Dataset Information

MiR-335-5p inhibits TGF-?1-induced epithelial-mesenchymal transition in non-small cell lung cancer via ROCK1.

ABSTRACT: BACKGROUND:Significant evidence has shown that the miRNA pathway is an important component in the downstream signaling cascades of TGF-?1 pathway. Our previous study has indicated that miR-335-5p expression was significantly down-regulated and acted as a vital player in the metastasis of non-small cell lung cancer (NSCLC), however the underlying mechanism remained unclear. METHODS:The differential expression level of miR-335-5p and ROCK1 were determined by qRT-PCR and IHC analysis in human tissue samples with or without lymph node metastasis. Transwell assay was conducted to determine cell ability of migration and invasion. SiRNA interference, microRNA transfection and western blot analysis were utilized to clarify the underlying regulatory mechanism. RESULTS:We showed that down-regulated expression of miR-335-5p and up-regulated expression of ROCK1 in NSCLC tissues were associated with lymph node metastasis. Over-expresion of miR-335-5p significantly inhibited TGF-?1-mediated NSCLC migration and invasion. Furthermore, luciferase reporter assays proved that miR-335-5p can bind to 3'-UTR of ROCK1 directly. Moreover, we confirmed that siRNA-mediated silencing of ROCK1 significantly diminished TGF-?1-mediated EMT and migratory and invasive capabilities of A549 and SPC-A1 cells. CONCLUSION:This is the first time to report that miR-335-5p regulates ROCK1 and impairs its functions, thereby playing a key role in TGF-?1-induced EMT and cell migration and invasion in NSCLC.

SUBMITTER: Du W

PROVIDER: S-EPMC6805547 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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miR-335-5p inhibits TGF-β1-induced epithelial-mesenchymal transition in non-small cell lung cancer via ROCK1.

Du Wenwen W Tang Haicheng H Lei Zhe Z Zhu Jianjie J Zeng Yuanyuan Y Liu Zeyi Z Huang Jian-An JA

Respiratory research 20191021 1

<h4>Background</h4>Significant evidence has shown that the miRNA pathway is an important component in the downstream signaling cascades of TGF-β1 pathway. Our previous study has indicated that miR-335-5p expression was significantly down-regulated and acted as a vital player in the metastasis of non-small cell lung cancer (NSCLC), however the underlying mechanism remained unclear.<h4>Methods</h4>The differential expression level of miR-335-5p and ROCK1 were determined by qRT-PCR and IHC analysis ...[more]

PMID: 31638991

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Project description:Inflammatory bowel disease (IBD), which consists of Crohn's disease (CD) and ulcerative colitis (UC), is a chronic, inflammatory disorder of the gastro-intestinal tract with unknown etiology. Current evidence suggests that intestinal epithelial cells (IECs) is prominently linked to the pathogenesis of IBD. Therefore, maintaining the intact of epithelium has potential roles in improving pathophysiology and clinical outcomes of IBD. MicroRNAs (miRNAs) act as post-transcriptional gene regulators and regulate many biological processes, including embryonal development, cell differentiation, apoptosis and proliferation. In this study, we found that miR-200b decreased significantly in inflamed mucosa of IBD, especially for UC, when compared with their adjacent normal tissue. Simultaneously, we also found that the genes of E-cadherin and cyclin D1 were reduced significantly and correlated positively to the miR-200b. In addition, the upregulation of transforming growth factor-beta 1 (TGF-?1) was inversely correlated to the miR-200b in IBD. To investigate the possible roles of miR-200b in IECs maintaining, we used TGF-?1 to induce epithelial-mesenchymal transition (EMT) in IEC-6 initially. After sustained over-expressing miR-200b in IEC-6, the EMT was inhibited significantly that was characterized by downregulation of vimentin and upregulation of E-cadherin. Furthermore, we found that miR-200b enhanced E-cadherin expression through targeting of ZEB1, which encode transcriptional repressors of E-cadherin. SMAD2 was found to act as a target of miR-200b with direct evidence that miR-200b binding to the 3' UTR of SAMD2 and the ability of miR-200b to repress SMAD2 protein expression. With SMAD2 depletion, the expression of vimentin decreased correspondingly, which suggested miR-200b might reduce vimentin through regulating the SMAD2. With endogenous over-expression of miR-200b, the proliferation of IEC-6 cells increased significantly by increasing S-phase entry and promoting expression of the protein cyclin D1. Summarily, our study suggested a potential role for mir-200b in maintaining intact of intestinal epithelium through inhibiting EMT and promoting proliferation of IECs.

Project description:Transforming growth factor (TGF)‑β1 is a key cytokine affecting the pathogenesis and progression of cervical cancer. Tumor‑derived exosomes contain microRNAs (miRNAs/miRs) that interact with cancer and stromal cells, thereby contributing to tissue remodeling in the tumor microenvironment (TME). The present study was designed to clarify how TGF‑β1 affects tumor biological functions through exosomes released by cervical cancer cells. Deep RNA sequencing found that TGF‑β1 stimulated cervical cancer cells to secrete more miR‑663b‑containing exosomes, which could be transferred into new target cells to promote metastasis. Further studies have shown that miR‑663b directly targets the 3'-untranslated regions (3'‑UTR) of mannoside acetylglucosaminyltransferase 3 (MGAT3) and is involved in the epithelial‑mesenchymal transition (EMT) process. Remarkably, the overexpression of MGAT3 suppressed cervical cancer cell metastasis promoted by exosomal miR‑663b, causing increased expression of epithelial differentiation marker E‑cadherin and decreased expression of mesenchymal markers N‑cadherin and β‑catenin. Throughout our study, online bioinformation tools and dual luciferase reporter assay were applied to identify MGAT3 as a novel direct target of miR‑663b. Exosome PKH67‑labeling experiment verified that exosomal miR‑663b could be endocytosed by cervical cancer cells and subsequently influence its migration and invasion functions which were measured by wound healing and Transwell assays. The expression of miR‑663b and MGAT3 and the regulation of the EMT pathway caused by MGAT3 were detected by quantitative real‑time transcription‑polymerase chain reaction (qPCR) and western blot analysis. These results, thus, provide evidence that cancer cell‑derived exosomal miR‑663b is endocytosed by cervical cancer cells adjacent or distant after TGF‑β1 exposure and inhibits the expression of MGAT3, thereby accelerating the EMT process and ultimately promoting local and distant metastasis.

Project description:BackgroundGlioma metastasis, invasion, epithelial-mesenchymal transition (EMT) and chemoresistance indicate poor prognosis. Accumulating evidence reveals that Notch1 is an important factor in tumour progression. However, the role of Notch1 in glioma EMT and associated microRNAs (miRNAs) with the Notch pathway remain controversial.MethodsUtilizing cBioPortal database to examine the gene signature of NOTCH1 (encoding Notch1), CDH2 (encoding N-cadherin) and SNAI1 (encoding Snail-1) in disease-free survival (DFS) and overall survival (OS). We analyzed the Notch1 expression from Oncomine. We used Western blot (WB), immunohistochemistry (IHC) and immunofluorescence to determine protein levels. Transcription was evaluated by quantitative real-time (qRT)-PCR. siRNA and lentivirus were used to knock down Notch1 and overexpress miR-139-5p, respectively. The migration and invasion of glioma cells were assessed by wound healing and transwell assays. Luciferase reporter assays were utilized to verify the relationship between Notch1 and miR-139-5p. A U87-implanted intracranial model was used to study the effect of miR-139-5p on tumour growth and Notch1 suppression efficacy or EMT reversion.ResultsIt revealed the association of NOTCH1, CDH2, SNAI1 genomic alterations with decreases in DFS and OS. Notch1 was upregulated in classical and proneural subtypes of GBM, and associated with tumour grade. Notch1 inhibition suppressed the biological behaviours of metastasis, invasion and EMT. Notch1 was identified as a novel direct target of miR-139-5p. MiR-139-5p overexpression partially phenocopied Notch1 siRNA, whereas the forced expression of Notch1 reversed the effects of miR-139-5p on the invasion of glioma. Moreover, intracranial tumourigenicity and EMT behaviours were reduced by the introduction of miR-139-5p and partially mediated by the decreased Notch1 expression.ConclusionsmiR-139-5p was identified as a tumour suppressor by negatively targeting Notch1, and this work suggests a possible molecular mechanism of the miR-139/Notch1/EMT axis for glioma treatment.

Dataset Information

MiR-335-5p inhibits TGF-?1-induced epithelial-mesenchymal transition in non-small cell lung cancer via ROCK1.

Publications

miR-335-5p inhibits TGF-β1-induced epithelial-mesenchymal transition in non-small cell lung cancer via ROCK1.

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