Circ_0006174 Upregulates IGF1R to Enhance Radioresistance and Tumorigenesis in Colorectal Cancer via miR-940 Suppression.

Colorectal cancer (CRC) is one of the most common malignancies all over the world. Increasing evidence has revealed that circular RNAs (circRNAs) are involved in the progression of CRC. In this study, we aimed to investigate the role and underlying mechanism of circ_0006174 in the development and radiosensitivity of CRC. Circ_0006174, microRNA-940 (miR-940), and insulin-like growth factor 1 receptor (IGF1R) expression levels were evaluated by real-time quantitative polymerase chain reaction (RT-qPCR). The radiosensitivity of cells also was assessed using colony formation assay. Besides, cell proliferation, apoptosis, migration, and invasion were detected by cell counting kit-8 (CCK-8), flow cytometry, and transwell assays. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were performed to verify the relationship between miR-940 and circ_0006174 or IGF1R. IGF1R protein level was examined using western blot. A xenograft tumor model was used to verify the function of circ_0006174 in CRC tumor growth in vivo. Circ_0006174 and IGF1R levels were elevated and miR-940 expression was decreased in CRC tissues and cells. Circ_0006174 knockdown enhanced the radiosensitivity of CRC cells by regulating cell proliferation, apoptosis, migration, and invasion in vitro. In mechanism, circ_0006174 served as a sponge for miR-940 to upregulate IGF1R expression. Moreover, circ_0006174 silencing suppressed CRC growth in vivo. Circ_0006174 boosts radioresistance of CRC cells at least partly through upregulating IGF1R expression by sponging miR-940, providing a novel theoretical basis for CRC therapy.

miR-940 modulates CD47 to suppress biological functions of lung adenocarcinoma cells.

OBJECTIVE: mir-940 and CD47 play regulatory and immunoregulatory roles in lung cancer. While previous study found that the expression of mir-940 decreased, associated with the increasing of CD47 in lung adenocarcinoma. However, their inherent correlations remain elusive. Herein, this experiment intends to search for the relevant molecular mechanisms regulating the biological function of non-small cell lung cancer. METHODS: The cancer and adjacent tissue samples were collected from 20 pairs of newly diagnosed non-small cell lung cancer patients without applying radiotherapy and chemotherapy. We performed immunohistochemistry containing 45 lung adenocarcinoma tissues to investigate the relationship between the clinicopathological features and CD47 expression. The expressions of mir-940 and CD47 were detected by real-time quantitative polymerase chain reaction (qRT-PCR). Lung epithelial and lung adenocarcinoma (A549, H1299, GLC-82, PC-9) cell lines were cultured to detect the expression of mir-940 and CD47 molecules in each cell line. According to the expression situation, 2 cell lines were selected for mimic and siRNA transfection, and the transfection efficiency was also verified by qRT-PCR and western blot. CCK-8, transwell migration, transwell invasion, and colony formation assays were used to detect the changes in biological functions of lung adenocarcinoma cells after transfection, such as enhanced proliferation, migration, invasion, and cloning. The changes of related protein molecules after transfection were detected by western blot. The dual-luciferase experiment verified the targeting regulation relationship between mir-940 and CD47. Finally, flow cytometry analysis of apoptosis and cell cycle were carried out to detect apoptosis cells and change phase of cell cycle distribution. RESULTS: CD47 expression was not associated with clinicopathologic factors in lung adenocarcinoma. The proliferation, migration, invasion, and cloning abilities of lung adenocarcinoma cells were weakened after transfection with mir-940 mimic and siRNA-CD47. Overexpression of CD47 could promote proliferation, migration, invasion, cloning abilities, reduce apoptosis rate and attenuate the antitumor effect of mir-940 on lung adenocarcinoma. Dual luciferase experiments confirmed that mir-940 can target CD47 molecules. CONCLUSION: mir-940 can inhibit the biological function of lung adenocarcinoma cells by targeting CD47.

Integrated analysis of high­throughput sequencing reveals the regulatory potential of hsa_circ_0035431 in HNSCC.

Circular RNAs (circRNAs) are molecular sponges that are involved in regulation of multiple types of cancer. The present study aimed to screen and explore the key circRNA/microRNA (miRNA or miR)/mRNA interactions in head and neck squamous cell carcinoma (HNSCC) using bioinformatics. A total of six pairs of cancerous and adjacent healthy tissue were obtained from patients with HNSCC and genome-wide transcriptional sequencing was performed. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were performed on differentially expressed genes (DEGs). Moreover, expression levels of DEGs were verified in HNSCC cells and tissues using reverse transcription-quantitative (RT-q)PCR. A molecular regulatory network consisting of three circRNAs, seven miRNAs and seven mRNAs was constructed, resulting in identification of two signaling axes, hsa_circ_0035431/hsa-miR-940/fucosyltransferase 6 (FUT6) and hsa_circ_0035431/hsa-miR-940/cingulin-like 1 (CGNL1). FUT6 and CGNL1 were downregulated in HNSCC compared with adjacent healthy tissue and the expression levels of these genes were associated with tumor stage. Low FUT6 and CGNL1 expression levels were associated with lower overall survival rate and progression-free intervals in HNSCC. RT-qPCR demonstrated that hsa_circ_0035431, FUT6 and CGNL1 were downregulated in HNSCC cells and tissue and hsa-miR-940 was upregulated. Notably, these results were consistent with those obtained using high-throughput sequencing. In conclusion, hsa_circ_0035431 may participate in regulation of FUT6 and CGNL1 expression by sponging hsa-miR-940, thus, impacting the occurrence, development and prognosis of HNSCC.

Unique microRNA expression profiles in plasmic exosomes from intrahepatic cholestasis of pregnancy.

BACKGROUND: Intrahepatic cholestasis of pregnancy (ICP) is strongly associated with an increased risk of adverse perinatal outcomes. Total bile acid (TBA) levels in the late second or third trimester are a major factor in the diagnosis. Here, we sought to establish the miRNA expression profile of plasm exosomes of ICP and identify possible biomarkers for the diagnosis of ICP. METHODS: This case-control study involved 14 ICP patients as the experimental group and 14 healthy pregnant women as the control group. Electron microscopy was used to observe the presence of exosomes in plasma. Nanosight and Western blotting of CD63 was used to assess exosome quality. Among them, three ICP patients and three controls were used for isolation plasmic exosome and preliminary miRNA array analysis. The Agilent miRNA array was utilized to dynamically monitor the miRNA expression in plasmic exosomes of included patients in the first trimester(T1), second trimester (T2), third trimester (T3), and delivery (T4). Then, Quantitative real-time Polymerase chain reaction was used to identify and validate differentially expressed miRNAs in plasma-derived exosomes. RESULTS: The expression levels of hsa-miR-940, hsa-miR-636, and hsa-miR-767-3p in plasma-derived exosomes of ICP patients were significantly higher than those of healthy pregnant women. Besides, these three miRNAs were also significantly up-regulated at the plasma, placental, and cellular levels (P < 0.05). The diagnostic accuracy of hsa-miR-940, hsa-miR-636, and hsa-miR-767-3p was further evaluated by the ROC curve, the area under the curve (AUC) values for each were 0.7591, 0.7727, and 0.8955, respectively. CONCLUSIONS: We identified three differentially expressed miRNAs in the plasma exosomes of ICP patients. Hence, hsa-miR-940, hsa-miR-636, and hsa-miR-767-3p may be potential biomarkers for enhancing the diagnosis and prognosis of ICP.

Depletion of circ_0006459 protects human brain microvascular endothelial cells from oxygen-glucose deprivation-induced damage through the miR-940/FOXJ2 pathway.

BACKGROUND: Multiple circular RNAs (circRNAs) play important roles in ischemic stroke. The present study aims to reveal the role and the mechanism of circ_0006459 in ischemic stroke. METHODS: Human brain microvascular endothelial cells (HBMECs) were treated with oxygen-glucose deprivation (OGD) to mimic an in vitro ischemic stroke model. RNA expression of circ_0006459, microRNA-940 (miR-940), and forkhead box J2 (FOXJ2) was detected by quantitative real-time polymerase chain reaction. Cell proliferation was analyzed by cell counting kit-8 (CCK-8) and 5-Ethynyl-29-deoxyuridine (EdU) assays. Cell apoptotic rate was quantified by flow cytometry analysis. The protein expression of proliferating cell nuclear antigen (PCNA), clusters of differentiation 6 (CDK6), BCL2-associated x protein (Bax), B-cell lymphoma 2 (Bcl2), interleukin-1ß (IL-1ß), IL-8, IL-18 and tumor necrosis factor-α (TNF-α) was analyzed by Western blotting. The regulatory relationships among circ_0006459, miR-940, and F 《》 OXJ2 were identified by dual-luciferase reporter assay, RNA immunoprecipitation assay, and RNA pull-down assay. RESULTS: Circ_0006459 and FOXJ2 expression were significantly upregulated, whereas miR-940 expression was downregulated in HBMECs after OGD. Circ_0006459 depletion assuaged OGD-induced inhibition in cell proliferation and promotion in cell apoptosis and inflammation in HBMECs. Circ_0006459 acted as a sponge for miR-940, and miR-940 targeted FOXJ2 in HBMECs. Besides, miR-940 silencing or FOXJ2 overexpression relieved circ_0006459 knockdown-induced promotion in cell proliferation and inhibition in cell apoptosis and inflammation in OGD-induced HBMECs. Further, circ_0006459 depletion decreased FOXJ2 protein expression by interacting with miR-940. CONCLUSION: Depletion of circ_0006459 protected human brain microvascular endothelial cells from oxygen-glucose deprivation-induced damage through miR-940/FOXJ2 pathway, providing a promising therapeutic target for ischemic stroke.

hsa_circ_0038382 upregulates T-box transcription factor 5 to inhibit keloid formation by interacting with miR-940.

BACKGROUND: A keloid is a benign fibroproliferative skin tumor whose formation is regulated by circular RNAs (circRNAs). However, the effect and regulatory mechanism of hsa_circ_0038382 on keloid formation have not been investigated. OBJECTIVES: This study aimed to identify the function and mechanism of hsa_circ_0038382 in keloid formation. MATERIAL AND METHODS: The expression levels of hsa_circ_0038382, microRNA-940 (miR-940) and T-box transcription factor 5 (TBX5) were measured using real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). After cell transfection of keloid fibroblasts, the effect of the hsa_circ_0038382/miR-940/TBX5 axis on keloid formation was assessed using cell function tools such as the cell counting kit-8 (CCK-8) assay, transwell migration assay and transwell invasion assay. The binding sites among hsa_circ_0038382, miR-940 and TBX5 were predicted with CircInteractome and TargetScan, and further identified using luciferase assays. RESULTS: The levels of hsa_circ_0038382 and TBX5 were reduced, whereas the level of miR-940 was elevated in keloid samples. Cell function experiments confirmed that hsa_circ_0038382 can inhibit keloid formation by suppressing proliferation, migration and invasion of keloid fibroblasts. Luciferase assays proved that hsa_circ_0038382 can absorb miR-940 to regulate TBX5 expression in keloids. Additionally, the overexpression of TBX5 restored the effect of hsa_circ_0038382 knockdown on keloid fibroblasts. CONCLUSIONS: This study suggests that hsa_circ_0038382 attenuates keloid formation by regulating the miR-940/TBX5 axis, which might provide a potential therapeutic target in the treatment of keloid formation.

Optimization of a method for the clinical detection of serum exosomal miR-940 as a potential biomarker of breast cancer.

Serum exosomal microRNAs (miRNAs) are potential biomarkers for tumor diagnosis. Clinically, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) can be used to determine the expression of exosomal miRNAs in the serum of breast cancer patients. The prerequisites for obtaining meaningful serum exosomal miRNA data of breast cancer patients include a suitable extraction method for exosomes and RT-qPCR data standardized by internal reference genes. However, the appropriate methods for the extraction of exosomes and the applicability of reference genes for analyzing exosomal miRNAs in breast cancer patients remain to be studied. This study compared the effects of three exosome extraction methods as well as the expression of exosomal miRNA in different initial serum amounts and at different serum states to identify the selection of the best method for serum exosome extraction. Five candidate reference genes including miR-16, miR-484, miR-1228, miR-191 and miR-423 for standardizing serum exosomal miRNAs were screened using five algorithms and were used for the quantification of serum exosomal miR-940. Significant downregulation of serum exosomal miR-940 expression in breast cancer was detected using miR-191 and miR-1228, whereas no significant down or up regulation was observed with miR-484, miR-423 and miR-16. Previous studies have shown that the expression level of miR-940 is downregulated in breast cancer tissues. The absolute quantitative results showed that miR-940 was significantly downregulated in breast cancer serum exosomes, which was consistent with the results from the analysis using miR-191 or miR-1228 as reference genes. Therefore, miR-191 and miR-1228 could serve as reference genes for the relative quantification of serum exosomal miRNAs. This finding indicated the importance of rigorously evaluating the stability of reference genes and standardization for serum exosomal miRNA expression. Moreover, the level of serum exosomal miR-940 in breast cancer could reflect the presence of lymph node metastasis and the status of HER2/neu, which indicates its potential as a biomarker for breast cancer metastasis. In summary, an optimized protocol for the detection of serum exosomal miR-940 as a breast cancer marker was preliminarily established.

The circular RNA circ_0099630/miR-940/receptor-associated factor 6 regulation cascade modulates the pathogenesis of periodontitis.

Background/purpose: Periodontitis is one of the highly prevalent chronic inflammatory conditions in adults. The importance of circular RNAs (circRNAs) in the regulation of inflammation has been gradually reported in recent years, but the role of circRNA circ_0099630 in periodontitis has not been reported. Materials and methods: The contents of circ_0099630, microRNA-940 (miR-940) and tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) were measured using quantitative real-time polymerase chain reaction (qRT-PCR) or Western blot. Inflammatory factor secretion, cell proliferation, and apoptosis were analyzed under the application of Enzyme-linked immunosorbent assay (ELISA), Cell Counting Kit-8 (CCK8), 5-ethynyl-2'-deoxyuridine (EdU) and flow cytometry, respectively. The Western blot also analyzed the phosphorylation levels of RELA proto-oncogene (P65) and IkappaBalpha (IκBα), key molecules of the nuclear factor kappa-B (NF-κB) pathway. The relationship between miR-940 and circ_0099630 or TRAF6 was verified by luciferase reporter system and RNA immunoprecipitation (RIP) assay. Results: Higher abundance of circ_0099630 and TRAF6 and lower miR-940 expression were observed in periodontitis, and circ_0099630 knockdown attenuated the damage of human PDL cells (PDLCs) induced by lipopolysaccharides (LPS). The relationship between miR-940 and circ_0099630 or TRAF6 was evidenced, while miR-940 downregulation diminished the repair effect of si-circ_0099630 on overexpression LPS-induced damage in PDLCs. Similarly, TRAF6 upregulation impaired the mitigating effect of miR-940 overexpression on LPS-induced injury in PDLCs. Circ_0099630 silencing evidently curbed the phosphorylation levels of P65 and IκBα and thus attenuating the inflammatory response by acting on the miR-940/TRAF6 axis. Conclusion: Silencing circ_0099630 alleviates LPS-induced periodontal ligament cell injury via targeting miR-940/TRAF6/NF-κB in periodontitis.

Depletion of circ_0128846 ameliorates interleukin-1ß-induced human chondrocyte apoptosis and inflammation through the miR-940/PTPN12 pathway.

BACKGROUND: Previous evidence has suggested that circular RNA (circRNA) is abnormally expressed in osteoarthritis (OA). However, the underlying mechanism of circRNA in OA progression remains unclear. The study aims to reveal the mechanism of circ_0128846 regulating OA. METHODS: Human chondrocytes (C28/I2 cells) were treated with interleukin-1ß (IL-1ß) to mimic an OA cell model. The expression levels of circ_0128846, miR-940 and protein tyrosine phosphatase 12 (PTPN12) were detected by qRT-PCR. Protein expression was checked by Western blotting. Cell viability, proliferation, and apoptosis were analyzed by a cell counting kit-8 assay, 5-Ethynyl-2'-deoxyuridine (EdU) assay and flow cytometry analysis, respectively. The production of tumor necrosis factor-α (TNF-α) and IL-6 was determined by an enzyme-linked immunosorbent assay (ELISA). The binding relationship between miR-940 and circ_0128846 or PTPN12 was identified by dual-luciferase reporter assay and RNA immunoprecipitation assay. RESULTS: Circ_0128846 and PTPN12 expression were significantly upregulated, whereas miR-940 was downregulated in the cartilage tissues of OA patients and IL-1ß-treated C28/I2 cells compared with controls. IL-1ß treatment inhibited C28/I2 cell proliferation and induced cell apoptosis and the production of inflammatory factors, TNF-α and IL-6; however, these effects were partly reversed after circ_0128846 depletion. In terms of mechanism, circ_0128846 acted as a miR-940 sponge, and miR-940 combined with PTPN12. Also, circ_0128846 depletion partly ameliorated IL-1ß-induced C28/I2 cell disorders through miR-940. PTPN12 overexpression also partly relieved miR-940-mediated effects in IL-1ß-treated C28/I2 cells. Further, circ_0128846 induced PTPN12 expression by interacting with miR-940. CONCLUSION: Circ_0128846 regulated human chondrocyte proliferation, apoptosis and inflammation through the miR-940/PTPN12 pathway in OA.

LncRNA GATA2-AS1 suppresses esophageal squamous cell carcinoma progression via the mir-940/PTPN12 axis.

Esophageal squamous cell carcinoma (ESCC) is a common malignant tumor worldwide. Long noncoding RNAs (lncRNAs) exhibit a regulatory role in the progression of ESCC. Our research was performed to investigate the potential molecular mechanism of lncRNA GATA2-AS1 in ESCC. METHODS: The expression of GATA2-AS1 was identified by qRT-PCR. Cell function assays explored the potential effect of GATA2-AS1 on ESCC progression. The subcellular hierarchical localization method was executed to identify the subcellular localization of GATA2-AS1 in ESCC cells. A prediction website was utilized to discover the relationships among GATA2-AS1, miR-940 and PTPN12. Dual luciferase reporter gene, pull-down assays and RIP assays were executed to verify the binding activity among GATA2-AS1, miR-940 and PTPN12. Xenograft tumor experiments were performed to evaluate ESCC cell growth in vivo. RESULTS: The expression of GATA2-AS1 and PTPN12 was reduced, while miR-940 expression was enhanced in ESCC tissues and cell lines. In vivo experiments showed that GATA2-AS1 inhibited the progression of ESCC cells toward malignancy. Bioinformatics analysis, dual luciferase and RIP assays revealed that GATA2-AS1 upregulated PTPN12 expression by competitively targeting miR-940. miR-940 reversed the inhibitory effect of GATA2-AS1 on the biological behavior of ESCC cells. CONCLUSION: Our findings suggested that GATA2-AS1, expressed at low levels in ESCC, plays a crucial role in the progression of ESCC by targeting the miR-940/PTPN12 axis and could be a potential drug target to treat ESCC patients.

Zinc finger Asp-His-His-Cys palmitoyl -acyltransferase 19 accelerates tumor progression through wnt/ß-catenin pathway and is upregulated by miR-940 in osteosarcoma.

Osteosarcoma (OS) is the most frequent malignant primary bone tumor in children and young adults. Zinc finger Asp-His-His-Cys palmitoyl-acyltransferase 19 (ZDHHC19) is a key enzyme in protein palmitoylation and plays crucial roles in tumor progression. However, its expression profile and biological function in OS have been unclear. In the present study, the expression level of ZDHHC19 in OS cell lines was determined by qRT-PCR and Western blot. The effect of ZDHHC19 in cell growth, invasion and migration was analyzed by CCK8, EDU, transwell, wound healing assay in vitro, and xenograft tumor model in vivo. In addition, bioinformatics analysis was used to explore the potential mechanism of ZDHHC19 in OS. Furthermore, the luciferase reporter assay was conducted to determine the direct binding between miR-940 and ZDHHC19. We discovered that ZDHHC19 was overexpressed in OS cells compared with the normal cells. The functional investigation demonstrated that ZDHHC19 silencing could inhibit proliferation, invasion and migration of OS in vitro and suppress tumorigenicity and lung metastasis in a xenograft model in vivo. Mechanistically, we identified that ZDHHC19 was a direct target of miR-940 and forced ZDHHC19 expressions partially rescue the suppression of proliferation, migration and invasion induced by miR-940. Moreover, bioinformatics analysis combined with validation experiments revealed that activating wnt/ß-catenin pathway contributed to the pro-oncogenic effect induced by ZDHHC19. Furthermore, rescue experiments further verified that miR-940/ZDHHC19 axis regulated wnt/ß-catenin pathway. Overall, these findings indicated that miR-940/ZDHHC19 axis played a significant role in OS progression and might be considered as a novel target for OS treatment.Abbreviations: OS, osteosarcoma; miRNAs, microRNAs; 3'-UTR, 3'- untranslated region; TARGET, Therapeutically Applicable Research To Generate Effective Treatments; qRT-PCR, quantitative real-time PCR; IHC, Immunohistochemistry; GSVA, Gene Set Variation Analysis; GSEA, Gene Set Enrichment Analysis; KEGG, Kyoto Encyclopedia of Genes and Genomes.

Circ_0003340 downregulation mitigates esophageal squamous cell carcinoma progression by targeting miR-940/PRKAA1 axis.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is a highly prevalent type of esophageal cancer (EC), usually found at an advanced stage with a high mortality rate, and it is now crucial to find new ways to diagnose and treat ESCC. This study analyzed the function of circular RNA_0003340 (circ_0003340)/microRNA-940 (miR-940)/protein kinase AMP-activated alpha 1 catalytic subunit (PRKAA1) axis in ESCC. METHODS: Circ_0003340, miR-940 and PRKAA1 contents were measured with the application of real-time quantitative polymerase chain reaction (RT-qPCR) and western blot. Cell proliferation, cell cycle, apoptosis, migration, invasion and angiogenesis were assessed with a cell counting kit-8 (CCK8), 5-ethynyl-2'-deoxyuridine (EdU), flow cytometry, wound healing, transwell and tube formation assays. We used both the luciferase reporter system and RNA immunoprecipitation (RIP) to analyze the relationship between miR-940 and circ_0003340 or PRKAA1. Finally, xenograft models were applied to analyze the effect of circ_0003340 on tumor growth in vivo. RESULTS: Upregulated circ_0003340 and PRKAA1, and downregulated miR-940 levels were detected in ESCC. Meanwhile, ESCC progression was apparently restrained by circ_0003340 knockdown in vitro. Circ_0003340 acted as a ceRNA for miR-940 in regulating ESCC progression and miR-940 was proved to target PRKAA1 to arrest ESCC progression in vitro. Finally, in vivo experiments established that silencing of circ_0003340 slowed tumor growth in vivo. CONCLUSION: Circ_0003340 downregulation mitigated esophageal squamous cell carcinoma progression by targeting miR-940/PRKAA1 axis.

Circ_0020123 promotes NSCLC tumorigenesis via up-regulating KIAA1522 expression through miR-940.

Circ_0020123 was highly expressed in NSCLC tissues and cell lines, and knockdown of circ_0020123 abolished cell growth, migration and invasion in vitro and hindered tumor growth in nude mice. Mechanically, circ_0020123 directly targeted miR-940, and KIAA1522 was a target of miR-940. Thereafter, a series of rescue experiments showed that circ_0020123 served its biological functions by miR-940/KIAA1522 axis. In all, circ_0020123 acted as an oncogene to promote the tumorigenesis of NSCLC via miR-940/KIAA1522 axis, suggesting a potential therapeutic target for NSCLC treatment.

LncRNA THEMIS2-211, a tumor-originated circulating exosomal biomarker, promotes the growth and metastasis of hepatocellular carcinoma by functioning as a competing endogenous RNA.

Hepatocellular carcinoma (HCC) is a major challenge for human health. Finding reliable diagnostic biomarkers and therapeutic targets for HCC is highly desired in the clinic. Currently, circulating exosomal lncRNA is a promising biomarker for the diagnosis of cancer and lncRNA is also a potential target in cancer therapy. Here, the diagnostic value of a panel based on exosomal lncRNA THEMIS2-211 and PRKACA-202, superior to that of AFP, was identified for diagnosing human HCC. Besides, the performance of exosomal lncRNA THEMIS2-211 alone exceeds that of AFP in diagnosing early-stage HCC patients (stage I). Furthermore, lncRNA THEMIS2-211 is highly expressed in HCC tissues and correlated with the poor prognosis of HCC patients. LncRNA THEMIS2-211 is upregulated and localized in the cytoplasm of HCC cells. LncRNA THEMIS2-211 exerts its biological function as an oncogene that promotes the proliferation, migration, invasion, EMT of HCC cells by physically interacting with miR-940 and therefore promoting SPOCK1 expressions. Rescue assays show the regulation of SPOCK1 by lncRNA THEMIS2-211 dependents on miR-940. The discovery of lncRNA THEMIS2-211 further illuminates the molecular pathogenesis of HCC and the THEMIS2-211/miR-940/SPOCK1 axis may act as a potential therapeutic target for HCC.

CircMAN1A2 promotes vasculogenic mimicry of nasopharyngeal carcinoma cells through upregulating ERBB2 via sponging miR-940.

Nasopharyngeal carcinoma (NPC) is the most prevalent human primary malignancy of the head and neck, and the presence of vasculogenic mimicry (VM) renders anti-angiogenic therapy ineffective and poorly prognostic. However, the underlying mechanisms are unclear. In the present study, we used miR-940 silencing and overexpression for in vitro NPC cell EdU staining, wound healing assay and 3D cell culture assay, and in vivo xenograft mouse model and VM formation to assess miR-940 function. We found that ectopic miR-940 expression reduced NPC cell proliferation, migration and VM, as well as tumorigenesis in vivo. By bioinformatic analysis, circMAN1A2 was identified as a circRNA that binds to miR-940. Mechanistically, we confirmed that circMAN1A2 acts as a sponge for miR-940, impairs the inhibitory effect of miR-940 on target ERBB2, and then activates the PI3K/AKT/mTOR signaling pathway using RNA-FISH, dual luciferase reporter gene and rescue analysis assays. In addition, upregulation of ERBB2 expression is associated with clinical staging and poor prognosis of NPC. Taken together, the present findings suggest that circMAN1A2 promotes VM formation and progression of NPC through miR-940/ERBB2 axis and further activates the PI3K/AKT/mTOR pathway. Therefore, circMAN1A2 may become a biomarker and therapeutic target for anti-angiogenic therapy in patients with nasopharyngeal carcinoma.

Hsa_circ_0062682 Promotes Serine Metabolism and Tumor Growth in Colorectal Cancer by Regulating the miR-940/PHGDH Axis.

Colorectal cancer (CRC) is one of the most common malignancies globally. Increasing evidence indicates that circular RNAs (circRNAs) play a pivotal role in various cancers. The present study focused on exploring the role of a functionally unknown circRNA, hsa_circ_0062682 (circ_0062682), in CRC. By online analyses and experimental validations, we showed that circ_0062682 expression was aberrantly increased in CRC tissues compared with paired normal tissues. Increased expression of circ_0062682 in CRC notably correlated with a poor prognosis and advanced tumor stage. Functional experiments showed that circ_0062682 knockdown reduced CRC growth both in vitro and in vivo. Mechanistically, we revealed that circ_0062682 could sponge miR-940 and identified D-3-phosphoglycerate dehydrogenase (PHGDH), a key oxidoreductase involved in serine biosynthesis, as a novel target of miR-940. Silencing miR-940 expression could mimic the inhibitory effect of circ_0062682 knockdown on CRC proliferation. The expression of PHGDH was downregulated in circ_0062682-depleted or miR-940 overexpressing CRC cells at both the mRNA and protein levels. Circ_0062682 knockdown suppressed CRC growth by decreasing PHGDH expression and serine production via miR-940. Taken together, these data demonstrate, for the first time, that circ_0062682 promotes serine metabolism and tumor growth in CRC by regulating the miR-940/PHGDH axis, suggesting circ_0062682 as a potential novel therapeutic target for CRC.

MiR-940 Serves as a Diagnostic Biomarker in Patients with Sepsis and Regulates Sepsis-Induced Inflammation and Myocardial Dysfunction.

INTRODUCTION: Sepsis is a heterogeneous syndrome with a life-long threat caused by infection. This study aimed to investigate the clinical function of miR-940 and its influence on cardiomyocyte models. METHODS: The relative expression of miR-940 was assessed by qRT-PCR and the roles in the clinical diagnosis of miR-940 were revealed by the ROC curve. The relationship between miR-940 and clinical parameters was validated by Pearson analysis. The sepsis rat models were established by treatment with cecal ligation and perforation (CLP) and clinical items including left ventricular systolic pressure (LVSP), left ventricular and end-diastolic pressure (LVEDP), maximum rate of increase/decrease in left ventricular blood pressure (± dp/dtmax) as well as troponin (cTnl), creatine kinase isoenzyme (CK-MB), TNF-α, IL-1ß, and IL-6 were detected. RESULTS: The finding of qRT-PCR accentuated that the relative expression of miR-940 was significantly decreased in sepsis patients and CLP-stimulated models. The ROC curve proposed that miR-940 could be a satisfactory diagnostic biomarker for sepsis patients. Pearson analysis reinforced the expression of miR-940 was negatively associated with the PCT, WBC, CRP, Scr, SOFA score, and APACHE II score. The outcome of CLP-steered rat verified that overexpression of miR-940 inhibited the detrimental effects of CLP on myocardial dysfunction and inflammation reactions. CONCLUSION: The downregulation of miR-940 was reported and it might be an underlying diagnostic marker in sepsis patients. Overexpression of miR-940 protected myocardial function from damage and inflammation induced by CLP.

miR-940 is a new biomarker with tumor diagnostic and prognostic value.

miR-940 is a microRNA located on chromosome 16p13.3, which has varying degrees of expression imbalance in many diseases. It binds to the 3' untranslated region (UTR) and affects the transcription or post-transcriptional regulation of target protein-coding genes. For a diversity of cellular processes, including cell proliferation, migration, invasion, apoptosis, epithelial-to-mesenchymal transition (EMT), cell cycle, and osteogenic differentiation, miR-940 can affect them not only by regulating protein-coding genes but also long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) in pathways. Intriguingly, miR-940 participates in four pathways that affect cancer development, including the Wnt/ß-catenin pathway, mitogen-activated protein kinase (MAPK) pathway, PD-1 pathway, and phosphatidylinositol 3-kinase (PI3K)-Akt pathway. Importantly, the expression of miR-940 is intimately correlated with the diagnosis and prognosis of tumor patients, as well as to the efficacy of tumor chemotherapy drugs. In conclusion, our main purpose is to outline the expression of miR-940 in various diseases and the molecular biological and cytological functions of target genes in order to reveal its potential diagnostic and prognostic value as well as its predictive value of drug efficacy.

MicroRNA-940 as a Potential Serum Biomarker for Prostate Cancer.

Prostate cancer is one of the leading causes of death despite an astoundingly high survival rate for localized tumors. Though prostate specific antigen (PSA) test, performed in conjunction with digital rectal examinations, is reasonably accurate, there are major caveats requiring a thorough assessment of risks and benefits prior to conducting the test. MicroRNAs, a class of small non-coding RNAs, are stable molecules that can be detected in circulation by non-invasive methods and have gained importance in cancer prognosis and diagnosis in the recent years. Here, we investigate circulating miR-940, a miRNA known to play a role in prostate cancer progression, in both cell culture supernatants as well as patient serum and urine samples to determine the utility of miR-940 as a new molecular marker for prostate cancer detection. We found that miR-940 was significantly higher in serum from cancer patients, specifically those with clinically significant tumors (GS ≥ 7). Analysis of receiver operating characteristic curve demonstrated that miR-940 in combination with PSA had a higher area under curve value (AUC: 0.818) than the miR-940 alone (AUC: 0.75) for the diagnosis of prostate cancer. This study provides promising results suggesting the use of miR-940 for prostate cancer diagnosis.

Hsa_circ_0046263 Drives the Carcinogenesis and Metastasis of Non-Small Cell Lung Cancer Through the Promotion of NOVA2 by Absorbing Mir-940 as a Molecular Sponge.

BACKGROUND: Circular RNAs (circRNAs) have increasingly been investigated in different cancers due to their regulatory roles. In this study, hsa_circ_0046263 will be detailedly researched in non-small cell lung cancer (NSCLC). METHODS: The analyses of hsa_circ_0046263, microRNA-940 (miR-940), and neuro-oncological ventral antigen 2 (NOVA2) levels were administrated by quantitative real-time polymerase chain reaction (qRT-PCR). The proliferation detection was conducted using Cell Counting Kit-8 (CCK-8) and colony formation assays. Cell cycle and apoptosis were evaluated by flow cytometry. Transwell assay for migration and invasion was used to determine cell metastatic capacity. Overall protein levels were examined adopting Western blot. Target binding analysis was completed via dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. The effect of hsa_circ_0046263 on NSCLC in vivo was studied by xenograft model in mice. RESULTS: Hsa_circ_0046263 was overtly upregulated in NSCLC with important prognostic value. In vitro experiments indicated that hsa_circ_0046263 knockdown caused inhibitory effects on NSCLC cell proliferation, cell cycle, and metastasis but stimulative effect on apoptosis. Molecular mechanism analysis demonstrated that hsa_circ_0046263 served as a miR-940 sponge to act in the development of NSCLC. Moreover, miR-940 targeted NOVA2 and NOVA2 was regulated by hsa_circ_0046263/miR-940 axis. NOVA2 overexpression also neutralized the miR-940-mediated progression inhibition of NSCLC cells. In vivo assays suggested that hsa_circ_0046263 enhanced NSCLC tumorigenesis by targeting miR-940/NOVA2 axis. CONCLUSION: Hsa_circ_0046263 was identified as a cancer-promoting factor in NSCLC via sponging miR-940 and upregulating NOVA2, which presented a clear mechanism of NSCLC occurrence and progression.