Chr9∶52042693|52047844⁃miR⁃351⁃5p⁃Pten Is Involved in the Development and Progression of Obstructive Sleep Apnea Syndrome through a ceRNA Mechanism / 中国生物化学与分子生物学报

Intermittent hypoxia (IH) is an important pathophysiological feature of obstructive sleep apnea (OSA), but its molecular mechanism is still unclear. We aim to investigate the role of endogenous competing endogenous RNA (ceRNA) regulatory network in the development of IH in OSA rats. An intermittent hypoxic rat model of OSA was constructed by hypoxic and reoxygenation cycles. CircRNAs and mRNAs were detected in rat bronchial tissues, and 230 up-regulated and 181 down-regulated circRNAs and 1238 up-regulated and 608 down-regulated mRNAs were analyzed and screened. The results of Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of the differential circRNAs and mRNAs suggested that they were mainly associated with metabolic pathways and PI3K-Akt signaling pathways. The key circRNAs (the top six circRNAs with the largest differences) were further validated by quantitative real-time polymerase chain reaction (qRT-PCR), chr9:52042693| 52047844 and chr4: 64889575|64899587 were expressed in bronchial tissues consistent with the sequencing results, which were used to further construct the ceRNA regulatory network. Four potential ceRNA regulatory networks were identified by TargetScan and miRanda database, combined with the results of differential circRNA and mRNA. The expression of molecules in the four potential ceRNA regulatory networks was detected by qRT-PCR in bronchial and lung tissues, and the results suggested that the expression of this regulatory network, chr9:52042693|52047844-miR-351-5p-Pten, was consistent with the sequencing results. The findings indicate that chr9:52042693 | 52047844-miR-351-5p-Pten may be involved in the development and progression of obstructive sleep apnea syndrome through a ceRNA mechanism.

Circular RNA ame_circ_000115 regulates expression of genes in larval gusts of Apis mellifera ligustica stressed by Ascosphaera apis / 生物工程学报

Circular RNAs (circRNAs) are a new class of non-coding RNAs, which have been confirmed to regulate insect gene expression and immune response through multiple manners such as competing endogenous RNA (ceRNA) regulatory network. Currently, function of circRNA in honey bee immune response remains unclear. In this study, PCR and Sanger sequencing were performed to validate the back splicing (BS) site of ame_circ_000115 (in short ac115). RT-qPCR was used to detect the expression profile of ac115 in larval guts of Apis mellifera ligustica stressed by Ascosphaera apis. Dual-luciferase reporter gene assay was conducted to verify the binding relationship between ac115 and ame-miR-13b. Interference of ac115 in larval guts was carried out by feeding specific siRNA, followed by determination of the effect of ac115 interference on expression of six genes relevant to host immune response. The results confirmed the existence of BS site within ac115. Compared with the un-inoculated group, the expression of ac115 in 4-day-old larval gut of the A. apis-inoculated group was up-regulated with extreme significance (P < 0.000 1), while that in 5- and 6-day-old larval guts were significantly up-regulated (P < 0.05). The brightness of specific band for ac115 in 4-, 5- and 6-day-old larval guts of the siRNA-circ_000115-fed group gradually became weak, whereas that of the siRNA-scrambl-fed group was pretty high without obvious variation. Compared with that of the siRNA-scramble-fed group, the expression of ac115 in 4-day-old larval gut of the siRNA-circ_000115-fed group was significantly down-regulated (P < 0.05), whereas that of the 5- and 6-day-old larval guts were down-regulated with extreme significance (P < 0.001). Ame-miR-13b was truly existed and expressed in A. m. ligustica larval guts, and there was true binding relationship between ac115 and ame-miR-13b. Compared with that of the siRNA-scramble-fed group, the expression of antimicrobial peptide genes hymenoptaecin and abaecin in 6-day-old larval gut of the siRNA-circ_000115-fed group was significantly up-regulated (P < 0.05), while that of ecdysone receptor (Ecr) was down-regulated with extreme significance (P < 0.01). These results indicate that ac115 is truly expressed in A. m. ligustica larval guts, BS site truly exists within ac115, and effective interference of ac115 in A. m. ligustica larval guts can be achieved via feeding siRNA. Moreover, ac115 potentially regulates Ecr expression through adsorption of ame-miR-13b and expression of hymenoptaecin and abaecin using a non-ceRNA manner, further participating in host stress-response.

Effects of Competing Endogenous RNA on Proliferation and Differentiation of Skeletal Muscle Cells / 中国生物化学与分子生物学报

The skeletal muscle is an important part of the animal body, which is closely related to body movement, energy consumption, production performance and its development process is regulated by many factors. The molecular mechanism of skeletal muscle growth and development is not only closely related to animal husbandry production, but also provides a theoretical basis for the treatment of muscle dis-eases, such as amyotrophic muscular dystrophy, and so on. Non-coding RNA (ncRNA) is a kind of RNA without coding ability, in which circular RNA (circRNA), long non-coding RNA (lncRNA) and micro RNA (miRNA) have been proved to play an important role in the development of skeletal muscles. miR-NA can specifically bind to the 3′ untranslated region (3′ UTR) of the target gene through its seed se-quence, so as to inhibit the translation process of the target gene or directly degrade the target gene, and become a prominent participant in a variety of biological processes. Studies have shown that circRNA, ln-cRNA, pseudogenes and mRNA with miRNA response elements (MRE) can competitively combine miRNA to regulate gene expression, forming a competing endogenous RNA (ceRNA) regulatory network model. This hypothesis subverts the previous concept of unidirectional regulation of target genes by miRNA, and endows them with new biological functions in transcriptome, which is of great biological significance. In recent years, it has been found that ceRNA plays an important regulatory role in the growth and development of skeletal muscles. This paper reviews the role of miRNA, lncRNA and circRNA in the proliferation and differentiation of animal skeletal muscle cells, which is an important member of ceRNA mechanism, in order to broaden the research direction of skeletal muscle mechanism and provide new ideas for the development of animal husbandry and the treatment of muscle diseases.

lncRNA LINC00308 acts as a competing endogenous RNA to promote the proliferation and invasion of prostate cancer PC3 cell through regulating the miR-361-5p/TRIP13 axis / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To investigate the effect of long non coding RNA (lncRNA) LINC00308 on proliferation, invasion and migration of prostate cancer cells and its related mechanism. Methods: lncRNAs and mRNAs differentially expressed in prostate cancer tissues and adjacent control tissues were screened by gene chip, and LINC00308 and TRIP13 (thyroid hormone receptor interactor13) were identified as the research objects. The effects of LINC00308 on the proliferation, invasion and migration of prostate cancer cells were detected by MTT assay, plate cloning, Transwell and scratch test. The above effects were verified in nude mice xenografts. The effect of LINC00308 on expression of TRIP13 in tumor tissues and cancer cells was detected by Western blotting and immunohistochemistry. Bioinformatics analysis, RIP (RNA immunoprecipitation), qPCR and Double luciferase gene reporter experiments were used to predict and explore the interaction mechanism between miR-361-5p and LINC00308 as well as TRIP13, and plate cloning and Transwell invasion test were used to verify the biological behaviors of cancer cells. Results: Both the microarray results and qPCR confirmed that the expressions of LINC00308 (P<0.01) and TRIP13 (P<0.05) were abnormally high in prostate cancer tissues and four cell lines; cell function test results showed that overexpression of LINC00308 could promote the proliferation, invasion and migration of prostate cancer PC3 cells (all P<0.05), while down-regulation of LINC00308 in prostate cancer cells had the opposite effect. In nude mice. LINC00308 could promote the tumorigenesis of prostate cancer cells in vivo, and increase the expression of TRIP13 both in vivo and in vitro (P<0.05). Bioinformatics analysis, RIP, qPCR and Double luciferase gene reporter results confirmed that miR-361-5p could bind to 3'-UTR of LINC00308 and TRIP13 respectively, and LINC00308 could act as a competing endogenous RNA (ceRNA) by sponging miR-361-5p to regulate the expression of TRIP13. In addition, MTT, plate cloning and Transwell assay confirmed the regulatory interaction among LINC00308 miR-361-5p and TRIP13 from the levels of proliferation, colony formation and invasion in cancer cells. Conclusion: LINC00308, which is abnormally highly expressed in prostate cancer tissues and cells, can inhibit the expression of miR-361-5p and enhance the expression of TRIP13 by exerting its ceRNA function, thus promoting the proliferation, invasion and migration of prostate cancer.

Screening prognosis-related molecules in hepatocellular carcinoma by competing endogenous RNAs regulatory network / 中国肿瘤生物治疗杂志

@#[ [Abstract] ] Objective: To identify the molecules related to the occurrence, development and prognosis of hepatocellular carcinoma (HCC) by constructing ceRNA regulatory network of HCC. Methods: Data of HCC transcription group were downloaded from TCGA database. We processed the original data into expression matrix of mRNA, lncRNA and miRNA via Perl language. DGEs of RNA and microRNA were extracted and analyzed from the“Edge”package of R language with the threshold of (|log FC|>2.0 and P<0.01). Through the database comparison, the relationship pairs of different lncRNAs-different miRNAs, different miRNAs-different mRNAs were obtained, and then imported them into the Cytoscape software to construct the ceRNA regulatory network diagram. The survival data of three DGEs were collected and analyzed by“survival”package and Kaplan Meier plotter analysis software. The survival curves were drawn and the genes were obtained by survival analysis. Results: The lncRNArelated ceRNA regulatory network of HCC was successfully constructed. Three regulatory pairs of lncRNA-miRNA-mRNA were obtained by analyzing the interaction and regulatory relationship between DGEs via ceRNA network. Among them, one regulatory pathway (CCDC26-hsa-mir-141-EPHA2) was in accordance with ceRNA theory. The prognostic analyses showed that the survival rate of patients with high expression of 14 mRNAs was lower than those with low expression, which could be used as biomarkers of adverse prognosis of HCC. The survival rate of patients with low expression of 1 lncRNA (TSPEAR-AS1) and 2 mRNA (CPEB3 and PROK2) was lower than those with high expression, which may be the protective gene of HCC. Conclusions:Through screening of HCC lncRNA related ceRNA regulatory network,14mRNAswithhighexpressionmaybetherelevant molecules related to poor prognosisofHCC,while1lncRNAand2 mRNAs with low expression may be the molecules related to good prognosis of HCC, providing reference for HCC treatment and prognosis evaluation.

Bioinformatics analysis of differentially expressed genes in ischemic cardiomyopathy / 上海交通大学学报（医学版）

Objective: To screen differentially expressed genes in ischemic cardiomyopathy (ICM) by bioinformatics analysis and construct the regulatory network of competing endogenous RNA (ceRNA). Methods: Data sets were downloaded from Gene Expression Omnibus (GEO) database to screen the differentially expressed mRNAs and lncRNAs between normal samples and ICM ones. Then, GO (Gene Ontology) analysis and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis were performed. The differentially expressed mRNAs and lncRNAs were analyzed to predict related miRNAs by bioinformatics methods, and then to construct ceRNA regulatory network. Results: GO analysis and KEGG pathways analysis results showed that the differentially expressed genes were enriched in the pathways such as metabolic pathway, oxidative phosphorylation, and extracellular matrix receptor interaction, and they were closely related to the functions such as endoplasmic reticulum stress, fibrosis, collagen catabolic process, and inflammatory response. A ceRNA regulatory network containing 26 mRNAs, 2 lncRNAs and 15 miRNAs was constructed. Conclusion: The bioinformatics method can be used to analyze the differentially expressed genes of ICM, and the regulatory network of ceRNA of ICM has been successfully constructed.