Reciprocal regulation of miR‐206 and IL‐6/STAT3 pathway mediates IL6‐induced gefitinib resistance in EGFR‐mutant lung cancer cells

Persistently activated IL‐6/STAT3 pathway promotes acquired resistance to targeted therapy with epidermal growth factor receptor‐tyrosine kinase inhibitors (EGFR‐TKIs) in non–small‐cell lung cancer (NSCLC) treatment. miR‐206 has been verified to be dysregulated and plays as a negative regulator in lung cancer. However, whether miR‐206 may overcome IL6‐induced gefitinib resistance in EGFR‐mutant lung cancer remains elusive. In this study, we investigated the role of miR‐206 in IL6‐induced gefitinib‐resistant EGFR‐mutated lung cancer cell lines. We showed that forced miR‐206 expression restored gefitinib sensitivity in IL6‐induced gefitinib‐resistant EGFR‐mutant lung cancer cells by inhibiting IL6/JAK1/STAT3 pathway. Specifically, mechanistic investigations revealed that miR‐206 blocked IL‐6/STAT3 signalling via directly targeting the 3'‐UTR of intracellular IL‐6 messenger RNA. Moreover, IL‐6 induced miR‐206 down‐regulation by reducing the cropping process of primary miR‐206 (pri‐miR‐206) into the Drosha/DGCR8 complex. Taken together, our findings reveal a direct role of miR‐206 in regulating IL‐6/STAT3 pathway and contrarily activated IL‐6/STAT3 signalling mediates the miR‐206 maturation process in gefitinib‐resistant EGFR‐mutant lung cancer cells.     

miR‐134 inhibits non‐small cell lung cancer growth by targeting the epidermal growth factor receptor

The epidermal growth factor receptor (EGFR) is frequently activated in a wide range of solid tumours and represents an important therapeutic target. MicroRNAs (miRNAs) have recently been recognized as a rational and potential modality for anti‐EGFR therapies. However, more EGFR‐targeting miRNAs need to be explored. In this study, we identified a novel EGFR‐targeting miRNA, miRNA‐134 (miR‐134), in non‐small‐cell lung cancer (NSCLC) cell lines. Luciferase assays confirmed that EGFR is a direct target of miR‐134. In addition, the overexpression of miR‐134 inhibited EGFR‐related signaling and suppressed NSCLC cells proliferation by inducing cell cycle arrest and/or apoptosis, suggesting that miR‐134 functions as a tumour suppressor in NSCLC. Further mechanistic investigation including RNAi and rescue experiments suggested that the down‐regulation of EGFR by miR‐134 partially contributes to the antiproliferative role of miR‐134. Last, in vivo experiments demonstrated that miR‐134 suppressed tumour growth of A549 xenograft in nude mice. Taken together, our findings suggest that miR‐134 inhibits non‐small cell lung cancer growth by targeting the EGFR.     

miR‐1‐3p and miR‐206 sensitizes HGF‐induced gefitinib‐resistant human lung cancer cells through inhibition of c‐Met signalling and EMT

Hepatocyte growth factor (HGF) overexpression is an important mechanism in acquired epidermal growth factor receptor (EGFR) kinase inhibitor gefitinib resistance in lung cancers with EGFR activating mutations. MiR‐1‐3p and miR‐206 act as suppressors in lung cancer proliferation and metastasis. However, whether miR‐1‐3p and miR‐206 can overcome HGF‐induced gefitinib resistance in EGFR mutant lung cancer is not clear. In this study, we showed that miR‐1‐3p and miR‐206 restored the sensitivities of lung cancer cells PC‐9 and HCC‐827 to gefitinib in present of HGF. For the mechanisms, we demonstrated that both miR‐1‐3p and miR‐206 directly target HGF receptor c‐Met in lung cancer. Knockdown of c‐Met mimicked the effects of miR‐1‐3p and miR‐206 transfections Meanwhile, c‐Met overexpression attenuated the effects of miR‐1‐3p and miR‐206 in HGF‐induced gefitinib resistance of lung cancers. Furthermore, we showed that miR‐1‐3p and miR‐206 inhibited c‐Met downstream Akt and Erk pathway and blocked HGF‐induced epithelial‐mesenchymal transition (EMT). Finally, we demonstrated that miR‐1‐3p and miR‐206 can increase gefitinib sensitivity in xenograft mouse models in vivo. Our study for the first time indicated the new function of miR‐1‐3p and miR‐206 in overcoming HGF‐induced gefitinib resistance in EGFR mutant lung cancer cell.     

Exosomes transmit T790M mutation‐induced resistance in EGFR‐mutant NSCLC by activating PI3K/AKT signalling pathway

Emerging evidence has shown that exosomes derived from drug‐resistant tumour cells are able to horizontally transmit drug‐resistant phenotype to sensitive cells. However, whether exosomes shed by EGFR T790M‐mutant–resistant NSCLC cells could transfer drug resistance to sensitive cells has not been investigated. We isolated exosomes from the conditioned medium (CM) of T790M‐mutant NSCLC cell line H1975 and sensitive cell line PC9. The role and mechanism of exosomes in regulating gefitinib resistance was investigated both in vitro and in vivo. Exosome‐derived miRNA expression profiles from PC9 and H1975 were analysed by small RNA sequencing and confirmed by qRT‐PCR. We found that exosomes shed by H1975 could transfer gefitinib resistance to PC9 both in vitro and in vivo through activating PI3K/AKT signalling pathway. Small RNA sequencing and RT‐PCR confirmed that miR‐3648 and miR‐522‐3p were the two most differentially expressed miRNAs and functional study showed that up‐regulation of miR‐522‐3p could induce gefitinib resistance in PC9 cell. The findings of our study reveal an important mechanism of acquired resistance to EGFR‐TKIs in NSCLC.     

MiR‐139‐5p,miR‐940 and miR‐193a‐5p inhibit the growth of hepatocellular carcinoma by targeting SPOCK1

The study was aimed to screen out miRNAs with differential expression in hepatocellular carcinoma (HCC), and to explore the influence of the expressions of these miRNAs and their target gene on HCC cell proliferation, invasion and apoptosis. MiRNAs with differential expression in HCC were screened out by microarray analysis. The common target gene of these miRNAs (miR‐139‐5p, miR‐940 and miR‐193a‐5p) was screened out by analysing the target genes profile (acquired from Targetscan) of the three miRNAs. Expression levels of miRNAs and SPOCK1 were determined by quantitative real time polymerase chain reaction (qRT‐PCR). The target relationships were verified by dual luciferase reporter gene assay and RNA pull‐down assay. Through 3‐(4,5‐dimethyl‐2‐thiazolyl)‐2,5‐diphenyl‐2‐H‐tetrazolium bromide,thiazolyl blue tetrazolium bromide (MTT) and transwell assays and flow cytometry, HCC cell viability, invasion and apoptosis were determined. In vivo experiment was conducted in nude mice to investigate the influence of three miRNAs on tumour growth. Down‐regulation of miR‐139‐5p, miR‐940 and miR‐193a‐5p was found in HCC. Overexpression of these miRNAs suppressed HCC cell viability and invasion, promoted apoptosis and inhibited tumour growth. SPOCK1, the common target gene of miR‐139‐5p, miR‐940 and miR‐193a‐5p, was overexpressed in HCC. SPOCK1 overexpression promoted proliferation and invasion, and restrained apoptosis of HCC cells. MiR‐139‐5p, miR‐940 and miR‐193a‐5p inhibited HCC development through targeting SPOCK1.     

Linc01014 regulates gefitinib resistance in oesophagus cancer via EGFR‐PI3K‐AKT‐mTOR signalling pathway

This study aimed to explore the underlying mechanism of linc01014 in oesophagus cancer gefitinib resistance. Gefitinib‐resistant oesophagus squamous cell carcinoma (ESCC gefitinibR) cell lines were constructed by using different gefitinib treatment in FLO‐1, KYAE‐1, TE‐8 and TE‐5 cell lines and confirmed by MTS50 and proliferation assays. Expression of linc01014 was overexpressed/silenced in FLO‐1 cells followed by gefitinib treatment, and then, the apoptosis‐associated markers Bax and Bcl‐2, and PI3KCA in PI3K signalling pathway were determined using Western blotting. MST50 and morphology analyses showed that ESCC gefitinibR cell lines presented obvious gefitinib resistance than their parental ESCC cell lines. ESCC gefitinibR cell lines showed significantly higher proliferation abilities than their parental ESCC cell lines after treating with gefitinib. Overexpression of linc01014 significantly inhibited the apoptosis of FLO‐1 cells induced by gefitinib and silencing linc01014 obviously promoted the apoptosis of FLO‐1 cells induced by gefitinib. Silencing linc01014 could significantly increase the gefitinib chemotherapy sensitivity of oesophagus cancer via PI3K‐AKT‐mTOR signalling pathway.     

MicroRNA‐20a‐5p contributes to hepatic glycogen synthesis through targeting p63 to regulate p53 and PTEN expression

Recently, it is implicated that aberrant expression of microRNAs (miRs) is associated with insulin resistance. However, the role of miR‐17 family in hepatic insulin resistance and its underlying mechanisms remain unknown. In this study, we provided mechanistic insight into the effects of miR‐20a‐5p, a member of miR‐17 family, on the regulation of AKT/GSK pathway and glycogenesis in hepatocytes. MiR‐20a‐5p was down‐regulated in the liver of db/db mice, and NCTC1469 cells and Hep1‐6 cells treated with high glucose, accompanied by reduced glycogen content and impaired insulin signalling. Notably, inhibition of miR‐20a‐5p significantly reduced glycogen synthesis and AKT/GSK activation, whereas overexpression of miR‐20a‐5p led to elevated glycogenesis and activated AKT/GSK signalling pathway. In addition, miR‐20a‐5p mimic could reverse high glucose‐induced impaired glycogenesis and AKT/GSK activation in NCTC1469 and Hep1‐6 cells. P63 was identified as a target of miR‐20a‐5p by bioinformatics analysis and luciferase reporter assay. Knockdown of p63 in the NCTC1469 cells and the Hep1‐6 cells by transfecting with siRNA targeting p63 could increase glycogen content and reverse miR‐20a‐5p inhibition‐induced reduced glycogenesis and activation of AKT and GSK, suggesting that p63 participated in miR‐20a‐5p‐mediated glycogenesis in hepatocytes. Moreover, our results indicate that p63 might directly bind to p53, thereby regulating PTEN expression and in turn participating in glycogenesis. In conclusion, we found novel evidence suggesting that as a member of miR‐17 family, miR‐20a‐5p contributes to hepatic glycogen synthesis through targeting p63 to regulate p53 and PTEN expression.     

LncRNA MEG3 inhibits the progression of prostate cancer by modulating miR‐9‐5p/QKI‐5 axis

This study was designed to detecting the influences of lncRNA MEG3 in prostate cancer. Aberrant lncRNAs expression profiles of prostate cancer were screened by microarray analysis. The qRT‐PCR and Western blot were employed to investigating the expression levels of lncRNA MEG3, miR‐9‐5p and QKI‐5. The luciferase reporter assay was utilized to testifying the interactions relationship among these molecules. Applying CCK‐8 assay, wound healing assay, transwell assay and flow cytometry in turn, the cell proliferation, migration and invasion abilities as well as apoptosis were measured respectively. LncRNA MEG3 was a down‐regulated lncRNA in prostate cancer tissues and cells and could inhibit the expression of miR‐9‐5p, whereas miR‐9‐5p down‐regulated QKI‐5 expression. Overexpressed MEG3 and QKI‐5 could decrease the abilities of proliferation, migration and invasion in prostate cancer cells effectively and increased the apoptosis rate. On the contrary, miR‐9‐5p mimics presented an opposite tendency in prostate cancer cells. Furthermore, MEG3 inhibited tumour growth and up‐regulated expression of QKI‐5 in vivo. LncRNA MEG3 was a down‐regulated lncRNA in prostate cancer and impacted the abilities of cell proliferation, migration and invasion, and cell apoptosis rate, this regulation relied on regulating miR‐9‐5p and its targeting gene QKI‐5.     

Knockdown of LINC01614 inhibits lung adenocarcinoma cell progression by up‐regulating miR‐217 and down‐regulating FOXP1

We tried to identify the function of LINC01614 in lung adenocarcinoma (LUAD) and reveal its underlying mechanisms. qRT‐PCR was applied to assess the expression of LINC016014 in LUAD tissues, noncancerous tissues and cells. Through colony formation assay, MTT assay and apoptosis analysis, we examined the variation of cell proliferation and apoptosis ability after silencing LINC01614. Moreover, the targeting interactions among LINC01614, miR‐217 and FOXP1 were validated via luciferase reporter assay, and then, we regulated the expression of miR‐217 and FOXP1 to ascertain their importance in cell proliferation and apoptosis. LINC01614 and FOXP1 were found to be up‐regulated in LUAD tumours and cells, whereas miR‐217 was down‐regulated. The experiment showed that target‐specific selectivity exists between LINC01614‐miR‐217 and miR‐217‐FOXP1 3′UTR. Furthermore, we disclosed that inhibition of LINC01614 could activate miR‐217, which subsequently restrained FOXP1. It was proved that LINC01614 promoted FOXP1 by inhibiting miR‐217, which ultimately stimulated the development of LUAD.     

miR‐200c/Bmi1 axis and epithelial–mesenchymal transition contribute to acquired resistance to BRAF inhibitor treatment

Resistance to BRAF inhibitors (BRAFi) is one of the major challenges for targeted therapies for BRAF‐mutant melanomas. However, little is known about the role of microRNAs in conferring BRAFi resistance. Herein, we demonstrate that miR‐200c expression is significantly reduced whereas miR‐200c target genes including Bmi1, Zeb2, Tubb3, ABCG5, and MDR1 are significantly increased in melanomas that acquired BRAFi resistance compared to pretreatment tumor biopsies. Similar changes were observed in BRAFi‐resistant melanoma cell lines. Overexpression of miR‐200c or knock‐down of Bmi1 in resistant melanoma cells restores their sensitivities to BRAFi, leading to deactivation of the PI3K/AKT and MAPK signaling cascades, and acquisition of epithelial–mesenchymal transition‐like phenotypes, including upregulation of E‐cadherin, downregulation of N‐cadherin, and ABCG5 and MDR1 expression. Conversely, knock‐down of miR‐200c or overexpression of Bmi1 in BRAFi‐sensitive melanoma cells activates the PI3K/AKT and MAPK pathways, upregulates N‐cadherin, ABCG5, and MDR1 expression, and downregulates E‐cadherin expression, leading to BRAFi resistance. Together, our data identify miR‐200c as a critical signaling node in BRAFi‐resistant melanomas impacting the MAPK and PI3K/AKT pathways, suggesting miR‐200c as a potential therapeutic target for overcoming acquired BRAFi resistance.     

Down‐regulated lncRNA SLC25A5‐AS1 facilitates cell growth and inhibits apoptosis via miR‐19a‐3p/PTEN/PI3K/AKT signalling pathway in gastric cancer

Mounting evidence has illustrated the vital roles of long non‐coding RNAs (lncRNAs in gastric cancer (GC). Nevertheless, the majority of their roles and mechanisms in GC are still largely unknown. In this study, we investigate the roles of lncRNA SLC25A5‐AS1 on tumourigenesis and explore its potential mechanisms in GC. The results showed that the expressions of SLC25A5‐AS1 in GC were significantly lower than that of adjacent normal tissues, which were significantly associated with tumour size, TNM stage and lymph node metastasis. Moreover, SLC25A5‐AS1 could inhibit GC cell proliferation, induce G1/G1 cell cycle arrest and cell apoptosis in vitro, as well as GC growth in vivo. Dual‐luciferase reporter assay confirmed the direct interaction between SLC25A5‐AS1 and miR‐19a‐3p, rescue experiment showed that co‐transfection miR‐19a‐3p mimics and pcDNA‐SLC25A5‐AS1 could partially restore the ability of GC cell proliferation and the inhibition of cell apoptosis. The mechanism analyses further found that SLC25A5‐AS1 might act as a competing endogenous RNAs (ceRNA), which was involved in the derepression of PTEN expression, a target gene of miR‐19a‐3p, and regulate malignant phenotype via PI3K/AKT signalling pathway in GC. Taken together, this study indicated that SLC25A5‐AS1 was down‐regulated in GC and functioned as a suppressor in the progression of GC. Moreover, it could act as a ceRNA to regulate cellular behaviours via miR‐19a‐3p/PTEN/PI3K/AKT signalling pathway. Thus, SLC25A5‐AS1 might be served as a potential target for cancer therapeutics in GC.     

LncRNA SNHG1 functions as a ceRNA to antagonize the effect of miR‐145a‐5p on the down‐regulation of NUAK1 in nasopharyngeal carcinoma cell

How lncRNA SNHG1 influences the aggressiveness of nasopharyngeal carcinoma cells as well as the underlying mechanism was studied. The lncRNA differences were analysed by GSE12452 gene microarray. The expression of SNHG1, MiR‐145‐5p and NUAK1 was identified by qRT‐PCR and western blot. Transfection was conducted to construct nasopharyngeal carcinoma cells with different expressions of SNHG1, miR‐145‐5p and NUAK1. Dual‐luciferase reporter assay was performed to explore the relationship between SNHG1, miR‐145‐5p and NUAK1. Wound‐healing assay and transwell invasion experiments were employed to study changes in cell migration capacity and cell invasion, respectively. Tumour xenografts were performed to observe lung metastasis of nude mice inoculated with transfected CNE cells. SNHG1 is highly expressed in nasopharyngeal carcinoma tissues and in cell lines. Down‐regulation of SNHG1 facilitated the expression of miR‐145‐5p and further suppressed the level of NAUK1 in CNE and HNE‐1 cells. Silencing of SNHG1, up‐regulation of miR‐145‐5p and inhibition of NAUK1 by relative transfection all attenuated the aggressiveness of CNE and HNE‐1 cells both in vivo and in vitro. Moreover, the impaired cell migration and invasion by SNHG1 siRNA could be rescued by cotransfection of miR‐145‐5p in CNE and HNE‐1 cells. LncRNA SNHG1 promoted the expression of NUAK1 by down‐regulating miR‐145‐5p and thus promoted the aggressiveness of nasopharyngeal carcinoma cells through AKT signalling pathway and induced epithelial‐mesenchymal transition (EMT).     

CiRS‐7 targeting miR‐7 modulates the progression of non‐small cell lung cancer in a manner dependent on NF‐κB signalling

The purpose of this study was to figure out the effect of ciRS‐7/miR‐7/NF‐κB axis on the development of non‐small cell lung cancer (NSCLC). In response, the expressions of ciRS‐7, miR‐7 and NF‐κB subunit (ie RELA) within NSCLC tissues and cell lines were determined with real‐time polymerase chain reaction (RT‐PCR) and Western blot. Moreover, the NSCLC cells were transfected with pcDNA3‐ciRS‐7‐ir, pcDNA3‐ciRS‐7, miR‐NC and miR‐7 mimic. Furthermore, the targeted relationships between ciRS‐7 and miR‐7, as well as between miR‐7 and RELA, were confirmed by luciferase reporter assay. The proliferation, migration and apoptosis of NSCLC cells were, successively, measured using CCK‐8 assay, wound‐healing assay and flow cytometry test. Consequently, ciRS‐7, miR‐7, histopathological grade, lymph node metastasis and histopathological stage could independently predict the prognosis of patients with NSCLC (all P < .05). Moreover, remarkably up‐regulated ciRS‐7 and RELA expressions, as along with down‐regulated miR‐7 expressions, were found within NSCLC tissues and cells in comparison with normal ones (P < .05). Besides, overexpressed ciRS‐7 and underexpressed miR‐7 were correlated with increased proliferation, migration and invasion, yet reduced apoptosis rate of NSCLC cells (P < .05). More than that, ciRS‐7 specifically targeted miR‐7 to reduce its expressions (P < .05). Ultimately, the NSCLC cells within miR‐7 + RELA group were observed with superior proliferative, migratory and invasive capabilities than those within miR‐7 group (P < .05), and RELA expression was also significantly modified by both ciRS‐7 and miR‐7 (P < .05). In conclusion, the ciRS‐7/miR‐7/NF‐kB axis could exert pronounced impacts on the proliferation, migration, invasion and apoptosis of NSCLC cells.     

Long noncoding RNA MALAT1 enhances the docetaxel resistance of prostate cancer cells via miR‐145‐5p‐mediated regulation of AKAP12

Our present work was aimed to study on the regulatory role of MALAT1/miR‐145‐5p/AKAP12 axis on docetaxel (DTX) sensitivity of prostate cancer (PCa) cells. The microarray data (GSE33455) to identify differentially expressed lncRNAs and mRNAs in DTX‐resistant PCa cell lines (DU‐145‐DTX and PC‐3‐DTX) was retrieved from the Gene Expression Omnibus (GEO) database. QRT‐PCR analysis was performed to measure MALAT1 expression in DTX‐sensitive and DTX‐resistant tissues/cells. The human DTX‐resistant cell lines DU145‐PTX and PC3‐DTX were established as in vitro cell models, and the expression of MALAT1, miR‐145‐5p and AKAP12 was manipulated in DTX‐sensitive and DTX‐resistant cells. Cell viability was examined using MTT assay and colony formation methods. Cell apoptosis was assessed by TUNEL staining. Cell migration and invasion was determined by scratch test (wound healing) and Transwell assay, respectively. Dual‐luciferase assay was applied to analyse the target relationship between lncRNA MALAT1 and miR‐145‐5p, as well as between miR‐145‐5p and AKAP12. Tumour xenograft study was undertaken to confirm the correlation of MALAT1/miR‐145‐5p/AKAP12 axis and DTX sensitivity of PCa cells in vivo. In this study, we firstly notified that the MALAT1 expression levels were up‐regulated in clinical DTX‐resistant PCa samples. Overexpressed MALAT1 promoted cell proliferation, migration and invasion but decreased cell apoptosis rate of PCa cells in spite of DTX treatment. We identified miR‐145‐5p as a target of MALAT1. MiR‐145‐5p overexpression in PC3‐DTX led to inhibited cell proliferation, migration and invasion as well as reduced chemoresistance to DTX, which was attenuated by MALAT1. Moreover, we determined that AKAP12 was a target of miR‐145‐5p, which significantly induced chemoresistance of PCa cells to DTX. Besides, it was proved that MALAT1 promoted tumour cell proliferation and enhanced DTX‐chemoresistance in vivo. There was an lncRNA MALAT1/miR‐145‐5p/AKAP12 axis involved in DTX resistance of PCa cells and provided a new thought for PCa therapy.     

LncRNA HOTAIR/miR‐613/c‐met axis modulated epithelial‐mesenchymal transition of retinoblastoma cells

Since lncRNAs could modulate neoplastic development by modulating downstream miRNAs and genes, this study was carried out to figure out the synthetic contribution of HOTAIR, miR‐613 and c‐met to viability, apoptosis and proliferation of retinoblastoma cells. Totally 276 retinoblastoma tissues and tumour‐adjacent tissues were collected, and human retinoblastoma cell lines (ie, Y79, HXO‐Rb44, SO‐Rb50 and WERI‐RB1) were also gathered. Moreover, transfections of pcDNA3.1‐HOTAIR, si‐HOTAIR, miR‐613 mimic, miR‐613 inhibitor, pcDNA3.1/c‐met were performed to evaluate the influence of HOTAIR, miR‐613 and c‐met on viability, apoptosis and epithelial‐mesenchymal transition (EMT) of retinoblastoma cells. Dual‐luciferase reporter gene assay was also arranged to confirm the targeted relationship between HOTAIR and miR‐613, as well as between miR‐613 and c‐met. Consequently, up‐regulated HOTAIR and down‐regulated miR‐613 expressions displayed associations with poor survival status of retinoblastoma patients (P < 0.05). Besides, inhibited HOTAIR and promoted miR‐613 elevated E‐cadherin expression, yet decreased Snail and Vimentin expressions (P < 0.05). Simultaneously, cell proliferation and cell viability were also less‐motivated (P < 0.05). Nonetheless, c‐met prohibited the functioning of miR‐613, resulting in promoted cell proliferation and viability, along with inhibited cell apoptosis (P < 0.05). Finally, HOTAIR was verified to directly target miR‐613, and c‐met was the direct target gene of miR‐613 (P < 0.05). In conclusion, the role of lncRNA HOTAIR/miR‐613/c‐met signalling axis in modulating retinoblastoma cells’ viability, apoptosis and expressions of EMT‐specific proteins might provide evidences for developing appropriate diagnostic and treatment strategies for retinoblastoma.     

Circ‐AKT3 inhibits the accumulation of extracellular matrix of mesangial cells in diabetic nephropathy via modulating miR‐296‐3p/E‐cadherin signals

Diabetic nephropathy is a leading cause of end‐stage renal disease globally. The vital role of circular RNAs (circRNAs) has been reported in diabetic nephropathy progression, but the molecular mechanism linking diabetic nephropathy to circRNAs remains elusive. In this study, we investigated the significant function of circ‐AKT3/miR‐296‐3p/E‐cadherin regulatory network on the extracellular matrix accumulation in mesangial cells in diabetic nephropathy. The expression of circ‐AKT3 and fibrosis‐associated proteins, including fibronectin, collagen type I and collagen type IV, was assessed via RT‐PCR and Western blot analysis in diabetic nephropathy animal model and mouse mesangial SV40‐MES13 cells. Luciferase reporter assays were used to investigate interactions among E‐cadherin, circ‐AKT3 and miR‐296‐3p in mouse mesangial SV40‐MES13 cells. Cell apoptosis was evaluated via flow cytometry. The level of circ‐AKT3 was significantly lower in diabetic nephropathy mice model group and mouse mesangial SV40‐MES13 cells treated with high‐concentration (25 mmol/L) glucose. In addition, circ‐AKT3 overexpression inhibited the level of fibrosis‐associated protein, such as fibronectin, collagen type I and collagen type IV. Circ‐AKT3 overexpression also inhibited the apoptosis of mouse mesangial SV40‐MES13 cells treated with high glucose. Luciferase reporter assay and bioinformatics tools identified that circ‐AKT3 could act as a sponge of miR‐296‐3p and E‐cadherin was the miR‐296‐3p direct target. Moreover, circ‐AKT3/miR‐296‐3p/E‐cadherin modulated the extracellular matrix of mouse mesangial cells in high‐concentration (25 mmol/L) glucose, inhibiting the synthesis of related extracellular matrix protein. In conclusion, circ‐AKT3 inhibited the extracellular matrix accumulation in diabetic nephropathy mesangial cells through modulating miR‐296‐3p/E‐cadherin signals, which might offer novel potential opportunities for clinical diagnosis targets and therapeutic biomarkers for diabetic nephropathy.     

MiR‐376a suppresses the proliferation and invasion of non‐small‐cell lung cancer by targeting c‐Myc

It has been reported that miR‐376a is involved in the formation and progression of several types of cancer. However, the expression and function of miR‐376a is still unknown in non‐small cell lung carcinomas (NSCLC). In this study, the expression of miR‐376a in NSCLC tissues and cell lines were examined by real‐time PCR, the effects of miR‐376a on cell proliferation, apoptosis and invasion were evaluated in vitro. Luciferase reporter assay was performed to identify the targets of miR‐376a. The results showed that miR‐376a was significantly downregulated in NSCLC tissues and cell lines. Restoration of miR‐376a in NSCLC cell line A549 significantly inhibited cell proliferation, increased cell apoptosis and suppressed cell invasion, compared with control‐transfected A549 cells. Luciferase reporter assay showed that c‐Myc, an oncogene that regulating cell survival, angiogenesis and metastasis, was a direct target of miR‐376a. Over‐expression of miR‐376a decreased the mRNA and protein levels of c‐Myc in A549 cells. In addition, upregulation of c‐Myc inhibited miR‐376a‐induced inhibition of cell proliferation and invasion in A549 cells. Therefore, our results indicate a tumor suppressor role of miR‐376a in NSCLC by targeting c‐Myc. miR‐376a may be a promising therapeutic target for NSCLC.     

Circular RNA ciRS‐7 promotes tube formation in microvascular endothelial cells through downregulation of miR‐26a‐5p

Atherosclerosis is one of the most common and crucial heart diseases involving the heart and brain. At present, atherosclerosis and its major complications comprise the leading causes of death worldwide. Our purpose was to identify the role of ciRS‐7 in atherosclerosis. Tubulogenesis of HMEC‐1 cell was evaluated utilizing tube formation assay. Cell Counting Kit‐8 assay and flow cytometry were utilized to test viability and apoptosis. Migration assay was utilized to determine the migration capacity of experimental cells. Western blot was applied to examine apoptosis and tube formation‐associated protein expression. In addition, the above experiments were repeated when silencing ciRS‐7, overexpressing ciRS‐7, and upregulating miR‐26a‐5p. HMEC‐1 cells formed tube‐like structures over time. Silencing ciRS‐7 suppressed viability, migration, and tube formation but promoted apoptosis. Oppositely, overexpressing ciRS‐7 reversed the effect in HMEC‐1 cells. miR‐26a‐5p expression was elevated by silencing ciRS‐7 and reduced by overexpressing ciRS‐7. Moreover, overexpressing ciRS‐7 facilitated viability, migration, and tube formation via upregulating miR‐26a‐5p. Conclusively, overexpressing ciRS‐7 mobilized phosphoinositide 3‐kinase/protein kinase B (PI3K/AKT) pathway and suppressed c‐Jun N‐terminal kinase (JNK)/p38 pathway. ciRS‐7 exerted influence on apoptosis, viability, migration, and tube formation through mediating PI3K/AKT and JNK/p38 pathways by miR‐26a‐5p downregulation in HMEC‐1 cells.     

MicroRNA‐130b targets PTEN to induce resistance to cisplatin in lung cancer cells by activating Wnt/β‐catenin pathway

More and more studies indicate the relevance of miRNAs in inducing certain drug resistance. Our study aimed to investigate whether microRNA‐130b‐3p (miR‐130b) mediates the chemoresistance as well as proliferation of lung cancer (LC) cells. MTS assay and apoptosis analysis were conducted to determine cell proliferation and apoptosis, respectively. Binding sites were identified using a luciferase reporter system, whereas mRNA and protein expression of target genes was determined by RT‐PCR and immunoblot, respectively. Mouse xenograft model was used to evaluate the role of miR‐130b in cisplatin resistance in vivo. The rising level of miR‐130b in cisplatin resistance LC cell lines (A549/CR and H446/CR ) versus its parental cell lines, indicated its crucial relevance for LC biology. We identified PTEN as miR‐130b's major target and inversely correlated with miR‐130b expression in LC. Moreover, excessive miR‐130b expression promoted drug resistance and proliferation, decreased apoptosis of A549 cells. Suppression of miR‐130b enhanced drug cytotoxicity and reduced proliferation of A549/CR cells both internally and externally. Particularly, miR‐130b mediated Wnt/β‐catenin signalling pathway activities, chemoresistance and proliferation in LC cell, which was partially blocked following knockdown of PTEN. These findings suggest that miR‐130b targets PTEN to mediate chemoresistance, proliferation, and apoptosis via Wnt/β‐catenin pathway. The rising level of miR‐130b in cisplatin resistance LC cell lines (A549/CR and H446/CR) versus its parental cell lines, indicated its crucial relevance for LC biology. Moreover, excessive miR‐130b expression promoted drug resistance and proliferation, decreased apoptosis of A549 cells. These findings suggest that miR‐130b targets PTEN to mediate chemoresistance, proliferation, and apoptosis via Wnt/β‐catenin pathway.     

MiR‐3116 sensitizes glioma cells to temozolomide by targeting FGFR1 and regulating the FGFR1/PI3K/AKT pathway

Glioma is a brain tumour that is often diagnosed, and temozolomide (TMZ) is a common chemotherapeutic drug used in glioma. Yet, resistance to TMZ is a chief hurdle towards curing the malignancy. The current work explores the pathways and involvement of miR‐3116 in the TMZ resistance. miR‐3116 and FGFR1 mRNA were quantified by real‐time PCR in malignant samples and cell lines. Appropriate assays were designed for apoptosis, viability, the ability to form colonies and reporter assays to study the effects of the miR‐3116 or FGFR1. The involvement of PI3K/AKT signalling was assessed using Western blotting. Tumorigenesis was evaluated in an appropriate xenograft mouse model in vivo. This work revealed that the levels of miR‐3116 dipped in samples resistant to TMZ, while increased miR‐3116 caused an inhibition of the tumour features mentioned above to hence augment TMZ sensitivity. miR‐3116 was found to target FGFR1. When FGFR1 was overexpressed, resistance to TMZ was augmented and reversed the sensitivity caused by miR‐3116. Our findings further confirmed PI3K/AKT signalling pathway is involved in this action. In conclusion, miR‐3116 sensitizes glioma cells to TMZ through FGFR1 downregulation and the PI3K/AKT pathway inactivation. Our results provide a strategy to overcome TMZ resistance in glioma treatment.