Exosome-mediated transfer of miR-1260b promotes cell invasion through Wnt/ß-catenin signaling pathway in lung adenocarcinoma.

Increasing evidence confirms that exosome-mediated transfer of microRNAs can influence cancer progression including tumor cell invasion, cell proliferation, and drug resistance via cell-cell communication. However, the potential role of exosomal-miR-1260b in lung adenocarcinoma (LAC) remains poorly understood. Thus, this study focused on investigating the function of exosomal-miR-1260b on cell invasion. Exosomal-miR-1260b was found to be higher in plasma of patients with LAC than that of healthy persons via quantitative real-time polymerase chain reaction assay. The sensitivity and specificity of exosomal-miR-1260b (cutoff point: 2.027) were 72% and 86%, and area under the curve of 0.845 (95% CI = 0.772-0.922). Elevated expression of miR-1260b in LAC tissues was positively correlated with exosomal-miR-1260b in plasma (r = .642, p < .05). Furthermore, ceramide biosynthesis regulated exosomal-miR-1260b secretion. Exosome-mediated transfer of miR-1260b promoted A549 cell invasion and was still functional inside A549 cells. Moreover, exosomal-miR-1260b regulated Wnt/ß-catenin signaling pathway by inhibiting sFRP1 and Smad4. This study identified a new regulation mechanism involving in cell invasion by exosome-mediated tumor-cell-to-tumor-cell communication. Targeting exosome-microRNAs may provide new insights into the diagnosis and treatment of LAC.

The long non-coding RNA AK001796 contributes to tumor growth via regulating expression of p53 in esophageal squamous cell carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is one of the prevalent and deadly cancers worldwide, especially in China. Considering the poor prognosis of ESCC, the aim of this study is to dissect the effects of long non-coding RNA (lncRNA) AK001796 on cell proliferation and cell cycle in vitro and tumorigenicity in vivo, providing therapeutic targets for ESCC. METHODS: We conducted quantitative real time PCR to detect the expression level of lncRNA AK001796 in human ESCC tumor and adjacent non-tumor tissues, and analyzed the correlation between lncRNA AK001796 expression and clinicopathologic feature of ESCC patients. Then we knocked down the expression of lncRNA AK001796 in human ESCC cell lines Eca-109 and TE-1, and next inspected cell cycle and apoptosis condition in these cells using flow cytometry. Subsequently, we used CCK-8 assay to test proliferation ability of the lncRNA AK001796-silenced ESCC cells, and the MDM2/p53 signaling pathway in these cells was analyzed by western blot analysis. At last, the ESCC xenograft models were established to verify the role of lncRNA AK001796 on the occurrence and development of ESCC. RESULTS: In this study, we demonstrated that lncRNA AK001796 was significantly upregulated in ESCC tumor tissues compared to adjacent non-tumor tissues. Knockdown of lncRNA AK001796 inhibited ESCC cell growth, cell cycle, and tumor growth in a xenograft mouse model via regulating MDM2/p53 signal pathway. The expression of lncRNA AK001796 was positively correlated with MDM2 levels in human ESCC samples. CONCLUSIONS: Overall, lncRNA AK001796 regulates cell proliferation and cell cycle via modulating MDM2/p53 signaling in ESCC, which provides a new insight into the treatment targets for ESCC.Trial registration This study was registrated in the Ethics Committee of the First Affiliated Hospital of Nanjing Medical University (Trial registration: 2012-SR-127, Registered 20 January 2012).

CircP4HB regulates ferroptosis via SLC7A11-mediated glutathione synthesis in lung adenocarcinoma.

Background: Circular ribonucleic acids (circRNAs) play a key role in the development of different types of cancer. Ferroptosis is a type of programmed cell death that contributes to cancer progression. However, the role of circRNAs in lung adenocarcinoma (LUAD) ferroptosis remains unclear. Methods: The gene expression levels of circRNA P4HB (circP4HB), microRNA-1184 (miR-1184) and Solute carrier family 7 member 11 (Slc7a11), also known as Xct were detected using quantitative real-time polymerase chain reaction (qRT-PCR). Ferroptosis of established LUAD cells was induced by erastin. Cell viability was examined via Cell Counting Kit 8 assays. Ferroptosis was evaluated by malondialdehyde (MDA), Prostaglandin-endoperoxide Synthase 2 (Ptgs2), lipid reactive oxygen species (lipid ROS), and JC-1 detection. The mechanism of circP4HB/miR-1184/SLC7A11 was investigated by luciferase reporter assays, RNA immunoprecipitation, RNA pull-down, and western blot assays. A functional for circP4HB in vivo was determined using xenograft nude mice models. Results: CircP4HB expression levels were increased in LUAD. It triggered glutathione (GSH) synthesis and, therefore protected LUAD cells from ferroptosis induced by erastin. CircP4HB may function as a competing endogenous RNA by modulating miR-1184 to regulate SLC7A11. CircP4HB inhibited ferroptosis by regulating miR-1184/ SLC7A11-mediated GSH synthesis. In vivo, overexpression of circP4HB promoted tumor growth and inhibited ferroptosis. Conclusions: The circRNA, circP4HB acts as a novel ferroptosis suppressor in LUAD. Furthermore, circP4HB protects LUAD from ferroptosis via modulation of the miR-1184/SLC7A11 axis. Our findings identified circP4HB as a novel biomarker in LUAD and warrants further investigation in the early diagnosis and treatment of LUAD.

miR-550a-5p Functions as a Tumor Promoter by Targeting LIMD1 in Lung Adenocarcinoma.

Lung adenocarcinoma accounts for half of all lung cancer cases in most countries. Mounting evidence has demonstrated that microRNAs play important roles in cancer progression, and some of them can be identified as potential biomarkers. This study aimed to explore the role of miR-550a-5p, a lung adenocarcinoma-associated mature microRNA screened out from the TCGA database via R-studio and Perl, with abundant expression in samples and with 5-year survival prognosis difference, as well as having not been studied in lung cancer yet. Potential target genes were predicted by the online database. Gene ontology enrichment, pathway enrichment, protein-protein interaction network, and hub genes-microRNA network were constructed by FunRich, STRING database, and Cytoscape. Then, LIMD1, a known tumor suppressor gene reported by multiple articles, was found to have a negative correlation with miR-550a-5p. The expression of miR-550a-5p was up-regulated in tumor samples and tumor-associated cell lines. Its high expression was also correlated with tumor size. Cell line A549 treated with miR-550a-5p overexpression promoted tumor proliferation, while H1299 treated with miR-550a-5p knockdown showed the opposite result. Mechanically, miR-550a-5p negatively regulated LIMD1 by directly binding to its 3'-UTR validated by dual luciferase assay. In summary, a new potential prognostic and therapeutic biomarker, miR-550a-5p, has been identified by bioinformatics analysis and experimental validation in vitro and in vivo, which promotes lung adenocarcinoma by silencing a known suppressor oncogene LIMD1.

Correction: miR-1260b, mediated by YY1, activates KIT signaling by targeting SOCS6 to regulate cell proliferation and apoptosis in NSCLC.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Moxifloxacin suppresses airway inflammation and modulates expression of caveolin-1 and flotillin-1 in airway smooth muscle cells of asthmatic rats.

BACKGROUND: Moxifloxacin (MXF) possesses anti-inflammatory properties on asthmatic airway smooth muscle cells (ASMCs) beyond their antimicrobial effects, but the mechanisms are still unknown. This study was to investigate effects of MXF on expression of caveolin-1 (Cav-1) and flotillin-1 (FLOT1) in ASMCs in asthmatic rats. METHODS: ASMCs were collected from the airway and cultured in vitro. Cells from normal rats were treated with normal saline (Group N); cells from asthmatic rats were incubated with normal saline (Group A) or MXF (20 mg/L) (Group M); Cav-1 expression was up-regulated by transferring Cav-1 expressing lentivirus (Group L) and FLOT1 expression down-regulated by using siRNA in cells from asthmatic rats (Group S). The expressions of Cav-1, FLOT1 and p65 NF-κB were measured by Western blotting and quantificational real-time polymerase chain reaction (qRT-PCR), and interleukin-8 (IL-8) and eotaxin contents were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Compared with normal control, Cav-1 expression significantly decreased in asthmatic groups (P<0.01); MXF up-regulated Cav-1 expression in asthmatic groups (P<0.01). However, compared with normal control, the expression of FLOT1 and p65 NF-κB dramatically increased in asthmatic groups (P<0.01); MXF down-regulated the expression of FLOT1 and p65 NF-κB in asthmatic groups (P<0.01); meanwhile, the expressions of FLOT1 and p65 NF-κB decreased after up-regulation of Cav-1 expression in asthmatic groups (P=0.01). Compared with asthmatic groups, the IL-8 and eotaxin contents significantly decreased in MXF Groups, Cav-1 up-regulation asthmatic groups and FLOT1 down-regulation asthmatic groups (P<0.01). CONCLUSIONS: MXF can modulate the airway inflammation, upregulate Cav-1 expression, downregulate the expression of FLOT1 and p65 NF-κB in asthmatic rat ASMCs, which may be related to the anti-inflammatory effects of MXF in asthmatic ASMCs.

miR-1260b, mediated by YY1, activates KIT signaling by targeting SOCS6 to regulate cell proliferation and apoptosis in NSCLC.

Non-small cell lung cancer (NSCLC) is one of the most common aggressive malignancies. miRNAs have been identified as important biomarkers and regulators of NSCLC. However, the functional contributions of miR-1260b to NSCLC cell proliferation and apoptosis have not been studied. In this study, miR-1260b was upregulated in NSCLC plasma, tissues, and cell lines, and its high expression was correlated with tumor size and progression. Functionally, miR-1260b overexpression promoted cell proliferation and cell cycle, conversely inhibited cell apoptosis and senescence. Mechanically, miR-1260b negatively regulated SOCS6 by directly binding to its 3'-UTR. Furthermore, miR-1260b-mediated suppression of SOCS6 activated KIT signaling. Moreover, YY1 was an upstream regulator of miR-1260b. This study is the first to illustrate that miR-1260b, mediated by YY1, activates KIT signaling by targeting SOCS6 to regulate NSCLC cell proliferation and apoptosis, and is a potential biomarker and therapeutic target for NSCLC. In sum, our work provides new insights into the molecular mechanisms of NSCLC involved in cell proliferation and apoptosis.

Long noncoding RNA RP11-766N7.4 functions as a tumor suppressor by regulating epithelial-mesenchymal transition in esophageal squamous cell carcinoma.

BACKGROUND: Numerous studies have proved that long non-coding RNAs participate in the initiation and metastasis of various cancers including esophageal squamous cell carcinoma (ESCC). Recently, a novel long non-coding RNA RP11-766N7.4 was discovered in a variety of human tissues. However, its role in oncogenesis and tumor metastasis remains unknown. METHODS: To investigate the function of long noncoding RNA RP11-766N7.4 in ESCC, RT-qPCR was used to monitor the expression level of long non-coding RNA RP11-766N7.4 in ESCC cell lines and 50 paired ESCC tissues. Moreover, the association between long non-coding RNA RP11-766N7.4 expression level and clinicopathological characteristics as well as 5-year survival rate of ESCC patients was evaluated. Furthermore, function assays containing cell proliferation assay, flow cytometry, Colony Formation, wound healing assay and Transwell assays were conducted to investigate the role of long noncoding RNA RP11-766N7.4 in ESCC. Western blotting assay were used to explore the regulation mechanism. RESULTS: In this study, we found that long noncoding RNA RP11-766N7.4 was downregulated in ESCC tissues and cell lines and correlated with lymph node metastasis, tumor stage and survival rate. Results also revealed that long noncoding RNA RP11-766N7.4 had no significant effect on cell proliferation, cell cycle or cell apoptosis of ESCC cells. In addition, long noncoding RNA RP11-766N7.4 knockdown promoted cellular migration and invasion via inducing EMT process, and overexpression of long noncoding RNA RP11-766N7.4 inhibited cellular migration and invasion by suppressing EMT process. CONCLUSION: Our study suggested that long noncoding RNA RP11-766N7.4 acts as a tumor suppressor in ESCC carcinogenesis and metastasis, and may be a potential prognostic mark and a therapeutic target for ESCC.

Long non­coding RNA AK001796 contributes to cisplatin resistance of non­small cell lung cancer.

Cisplatin (DDP)­based chemotherapy is the most widely used therapy for non­small cell lung cancer (NSCLC). However, the existence of chemoresistance has become a major limitation in its efficacy. Long non­coding RNAs (lncRNAs) have been shown to be involved in chemotherapy drug resistance. The aim of the present study was to investigate the biological role of lncRNA AK001796 in cisplatin­resistant NSCLC A549/DDP cells. Reverse transcription­quantitative polymerase chain reaction (RT­qPCR) analysis was performed to monitor the differences in the expression of AK001796 in cisplatin-resistant (A549/DDP) cells and parental A549 cells. Cellular sensitivity to cisplatin and cell viability were examined using an MTT assay. Cell apoptosis and cell cycle distribution were measured using flow cytometry. The expression levels of cell cycle proteins cyclin C (CCNC), baculoviral IAP repeat containing 5 (BIRC5), cyclin­dependent kinase 1 (CDK1) and G2 and S phase­expressed 1 (GTSE1) were assessed using RT­qPCR and western blot analyses. It was found that the expression of AK001796 was increased in A549/DDP cells, compared with that in A549 cells. The knockdown of AK001796 by small interfering RNA reduced cellular cisplatin resistance and cell viability, and resulted in cell­cycle arrest, with a marked increase in the proportion of A549/DDP cells in the G0/G1 phase. By contrast, the knockdown of AK001796 increased the number of apoptotic cancer cells during cisplatin treatment. It was also shown that the knockdown of AK001796 positively induced the expression of cell apoptosis­associated factors, CCNC and BIRC5, and suppressed the expression of cell cycle­associated factors, CDK1 and GTSE5. Taken together, these findings indicated that lncRNA AK001796 increased the resistance of NSCLC cells to cisplatin through regulating cell apoptosis and cell proliferation, and thus provides an attractive therapeutic target for NSCLC.

Long non-coding RNA-Low Expression in Tumor inhibits the invasion and metastasis of esophageal squamous cell carcinoma by regulating p53 expression.

Long non-coding RNAs (lncRNAs) are involved in governing fundamental biological processes, and, in many lncRNAs, the expression level is altered and likely to have a functional role in tumorigenesis, including apoptosis, migration and invasion. The lncRNA­Low Expression in Tumor (LET), a recently identified lncRNA, was demonstrated to be downregulated in hepatocellular and gallbladder cancer. However, its role in esophageal squamous cell carcinoma (ESCC) requires investigation. The expression level of lncRNA­LET mRNA in primary ESCC and matched healthy tissues (48 cases) was determined by reverse transcription­quantitative polymerase chain reaction. In addition, the effects of lncRNA-LET on cell apoptosis were evaluated by flow cytometric analysis, the regulatory effect of lncRNA­LET on migration was detected using a wound healing assay and cellular invasion was analyzed by Matrigel­coated transwell assay. Furthermore, the effect of lncRNA­LET on cell proliferation was investigated by 5­ethynyl­2'-deoxyuridine cell proliferation assay and protein levels of lncRNA-LET targets were analyzed by western blotting. lncRNA-LET expression was decreased in primary ESCC tissues when compared with paired healthy tissues, and was identified to be associated with the clinical features. Overexpression of lncRNA­LET was observed to inhibit the migration and invasion of ESCC cells, and modulate p53 expression levels in human ESCC cell lines in vitro. These results establish that lncRNA-LET is significant in the regulation of tumor progression and metastasis, and serves as a tumor suppressor in, and therefore has therapeutic potential for, the treatment of human ESCC.

Long Noncoding RNA-LET Suppresses Tumor Growth and EMT in Lung Adenocarcinoma.

Recently, many studies showed that long noncoding RNAs (lncRNAs) are involved in tumor progression. It is reported that lncRNA-LET is downregulated and has antitumor effect on several types of cancer. This study focuses on the role of lncRNA-LET on lung adenocarcinoma (LAC) progression. RT-PCR results indicated that frequent downregulation of lncRNA-LET in LAC tissues was related to clinicopathologic factors. lncRNA-LET knockdown significantly promoted LAC cell proliferation, invasion, and migration while lncRNA-LET overexpression obviously inhibited LAC cell proliferation, invasion, and migration, indicating a tumor inhibition of lncRNA-LET in LAC progression. Besides, lncRNA-LET inhibited EMT and negatively regulated Wnt/ß-catenin pathway in part. Our study suggests that lncRNA-LET exhibits an important tumor-suppressive effect on LAC progression by inhibiting EMT and Wnt/ß-catenin pathway, which provides potential therapeutic targets for LAC.

An experimental model for induction of lung cancer in rats by Chlamydia pneumoniae.

OBJECTIVE: To assess induction effects of Chlamydia pneumoniae (Cpn) on lung cancer in rats. METHODS: A lung cancer animal model was developed through repeated intratracheal injection of Cpn (TW-183) into the lungs of rats, with or without exposure to benzo(a)pyrene (Bp). Cpn antibodies (Cpn-IgA, -IgG, and -IgM) in serum were measured by microimmunofluorescence. Cpn-DNA or Cpn-Ag of rat lung cancer was detected through polymerase chain reaction or enzyme-linked immunosorbent assay. RESULTS: The prevalence of Cpn infection was 72.9% (35/48) in the Cpn group and 76.7% (33/43) in the Cpn plus benzo(a)pyrene (Bp) group, with incidences of lung carcinomas in the two groups of 14.6% (7/48) and 44.2% (19/43), respectively (P-values 0.001 and <0.001 compared with normal controls). CONCLUSIONS: A rat model of lung carcinoma induced by Cpn infection was successfully established in the laboratory for future studies on the treatment, prevention, and mechanisms of the disease.