406991

MIR21

The small regulatory RNA microRNA-21 (miR-21) plays a crucial role in a plethora of biological functions and diseases including development, cancer, cardiovascular diseases and inflammation. The gene coding for pri-miR-21 (primary transcript containing miR-21) is located within the intronic region of the TMEM49 gene. Despite pri-miR-21 and TMEM49 are overlapping genes in the same direction of transcription, pri-miR-21 is independently transcribed by its own promoter regions and terminated with its own poly(A) tail. After transcription, primiR- 21 is finally processed into mature miR-21. expression of miR-21 has been found to be deregulated in almost all types of cancers and therefore was classified as an oncomiR. During recent years, additional roles of miR-21 in cardiovascular and pulmonary diseases, including cardiac and pulmonary fibrosis as well as myocardial infarction have been described. MiR-21 additionally regulates various immunological and developmental processes. Due to the critical functions of its target proteins in various signaling pathways, miR-21 has become an attractive target for genetic and pharmacological modulation in various disease conditions.

The transforming growth factor-beta (TGF-beta) signaling pathway plays important roles in maintaining normal tissue homeostasis, and is tightly controlled by a network of biomolecules. MicroRNAs (miRNAs) are small noncoding RNAs of approximately 22 nucleotides that regulate gene expression at posttranscriptional levels. Increasing evidence points to the important role of miRNAs in TGF-beta signaling. OncomicroRNA miR-21 has been established as a key regulator of mesenchymal phenotype transition induced by TGF-beta. However, the effects of miR-21 on epithelial biology involved in TGF-beta signaling pathway such as cytostatic program and epithelial to mesenchymal transition (EMT) processes are unclear. Here we show that miR-21 is upregulated after TGF-beta exposure in both growth inhibition and EMT models of HaCaT keratinocytes. To determine the potential roles of miR-21 in TGF-beta-induced growth-arrest and EMT models, we showed that ectopic expression of miR-21 overcame TGF-beta' growth-inhibitory effect and the knockdown of miR-21 potentialized this effect, but perturbation of miR-21 levels had little effect on EMT. Moreover, TGFBR2, PTEN, PDCD4, and TAp63 were identified as targets of miR-21 in HaCaT cells. And among them, TGFBR2, PTEN, and TAp63 were associated with TGF-beta-induced cytostatic program. Thus, our results suggest that miR-21 regulates the ability of epithelial cells to respond to TGF-beta, with potential impact on epithelium homeostasis, wound-healing and tumorigenesis.CI - Copyright (c) 2011 Elsevier Ltd. All rights reserved.

MiR-21 is known to play an important role in the development and progression, including migration and invasion, of many malignancies including breast cancer. Accumulating evidence suggest that the induction of epithelial-mesenchymal transition (EMT) phenotype and acquisition of cancer stem cell (CSC) characteristics are highly interrelated, and contribute to tumorigenesis, tumor progression, metastasis, and relapse. The molecular mechanisms underlying EMT and CSC characteristics during miR-21 contributes to cell migration and invasion of breast cancer are poorly understood. Therefore, we established miR-21 re-expressing breast cancer MCF-7 (MCF-7/miR-21) cells, which showed increasing cell growth, migration and invasion, self-renewal and clonogenicity. Our data showed that re-expression of miR-21 induced the acquisition of EMT phenotype by activation of mesenchymal cell markers (N-cadherin, Vimentin, alpha-SMA) and inhibition of epithelial cell marker (E-cadherin) in MCF-7/miR-21 cells, which consistent with increased cell subpopulation expressing CSC surface markers (ALDH1(+) and CD44(+)/CD24(-/low)) and the capacity of sphereforming (mammospheres). Our results demonstrated that re-expression of miR-21 is responsible for migration and invasion by activating the EMT process and enhancing the characteristics of CSCs in MCF-7 cells.

cancer stem cells (CSCs) are predicted to be critical drivers of tumor progression due to their "stemness", but the molecular mechanism of CSCs in regulating metastasis remains to be elucidated. epithelial-mesenchymal transition (EMT), hypoxia-inducible factor (HIF)-1alpha, and miR-21, all of which contribute to cell migration for metastasis, are interrelated with CSCs. In the present study, third-sphere forming (3-S) CSC-like cells, which showed elevated CSC surface markers (ALDH1(+) and CD44(+)/CD24(-/low)) and sphereforming capacity as well as migration and invasion capacities, were cultured and isolated from breast cancer MCF-7 parental cells, to evaluate the role of miR-21 in regulating the CSC-like cell biological features, especially EMT. EMT, which was assessed by overexpression of mesenchymal cell markers (N-cadherin, Vimentin, alpha-smooth muscle actin [alpha-SMA]) and suppression of epithelial cell marker (E-cadherin), was induced in 3-S CSC-like cells. Moreover, both of HIF-1alpha and miR-21 were upregulated in the CSC-like cells. Interestingly, antagonism of miR-21 by antagomir led to reversal of EMT, downexpression of HIF-1alpha, as well as suppression of invasion and migration, which indicates a key role of miR-21 involved in regulate CSC-associated features. In conclusion, we demonstrated that the formation of CSC-like cells undergoing process of EMT-like associated with overexpression of HIF-1alpha, both of which are regulated by miR-21.CI - (c) 2012 Japanese cancer Association.

The flavolignan silibinin was studied for its ability to restore drug sensitivity to EGFR-mutant NSCLC xenografts with epithelial-to-mesenchymal transition (EMT)-driven resistance to erlotinib. As a single agent, silibinin significantly decreased the tumor volumes of erlotinib-refractory NSCLC xenografts by approximately 50%. Furthermore, the complete abrogation of tumor growth was observed with the co-treatment of erlotinib and silibinin. Silibinin fully reversed the EMT-related high miR-21/low miR-200c microRNA signature and repressed the mesenchymal markers SNAIL, ZEB, and N-cadherin observed in erlotinib-refractory tumors. Silibinin was sufficient to fully activate a reciprocal mesenchymal-to-epithelial transition (MET) in erlotinib-refractory cells and prevent the highly migratogenic phenotype of erlotinib-resistant NSCLC cells. Given that the various mechanisms of resistance to erlotinib result from EMT, regardless of the EGFR mutation status, a water-soluble, silibinin-rich milk thistle extract might be a suitable candidate therapy for upcoming clinical trials aimed at preventing or reversing NSCLC progression following erlotinib treatment.

BACKGROUND: The excess and persistent accumulation of fibroblasts due to aberrant tissue repair results in fibrotic diseases such as idiopathic pulmonary fibrosis. Recent reports have revealed significant changes in microRNAs during idiopathic pulmonary fibrosis and evidence in support of a role for microRNAs in myofibroblast differentiation and the epithelial-mesenchymal transition in the context of fibrosis. It has been reported that microRNA-21 is up-regulated in myofibroblasts during fibrosis and promotes transforming growth factor-beta signaling by inhibiting Smad7. However, expression changes in microRNA-21 and the role of microRNA-21 in epithelial-mesenchymal transition during lung fibrosis have not yet been defined. METHODS: Lungs from saline- or bleomycin-treated C57BL/6 J mice and lung specimens from patients with idiopathic pulmonary fibrosis were analyzed. Enzymatic digestions were performed to isolate single lung cells. Lung epithelial cells were isolated by flow cytometric cell sorting. The expression of microRNA-21 was analyzed using both quantitative PCR and in situ hybridization. To induce epithelial-mesenchymal transition in culture, isolated mouse lung alveolar type II cells were cultured on fibronectin-coated chamber slides in the presence of transforming growth factor-beta, thus generating conditions that enhance epithelial-mesenchymal transition. To investigate the role of microRNA-21 in epithelial-mesenchymal transition, we transfected cells with a microRNA-21 inhibitor. Total RNA was isolated from the freshly isolated and cultured cells. MicroRNA-21, as well as mRNAs of genes that are markers of alveolar epithelial or mesenchymal cell differentiation, were quantified using quantitative PCR. RESULTS: The lung epithelial cells isolated from the bleomycin-induced lung fibrosis model system had decreased expression of epithelial marker genes, whereas the expression of mesenchymal marker genes was increased. MicroRNA-21 was significantly upregulated in isolated lung epithelial cells during bleomycin-induced lung fibrosis and human idiopathic pulmonary fibrosis. MicroRNA-21 was also upregulated in the cultured alveolar epithelial cells under the conditions that enhance epithelial-mesenchymal transition. Exogenous administration of a microRNA-21 inhibitor prevented the increased expression of vimentin and alpha-smooth muscle actin in cultured primary mouse alveolar type II cells under culture conditions that induce epithelial-mesenchymal transition. CONCLUSIONS: Our experiments demonstrate that microRNA-21 is increased in lung epithelial cells during lung fibrosis and that it promotes epithelial-mesenchymal transition.

BACKGROUND: Late kidney allograft dysfunction is becoming a significant problem and tubular atrophy and interstitial fibrosis are main causes. It was reported that hypoxia could induce epithelial-mesenchymal transition (EMT) in renal tubular epithelial cells (TECs), and hypoxia-inducible factor-1 (HIF-1) is one of the important regulators of cellular adaptive to hypoxia. METHODS: In this study, we used an HIF-1alphaDeltaODD-expressing adenovirus, which could stably and functionally express HIF-1alpha under normoxia conditions, and used a hypoxia/reoxygenation cell model to simulate ischemia/reperfusion (I/R) injury in vitro, to investigate the effect of HIF-1alpha on EMT-related gene expressions. RESULTS: Our results demonstrated that HIF-1alpha could significantly upregulate alpha-smooth muscle actin expression, and reduced the E-cadherin expression in HK-2 cells during I/R injury. Moreover, miR-21 expression had a positive correlation with HIF-1alpha in this process. CONCLUSION: These results suggest that HIF-1alpha may promote the EMT development through regulating fibrotic gene expression during I/R injury in human renal TECs, and miR-21 could be among the important regulatory pathways in the process.CI - Copyright (c) 2014 Elsevier Inc. All rights reserved.

A consistent positive association between cigarette smoking and the human esophageal cancer has been confirmed all over the world. However, details in the association need to be more focused on and be identified. Recently, aberrantly expressed microRNAs (miRNAs) have been shown to be promising biomarkers for understanding the tumorigenesis of a wide array of human cancers, including the esophageal cancer, and the deregulation on the epithelial to mesenchymal transition (EMT) by miRNAs is involved in the tumorigenesis. In present study, we were going to identify the role of nicotine-induced miR-21 in the EMT of esophageal cells. We found that there was an overexpression of miR-21 in esophageal specimens, having an association with cigarette smoking, and the upregulation of miR-21 was also induced by nicotine in esophageal carcinoma cell line, EC9706. Moreover, the upregulated miR-21 by nicotine promoted EMT transforming growth factor beta (TGF-beta) dependently. Thus, the present study reveals a novel oncogenic role of nicotine in human esophageal cancer.

epithelial-to-mesenchymal transition (EMT) plays an important role in renal interstitial fibrosis (RIF) with diabetic nephropathy (DN). Smad7 (a inhibitory smad), a downstream signaling molecules of TGF-beta1, represses the EMT. The physiological function of miR-21 is closely linked to EMT and RIF. However, it remained unclear whether miR-21 over-expression affected TGF-beta1-induced EMT by regulating smad7 in DN. In this study, real-time RT-PCR, cell transfection, luciferase reporter gene assays, western blot and confocal microscope were used, respectively. Here, we found that miR-21 expression was upregulated by TGF-beta1 in time- and concentration -dependent manner. Moreover, miR-21 over-expression enhanced TGF-beta1-induced EMT(upregulation of a-SMA and downregulation of E-cadherin) by directly down-regulating smad7/p-smad7 and indirectly up-regulating smad3/p-smad3, accompanied by the decrease of Ccr and the increase of col-IV, FN, the content of collagen fibers, RTBM, RTIAW and ACR. Meantime, the siRNA experiment showed that smad7 can directly regulate a-SMA and E-cadherin expression. More importantly, miR-21 inhibitor can not only inhibit EMT and fibrosis but also ameliorate renal structure and function. In conclusion, our results demonstrated that miR-21 overexpression can contribute to TGF-beta1-induced EMT by inhibiting target smad7, and that targeting miR-21 may be a better alternative to directly suppress TGF-beta1-mediated fibrosis in DN.CI - Copyright (c) 2014 Elsevier Ireland Ltd. All rights reserved.

BACKGROUND: MicroRNA-21 (miR-21) plays an important role in the pathogenesis and progression of liver fibrosis. Here, we determined the serum and hepatic content of miR-21 in patients with liver cirrhosis and rats with dimethylnitrosamine-induced hepatic cirrhosis and examined the effects of miR-21 on SPRY2 and HNF4alpha in modulating ERK1 signaling in hepatic stellate cells (HSCs) and epithelial-mesenchymal transition (EMT) of hepatocytes. METHODS: Quantitative RT-PCR was used to determine miR-21 and the expression of SPRY2, HNF4alpha and other genes. Immunoblotting assay was carried out to examine the expression of relevant proteins. Luciferase reporter assay was performed to assess the effects of miR-21 on its predicted target genes SPRY2 and HNF4alpha. Primary HSCs and hepatocytes were treated with miR-21 mimics/inhibitors or appropriate adenoviral vectors to examine the relation between miR-21 and SPRY2 or HNF4alpha. RESULTS: The serum and hepatic content of miR-21 was significantly higher in cirrhotic patients and rats. SPRY2 and HNF4alpha mRNA levels were markedly lower in the cirrhotic liver. MiR-21 overexpression was associated with enhanced ERK1 signaling and EMT in liver fibrosis. Luciferase assay revealed suppressed SPRY2 and HNF4alpha expression by miR-21. Ectopic miR-21 stimulated ERK1 signaling in HSCs and induced hepatocyte EMT by targeting SPRY2 or HNF4alpha. Downregulating miR-21 suppressed ERK1 signaling, inhibited HSC activation, and blocked EMT in TGFbeta1-treated hepatocytes. CONCLUSIONS: MiR-21 modulates ERK1 signaling and EMT in liver fibrosis by regulating SPRY2 and HNF4alpha expression. MiR-21 may serve as a potentially biomarker as well as intervention target for hepatic cirrhosis.

Arsenic is well established as a human carcinogen, but the molecular mechanisms leading to arsenic-induced carcinogenesis are complex and elusive. It is not been determined if the epithelial-mesenchymal transition (EMT) contributes to carcinogen-induced malignant transformation and subsequent tumor formation. We have found that, during the neoplastic transformation induced in human bronchial epithelial (HBE) cells by a low concentration (1.0muM) of arsenite, the cells undergo an EMT and show enhanced invasion and migration. With longer times for transformation of HBE cells, there was increased miR-21 expression. Further, during the transformation of HBE cells, inhibition of miR-21 with an miR-21 inhibitor increased levels of PDCD4, an inhibitor of neoplastic transformation; reduced Twist1, a transcription factor involved in cell differentiation; and inhibited cell invasion and migration. In addition, PDCD4 interacted with Twist1 and inhibited its expression function, which is involved in arsenite-induced EMT. Thus, miR-21, acting on PDCD4, which interacts with Twist1 and represses the expression of Twist1, contributes to the EMT induced by arsenite. These observations add to an understanding of the processes involved in arsenite-induced carcinogenesis.CI - Copyright (c) 2014 Elsevier Ireland Ltd. All rights reserved.

miR-21 has been shown to play fundamental role in diverse biological and pathological processes, including fibrotic diseases. In the present study, we investigated whether miR-21 regulated the fibrogenic epithelial-mesenchymal transition (EMT) in human hepatocytes QSG-7701 and explored underlying mechanisms. The results showed that treatment of QSG-7701 cells with pro-fibrogenic factor TGF-beta1 resulted in increased expression of miR-21 and promoted fibrogenic EMT in hepatocytes. Downregulation of miR-21 expression by transfection of anti-miR-21 into QSG-7701 cells inhibited fibrogenic EMT induced by TGF-beta1. Furthermore, overexpression of miR-21 alone also resulted in EMT-like transformation in QSG-7701 cells. TGF-beta1 treatment resulted in decreased PTEN and increased Akt phosphorylation and anti-miR-21 abolished this effect. Overexpression of miR-21 in QSG-7701 cells also downregulated PTEN and upregulated Akt phosphralation. Inhibition of Akt signaling by specific inhibitor Akt inhibitor IV blocked TGF-beta1 and miR-21-induced fibrogenic EMT. In summary, our results identify miR-21 as a key regulator of fibrogenic EMT in hepatocytes via PTEN/Akt pathway. Targeting miR-21 may provide a new therapeutic strategy against hepatic fibrosis.CI - Copyright (c) 2014 Elsevier Masson SAS. All rights reserved.

Inflammation and microRNAs are involved in human skin cancer; however, their molecular mechanisms remain unclear. Further, a concern in skin cancer research is the identification of biomarkers for early diagnosis and accurate prognosis. To explore new biomarkers of chemical exposure in risk assessment of chemical carcinogenesis and arsenite-induced skin cancer, we investigated the roles of interleukin-6 (IL-6) regulation of microRNA-21 (miR-21), functioning via activation of signal transducers and activators of transcription 3 (STAT3), in neoplastic and metastatic properties of immortalized human keratinocytes (HaCaT cells) transformed by arsenite. In HaCaT cells, arsenite caused increases of IL-6 and miR-21 levels and activation of STAT3, which induced the epithelial-mesenchymal transition (EMT). Blocking IL-6 with anti-IL-6 antibody inhibited the activation of STAT3 and increases of miR-21 levels. Knock-down of STAT3 by siRNA blocked the increases of miR-21. In arsenite-transformed HaCaT (HaCaT-30T) cells, down-regulation of STAT3 by siRNA blocked the process of EMT and decreased their neoplastic properties and migratory capacity, effects that were antagonized by over-expression of miR-21.Thus, the IL-6/STAT3 pathway via miR-21 is involved in EMT, neoplastic properties, and migratory capacity of arsenite-transformed HaCaT cells. The results may lead to development of biomarkers for early diagnosis and accurate prognosis of arsenite-induced skin cancer.CI - Copyright (c) 2015 Elsevier Ireland Ltd. All rights reserved.

MicroRNAs (miRNAs) have recently been demonstrated to play a crucial role in malignant progression including differentiation, proliferation, metastasis and invasion, MicroRNA-21 (mir-21) also has been reported to have association with tumor invasion and metastasis in some tumors including cholangiocarcinoma (CCA). In this study, we further investigated the association of mir-21 with CCA biological behavior by transfecting miR-21 mimics or mir-21 inhibitor into QBC939 and RBE cells accompanied with the tumor xenografts experiment. Results indicated that over-expression of miR-21 significantly promoted cell migration, invasion and xenografts growth, whereas contrary phenomenon was observed in mir-21 inhibitor group. Furthermore, we explored the expression of EMT related proteins in CCA cells and tumor xenografts. Results showed that E-cadherin was decreased and N-cadherin, Vimentin were up-regulated significantly when miR-21 was over-expressed. In conclusion, microRNA-21 is crucial for CCA carcinogenesis and metastasis, which could induce EMT process, thereby promote the invasion and migration of CCA cells. These findings may provide new strategy for prevention and treatment of CCA in the future.

Patients with primary HER2-positive breast cancer benefit from HER2-targeted therapies. Nevertheless, a significant proportion of these patients die of disease progression due to mechanisms of drug resistance. MicroRNAs (miRNAs) are emerging as critical core regulators of drug resistance that act by modulating the epithelial-to-mesenchymal transition (EMT) and cancer-related immune responses. In this study, we investigated the association between the expression of a specific subset of 14 miRNAs involved in EMT processes and immune functions and the response to neoadjuvant trastuzumab and chemotherapy in 52 patients with HER2-overexpressing breast tumors. The expression of only a single miRNA, miR-21, was significantly associated with residual disease (p = 0.030) and increased after trastuzumab-chemotherapy (p = 0.012). A target prediction analysis coupled with in vitro and in vivo validations revealed that miR-21 levels inversely correlated with the expression of PTEN (rs = -0.502; p = 0.005) and PDCD4 (rs = -0.426; p = 0.019), which differentially influenced the drug sensitivity of HER2-positive breast cancer cells. However, PTEN expression was only marginally associated with residual disease. We further demonstrated that miR-21 was able to affect the response to both trastuzumab and chemotherapy, triggering an IL-6/STAT3/NF-kappaB-mediated signaling loop and activating the PI3K pathway. Our findings support the ability of miR-21 signaling to sustain EMT and shape the tumor immune microenvironment in HER2-positive breast cancer. Collectively, these data provide a rationale for using miR-21 expression as a biomarker to select trastuzumab-chemotherapy-resistant HER2-positive breast cancer patients who may benefit from treatments containing PI3K inhibitors or immunomodulatory drugs.

Acquisition of gemcitabine resistance in breast cancer has not been fully clarified. Prior studies suggest that miRNAs are important to chemoresistance in solid tumors and we confirmed that miR-21 is involved in the development of gemcitabine resistance. epithelial-to-mesenchymal transition (EMT) and AKT pathway activation were noted to be important to this resistance as well. PTEN, a direct target gene of miR-21, was significantly downregulated in gemcitabine-resistant breast cancer cells and restoration of PTEN expression blocked miR-21-induced EMT and gemcitabine resistance. Our data offer novel insight into gemcitabine resistance in breast cancer and suggest that miR-21 may be used to predict optimal breast cancer therapy and may be a potential therapeutic target for reversing gemcitabine resistance.

Glyoxalase I (Glo1) is the main scavenging enzyme of methylglyoxal (MG), a potent precursor of advanced glycation end products (AGEs). AGEs are known to control multiple biological processes, including epithelial to mesenchymal transition (EMT), a multistep phenomenon associated with cell transformation, playing a major role in a variety of diseases, including cancer. Crystalline silica is a well-known occupational health hazard, responsible for a great number of human pulmonary diseases, such as silicosis. There is still much debate concerning the carcinogenic role of crystalline silica, mainly due to the lack of a causal demonstration between silica exposure and carcinogenesis. It has been suggested that EMT might play a role in crystalline silica-induced lung neoplastic transformation. The aim of this study was to investigate whether, and by means of which mechanism, the antiglycation defence Glo1 is involved in Min-U-Sil 5 (MS5) crystalline silica-induced EMT in BEAS-2B human bronchial epithelial cells chronically exposed, and whether this is associated with the beginning of a neoplastic-like transformation process. By using gene silencing/overexpression and scavenging/inhibitory agents, we demonstrated that MS5 induced hydrogen peroxide-mediated c-Jun-dependent Glo1 up-regulation which resulted in a decrease in the Argpyrimidine-modified Hsp70 protein level which triggered EMT in a novel mechanism involving miR-21 and SMAD signalling. The observed EMT was associated with a neoplastic-like phenotype. The results obtained provide a causal in vitro demonstration of the MS5 pro-carcinogenic transforming role and more importantly they provide new insights into the mechanisms involved in this process, thus opening new paths in research concerning the in vivo study of the carcinogenic potential of crystalline silica.CI - Copyright (c) 2016 Elsevier Inc. All rights reserved.

OBJECTIVE: The aims of this study were to discover if increased circulating microRNA-21 (miR-21) expression in serum is associated with lymph node metastasis in patients with cervical cancer and look further into the molecular mechanism of these. Whole-blood samples from 89 patients who have been histopathologically confirmed as having cervical cancer and 20 control subjects were collected, and then the association between lymph node metastasis and the level of circulating miR-21 was compared. Then cervical cancer cell lines HeLa (HPV-18 DNA, E6/E7RNA) and HT-3 (HPV DNA, E6/E7RNA) were used to confirm the interaction between miR-21 and RASA1. The role of RASA1 in cervical cancer cell migration was also studied in HeLa. Increased circulating miR-21 expression in serum is associated with lymph node metastasis in patients with cervical cancer. MicroRNA-21 reduces RASA1 expression in cervical cancer cell lines and promotes cervical cancer cell migration via RASA1. Furthermore, Ras-induced epithelial-mesenchymal transition contributes to miR-21/RASA1 axis promoting cervical cancer cell migration. Circulating miR-21 in serum could be a promising biomarker in auxiliary diagnosis of lymph node metastasis in cervical cancer, and inhibition of miR-21/RASA1 axis could be a possible strategy to restrain migration of cervical cancer.

MicroRNA-21 (miR-21) has emerged as a critical regulatory molecule and an important serum marker in hepatic fibrogenesis. The aim of the present study was to investigate the role of inhibiting miR-21 on hepatic fibrosis treatment. Serum miR-21 levels in 60 healthy individuals and 180 patients with different stages of liver cirrhosis were examined, miR-21 levels in normal or cirrhotic human liver tissues (n=10 each) were also detected. An adenoviral vector (Ad-TuD-21) carrying the sponging ToughDecoy (TuD)-RNA sequence against miR-21 was constructed to reduce miR-21 expression efficiently in vitro and in vivo Histological and immunohistological examinations were performed to evaluate the inhibitory effects and mechanism of Ad-TuD-21 delivery into carbon tetrachloride (CCl4) induced hepatic fibrosis rats by targeting extracellular signal-regulated kinase 1 (ERK1) signalling in hepatic stellate cells (HSC) and hepatocyte epithelial-mesenchymal transition (EMT). Our results revealed that enhanced miR-21 levels in cirrhotic patients were related to the severity and activity of liver cirrhosis. Ad-TuD-21 administered to liver fibrosis rats could remarkably suppress profibrotic gene expression, cause histological improvements in liver and attenuate hepatic fibrosis significantly. More importantly, after Ad-TuD-21 treatment, inhibition of both the ERK1 signalling pathway in HSC and hepatocyte EMT was confirmed, which paralleled the enhancement of miR-21 target genes-sprouty2 (SPRY2) and hepatocyte nuclear factor 4alpha (HNF4alpha)-expression in vivo These data demonstrated that miR-21 is a key regulator to promote hepatic fibrogenesis, and sponging miR-21 expression may present a novel potentially therapeutic option for hepatic fibrosis.CI - (c) 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

Keloid is the abnormal wound healing puzzled by the aggressive growth and high recurrence rate due to its unrevealed key pathogenic mechanism. MicroRNAs contribute to a series of biological processes including epithelial-mesenchymal transition (EMT) and cells stemness involved in fibrotic disease. Here, using microRNAs microarray analysis we found mir-21-5p was significantly up-regulated in keloid epidermis. To investigate the role of miR-21-5p in keloid pathogenesis, we transfected miR-21-5p mimic or inhibitor in keloid keratinocytes and examined the abilities of cell proliferation, apoptosis, migration and invasion, the expressions of EMT-related markers vimentin and E-cadherin and stem-like cells-associated markers CD44 and ALDH1, and the involvement of PTEN and the signaling of AKT and ERK. Our results demonstrated that up-regulation or knockdown of miR-21-5p significantly increased or decreased the migration, invasion and sphere-forming abilities of keloid keratinocytes, and the phenotype of EMT and cells stemness were enhanced or reduced as well. Furthermore, PTEN and p-AKT were shown to participate in the regulation of miR-21-5p on EMT phenotypes and stemness signatures of keloid keratinocytes, which might account for the invasion and recurrence of keloids. This molecular mechanism of miR-21-5p on keloid keratinocytes linked EMT with cells stemness and implicated novel therapeutic targets for keloids.

This study aimed to explore the effects of miR-21 and PTEN/Akt signaling pathway on TGF-beta1-induced epithelial-mesenchymal transition (EMT) in gastric cancer (GC). GC tissues and adjacent tissues were collected from 83 patients. The qRT-PCR assay was performed to detect miR-21 expression. The expressions of PTEN, Akt and p-Akt were detected by immunohistochemistry. After 48 h of treatment with TGF-beta1 (10 ng/mL), the SGC-7901 and KATO-III cells were divided into the blank, negative control (NC), miR-21 inhibitors, PTEN-siRNA and miR-21 inhibitors + PTEN-siRNA groups. EMT related factors and PTEN expressions were detected by qRT-PCR assay and Western blotting. The scratch test was conducted to observe cell migration. Xenograft tumor model in nude mice was used to evaluate the effects of miR-21 on EMT of GC cells in vivo. In GC tissues, the expressions of miR-21, Akt and p-Akt were up-regulated, while PTEN expression was down-regulated. Gene and protein expressions of E-cadherin and PTEN in the miR-21 inhibitors group were higher than the blank, NC, PTEN-siRNA and miR-21 inhibitors + PTEN-siRNA groups, while the expressions of N-cadherin, beta-catenin, Vimentin and Slug in the miR-21 inhibitors group were lower than other groups. MiR-21 inhibitors significantly inhibit cell migration and invasion in GC cell lines. In vivo xenograft experiment revealed that miR-21 inhibitor inhibits the growth of transplanted tumor through up-regulating E-cadherin and PTEN expressions and down-regulating the expressions of N-cadherin, beta-catenin, Vimentin and Slug. These results suggest that miR-21 could promote TGF-beta1-induced EMT in GC cells through up-regulating PTEN expression.

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-beta induced EMT in CRC remain largely unknown. In the current study, we were interested to examine the role of NR2F2 in the TGF-beta-induced EMT in CRC. Here, we found NR2F2 was upregulated in CRC cells and promotes TGF-beta-induced EMT in CRC. Using comparative miRNA profiling TGF-beta 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-beta 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-beta-induced EMT and inhibit Smad7 expression via transactivation of miR-21, and NR2F2 may be a new common therapeutic target for CRC.CI - Copyright (c) 2017 Elsevier Inc. All rights reserved.

miR-21 induces epithelial-mesenchymal transition (EMT) of human cholangiocarcinoma (CCA) cells. However, the mechanism by which this occurs remains unclear. In the present study, high throughput platform was employed to detect the genes that are differential expressed in QBC939 cells transfected with a hsa-miR-21 antagomir or control vectors. The EMT-related Kruppel-like factor 4 (KLF4) gene was downregulated after miR-21 was knocked down. Overexpression of miR-21 upregulated KLF4, Akt, ERK and mesenchymal cell markers (N-cadherin and vimentin), downregulated the expression of epithelial cell marker E-cadherin and reduced cell migration and invasion. Immunohistochemistry showed that KLF4, pAkt and pERK were upregulated in tumor xenografts transfected with miR-21 mimics. Inhibitors of the PI3K-Akt and ERK1/2 pathways, LY294002 and U0126, significantly suppressed the EMT phenotype. The present data demonstrated that overexpression of miR-21, accompanied with KLF4, augmented the EMT via inactivation of Akt and ERK1/2 pathways. In conclusion, we have identified a novel mechanism that may be targeted in an attempt to relieve the malignant biological behavior of CCA cells.

BACKGROUND/AIMS: Lung cancer is one of the most common malignancies in the world. Apoptosis-stimulating protein of p53 (ASPP2), a tumorigenesis related protein, plays a critical role in the initiation and development of various types of cancers. However, the effect of ASPP2 on lung cancer remains unknown. The purpose of this study aims to investigate the mechanism of ASPP2 regulated by miR-21 in lung cancer in vitro and in vivo. METHODS: In the study, migration and invasion assays, apoptosis assay, caspase activity assay, TUNEL staining, real time PCR and western blot were used to investigate the mechanism of ASPP2 regulated by miR-21 in lung cancer in vitro and in vivo. RESULTS: We demonstrated that the miR-21 inhibitor induced apoptosis through inhibiting the PI3K/Akt/NF-kappaB signaling pathway in non-small cell lung carcinoma (NSCLC). Moreover, ASPP2 was directly targeted by miR-21 in NSCLC cells. Down-regulation of miR-21 suppressed cell migration and invasion, as well as the EMT signaling pathway in NSCLC cells. Furthermore, the miR-21 inhibitor induced cell apoptosis via the caspase dependent pathway in NSCLC cells. The miR-21 inhibitor enhanced caspase-3, 8, 9 activity in NSCLC cells. In addition, the caspase inhibitor significantly reduced the apoptosis induced by the miR-21 inhibitor in NSCLC cells. CONCLUSIONS: Our results revealed that the miR-21 inhibitor could induce apoptosis through inhibiting the PI3K/Akt/NF-kappaB signaling pathway in human NSCLC cells, and might serve as a therapeutic strategy to treat NSCLC.CI - (c) 2018 The Author(s). Published by S. Karger AG, Basel.

The prognosis of patients with gastric cancer remains poor mainly due to distant metastasis. Maternally expressed gene 3 (MEG3), a long noncoding RNA (lncRNA), is downregulated in various tumor tissues and suppresses tumor progression. miR21 is a microRNA which is expressed highly in tumor tissues. In the present study, we investigated the relationship between MEG3 and miR21 in regards to the cell mobility of gastric cancer. Our data demonstrated that MEG3 was downregulated while miR21 was upregulated in gastric cancer tissues and cell lines by qRTPCR. Overexpression of MEG3 suppressed cell mobility of gastric cancer cells (AGS) by downregulating the expression of MMP3, MMP9 and VEGF. As shown by western blot analysis, overexpression of MEG3 also suppressed epithelialmesenchymal transition (EMT) by increasing the expression of an epithelial marker (Ecadherin) and downregulating the expression of mesenchymal markers (Ncadherin, Snail and betacatenin), indicating that MEG3 suppressed cell mobility through the inhibition of EMT in gastric cancer. The expression of miR21 was negatively regulated by MEG3 and overexpression of miR21 promoted cell mobility of AGS through activation of EMT. Cotransfection of lncRNAMEG3 and miR21 mimic counteracted the inhibitory effect on the cell mobility attributed to MEG3, suggesting that the MEG3/miR21 axis affects cell mobility by suppressing EMT in gastric cancer. Using a mouse xenograft tumor model, we found that the overexpression of MEG3 suppressed tumor growth and metastasis while overexpression of miR21 had the opposite effects. The MEG3/miR21 axis affected gastric cancer growth and metastasis through inhibition of EMT in vivo. In conclusion, we demonstrated that the MEG3/miR21 axis participates in the tumor progression and metastasis of gastric cancer through the regulation of EMT.