Expression of miR-34a-5p and AKT1 gene in endometrial tissue of endometriosis and its effect on migration and invasion of endometrial stromal cells / 解放军医学杂志

Objective To investigate the expression of miR-34a-5p and AKT1 genes in endometrium tissues of patients with endometriosis (EM) and their effects on migration and invasion of endometrial stromal cells (ESCs). Methods A total of 91 patients, undergone hysterectomy in the Department of Obstetrics and Gynecology of Zhujiang Hospital of Southern Medical University from Jan. 2018 to Jun. 2019 due to benign gynecological diseases, were collected and divided into EM group (68 cases) and non-EM group (23 cases). The expressions of miR-34a-5p and AKT1 genes in endometrium tissues of patients were detected by in situ hybridization and immunohistochemistry. ESCs were transfected with miR-34a-5p mimic and negative control RNA (miR-NC) using liposome-3000 to construct the cell models of miR-34a-5p mimic group and miR-NC group. The cell proliferation rate was detected by CCK-8 method, and cell migration, invasion, apoptosis and autophagy ability experiments were performed to determine the effect of miR-34a-5p on ESCs' proliferation, migration, invasion, apoptosis and autophagy. Results The positive expression rate of miR-34a-5p was lower, and of AKT1 was higher in EM group than those in non-EM group (16.2% vs. 82.6%, X2=34.323; 72.1% vs. 30.4%, X2=12.581, P<0.001). After culturing for 12, 24 and 48 h, the cell proliferation rate was higher in miR-NC group than that in miR-34a-5p mimic group (P<0.05). The cell migration ability and invasion ability were higher in miR-NC group than those in miR-34a-5p mimic group with statistically significant difference [(65.00%±5.00%) vs. (30.67%±4.04%); (88.0±8.5) vs. (32.3±6.1), t=9.179, P<0.05]. The cell apoptosis rate and the expression level of LC3 gene were obviously lower in miR-NC group than those in miR-34a-5p mimic group [(9.33%±3.51%) vs. (18.00%±2.00%); (0.19±0.04) vs. (0.39±0.03), t=8.02, P<0.05]. Conclusion miR-34a-5p may be involved in the pathogenesis of EM by targeting AKT1 genes to affect the proliferation, migration, invasion, apoptosis and autophagy function of ESC.

LncRNA SNHG7/miR-34a-5p/SYVN1 axis plays a vital role in proliferation, apoptosis and autophagy in osteoarthritis

BACKGROUND: Osteoarthritis (OA) is one of the most common rheumatic diseases of which clinical symptoms includes swelling, synovitis and inflammatory pain, affect patients' daily life. It was reported that non-coding RNAs play vital roles in OA. However, the regulation mechanism of ncRNA in OA pathogenesis has not been fully elucidated. METHODS: The expression of SNHG7, miR-34a-5p and SYVN1 was detected using qRT-PCR in tissues, serum and cells. The protein expression of SYVN1, PCNA, cleavage-caspase 3, beclinl and LC3 were measured using western blot. The RNA immunoprecipitation (RIP), RNA pulldown, and luciferase reporter assays were used to verify the relationship between SNHG7, miR-34a-5p and SYVN1. The MTT and flow cytometry assay was performed to detected cell proliferation and cell apoptosis respectively. RESULTS: In this study, SNHG7 and SYVN1 expression were down-regulated, but miR-34a-5p was up-regulated in OA tissues and IL-1P treated cells compared with normal tissues and chondrocyte. Functional investigation revealed that up-regulated SNHG7 or down-regulated miR-34a-5p could promote cell proliferation and inhibit cell apoptosis and autophagy in OA cells. More than that, RIP, pulldown and luciferase reporter assay was applied to determine that miR-34a-5p was a target miRNA of SNHG7 and SYVN1 was a target mRNA of miR-34-5p. Rescue experiments showed that overexpression of miR-34a reversed high expression of SNHG7-mediated suppression of apoptosis and autophagy as well as promotion of proliferation, while its knockdown inhibited cell apoptosis and autophagy and promoted cell proliferation which could be impaired by silencing SYVN1. In addition, SNHG7 regulated SYVN1 through sponging miR-34a-5p. CONCLUSION: SNHG7 sponged miR-34a-5p to affect cell proliferation, apoptosis and autophagy through targeting SYVN1 which provides a novel sight into the pathogenesis of OA.

The Long Noncoding RNA NEAT1 Targets miR-34a-5p and Drives Nasopharyngeal Carcinoma Progression via Wnt/β-Catenin Signaling

PURPOSE: Long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) has been deemed an oncogene in many human cancers. However, the underlying mechanism of NEAT1 in nasopharyngeal carcinoma (NPC) progression remains largely unclear. MATERIALS AND METHODS: Quantitative real-time PCR assay was performed to assess the expression of NEAT1 and miR-34a-5p in NPC tissues and cells. Western blot analysis was used to observe cell epithelial to mesenchymal transition (EMT) and the activation of Wnt/β-catenin signaling in 5-8F cells. MiRNA directly interacting with NEAT1 were verified by dual-luciferase reporter assay and RNA immunoprecipitation. Cell proliferation ability was determined by CCK-8 assay, and cell migration and invasion capacities were assessed by transwell assays. An animal model was used to investigate the regulatory effect of NEAT1 on tumor growth in vivo. RESULTS: Our data revealed that NEAT1 is upregulated, while miR-34a-5p is downregulated in NPC tissues and cell lines. NEAT1 knockdown repressed tumor growth in vitro and in vivo. Additionally, we discovered that NEAT1 directly binds to miR-34a-5p and suppresses miR-34a-5p expression. Moreover, NEAT1 knockdown exerted suppression effects on cell proliferation, migration, invasion, and EMT by miR-34a-5p. NEAT1 knockdown blocked Wnt/β-catenin signaling via miR-34a-5p. CONCLUSION: Our study demonstrated that NEAT1 targets miR-34a-5p at least partly to drive NPC progression by regulating Wnt/β-catenin signaling, suggesting a potential therapeutic target for NPC.

lncRNA XIST/miR-34a-5p/SIRT6 axis regulates proliferation and metastasis of oral squamous cell carcinoma cells / 中国肿瘤生物治疗杂志

@# Objective: To investigate the molecular mechanisms of lncRNA XIST/miR-34a-5p/SIRT6 axis regulating the proliferation and metastasis of oral squamous cell carcinoma (OSCC) cells. Methods: Specimens of tumor tissues and paracancer tissues of 47 patients with OSCC, who visited the Qingdao Stomatological Hospital from March 2013 to March 2018, were enrolled in this study. qPCR was performed to measure the mRNA expressions of lncRNA XIST, miR-34a-5p and SIRT6 in OSCC tissues and cell lines. WB was performed to measure the protein levels of SIRT6, Ki67, pcDNA, cleaved-caspase3, cleaved-caspase8, E-cadherin and vimentin in OSCC tissues and cell lines. CCK-8 assay was performed to observe the effect of lncRNA XIST knockdown on proliferation of Cal-27 and Tca-8113 cells; Tanswell assay was performed to detect migration and invasion of Cal-27 and Tca-8113 cells; flow cytometry was performed to detect the apoptosis of Cal-27 and Tca-8113 cells; and dual luciferase reporter assay was performed to verify the relationship between lncRNAXIST and miR-34a, or miR-34a and SIRT6. Results: Expressions of lncRNAXIST and SIRT6 were up-regulated in OSCC tissues and cell lines (all P<0.05), reversely, miR-34a-5p was down-regulated in OSCC tissues and cell lines (P<0.01). lncRNA XIST knockdown significantly suppressed OSCC cells proliferation, migration and invasion, and induced apoptosis of OSCC cells (all P<0.01); however, simultaneous transfection with miR-34a-5p inhibitor or pcDNA-SIRT6 vector exerted opposite effect. lncRNA XIST knockdown significantly increased cell proliferation and metastasis related protein expression in OSCC cells (all P< 0.01), and simultaneous transfection with miR-34a-5p inhibitor or pcDNA-SIRT6 vector exerted opposite effect. In addition, lncRNA XIST directly targeted miR-34a-5p, and miR-34a-5p targeted SIRT6, lncRNA XIST upregulated SIRT6 expression via inhibiting miR34a-5p (P<0.01). Conclusion: Taken together, lncRNA XIST/miR-34a-5p/SIRT6 axis regulates the proliferation and metastasis of OSCC cells and provides potential therapeutic targets for OSCC.

miR-34a-5p inhibits the erythroid differentiation of K562 cells / 基础医学与临床

Objective To study the effects of microRNA-34a-5p on erythroid differentiation of K562 cells.Methods K562 cells were transfected with the microRNA-34a-5p mimics and antisense inhibitors specifically targeting mi-croRNA-34a-5p, respectively.The effects of over-expression or knocking-down of microRNA-34a-5p were exam-ined by Quantitative RT-PCR.Flow cytometry was performed to detect specific surface marker of erythroid cells . The benzidine staining assay was used to access the differentiation of K 562 cells.Western blot was performed to de-tect miRNA targets.Results microRNA-34a-5p was down-regulated at the early stage of K562 erythroid differenti-ation.Over-expression of microRNA-34a-5p in K562 cells attenuates erythroid differentiation , in contrast, inhibi-tion of microRNA-34a-5p accelerates erythroid pheotypes in K562 cells.c-MYB was found to be the direct target of microRNA-34 a-5 p in erythroid cells .Conclusions microRNA-34 a-5 p regulates early erythroid differentiation of K562 cells via repressing c-MYB.