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Argonaute 2 promotes myeloma angiogenesis via microRNA dysregulation.

ABSTRACT: BACKGROUND:Dysregulated microRNA (miRNA) expression contributes to cancer cell proliferation, apoptosis and angiogenesis. Angiogenesis is a hallmark of multiple myeloma (MM) development and progression. Argonaute 2 (AGO2) protein, a core component of the RNA-induced silencing complex (RISC), can directly bind to miRNAs and mediate target messenger RNA (mRNA) degradation. A previous study showed that AGO2 knockdown suppressed human umbilical vein endothelial cell (HUVEC) growth and tube formation. However, the roles and molecular mechanisms of AGO2-induced myeloma angiogenesis are not yet fully understood. The aim of this study was to characterize these roles and effects and their associated mechanisms. RESULTS:Supernatants from AGO2-overexpressing MM lines induced HUVEC migration and accelerated tube formation. Conversely, supernatants from AGO2-knockdown MM lines suppressed HUVEC cell migration and tube formation. Moreover, a chick chorioallantoic membrane (CAM) assay was used to demonstrate that AGO2 could drive neovessel formation in MM lines in vivo. Using an miRNA microarray, we observed that 25 miRNAs were upregulated and 7 were downregulated in response to AGO2. Most let-7 family members and 2 miR-17/92 cluster members (miR-17a and miR-92-1), all known pro-angiogenic miRNAs, were positively regulated by AGO2 whereas anti-angiogenic miRNAs such as miR-145 and miR-361 were negatively regulated by AGO2. CONCLUSIONS:We conclude that AGO2 can drive neovessel formation in vitro and in vivo by dysregulating the expression of some angiogenic miRNAs. The pro-angiogenic miRNAs of the let-7 family and the miR-17/92 cluster, along with the anti-angiogenic miRNA miR-145, play crucial roles in AGO2-mediated angiogenesis by targeting angiogenesis-related genes.

SUBMITTER: Wu S

PROVIDER: S-EPMC4108130 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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Argonaute 2 promotes myeloma angiogenesis via microRNA dysregulation.

Wu Shuang S Yu Wenjun W Qu Xiaoyan X Wang Rong R Xu Ji J Zhang Qiguo Q Xu Jiaren J Li Jianyong J Chen Lijuan L

Journal of hematology & oncology 20140507

<h4>Background</h4>Dysregulated microRNA (miRNA) expression contributes to cancer cell proliferation, apoptosis and angiogenesis. Angiogenesis is a hallmark of multiple myeloma (MM) development and progression. Argonaute 2 (AGO2) protein, a core component of the RNA-induced silencing complex (RISC), can directly bind to miRNAs and mediate target messenger RNA (mRNA) degradation. A previous study showed that AGO2 knockdown suppressed human umbilical vein endothelial cell (HUVEC) growth and tube f ...[more]

PMID: 24886719

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Project description:AimArgonaute2 (AGO2) protein is the active part of RNA-induced silencing complex, cleaving the target mRNA strand complementary to their bound siRNA. An increasing number of miRNAs has been identified as essential to angiogenesis of hepatocellular carcinoma (HCC). In this study we investigated how AGO2 affected HCC angiogenesis.MethodsHuman HCC cell lines HepG2, Hep3B, Huh7, SMMC-7721, Bel-7404, MHCC97-H and LM-3, and human umbilical vein endothelial cells (HUVEC) were tested. The expression of AGO2 in HCC cells was knocked down with siRNA and restored using recombinant adenovirus expressing Ago2. The levels of relevant mRNAs and proteins were examined using RT-PCR, Western blot and EILSA. Nude mice were implanted with Huh7 or SMMC-7721 cells, and tumor volumes were measured. After the mice were euthanized, the xenograft tumors were used for immunohistological analysis.ResultsIn 6 HCC cell lines, AGO2 protein expression was significantly correlated with VEGF expression (r=+0.79), and with VEGF secretion (r=+0.852). Knockdown of Ago2 in Huh7 cells and SMMC-7721 cells substantially decreased VEGF expression, whereas the restoration of AGO2 reversed both VEGF expression and secretion. Furthermore, knockdown of Ago2 significantly up-regulated the expression of PTEN (a tumor suppressor involved in the inhibition of HCC angiogenesis), and vice versa. Moreover, the specific PTEN inhibitor bisperoxovanadate (7, 14, 28 nmol/L) dose-dependently restored the expression of VEGF and the capacity of HCC cells to induce HUVECs to form capillary tubule structures. In the xenograft nude mice, knockdown of Ago2 markedly suppressed the tumor growth and decreased PTEN expression and CD31-positive microvascular in the xenograft tumors.ConclusionA direct relationship exists between the miRNA processing machinery AGO2 and HCC angiogenesis that is mediated by the AGO2/PTEN/VEGF signaling pathway. The results suggest the high value of Ago2 knockdown in anti-angiogenesis therapy for HCC.

Project description:BackgroundMicroRNAs (miRs) are small non-coding RNAs that play an important role in cancer development where they can act as oncogenes or as tumor-suppressors. miR-34a is a tumor-suppressor that is frequently downregulated in a number of tumor types. However, little is known about the role of miR-34a in head and neck squamous cell carcinoma (HNSCC).Methods and resultsmiR-34a expression in tumor samples, HNSCC cell lines and endothelial cells was examined by real time PCR. Lipofectamine-2000 was used to transfect miR-34a in HNSCC cell lines and human endothelial cells. Cell-proliferation, migration and clonogenic survival was examined by MTT, Xcelligence system, scratch assay and colony formation assay. miR-34a effect on tumor growth and tumor angiogenesis was examined by in vivo SCID mouse xenograft model. Our results demonstrate that miR-34a is significantly downregulated in HNSCC tumors and cell lines. Ectopic expression of miR-34a in HNSCC cell lines significantly inhibited tumor cell proliferation, colony formation and migration. miR-34a overexpression also markedly downregulated E2F3 and survivin levels. Rescue experiments using microRNA resistant E2F3 isoforms suggest that miR-34a-mediated inhibition of cell proliferation and colony formation is predominantly mediated by E2F3a isoform. In addition, tumor samples from HNSCC patients showed an inverse relationship between miR-34a and survivin as well as miR-34a and E2F3 levels. Overexpression of E2F3a completely rescued survivin expression in miR-34a expressing cells, thereby suggesting that miR-34a may be regulating survivin expression via E2F3a. Ectopic expression of miR-34a also significantly inhibited tumor growth and tumor angiogenesis in a SCID mouse xenograft model. Interestingly, miR-34a inhibited tumor angiogenesis by blocking VEGF production by tumor cells as well as directly inhibiting endothelial cell functions.ConclusionsTaken together, these findings suggest that dysregulation of miR-34a expression is common in HNSCC and modulation of miR34a activity might represent a novel therapeutic strategy for the treatment of HNSCC.

Project description:BackgroundGlioma is the most frequent and lethal primary brain malignancy. Amounting evidence has highlighted the importance of exosomal microRNAs (miRNAs or miRs) in this malignancy. This study aimed to investigate the regulatory role of exosomal miR-148a-3p in glioma.MethodsBioinformatics analysis was firstly used to predict the target genes of miR-148a-3p. Exosomes were then extracted from normal human astrocytes and glioma cells. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was applied to determine the expression patterns of miR-148a-3p and ERBB receptor feedback inhibitor 1 (ERRFI1). Dual-luciferase reporter gene assay was applied to verify the direct binding between miR-148a-3p and ERRFI1. Cell counting kit-8 and tube formation assays were further conducted to assess the proliferation and angiogenic properties of human umbilical vein endothelial cells (HUVECs) in the co-culture system with exosomes. Lastly, glioma tumor models were established in BALB/c nude mice to study the role of exosomal miR-148a-3p in vivo.ResultsmiR-148a-3p was highly expressed, while ERRFI1 was poorly expressed in glioma. miR-148a-3p was found to be enriched in glioma cells-derived exosomes and could be transferred to HUVECs via exosomes to promote their proliferation and angiogenesis. ERRFI1 was identified as a target gene of miR-148a-3p. In addition, miR-148a-3p activated the epidermal growth factor receptor (EGFR)/mitogen-activated protein kinase (MAPK) signaling pathway by inhibiting ERRFI1. In the co-culture system, our data demonstrated that glioma cells-derived exosomal miR-148a-3p down-regulated ERRFI1 and activated the EGFR/MAPK signaling pathway, so as to promote cell proliferation and angiogenesis. In vivo experimentation further demonstrated that this mechanism was responsible for the promotive role of exosomal miR-148a-3p in tumorigenesis and angiogenesis.ConclusionTaken together, glioma-derived exosomal miR-148a-3p promoted tumor angiogenesis through activation of the EGFR/MAPK signaling pathway by ERRFI1 inhibition.

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Argonaute 2 promotes myeloma angiogenesis via microRNA dysregulation.

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Argonaute 2 promotes myeloma angiogenesis via microRNA dysregulation.

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