LINC02605 involved in paediatric Mycoplasma pneumoniae pneumonia complicated with diarrhoea via miR-539-5p/CXCL1 axis.

BACKGROUND: To investigate the involvement of LINC02605 in the progression of paediatric Mycoplasma pneumoniae pneumonia (MPP). METHODS: One hundred and thirty-two children with MPP (90 simple MPP and 42 MPP + diarrhoea) were enrolled, and their plasma was collected for detection of LINC026505 expression. CCK-8 kit and commercial apoptosis kit were introduced to determine cell growth and apoptosis. In silico prediction analyses were conducted to predict the downstream miRNA for LINC02605, following verification by dual luciferase reporter assay. The lipid-associated membrane proteins (LAMPs) were used to treat A549 and Coca-2 cells. RESULTS: LIN02605 was highly expressed in the MPP, especially in MPP complicated with diarrhoea. LINC02605 downregulation in A549 cells correlated with significant suppression of cell apoptosis rate and growth inhibition rate in vitro. Introduction of miR-539-5p inhibited luciferase activity in a reporter system containing the wild-type LINC02605 and CXCL1. After stimulation with LAMPs, overexpression of LINC02605 and CXCL1 and inhibition of miR-539-5p were found. miR-539-5p and CXCL1 knockdown resulted in a rescue effect on the LINC02605-inhibited cell apoptosis. LAMPs induced IL-1ß in intestinal epithelial cells and IL-1ß induced LINC02605 expression in A549 cells. CONCLUSIONS: LINC02605 was upregulated in MPP and miR-539-5p was a target for LINC02605. LINC02605 may be involved in the crosstalk between the gastrointestinal tract and the respiratory tract.

Bone marrow mesenchymal stem cells-derived exosomes containing miR-539-5p inhibit pyroptosis through NLRP3/caspase-1 signalling to alleviate inflammatory bowel disease.

BACKGROUND: Exosomes derived from bone mesenchymal stem cells (BMSCs) are potential candidates for inflammatory bowel disease (IBD) treatment. The present study investigated the therapeutic effect and potential mechanism of BMSCs-derived exosomes on pyroptosis in IBD. METHODS: We induced IBD in mice and cell models through dextran sulfate sodium (DSS) and LPS, respectively. The mRNA and protein expression levels were assessed by qRT-PCR, Western blotting, IF and IHC. The concentrations of IL-1ß, IL-18 and TNFα were assessed using ELISA. ROS levels were determined using DCFH-DA staining. Cell proliferation of mIECs was analysed using an MTT assay. In addition, a flow cytometry assay was performed to detect pyroptosis. Finally, the binding relationship between miR-539-5p and NLRP3 was verified by a dual luciferase reporter gene assay. RESULTS: Our results revealed that intraperitoneal injection of BMSCs-derived exosomes inhibited DSS-induced pyroptosis as well as IBD symptoms in mice. In addition, BMSCs-derived exosome treatment suppressed pyroptosis, ROS levels and the concentrations of proinflammatory cytokines (IL-1ß, IL-18 and TNFα) in LPS-treated mIECs in a miR-539-5p-dependent manner. Further research found that miR-539-5p suppressed NLRP3 expression in mIECs by directly targeting NLRP3. As expected, pyroptosis in LPS-treated mIECs was significantly reduced by NLRP3 knockdown. In addition, NLRP3 silencing restored the inhibitory effect of exosomes derived from BMSCs transfected with miR-539-5p inhibitor on pyroptosis in LPS-treated mIECs. CONCLUSION: The present study demonstrated that BMSCs-derived exosomal miR-539-5p suppresses pyroptosis through NLRP3/caspase-1 signalling to inhibit IBD progression.

lncRNA CRNDE promotes the proliferation and metastasis by acting as sponge miR-539-5p to regulate POU2F1 expression in HCC.

BACKGROUND: This article focuses on the roles and mechanism of lncRNA CRNDE on the progression of HCC. METHODS: We used qRT-PCR to detect the expression of lncRNA CRNDE in HCC cells, normal cells and clinical tissues. MTT assay, FCM analysis, Transwell migration and invasion assay were used to detect the effects of lncRNA CRNDE on cell viability, apoptosis, migration and invasion of HCC cells. The expression of apoptosis-related proteins Bcl-2, Bax, Cleaved Caspase 3, Cleaved Caspase 9, EMT epithelial marker E-cadherin and mesothelial marker Vimentin were analyzed by Western blot. Online prediction software was used to predict the binding sites between lncRNA CRNDE and miR-539-5p, or miR-539-5p and POU2F1 3'UTR. Dual luciferase reporter assay, qRT-PCR and RNA pulldown were used to detect target-relationship between lncRNA CRNDE and miR-539-5p. Dual luciferase reporter assay, qRT-PCR, Western blot and Immunofluorescence were used to detect target-relationship between miR-539-5p and POU2F1. qRT-PCR was used to detect the expression of miR-539-5p and POU2F1 in clinical tissues. Rescue experiments was used to evaluate the association among lncRNA CRNDE, miR-539-5p and POU2F1. Finally, we used Western blot to detect the effects of lncRNA CRNDE, miR-539-5p and POU2F1 on NF-κB and AKT pathway. RESULTS: lncRNA CRNDE was highly expressed in HCC cells and HCC tissues compared with normal cells and the corresponding adjacent normal tissues. lncRNA CRNDE promoted the cell viability, migration and invasion of HCC cells, while inhibited the apoptosis and promoted the EMT process of HCC cells. lncRNA CRNDE adsorbed miR-539-5p acts as a competitive endogenous RNA to regulate POU2F1 expression indirectly. In HCC clinical tissues, miR-539-5p expression decreased and POU2F1 increased compared with the corresponding adjacent normal tissues. lncRNA CRNDE/miR-539-5p/POU2-F1 participated the NF-κB and AKT pathway in HCC. CONCLUSION: lncRNA CRNDE promotes the expression of POU2F1 by adsorbing miR-539-5p, thus promoting the progression of HCC.

MiR-539-5p negatively regulates migration of rMSCs induced by Bushen Huoxue decoction through targeting Wnt5a.

Bone fractures are very common, and above 5% of the fractures are impaired, leading to nonunions and severe disablilities. The traditional Chinese medicine Bushen Huoxue decoction (BHD) has been used to treat fracture in China. Our previous report has found that BHD promotes migration of rat mesenchymal stem cells (rMSCs) by activating Wnt5a signaling pathway. However, whether and how miRNAs are involved in modulating rMSCs migration induced by BHD has not been explored. In the present study, miRNA microarray analysis and further validation by real-time quantitative RT-PCR revealed that miR-539-5p was down-regulated in BHD-induced rMSCs. Transfection of miR-539-5p mimics suppressed rMSCs migration while the miR-539-5p inhibitor promoted rMSCs migration. Our results suggested that miR-539-5p was a negative regulator of migration of rMSCs induced by BHD. Target prediction analysis tools and Dual-luciferase reporter gene assay identified Wnt5a as a direct target of miR-539-5p. MiR-539-5p inhibited the expression of the Wnt5a and its downstream signaling molecules including JNK, PKC and CaMKII, which played a critical role in regulating migration of rMSCs. Taken together, our results demonstrate that miR-539-5p negatively regulates migration of rMSCs induced by BHD through targeting Wnt5a. These findings provide evidence that miR-539-5p should be considered as an important candidate target for the development of preventive or therapeutic approaches against bone nonunions.

TsMS combined with EA promotes functional recovery and axonal regeneration via mediating the miR-539-5p/Sema3A/PlexinA1 signalling axis in sciatic nerve-injured rats.

Enhancing axonal regeneration is one of the most important processes in treating nerve injuries. Both magnetic and electrical stimulation have the effect of promoting nerve axon regeneration. But few study has investigated the effects of trans-spinal magnetic stimulation (TsMS) combined with electroacupuncture (EA) on nerve regeneration in rats with sciatic nerve injury. In this study, we compared the improvement of neurological function in rats with sciatic nerve crush injuries after 4 weeks of different interventions (EA, TsMS, or TsMS combined with EA). We further explored the morphological and molecular biological alterations following sciatic nerve injury by HE, Masson, RT-PCR, western blotting, immunofluorescence staining and small RNA transcriptome sequencing. The results showed that TsMS combined with EA treatment significantly promoted axonal regeneration, increased the survival rate of neurons, and suppressed denervation atrophy of the gastrocnemius muscle. Subsequent experiments suggested that the combination treatment may play an active role by mediating the miR-539-5p/Sema3A/PlexinA1 signaling axis.

miR-539-5p targets BMP2 to regulate Treg activation in B-cell acute lymphoblastic leukemia through TGF-ß/Smads/MAPK.

MicroRNAs (mRNAs) were believed to play an important role in cancers, and this study aimed to explore the mechanism of miRNA regulating Treg in B-cell acute lymphoblastic leukemia (B-ALL). Firstly, the differentially expressed miRNAs and target genes significantly associated with Tregs were screened out by high-throughput sequencing, and their enrichment pathways were analyzed. The binding relationship between miRNA and target genes was further verified, and the effects of miRNA on the proliferation and apoptosis of B-ALL Nalm-6 cells and Treg activation were analyzed. Results showed that differentially expressed miR-539-5p was significantly under-expressed, and its target gene BMP2 was significantly over-expressed in B-ALL, and significantly enriched in the TGF-ß1 pathway. In addition, both miR-539-5p and BMP2 were significantly correlated with Treg activity in B-ALL. In vitro experiments further confirmed that miR-539-5p could directly target BMP2. The low expression of miR-539-5p in B-ALL significantly promoted BMP2 expression to promote the proliferation and inhibit apoptosis of Nalm-6 cells. Furthermore, the high expression of BMP2 in B-ALL could cooperate with TGF-ß1 to promote the activation of human CD4+CD25-T cells to Treg, and significantly activate the TGF-ß/Smads/MAPK pathway. In vivo experiments also confirmed that overexpression of miR-539-5p significantly inhibited BMP2 to suppress Treg activation and Smad1 and Smad2 phosphorylation, and finally inhibit the B-ALL process. In conclusion, miR-539-5p was significantly under-expressed in B-ALL and could target BMP2 to promote its expression, and the overexpressed BMP2 further promoted Treg activation in B-ALL by regulating TGF-ß/Smads/MAPK pathway.

LncRNA TP73-AS1 enhances the malignant properties of colorectal cancer by increasing SPP-1 expression through miRNA-539-5p sponging.

Colorectal cancer (CC) is one of the most aggressive cancers, with a high mortality rate worldwide. This study focuses on the mechanism of CC to explore the effective therapeutic targets. We determined that LncRNA TP73-AS1 (TP-73-AS1) expression was significantly increased in CC tissues. TP73-AS1 silencing dynamically inhibited the proliferation, migratory and invasive capacity in CC cells. Mechanistically, we found that TP73-AS1 targeted miR-539-5p and miR-539-5p silencing could promote the migratory and invasive capacity in CC cells. Further study also confirmed that SPP-1 expression significantly increased after co-transfection of miR-539-5p inhibitors. Knockdown the SPP-1 can reverse malignant properties of CC cells. Si-TP73-AS1 suppressed the tumor growth of CC cells in vivo. In a word, we found that TP73-AS1 enhances the malignant properties of colorectal cancer by increasing SPP-1 expression through miRNA-539-5p sponging. And our study provides a potential therapeutic target of CC.

miR-539-5p regulates Srebf1 transcription in the skeletal muscle of diabetic mice by targeting DNA methyltransferase 3b.

Aberrant DNA methylation is associated with diabetes, but the precise regulatory events that control the levels and activity of DNA methyltransferases (DNMTs) is not well understood. Here we show that miR-539-5p targets Dnmt3b and regulates its cellular levels. miR-539-5p and Dnmt3b show inverse patterns of expression in skeletal muscle of diabetic mice. By binding to the 3' UTR of Dnmt3b, miR-539-5p downregulates its levels in C2C12 cells and in human primary skeletal muscle cells. miR-539-5p-Dnmt3b interaction regulates Srebf1 transcription by altering methylation at CpG islands within Srebf1 in C2C12 cells. Dnmt3b inhibition alone was sufficient to upregulate Srebf1 transcription. In vivo antagonism of miR-539-5p in normal mice induced hyperglycemia and hyperinsulinemia and impaired oral glucose tolerance. These mice had elevated Dnmt3b and decreased Srebf1 levels in skeletal muscle. db/db mice injected with miR-539-5p mimics showed improved circulatory glucose and cholesterol levels. Oral glucose tolerance improved together with normalization of Dnmt3b and Srebf1 levels in skeletal muscle. Our results support a critical role of miR-539-5p and Dnmt3b in aberrant skeletal muscle metabolism during diabetes by regulating Srebf1 transcription; modulating the miR-539-5p-Dnmt3b axis might have therapeutic potential for addressing altered skeletal muscle physiology during insulin resistance and type 2 diabetes.

LncRNA PCGEM1 contributes to malignant behaviors of glioma by regulating miR-539-5p/CDK6 axis.

BACKGROUND: Glioma, one of the most prevalent and aggressive cancers, is regulated by long noncoding RNAs (lncRNAs). This study aims to research the functional mechanism of lncRNA PCGEM1 involved in glioma progression. METHODS: Expression levels of PCGEM1, miR-539-5p and CDK6 were analyzed by qRT-PCR in NHA, U251, U87, and LN229 cells or glioma tissues. shRNAs were used to knock down PCGEM1 in U251 and LN229 cells. Kaplan-Meier curve and log rank test were utilized to examine survival rate. CCK8 (Cell Counting Kit-8) assay, colony formation assay and EdU staining were conducted to detect cell proliferation. Transwell assay was performed to evaluate cell migration and invasion. Luciferase reporter assay was conducted to assess RNA interaction between PCGEM1 and miR-539-5p. Nude mice were used for tumor xenograft assay. RESULTS: LncRNA PCGEM1 was upregulated in glioma tissues and tumor cell lines. PCGEM1 upregulation predicted unsatisfactory prognosis. PCGEM1 knockdown inhibited proliferation, colony formation, migration and invasion. PCGEM1 knockdown delayed tumor growth in vivo. PCGEM1 played as a competing endogenous RNA (ceRNA) for miR-539-5p to promote CDK6 expression. MiR-539-5p mimics repressed glioma progression while CDK6 overexpression reversed the roles of PCGEM1 knockdown. CONCLUSION: PCGEM1 knockdown suppressed glioma progression through sponging miR-539-5p and regulating CDK6 expression, implying PCGEM1 as a potential therapeutic target.

CASC21, a FOXP1 induced long non-coding RNA, promotes colorectal cancer growth by regulating CDK6.

Emerging studies indicate that long non-coding RNAs (lncRNAs) play crucial roles in colorectal cancer (CRC). Here, we reported lncRNA CASC21, which is induced by FOXP1, functions as an oncogene in CRC. We systematically elucidated its clinical significance and possible molecular mechanism in CRC. LncRNA expression in CRC was analyzed by RNA-sequencing data in TCGA. The expression level of CASC21 in tissues was determined by qRT-PCR. The functions of CASC21 was investigated by in vitro and in vivo assays (CCK8 assay, colony formation assay, EdU assay, xenograft model, flow cytometry assay, immunohistochemistry (IHC) and Western blot). Chromatin immunoprecipitation (ChIP), RNA immunoprecipitation (RIP) and luciferase reporter assays were utilized to demonstrate the potential mechanisms of CASC21. CASC21 is overexpressed in CRC and high CASC21 expression is associated with poor survival. Functional experiments revealed that CASC21 promotes CRC cell growth. Mechanistically, we found that CASC21 expressed predominantly in the cytoplasm. CASC21 could interact with miR-539-5p and regulate its target CDK6. Together, our study elucidated that CASC21 acted as an oncogene in CRC, which might serve as a novel target for CRC diagnosis and therapy.

MiR-539-5p Decreases amyloid ß-protein production, hyperphosphorylation of Tau and Memory Impairment by Regulating PI3K/Akt/GSK-3ß Pathways in APP/PS1 Double Transgenic Mice.

The production of amyloid ß (Aß) and tau hyperphosphorylation have been identified as key processes in Alzheimer's disease (AD) pathogenesis. MiR-539-5p has been found to be abnormally expressed in brain tissue; however, the functional role of miR-539-5p in the pathogenesis of AD remains unclear. In our study, we found that the expression of miR-539-5p was significantly downregulated in humans and mice with AD and was negatively correlated with expression of APP, caveolin 1, and GSK-3ß. Moreover, upregulation of miR-539-5p inhibited Aß accumulation, tau phosphorylation, oxidative stress, and apoptosis and improved memory ability in AD mice. Furthermore, by using bioinformatics tool and dual-luciferase reporter assay, APP, Caveolin 1, and GSK-3ß were confirmed as direct targets of miR-539-5p. In addition, the PI3K/AKT/GSK-3ß signaling pathway can be regulated by miR-539-5p. In conclusion, this study provided a novel insight into the pathologic mechanism of AD by identifying that miR-539-5p plays a neuroprotective role in AD.

Long Non-coding RNA LINC00339 Stimulates Glioma Vasculogenic Mimicry Formation by Regulating the miR-539-5p/TWIST1/MMPs Axis.

Glioma is recognized as a highly angiogenic malignant brain tumor. Vasculogenic mimicry (VM) greatly restricts the therapeutic effect of anti-angiogenic tumor therapy for glioma patients. However, the molecular mechanisms of VM formation in glioma remain unclear. Here, we demonstrated that LINC00339 was upregulated in glioma tissue as well as in glioma cell lines. The expression of LINC00339 in glioma tissues was positively correlated with glioma VM formation. Knockdown of LINC00339 inhibited glioma cell proliferation, migration, invasion, and tube formation, meanwhile downregulating the expression of VM-related molecular MMP-2 and MMP-14. Furthermore, knockdown of LINC00339 significantly increased the expression of miR-539-5p. Both bioinformatics and luciferase reporter assay revealed that LINC00339 regulated the above effects via binding to miR-539-5p. Besides, overexpression of miR-539-5p resulted in decreased expression of TWIST1, a transcription factor known to play an oncogenic role in glioma and identified as a direct target of miR-539-5p. TWIST1 upregulated the promoter activities of MMP-2 and MMP-14. The in vivo study showed that nude mice carrying tumors with knockdown of LINC00339 and overexpression of miR-539-5p exhibited the smallest tumor volume through inhibiting VM formation. In conclusion, LINC00339 may be used as a novel therapeutic target for VM formation in glioma.

Long non-coding RNA LOC100129148 functions as an oncogene in human nasopharyngeal carcinoma by targeting miR-539-5p.

Emerging studies have shown that long noncoding RNAs (lncRNAs) play critical roles in carcinogenesis and progression, including human nasopharyngeal carcinoma (NPC). The correlation between lncRNAs expression and NPC development has not been well identified in the recent literature. Recently, high-through put analysis reveals that LOC100129148 is highly expressed in NPC. However, whether the aberrant expression of LOC100129148 in NPC is corrected with tumorigenesis or prognosis has not been investigated. Herein, we identified that LOC100129148 was up-regulated in NPC tissues and cell lines, and higher expression of LOC100129148 resulted in a markedly poorer survival time. Over-expressed LOC100129148 favored, but silenced LOC100129148 hampered cell proliferation in NPC cells. Additionally, LOC100129148 enhanced the KLF12 expression through functioning as a competitive 'sponge' for miR-539-5p. Thus, our study reports a novel mechanism underlying NPC carcinogenesis, and provides a potential novel diagnosis and treatment biomarker for NPC.