Direct Target Of miR-124 Research Articles

MicroRNA-145 targets TRIM2 and exerts tumor-suppressing functions in epithelial ovarian cancer

ObjectiveEpithelial ovarian cancer (EOC) is one of the most common cancers in women worldwide but relatively little is known about its molecular pathogenesis. MicroRNAs, which regulate gene expression post-transcriptionally, have emerged as key players in tumorigenesis. The present study aims to investigate the dysregulation of miR-145 in EOC. MethodsmiRNA expression was assessed in EOC tissues and cell lines by quantitative reverse transcription (RT)-PCR. Xenograft mouse model was used for evaluation of the effect of miR-145 on tumor growth. Cell proliferation, colony formation assays, invasion assay, flow cytometry, Western blot and gene expression analysis were used for identification of the functional role of miR-145 in EOC cells. Luciferase reporter assay was used to confirm the interaction between miR-145 and its target mRNA 3′-UTR. ResultsmiR-145 expression was downregulated in EOC tissues and cell lines as compared with normal ovarian tissues. Transfection of miR-145 agomiR significantly inhibited the proliferation, colony forming ability, invasiveness and in vivo tumorigenicity of EOC cells. Transfection of agomiR-145 into EOC cells also markedly induced apoptosis. Furthermore, computational algorithm combined with luciferase reporter assays identified TRIM2 as the direct target of miR-145 in EOC cells. To this end, agomiR-145 downregulated TRIM2 to derepress Bim (a pro-apoptotic Bcl-2 family member degraded by TRIM2). ConclusionsThese data confirmed the tumor-suppressing function of miR-145 in EOC and identified TRIM2 as a new miR-145 target. In vivo delivery of agomiR-145 might be a feasible approach for miRNA-directed cancer therapy.

The candidate oncogene (MCRS1) promotes the growth of human lung cancer cells via the miR-155-Rb1 pathway.

BackgroundMicrospherule protein 1 (MCRS1) is a candidate oncogene and participates in various cellular processes, including growth, migration, senescence and transformation. MCRS1 is overexpressed in non-small cell lung cancer (NSCLC) and promotes the growth of cancer cells. However, the mechanisms driving these processes are not fully understood.MethodsRetrovirus-mediated RNA interference was employed to knockdown MCRS1 expression in cell lines. Cell proliferation assays and animal experiments were respectively performed to evaluate the growth of NSCLC cells in vitro and in vivo. Microarray analysis was carried out for mRNA profiling. Luciferase reporter assay and microRNA (miRNA) transfection were used to investigate the interaction between miRNA and gene.ResultsStably knocking down MCRS1 expression inhibited the proliferation of NSCLC cells in vitro and in vivo. By comparing the mRNA expression profiles of NSCLC cells with or without MCRS1 silencing, we found that MCRS1 regulated expressions of various genes related to cell proliferation, including Rb1, TP53, cell cycle-related genes, MYC, E2F2, PCNA, and Ki67. However, MCRS1 did not directly bind to these differentially expressed genes. Here, we confirmed that Rb1, an important tumor suppression gene (TSG), is a direct target of miR-155 which is directly up-regulated by MCRS1. Furthermore, the level of Rb1 expression in NSCLC tissues was inversely correlated with those of miR-155 and MCRS1, and MCRS1 regulated expression of Rb1 via miR-155. Additionally, we found that the DNA copy number of the MCRS1 gene played a role in MCRS1 overexpression in NSCLCs.ConclusionMCRS1 overexpression induced NSCLC proliferation through the miR-155–Rb1 pathway and DNA copy-number amplification is one of the mechanisms underlying MCRS1 overexpression in NSCLC. Moreover, we put forward the hypothesis that there are regulatory relationships between oncogenes and TSGs apart from the functional synergy of both; the oncogene-miRNA-TSG networks are one of mechanisms among the regulatory relationships; the regulatory relationships and the networks might play active roles in the development and progression of cancer.Electronic supplementary materialThe online version of this article (doi:10.1186/s13046-015-0235-5) contains supplementary material, which is available to authorized users.

MiR-124 Regulates the Epithelial-Restricted with Serine Box/Epidermal Growth Factor Receptor Signaling Axis in Head and Neck Squamous Cell Carcinoma.

Epithelial-restricted with serine box (ESX), a member of the ETS transcription factor family, is elevated and regulates EGFR in head and neck squamous cell carcinoma (HNSCC). However, the molecular mechanisms that contribute to ESX dysregulation remain to be elucidated. In this study, in silico analysis of the 3'-untranslated region (UTR) of ESX predicted two miR-124-binding sites. Delivery of miR-124 inhibited the 3'UTR ESX-driven reporter activity by 50% (P < 0.05) confirming ESX as a direct target of miR-124. Loss of miR-124 was found to be a frequent event in HNSCC. miR-124 expression was significantly depleted in the primary tumor compared with matched normal tissue in 100% (12/12) of HNSCC patients; relative mean miR-124 expression of 0.01197 and 0.00118 (P < 0.001, n = 12) in matched normal adjacent tissue and primary HNSCC tumor, respectively. Overexpression of miR-124 decreased ESX and EGFR levels in miR-124(low)/ESX(high)/EGFR(high) SCC15 HNSCC cells and reduced cell invasion, migration, proliferation, and colony formation. SCC15 cells with miR-124 restoration were less tumorigenic in vivo than miR-control SCC15 cells (70% inhibition, P < 0.01). Restoration of miR-124 in SCC15 cells enhanced the antiproliferative efficacy of the EGFR/Her2 tyrosine kinase inhibitors. Furthermore, recapitulation of EGFR in miR-124-overexpressing SCC15 cells was sufficient to completely block the antiproliferative effects of lapatinib and afatinib. Taken together, our work provides intriguing evidence that miR-124 is a novel therapeutic approach to reduce ESX/EGFR, and may be a tractable strategy to enhance the response rate of HNSCC patients to current anti-EGFR/Her2 therapies.

MiR-203 Acts as a Tumor Suppressor Gene in Osteosarcoma by Regulating RAB22A.

microRNAs (miRNAs), small noncoding RNAs of 19–25 nt, play an important roles in the pathological processes of tumorigenesis. The object of this study was to study the expression and function of miR-203 and to found its target gene in osteosarcoma. In our study, we found the expression level of miR-203 was significantly downregulated in osteosarcoma cell lines and tissues. In addition, overexpression of miR-203 inhibited the osteosarcoma cell proliferation and migration and inhibited Mesenchymal-to-Epithelial reversion Transition (MErT). Moreover, we identified RAB22A as a direct target of miR-203 and RAB22A overexpression blocks the roles of miR-203 in osteosarcoma cell. Furthermore, we demonstrated that RAB22A expression was upregulated in human osteosarcoma cell lines and tissues. Take together, our results demonstrated that miR-203 act as a tumor suppressor miRNA through regulating RAB22A expression and suggested its involvement in osteosarcoma progression and carcinogenesis.

MiR-128 downregulation promotes growth and metastasis of bladder cancer cells and involves VEGF-C upregulation

MicroRNA-128 (miR-128) serves an important role in regulating growth, invasiveness, stem cell-like traits, differentiation and apoptosis of different types of tumor cells. Vascular endothelial growth factor-C (VEGF-C) has been associated with angiogenesis, lymphangiogenesis and regional lymph node metastasis and has previously been reported to have an anti-apoptotic and proliferative role in bladder cancer (BC). To investigate the regulation of miR-128 on VEGF-C expression and their effects on proliferation and metastasis of bladder cancer, T24 and 5637 BC cells were transfected with pre-miR-128, anti-miR-128 and their respective negative control. miR-128 was downregulated in BC tissues and cell lines, while the expression levels of VEGF-C were upregulated. The present results indicated that miR-128 negatively regulated VEGF-C expression in BC T24 and 5637 BC cells. VEGF-C is a direct target of miR-128 in BC cells. Overexpression of miR-128 inhibited cell proliferation, migration and invasion. Knockdown of miR-128 promoted proliferation, migration and invasion in BC cells. Therefore, downregulation mediated malignant progression of BC may be partly attributed to increased VEGF-C expression. Consequently, the findings of the present study provide a molecular basis for the role of miR-128/VEGF-C in the progression of human BC and indicate a novel target for treatment of BC.

MicroRNA-128 suppresses cell growth and metastasis in colorectal carcinoma by targeting IRS1

Evidence has shown that microRNAs play important roles in tumor development, progression, and metastasis. miR-128 has been reported to be deregulated in different tumor types, whereas the function of miR-128 in colorectal carcinoma (CRC) largely remains to be elucidated. The aim of the present study was to investigate the clinical significance, biological effects and underlying mechanisms of miR-128 in CRC using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blotting. It was found that the expression of miR-128 was downregulated in CRC tissues and cell lines as determined by RT-qPCR. Furthermore, the expression of miR-128 in tumor tissues was significantly negatively correlated with TNM stage and lymph node metastasis in CRC patients. Functional assay revealed that the overexpression of miR-128 inhibited CRC cell proliferation, colony formation, migration and invasion and promoted apoptosis invitro, and suppressed CRC xenograft tumor growth invivo. In addition, insulin receptor substrate1 (IRS1), a key mediator in oncogenic insulin-like growth factor (IGF) signaling, was confirmed as a direct target of miR-128 by a luciferase reporter assay. Western blot analysis indicated that the overexpression of miR-128 significantly downregulated IRS1 expression and its downstream Akt signaling in CRCcells. Moreover, miR-128 was negatively associated with IRS1 in CRC tissues compared to adjacent non-tumor tissues. Taken together, these data suggested that miR-128 serves as a tumor suppressor and blocks CRC growth and metastasis by targeting IRS1.

MicroRNA-365 inhibits growth, invasion and metastasis of malignant melanoma by targeting NRP1 expression.

The role of miR-365 in cancer cells seemed controversial in previous studies. We thereby in this article aimed to define the role of miR-365 in malignant melanoma (MM) pathogenesis. We detected miR-365 expression in malignant melanoma cell lines and then investigated the effects of miR-365 on the metastasis and malignancy of melanoma cells. The correlation between miR-365 level and NRP1 (neuropilin1) was further investigated in clinical malignant melanoma specimens. MiR-365 was strongly down-regulated in malignant melanoma (MM) tissues and cell lines, and its expression levels were associated with lymph node metastasis and clinical stage, as well as overall survival and replase-free survival of MM. We also found that ectopic expression of miR-365 inhibited MM cell proliferation and MM metastasis in vitro and in vivo. We further identified a novel mechanism of miR-365 to suppress MM growth and metastasis. NRP1 was proved to be a direct target of miR-365, using luciferase assay and western blot. NRP1 over-expression in miR-365 expressing cells could rescue invasion and growth defects of miR-365. In addition, miR-365 expression inversely correlated with NRP1 protein levels in MM. Our data suggest that miR-365 functions as a tumor suppressor in MM development and progression, and holds promise as a prognostic biomarker and potential therapeutic target for MM.

Abstract LB-292: Epigenetic downregulation of ABCG2 in colorectal cancer by a novel microRNA-DNA methylation mechanism - an attractive target for cancer prevention

Abstract Colorectal cancer (CRC) is a major cause of mortality and morbidity worldwide. It develops in a complex multi-step process. The majority of studies to date have focused on genetic mutations and epigenetic changes that drive the CRC carcinogenesis process. Xenobiotic transporters play an important role in safeguarding our body from external toxic substances. These transporters lining the gastrointestinal tract may thus protect us from the dietary carcinogens. This study aims to investigate the downregulation of an efflux transporter ABCG2 in colorectal cancer versus normal colon mucosa, so as to shed some light on its relevance to CRC initiation and progression. We found that ABCG2 expression is at least 50-fold lower in adenomatous polyps and colon carcinoma specimens obtained from CRC patients than in their matched pair of adjacent normal colon mucosa. This may facilitate the accumulation of genotoxins to propel tumor initiation and progression. The underlying mechanism(s) for ABCG2 under-expression in CRC is currently not known. To this end, aberrant promoter methylation of ABCG2 has been reported to cause its repression in a few cancer types including renal carcinoma and multiple myeloma. On the other hand, ABCG2 promoter demethylation has been reported to upregulate the transporter to mediate multidrug resistance in lung cancer and acute lymphoblastic leukemia. In this study, miR-203 was found to be downregulated in all polyps and CRC specimens, relative to adjacent normal colon mucosa. We demonstrated that the de novo DNA methyltransferase DNMT3 is a direct target of miR-203. Importantly, by relieving the repression on DNMT3, the lower expression of miR-203 in CRC caused remarkable ABCG2 promoter methylation and lower ABCG2 expression in colon cancer cell lines and the patient CRC specimens. In spite of the complex sequential genetic mutations and epigenetic changes leading to CRC formation, we proposed that the restoration of the xenobiotic transporter ABCG2 function via modulating this new microRNA-DNA methylation mechanism in precancerous cells may represent an attractive strategy to stop or delay the carcinogenesis process. Citation Format: Kenneth K.W. To, Wing Wah Leung, Simon S.M. Ng. Epigenetic downregulation of ABCG2 in colorectal cancer by a novel microRNA-DNA methylation mechanism - an attractive target for cancer prevention. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-292. doi:10.1158/1538-7445.AM2015-LB-292

MicroRNA-145 inhibits migration and invasion via inhibition of fascin 1 protein expression in non-small-cell lung cancer cells.

MicroRNA (miR)-145 has been shown to act as a suppressor in numerous cancer types, including non-small-cell lung cancer (NSCLC). Fascin 1 (FSCN1), an actin bundling protein, has been implicated in NSCLC. However, the detailed role of miR-145 as well as the association between miR-145 and FSCN1 in the regulation of migration and invasion in NSCLC cells has remained elusive. The present study revealed that miR-145 was downregulated and FSCN1 was upregulated in NSCLC tissues and cell lines. Further investigation showed that overexpression of miR-145 markedly inhibited the protein expression of FSCN1, while knockdown of miR-145 upregulated the protein (but not mRNA) levels of FSCN1 in the NSCLC cell line H129. Moreover, a luciferase reporter assay indicated that FSCN1 is a direct target of miR-145 in NSCLC H129 cells. Furthermore, overexpression of miR-145 markedly inhibited the migration and invasion of NSCLC cells, similar to the effect of small interfering RNA-mediated FSCN1 inhibition in H129 cells. In addition, the inhibitory effect of miR-145 overexpression on migration and invasion was reversed by FSCN1 upregulation in H129 cells. These findings suggested that miR-145 has an inhibitory effect on the migration and invasion in NSCLC cells, at least in part through suppressing the protein expression of its target FSCN1. Therefore, miR-145/FSCN1 may be used as a potential target for the treatment of NSCLC.

Hypoxia-inducible MiR-182 promotes angiogenesis by targeting RASA1 in hepatocellular carcinoma.

BackgroundHypoxia is a common feature of solid tumors, including HCC. And hypoxia has been reported to play an important role in HCC progression. However, the potential mechanism of miRNAs in hypoxia mediating HCC progression still remains unclear.MethodsThe HCC cells were cultured in the atmosphere of 1 % oxygen to induce hypoxia. The microRNA microarray was employed to search for the hypoxia-inducible miRNAs. RT-PCR, western blot and immunohistochemistry were used to detect the RNA and protein levels. HUVEC were applied to explore the angiogenesis level.ResultsWe found that miR-182 was upregulated in the hypoxia-based microarray. We then revealed that miR-182 was also significantly increased in the HCC tissues compared to the corresponding normal tissues. In vitro capilliary tube formation assays showed that the miR-182 promoted angiogenesis. RASA1 was demonstrated as the direct target of miR-182. In addition, the suppression of RASA1 phenocopied the pro-angiogenesis effects of miR-182. Besides, RASA1 was also decreased in the hypoxia HCC cells while the inhibition of miR-182 partially restored the level of RASA1.ConclusionsOur data showed that hypoxia regulated the expression of miR-182 and RASA1 to promote HCC angiogenesis.Electronic supplementary materialThe online version of this article (doi:10.1186/s13046-015-0182-1) contains supplementary material, which is available to authorized users.

MiR-124 regulates fetal pulmonary epithelial cell maturation.

MicroRNAs are a family of small noncoding RNAs that regulate the expression of their target proteins at the posttranscriptional level. Their functions cover almost every aspect of cell physiology. However, the roles of microRNAs in fetal lung development are not completely understood. The objective of this study is to investigate the regulation and molecular mechanisms of alveolar epithelial cell maturation during fetal lung development by miR-124. We discovered that miR-124 was downregulated during rat fetal lung development and predominantly expressed in the epithelial cells at late stage of the lung development. Overexpression of miR-124 with an adenovirus vector led to the inhibition of epithelial maturation in rat fetal lung organ cultures and fetal alveolar epithelial type II cells, as demonstrated by a decrease in the type II cell marker expression and an increase in glycogen content. We further demonstrated by luciferase reporter assays that miR-124 inhibited the NF-κB, cAMP/PKA, and MAPK/ERK pathways. In addition, nuclear factor I/B (NFIB), a critical protein in fetal lung maturation, was validated as a direct target of miR-124. Furthermore, miR-124 expression was induced by the Wnt/β-catenin signaling pathway through a direct interaction of LEF1 and the miR-124 promoter region. We concluded that miR-124 downregulation is critical to fetal lung epithelial maturation and miR-124 inhibits this maturation process at least partially through the inhibition of NFIB.

Loss of Androgen-Regulated MicroRNA 1 Activates SRC and Promotes Prostate Cancer Bone Metastasis.

Bone metastasis is the hallmark of progressive and castration-resistant prostate cancers. MicroRNA 1 (miR-1) levels are decreased in clinical samples of primary prostate cancer and further reduced in metastases. SRC has been implicated as a critical factor in bone metastasis, and here we show that SRC is a direct target of miR-1. In prostate cancer patient samples, miR-1 levels are inversely correlated with SRC expression and a SRC-dependent gene signature. Ectopic miR-1 expression inhibited extracellular signal-regulated kinase (ERK) signaling and bone metastasis in a xenograft model. In contrast, SRC overexpression was sufficient to reconstitute bone metastasis and ERK signaling in cells expressing high levels of miR-1. Androgen receptor (AR) activity, defined by an AR output signature, is low in a portion of castration-resistant prostate cancer. We show that AR binds to the miR-1-2 regulatory region and regulates miR-1 transcription. Patients with low miR-1 levels displayed correlated low canonical AR gene signatures. Our data support the existence of an AR-miR-1-SRC regulatory network. We propose that loss of miR-1 is one mechanistic link between low canonical AR output and SRC-promoted metastatic phenotypes.

MicroRNA-155 aggravates ischemia-reperfusion injury by modulation of inflammatory cell recruitment and the respiratory oxidative burst.

The inflammatory sequelae of ischemia-reperfusion injury (IRI) are a major causal factor of tissue injury in various clinical settings. MicroRNAs (miRs) are short, non-coding RNAs, which regulate protein expression. Here, we investigated the role of miR-155 in IR-related tissue injury. Quantifying microRNA-expression levels in a human muscle tissue after IRI, we found miR-155 expression to be significantly increased and to correlate with the increased expression of TNF-α, IL-1β, CD105, and Caspase3 as well as with leukocyte infiltration. The direct miR-155 target gene SOCS-1 was downregulated. In a mouse model of myocardial infarction, temporary LAD ligation and reperfusion injury resulted in a smaller area of necrosis in miR-155-/- animals compared to wildtype animals. To investigate the underlying mechanisms, we evaluated the effect of miR-155 on inflammatory cell recruitment by intravital microscopy and on the generation of reactive oxygen species (ROS) of macrophages. Our intravital imaging results demonstrated a decreased recruitment of inflammatory cells in miR-155-/- animals during IRI. The generation of ROS in leukocytic cells of miR-155-/- animals was also reduced. RNA silencing of the direct miR-155 target gene SOCS-1 abrogated this effect. In conclusion, miR-155 aggravates the inflammatory response, leukocyte infiltration and tissue damage in IRI via modulation of SOCS-1-dependent generation of ROS. MiR-155 is thus a potential target for the treatment or prevention of IRI.

Tumor suppressor miR-145 reverses drug resistance by directly targeting DNA damage-related gene RAD18 in colorectal cancer.

Colorectal cancer (CRC) is one of the most common cancers worldwide. Although chemotherapy is used as a palliative treatment, ultimately, nearly all patients develop drug resistance. Therefore, the cell-inherent DNA repair pathway must reverse the DNA-damaging effect of cytotoxic drugs that mediates therapeutic resistance to chemotherapy. RAD18, a DNA damage-activated E3 ubiquitin ligase, is known to play a critical role in DNA damage repair in cancer cells. Here, we show that RAD18 is highly expressed in human 5-fluorouracil (5-FU)-resistant cancer cells after 5-FU treatment. In addition, RAD18 increases in CRC cells could induce DNA damage repair, suggesting that RAD18 might be a possible target for overcoming drug resistance. Moreover, the expression of tumor suppressor microRNA-145 (miR-145) was negatively correlated with RAD18 expression in CRC tissues of 140 patients. Using luciferase reporters carrying the 3'-untranslated region of RAD18 combined with Western blotting, we identified RAD18 as a direct target of miR-145. Also of interest, suppression of RAD18 by miR-145 enhanced DNA damage in CRC cells after 5-FU treatment. Finally, the 5-FU-resistant cancer cells could be selectively ablated by treatment with miR-145. Taken together, these results suggest that miR-145 can act as an RAD18 inhibitor and contribute as an important factor in reversing drug resistance after chemotherapy.

MiR-223 Regulates Adipogenic and Osteogenic Differentiation of Mesenchymal Stem Cells Through a C/EBPs/miR-223/FGFR2 Regulatory Feedback Loop.

Several miRNAs have recently been identified to regulate adipocyte or osteoblast differentiation or both. In this study, miR-223 was found to be involved in the reciprocal regulation of adipocyte and osteoblast differentiation. miR-223 was induced in primary cultured mouse marrow stromal cell, mesenchymal line C3H10T1/2 and stromal line ST2 after adipogenic treatment. Conversely, it was reduced in preosteoblast MC3T3-E1 after osteogenic treatment. Supplementing miR-223 levels using synthetic miR-223 mimics significantly suppressed the growth of the C3H10T1/2 and ST2 cells and induced the progenitor cells to fully differentiate into adipocytes, along with induction of adipocyte-specific transcription factors peroxisome proliferator-activated receptor γ, CCAAT/enhancer binding protein-α (C/EBPα), and marker genes aP2 and adipsin. By contrast, depletion of the endogenous miR-223 using synthetic miR-223 inhibitor repressed the progenitor cells to differentiate. The effects of miR-223 on adipocyte formation from ST2 cells were also demonstrated by using lentivirus that overexpresses miR-223. Conversely, supplementing miR-223 blocked ST2 to differentiate into osteoblasts. Fibroblast growth factor receptor 2 (Fgfr2), a critical regulator of osteoblast, was shown to be a direct target of miR-223 by using dual luciferase reporter assay. Knockdown of Fgfr2 in C3H10T1/2 downregulated phosphorylation of ERK1/2 and upregulated expression of C/EBPα and dramatically enhanced the differentiation of the cells into adipocytes. Further investigation of mechanisms that control miR-223 expression demonstrated that C/EBPs induced miR-223 expression through binding to the promoter regions of the miR-223. Taken together, our study provides evidences that miR-223 regulates adipocyte and osteoblast differentiation through a novel C/EBPs/miR-223/FGFR2 regulatory feedback loop.

Down-regulation of microRNA-223 promotes degranulation via the PI3K/Akt pathway by targeting IGF-1R in mast cells.

BackgroundMast cells play a central role in allergic and inflammatory disorders by inducing degranulation and inflammatory mediator release. Recent reports have shown that miRNAs play an important role in inflammatory response regulation. Therefore, the role of miR-223 in mast cells was investigated.MethodsThe expression of miR-223 was quantified by quantitative real-time polymerase chain reaction (qRT-PCR) in immunoglobulin E (IgE)-mediated mast cells. After successful miR-223 inhibition by transfection, degranulation was detected in IgE-mediated mast cells. The phosphorylation of IκB-α and Akt were examined using western blotting. NF-κB was tested using electrophoretic mobility shift assay. PI3K-inhibitor (LY294002) was used to investigate whether the PI3K/Akt pathway was essential for mast cell activation. The TargetScan database and a luciferase reporter system were used to identify whether insulin-like growth factor 1 receptor (IGF-1R) is a direct target of miR-223.ResultsMiR-223 expression was up-regulated in IgE-mediated mast cells, whereas its down-regulation promoted mast cell degranulation. Levels of IκB-α and Akt phosphorylation as well as NF-κB were increased in miR-223 inhibitor cells. LY294002 could block the PI3K/Akt signaling pathway and rescue the promotion caused by suppressing miR-223 in mast cells. IGF-1R was identified as a direct target of miR-223.ConclusionsThese findings suggest that down-regulation of miR-223 promotes degranulation via the PI3K/Akt pathway by targeting IGF-1R in mast cells.

MiR-203 inhibition of renal cancer cell proliferation, migration and invasion by targeting of FGF2.

BackgroundRenal cell carcinoma (RCC) is one of the leading causes of cancer related mortality worldwide. Increasing evidence has shown that microRNAs function as oncogenes or tumor suppressors in human malignancies, but the roles of miR-203 in human RCC is still unclear.MethodsFirst, quantitative real-time PCR (qRT-PCR) was performed to detect miR-203 expression in renal cancer cell lines and clear cell RCC (ccRCC) specimens. Then, the association of miR-203 expression with clinicopathological features and survival was later analyzed. Finally, the roles of miR-203 in regulation of tumor proliferation, migration, invasion, and target gene expression were further investigated.ResultsOur study showed miR-203 was down-regulated in renal cancer cell lines and ccRCC specimens (P < 0.05). Respectively, the low miR-203 expression in ccRCC specimens was associated with advanced clinical features and poorer prognosis (P < 0.05). miR-203 expression was an independent prognostic marker of overall ccRCC patient survival in a multivariate analysis (P < 0.05). Transient forced expression of miR-203 inhibited renal cancer cell growth and metastasis (P < 0.05). In contrast, down-regulation of miR-203 expression promoted renal cancer cell growth and metastasis (P < 0.05). Mechanistic investigations confirmed FGF2 as a direct target of miR-203, and up-regulation of miR-203 could decrease expression of FGF2. Further investigation showed that ectopic expression of FGF2 partially reversed the inhibition effect of enforced miR-203 expression on the malignant phenotypes of renal cancer cells.ConclusionsOur study suggested that miR-203 could be a potential prognostic marker and functions as a tumor suppressor in human renal cancer by post-transcriptionally targeting FGF2.Virtual slidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/6828145701534108.

MicroRNA-155 Exerts Cell-Specific Antiangiogenic but Proarteriogenic Effects During Adaptive Neovascularization

Adaptive neovascularization after arterial occlusion is an important compensatory mechanism in cardiovascular disease and includes both the remodeling of pre-existing vessels to collateral arteries (arteriogenesis) and angiogenic capillary growth. We now aimed to identify regulatory microRNAs involved in the modulation of neovascularization after femoral artery occlusion in mice. Using microRNA-transcriptome analysis, we identified miR-155 as a downregulated microRNA during hindlimb ischemia. Correspondingly, inhibition of miR-155 in endothelial cells had a stimulatory effect on proliferation and angiogenic tube formation via derepression of its direct target gene angiotensin II type 1 receptor. Surprisingly, miR-155-deficient mice showed an unexpected phenotype in vivo, with a strong reduction of blood flow recovery after femoral artery ligation (arteriogenesis) dependent on the attenuation of leukocyte-endothelial interaction and a reduction of proarteriogenic cytokine expression. Consistently, miR-155-deficient macrophages exhibit a specific alteration of the proarteriogenic cytokine expression profile, which is partly mediated by the direct miR-155 target gene SOCS-1. Our data demonstrate that miR-155 exerts an antiangiogenic but proarteriogenic function in the regulation of neovascularization via the suppression of divergent cell-specific target genes and that its expression in both endothelial and bone marrow-derived cells is essential for arteriogenesis in response to hindlimb ischemia in mice.

Overexpression of miR-155 in the Liver of Transgenic Mice Alters the Expression Profiling of Hepatic Genes Associated with Lipid Metabolism

Hepatic expression profiling has revealed miRNA changes in liver diseases, while hepatic miR-155 expression was increased in murine non-alcoholic fatty liver disease, suggesting that miR-155 might regulate the biological process of lipid metabolism. To illustrate the effects of miR-155 gain of function in transgenic mouse liver on lipid metabolism, transgenic mice (i.e., Rm155LG mice) for the conditional overexpression of mouse miR-155 transgene mediated by Cre/lox P system were firstly generated around the world in this study. Rm155LG mice were further crossed to Alb-Cre mice to realize the liver-specific overexpression of miR-155 transgene in Rm155LG/Alb-Cre double transgenic mice which showed the unaltered body weight, liver weight, epididymal fat pad weight and gross morphology and appearance of liver. Furthermore, liver-specific overexpression of miR-155 transgene resulted in significantly reduced levels of serum total cholesterol, triglycerides (TG) and high-density lipoprotein (HDL), as well as remarkably decreased contents of hepatic lipid, TG, HDL and free fatty acid in Rm155LG/Alb-Cre transgenic mice. More importantly, microarray data revealed a general downward trend in the expression profile of hepatic genes with functions typically associated with fatty acid, cholesterol and triglyceride metabolism, which is likely at least partially responsible for serum cholesterol and triglyceride lowering observed in Rm155LG/Alb-Cre mice. In this study, we demonstrated that hepatic overexpression of miR-155 alleviated nonalcoholic fatty liver induced by a high-fat diet. Additionally, carboxylesterase 3/triacylglycerol hydrolase (Ces3/TGH) was identified as a direct miR-155 target gene that is potentially responsible for the partial liver phenotypes observed in Rm155LG/Alb-Cre mice. Taken together, these data from miR-155 gain of function study suggest, for what we believe is the first time, the altered lipid metabolism and provide new insights into the metabolic state of the liver in Rm155LG/Alb-Cre mice.

Deletion of Mir155 Prevents Fas-Induced Liver Injury through Up-Regulation of Mcl-1

Fas-induced apoptosis is involved in diverse liver diseases. Herein, we investigated the effect of Mir155 deletion on Fas-induced liver injury. Wild-type (WT) mice and Mir155 knockout (KO) mice were i.p. administered with the anti-Fas antibody (Jo2) to determine animal survival and the extent of liver injury. After Jo2 injection, the Mir155 KO mice exhibited prolonged survival versus the WT mice (P < 0.01). The Mir155 KO mice showed lower alanine aminotransferase and aspartate aminotransferase levels, less liver tissue damage, fewer apoptotic hepatocytes, and lower liver tissue caspase 3/7, 8, and 9 activities compared with the WT mice, indicating that Mir155 deletion prevents Fas-induced hepatocyte apoptosis and liver injury. Hepatocytes isolated from Mir155 KO mice also showed resistance to Fas-induced apoptosis, in vitro. Higher protein level of myeloid cell leukemia-1 (Mcl-1) was also observed in Mir155 KO hepatocytes compared to WT hepatocytes. A miR-155 binding site was identified in the 3'-untranslated region of Mcl-1 mRNA; Mcl1 was identified as a direct target of miR-155 in hepatocytes. Consistently, pretreatment with a siRNA specific for Mcl1 reversed Mir155 deletion-mediated protection against Jo2-induced liver tissue damage. Finally, restoration of Mir155 expression in Mir155 KO mice abolished the protection against Fas-induced hepatocyte apoptosis. Taken together, these findings demonstrate that deletion of Mir155 prevents Fas-induced hepatocyte apoptosis and liver injury through the up-regulation of Mcl1.