China); Zhang, Weiying, E-mail: zhwybao@nankai.edu.cn [State Key Laboratory of Medicinal Chemical Biology, Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin (China); Zhang, Xiaodong, E-mail: zhangxd@nankai.edu.cn [State Key Laboratory of Medicinal Chemical Biology, Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin (China)

2016-04-08

Aberrant microRNA expression has been shown to be characteristic of many cancers. It has been reported that the expression levels of miR-30e are decreased in liver cancer tissues. However, the role of miR-30e in hepatocellular carcinoma remains poorly understood. In the present study, we investigated the significance of miR-30e in hepatocarcinogenesis. Bioinformatics analysis reveals a putative target site of miR-30e in the 3′-untranslated region (3′UTR) of prolyl 4-hydroxylase subunit alpha-1 (P4HA1) mRNA. Moreover, luciferase reporter gene assays verified that miR-30e directly targeted 3′UTR of P4HA1 mRNA. Then, we demonstrated that miR-30e was able to reduce the expression of P4HA1 at the levels of mRNA and protein using reverse transcription-polymerase chain reaction and Western blot analysis. Enforced expression of miR-30e suppressed proliferation of HepG2 cells by 5-ethynyl-2-deoxyuridine (EdU) assay and reduced colony formation of these cells by colony formation analysis. Conversely, anti-miR-30e enhanced the proliferation of hepatoma cells in vitro. Interestingly, the ectopic expression of P4HA1 could efficiently rescue the inhibition of cell proliferation mediated by miR-30e in HepG2 cells. Meanwhile, silencing of P4HA1 abolished the anti-miR-30e-induced proliferation of cells. Clinically, quantitative real-time PCR showed that miR-30e was down-regulated in liver tumor tissues relative to their peritumor tissues. The expression levels of miR-30e were negatively correlated to those of P4HA1 mRNA in clinical liver tumor tissues. Thus, we conclude that miR-30e suppresses proliferation of hepatoma cells through targeting P4HA1 mRNA. Our finding provides new insights into the mechanism of hepatocarcinogenesis. - Highlights: • P4HA1 is a novel target gene of miR-30e. • P4HA1 is increased in clinical HCC tissues. • MiR-30e is negatively correlated with P4HA1 in clinical HCC tissues. • MiR-30e suppresses the proliferation of HCC cells through

miR-320 enhances the sensitivity of human colon cancer cells to chemoradiotherapy in vitro by targeting FOXM1

Energy Technology Data Exchange (ETDEWEB)

Wan, Lu-Ying; Deng, Jun; Xiang, Xiao-Jun; Zhang, Ling; Yu, Feng; Chen, Jun; Sun, Zhe; Feng, Miao; Xiong, Jian-Ping, E-mail: jpxiong@medmail.com.cn

2015-02-06

Highlights: • miR-320 plays a significant role in chemoresistance. • This role might be attribute to targeting FOXM1. • The Wnt/β-catenin pathway also involves in this chemotherapy sensitivity. - Abstract: miR-320 expression level is found to be down-regulated in human colon cancer. To date, however, its underlying mechanisms in the chemo-resistance remain largely unknown. In this study, we demonstrated that ectopic expression of miR-320 led to inhibit HCT-116 cell proliferation, invasion and hypersensitivity to 5-Fu and Oxaliplatin. Also, knockdown of miR-320 reversed these effects in HT-29 cells. Furthermore, we identified an oncogene, FOXM1, as a direct target of miR-320. In addition, miR-320 could inactive the activity of Wnt/β-catenin pathway. Finally, we found that miR-320 and FOXM1 protein had a negative correlation in colon cancer tissues and adjacent normal tissues. These findings implied that miR-320–FOXM1 axis may overcome chemo-resistance of colon cancer cells and provide a new therapeutic target for the treatment of colon cancer.

MiR-181b targets Six2 and inhibits the proliferation of metanephric mesenchymal cells in vitro

International Nuclear Information System (INIS)

Lyu, Zhongshi; Mao, Zhaomin; Wang, Honglian; Fang, Yin; Chen, Tielin; Wan, Qianya; Wang, Ming; Wang, Nian; Xiao, Jiangming; Wei, Hongyuan; Li, Xun; Liu, Yi; Zhou, Qin

2013-01-01

Highlights: •We do bio-informatics websites to analysis of Six2 3′UTR. •MiR181b is a putative miRNA which can targets Six2 3′UTR. •MiR-181b binding site in the 3′UTR of Six2 is functional. •MiR-181b suppresses MK3 cells cell proliferation by targeting Six2. -- Abstract: MicroRNAs (miRNAs) are small non-coding RNAs that down-regulate gene expression by binding to target mRNA for cleavage or translational repression, and play important regulatory roles in renal development. Despite increasing genes have been predicted to be miRNA targets by bioinformatic analysis during kidney development, few of them have been verified by experiment. The objective of our study is to identify the miRNAs targeting Six2, a critical transcription factor that maintains the mesenchymal progenitor pool via self-renewal (proliferation) during renal development. We initially analyzed the 3′UTR of Six2 and found 37 binding sites targeted by 50 putative miRNAs in the 3′UTR of Six2. Among the 50 miRNAs, miR-181b is the miRNAs predicted by the three used websites. In our study, the results of luciferase reporter assay, realtime-PCR and Western blot demonstrated that miR-181b directly targeted on the 3′UTR of Six2 and down-regulate the expression of Six2 at mRNA and protein levels. Furthermore, EdU proliferation assay along with the Six2 rescue strategy showed that miR-181b suppresses the proliferation of metanephric mesenchymal by targeting Six2 in part. In our research, we concluded that by targeting the transcription factor gene Six2, miR-181b inhibits the proliferation of metanephric mesenchymal cells in vitro and might play an important role in the formation of nephrons

MiR-181b targets Six2 and inhibits the proliferation of metanephric mesenchymal cells in vitro

Energy Technology Data Exchange (ETDEWEB)

Lyu, Zhongshi; Mao, Zhaomin; Wang, Honglian; Fang, Yin; Chen, Tielin [The Division of Molecular Nephrology and the Creative Training Center for Undergraduates, The M.O.E. Key Laboratory of Laboratory Medical Diagnostics, The College of Laboratory Medicine, Chongqing Medical University, Chongqing 400016 (China); Wan, Qianya [The Undergraduates Class of 2011 entry, The College of Laboratory Medicine, Chongqing Medical University, Chongqing 400016 (China); Wang, Ming; Wang, Nian; Xiao, Jiangming; Wei, Hongyuan; Li, Xun; Liu, Yi [The Division of Molecular Nephrology and the Creative Training Center for Undergraduates, The M.O.E. Key Laboratory of Laboratory Medical Diagnostics, The College of Laboratory Medicine, Chongqing Medical University, Chongqing 400016 (China); Zhou, Qin, E-mail: zhouqin@cqmu.edu.cn [The Division of Molecular Nephrology and the Creative Training Center for Undergraduates, The M.O.E. Key Laboratory of Laboratory Medical Diagnostics, The College of Laboratory Medicine, Chongqing Medical University, Chongqing 400016 (China)

2013-11-01

Highlights: •We do bio-informatics websites to analysis of Six2 3′UTR. •MiR181b is a putative miRNA which can targets Six2 3′UTR. •MiR-181b binding site in the 3′UTR of Six2 is functional. •MiR-181b suppresses MK3 cells cell proliferation by targeting Six2. -- Abstract: MicroRNAs (miRNAs) are small non-coding RNAs that down-regulate gene expression by binding to target mRNA for cleavage or translational repression, and play important regulatory roles in renal development. Despite increasing genes have been predicted to be miRNA targets by bioinformatic analysis during kidney development, few of them have been verified by experiment. The objective of our study is to identify the miRNAs targeting Six2, a critical transcription factor that maintains the mesenchymal progenitor pool via self-renewal (proliferation) during renal development. We initially analyzed the 3′UTR of Six2 and found 37 binding sites targeted by 50 putative miRNAs in the 3′UTR of Six2. Among the 50 miRNAs, miR-181b is the miRNAs predicted by the three used websites. In our study, the results of luciferase reporter assay, realtime-PCR and Western blot demonstrated that miR-181b directly targeted on the 3′UTR of Six2 and down-regulate the expression of Six2 at mRNA and protein levels. Furthermore, EdU proliferation assay along with the Six2 rescue strategy showed that miR-181b suppresses the proliferation of metanephric mesenchymal by targeting Six2 in part. In our research, we concluded that by targeting the transcription factor gene Six2, miR-181b inhibits the proliferation of metanephric mesenchymal cells in vitro and might play an important role in the formation of nephrons.

Ciculating miRNA-21 as a Biomarker Predicts Polycystic Ovary Syndrome (PCOS) in Patients.

Science.gov (United States)

Jiang, Liyan; Li, Wei; Wu, Minmin; Cao, Sifan

2015-01-01

Polycystic ovary syndrome (PCOS) is characterized by hyperandrogenism, hyperinsulinemia, and infertility. In PCOS, abnormal regulation of relevant genes is required for follicular development. By binding to the 3' untranslated region (3'URT), microRNAs (miRNAs) are widely involved in posttranscriptional gene regulation. However, few studies have been conducted on circulating miRNA expression in PCOS. This study aims to describe altered expression of circulating miR-21 in PCOS. The expression of serum miRNAs of PCOS patients were explored using the TaqMan Low Density Array followed by individual quantitative reverse transcription polymerase chain reaction assays. The protein level of LATS1 was determined using Western blot. To validate whether miR-21 targeted LATS1, the luciferase assay was applied. In comparison with normal subjects, the circulating level of miRNA-21 was significantly enhanced in PCOS patients. In PCOS patients, the expression levels of MST1/2, LATS1/2, TAZ were much lower than the control subjects. Luciferase reporter assay revealed that LATS1 was a downstream target of miR-21. In comparison with normal subjects, serum miR-21 is obviously increased in PCOS patients. Through targeting LATS1, miR-21 could prompt PCOS progression and could act as a novel non-invasive biomarker for diagnosis of PCOS.

TSA suppresses miR-106b-93-25 cluster expression through downregulation of MYC and inhibits proliferation and induces apoptosis in human EMC.

Science.gov (United States)

Zhao, Zhi-Ning; Bai, Jiu-Xu; Zhou, Qiang; Yan, Bo; Qin, Wei-Wei; Jia, Lin-Tao; Meng, Yan-Ling; Jin, Bo-Quan; Yao, Li-Bo; Wang, Tao; Yang, An-Gang

2012-01-01

Histone deacetylase (HDAC) inhibitors are emerging as a novel class of anti-tumor agents and have manifested the ability to decrease proliferation and increase apoptosis in different cancer cells. A significant number of genes have been identified as potential effectors responsible for the anti-tumor function of HDAC inhibitor. However, the molecular mechanisms of these HDAC inhibitors in this process remain largely undefined. In the current study, we searched for microRNAs (miRs) that were affected by HDAC inhibitor trichostatin (TSA) and investigated their effects in endometrial cancer (EMC) cells. Our data showed that TSA significantly inhibited the growth of EMC cells and induced their apoptosis. Among the miRNAs that altered in the presence of TSA, the miR-106b-93-25 cluster, together with its host gene MCM7, were obviously down-regulated in EMC cells. p21 and BIM, which were identified as target genes of miR-106b-93-25 cluster, increased in TSA treated tumor cells and were responsible for cell cycle arrest and apoptosis. We further identified MYC as a regulator of miR-106b-93-25 cluster and demonstrated its down-regulation in the presence of TSA resulted in the reduction of miR-106b-93-25 cluster and up-regulation of p21 and BIM. More important, we found miR-106b-93-25 cluster was up-regulated in clinical EMC samples in association with the overexpression of MCM7 and MYC and the down-regulation of p21 and BIM. Thus our studies strongly indicated TSA inhibited EMC cell growth and induced cell apoptosis and cell cycle arrest at least partially through the down-regulation of the miR-106b-93-25 cluster and up-regulation of it's target genes p21 and BIM via MYC.

MiR-520b suppresses proliferation of hepatoma cells through targeting ten-eleven translocation 1 (TET1) mRNA

Energy Technology Data Exchange (ETDEWEB)

Zhang, Weiying; Lu, Zhanping; Gao, Yuen [State Key Laboratory of Medicinal Chemical Biology, Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin (China); Ye, Lihong [State Key Laboratory of Medicinal Chemical Biology, Department of Biochemistry, College of Life Sciences, Nankai University, Tianjin (China); Song, Tianqiang, E-mail: tjchi@hotmai.com [Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Tianjin (China); Zhang, Xiaodong, E-mail: zhangxd@nankai.edu.cn [State Key Laboratory of Medicinal Chemical Biology, Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin (China)

2015-05-08

Accumulating evidence indicates that microRNAs are able to act as oncogenes or tumor suppressor genes in human cancer. We previously reported that miR-520b was down-regulated in hepatocellular carcinoma (HCC) and its deregulation was involved in hepatocarcinogenesis. In the present study, we report that miR-520b suppresses cell proliferation in HCC through targeting the ten-eleven translocation 1 (TET1) mRNA. Notably, we identified that miR-520b was able to target 3′-untranslated region (3′UTR) of TET1 mRNA by luciferase reporter gene assays. Then, we revealed that miR-520b was able to reduce the expression of TET1 at the levels of mRNA and protein using reverse transcription-polymerase chain reaction and Western blotting analysis. In terms of function, 5-ethynyl-2-deoxyuridine (EdU) incorporation and colony formation assays demonstrated that the forced miR-520b expression remarkably inhibited proliferation of hepatoma cells, but TET1 overexpression could rescue the inhibition of cell proliferation mediated by miR-520b. Furthermore, anti-miR-520b enhanced proliferation of hepatoma cells, whereas silencing of TET1 abolished anti-miR-520b-induced acceleration of cell proliferation. Then, we validated that the expression levels of miR-520b were negatively related to those of TET1 mRNA in clinical HCC tissues. Thus, we conclude that miR-520b depresses proliferation of liver cancer cells through targeting 3′UTR of TET1 mRNA. Our finding provides new insights into the mechanism of hepatocarcinogenesis. - Highlights: • TET1 is a novel target gene of miR-520b. • TET1 is upregulated in clinical HCC tissues. • MiR-520b is negatively correlated with TET1 in clinical HCC tissues. • MiR-520b depresses the proliferation of HCC cells through targeting TET1 mRNA.

Down-regulation of the miR-543 alleviates insulin resistance through targeting the SIRT1.

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Hu, Xiaojing; Chi, Liyi; Zhang, Wentao; Bai, Tiao; Zhao, Wei; Feng, Zhanbin; Tian, Hongyan

2015-12-25

Insulin resistance plays an important role in the development of hypertension, which is seriously detrimental to human health. Recently, Sirtuin-1 (SIRT1) has been found to participate in regulation of insulin resistance. Therefore, further studies focused on the SIRT1 regulators might provide a potential approach for combating insulin resistance and hypertension. Interestingly, in this study, we found that SIRT1 was the target gene of the miR-543 by the Dual-Luciferase Reporter Assay. Moreover, the miR-543 expression notably increased in the insulin-resistant HepG2 cells induced by TNF-α. Further analysis showed that the overexpression of the miR-543 lowered the SIRT1 mRNA and protein levels, resulting in the insulin resistance in the HepG2 cells; the inhibition of miR-543, however, enhanced the mRNA and protein expression of the SIRT1, and alleviated the insulin resistance. Furthermore, the SIRT1 overexpression abrogated the effect of miR-543 on insulin resistance. In addition, the overexpression of the miR-543 by the lentivirus-mediated gene transfer markedly impaired the insulin signaling assessed by the Western blot analysis of the glycogen synthesis and the phosphorylation of Akt and GSK3β. In summary, our study suggested that the downregulation of the miR-543 could alleviate the insulin resistance via the modulation of the SIRT1 expression, which might be a potential new strategy for treating insulin resistance and a promising therapeutic method for hypertension. Copyright © 2015 Elsevier Inc. All rights reserved.

Neurophysiological defects and neuronal gene deregulation in Drosophila mir-124 mutants.

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Kailiang Sun

2012-02-01

Full Text Available miR-124 is conserved in sequence and neuronal expression across the animal kingdom and is predicted to have hundreds of mRNA targets. Diverse defects in neural development and function were reported from miR-124 antisense studies in vertebrates, but a nematode knockout of mir-124 surprisingly lacked detectable phenotypes. To provide genetic insight from Drosophila, we deleted its single mir-124 locus and found that it is dispensable for gross aspects of neural specification and differentiation. On the other hand, we detected a variety of mutant phenotypes that were rescuable by a mir-124 genomic transgene, including short lifespan, increased dendrite variation, impaired larval locomotion, and aberrant synaptic release at the NMJ. These phenotypes reflect extensive requirements of miR-124 even under optimal culture conditions. Comparison of the transcriptomes of cells from wild-type and mir-124 mutant animals, purified on the basis of mir-124 promoter activity, revealed broad upregulation of direct miR-124 targets. However, in contrast to the proposed mutual exclusion model for miR-124 function, its functional targets were relatively highly expressed in miR-124-expressing cells and were not enriched in genes annotated with epidermal expression. A notable aspect of the direct miR-124 network was coordinate targeting of five positive components in the retrograde BMP signaling pathway, whose activation in neurons increases synaptic release at the NMJ, similar to mir-124 mutants. Derepression of the direct miR-124 target network also had many secondary effects, including over-activity of other post-transcriptional repressors and a net incomplete transition from a neuroblast to a neuronal gene expression signature. Altogether, these studies demonstrate complex consequences of miR-124 loss on neural gene expression and neurophysiology.

Overexpression of miR-19b Impairs Cardiac Development in Zebrafish by Targeting ctnnb1

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Mengmeng Li

2014-07-01

Full Text Available Background: MicroRNAs are broadly accepted as crucial regulators of cardiovascular development, and dysregulation of their expression has been linked to cardiac disease. MicroRNA cluster miR-17-92 has been implicated in cardiac development and function, yet its defined mechanisms of action in this context are uncertain. Here, we focused on miR-19b, a key component of the miR-17-92 cluster proven to induce cardiomyocyte proliferation in vitro. We aimed to identify the biological significance of miR-19b in cardiac development and its underlying molecular mechanism of action in vivo. Methods: We micro-injected zebrafish embryos with different concentrations (0, 2, 4 and 8 μm of miR-19b mimics or a negative control, and assessed the embryo malformation rate, mortality rate, hatching rate and heart abnormalities at 72 hours post-fertilization (72 hpf. Results: We found that overexpression of miR-19b impacted left-right symmetry and cardiac development of zebrafish embryos, characterized by pericardial edema, slower heart rate and cardiac looping defects in a dose-dependent manner. Moreover, several important signaling molecules in the Wnt signaling pathway were abnormally expressed, suggesting that overexpression of miR-19b induces the inhibition of the Wnt signaling pathway by directly targeting ctnnb1. Interestingly, the deformed cardiac phenotype was partially rescued by treatment with the GSK3β inhibitor lithium chloride. Conclusion: Our findings suggest that miR-19b regulates laterality development and heart looping in zebrafish embryos by targeting ctnnb1.

-5p and -3p strands of miR-145 and miR-140 during mesenchymal stem cell chondrogenic differentiation.

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Kenyon, Jonathan D; Sergeeva, Olga; Somoza, Rodrigo A; Li, Ming; Caplan, Arnold I; Khalil, Ahmad M; Lee, Zhenghong

2018-04-20

The chondrogenic differentiation of mesenchymal stem cells (MSCs) is mediated by transcription factors and small non-coding RNAs such as micro-RNAs (miRNAs). Each miRNA is initially transcribed as a long transcript, which matures to produce -5p and -3p strands. It is widely believed that the mature and functional miRNA from any given pre-miRNA, usually the -5p strand, is functional, while the opposing -3p strand is degraded. However, recent cartilage literature started to show functional -3p stands for a few miRNAs. This study aimed at examining both -5p and -3p strands of two key miRNAs miR-140 and miR-145 that are known to be involved in the chondrogenic differentiation of MSCs. The level (copy number) of both -5p and -3p strands of miR-145 and miR-140 along the timeline of MSC chondrogenic differentiation was determined by PCR. The gene expression profiles of several genes related to MSC chondrogenesis were compared with these miRNA profiles along the same timeline. While miR-145-3p is declining in step with miR-145-5p in pellet cultures during the process, the -3p strand is only 1% - 2% of the total miR-145 products. In contrast, the mature -3p and -5p products of miR-140 are found to increase with near equal molar expression throughout chondrogenic differentiation. Numerous genes are expressed by cartilage progenitor cells during development. One such target gene, Sox9 is a regulatory target of the dominant miR-145-5p, consistent with the data. Further experimental validations are warranted to confirm that ACAN, FOXO1 and RUNX3 as direct targets of miR-145-5p in the context of MSC chondrogenesis. Similarly, TRSP1 and ACAN are worth further validation as direct targets of miR-145-3p. For miR-140, SOX4 shall be further validated as a direct target of miR-140-5p while KLF4, PTHLH, and WNT5A can be validated as direct targets of miR-140-3p.

Novel Mad2-targeting miR-493-3p controls mitotic fidelity and cancer cells' sensitivity to paclitaxel.

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Tambe, Mahesh; Pruikkonen, Sofia; Mäki-Jouppila, Jenni; Chen, Ping; Elgaaen, Bente Vilming; Straume, Anne Hege; Huhtinen, Kaisa; Cárpen, Olli; Lønning, Per Eystein; Davidson, Ben; Hautaniemi, Sampsa; Kallio, Marko J

2016-03-15

The molecular pathways that contribute to the proliferation and drug response of cancer cells are highly complex and currently insufficiently characterized. We have identified a previously unknown microRNA-based mechanism that provides cancer cells means to stimulate tumorigenesis via increased genomic instability and, at the same time, evade the action of clinically utilized microtubule drugs. We demonstrate miR-493-3p to be a novel negative regulator of mitotic arrest deficient-2 (MAD2), an essential component of the spindle assembly checkpoint that monitors the fidelity of chromosome segregation. The microRNA targets the 3' UTR of Mad2 mRNA thereby preventing translation of the Mad2 protein. In cancer cells, overexpression of miR-493-3p induced a premature mitotic exit that led to increased frequency of aneuploidy and cellular senescence in the progeny cells. Importantly, excess of the miR-493-3p conferred resistance of cancer cells to microtubule drugs. In human neoplasms, miR-493-3p and Mad2 expression alterations correlated with advanced ovarian cancer forms and high miR-493-3p levels were associated with reduced survival of ovarian and breast cancer patients with aggressive tumors, especially in the paclitaxel therapy arm. Our results suggest that intratumoral profiling of miR-493-3p and Mad2 levels can have diagnostic value in predicting the efficacy of taxane chemotherapy.

miR-124-3p functions as a tumor suppressor in breast cancer by targeting CBL

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Wang, Yanbo; Chen, Luxiao; Wu, Zhenyu; Wang, Minghai; Jin, Fangfang; Wang, Nan; Hu, Xiuting; Liu, Zhengya; Zhang, Chen-Yu; Zen, Ke; Chen, Jiangning; Liang, Hongwei; Zhang, Yujing; Chen, Xi

2016-01-01

The origin and development of breast cancer remain complex and obscure. Recently, microRNA (miRNA) has been identified as an important regulator of the initiation and progression of breast cancer, and some studies have shown the essential role of miR-124-3p as a tumor suppressor in breast tumorigenesis. However, the detailed role of miR-124-3p in breast cancer remains poorly understood. Quantitative RT-PCR and western blotting assays were used to measure miR-124-3p and CBL expression levels in breast cancer tissues, respectively. Luciferase reporter assay was employed to validate the direct targeting of CBL by miR-124-3p. Cell proliferation and invasion assays were performed to analyze the biological functions of miR-124-3p and CBL in breast cancer cells. In the present study, we found that miR-124-3p was consistently downregulated in breast cancer tissues. Moreover, we showed that miR-124-3p significantly suppressed the proliferation and invasion of breast cancer cells. In addition, we investigated the molecular mechanism through which miR-124-3p contributes to breast cancer tumorigenesis and identified CBL (Cbl proto-oncogene, E3 ubiquitin protein ligase) as a direct target gene of miR-124-3p. Moreover, we found that ectopic expression of CBL can attenuate the inhibitory effect of miR-124-3p on cell proliferation and invasion in breast cancer cells. This study identified a new regulatory axis in which miR-124-3p and CBL regulate the proliferation and invasion of breast cancer cells. The online version of this article (doi:10.1186/s12885-016-2862-4) contains supplementary material, which is available to authorized users

Potential Role of Circulating MiR-21 in the Diagnosis and Prognosis of Digestive System Cancer: A Systematic Review and Meta-Analysis.

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Yin, Chengqiang; Zhou, Xiaoying; Dang, Yini; Yan, Jin; Zhang, Guoxin

2015-12-01

Recent evidences indicate that circulating microRNAs (miRNAs) exhibit aberrant expression in the plasma of patients suffering from cancer compared to normal individuals, suggesting that it may be a useful noninvasion diagnostic method. MiR-21 plays crucial roles in carcinogenesis and can be served as a biomarker for the detection of various cancers. Therefore, the aim of this meta-analysis is to assess the potential role of miR-21 for digestive system cancer. By searching the PubMed, Embase, and Web of Science for publications concerning the diagnostic value of miR-21 for digestive system cancer, total of 23 publications were included in this meta-analysis. Receiver operating characteristic curves (ROC) were used to check the overall test performance. For prognostic meta-analysis, pooled hazard ratios (HRs) of circulating miR-21 for survival were calculated. Totally 23 eligible publications were included in this meta-analysis (15 articles for diagnosis and 8 articles for prognosis). For diagnostic meta-analysis, the summary estimates revealed that the pooled sensitivity and specificity were 0.76 (95% CI = 0.70-0.82) and 0.84 (95% CI = 0.78-0.89). Besides, the area under the summary ROC curve (AUC) is 0.87. For prognostic meta-analysis, the pooled HR of higher miR-21 expression in circulation was 1.94 (95% CI = 0.99-3.82, P = 0.055), which indicated higher miR-21 expression could be likely to predict poorer survival in digestive system cancer. The subgroup analysis implied the higher expression of miR-21 was correlated with worse overall survival in the Asian population in digestive system cancer (HR = 2.41, 95% CI = 1.21-4.77, P = 0.012). The current evidence suggests circulating miR-21 may be suitable to be a diagnostic and prognostic biomarker for digestive system cancer in the Asians.

Decreased miR-106a inhibits glioma cell glucose uptake and proliferation by targeting SLC2A3 in GBM.

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Dai, Dong-Wei; Lu, Qiong; Wang, Lai-Xing; Zhao, Wen-Yuan; Cao, Yi-Qun; Li, Ya-Nan; Han, Guo-Sheng; Liu, Jian-Min; Yue, Zhi-Jian

2013-10-14

MiR-106a is frequently down-regulated in various types of human cancer. However the underlying mechanism of miR-106a involved in glioma remains elusive. The association of miR-106a with glioma grade and patient survival was analyzed. The biological function and target of miR-106a were determined by bioinformatic analysis and cell experiments (Western blot, luciferase reporter, cell cycle, ntracellular ATP production and glucose uptake assay). Finally, rescue expression of its target SLC2A3 was used to test the role of SLC2A3 in miR-106a-mediated cell glycolysis and proliferation. Here we showed that miR-106a was a tumor suppressor miRNA was involved in GBM cell glucose uptake and proliferation. Decreased miR-106a in GBM tissues and conferred a poor survival of GBM patients. SLC2A3 was identified as a core target of miR-106a in GBM cells. Inhibition of SLC2A3 by miR-106a attenuated cell proliferation and inhibited glucose uptake. In addition, for each biological process we identified ontology-associated transcripts that significantly correlated with SLC2A3 expression. Finally, the expression of SLC2A3 largely abrogated miR-106a-mediated cell proliferation and glucose uptake in GBM cells. Taken together, miR-106a and SLC2A3 could be potential therapeutic approaches for GBM.

MiR-142 modulates human pancreatic cancer proliferation and invasion by targeting hypoxia-inducible factor 1 (HIF-1α in the tumor microenvironments

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Yebin Lu

2017-02-01

Full Text Available MicroRNAs regulate most protein-coding genes, including genes important in cancer and other diseases. In this study, we demonstrated that the expression of miR-142 could be significantly suppressed in pancreatic cancer specimens and cell lines compared to their adjacent tissues and normal pancreatic cells. Growth and invasion of PANC-1 and SW1990 cells were attenuated by overexpression of miR-142 in vitro. With the help of bioinformatics analysis, hypoxia-inducible factor 1 (HIF-1α was identified to be a direct target of miR-142, and a luciferase reporter experiment confirmed this discovery. Overexpression of miR-142 decreases protein expression of HIF-1α. In the hypoxic microenvironment, HIF-1α was up-regulated while miR-142 was down-regulated. The invaded cells significantly increased in the hypoxic microenvironment compared to the normoxic microenvironment. The hypoxia treatment induced cells’ proliferation, and invasion could be inhibited by miR-142 overexpression or HIF-1α inhibition. Moreover, expression of epithelial-mesenchymal transition (EMT markers, Vimentin, VEGF-C and E-cad, was altered under hypoxia conditions and regulated by miR-142/HIF-1α. Above all, these findings provided insights on the functional mechanism of miR-142, suggesting that the miR-142/HIF-1α axis may interfere with the proliferative and invasive properties of pancreatic cancer cells, and indicated that miR-142 could be a potential therapeutic target for pancreatic cancer.

miR-24-2 controls H2AFX expression regardless of gene copy number alteration and induces apoptosis by targeting antiapoptotic gene BCL-2: a potential for therapeutic intervention.

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Srivastava, Niloo; Manvati, Siddharth; Srivastava, Archita; Pal, Ranjana; Kalaiarasan, Ponnusamy; Chattopadhyay, Shilpi; Gochhait, Sailesh; Dua, Raina; Bamezai, Rameshwar N K

2011-04-04

New levels of gene regulation with microRNA (miR) and gene copy number alterations (CNAs) have been identified as playing a role in various cancers. We have previously reported that sporadic breast cancer tissues exhibit significant alteration in H2AX gene copy number. However, how CNA affects gene expression and what is the role of miR, miR-24-2, known to regulate H2AX expression, in the background of the change in copy number, are not known. Further, many miRs, including miR-24-2, are implicated as playing a role in cell proliferation and apoptosis, but their specific target genes and the pathways contributing to them remain unexplored. Changes in gene copy number and mRNA/miR expression were estimated using real-time polymerase chain reaction assays in two mammalian cell lines, MCF-7 and HeLa, and in a set of sporadic breast cancer tissues. In silico analysis was performed to find the putative target for miR-24-2. MCF-7 cells were transfected with precursor miR-24-2 oligonucleotides, and the gene expression levels of BRCA1, BRCA2, ATM, MDM2, TP53, CHEK2, CYT-C, BCL-2, H2AFX and P21 were examined using TaqMan gene expression assays. Apoptosis was measured by flow cytometric detection using annexin V dye. A luciferase assay was performed to confirm BCL-2 as a valid cellular target of miR-24-2. It was observed that H2AX gene expression was negatively correlated with miR-24-2 expression and not in accordance with the gene copy number status, both in cell lines and in sporadic breast tumor tissues. Further, the cells overexpressing miR-24-2 were observed to be hypersensitive to DNA damaging drugs, undergoing apoptotic cell death, suggesting the potentiating effect of mir-24-2-mediated apoptotic induction in human cancer cell lines treated with anticancer drugs. BCL-2 was identified as a novel cellular target of miR-24-2. mir-24-2 is capable of inducing apoptosis by modulating different apoptotic pathways and targeting BCL-2, an antiapoptotic gene. The study suggests

miR-449 overexpression inhibits papillary thyroid carcinoma cell growth by targeting RET kinase-β-catenin signaling pathway.

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Li, Zongyu; Huang, Xin; Xu, Jinkai; Su, Qinghua; Zhao, Jun; Ma, Jiancang

2016-10-01

Papillary thyroid carcinoma (PTC) is the most common thyroid cancer and represent approximately 80% of all thyroid cancers. The present study is aimed to investigate the role of microRNA (miR)-449 in the progression of PTC. Our results revealed that miR-449 was underexpressed in the collected PTC specimens compared with non-cancerous PTC tissues. Overexpression of miR-449 induced a cell cycle arrest at G0/G1 phase and inhibited PTC cell growth in vitro. Further studies revealed that RET proto-oncogene (RET) is a novel miR-449 target, due to miR-449 bound directly to its 3'-untranslated region and miR-449 mimic reduced the protein expression of RET. Similar to the effects of miR-449 overexpression, RET downregulation inhibited cell growth, whereas RET overexpression reversed the inhibitive effect of miR-449 mimic. Furthermore, miR-449 overexpression inhibited the nuclear translocation of β-catenin and reduced the expression of several downstream genes, including c-Myc, cyclin D1, T cell-specific transcription factor (TCF) and lymphoid enhancer-binding factor 1 (LEF-1), and inactivated the β-catenin pathway in TPC-1 cells. Moreover, overexpression of β-catenin prevented miR-449-reduced cell cycle arrest and cell viability. In xenograft animal experiments, miR-449 overexpression effectively suppressed the tumor growth of PTC. Taken together, our research indicated that miR-449 functions as an anti-oncogene by targeting RET, and that miR-449 overexpression inhibited the growth of PTC by inactivating the β-catenin pathway. Thus, miR-449 may serve as a potential therapeutic strategy for the treatment of PTC.

Coronary heart disease-associated variation in TCF21 disrupts a miR-224 binding site and miRNA-mediated regulation.

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Miller, Clint L; Haas, Ulrike; Diaz, Roxanne; Leeper, Nicholas J; Kundu, Ramendra K; Patlolla, Bhagat; Assimes, Themistocles L; Kaiser, Frank J; Perisic, Ljubica; Hedin, Ulf; Maegdefessel, Lars; Schunkert, Heribert; Erdmann, Jeanette; Quertermous, Thomas; Sczakiel, Georg

2014-03-01

Genome-wide association studies (GWAS) have identified chromosomal loci that affect risk of coronary heart disease (CHD) independent of classical risk factors. One such association signal has been identified at 6q23.2 in both Caucasians and East Asians. The lead CHD-associated polymorphism in this region, rs12190287, resides in the 3' untranslated region (3'-UTR) of TCF21, a basic-helix-loop-helix transcription factor, and is predicted to alter the seed binding sequence for miR-224. Allelic imbalance studies in circulating leukocytes and human coronary artery smooth muscle cells (HCASMC) showed significant imbalance of the TCF21 transcript that correlated with genotype at rs12190287, consistent with this variant contributing to allele-specific expression differences. 3' UTR reporter gene transfection studies in HCASMC showed that the disease-associated C allele has reduced expression compared to the protective G allele. Kinetic analyses in vitro revealed faster RNA-RNA complex formation and greater binding of miR-224 with the TCF21 C allelic transcript. In addition, in vitro probing with Pb2+ and RNase T1 revealed structural differences between the TCF21 variants in proximity of the rs12190287 variant, which are predicted to provide greater access to the C allele for miR-224 binding. miR-224 and TCF21 expression levels were anti-correlated in HCASMC, and miR-224 modulates the transcriptional response of TCF21 to transforming growth factor-β (TGF-β) and platelet derived growth factor (PDGF) signaling in an allele-specific manner. Lastly, miR-224 and TCF21 were localized in human coronary artery lesions and anti-correlated during atherosclerosis. Together, these data suggest that miR-224 interaction with the TCF21 transcript contributes to allelic imbalance of this gene, thus partly explaining the genetic risk for coronary heart disease associated at 6q23.2. These studies implicating rs12190287 in the miRNA-dependent regulation of TCF21, in conjunction with

miR-371, miR-138, miR-544, miR-145, and miR-214 could modulate Th1/Th2 balance in asthma through the combinatorial regulation of Runx3.

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Qiu, Yu-Ying; Zhang, Ying-Wei; Qian, Xiu-Fen; Bian, Tao

2017-01-01

Asthma is tightly related to the imbalance of Th1/Th2 cells, and Runx3 plays a pivotal role in the differentiation of T helper cells. The present study aimed to investigate dysregulated microRNAs that may target Runx3 in CD4 + T cells from asthmatic patients and reveal Runx3 function in Th1/Th2 balance regulation. We detected the levels of Th1- and Th2-related cytokines by ELISA and analyzed the differentiation marker gene of T helper cells by qRT-PCR. Results indicated that an imbalance of Th1/Th2 cells was present in our asthmatic subject. Runx3 expression was reduced in the CD4 + T cells from asthmatic patients. Overexpression of Runx3 could restore the Th1/Th2 balance. After performing microRNA microarray assay, we found a series of microRNAs that were considerably altered in the CD4 + T cells from asthmatic patients. Among these upregulated microRNAs, eight microRNAs that may target Runx3 were selected by bioinformatics prediction. Five microRNAs, namely miR-371, miR-138, miR-544, miR-145, and miR-214, were confirmed by qRT-PCR and selected as candidate microRNAs. Luciferase reporter assay showed that these five microRNAs could directly target the 3'-UTR of Runx3. However, only simultaneous inhibition of these five microRNAs could alter the expression of Runx3. Most importantly, only simultaneous inhibition could improve the Th1/Th2 balance. Thus, we suggest that miR-371, miR-138, miR-544, miR-145, and miR-214 can modulate the Th1/Th2 balance in asthma by regulating Runx3 in a combinatorial manner.

Expression of programmed cell death protein 4 (PDCD4) and miR-21 in urothelial carcinoma

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Fischer, Nicolas, E-mail: simplissimus@gmx.de [Department of Urology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany); Goeke, Friederike, E-mail: Friederike.goeke@ukb.uni-bonn.de [Department of Pathology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany); Splittstoesser, Vera, E-mail: Veri.sp@web.de [Department of Urology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany); Lankat-Buttgereit, Brigitte, E-mail: Lankatbu@staff.uni-marburg.de [Department of Internal Medicine, Philipps-University of Marburg, Baldingerstrasse, 35043 Marburg (Germany); Mueller, Stefan C., E-mail: Stefan.mueller@ukb.uni-bonn.de [Department of Urology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany); Ellinger, Joerg, E-mail: Joerg.ellinger@ukb.uni-bonn.de [Department of Urology, University of Bonn, Sigmund-Freud-Strasse 25, 53127 Bonn (Germany)

2012-01-06

Highlights: Black-Right-Pointing-Pointer The tumor suppressor gene PDCD4 is down-regulated in many tumorous entities. Black-Right-Pointing-Pointer We investigate the impact of PDCD4 and its regulating factor miR-21 in urothelial carcinoma. Black-Right-Pointing-Pointer We confirm PDCD4 as a tumor suppressor gene and it could be a diagnostic marker for this tumor. -- Abstract: Background: We investigated the role of the programmed cell death 4 (PDCD4) tumor suppressor gene in specimens of transitional cell carcinoma and of healthy individuals. Methods: PDCD4 immunohistochemical expression was investigated in 294 cases in histologically proven transitional cell carcinoma in different tumorous stages (28 controls, 122 non-muscle invasive urothelial carcinoma, stages Tis-T1, 119 invasive transitional cell carcinoma stages T2-T4 and 25 metastases). MiR-21 expression, an important PDCD4 regulator, was assessed with real-time PCR analysis and showed inverse correlation to tissue PDCD4 expression. Results: Nuclear and cytoplasmatic PDCD4 immunostaining decreased significantly with histopathological progression of the tumor (p < 0001). Controls showed strong nuclear and cytoplasmatic immunohistochemical staining. MiR-21 up regulation in tissue corresponded to PDCD4 suppression. Conclusions: These data support a decisive role for PDCD4 down regulation in transitional cell carcinoma and confirm miR-21 as a negative regulator for PDCD4. Additionally, PDCD4 immunohistochemical staining turns out to be a possible diagnostic marker for transitional cell carcinoma.

Expression of programmed cell death protein 4 (PDCD4) and miR-21 in urothelial carcinoma

International Nuclear Information System (INIS)

Fischer, Nicolas; Göke, Friederike; Splittstößer, Vera; Lankat-Buttgereit, Brigitte; Müller, Stefan C.; Ellinger, Jörg

2012-01-01

Highlights: ► The tumor suppressor gene PDCD4 is down-regulated in many tumorous entities. ► We investigate the impact of PDCD4 and its regulating factor miR-21 in urothelial carcinoma. ► We confirm PDCD4 as a tumor suppressor gene and it could be a diagnostic marker for this tumor. -- Abstract: Background: We investigated the role of the programmed cell death 4 (PDCD4) tumor suppressor gene in specimens of transitional cell carcinoma and of healthy individuals. Methods: PDCD4 immunohistochemical expression was investigated in 294 cases in histologically proven transitional cell carcinoma in different tumorous stages (28 controls, 122 non-muscle invasive urothelial carcinoma, stages Tis-T1, 119 invasive transitional cell carcinoma stages T2–T4 and 25 metastases). MiR-21 expression, an important PDCD4 regulator, was assessed with real-time PCR analysis and showed inverse correlation to tissue PDCD4 expression. Results: Nuclear and cytoplasmatic PDCD4 immunostaining decreased significantly with histopathological progression of the tumor (p < 0001). Controls showed strong nuclear and cytoplasmatic immunohistochemical staining. MiR-21 up regulation in tissue corresponded to PDCD4 suppression. Conclusions: These data support a decisive role for PDCD4 down regulation in transitional cell carcinoma and confirm miR-21 as a negative regulator for PDCD4. Additionally, PDCD4 immunohistochemical staining turns out to be a possible diagnostic marker for transitional cell carcinoma.

miR-196a targets netrin 4 and regulates cell proliferation and migration of cervical cancer cells

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Zhang, Jie; Zheng, Fangxia; Yu, Gang; Yin, Yanhua; Lu, Qingyang

2013-01-01

Highlights: •miR-196a was overexpressed in cervical cancer tissue compared to normal tissue. •miR-196a expression elevated proliferation and migration of cervical cancer cells. •miR-196a inhibited NTN4 expression by binding 3′-UTR region of NTN4 mRNA. •NTN4 inversely correlated with miR-196a expression in cervical tissue and cell line. •NTN4 expression was low in cervical cancer tissue compared to normal tissue. -- Abstract: Recent research has uncovered tumor-suppressive and oncogenic potential of miR-196a in various tumors. However, the expression and mechanism of its function in cervical cancer remains unclear. In this study, we assess relative expression of miR-196a in cervical premalignant lesions, cervical cancer tissues, and four cancer cell lines using quantitative real-time PCR. CaSki and HeLa cells were treated with miR-196a inhibitors, mimics, or pCDNA/miR-196a to investigate the role of miR-196a in cancer cell proliferation and migration. We demonstrated that miR-196a was overexpressed in cervical intraepithelial neoplasia 2–3 and cervical cancer tissue. Moreover, its expression contributes to the proliferation and migration of cervical cancer cells, whereas inhibiting its expression led to a reduction in proliferation and migration. Five candidate targets of miR-196a chosen by computational prediction and Cervical Cancer Gene Database search were measured for their mRNA in both miR-196a-overexpressing and -depleted cancer cells. Only netrin 4 (NTN4) expression displayed an inverse association with miR-196a. Fluorescent reporter assays revealed that miR-196a inhibited NTN4 expression by targeting one binding site in the 3′-untranslated region (3′-UTR) of NTN4 mRNA. Furthermore, qPCR and Western blot assays verified NTN4 expression was downregulated in cervical cancer tissues compared to normal controls, and in vivo mRNA level of NTN4 inversely correlated with miR-196a expression. In summary, our findings provide new insights about the

Downregulation of miR-29a/b/c in placenta accreta inhibits apoptosis of implantation site intermediate trophoblast cells by targeting MCL1.

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Gu, Yongzhong; Bian, Yuehong; Xu, Xiaofei; Wang, Xietong; Zuo, Changting; Meng, Jinlai; Li, Hongyan; Zhao, Shigang; Ning, Yunnan; Cao, Yongzhi; Huang, Tao; Yan, Junhao; Chen, Zi-Jiang

2016-12-01

Placenta accreta is defined as abnormal adhesion of placental villi to the uterine myometrium. Although this condition has become more common as a result of the increasing rate of cesarean sections, the underlying causative mechanism(s) remain elusive. Because microRNA-29a/b/c (miR-29a/b/c) have been shown to play important roles in placental development, this study evaluated the roles of these microRNAs in placenta accreta. Expression of miR-29a/b/c and myeloid cell leukemia-1 (MCL1) were quantified in patient tissues and HTR8/SVneo trophoblast cells using the real-time quantitative polymerase chain reaction. Western blotting was used to analyze expression of the MCL1 protein in HTR8/SVneo trophoblast cells with altered expression of miR-29a/b/c. To determine their role in apoptosis, miR-29a/b/c were overexpressed in HTR-8/SVneo cells, and levels of apoptosis were analyzed by flow cytometry. Luciferase activity assays were used to determine whether MCL1 is a target gene of miR-29a/b/c. Expression of miR-29a/b/c was significantly lower in creta sites compared to noncreta sites (p = 0.018, 0.041, and 0.022, respectively), but expression of MCL1 was upregulated in creta sites (p = 0.039). MCL1 expression was significantly downregulated in HTR-8/SVneo cells overexpressing miR-29a/b/c (p = 0.002, 0.008, and 0.013, respectively). Luciferase activity assays revealed that miR-29a/b/c directly target the 3' untranslated region of MCL1 in 293T cells. Over-expression of miR-29a/b/c induced apoptosis in the HTR-8/SVneo trophoblast cell line. Moreover, histopathological evaluation revealed that the number of implantation site intermediate trophoblast (ISIT) cells was increased in creta sites and that these cells were positive for MCL1. Our results demonstrate that in placenta accreta, miR-29a/b/c inhibits apoptosis of ISIT cells by targeting MCL1. These findings provide new insights into the pathogenesis of placenta accreta. Copyright © 2016 Elsevier Ltd. All rights

Involvement of miR160/miR393 and their targets in cassava responses to anthracnose disease.

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Pinweha, Nattaya; Asvarak, Thipa; Viboonjun, Unchera; Narangajavana, Jarunya

2015-02-01

Cassava is a starchy root crop for food and industrial applications in many countries around the world. Among the factors that affect cassava production, diseases remain the major cause of yield loss. Cassava anthracnose disease is caused by the fungus Colletotrichum gloeosporioides. Severe anthracnose attacks can cause tip die-backs and stem cankers, which can affect the availability of planting materials especially in large-scale production systems. Recent studies indicate that plants over- or under-express certain microRNAs (miRNAs) to cope with various stresses. Understanding how a disease-resistant plant protects itself from pathogens should help to uncover the role of miRNAs in the plant immune system. In this study, the disease severity assay revealed different response to C. gloeosporioides infection in two cassava cultivars. Quantitative RT-PCR analysis uncovered the differential expression of the two miRNAs and their target genes in the two cassava cultivars that were subjected to fungal infection. The more resistant cultivar revealed the up-regulation of miR160 and miR393, and consequently led to low transcript levels in their targets, ARF10 and TIR1, respectively. The more susceptible cultivar exhibited the opposite pattern. The cis-regulatory elements relevant to defense and stress responsiveness, fungal elicitor responsiveness and hormonal responses were the most prevalent present in the miRNAs gene promoter regions. The possible dual role of these specific miRNAs and their target genes associated with cassava responses to C. gloeosporioides is discussed. This is the first study to address the molecular events by which miRNAs which might play a role in fungal-infected cassava. A better understanding of the functions of miRNAs target genes should greatly increase our knowledge of the mechanism underlying susceptibility and lead to new strategies to enhance disease tolerance in this economically important crop. Copyright © 2014 Elsevier GmbH. All

miR-338-3p functions as a tumor suppressor in gastric cancer by targeting PTP1B.

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Sun, Feng; Yu, Mengchao; Yu, Jing; Liu, Zhijian; Zhou, Xinyan; Liu, Yanqing; Ge, Xiaolong; Gao, Haidong; Li, Mei; Jiang, Xiaohong; Liu, Song; Chen, Xi; Guan, Wenxian

2018-05-09

Gastric cancer (GC) is one of the most common malignant tumors and peritoneal metastasis is the primary cause for advanced GC's mortality. Protein-tyrosine phosphatase 1B (PTP1B) functions as an oncogene and involves in carcinogenesis and cancer dissemination. However, the function and regulation of PTP1B in GC remain poorly understood. In this study, we found that PTP1B was upregulated in GC tissues and overexpression of PTP1B in vitro promoted cell migration and prevented apoptosis. Then, we predicted that PTP1B was a target of miR-338-3p and we revealed an inverse correlation between miR-338-3p levels and PTP1B protein levels in GC tissues. Next, we verified that PTP1B was inhibited by miR-338-3p via direct targeting to its 3'-untranslated regions. Moreover, overexpression of miR-338-3p in vitro attenuated GC cell migration and promoted apoptosis, and these effects could be partially reversed by reintroduction of PTP1B. Finally, we established an orthotopic xenograft model and a peritoneal dissemination model of GC to demonstrate that miR-338-3p restrained tumor growth and dissemination in vivo by targeting PTP1B. Taken together, our results highlight that PTP1B is an oncogene and is negatively regulated by miR-338-3p in GC, which may provide new insights into novel molecular therapeutic targets for GC.

Downregulation of miR-221-3p contributes to IL-1β-induced cartilage degradation by directly targeting the SDF1/CXCR4 signaling pathway.

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Zheng, Xin; Zhao, Feng-Chao; Pang, Yong; Li, Dong-Ya; Yao, Sheng-Cheng; Sun, Shao-Song; Guo, Kai-Jin

2017-06-01

Osteoarthritis (OA) is characterized by degradation of chondrocyte extracellular matrix (ECM). Accumulating evidence suggests that microRNAs (miRNAs) are associated with OA, but little is known of their function in chondrocyte ECM degradation. The objective of this study was to investigate the expression and function of miRNAs in OA. miRNA expression profile was determined in OA cartilage tissues and controls, employing Solexa sequencing and reverse transcription quantitative PCR (RT-qPCR). According to a modified Mankin scale, cartilage degradation was evaluated. Functional analysis of the miRNAs on chondrocyte ECM degradation was performed after miRNA transfection and IL-1β treatment. Luciferase reporter assays and western blotting were employed to determine miRNA targets. Expression of miR-221-3p was downregulated in OA cartilage tissues, which was significantly correlated with a modified Mankin scale. Through gain-of-function and loss-of-function studies, miR-221-3p was shown to significantly affect matrix synthesis gene expression and chondrocyte proliferation and apoptosis. Using SW1353 and C28I2 cells, SDF1 was identified as a target of miR-221-3p. SDF1 overexpression resulted in increased expression of catabolic genes such as MMP-13 and ADAMTS-5 in response to IL-1β, but these effects were moderated by miR-221-3p. SDF1 treatment antagonized this effect, while knockdown of SDF1 by shSDF1 induced inhibitory effects on the expression of CXCR4 and its main target genes, similar to miR-221-3p. The results indicate that upregulation of miR-221-3p could prevent IL-1β-induced ECM degradation in chondrocytes. Targeting the SDF1/CXCR4 signaling pathway may be used as a therapeutic approach for OA. miR-221-3p is downregulated in human cartilage tissues. miR-221-3p levels are associated with cartilage degeneration grade. miR-221-3p upregulation prevents IL-1β-induced ECM degradation in chondrocytes. Protection of ECM degradation by miR-223-3p occurs via SDF1/CXCR4

miR-126 Functions as a Tumor Suppressor in Osteosarcoma by Targeting Sox2

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Chenglin Yang

2013-12-01

Full Text Available Osteosarcoma (OS is the most common malignant bone tumor in children and young adults, the early symptoms and signs of which are non-specific. The discovery of microRNAs (miRNAs provides a new avenue for the early diagnosis and treatment of OS. miR-126 has been reported to be highly expressed in vascularized tissues, and is recently widely studied in cancers. Herein, we explored the expression and significance of miR-126 in OS. Using TaqMan RT-PCR analysis, we analyzed the expression of miR-126 in 32 paired OS tumor tissues and 4 OS cell lines and found that miR-126 was consistently under-expressed in OS tissues and cell lines compared with normal bone tissues and normal osteoblast cells (NHOst, respectively. As miR-126 is significantly decreased in OS tissues and cell lines, we sought to compensate for its loss through exogenous transfection into MG-63 cells with a miR-126 mimic. Ectopic expression of miR-126 inhibited cell proliferation, migration and invasion, and induced apoptosis of MG-63 cells. Moreover, bioinformatic prediction suggested that the sex-determining region Y-box 2 (Sox2 is a target gene of miR-126. Using mRNA and protein expression analysis, luciferase assays and rescue assays, we demonstrate that restored expression of Sox2 dampened miR-126-mediated suppression of tumor progression, which suggests the important role of miR-126/Sox2 interaction in tumor progression. Taken together, our data indicate that miR-126 functions as a tumor suppressor in OS, which exerts its activity by suppressing the expression of Sox2.

Hepatitis B virus X protein mutant HBxΔ127 promotes proliferation of hepatoma cells through up-regulating miR-215 targeting PTPRT

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Liu, Fabao [Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin 300071 (China); Department of Biochemistry, College of Life Sciences, Nankai University, Tianjin 300071 (China); You, Xiaona [Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin 300071 (China); Chi, Xiumei [Department of Hepatology, The First Hospital, Jilin University, Changchun 130021 (China); Wang, Tao [Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin 300071 (China); Ye, Lihong [Department of Biochemistry, College of Life Sciences, Nankai University, Tianjin 300071 (China); Niu, Junqi, E-mail: junqiniu@yahoo.com.cn [Department of Hepatology, The First Hospital, Jilin University, Changchun 130021 (China); Zhang, Xiaodong, E-mail: zhangxd@nankai.edu.cn [Department of Cancer Research, College of Life Sciences, Nankai University, Tianjin 300071 (China)

2014-02-07

Highlights: • Relative to wild type HBx, HBX mutant HBxΔ127 strongly enhances cell proliferation. • Relative to wild type HBx, HBxΔ127 remarkably up-regulates miR-215 in hepatoma cells. • HBxΔ127-elevated miR-215 promotes cell proliferation via targeting PTPRT mRNA. - Abstract: The mutant of virus is a frequent event. Hepatitis B virus X protein (HBx) plays a vital role in the development of hepatocellular carcinoma (HCC). Therefore, the identification of potent mutant of HBx in hepatocarcinogenesis is significant. Previously, we identified a natural mutant of the HBx gene (termed HBxΔ127). Relative to wild type HBx, HBxΔ127 strongly enhanced cell proliferation and migration in HCC. In this study, we aim to explore the mechanism of HBxΔ127 in promotion of proliferation of hepatoma cells. Our data showed that both wild type HBx and HBxΔ127 could increase the expression of miR-215 in hepatoma HepG2 and H7402 cells. However, HBxΔ127 was able to significantly increase miR-215 expression relative to wild type HBx in the cells. We identified that protein tyrosine phosphatase, receptor type T (PTPRT) was one of the target genes of miR-215 through targeting 3′UTR of PTPRT mRNA. In function, miR-215 was able to promote the proliferation of hepatoma cells. Meanwhile anti-miR-215 could partially abolish the enhancement of cell proliferation mediated by HBxΔ127 in vitro. Knockdown of PTPRT by siRNA could distinctly suppress the decrease of cell proliferation mediated by anti-miR-215 in HepG2-XΔ127/H7402-XΔ127 cells. Moreover, we found that anti-miR-215 remarkably inhibited the tumor growth of hepatoma cells in nude mice. Collectively, relative to wild type HBx, HBxΔ127 strongly enhances proliferation of hepatoma cells through up-regulating miR-215 targeting PTPRT. Our finding provides new insights into the mechanism of HBx mutant HBxΔ127 in promotion of proliferation of hepatoma cells.

Control of sulfate concentration by miR395-targeted APS genes in Arabidopsis thaliana

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Qin Ai

2016-04-01

Full Text Available Sulfur nutrition is crucial for plant growth and development, as well as crop yield and quality. Inorganic sulfate in the soil is the major sulfur source for plants. After uptake, sulfate is activated by ATP sulfurylase, and then gets assimilated into sulfur-containing metabolites. However, the mechanism of regulation of sulfate levels by ATP sulfurylase is unclear. Here, we investigated the control of sulfate levels by miR395-mediated regulation of APS1/3/4. Sulfate was over-accumulated in the shoots of miR395 over-expression plants in which the expression of the APS1, APS3, and APS4 genes was suppressed. Accordingly, reduced expression of miR395 caused a decline of sulfate concentration. In agreement with these results, over-expression of the APS1, APS3, and APS4 genes led to the reduction of sulfate levels. Differential expression of these three APS genes in response to sulfate starvation implied that they have different functions. Further investigation revealed that the regulation of sulfate levels mediated by miR395 depends on the repression of its APS targets. Unlike the APS1, APS3, and APS4 genes, which encode plastid-localized ATP sulfurylases, the APS2 gene encodes a cytosolic version of ATP sulfurylase. Genetic analysis indicated that APS2 has no significant effect on sulfate levels. Our data suggest that miR395-targeted APS genes are key regulators of sulfate concentration in leaves.

Evaluation of miR-21 Inhibition and its Impact on Cancer Susceptibility Candidate 2 Long Noncoding RNA in Colorectal Cancer Cell Line

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Miganoosh Simonian

2018-01-01

Full Text Available Background: Both microRNAs (miRNAs and long noncoding RNAs (lncRNAs have been shown to have a critical role in the regulation of cellular processes such as cell growth and apoptosis, as well as cancer progression and metastasis. lncRNAs are aberrantly expressed in many diseases including cancer. Although it is well known that miRNAs can target a large number of protein-coding genes, little is known whether miRNAs can also target lncRNAs. In the present study, we determine whether miR-21 can regulate lncRNA cancer susceptibility candidate 2 (CASC2 in colorectal cancer. Materials and Methods: LS174T cells were transfected with locked nucleic acid (LNA-anti-miR-21 and scrambled LNA for 24, 48 and 72 h. The expression of miR-21 and lncCASC2 was evaluated by quantitative reverse transcriptase polymerase chain reaction. Results: However, contrary to what we expected and reported by others, lncCASC2 quantity was significantly reduced in LNA treated LS174T cells compared to the scrambled treated and normal untreated cells (P < 0.05. Conclusion: The interaction of miRNA and lncRNA are not as simple as suggested by other reports. Moreover, it could be complex molecular mechanisms underlying the communication of various noncoding RNA elements.

Identification of miR-2400 gene as a novel regulator in skeletal muscle satellite cells proliferation by targeting MYOG gene

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Zhang, Wei Wei [The Laboratory of Cell and Development, Northeast Agricultural University, Harbin, Heilongjiang 150030 (China); College of Life Sciences and Agriculture & Forestry, Qiqihar University, Qiqihar, Heilongjiang 161006 (China); Tong, Hui Li; Sun, Xiao Feng; Hu, Qian; Yang, Yu; Li, Shu Feng [The Laboratory of Cell and Development, Northeast Agricultural University, Harbin, Heilongjiang 150030 (China); Yan, Yun Qin, E-mail: yanyunqin@sohu.com [The Laboratory of Cell and Development, Northeast Agricultural University, Harbin, Heilongjiang 150030 (China); Li, Guang Peng [The Key Laboratory of Mammal Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Hohhot 010021 (China)

2015-08-07

MicroRNAs play critical roles in skeletal muscle development as well as in regulation of muscle cell proliferation and differentiation. Previous study in our laboratory showed that the expression level of miR-2400, a novel and unique miRNA from bovine, had significantly changed in skeletal muscle-derived satellite cells (MDSCs) during differentiation, however, the function and expression pattern for miR-2400 in MDSCs has not been fully understood. In this report, we firstly identified that the expression levels of miR-2400 were down-regulated during MDSCs differentiation by stem-loop RT-PCR. Over-expression and inhibition studies demonstrated that miR-2400 promoted MDSCs proliferation by EdU (5-ethynyl-2′ deoxyuridine) incorporation assay and immunofluorescence staining of Proliferating cell nuclear antigen (PCNA). Luciferase reporter assays showed that miR-2400 directly targeted the 3′ untranslated regions (UTRs) of myogenin (MYOG) mRNA. These data suggested that miR-2400 could promote MDSCs proliferation through targeting MYOG. Furthermore, we found that miR-2400, which was located within the eighth intron of the Wolf-Hirschhorn syndrome candidate 1-like 1 (WHSC1L1) gene, was down-regulated in MDSCs in a direct correlation with the WHSC1L1 transcript by Clustered regularly interspaced palindromic repeats interference (CRISPRi). In addition, these observations not only provided supporting evidence for the codependent expression of intronic miRNAs and their host genes in vitro, but also gave insight into the role of miR-2400 in MDSCs proliferation. - Highlights: • miR-2400 is a novel and unique miRNA from bovine. • miR-2400 could promote skeletal muscle satellite cells proliferation. • miR-2400 directly targeted the 3′ untranslated regions of MYOG mRNA. • miR-2400 could be coexpressed together with its host gene WHSC1L1.

PSMB4 promotes multiple myeloma cell growth by activating NF-κB-miR-21 signaling

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Zheng, Peihao; Guo, Honggang [Department of Hematology, Navy General Hospital, Beijing 100048 (China); Li, Guangchao [School of Bioscience and Bioengineering, South China University of Technology, Guangzhou 510006 (China); Han, Siqi [Department of Medical Oncology, Jinling Hospital, Nanjing 210002 (China); Luo, Fei [Department of Stomatology, Jinling Hospital, Nanjing 210002 (China); Liu, Yi, E-mail: liuyi2033@163.com [Department of Hematology, Navy General Hospital, Beijing 100048 (China)

2015-03-06

Proteasomal subunit PSMB4, was recently identified as potential cancer driver genes in several tumors. However, the regulatory mechanism of PSMB4 on carcinogenesis process remains unclear. In this study, we investigated the expression and roles of PSMB4 in multiple myeloma (MM). We found a significant up-regulation of PSMB4 in MM plasma and cell lines. Ectopic overexpression of PSMB4 promoted cell growth and colony forming ability of MM cells, whereas inhibition of PSMB4 led to a decrease of such events. Furthermore, our results demonstrated the up-regulation of miR-21 and a positive correlation between the levels of miR-21 and PSMB4 in MM. Re-expression of miR-21 markedly rescued PSMB4 knockdown-mediated suppression of cell proliferation and clone-formation. Additionally, while enforced expression of PSMB4 profoundly increased NF-κB activity and the level of miR-21, PSMB4 knockdown or NF-κB inhibition suppressed miR-21 expression in MM cells. Taken together, our results demonstrated that PSMB4 regulated MM cell growth in part by activating NF-κB-miR-21 signaling, which may represent promising targets for novel specific therapies. - Highlights: • First reported upregulation of PSMB4 in MM plasma and cell lines. • PSMB4 promoted MM cell growth and colony forming ability. • Further found miR-21 was up-regulated by PSMB4 in MM plasma and cell lines. • PSMB4-induced miR-21 expression was modulated by NF-κB. • PSMB4-NF-κB-miR-21 axis may be potential therapeutic targets of MM.

Altered regulation of miR-34a and miR-483-3p in alcoholic hepatitis and DDC fed mice.

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Liu, Hui; French, Barbara A; Li, Jun; Tillman, Brittany; French, Samuel W

2015-12-01

MicroRNAs are small noncoding RNAs that negatively regulate gene expression by binding to the untranslated regions of their target mRNAs. Deregulation of miRNAs is shown to play pivotal roles in tumorigenesis and progression. Mallory-Denk Bodies (MDBs) are prevalent in various liver diseases including alcoholic hepatitis (AH) and are formed in mice livers by feeding DDC. By comparing AH livers where MDBs had formed with normal livers, there were significant changes of miR-34a and miR-483-3p by RNA sequencing (RNA-Seq) analyses. Real-time PCR further shows a 3- and 6-fold upregulation (respectively) of miR-34a in the AH livers and in the livers of DDC re-fed mice, while miR-483-3p was significantly downregulated in AH and DDC re-fed mice livers. This indicates that miR-34a and miR-483-3p may be crucial for liver MDB formation. P53 mRNA was found to be significantly downregulated both in the AH livers and in the livers of DDC re-fed mice, indicating that the upregulation of miR-34a is permitted by the decrease of p53 in AH since miR-34a is a main target of p53. Overexpression of miR-34a leads to an increase of p53 targets such as p27, which inhibits the cell cycle leading to cell cycle arrest. Importantly, BRCA1 is a target gene of miR-483-3p by RNA-Seq analyses and the downregulation of miR-483-3p may be the mechanism for liver MDB formation since the BRCA1 signal was markedly upregulated in AH livers. These results constitute a demonstration of the altered regulation of miR-34a and miR-483-3p in the livers of AH and mice fed DDC where MDBs formed, providing further insight into the mechanism of MDB formation mediated by miR-34a and miR-483-3p in AH. Copyright © 2015 Elsevier Inc. All rights reserved.

[miR-143 inhibits cell proliferation through targeted regulating the expression of K-ras gene in HeLa cells].

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Qin, H X; Cui, H K; Pan, Y; Hu, R L; Zhu, L H; Wang, S J

2016-12-23

Objective: To explore the effect of microRNA miR-143 on the proliferation of cervical cancer HeLa cells through targeted regulating the expression of K-ras gene. Methods: The luciferase report carrier containing wild type 3'-UTR of K-ras gene (K-ras-wt) or mutated 3'-UTR of the K-ras (K-ras-mut) were co-transfected with iR-143 mimic into the HeLa cells respectively, and the targeting effect of miR-143 in the transfectants was verified by the dual luciferase report system. HeLa cells were also transfected with miR-143 mimic (miR-143 mimic group), mimic control (negative control group), and miR-143 mimic plus K-ras gene (miR-143 mimic+ K-ras group), respectively. The expression of miR-143 in the transfected HeLa cells was detected by real-time PCR (RT-PCR), and the expression of K-ras protein was detected by Western blot. The cell proliferation activity of each group was examined by MTT assay. In addition, human cervical cancer tissue samples ( n =5) and cervical intraepithelial neoplasia tissue samples ( n =5) were also examined for the expression of miR-143 and K-ras protein by RT-PCR and Western blot, respectively. Results: The luciferase report assay showed that co-transfection with miR-143 mimic decreased the luciferase activity of the K-ras-wt significantly, but did not inhibit the luciferase activity of the K-ras-mut. The expression of miR-143 in the HeLa cells transfected with miR-143 mimic was significantly higher than that in the HeLa cells transfected with the mimic control (3.31±0.45 vs 0.97±0.22, P cell proliferative activity of the miR-143 mimic group was significantly lower than that of the negative control group ( P cell proliferative activity of the miR-143 mimic+ K-ras group was also significantly lower than the control group ( P HeLa cells through targeted regulating the expression of K-ras gene. In human cervical cancer tissues of a small sample set, the expression of miR-143 is downregulated, and the expression of K-ras is upregulated.

MiR-130a regulates neurite outgrowth and dendritic spine density by targeting MeCP2

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Yunjia Zhang

2016-06-01

Full Text Available ABSTRACT MicroRNAs (miRNAs are critical for both development and function of the central nervous system. Significant evidence suggests that abnormal expression of miRNAs is associated with neurodevelopmental disorders. MeCP2 protein is an epigenetic regulator repressing or activating gene transcription by binding to methylated DNA. Both loss-of-function and gain-of-function mutations in the MECP2 gene lead to neurodevelopmental disorders such as Rett syndrome, autism and MECP2 duplication syndrome. In this study, we demonstrate that miR-130a inhibits neurite outgrowth and reduces dendritic spine density as well as dendritic complexity. Bioinformatics analyses, cell cultures and biochemical experiments indicate that miR-130a targets MECP2 and down-regulates MeCP2 protein expression. Furthermore, expression of the wild-type MeCP2, but not a loss-of-function mutant, rescues the miR-130a-induced phenotype. Our study uncovers the MECP2 gene as a previous unknown target for miR-130a, supporting that miR-130a may play a role in neurodevelopment by regulating MeCP2. Together with data from other groups, our work suggests that a feedback regulatory mechanism involving both miR-130a and MeCP2 may serve to ensure their appropriate expression and function in neural development.

miR-150-5p inhibits hepatoma cell migration and invasion by targeting MMP14.

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Tao Li

Full Text Available Hepatocellular carcinoma (HCC is one of the leading causes of cancer-related mortality worldwide. Despite progress in diagnostics and treatment of HCC, its prognosis remains poor because the molecular mechanisms underlying hepatocarcinogenesis are not well understood. In the study, we focused on identifying the role of miRNAs in HCC progression. miRNA microarray was used to analyze the differentially expressed miRNAs, and the results were validated by qPCR. We found that the miR-150-5p expression is down-regulated in HCC tissues compared with pair non-tumor tissues. miR-150-5p expression is also decreased in metastatic cancer tissues compared with pair primary tissues, indicating that miR-150-5p may be involved in HCC metastasis. Functionally, miR-150-5p inhibition significantly promotes hepatoma cell migration and invasion, whereas miR-150-5p overexpression suppresses cancer cell migration and invasion in vitro. The matrix metalloproteinase 14 (MMP14 is identified as a new target gene of miR-150-5p. miR-150-5p markedly inhibits MMP14 expression in hepatoma cells, and miR-150-5p expression is negative correlation with MMP14 expression in vivo. More important, re-expression of MMP14 in hepatoma cells partially reverses the effect of miR-150-5p in inhibiting cell invasion.

miR-885-5p upregulation promotes colorectal cancer cell proliferation and migration by targeting suppressor of cytokine signaling.

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Su, Meng; Qin, Baoli; Liu, Fang; Chen, Yuze; Zhang, Rui

2018-07-01

The aim of the present study was to investigate the role of microRNA (miR)-885-5p in colorectal cancer cell proliferation and migration, and to determine the possible underlying molecular mechanisms. The expression of miR-885-5p in colorectal cancer tissue and cells was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The expression levels of three suppressor of cytokine signaling (SOCS) factors were detected by RT-qPCR and western blotting. The effects of miR-885-5p on tumor cell proliferation and migration were studied using MTT and Transwell assays, respectively. Additionally, the expression levels of epithelial-mesenchymal transition (EMT)-related proteins (N-cadherin, E-cadherin, vimentin and Snail) were detected by RT-qPCR and western blot analysis. Furthermore, the target of miR-885-5p was predicted and confirmed using a luciferase reporter assay. miR-885-5p was demonstrated to be upregulated and SOCS was downregulated in colorectal cancer tissue, and cells. miR-885-5p suppression significantly inhibited tumor cell proliferation and migration, promoted E-cadherin expression, and inhibited the expression levels of N-cadherin, vimentin and Snail. Further studies showed that SOCS5, SOCS6 and SOCS7 were direct targets of miR-885-5p. The results suggest that miR-885-5p suppression inhibited cell proliferation and migration, and the EMT process by targeting SOCS5, SOCS6 and SOCS7 genes in colorectal cancer. miR-885-5p and SOCS may be used for the diagnosis and treatment of colorectal cancer.

MYC through miR-17-92 Suppresses Specific Target Genes to Maintain Survival, Autonomous Proliferation, and a Neoplastic State

KAUST Repository

Li, Yulin; Choi, Peter  S.; Casey, Stephanie  C.; Dill, David  L.; Felsher, Dean  W.

2014-01-01

The MYC oncogene regulates gene expression through multiple mechanisms, and its overexpression culminates in tumorigenesis. MYC inactivation reverses turmorigenesis through the loss of distinguishing features of cancer, including autonomous proliferation and survival. Here we report that MYC via miR-17-92 maintains a neoplastic state through the suppression of chromatin regulatory genes Sin3b, Hbp1, Suv420h1, and Btg1, as well as the apoptosis regulator Bim. The enforced expression of miR-17-92 prevents MYC suppression from inducing proliferative arrest, senescence, and apoptosis and abrogates sustained tumor regression. Knockdown of the five miR-17-92 target genes blocks senescence and apoptosis while it modestly delays proliferative arrest, thus partially recapitulating miR-17-92 function. We conclude that MYC, via miR-17-92, maintains a neoplastic state by suppressing specific target genes.

MYC through miR-17-92 Suppresses Specific Target Genes to Maintain Survival, Autonomous Proliferation, and a Neoplastic State

KAUST Repository

Li, Yulin

2014-08-01

The MYC oncogene regulates gene expression through multiple mechanisms, and its overexpression culminates in tumorigenesis. MYC inactivation reverses turmorigenesis through the loss of distinguishing features of cancer, including autonomous proliferation and survival. Here we report that MYC via miR-17-92 maintains a neoplastic state through the suppression of chromatin regulatory genes Sin3b, Hbp1, Suv420h1, and Btg1, as well as the apoptosis regulator Bim. The enforced expression of miR-17-92 prevents MYC suppression from inducing proliferative arrest, senescence, and apoptosis and abrogates sustained tumor regression. Knockdown of the five miR-17-92 target genes blocks senescence and apoptosis while it modestly delays proliferative arrest, thus partially recapitulating miR-17-92 function. We conclude that MYC, via miR-17-92, maintains a neoplastic state by suppressing specific target genes.

Threshold-dependent repression of SPL gene expression by miR156/miR157 controls vegetative phase change in Arabidopsis thaliana.

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Jia He

2018-04-01

Full Text Available Vegetative phase change is regulated by a decrease in the abundance of the miRNAs, miR156 and miR157, and the resulting increase in the expression of their targets, SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL transcription factors. To determine how miR156/miR157 specify the quantitative and qualitative changes in leaf morphology that occur during vegetative phase change, we measured their abundance in successive leaves and characterized the phenotype of mutations in different MIR156 and MIR157 genes. miR156/miR157 decline rapidly between leaf 1&2 and leaf 3 and decrease more slowly after this point. The amount of miR156/miR157 in leaves 1&2 greatly exceeds the threshold required to specify their identity. Subsequent leaves have relatively low levels of miR156/miR157 and are sensitive to small changes in their abundance. In these later-formed leaves, the amount of miR156/miR157 is close to the threshold required to specify juvenile vs. adult identity; a relatively small decrease in the abundance of miR156/157 in these leaves produces a disproportionately large increase in SPL proteins and a significant change in leaf morphology. miR157 is more abundant than miR156 but has a smaller effect on shoot morphology and SPL gene expression than miR156. This may be attributable to the inefficiency with which miR157 is loaded onto AGO1, as well as to the presence of an extra nucleotide at the 5' end of miR157 that is mis-paired in the miR157:SPL13 duplex. miR156 represses different targets by different mechanisms: it regulates SPL9 by a combination of transcript cleavage and translational repression and regulates SPL13 primarily by translational repression. Our results offer a molecular explanation for the changes in leaf morphology that occur during shoot development in Arabidopsis and provide new insights into the mechanism by which miR156 and miR157 regulate gene expression.

Lower expressed miR-198 and its potential targets in hepatocellular carcinoma: a clinicopathological and in silico study

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Huang WT

2016-08-01

Full Text Available Wen-Ting Huang,1,* Han-Lin Wang,1,* Hong Yang,2 Fang-Hui Ren,1 Yi-Huan Luo,1 Chun-Qin Huang,1 Yue-Ya Liang,1 Hai-Wei Liang,1 Gang Chen,1 Yi-Wu Dang1 1Department of Pathology, 2Department of Ultrasonography, First Affiliated Hospital of Guangxi Medical University, Nanning, People’s Republic of China *These authors contributed equally to this work Purpose: To investigate the clinicopathological value and potential roles of microRNA-198 (miR-198 in hepatocellular carcinoma (HCC.Methods: Ninety-five formalin-fixed paraffin-embedded HCC and the para-cancerous liver tissues were gathered. Real-time reverse transcription quantitative polymerase chain reaction was applied to determine the miR-198 expression. The association between the miR-198 expression and clinicopathological features was examined. Meanwhile, potential target messenger RNAs of miR-198 in HCC were obtained from 14 miRNA prediction databases and natural language processing method, in which we pooled the genes related to the tumorigenesis and progression of HCC and classified them by their frequency. The selected target genes were finally analyzed in the Gene Ontology analysis and Kyoto Encyclopedia of Genes and Genomes pathway.Results: miR-198 expression was significantly lower in HCC than that in adjacent noncancerous liver tissues (1.30±0.72 vs 2.01±0.58, P<0.001. Low miR-198 expression was also correlated to hepatitis C virus infection (r=-0.48, P<0.001, tumor capsular infiltration (r=-0.43, P<0.001, metastasis (r=-0.26, P<0.010, number of tumor nodes (r=-0.25, P=0.013, vaso-invasion (r=-0.24, P=0.017, and clinical tumor node metastasis stage (r=-0.23, P=0.024. Altogether, 1,048 genes were achieved by the concurrent prediction from at least four databases and natural language processing indicated 1,800 genes for HCC. Further, 127 overlapping targets were further proceeded with for pathway analysis. The most enriched Gene Ontology terms in the potential target messenger RNAs of miR

MiR-17/20/93/106 promote hematopoietic cell expansion by targeting sequestosome 1-regulated pathways in mice

NARCIS (Netherlands)

J.C.M. Meenhuis (Annemarie); P. van Veelen (Peter); H. de Looper (Hans); N. van Boxtel (Nicole); I.J. van den Berge (Iris); S.M. Sun; E. Taskesen (Erdogan); P. Stern (Patrick); A. de Ru (Arnoud); A.J. van Adrichem (Arjan); J.A.A. Demmers (Jeroen); M. Jongen-Lavrencic (Mojca); B. Löwenberg (Bob); I.P. Touw (Ivo); P.A. Sharp (Phillip); S.J. Erkeland (Stefan)

2011-01-01

textabstractMicroRNAs (miRNAs) are pivotal for regulation of hematopoiesis but their critical targets remain largely unknown. Here, we show that ectopic expression of miR-17, -20,-93 and -106, all AAAGUGC seed-containing miRNAs, increases proliferation, colony outgrowth and replating capacity of

Increased Expression of miR-146a in Children With Allergic Rhinitis After Allergen-Specific Immunotherapy.

Science.gov (United States)

Luo, Xi; Hong, Haiyu; Tang, Jun; Wu, Xingmei; Lin, Zhibin; Ma, Renqiang; Fan, Yunping; Xu, Geng; Liu, Dabo; Li, Huabin

2016-03-01

MicroRNAs (miRs) were recently recognized to be important for immune cell differentiation and immune regulation. However, whether miRs were involved in allergen-specific immunotherapy (SIT) remains largely unknown. This study sought to examine changes in miR-146a and T regulatory cells in children with persistent allergic rhinitis (AR) after 3 months of subcutaneous immunotherapy (SCIT) and sublingual immunotherapy (SLIT). Twenty-four HDM-sensitized children with persistent AR were enrolled and treated with SCIT (n=13) or SLIT (n=11) for 3 months. Relative miR-146a and Foxp3 mRNA expression, the TRAF6 protein level, and the ratio of post-treatment to baseline IL-10+CD4+ T cells between the SCIT and SLIT groups were examined in the peripheral blood mononuclear cells (PBMCs) of AR patients using quantitative reverse transcription polymerase chain reaction (qRT-PCR), flow cytometry, and Western blot analysis, respectively. Serum levels of IL-5 and IL-10 were determined using ELISA. After 3 months of SIT, both the TNSS and INSS scores were significantly decreased compared to the baseline value (P<0.01). The relative expression of miR-146a and Foxp3 mRNA was significantly increased after both SCIT and SLIT (P<0.01). The ratio of post-treatment to baseline IL-10⁺CD4⁺ T cells and the serum IL-10 level were significantly increased in both the SCIT and SLIT groups (P<0.01), whereas the TRAF6 protein level and serum IL-5 level were significantly decreased (P<0.01). No significant differences in these biomarkers were observed between the SCIT and SLIT groups. Our findings suggest that miR-146a and its related biomarkers may be comparably modulated after both SCIT and SLIT, highlighting miR-146a as a potential therapeutic target for the improved management of AR.

Mel-18 negatively regulates stem cell-like properties through downregulation of miR-21 in gastric cancer.