Search

Background: In recent years, several giant pandas have suffered from testicular tumor, which has seriously affected giant panda health. However, the pathogenesis of testicular tumor in giant panda is still unclear. Studies have shown that miRNAs are involved in the occurrence and development of a variety of cancers. However, the effect of miRNAs on giant panda testicular tumor has been little studied. Therefore, this study explored the pathogenesis of giant panda testicular tumor through miRNA and mRNA sequencing, and screened out diagnostic markers of testicular tumor. Results: Combined with phenotypic symptoms and pathological section results, three giant pandas were diagnosed with testicular tumor and divided into tumor group, and three other giant pandas were divided into normal group. A total of 29 differentially expressed miRNAs (DEmiRNAs) were screened by blood miRNA-seq, and 3149 target gene candidates were predicted. Functional enrichment analysis showed that the target genes were mainly involved in intermembrane lipid transfer and ATP-dependent chromatin remodeling. However, only 5 DEmiRNAs were screened by miRNA-seq of blood-derived exosomes and 364 target genes were predicted, which were mainly involved in antigen processing and presentation. In addition, 216 differentially expressed genes (DEGs) were screened by RNA-seq, and functional enrichment analysis showed that tumor-specific DEGs significantly enriched to protein phosphorylation. Spearman correlation analysis of miRNA-mRNA showed that the expressions of miR-331-3p and PKIG were significantly positively correlated (spearman = 0.943, p < 0.01), while the expressions of miR-331-3p and ENSAMEG00000013628 were significantly negatively correlated (spearman= -0.829, p < 0.05). RT-PCR showed that the expression of miR-331-3p was significantly decreased in giant panda with tumor (p < 0.01). Conclusions: blood miRNAs and exosomal miRNAs exhibit distinct regulatory patterns concerning giant panda testicular tumor, potentially reflecting divergent biological processes in the disease's etiology. Meanwhile, miR-331-3p could be used as a potential diagnostic marker for giant panda testicular tumor. Our findings are conducive to the rapid clinical diagnosis of testicular tumor in giant pandas, and are also expected to provide scientific reference for further research on the pathogenesis of testicular tumor. [ABSTRACT FROM AUTHOR]

Abstract Background In recent years, several giant pandas have suffered from testicular tumor, which has seriously affected giant panda health. However, the pathogenesis of testicular tumor in giant panda is still unclear. Studies have shown that miRNAs are involved in the occurrence and development of a variety of cancers. However, the effect of miRNAs on giant panda testicular tumor has been little studied. Therefore, this study explored the pathogenesis of giant panda testicular tumor through miRNA and mRNA sequencing, and screened out diagnostic markers of testicular tumor. Results Combined with phenotypic symptoms and pathological section results, three giant pandas were diagnosed with testicular tumor and divided into tumor group, and three other giant pandas were divided into normal group. A total of 29 differentially expressed miRNAs (DEmiRNAs) were screened by blood miRNA-seq, and 3149 target gene candidates were predicted. Functional enrichment analysis showed that the target genes were mainly involved in intermembrane lipid transfer and ATP-dependent chromatin remodeling. However, only 5 DEmiRNAs were screened by miRNA-seq of blood-derived exosomes and 364 target genes were predicted, which were mainly involved in antigen processing and presentation. In addition, 216 differentially expressed genes (DEGs) were screened by RNA-seq, and functional enrichment analysis showed that tumor-specific DEGs significantly enriched to protein phosphorylation. Spearman correlation analysis of miRNA-mRNA showed that the expressions of miR-331-3p and PKIG were significantly positively correlated (spearman = 0.943, p

Abstract Background Osteoporosis (OP) is a high-incidence bone disease that is prone to osteoporotic fractures (OF), so it has attracted widespread attention. Aim This study investigated the specific expression and role of miR-331 in patients with OP and OF. The findings have profound implications for the clinical prevention and treatment of these conditions. Methods The study included 60 OP patients, 46 OF patients, and 40 healthy controls. The expression level of miR-331-3p was detected using RT-qPCR. BMP2 was used to stimulate differentiation in MC3T3-E1 cells. After induction, the expression activity of osteogenic differentiation-related gene markers was detected using RT-qPCR. The target gene analysis was conducted using a luciferase reporter assay. Results The levels of miR-331-3p were significantly elevated, while NRP2 levels were significantly reduced in OF patients. Post-surgery, miR-331-3p levels decreased over time. MiR-331-3p was found to negatively regulate the luciferase activity of NPR2 in MC3T3-E1 cells. Furthermore, overexpression of miR-331-3p inhibited cell proliferation and decreased the levels of osteoblast differentiation markers. Conclusion The up-regulation of miR-331-3p can promote OP and might also encourage the occurrence of OF by regulating NRP2. However, this needs further verification.

Background: Osteoporosis (OP) is a high-incidence bone disease that is prone to osteoporotic fractures (OF), so it has attracted widespread attention. Aim: This study investigated the specific expression and role of miR-331 in patients with OP and OF. The findings have profound implications for the clinical prevention and treatment of these conditions. Methods: The study included 60 OP patients, 46 OF patients, and 40 healthy controls. The expression level of miR-331-3p was detected using RT-qPCR. BMP2 was used to stimulate differentiation in MC3T3-E1 cells. After induction, the expression activity of osteogenic differentiation-related gene markers was detected using RT-qPCR. The target gene analysis was conducted using a luciferase reporter assay. Results: The levels of miR-331-3p were significantly elevated, while NRP2 levels were significantly reduced in OF patients. Post-surgery, miR-331-3p levels decreased over time. MiR-331-3p was found to negatively regulate the luciferase activity of NPR2 in MC3T3-E1 cells. Furthermore, overexpression of miR-331-3p inhibited cell proliferation and decreased the levels of osteoblast differentiation markers. Conclusion: The up-regulation of miR-331-3p can promote OP and might also encourage the occurrence of OF by regulating NRP2. However, this needs further verification. [ABSTRACT FROM AUTHOR]

The excessive proliferation of keloid fibroblasts is one of the important reasons leading to the formation of keloids. Circular RNA (circRNA) is an important regulator that regulates the biological functions of cells. However, the role and mechanism of circ‐PDE7B in keloid formation have not been studied yet. QRT‐PCR was used to detect the circ‐PDE7B, miR‐331‐3p and cyclin‐dependent kinase 6 (CDK6) expression. The biological functions of keloid fibroblasts were determined by MTT assay, flow cytometry, transwell assay and wound healing assay. Western blot analysis was used to measure the protein levels of extracellular matrix (ECM) markers and CDK6. The interaction between miR‐331‐3p and circ‐PDE7B or CDK6 was confirmed by dual‐luciferase reporter assay and RIP assay. Circ‐PDE7B was found to be upregulated in keloid tissues and fibroblasts. Downregulation of circ‐PDE7B could suppress the proliferation, invasion, migration, ECM accumulation and accelerate the apoptosis of keloid fibroblasts. Circ‐PDE7B could serve as a sponge of miR‐331‐3p, and the regulation of silenced circ‐PDE7B on the biological functions of keloid fibroblasts could be abolished by miR‐331‐3p inhibitor. Additionally, CDK6 was a target of miR‐331‐3p, and its overexpression could reverse the negative regulation of miR‐331‐3p on the biological functions of keloid fibroblasts. Circ‐PDE7B sponged miR‐331‐3p to positively regulate CDK6 expression. Taken together, circ‐PDE7B promoted the proliferation, invasion, migration and ECM accumulation of keloid fibroblasts by regulating the miR‐331‐3p/CDK6 axis, suggesting that circ‐PDE7B might be a potential target for keloid treatment. [ABSTRACT FROM AUTHOR]

Abstract Background Circular RNAs are implicated in modulating the progression of various malignant tumors. However, the function and underlying mechanisms of circ_0005615 in multiple myeloma (MM) remain unclear. Methods The expression levels of circ_0005615, miR-331-3p and IGF1R were tested by quantitative real-time polymerase chain reaction or western blot assay. Cell counting kit-8 and 5-ethynyl-2′-deoxyuridine (EdU) assay were performed for cell proliferation detection. Cell apoptosis and cell cycle were measured by flow cytometry. The protein expressions of Bax and Bcl-2 were detected by western blot assay. Glucose consumption, lactate production and ATP/ADP ratios were estimated to disclose cell glycolysis. The interaction relationship among miR-331-3p and circ_0005615 or IGF1R was proved by dual-luciferase reporter assay. Results The abundance of circ_0005615 and IGF1R was increased in MM patients and cells, while the expression of miR-331-3p was decreased. Circ_0005615 inhibition retarded the proliferation and cell cycle progression, while reinforced the apoptosis of MM cells. Molecularly, circ_0005615 could sponge miR-331-3p, and the repressive trends of circ_0005615 deficiency on MM progression could be alleviated by anti-miR-331-3p introduction. Additionally, IGF1R was validated to be targeted by miR-331-3p, and IGF1R overexpression mitigated the suppressive function of miR-331-3p on MM development. Furthermore, IGF1R was mediated by circ_0005615/miR-331-3p axis in MM cells. Conclusion Circ_0005615 downregulation blocked MM development by targeting miR-331-3p/IGF1R axis.

microRNA-331-3p (miR-331-3p) has been displayed as an oncogene in pancreatic cancer (PC). The current research set out to elucidate how miR-331-3p in carcinoma-associated fibroblasts (CAFs)-derived extracellular vesicles (EVs) facilitated the tumorigenesis in PC. First, a dual-luciferase reporter assay was adopted to investigate the relationship between miR-331-3p and SCARA5. In addition, EVs were isolated normal fibroblasts and CAFs, and these isolated EVs were co-cultured with PC cells. Cell proliferative and migrating/invasive potentials were further evaluated with the help of a CCK-8 and Transwell assays, respectively. Gain- and loss-of-function assays were also implemented to assess the role of miR-331-3p, SCARA5, and FAK pathway in PC cells. Lastly, xenograft nude mice were established to investigate the role of miR-331-3p in vivo. miR-331-3p negatively targeted SCARA5 and was highly expressed in CAFs-derived EVs, which accelerated the proliferative, migrating, and invasive potentials of PC cells. Meanwhile, over-expression of miR-331-3p enhanced the proliferative, migrating, and invasive properties of PC cells and promoted tumor growth in vivo by manipulating SCARA5/FAK axis, whereas SCARA5 countered the oncogenic effects of miR-331-3p. Overall, miR-331-3p in CAFs-derived EVs inhibits SCARA5 expression and activates the FAK pathway, thereby augmenting the progression of PC. Our study provides a potential therapeutic target for the treatment of PC.

Zhao Zhou,1,2,&ast; Xiaohan Cui,1,2,&ast; Peng Gao,2 Xudong Zhang,2 Chunfu Zhu,2 Beicheng Sun1 1Department of Hepatobiliary Surgery of Nanjing Drum Tower Hospital, Clinical College of Nanjing Medical University, Nanjing, People’s Republic of China; 2The Affiliated Changzhou NO.2 People’s Hospital of Nanjing Medical University, Changzhou, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Chunfu Zhu, The Affiliated Changzhou NO.2 People’s Hospital of Nanjing Medical University, Changzhou, People’s Republic of China, Email zcfmlm@njmu.edu.cn Beicheng Sun, Department of Hepatobiliary Surgery of Nanjing Drum Tower Hospital, Clinical College of Nanjing Medical University, Nanjing, People’s Republic of China, Email sunbc@nju.edu.cnObjective: Recently, emerging studies have validated that circular RNAs participate in multiple biological progresses in various human malignant tumors, including hepatocellular carcinoma (HCC). However, until now, the elucidated mechanism of circular RNAs is only the tip of the iceberg. In this study, we firstly identify a novel circular RNA circRASSF5 (the only circular RNA derived from the RASSF5 gene), and attempt to investigate its biological function and underlying mechanism in HCC.Methods: qRT-PCR, Western blotting and IHC were applied to detect the expression of related genes. CCK-8 assay, EdU staining, wound healing and transwell assays were used to investigate HCC proliferation, migration and invasion abilities. Animal model studies were included to investigate the function of circRASSF5 in HCC tumorigenesis and metastasis. RNA pull-down assay, luciferase reporter assay and FISH (fluorescence in situ hybridization) assay were performed to explore the potential biological mechanism underlying circRASSF5 function in HCC.Results: CircRASSF5 is obviously downregulated in both HCC tissues and cell lines. Low level of circRASSF5 is negatively associated with larger tumor size, severe vascular invasion, more portal vein tumor embolus and unfavorable prognosis. Loss-of-function assay reveals that circRASSF5 remarkably impedes the growth and metastasis of HCC cells in vitro and in vivo. Mechanistically, circRASSF5 directly interacts with miR-331-3p as a sponge, and then enhances the expression of PH domain and leucine-rich repeat protein phosphatase (PHLPP), thus restraining the progression of HCC cells.Conclusion: Altogether, we validate that circRASSF5 is a tumor suppressor in HCC, which competitively sponges with miR-331-3p and then enhances the tumor inhibitory effect of PHLPP, indicating the potential application value of circRASSF5 for HCC diagnosis and clinical treatment.Keywords: circRASSF5, miR-331-3p, PHLPP, HCC

Purpose: Long noncoding RNAs (lncRNAs) have been gradually regarded as influential indicators of various cancers. The present study aimed to identify the effects of lncRNA HOTAIR on cervical cancer progression. Methods: RNA and protein expressions were quantified by RT-qPCR and western blot assays. Fluorescence in situ hybridization (FISH) assay was carried out to examine the intracellular location of HOTAIR. Cancer cell viability and mobility were detected by CCK-8, colony formation, transwell and wound healing assays. Binding relationships between miR-331-3p and HOTAIR/RCC2 were validated by luciferase reporter assay. Results: RT-qPCR assays showed that HOTAIR levels were notably upregulated in cervical cancer tissues and cell lines. Furthermore, a fluorescence in situ hybridization (FISH) assay suggested that HOTAIR was mostly located in the cytoplasm of cancer cells, indicating a sponging function. CCK-8, colony formation, Transwell and wound-healing assays indicated that knockdown of HOTAIR in HeLa and SiHa cells significantly reduced cell growth, migration and invasion. Subsequently, miR-331-3p was proven to be the target molecule of HOTAIR. In addition, results from Pearson's correlation analysis indicated negative correlation between HOTAIR and miR-331-3p in cervical cancer tissues. HOTAIR negatively modulated miR-331-3p expression. Ultimately, the target gene of miR-331-3p was verified to be RCC2, and miR-331-3p negatively modulated RCC2 expression. In addition, analysis on clinical cervical cancer tissues confirmed the negative correlation between miR-331-3p and RCC2. HOTAIR and RCC2 showed oncogenic functions in HeLa and SiHa cells, while miR-331-3p exerted the reverse effect. Conclusions: HOTAIR plays a carcinogenic role in cervical cancer by targeting the miR-331-3p/RCC2 axis. Moreover, clinical cervical cancer tissues confirmed the negative correlation between miR-331-3p with lncRNA HOTAIR and RCC2. These data suggested an underlying therapeutic target for cervical cancer. [ABSTRACT FROM AUTHOR]

Background: Circular RNAs are implicated in modulating the progression of various malignant tumors. However, the function and underlying mechanisms of circ_0005615 in multiple myeloma (MM) remain unclear. Methods: The expression levels of circ_0005615, miR-331-3p and IGF1R were tested by quantitative real-time polymerase chain reaction or western blot assay. Cell counting kit-8 and 5-ethynyl-2′-deoxyuridine (EdU) assay were performed for cell proliferation detection. Cell apoptosis and cell cycle were measured by flow cytometry. The protein expressions of Bax and Bcl-2 were detected by western blot assay. Glucose consumption, lactate production and ATP/ADP ratios were estimated to disclose cell glycolysis. The interaction relationship among miR-331-3p and circ_0005615 or IGF1R was proved by dual-luciferase reporter assay. Results: The abundance of circ_0005615 and IGF1R was increased in MM patients and cells, while the expression of miR-331-3p was decreased. Circ_0005615 inhibition retarded the proliferation and cell cycle progression, while reinforced the apoptosis of MM cells. Molecularly, circ_0005615 could sponge miR-331-3p, and the repressive trends of circ_0005615 deficiency on MM progression could be alleviated by anti-miR-331-3p introduction. Additionally, IGF1R was validated to be targeted by miR-331-3p, and IGF1R overexpression mitigated the suppressive function of miR-331-3p on MM development. Furthermore, IGF1R was mediated by circ_0005615/miR-331-3p axis in MM cells. Conclusion: Circ_0005615 downregulation blocked MM development by targeting miR-331-3p/IGF1R axis. [ABSTRACT FROM AUTHOR]

Abstract Background Prostate cancer (PCa) is the most frequently diagnosed malignancy in men, and its mechanism remains poorly understood. Therefore, it is urgent to discover potential novel diagnostic biomarkers and therapeutic targets that can potentially facilitate the development of efficient anticancer strategies. Methods A series of functional in vitro and in vivo experiments were conducted to evaluate the biological behaviors of PCa cells. RNA pulldown, Western blot, luciferase reporter, immunohistochemistry and chromatin immunoprecipitation assays were applied to dissect the detailed underlying mechanisms. High-throughput sequencing was performed to screen for differentially expressed circRNAs in PCa and adjacent normal tissues. Results Upregulation of protein arginine methyltransferase 5 (PRMT5) is associated with poor progression-free survival and the activation of multiple signaling pathways in PCa. PRMT5 inhibits the transcription of CAMK2N1 by depositing the repressive histone marks H4R3me2s and H3R8me2s on the proximal promoter region of CAMK2N1, and results in malignant progression of PCa both in vitro and in vivo. Moreover, the expression of circSPON2, a candidate circRNA in PCa tissues identified by RNA-seq, was found to be associated with poor clinical outcomes in PCa patients. Further results showed that circSPON2 induced PCa cell proliferation and migration, and that the circSPON2-induced effects were counteracted by miR-331-3p. Particularly, circSPON2 acted as a competitive endogenous RNA (ceRNA) of miR-331-3p to attenuate the repressive effects of miR-331-3p on its downstream target PRMT5. Conclusions Our findings showed that the epigenetic regulator PRMT5 aggravates PCa progression by inhibiting the transcription of CAMK2N1 and is modulated by the circSPON2/miR-331-3p axis, which may serve as a potential therapeutic target for patients with aggressive PCa.

Background: Multiple myeloma (MM) is a malignant tumor of plasma cells in the bone marrow. Circular RNAs (circRNAs) exert important activity in the tumorigenesis and chemoresistance of MM. In the current work, we sought to identify the expression, activity, and mechanism of circPSAP activity in MM. Methods: CircPSAP, microRNA (miR)-331-3p, and histone deacetylase 4 (HDAC4) were quantified by qRT-PCR and immunoblotting assays. Cell proliferation and survival were assessed by CCK-8 assay. Cell cycle and apoptosis were detected by flow cytometry. The direct relationship between miR-331-3p and circPSAP or HDAC4 3′UTR was validated by dual-luciferase reporter, RNA pull-down, and RNA immunoprecipitation (RIP) assays. Results: CircPSAP was overexpressed in human MM and high levels of circPSAP predicted poor prognosis in MM patients. CircPSAP depletion repressed cell proliferation and promoted apoptosis and BTZ sensitivity. Mechanistically, circPSAP functioned as a miR-331-3p sponge, and circPSAP regulated cell proliferation, apoptosis and BTZ sensitivity by sponging miR-331-3p. MiR-331-3p directly targeted and inhibited HDAC4. MiR-331-3p-mediated inhibition of HDAC4 impaired cell proliferation and enhanced cell apoptosis and BTZ sensitivity. Moreover, circPSAP modulated HDAC4 expression by acting as a miR-331-3p sponge. Conclusion: Our findings highlight a novel mechanism, in which circPSAP functions as a miR-331-3p sponge to impact MM cell proliferation, apoptosis and BTZ sensitivity by regulating HDAC4 expression.

The dysregulation of circular RNAs (circRNAs) is closely associated with the pathogenesis of colorectal cancer (CRC). The present study aimed to elucidate the biological function and mechanism of circ_0060927 in CRC. 5-ethynyl-2'-deoxyuridine, Cell Counting Kit-8 (CCK-8), flow cytometry and transwell assays, as well as Xenograft tumor models were adopted for in vitro and in vivo analyses. The interaction between microRNA-331–3p (miR-331–3p) and circ_0060927 or T-box transcription factor 2 (TBX2) was verified by the dual-luciferase reporter and RNA pull-down assays. Circ_0060927 deficiency inhibited cell proliferation, autophagy, migration, and invasion and increased cell apoptosis and necrosis in CRC cells, as well as inhibited tumor growth in vivo. Circ_0060927 could bind to miR-331–3p, and circ_0060927 regulated CRC cell behaviors via sponging miR-331–3p. TBX2 was targeted by miR-331–3p, and miR-331–3p targeted TBX2 to exert the anti-cancer role in CRC cells. Mechanically, circ_0060927 regulated TBX2 expression by sequestering miR-331–3p in CRC cells. Circ_0060927 downregulation inhibited CRC progression by regulating the miR-331–3p/TBX2 axis, which might offer a potential treatment target for CRC. [ABSTRACT FROM AUTHOR]

Abstract Background Osteoarthritis (OA) is characterized by joint pain and joint function limitation. Hsa_circ_0045714 (circ_0045714) is a novel OA-related circular RNA. However, its repertoire remains to be further clarified in joint chondrocytes. Methods RNA and protein expression levels and inflammatory factor levels were detected by real-time quantitative polymerase chain reaction, western blotting and enzyme-linked immunosorbent assay. Cell proliferation and apoptosis were determined by colony formation assay, cell counting kit-8 assay and apoptosis assay. Direct interaction was predicted by bioinformatics method and confirmed by dual-luciferase reporter assay. Results Expression of circ_0045714 and phosphoinositide-3-kinase (PI3K) regulatory subunit 3 (PIK3R3) was declined, and microRNA (miR)-331-3p was promoted in knee articular cartilages and cells from OA patients, as well as interleukin (IL)-1β-challenged human articular chondrocytes (HAC) cell line. In stimulation of IL-1β, HAC cells showed a loss of colony formation ability, cell viability and expression of Bcl-2 and Collagen II, allied with an increase in apoptosis rate and levels of IL-6, IL-8 and tumor necrosis factor-α, Bcl-2-associated X protein, cleaved caspase-3, and ADAM with thrombospondin motif-5. Noticeably, overexpressing circ_0045714 and inhibiting miR-331-3p could suppress IL-1β-evoked these effects, and both were through up-regulating PIK3R3, a key gene in PI3K/AKT signaling pathway. Mechanically, circ_0045714 functioned as competing endogenous RNA (ceRNA) for miR-331-3p and further regulated expression of the downstream target gene PIK3R3. Conclusion There was a novel circ_0045714/miR-331-3p/PIK3R3 ceRNA axis in HAC, and its inhibition might be one mechanism of HAC injury in OA.

Introduction: Ulcerative colitis (UC) is a debilitating condition of the gastrointestinal system, and long non-coding RNA (lncRNA)-H19 emerges as a crucial player in inflammatory diseases. This study is designed to evaluate the mechanism of H19 in intestinal injury of UC mice and hint at a novel target for UC treatment. Methods: UC mouse model was established, followed by injection of shH19, antagomir-331-3p, and tumor necrosis factor receptor-associated factor 4 (TRAF4) overexpression vector. H19, miR-331-3p, and TRAF4 expressions were detected via reverse transcription quantitative polymerase chain reaction. Intestinal injury was appraised via disease activity index (DAI), hematoxylin-eosin staining, and histopathological scoring. Interleukin (IL)-1β, tumor necrosis factor (TNF)-α, and IL-10 levels were detected via enzyme-linked immunosorbent assay. Binding relationships of H19 and miR-331-3p and TRAF4 were verified. Results: H19 was highly expressed in colon tissues. Silencing H19 attenuated intestinal injury of UC mice, manifested by reductions in weight loss, DAI, histopathological scores, IL-1β and TNF-α, and increases in colon length and IL-10. Mechanically, lncRNA-H19 is bound to miR-331-3p to inhibit its expression. TRAF4 is a target of miR-331-3p. Inhibition of miR-331-3p or overexpression of TRAF4 could reverse the alleviating role of lncRNA-H19 in intestinal injury of UC mice. Conclusion: LncRNA-H19 was highly expressed in UC mice and bound to miR-331-3p to promote TRAF4 transcription, thereby aggravating intestinal injury. [ABSTRACT FROM AUTHOR]

Abstract Background Circular RNAs (circRNAs) are gradually reported to be implicated in the development of malignant tumors, including ovarian cancer (OC). This paper intended to explore the function and action mechanism of hsa_circ_0004712 in OC. Results In our results, hsa_circ_0004712 was aberrantly overexpressed in OC tissues and cells. Downregulation of hsa_circ_0004712 impaired OC cell proliferation, colony formation, invasion and migration, and accelerated apoptosis. Hsa_circ_0004712 directly targeted miR-331-3p whose inhibitors reversed the effects of hsa_circ_0004712 downregulation. FZD4 was targeted by miR-331-3p, and hsa_circ_0004712 could positively regulated FZD4 expression by targeting miR-331-3p. The anti-tumor effects of miR-331-3p restoration were reversed by FZD4 overexpression. Downregulation of hsa_circ_0004712 also impaired tumor development in vivo by regulating miR-331-3p and FZD4. Conclusion In conclusion, hsa_circ_0004712 deficiency repressed OC development by mediating the miR-331-3p/FZD4 pathway, predicting that hsa_circ_0004712 was a promising biomarker for OC diagnosis and therapy.

Jun Rao, Lihua Shao, Min Lin, Jin Huang, Li Fan Department of Gastroenterology, The Affiliated Changzhou No.2 People’s Hospital of Nanjing Medical University, Changzhou, People’s Republic of ChinaCorrespondence: Li FanDepartment of Gastroenterology, The Affiliated Changzhou No.2 People’s Hospital of Nanjing Medical University, No. 68 Gehu Road, Wujin District, Changzhou, Jiangsu, 213000, People’s Republic of ChinaEmail lifan226@163.comBackground: Ulcerative colitis (UC) has become one of the fastest-growing severe diseases worldwide with high morbidity. This research aimed to explore the function of lncRNA UCA1 in UC progression.Methods: RT-qPCR analysis was used to examine the expression of UCA1 level in colonic mucosa tissues of UC patients. Then, fetal human cells (FHCs) were stimulated by LPS to induce inflammatory injury. CCK-8, flow cytometry and ELISA were adopted to determine the influence of UCA1 depletion on cell viability, apoptosis and pro-inflammatory factors levels in LPS-induced FHCs. The interaction between UCA1 and miR-331-3p or BRD4 was confirmed by luciferase reporter assay. The expressions of key factors involved in NF-κB pathway were assessed by Western blotting.Results: LncRNA UCA1 level was elevated in colonic mucosa tissues of UC patients. LPS stimulation restrained cell viability and promoted the apoptosis and inflammatory factors levels, thus inducing FHCs inflammatory injury, while these effects were partially abolished by UCA1 knockdown. Moreover, it was found that UCA1 silence improved LPS-triggered cell injury via miR-331-3p. In addition, BRD4 was directly targeted by miR-331-3p, and BRD4 deficiency neutralized the effects of miR-331-3p repression on LPS-triggered injury in LPS-treated FHCs.Conclusion: Our data determined that UCA1 knockdown attenuated UC development via targeting the miR-331-3p/BRD4/NF-κB pathway.Keywords: UCA1, ulcerative colitis, miR-331-3p, BRD4

Abstract Background Hepatocellular carcinoma and hepatitis C are strongly associated. The current work aimed to study the expression levels of microRNA-331-3p and microRNA-23b-3p as propable biomarkers for detecting liver cancer (HCC) at its early stages in patients with HCV-related liver cirrhosis. The current prospective study included two hundred participants, divided into three groups: group I, 100 patients with HCV-related liver cirrhosis; group II, 50 HCC patients at early stages; and group III, 50 apparentlyhealthy controls. All patients had routine laboratory workup and ultrasound hepatic assessment. Values of microRNA-331-3p and microRNA-23b-3p were measured by real-time quantitative PCR. Results Levels of miR-331-3p were significantly higher in HCC patients than in cirrhotic patients and controls (p < 0.001), while levels of miR-23b-3p were significantly lower in HCC patients compared to cirrhotics and controls (p < 0.001). ROC curve revealed that miR-23b-3p had 80% sensitivity and 74% specificity, miR-331-3p had 66% sensitivity and 61% specificity, and AFP had 64% sensitivity and 61% specificity of 61% in discrimination between HCC patients from controls. Conclusion Serum miR-23b-3p is a more effective predictor than miR-331-3p and AFP for the development of hepatocellular carcinoma in hepatitis C (HCV)-related cirrhotic patients.

Background: Prostate cancer (PCa) is the most frequently diagnosed malignancy in men, and its mechanism remains poorly understood. Therefore, it is urgent to discover potential novel diagnostic biomarkers and therapeutic targets that can potentially facilitate the development of efficient anticancer strategies. Methods: A series of functional in vitro and in vivo experiments were conducted to evaluate the biological behaviors of PCa cells. RNA pulldown, Western blot, luciferase reporter, immunohistochemistry and chromatin immunoprecipitation assays were applied to dissect the detailed underlying mechanisms. High-throughput sequencing was performed to screen for differentially expressed circRNAs in PCa and adjacent normal tissues. Results: Upregulation of protein arginine methyltransferase 5 (PRMT5) is associated with poor progression-free survival and the activation of multiple signaling pathways in PCa. PRMT5 inhibits the transcription of CAMK2N1 by depositing the repressive histone marks H4R3me2s and H3R8me2s on the proximal promoter region of CAMK2N1, and results in malignant progression of PCa both in vitro and in vivo. Moreover, the expression of circSPON2, a candidate circRNA in PCa tissues identified by RNA-seq, was found to be associated with poor clinical outcomes in PCa patients. Further results showed that circSPON2 induced PCa cell proliferation and migration, and that the circSPON2-induced effects were counteracted by miR-331-3p. Particularly, circSPON2 acted as a competitive endogenous RNA (ceRNA) of miR-331-3p to attenuate the repressive effects of miR-331-3p on its downstream target PRMT5. Conclusions: Our findings showed that the epigenetic regulator PRMT5 aggravates PCa progression by inhibiting the transcription of CAMK2N1 and is modulated by the circSPON2/miR-331-3p axis, which may serve as a potential therapeutic target for patients with aggressive PCa. [ABSTRACT FROM AUTHOR]

Coxsackie virus B3 (CVB3), an enterovirus, is the main pathogen causing viral myocarditis, pericarditis, hepatitis and other inflammation-related diseases. Non-coding RNAs with a closed loop molecular structure, called circular RNAs (circRNAs), have been shown to be involved in multiple virus-related processes, but roles and mechanisms in CVB3 infection have not been systematically studied. In this study, when HeLa cells were infected with CVB3, the expression of hsa_circ_0000367 (circSIAE) was significantly decreased as demonstrated by real-time quantitative PCR assays. We found that circSIAE downregulated the expression of miR-331-3p through direct binding and inhibited the replication of CVB3 in HeLa and 293T cells. The analysis of signals downstream of miR-331-3p suggested that miR-331-3p promotes CVB3 replication, viral plaque formation and fluorescent virus cell production through interactions with the gene coding for thousand and one amino-acid kinase 2 (TAOK2). In conclusion, this study found that circSIAE can target TAOK2 through sponge adsorption of miR-331-3p to inhibit the replication and proliferation of CVB3 virus, providing an early molecular target for the diagnosis of CVB3 infection. [ABSTRACT FROM AUTHOR]

microRNA-331-3p (miR-331-3p) has been displayed as an oncogene in pancreatic cancer (PC). The current research set out to elucidate how miR-331-3p in carcinoma-associated fibroblasts (CAFs)-derived extracellular vesicles (EVs) facilitated the tumorigenesis in PC. First, a dual-luciferase reporter assay was adopted to investigate the relationship between miR-331-3p and SCARA5. In addition, EVs were isolated normal fibroblasts and CAFs, and these isolated EVs were co-cultured with PC cells. Cell proliferative and migrating/invasive potentials were further evaluated with the help of a CCK-8 and Transwell assays, respectively. Gain- and loss-of-function assays were also implemented to assess the role of miR-331-3p, SCARA5, and FAK pathway in PC cells. Lastly, xenograft nude mice were established to investigate the role of miR-331-3p in vivo. miR-331-3p negatively targeted SCARA5 and was highly expressed in CAFs-derived EVs, which accelerated the proliferative, migrating, and invasive potentials of PC cells. Meanwhile, over-expression of miR-331-3p enhanced the proliferative, migrating, and invasive properties of PC cells and promoted tumor growth in vivo by manipulating SCARA5/FAK axis, whereas SCARA5 countered the oncogenic effects of miR-331-3p. Overall, miR-331-3p in CAFs-derived EVs inhibits SCARA5 expression and activates the FAK pathway, thereby augmenting the progression of PC. Our study provides a potential therapeutic target for the treatment of PC. [ABSTRACT FROM AUTHOR]

Abstract Background We aimed to explore the role of long noncoding RNA urothelial carcinoma-associated 1 (lncRNA UCA1) and its underlying mechanism in the radioresistance of prostate cancer (PCa). Methods QRT-PCR was conducted to measure the expression of UCA1, microRNA-331-3p (miR-331-3p) and eukaryotic translation initiation factor 4 gamma 1 (EIF4G1) in PCa tissues and cells. The relative protein level was determined by western blot assay. Cell proliferation and apoptosis were detected by MTT, colony formation assay, and flow cytometry, respectively. The target interaction between miR-331-3p and UCA1 or EIF4G1 was predicted through bioinformatics analysis, and verified by dual-luciferase reporter gene assay system. Results The high levels of UCA1 and EIF4G1 as well as the low level of miR-331-3p were observed in PCa tissues and cell lines. UCA1 and EIF4G1 expression were significantly upregulated by Gy radiation treatement. UCA1 or EIF4G1 knockdown repressed cell growth and enhanced cell apoptosis in 22RV1 and DU145 cells under radiation. Moreover, overexpression of EIF4G1 abolished UCA1 knockdown-induced effect on 6 Gy irradiated PCa cells. UCA1 sponged miR-331-3p to regulate EIF4G1 expression. Conclusions LncRNA UCA1 deletion suppressed the radioresistance to PCa by suppressing EIF4G1 expression via miR-331-3p. UCA1 acted as a potential regulator of radioresistance of PCa, providing a promising therapeutic target for PCa.

Qing-Qing Tian,1 Jing Xia,2 Xin Zhang,3 Bao-Qin Gao,4 Wei Wang5 1Department of Pathology, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People’s Republic of China; 2General Department of Houhu, The Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People’s Republic of China; 3Department of Radiology, The Fourth People’s Hospital of Huai’an, Huai’an, Huai’an, People’s Republic of China; 4Operating Room, Huai’an Second People’s Hospital and the Affiliated Huai’an Hospital of Xuzhou Medical University, Huai’an, People’s Republic of China; 5Department of Oncology, Huai’an Second People’s Hospital and the Affiliated Huai’an Hospital of Xuzhou Medical University, Huai’an, People’s Republic of ChinaCorrespondence: Bao-Qin GaoOperating Room, Huai’an Second People’s Hospital and the Affiliated Huai’an Hospital of Xuzhou Medical University, Huai’an, People’s Republic of ChinaEmail hagbqsyh@163.comWei WangDepartment of Oncology, Huai’an Second People’s Hospital and the Affiliated Huai’an Hospital of Xuzhou Medical University, Huai’an, People’s Republic of ChinaEmail Wwei2003@126.comObjective: Mounting research has established the role of microRNAs (miRNAs) as oncogenes or anti-oncogenes (tumor suppressors) in the development and progression of several cancers. The purpose of our current study is to delineate the roles and functional mechanisms of miR-331-3p and MLLT10 in non-small cell lung cancer (NSCLC) tumorigenesis.Patients and Methods: Quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was employed to measure miR-331-3p expression levels in twenty-six matched tumor tissues and non-cancerous tissues collected from patients suffering from NSCLC, and from six NSCLC cell lines separately: A549, H1650, H292, H1299, H1944 and BEAS-2b. We employed the dual-luciferase activity assay to check whether the putative gene, MLLT10, was a downstream target of miR-331-3p in NSCLC pathogenesis and development. Western blot was conducted to analyze the protein expression levels of MLLT10 (AF10), E-cadherin, Vimentin, and GAPDH. CCK-8 assay, transwell migration assay, and transwell invasion assay were carried out to observe the functions of miR-331-3p and MLLT10 on NSCLC tumor cell proliferation, metastasis, and invasion, respectively. To identify whether the metastasis of NSCLC tumor cells was EMT-mediated, supplementary experiments involving E-cadherin and Vimentin were implemented.Results: miR-331-3p was downregulated in NSCLC, which promoted tumor cell proliferation, whereas the overexpression of miR-331-3p inhibited tumor cell proliferation. Being a direct target of miR-331-3p, MLLT10 was negatively modulated by miR-331-3p, which suppressed tumor cell proliferation, migration, and invasion in NSCLC. However, MLLT10 overexpression alleviated the above inhibitory effects. Furthermore, EMT-mediated metastasis was proved to be present in NSCLC.Conclusion: miR-331-3p played a suppressor role in NSCLC tumor cell proliferation, EMT-mediated metastasis, and invasion by targeting MLLT10. Our findings highlighted that miR-331-3p/MLLT10 axis could be useful as a clinical diagnostic marker and therapeutic target in NSCLC patients.Keywords: NSCLC, miR-331-3p, MLLT10, EMT-mediated metastasis

Coxsackie virus B3 (CVB3), an enterovirus, is the main pathogen causing viral myocarditis, pericarditis, hepatitis and other inflammation-related diseases. Non-coding RNAs with a closed loop molecular structure, called circular RNAs (circRNAs), have been shown to be involved in multiple virus-related processes, but roles and mechanisms in CVB3 infection have not been systematically studied. In this study, when HeLa cells were infected with CVB3, the expression of hsa_circ_0000367 (circSIAE) was significantly decreased as demonstrated by real-time quantitative PCR assays. We found that circSIAE downregulated the expression of miR-331-3p through direct binding and inhibited the replication of CVB3 in HeLa and 293T cells. The analysis of signals downstream of miR-331-3p suggested that miR-331-3p promotes CVB3 replication, viral plaque formation and fluorescent virus cell production through interactions with the gene coding for thousand and one amino-acid kinase 2 (TAOK2). In conclusion, this study found that circSIAE can target TAOK2 through sponge adsorption of miR-331-3p to inhibit the replication and proliferation of CVB3 virus, providing an early molecular target for the diagnosis of CVB3 infection.

Background: Osteoarthritis (OA) is characterized by joint pain and joint function limitation. Hsa_circ_0045714 (circ_0045714) is a novel OA-related circular RNA. However, its repertoire remains to be further clarified in joint chondrocytes. Methods: RNA and protein expression levels and inflammatory factor levels were detected by real-time quantitative polymerase chain reaction, western blotting and enzyme-linked immunosorbent assay. Cell proliferation and apoptosis were determined by colony formation assay, cell counting kit-8 assay and apoptosis assay. Direct interaction was predicted by bioinformatics method and confirmed by dual-luciferase reporter assay. Results: Expression of circ_0045714 and phosphoinositide-3-kinase (PI3K) regulatory subunit 3 (PIK3R3) was declined, and microRNA (miR)-331-3p was promoted in knee articular cartilages and cells from OA patients, as well as interleukin (IL)-1β-challenged human articular chondrocytes (HAC) cell line. In stimulation of IL-1β, HAC cells showed a loss of colony formation ability, cell viability and expression of Bcl-2 and Collagen II, allied with an increase in apoptosis rate and levels of IL-6, IL-8 and tumor necrosis factor-α, Bcl-2-associated X protein, cleaved caspase-3, and ADAM with thrombospondin motif-5. Noticeably, overexpressing circ_0045714 and inhibiting miR-331-3p could suppress IL-1β-evoked these effects, and both were through up-regulating PIK3R3, a key gene in PI3K/AKT signaling pathway. Mechanically, circ_0045714 functioned as competing endogenous RNA (ceRNA) for miR-331-3p and further regulated expression of the downstream target gene PIK3R3. Conclusion: There was a novel circ_0045714/miR-331-3p/PIK3R3 ceRNA axis in HAC, and its inhibition might be one mechanism of HAC injury in OA. [ABSTRACT FROM AUTHOR]

Background: Circular RNAs (circRNAs) are gradually reported to be implicated in the development of malignant tumors, including ovarian cancer (OC). This paper intended to explore the function and action mechanism of hsa_circ_0004712 in OC. Results: In our results, hsa_circ_0004712 was aberrantly overexpressed in OC tissues and cells. Downregulation of hsa_circ_0004712 impaired OC cell proliferation, colony formation, invasion and migration, and accelerated apoptosis. Hsa_circ_0004712 directly targeted miR-331-3p whose inhibitors reversed the effects of hsa_circ_0004712 downregulation. FZD4 was targeted by miR-331-3p, and hsa_circ_0004712 could positively regulated FZD4 expression by targeting miR-331-3p. The anti-tumor effects of miR-331-3p restoration were reversed by FZD4 overexpression. Downregulation of hsa_circ_0004712 also impaired tumor development in vivo by regulating miR-331-3p and FZD4. Conclusion: In conclusion, hsa_circ_0004712 deficiency repressed OC development by mediating the miR-331-3p/FZD4 pathway, predicting that hsa_circ_0004712 was a promising biomarker for OC diagnosis and therapy. [ABSTRACT FROM AUTHOR]

• Circ_0062270 silencing represses melanoma cell progression and tumor growth. • Circ_0062270 interacts with miR-331-3p. • EPHA2 is a target of miR-331-3p. Circular RNA (circRNA) has been confirmed to play a vital role in melanoma progression. The regulatory function of circ_0062270, a novel circRNA, in melanoma progression is unclear. Relative expression levels of circ_0062270 and microRNA (miR)-331-3p were determined using qRT-PCR. Cell counting kit 8 assay, EdU staining and flow cytometry were used to measure cell proliferation, cell cycle distribution and apoptosis. The protein levels of proliferation, apoptosis and metastasis-related markers, as well as EPH receptor A2 (EPHA2), were tested using western blot analysis. Besides, cell migration and invasion were evaluated using transwell assay. Meanwhile, the interaction between miR-331-3p and circ_0062270 or EPHA2 was confirmed by dual-luciferase reporter assay or RIP assay. Additionally, tumor xenograft models were constructed to investigate the function of circ_0062270 on melanoma tumor growth in vivo. The expression of circ_0062270 was increased in melanoma tissues and cells. Knockdown of circ_0062270 inhibited proliferation, promoted apoptosis, and repressed metastasis in melanoma. Moreover, circ_0062270 could serve as miR-331-3p sponge, and miR-331-3p could target EPHA2. Furthermore, miR-331-3p inhibitor and EPHA2 overexpression reversed the inhibitory effect of circ_0062270 silencing on melanoma progression. In addition, silenced circ_0062270 also could inhibit melanoma tumor growth in vivo. Circ_0062270 accelerated the progression of melanoma through regulating the miR-331-3p/EPHA2 axis, suggesting that circ_0062270 might be a novel potential therapeutic target for melanoma. [ABSTRACT FROM AUTHOR]

Peripheral nerve injury (PNI) is a common clinical problem, which can cause severe disability and dramatically affect a patient’s quality of life. Neural regeneration after PNI is a complex biological process that involves a variety of signaling pathways and genes. Emerging studies demonstrated that long non-coding RNAs (lncRNAs) were abnormally expressed after PNI and played pivotal roles in peripheral nerve regeneration. Based on the rat sciatic nerve injury model, we found that the expression levels of several lncRNAs were increased significantly in the sciatic nerve after injury. Software prediction prompted us to focus on one up-regulated lncRNA, MSTRG.24008.1. Dual-luciferase reporter assay, RNA pull-down assay and RNA interference approach verified that MSTRG.24008.1 regulated neuroregeneration via the miR-331-3p/nucleotide-binding oligomerization domain-like pyrin domain containing 3 (NLRP3)/myelin and lymphocyte protein (MAL) axis in vitro. Subsequently, we performed gastrocnemius muscle gravity and sciatic functional index experiments to evaluate the recovery of injured sciatic nerves after MSTRG.24008.1 siRNA interference in vivo. In conclusion, knockdown of MSTRG.24008.1 promotes the regeneration of the sciatic nerve via the miR-331-3p/NLRP3/MAL axis, which may provide a new strategy to evaluate and repair injured peripheral nerves clinically.

Purpose: To explore the effect of SEV on colon cancer cells through circ-PI4KA. Methods: The RNA level of circular RNA_0062389, microRNA-331-3p and LIM and SH3 protein 1 was determined by quantitative real-time polymerase chain reaction. Protein expression was detected by Western blot. Cell proliferation was investigated by 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide, cell colony formation and 5-ethynyl-29-deoxyuridine assays. Cell apoptosis was demonstrated using Annexin V-fluorescein isothiocyanate/propidium iodide double staining assay. Cell migration and invasion were detected by transwell assay. The target relationship between miR-331-3p and circ-PI4KA or LASP1 was predicted by starBase v2.0 online database, and identified by a dual-luciferase reporter assay. The effects between SEV treatment and circ-PI4KA knockdown on tumor formation were presented by in vivo tumor formation assay. Results: Circ-PI4KA and LASP1 expressions were dramatically upregulated, while miR-331-3p was downregulated in colon cancer tissues and cells, respectively. SEV exposure significantly decreased the expression of circ-PI4KA and LASP1, but increased miR-331-3p expression. SEV inhibited cell proliferation, migration and invasion, and induced cell apoptosis by regulating circ-PI4KA. Furthermore, circ-PI4KA interacted with miR-331-3p, and miR-331-3p interacted with LASP1. SEV inhibited tumor growth by controlling circ-PI4KA in vivo. Conclusion: Circ-PI4KA attenuated SEV-treated colon cancer cell malignancy by upregulating LASP1 through binding to miR-331-3p, which provided a new mechanism for studying surgery-mediated therapy of colon cancer. [ABSTRACT FROM AUTHOR]

Long noncoding RNAs (lncRNAs) serve as crucial regulators in the pathogenesis of spinal cord injury (SCI). However, the role of lncRNA SOX2 overlapping transcript (SOX2OT) in SCI remains to be well revealed. An SCI rat model was established and assessed by the Basso–Beattie–Bresnahan (BBB) method. An SCI PC12 cell model was established through lipopolysaccharide (LPS) treatment. Quantitative real-time polymerase chain reaction assay was used for SOX2OT, miR-331-3p, and neurogenic differentiation 1 (Neurod1) mRNA levels. Cell counting kit-8 assay and flow cytometry analysis were performed for cell viability and apoptosis, respectively. Enzyme-linked immunosorbent assay was performed for the levels of inflammatory cytokines. The production of superoxide dismutase and malondialdehyde was determined with relevant kits. Dual-luciferase reporter and RNA immunoprecipitation assays were conducted for the relationships among SOX2OT, miR-331-3p, and Neurod1. Western blot assay was employed for protein levels. SOX2OT was elevated in SCI rat and cell models. SOX2OT knockdown relieved the injury of SCI in SCI rat model. Moreover, the suppressive role in PC12 cell viability and the promotional roles in apoptosis, inflammation, and oxidative stress mediated by LPS were all restored by silencing SOX2OT. For mechanism analysis, SOX2OT was identified as a sponge of miR-331-3p to positively regulate Neurod1 expression. Inhibition of miR-331-3p reversed the effect of SOX2OT knockdown on LPS-induced PC12 damage. Overexpression of miR-331-3p protected PC12 cells from LPS-induced damage by binding to Neurod1. In addition, SOX2OT knockdown relieved PC12 cell injury by inactivation of Janus kinase–signal transducer and activator of transcription pathway. SOX2OT promoted PC12 cell injury through modulating miR-331-3p/Neurod1 axis and activating Janus kinase–signal transducer and activator of transcription pathway. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS) caused by a single-stranded RNA virus (PRRSV) is a highly infectious respiratory disease and leads to huge economic losses to the swine industry worldwide. To investigate the role of miRNAs in the infection and lung injury induced by PRRSV, the differentially expressed miRNAs (DE-miRs) were isolated from PRRSV-2 infected/mock-infected PAMs of Meishan, Landrace, Pietrain, and Qingping pigs at 9, 36, and 60 hpi. Mir-331-3p was the only common DE-miR in each set of miRNA expression profile at 36 hpi. Mir-210 was one of 7 common DE-miRs between PRRSV infected and mock-infected PAMs of Meishan, Pietrain, and Qingping pigs at 60 hpi. Mir-331-3p/mir-210 could target PRRSV-2 ORF1b, bind and downregulate porcine TNF-α/STAT1 expression, and inhibit PRRSV-2 replication, respectively. Furthermore, STAT1 and TNF-α could mediate the transcriptional activation of MCP-1, VCAM-1, and ICAM-1. STAT1 could also upregulate the expression of TNF-α by binding to its promoter region. In vivo, pEGFP-N1-mir-331-3p could significantly reduce viral replication and pathological changes in PRRSV-2 infected piglets. Taken together, Mir-331-3p/mir-210 have significant roles in the infection and lung injury caused by PRRSV-2, and they may be promising therapeutic targets for PRRS and lung injury/inflammation.

Purpose: Non-small cell lung cancer (NSCLC) accounts for more than 80% of lung cancer cases and remains the primary cause of cancer-related deaths worldwide. Fentanyl is a commonly utilized anesthetic during the process of tumor resection, and exhibits inhibitory effects on the progression of numerous cancer types, including pancreatic cancer, colorectal cancer and gastric cancer. However, the effects of fentanyl on the cell viability and invasion of NSCLC has not been investigated. Current study aimed to investigate the effects and the mechanisms underlying the effects of fentanyl on NSCLC. Methods: The expression of μ-opioid receptor (MOR) was proved by flow cytometry. The expression of microRNA-331-3p (miR-331-3p) and histone deacetylase 5 (HDAC5) in NSCLC tissues and cell lines are evaluated by reverse transcription-quantitative PCR (RT-qPCR) and Western blot, respectively. Cell viability and invasion are measured by cell counting kit-8 (CCK-8) assay and transwell assay, respectively. The interaction between miR-331-3p and 3ʹ-untranslated region (UTR) of HDAC5 is predicted by TargetScan 7.1 (http://www.targetscan.org/vert%5f71/), validated by dual luciferase assay, RT-qPCR and Western blot. Results: There was lower miR-331-3p expression and higher HDAC5 expression in NSCLC cell lines A549 and CALU-1 compared with BEAS-2B, which was reversed by fentanyl administration. miR-331-3p targeted 3ʹ-UTR of HDAC5 in NSCLC cell lines A549 and CALU-1. miR-331-3p inhibitor partially abrogated the inhibitory effects of fentanyl on NSCLC cell viability and invasion by targeting HDAC5. In addition, there was higher HDAC5 expression and lower miR-331-3p expression in tumor tissues which were isolated from patients with NSCLC compared to the adjacent normal tissues, and miR-331-3p was negatively correlated with HDAC5 in NSCLC tumor tissues. Conclusion: Fentanyl inhibits the viability and invasion of NSCLC cells by induction of miR-331-3p and reduction of HDAC5. [ABSTRACT FROM AUTHOR]

Purpose: To investigate the specific role of melanotransferrin antisense RNA (MFI2-AS1) in esophageal cancer (EC) progression. Methods: The differential expression of MFI2-AS1 in EC tissues and cells was determined using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Silencing MFI2-AS1 was performed by transfection with specific short hairpin RNAs targeting MFI2-AS1. The 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MTT) and flow cytometry (FC) were used to assess cell viability and apoptosis of EC cells, respectively. The sponging microRNA (miRNA) of MFI2-AS1 was validated using luciferase activity and RNA immunoprecipitation assays while the downstream target gene of the sponging miRNA was evaluated by luciferase activity assay. Results: MFI2-AS1 was significantly enhanced in EC tissues (p < 0.01) and indicated a poor prognosis in EC patients. Knockdown of MFI2-AS1 in EC cells decreased cell viability and promoted cell apoptosis of EC cells. Functionally, MFI2-AS1 targeted miR-331-3p, and sex-determining region on Ychromosome- related high-mobility-group box4 (SOX4) was identified as a target gene of miR-331-3p. Ectopic expression of SOX4 counteracted the suppressive effect of MFI2-AS1 knockdown on EC cell viability and stimulative effect on EC cell apoptosis. Conclusion: The pro-oncogenic effect of MFI2-AS1 on EC progression occurs via the regulation of the miR-331-3p/SOX4 axis, providing a new potential therapeutic target for EC. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS) caused by a single-stranded RNA virus (PRRSV) is a highly infectious respiratory disease and leads to huge economic losses to the swine industry worldwide. To investigate the role of miRNAs in the infection and lung injury induced by PRRSV, the differentially expressed miRNAs (DE-miRs) were isolated from PRRSV-2 infected/mock-infected PAMs of Meishan, Landrace, Pietrain, and Qingping pigs at 9, 36, and 60 hpi. Mir-331-3p was the only common DE-miR in each set of miRNA expression profile at 36 hpi. Mir-210 was one of 7 common DE-miRs between PRRSV infected and mock-infected PAMs of Meishan, Pietrain, and Qingping pigs at 60 hpi. Mir-331-3p/mir-210 could target PRRSV-2 ORF1b, bind and downregulate porcine TNF- α/ STAT1 expression, and inhibit PRRSV-2 replication, respectively. Furthermore, STAT1 and TNF- α could mediate the transcriptional activation of MCP-1, VCAM-1 , and ICAM-1. STAT1 could also upregulate the expression of TNF- α by binding to its promoter region. In vivo , pEGFP-N1-mir-331-3p could significantly reduce viral replication and pathological changes in PRRSV-2 infected piglets. Taken together, Mir-331-3p/mir-210 have significant roles in the infection and lung injury caused by PRRSV-2, and they may be promising therapeutic targets for PRRS and lung injury/inflammation. [ABSTRACT FROM AUTHOR]

Gastric cancer (GC) has emerged as one of the most common malignancies in gastrointestinal system. Inducible T-cell costimulator ligand (ICOSLG) was found to be highly expressed in various cancers, which contributes to disease progression. This study aims to investigate the role of ICOSLG and its potential mechanism of action in dictating the aggressiveness of GC cell. ICOSLG and miR-331–3p expression patterns in cancerous and para-cancerous tissues from GC patients were examined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The miRNAs targeting ICOSLG were predicted by "miRDB", "starBase," and "TargetScan" databases. The interplay of ICOSLG and miR-331–3p in dictating the aggressiveness and glycolysis of GC cells was investigated by CCK-8 proliferation assay and Transwell migration/invasion assays, as well as the detection of glucose uptake, lactate production and ATP levels. The tumorigenesis of GC cells after ICOSLG silencing was examined in the nude mice. ICOSLG was highly expressed in GC tissues, and GC patients with high ICOSLG expression showed a poorer prognosis than the low-expression group. Further, high ICOSLG level was correlated with more advanced TNM stages, more lymph-node metastases, and poorer tumor differentiation. ICOSLG knockdown inhibited the proliferation, migration, invasion and tumor formation of GC cells, which was concomitant with reduced glucose consumption, lactate production, and ATP levels. In contrast, ICOSLG overexpression enhanced the aggressiveness of GC cells, and this effect was abrogated after the treatment with glycolysis inhibitor. We further found that miR-331–3p was a negative regulator of ICOSLG4, and miR-331–3p overexpression reduced ICOSLG4 expression and suppressed the aggressive phenotype induced by ICOSLG4 in GC cells. Together, these findings indicate that ICOSLG4, as an oncogene, is upregulated to promote glycolysis and the malignant phenotype in GC cells. miR-331–3p, which is downregulated in GC tissues, functions as a negative regulator of ICOSLG4. Targeting miR-331–3p/ICOSLG4 axis could potentially suppress GC progression. miR-331-3p negatively regulates ICOSLG expression in GC cells. A) miRNA regulators of ICOSLG mRNA were predicted by "miRDB", "starBase," and "TargetScan" databases. B) qRT-PCR analysis of ICOSLG mRNA levels in GC cells after the transfection of miR-NC or miRNA mimics. C) qRT-PCR analysis of miR-331–3p expression in 70 pairs of GC tumor samples and para-neoplastic tissues. D) Spearman correlation coefficient analysis of ICOSLG4 and miR-331–3p expression levels in GC tumor tissues. E) qRT-PCR analysis of miR-331–3p levels in GC cells after the transfection with miRNA mimic or miRNA inhibitor. F) WB analysis of ICOSLG4 protein levels in GC cells after the transfection with miRNA mimic or miRNA inhibitor. G) The WT and mutated binding sites between miR-331–3p and ICOSLG mRNA 3′UTR. H) The dual-luciferase reporter assay using WT or MUT reporter in the presence of miR-331–3p mimic or miR-NC. [Display omitted] • The upregulation of ICOSLG in GC tissues and the association of its overexpression with the poor prognosis in GC patients. • ICOSLG knockdown attenuated the aggressiveness and glycolysis in GC cells. • MiR-331–3p served as a negative regulator of ICOSLG, which was downregulated in GC tissues. • Targeting miR-331–3p/ICOSLG axis could modulate the malignancy and glycolysis in GC cells. • Future efforts are required to decipher the mechanism by which ICOSLG regulates glycolysis in GC cells. [ABSTRACT FROM AUTHOR]

Colorectal carcinoma (CRC) is a common tumor of the digestive system with poor prognosis. Studies have shown that aberrant microRNA (miRNA) expression can affect CRC progression by regulating target genes. In the present study, we investigated the functional roles and potential mechanisms of miR-331-3p in CRC. The expression of miR-331-3p and neuropilin-2 (NRP2) in CRC was detected by RT-qPCR. Then, Transwell assays were conducted to investigate the influence of miR-331-3p on CRC cell invasion and migration abilities. Luciferase reporter assays were performed to determine the target gene of miR-331-3p. It was found that miR-331-3p expression was notably declined in CRC and inversely correlated with the NRP2 expression. miR-331-3p upregulation significantly inhibited CRC cell invasion and migration. Additionally, western blot analysis demonstrated that miR-331-3p restoration evidently suppressed CRC cell EMT. Moreover, NRP2 was conformed to be a novel target of miR-331-3p and knockdown of NRP2 partially inversed the effects of the miR-331-3p inhibitor on cell invasion and migration. These results suggested that miR-331-3p exerted tumor suppressive roles in CRC by targeting NRP2 and miR-331-3p/NRP2 may serve as a potential therapy for CRC. [ABSTRACT FROM AUTHOR]

Introduction: Epithelial ovarian carcinoma (EOC) is one of the most lethal gynecologic malignancies, with a poor 5-year survival rate. Numerous studies have shown that microRNAs participate in the malignant behavior of ovarian cancer cells by directly targeting multiple oncogenes or tumor suppressor genes.Material and Methods: Reverse transcription-PCR was used to determine the level of miR-331-3p in EOC. Cells proliferation was measured with the Cell Counting Kit-8. Cell mobility were measured by wound-healing assay. Cell migration and invasion were measured by transwell assay. Luciferase assays were used to demonstrate that RCC2 was a directed target of miR-331-3p in EOC. Western blots were used to measure the protein expression.Results: We found that the expression of microRNA-331-3p (miR-331-3p) in ovarian cancer cell lines is reduced (p < 0.01), and an increase of expression of miR-331-3p in ovarian cancer cells significantly inhibits cell proliferation (p < 0.001). Transwell and wound-healing assays showed that miR-331-3p inhibits the cell motility of ovarian cancer cells (p < 0.001). Regulator of chromosome condensation 2 (RCC2) was predicted to be a novel target for miR-331-3p. Our luciferase activity assay confirmed that RCC2 is directly targeted by miR-331-3p. RCC2 was negatively regulated by miR-331-3p (p < 0.001), and overexpression of RCC2 could restore the malignant behaviors of ovarian cancer cells, which was suppressed by miR-331-3p.Conclusions: These data indicate that miR-331-3p can inhibit proliferation, migration, and invasion of ovarian cancer cells via directly targeting RCC2. Our study provides potential therapeutic targets for the treatment of ovarian cancer. [ABSTRACT FROM AUTHOR]

OBJECTIVE: We attempted to clarify the regulatory mechanism of UCA1/miR- 331-3p/IL6R on cell progression in multiple myeloma (MM). PATIENTS AND METHODS: The expression of UCA1, miR-331-3p, and IL6R in tumor tissues and cells was measured by qRT-PCR. Cell Counting Kit-8 (CCK-8) was conducted to detect cell proliferation, and flow cytometry assay was applied to examine cell apoptosis. Protein expression of L6R, p-JAK2, p-STAT3, c-Myc, CyclinD1, Bcl-2, and Bax was detected by Western blot assay. The interaction among miR-331-3p, UCA1, and IL6R was determined by Luciferase reporter system. Murine xenograft assay was performed to confirm the biological function of UCA1 in vivo. RESULTS: The expression of UCA1 and IL6R was up-regulated, while miR-331-3p was down-regulated in MM tumors and cell lines compared with normal tissues and cells. By calculation, miR-331-3p was correlated with UCA1 or IL6R inversely. In addition, UCA1 knockdown suppressed cell proliferation and promoted apoptosis in vitro and in vivo. Luciferase reporter system confirmed the interaction between miR-331-3p and UCA1 or IL6R. More importantly, UCA1 restored miR-331- 3p mediated inhibition of proliferation and promotion on apoptosis of MM cells. Consistently, IL6R rescued UCA1 knockdown caused repression on MM cell growth and elevation on apoptosis. Besides, UCA1 facilitated the activation of the JAK2/STAT3 signaling pathway by enhancing IL6R expression via targeting miR-331-3p. CONCLUSIONS: UCA1 accelerates proliferation and suppresses apoptosis in MM by targeting miR-331-3p/IL6R axis to activate JAK2/STAT3 pathway, providing potential targets for the diagnosis and therapy of MM. [ABSTRACT FROM AUTHOR]

MicroRNAs affect various pathological pathways by binding to multiple target mRNAs. Recently, it was reported that eukaryotic RNA G-quadruplexes were enriched in mRNA 3′-UTR, where microRNAs can also bind. To determine the roles of the G-quadruplex within mRNA 3′-UTR in microRNA binding sites, a bioinformatics approach was utilized to identify candidate microRNA target mRNAs with potential G-quadruplex formation. Circular dichroism spectrometry demonstrated the formation of RNA G-quadruplexes in vitro in candidate G-rich sequences. Mutated guanosines that are critical for G-quadruplex formation significantly decreased luciferase activities. Moreover, a G-quadruplex ligand TMPyP4 was used to destabilize the KCNJ11 RNA G-quadruplex in cardiomyocytes, resulting in binding of the microRNA to mRNA and subsequent suppression of KCNJ11 expression. In conclusion, our study showed that the G-quadruplex structure affects microRNA binding to its target mRNA. Thus, our study reveals a novel mechanism for G-quadruplex-dependent regulation of microRNA-mRNA interaction, which is essential to maintain normal gene expression. [ABSTRACT FROM AUTHOR]

OBJECTIVE: Recent evidence shows that gastric adenocarcinoma predictive long intergenic noncoding RNA (GAPLINC) acts as a critical role in the proliferation and metastasis in several tumors. We aimed to explore the expression pattern, function, and potential mechanism of GAPLINC in glioblastoma multiforme (GBM). PATIENTS AND METHODS: The expression levels of GAPLINC and its clinical significance were determined by analyzing TCGA datasets. RT-PCR was performed to detect the levels of GAPLINC and miR-331-3p. CCK-8, colony formation, EdU assays, wound healing assay and transwell invasion assay were performed to analyze the effect of GAPLINC on GBM behaviors. MiRNAs that may interact with GAPLINC were predicted using StarBase and RegRNA 2.0. A luciferase reporter assay was used to detect the targeting effect of GAPLINC on miR-331-3p. RESULTS: We found that GAPLINC expression was significantly up-regulated in both GBM tissues and cell lines. The overexpression of GAPLINC was associated with shorter overall survival and disease-free survival. Functional assays indicated that GAPLINC silencing suppressed GBM cells proliferation, migration, and invasion, and promoted apoptosis. In the mechanism, we found that GAPLINC acted as a competing endogenous RNA to sponge miR-331-3p and knockdown of GAPLINC promoted the expression of miR-331-3p. CONCLUSIONS: Our findings suggested that GAPLINC served as an oncogenic lncRNA in GBM through negative modulation of miR-331-3p, providing a novel treatment targeting for GBM. [ABSTRACT FROM AUTHOR]

Accumulating evidence documented the key functions of circular RNAs (circRNAs) in various malignancies. However, the study regarding the involvement of circRNAs in non-small cell lung cancer (NSCLC) has just begun. In the present study, qRT-PCR was used to determine the expression of circ_0001649 in NSCLC tissues and cells. Its clinical significance was further assessed by Fisher's exact test, Kaplan-Meier analysis and Cox regression model. Additionally, loss-of-function and gain-of-function experiments were carried out to detect the functional role of circ_0001649 in cell proliferation, migration and invasion. Furthermore, animal study was performed to confirm the in vitro results. Importantly, luciferase reporter assay was induced to reveal the underlying mechanism of circ_0001649. As a result, circ_0001649 was decreased in NSCLC tissues and cells and this downregulation is correlated with advanced TNM stage, positive lymph node metastasis and unfavorable prognosis. Additionally, circ_0001649 inhibited cell growth and metastasis both in vitro and in vivo . In mechanism, circ_0001649 was identified as the sponge of miR-331-3p and miR-338-5p. Moreover, the biological functions of circ_0001649 is partly dependent on its regulation on miR-331-3p and miR-338-5p. Collectively, this study suggested that circ_0001649/miR-331-3p/miR-338-5p regulatory signaling might be a potential target for NSCLC therapy. [ABSTRACT FROM AUTHOR]

Copyright of Practical Oncology Journal is the property of Journal of Practical Oncology Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Aberrant expression of microRNAs (miRNAs) is involved in the development and progression of various types of cancers. In this study, we investigated the role of miR-331-3p in cell proliferation and the expression of keratinocyte differentiation markers of uterine cervical cancer cells. Moreover, we evaluated whether neuropilin 2 (NRP2) are putative target molecules that regulate the human papillomavirus (HPV) related oncoproteins E6 and E7. Cell proliferation in the human cervical cancer cell lines SKG-II, HCS-2, and HeLa was assessed using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay. Cellular apoptosis was measured using the TdT-mediated dUTP nick end labeling (TUNEL) and Annexin V assays. Quantitative RT-PCR was used to measure the messenger RNA (mRNA) expression of the NRP2, E6, E7, p63, and involucrin (IVL) genes. A functional assay for cell growth was performed using cell cycle analyses. Overexpression of miR-331-3p inhibited cell proliferation, and induced G2/M phase arrest and apoptosis in SKG-II, HCS-2 and HeLa cells. The luciferase reporter assay of the NRP2 30-untranslated region revealed the direct regulation of NRP2 by miR-331-3p. Gene expression analyses using quantitative RT-PCR in SKG-II, HCS-2, and HeLa cells overexpressing miR-331-3p or suppressing NRP2 revealed down-regulation of E6, E7, and p63 mRNA and up-regulation of IVL mRNA. Moreover, miR-331-3p overexpression was suppressed NRP2 expression in protein level. We showed that miR-331-3p and NRP2 were key effectors of cell proliferation by regulating the cell cycle, apoptosis. NRP-2 also regulates the expression of E6/E7 and keratinocyte differentiation markers. Our findings suggest that miR-331-3p has an important role in regulating cervical cancer cell proliferation, and that miR-331-3p may contribute to keratinocyte differentiation through NRP2 suppression. miR-331-3p and NRP2 may contribute to anti-cancer effects. [ABSTRACT FROM AUTHOR]