LncRNA MT1JP functions as a ceRNA in regulating FBXW7 through competitively binding to miR-92a-3p in gastric cancer.
ABSTRACT: BACKGROUND:Emerging evidence has shown that dysregulation function of long non-coding RNAs (lncRNAs) implicated in gastric cancer (GC). However, the role of the differentially expressed lncRNAs in GC has not fully explained. METHODS:LncRNA expression profiles were determined by lncRNA microarray in five pairs of normal and GC tissues, further validated in another 75 paired tissues by quantitative real-time PCR (qRT-PCR). Overexpression of lncRNA MT1JP was conducted to assess the effect of MT1JP in vitro and in vivo. The biological functions were demonstrated by luciferase reporter assay, western blotting and rescue experiments. RESULTS:LncRNA MT1JP was significantly lower in GC tissues than adjacent normal tissues, and higher MT1JP was remarkably related to lymph node metastasis and advance stage. Besides, GC patients with higher MT1JP expression had a well survival. Functionally, overexpression of lncRNA MT1JP inhibited cell proliferation, migration, invasion and promoted cell apoptosis in vitro, and inhibited tumor growth and metastasis in vivo. Functional analysis showed that lncRNA MT1JP regulated FBXW7 expression by competitively binding to miR-92a-3p. MiR-92a-3p and down-regulated FBXW7 reversed cell phenotypes caused by lncRNA MT1JP by rescue analysis. CONCLUSION:MT1JP, a down-regulated lncRNA in GC, was associated with malignant tumor phenotypes and survival of GC. MT1JP regulated the progression of GC by functioning as a competing endogenous RNA (ceRNA) to competitively bind to miR-92a-3p and regulate FBXW7 expression. Our study provided new insight into the post-transcriptional regulation mechanism of lncRNA MT1JP, and suggested that MT1JP may act as a potential therapeutic target and prognosis biomarker for GC.
Project description:BACKGROUND:Cancer associated fibroblasts (CAFs) are key stroma cells that play dominant roles in tumor progression. However, the CAFs-derived molecular determinants that regulate colorectal cancer (CRC) metastasis and chemoresistance have not been fully characterized. METHODS:CAFs and NFs were obtained from fresh CRC and adjacent normal tissues. Exosomes were isolated from conditioned medium and serum of CRC patients using ultracentrifugation method and ExoQuick Exosome Precipitation Solution kit, and characterized by transmission electronic microscopy, nanosight and western blot. MicroRNA microarray was employed to identify differentially expressed miRNAs in exosomes secreted by CAFs or NFs. The internalization of exosomes, transfer of miR-92a-3p was observed by immunofluorescence. Boyden chamber migration and invasion, cell counting kit-8, flow cytometry, plate colony formation, sphere formation assays, tail vein injection and primary colon cancer liver metastasis assays were employed to explore the effect of NFs, CAFs and exosomes secreted by them on epithelial-mesenchymal transition, stemness, metastasis and chemotherapy resistance of CRC. Luciferase report assay, real-time qPCR, western blot, immunofluorescence, and immunohistochemistry staining were employed to explore the regulation of CRC metastasis and chemotherapy resistance by miR-92a-3p, FBXW7 and MOAP1. RESULTS:CAFs promote the stemness, epithelial-mesenchymal transition (EMT), metastasis and chemotherapy resistance of CRC cells. Importantly, CAFs exert their roles by directly transferring exosomes to CRC cells, leading to a significant increase of miR-92a-3p level in CRC cells. Mechanically, increased expression of miR-92a-3p activates Wnt/β-catenin pathway and inhibits mitochondrial apoptosis by directly inhibiting FBXW7 and MOAP1, contributing to cell stemness, EMT, metastasis and 5-FU/L-OHP resistance in CRC. Clinically, miR-92a-3p expression is significantly increased in CRC tissues and negatively correlated with the levels of FBXW7 and MOAP1 in CRC specimens, and high expression of exosomal miR-92a-3p in serum was highly linked with metastasis and chemotherapy resistance in CRC patients. CONCLUSIONS:CAFs secreted exosomes promote metastasis and chemotherapy resistance of CRC. Inhibiting exosomal miR-92a-3p provides an alternative modality for the prediction and treatment of metastasis and chemotherapy resistance in CRC.
Project description:Objective:This study aimed to determine the effects of the long non-coding (lnc) RNA MT1JP on the apoptosis and migration of hepatocellular carcinoma cells. Patients and Methods:Patients with liver cancer admitted to the Second People's Hospital of Liaocheng were included in this study. We transfected hepatocellular carcinoma cells with MT1JP and miR-24-3p and assessed their expression and effects on apoptosis and migration. Correlations were verified using a dual-luciferase reporter and RNA-binding protein coimmunoprecipitation. Results:The expression of MT1JP was downregulated (P < 0.05), whereas that of miR-24-3p was upregulated in liver cancer. Serum MT1JP levels were correlated with tumor size, alpha-fetoprotein (AFP), TNM stage, differentiation, and lymph node metastasis. Both MT1JP overexpression and miR-24-3p inhibition inhibited cellular proliferation and migration and increased apoptosis rates. They significantly downregulated expression of the cell migration-associated proteins matrix metalloproteinase -2, -9 (MMP-2, MMP-9) (P < 0.05). They upregulated the expression of Bcl-2-related X protein (Bax) and cysteinyl aspartate-specific proteinases (Caspase-3 and -9) proteins that are involved in apoptosis. They decreased expression of B-cell lymphoma/leukemia-2 (Bcl-2; P < 0.05). A target relationship between MT1JP and miR-24-3p was identified using dual-luciferase gene reporter assays and RNA-binding protein coimmunoprecipitations. MT1JP overexpression significantly downregulated miR-24-3p expression (P < 0.05). MT1JP and miR-24-3p expression were negatively correlated in liver cancer tissues (r = -0.561, P < 0.001; Pearson ?2 tests). Rescue experiments showed that upregulating miR-24-3p expression could counteract MT1JP overexpression in hepatocellular carcinoma cells. Conclusion:MT1JP, even when expressed at low levels, participates in the proliferation, apoptosis, and migration of liver cancer cells by regulating miR-24-3p.
Project description:The role of LncRNA ADAMTS9-AS2 in the regulation of chemoresistance of gastric cancer (GC) is largely unknown. Here we found that LncRNA ADAMTS9-AS2 was low-expressed in GC tissues and cells compared to their normal counterparts. In addition, LncRNA ADAMTS9-AS2 inhibited miR-223-3p expressions in GC cells by acting as competing endogenous RNA, and the levels of LncRNA ADAMTS9-AS2 and miR-223-3p showed negative correlations in GC tissues. Of note, overexpression of LncRNA ADAMTS9-AS2 inhibited GC cell viability and motility by sponging miR-223-3p. In addition, the levels of LncRNA ADAMTS9-AS2 were lower, and miR-223-3p was higher in cisplatin-resistant GC (CR-GC) cells than their parental cisplatin-sensitive GC (CS-GC) cells. LncRNA ADAMTS9-AS2 overexpression enhanced the cytotoxic effects of cisplatin on CR-GC cells, which were reversed by overexpressing miR-223-3p. Furthermore, LncRNA ADAMTS9-AS2 increased NLRP3 expressions by targeting miR-223-3p, and upregulation of LncRNA ADAMTS9-AS2 triggered pyroptotic cell death in cisplatin treated CR-GC cells by activating NLRP3 inflammasome through downregulating miR-223-3p. Finally, the promoting effects of LncRNA ADAMTS9-AS2 overexpression on CR-GC cell death were abrogated by pyroptosis inhibitor Necrosulfonamide (NSA). Collectively, LncRNA ADAMTS9-AS2 acted as a tumor suppressor and enhanced cisplatin sensitivity in GC cells by activating NLRP3 mediated pyroptotic cell death through sponging miR-223-3p.
Project description:OBJECTIVE:Long non-coding RNA (lncRNA) has become an important regulator of many human malignancies. However, the biological role and clinical significance of most lncRNA in gastric cancer (GC) remain unclear. METHODS:We investigate the biological function, mechanism of action and clinical expression of lncRNA MYOSLID in GC. First, we analysed the differential expression of lncRNA MYOSLID in GC tissues and non-cancerous tissues by analysing the sequencing data obtained from The Cancer Genome Atlas. Subsequently, we verified that lncRNA MYOSLID regulates the proliferation and apoptosis of GC cells by acting as a ceRNA against miR-29c-3p. The nude mouse xenograft was used to further confirm the functional significance of lncRNA MYOSLID in vivo. RESULTS:We found for the first time that the expression of lncRNA MYOSLID was significantly up-regulated in GC tissues, and the up-regulation of lncRNA MYOSLID in GC was correlated with tumour size, AJCC stage, depth of invasion and survival time. In addition, apoptosis and growth arrest can be induced in vitro after knockdown of lncRNA MYOSLID, which inhibits tumorigenesis in mouse xenografts in vivo. Further in-depth studies revealed that lncRNA MYOSLID acts as a ceRNA of miR-29c-3p, resulting in de-repression of its downstream target gene MCL-1. CONCLUSION:The lncRNA MYOSLID-miR-29c-3p-MCL-1 axis plays a key role in the development of GC. Our findings may provide potential new targets for the diagnosis and treatment of human GC.
Project description:The clinical application of doxorubicin (Dox) is limited due to its undesirable cardiotoxicity side effects. Cellular senescence plays an important role in Dox-induced cardiotoxicity. Exosomes derived from stem cells showed a therapeutic effect in Dox-induced cardiomyopathy (DIC). Hypoxia-preconditioned exosomes (exosomeHypoxia) display pro-metabolism and pro-survival abilities. Several long-noncoding RNAs/microRNAs act as competing endogenous RNAs (ceRNAs) modulating DIC. No study investigated whether exosomeHypoxia could attenuate DIC through modulating ceRNAs.Treatment of the human adipose-derived mesenchymal stem cells with hypoxia induced lncRNA-MALAT1 accumulation in the secreted exosomes. In addition, the lncRNA-MALAT1 was identified as an exosomal transfer RNA to repress miR-92a-3p expression. Silencing the lncRNA-MALAT1 in MSCs or miR-92a-3p overexpression in cardiomyocytes significantly impaired the rejuvenation induced by exosomeHypoxia. TargetScan and luciferase assay showed that miR-92a-3p targeted the ATG4a 3' untranslated region. Silencing ATG4a blocked the anti-senescent effect of exosomeHypoxia.This study identified the lncRNA-MALAT1 that functioned as ceRNA binding to miR-92a-3p, leading to ATG4a activation, thus improving mitochondrial metabolism. LncRNA-MALAT1/miR-92a-3p/ATG4a partially mediates the cardioprotective roles of exosomeHypoxia in Dox-induced cardiac damage. ExosomeHypoxia may serve as a potential therapeutic target against DIC.
Project description:Background:Abnormally expressed microRNAs (miRNAs) contribute greatly to the initiation and development of human cancers, including cervical cancer, by regulating the target mRNAs. MiR-27a-3p was up-regulated and acted as an oncogene in multiple cancers. However, the function of miR-27a-3p in cervical cancer has not been fully understood. Methods:The expression of miR-27a-3p in cervical cancer tissues and cell lines was detected by RT-pPCR. MTT assay, colony formation assay and flow cytometry analysis were performed to determine the effects of miR-27a-3p on the growth of cervical cancer cells. The targets of miR-27a-3p were predicted using the miRDB database. Luciferase reporter assay was utilized to confirm the binding between miR-27a-3p and the 3'-untranslated region (UTR) of targets. The expression of target proteins was determined by RT-qPCR and Western blot. Results:Our results found that miR-27a-3p was overexpressed in cervical cancer tissues and cell lines. Down-regulation of miR-27a-3p significantly inhibited the proliferation, colony formation and promoted apoptosis of cervical cancer cells. Overexpression of miR-27a-3p enhanced the cell proliferation. miR-27a-3p was found to bind the 3'-UTR of F-box and WD repeat domain containing 7 (FBXW7) and resulted in the down-regulation of FBXW7. The up-regulated level of miR-27a-3p was inversely correlated with that of FBXW7 in cervical cancer tissues. Additionally, reintroducing of FBXW7 significantly attenuated the promoting effect of miR-27a-3p on the proliferation of cervical cancer cells. Conclusion:These results indicated the growth-promoting function of miR-27a-3p in cervical cancer via targeting FBXW7. Our finding suggested the potential application of miR-27a-3p/FBXW7 axis in the diagnosis and treatment of cervical cancer.
Project description:MicroRNA (miRNA/miR)?92a has been identified as being significantly downregulated in non?small cell lung cancer (NSCLC) tissues using a miRNA array. However, its biological function and molecular mechanisms in NSCLC have not been fully elucidated. The aim of the present study was to determine the role of miR?92a in NSCLC and the mechanisms by which it affects NSCLC cells. The expression levels of miR?92a in NSCLC tissues and cell lines were analyzed using reverse transcription?quantitative PCR. Cell viability and cell apoptosis were determined using an MTT assay and flow cytometry, respectively. It was observed that miR?92a was significantly upregulated in NSCLC tissues and cell lines. Inhibition of miR?92a significantly suppressed viability of NSCLC cells, with concomitant downregulation of key proliferative genes, such as proliferating cell nuclear antigen and Ki?67. miR?92a downregulation induced apoptosis of NSCLC cells, as evidenced by flow cytometry and apoptosis?related protein detection. Luciferase assays confirmed that miR?92a could directly bind to the 3'?untranslated region of tumor suppressor F?box/WD repeat?containing protein 7 (FBXW7) and suppress its translation. Furthermore, small interfering RNA?mediated FBXW7 inhibition partially attenuated the tumor suppressive effect of an miR?92a inhibitor on NSCLC cells. Collectively, these findings demonstrated that miR?92a might function as an oncogene in NSCLC by regulating FBXW7. In conclusion, miR?92a could serve as a potential therapeutic target in NSCLC treatment.
Project description:Dysregulation of miR‑92a‑3p has been shown to contribute to many tumorigenic processes, and is correlated with tumor progression and prognosis. However, the association between miR‑92a‑3p and the clinicopathological features of Wilms tumorand its regulatory mechanism remain unknown. In the present study, we demonstrated that miR‑92a‑3p was downregulated in Wilms tumor tissues and was signiﬁcantly correlated with the lung metastasis of patients with Wilms tumor. Furthermore, miR‑92a‑3p mimics suppressed Wilms tumor cell proliferation, migration and invasion by in vitro assays. In addition, miR‑92a‑3p knockdown promoted tumor progression. Moreover, miR‑92a‑3p was shown to target directly the 3'‑UTR of NOTCH1 mRNA by Dual‑Luciferase reporter assays in Wilm's tumor cells. miR‑92a‑3p mimics decreased the expression of mRNA and protein of NOTCH1. miR‑92a‑3p inhibitor enhanced NOTCH1 expression by using western blotting and qPCR. In Wilms tumor tissues, NOTCH1 was highly expressed when compared with that in adjacent non‑tumor tissues. NOTCH1 expression was found to be negatively correlated with miR‑92a‑3p in tumor tissues. Knockdown of NOTCH1 expression reversed the promotive effect of miR‑92a‑3p inhibitor on the cell proliferation, migration and invasion in Wilms tumor. In conclusion, miR‑92a‑3p blocks the progression of Wilms tumor by targeting NOTCH1.
Project description:Background:Dysregulation of long non-coding RNAs (lncRNAs) is involved in development of prostate cancer. However, the molecular mechanisms of many lncRNAs in prostate cancer have not been studied yet. Methods:The lncRNA Fer-1-like protein 4 (FER1L4) expression was explored in prostate tumors and normal prostate tissues by RT-qPCR and bioinformatic analysis. Overexpression of FER1L4 was performed to evaluate its role in prostate cancer cell proliferation and survival. The molecular mechanism of FER1L4 was investigated by dual luciferase reporter assay, RNA pull down assay, western blotting and RT-qPCR. Results:It was found that FER1L4 was lower in prostate cancer tissues than normal tissues. Higher expression of FER1L4 was associated with prostate cancer tissues of early stage (AJCC stage I/II). Overexpression of FER1L4 inhibited cell proliferation and promoted cell apoptosis in prostate cancer cells. Bioinformatic analysis, RT-qPCR, RNA pull down assay and dual luciferase assay showed that FER1L4 upregulated F-box/WD repeat-containing protein 7 (FBXW7) tumor suppressor via sponging miR-92a-3p. Silencing of FBXW7 reversed the cell phenotypes caused by FER1L4 overexpression in prostate cancer cells. Conclusion:The data demonstrated that FER1L4, a downregulated lncRNA in prostate cancer, was pivotal for cell proliferation and survival of prostate cancer. The study provided new sights into understanding of the signaling network in prostate cancer and implied that FER1L4 might be a biomarker for patients with prostate cancer.
Project description:BACKGROUND:Accumulating works show that lncRNAs play critical roles in the development of gastric cancer (GC). LncRNA HLA complex group 18 (HCG18) was implicated in the progression of bladder cancer and glioma, but its role in GC is unknown. METHODS:RT-PCR was used to detect HCG18 and miR-141-3p expression in GC specimen. GC cell lines (AGS and MKN-28) were exploited as cell model. The biological effect of HCG18 on cancer cells was probed by CCK-8, colony formation, flow cytometry, Transwell and wound-healing experiments in vitro, and subcutaneous xenotransplanted tumor model and tail vein injection model in vivo. Interaction between HCG18 and miR-141-3p was determined by bioinformatics analysis, RT-PCR, and luciferase reporter experiments. Downstream gene expression of miR-141-3p, including Wiskott-Aldrich syndrome protein interacting protein family member 1 (WIPF1), Yes associated protein 1 (YAP), and tafazzin (TAZ) were detected using Western blot. RESULTS:HCG18 was markedly up-regulated in GC specimens, while miR-141-3p was markedly down-regulated. Down-regulation of HCG18 inhibited viability, migration, and invasion of GC cells, while miR-141-3p transfection led to opposite effect. HCG18 could down-regulate miR-141-3p through adsorbing it, and a negative association between HCG18 and miR-141-3p was found in GC specimens. HCG18 promoted WIPF1, YAP and TAZ expression, nonetheless, such influence was reversed by co-transfecting with miR-141-3p. CONCLUSION:HCG18 was aberrantly up-regulated in GC tissues, and it indirectly regulated the activity of Hippo signaling through counteracting miR-141-3p expression.