scholarly journals Circ-0010928 Negatively Regulates Hypoxia-Induced Endothelial Cell Angiogenesis Through The miR-921/LSM14A axis.

2021 ◽  
Author(s):  
Tian Rong Zhang ◽  
WeiQiang Huang
Keyword(s):  
Endothelial Cells ◽  
Scratch Test ◽  
Tube Formation ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Downstream Target ◽  
Hypoxic Conditions ◽  
Downstream Target Gene ◽  
Reporter Assays

Abstract Background Angiogenesis is an important factor in promoting vascular repair and a valuable process in the treatment of cardiovascular diseases. Circular RNAs (circRNAs) are widely expressed in eukaryotic cells and play an important role in the regulation of endothelial cells (ECs). In our study, bioinformatics analysis and real-time fluorescent PCR detection revealed that circRNA 0010928 (circ-0010928) is differentially expressed in human cardiac microvascular endothelial cells (HCMECs). Material & Methods We evaluated the role of circ-0010928 in HCMECs. Then, we can verify the function of circ-0010928 in HCMECs by cell counting kit-8 (CCK8), scratch test, transwell experiment, tube forming experiment, flow cytometry. Use dual luciferase experiment to detect the binding relationship between circ-0010928, miR-921 and LSM14A. Results Overexpression of circ-0010928 inhibited the proliferation, migration and tube formation of HCMECs under hypoxic conditions and promoted their apoptosis. In addition, dual luciferase reporter assays confirmed that circ-0010928 acted as a sponge of miR-921 and LSM14A as a downstream target gene of miR-921. Silencing miR-921 could also inhibit the proliferation, migration and tube formation of HCMECs and negatively regulate angiogenesis. Conclusion CircRNA-0010928 may inhibit the function of miRNA-921by combining with miRNA-921, and then miRNA-921 plays a role in regulating LSM14A, thereby regulating the state of angiogenesis.

scholarly journals MiR-375-3p regulates rat pulmonary microvascular endothelial cell activity by targeting Notch1 during hypoxia

2020 ◽  
Vol 48 (7) ◽  
pp. 030006052092685
Author(s):  
Yuan An ◽  
Ziquan Liu ◽  
Hui Ding ◽  
Qi Lv ◽  
Haojun Fan ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Cell Activity ◽  
Tube Formation ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Microvascular Endothelial Cell ◽  
Adaptation To Hypoxia ◽  
Pulmonary Microvascular Endothelial Cell ◽  
Reporter Assays ◽  
Microvascular Endothelial

Objective Pulmonary microvascular endothelial cells (PMECs) exhibit specific responses in adaptation to hypoxia. However, the mechanisms regulating PMEC activities during hypoxia remain unclear. This study investigated the potential involvement of a microRNA, miR-375-3p, in the regulation of PMEC activities. Methods Primary PMECs were isolated from rats. The expression levels of miR-375-3p and Notch1 in the PMECs were detected by quantitative PCR and western blotting. Luciferase reporter assays were performed to explore the transcriptional regulation of Notch1 by miR-375-3p. The proliferation and chemotaxis of the PMECs were measured with the Cell Counting Kit-8 and Transwell invasion assays, respectively. Additionally, the capacity of hypoxia-treated PMECs for angiogenesis and inflammatory response was determined with tube formation assays and ELISA, respectively. Results The expression of miR-375-3p and Notch1 in the PMECs was significantly down-regulated and up-regulated during hypoxia, respectively. The results demonstrated that miR-375-3p directly targets Notch1 in PMECs, thereby suppressing the transcriptional expression of Notch1. It was further revealed that miR-375-3p regulates the proliferation, chemotaxis, angiogenesis, and inflammatory response of PMECs. Conclusions Our findings revealed the important role of miR-375-3p in the regulation of PMEC function and suggest the potential involvement of miR-375-3p in the development of lung diseases.

scholarly journals Novel circular RNA circNF1 acts as a molecular sponge, promoting gastric cancer by absorbing miR-16

2019 ◽  
Vol 26 (3) ◽  
pp. 265-277 ◽  
Author(s):  
Zhe Wang ◽  
Ke Ma ◽  
Steffie Pitts ◽  
Yulan Cheng ◽  
Xi Liu ◽  
...  
Keyword(s):  
Cell Lines ◽  
Circular Rna ◽  
Gastric Carcinogenesis ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Limited Information ◽  
Sequencing Data ◽  
Downstream Target ◽  
New Class ◽  
Reporter Assays

Circular RNAs (circRNAs) are a new class of RNA involved in multiple human malignancies. However, limited information exists regarding the involvement of circRNAs in gastric carcinoma (GC). Therefore, we sought to identify novel circRNAs, their functions and mechanisms in gastric carcinogenesis. We analyzed next-generation RNA sequencing data from GC tissues and cell lines, identifying 75,201 candidate circRNAs. Among these, we focused on one novel circRNA, circNF1 , which was upregulated in GC tissues and cell lines. Loss- and gain-of-function studies demonstrated that circNF1 significantly promotes cell proliferation. Furthermore, luciferase reporter assays showed that circNF1 binds to miR-16, thereby derepressing its downstream target mRNAs, MAP7 and AKT3. Targeted silencing or overexpression of circNF1 had no effect on levels of its linear RNA counterpart, NF1. Taken together, these results suggest that circNF1 acts as a novel oncogenic circRNA in GC by functioning as a miR-16 sponge.

scholarly journals Hsa_circRNA_103124 Upregulation in Crohn’s Disease Promotes Cell Proliferation and Inhibits Autophagy by Regulating the Hsa-miR-650/AKT2 Signaling Pathway

2021 ◽  
Vol 12 ◽  
Author(s):  
Juan Yin ◽  
Fuyi Tong ◽  
Yulan Ye ◽  
Tong Hu ◽  
Lijuan Xu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Signaling Pathway ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Downstream Target ◽  
Reverse Transcription Pcr ◽  
Reporter Assays ◽  
Pathway Analyses

Circular RNAs (circRNAs) play important roles in the pathogenesis of Crohn’s disease (CD). We discovered that hsa_circRNA_103124 was upregulated in CD patients in our previous study. Nonetheless, the function of hsa_circRNA_103124 is unclear. In this study, hsa_circRNA_103124 was predicted to interact with hsa-miR-650. Gene Ontology (GO) and pathway analyses identified AKT serine/threonine kinase 2 (AKT2) as the downstream target protein of hsa-miR-650. Activated AKT2 inhibits autophagy, but promotes cell proliferation. Recent studies suggest that the inhibition of autophagy is one of the mechanisms of CD pathogenesis. Therefore, we inferred that hsa_circRNA_103124 might regulate autophagy and proliferation by targeting AKT2 as a sponge for hsa-miR-650. Here, quantitative reverse transcription PCR (RT-QPCR) results revealed that upregulated hsa_circRNA_103124 expression in patients with CD was negatively correlated with hsa-miR-650 expression but positively correlated with the white blood cell count and calprotectin levels. TSC complex subunit 1 (TSC1), one of the proteins upstream of autophagy was downregulated in patients with CD. Consisting with the bioinformatics prediction, it was verified that hsa_circRNA_103124 targeted to hsa-miR650 by fluorescence in situ hybridization (FISH) and luciferase reporter assays. A hsa-miR-650 inhibitor reversed the promotion of rapamycin-induced autophagy and the inhibition of cell proliferation by the hsa_circRNA_103124 siRNA. However, hsa-miR-650 mimics reversed the inhibition of rapamycin-induced autophagy and the promotion of cell proliferation through hsa_circRNA_103124 overexpression. These results indicate that hsa_circRNA_103124 upregulation in patients with CD promotes cell proliferation and inhibits autophagy by regulating the hsa-miR-650/AKT2 signaling pathway.

scholarly journals CircRtn4 Acts as the Sponge of miR-24-3p to Promote Neurite Growth by Regulating CHD5

2021 ◽  
Vol 14 ◽  
Author(s):  
Yue Qi ◽  
Nana Ma ◽  
Xiaofan Chen ◽  
Yue Wang ◽  
Wei Zhang ◽  
...  
Keyword(s):  
Neurite Growth ◽  
Luciferase Reporter ◽  
Mammalian Brain ◽  
Circular Rnas ◽  
Neurite Length ◽  
Rna Molecules ◽  
Downstream Target ◽  
Downstream Target Gene ◽  
Precursor Mrna ◽  
Positive Effect

Circular RNAs (circRNAs) are covalently closed single-stranded RNA molecules. After derived from precursor mRNA back-splicing, circRNAs play important roles in many biological processes. Recently, it was shown that several circRNAs were enriched in the mammalian brain with unclear functions. The expression of circRtn4 in the mouse brain was increased with the differentiation of primary neurons. In our study, knockdown of circRtn4 inhibited neurite growth, while overexpression of circRtn4 significantly increased neurite length. By dual-luciferase reporter assay and RNA antisense purification assay, circRtn4 was identified as a miRNA sponge for miR-24-3p. Moreover, knockdown of miR-24-3p increased neurite length, while overexpression of miR-24-3p significantly inhibited neurite growth. Furthermore, CHD5 was confirmed to be a downstream target gene of miR-24-3p. And CHD5 silence counteracted the positive effect of circRtn4 overexpression on neurite growth. In conclusion, circRtn4 may act as the sponge for miR-24-3p to promote neurite growth by regulating CHD5.

scholarly journals Exosomal let-7f-5p Derived from Mineralized Osteoblasts Promotes the Angiogenesis of Endothelial Cells via DUSP1 / Erk1/2 Signaling Pathway

2021 ◽  
Author(s):  
Yiqun He ◽  
Hailong Li ◽  
Zuochong Yu ◽  
Linli Li ◽  
Xujun Chen ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Endothelial Cells ◽  
Bone Formation ◽  
Signaling Pathway ◽  
Target Genes ◽  
Tube Formation ◽  
Luciferase Reporter ◽  
Promising Target ◽  
Reporter Assays ◽  
Migration Assays

Abstract Background: Angiogenesis is essential for the tissue engineering bone formation, and osteoblasts (OBs) has been proved to play an important role in angiogenesis via various pro-angiogenic factors. However, whether the mineralized osteoblast derived exosomes (MOB-Exos) and containing let-7f-5p can promote the angiogenesis of endothelial cells (ECs) is still unknown.Methods: MOB-Exos, let-7f-5p mimicked MOB-Exos (miR mimic group) and let-7f-5p inhibited MOB-Exos (miR inhibitor group) were respectively harvested from mineralized osteoblasts (MOBs) and then co-cultured with bEnd.3. Besides, the Erk1/2 signaling pathway in ECs in miR mimic group was inhibited. Subsequently, CCK-8 assays, wound healing assays, transwell migration assays and tube formation assays were performed to detect the angiogenic capability of ECs. Dual luciferase reporter assays were conducted to verify the target genes of exosomal let-7f-5p. Results: The results showed that MOB-Exos could significantly promote the angiogenesis of ECs, which could be enhanced by mimicking exosomal let-7f-5p, and attenuated by inhibiting exosomal let-7f-5p. And the angiogenic capability of ECs was partly impaired after inhibiting the Erk1/2 signaling pathway despite co-cultured with let-7f-5p mimicked MOB-Exos. Moreover, let-7f-5p suppressed the luciferase activity of wide-type DUSP1, while mutation of DUSP1 abrogated the repressive ability of let-7f-5p. Conclusion: Based the results, our study concluded that exosomal let-7f-5p derived from MOBs could promote the angiogenesis of ECs via activating DUSP1/Erk1/2 signaling pathway, which might be a promising target for tissue engineering bone formation.

LncRNA MALAT1 inhibits hypoxia/reoxygenation-induced human umbilical vein endothelial cell injury via targeting the microRNA-320a/RAC1 axis

2020 ◽  
Vol 401 (3) ◽  
pp. 349-360 ◽  
Author(s):  
Rongrong Zhu ◽  
Xiao Hu ◽  
Wei Xu ◽  
Zhourui Wu ◽  
Yanjing Zhu ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Cell Injury ◽  
Umbilical Vein ◽  
Tube Formation ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Human Umbilical Vein ◽  
Rna Immunoprecipitation ◽  
Reporter Assays ◽  
Hypoxia Reoxygenation

AbstractAngiogenesis is believed to protect against hypoxia/reoxygenation (H/R)-induced cell injury. MALAT1 and microRNA-320a (miR-320a) are involved in cancer angiogenesis. To investigate the function of the MALAT1/miR-320a axis in H/R-induced cell injury, human umbilical vein endothelial cell (HUVEC) angiogenesis was detected using the Cell Counting Kit-8 (CCK-8), Transwell migration, cell adhesion and tube formation assays. The expression of MALAT1 and miR-320a was revealed by quantitative reverse transcription polymerase chain reaction (qRT-PCR). The direct binding relationship between miR-320a and MALAT1 was detected by RNA immunoprecipitation (RIP) and dual luciferase reporter assays. The data indicated that H/R induces angiogenesis injury and that the expression of MALAT1 was augmented in H/R-stimulated HUVECs. Overexpression of MALAT1 alleviated H/R-stimulated HUVEC dysfunction, whereas silencing of MALAT1 exerted the opposite effects. MALAT1 also reduced miR-320a levels in HUVECs. Overexpression of miR-320a repressed the function of MALAT1 on H/R-stimulated HUVECs, whereas inhibition of miR-320a exerted the opposite effect. Additionally, miR-320a inhibition alleviated H/R-stimulated HUVEC injury via RAC1. Taken together, this investigation concluded that MALAT1 represses H/R-stimulated HUVEC injury by targeting the miR-320a/RAC1 axis.

scholarly journals LncRNA HAS2-AS1 Promotes Glioblastoma Proliferation by Sponging miR-137

2021 ◽  
Vol 11 ◽  
Author(s):  
Yalin Lu ◽  
Gaochao Guo ◽  
Rujun Hong ◽  
Xingjie Chen ◽  
Yan Sun ◽  
...  
Keyword(s):  
Underlying Mechanism ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Tumor Type ◽  
Downstream Target ◽  
Qrt Pcr ◽  
Lysine Specific Demethylase ◽  
Reporter Assays

GBM (Glioblastoma multiform) is the most malignant tumor type of the central nervous system and has poor diagnostic and clinical outcomes. LncRNAs (Long non-coding RNAs) have been reported to participate in multiple biological and pathological processes, but their underlying mechanism remains poorly understood. Here, we aimed to explore the role of the lncRNA HAS2-AS1 (HAS2 antisense RNA 1) in GBM. GSE103227 was analyzed, and qRT-PCR was performed to measure the expression of HAS2-AS1 in GBM. FISH (Fluorescence in situ hybridization) was performed to verify the localization of HAS2-AS1. The interaction between HAS2-AS1 and miR-137 (microRNA-137) was predicted by LncBook and miRcode followed by dual‐luciferase reporter assays, and the relationships among HAS2-AS1, miR-137 and LSD1 (lysine-specific demethylase 1) were assessed by WB (western blot) and qRT-PCR. Colony formation and CCK-8 (cell counting kit-8) assays were performed as functional tests. In vivo, nude mice were used to confirm the function of HAS2-AS1. HAS2-AS1 expression was upregulated in GBM cell lines, and HAS2-AS1 was localized mainly in the cytoplasm. In vitro, high HAS2-AS1 expression promoted proliferation, and knockdown of HAS2-AS1 significantly inhibited proliferation. Furthermore, HAS2-AS1 functioned as a ceRNA (competing endogenous RNA) of miR-137, leading to the disinhibition of its downstream target LSD1. The miR-137 level was downregulated by HAS2-AS1 overexpression and upregulated by HAS2-AS1 knockdown. In a subsequent study, LSD1 expression was negatively regulated by miR-137, while miR-137 reversed the LSD1 expression levels caused by HAS2-AS1. These results were further supported by the nude mouse tumorigenesis experiment; compared with xenografts with high HAS2-AS1 expression, the group with low levels of HAS2-AS1 exhibited suppressed proliferation and better survival. We conclude that lncRNA HAS2-AS1 promotes proliferation by functioning as a miR‐137 decoy to increase LSD1 levels and thus might be a possible biomarker for GBM.

scholarly journals Knockdown of Circ_SLC39A8 protects against the progression of osteoarthritis by regulating miR-591/IRAK3 axis

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Jizhe Yu ◽  
Yushuang Qin ◽  
Naxin Zhou
Keyword(s):  
Cell Viability ◽  
Molecular Mechanisms ◽  
Interleukin 1 ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Circular Rnas ◽  
Oa Chondrocytes ◽  
Polymerase Chain ◽  
Apoptosis And Inflammation ◽  
The Relationship

Abstract Background The dysregulation of circular RNAs (circRNAs) has been identified in various human diseases, including osteoarthritis (OA). The purpose of this study was to identify the role and mechanism of circ_SLC39A8 in regulating the progression of OA. Methods The expression levels of circ_SLC39A8, miR-591, and its potential target gene, interleukin-1-receptor-associated kinase 3 (IRAK3), were identified by quantitative real-time polymerase chain reaction (qRT-PCR). Cell viability and apoptosis were determined by Cell Counting Kit-8 (CCK-8) assay and flow cytometry, respectively. The relationship between miR-591 and circ_SLC39A8 or IRAK3 was predicted by bioinformatics tools and verified by dual-luciferase reporter. Results Circ_SLC39A8 and IRAK3 were upregulated and miR-591 was downregulated in OA cartilage tissues. Knockdown of circ_SLC39A8 inhibited apoptosis and inflammation in OA chondrocytes, while these effects were reversed by downregulating miR-591. Promotion cell viability effects of miR-591 were partially reversed by IRAK3 overexpression. Conclusion Our findings indicated that knockdown of circ_SLC39A8 delayed the progression of OA via modulating the miR-591-IRAK3 axis, providing new insight into the molecular mechanisms of OA pathogenesis.

scholarly journals Warburg effect-promoted exosomal circ_0072083 releasing up-regulates NANGO expression through multiple pathways and enhances temozolomide resistance in glioma

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Chenyu Ding ◽  
Xuehan Yi ◽  
Xiangrong Chen ◽  
Zanyi Wu ◽  
Honghai You ◽  
...  
Keyword(s):  
Warburg Effect ◽  
Lactate Production ◽  
Xenograft Model ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Circular Rnas ◽  
Rna Immunoprecipitation ◽  
Resistant Cells

Abstract Background Temozolomide (TMZ) resistance limits its application in glioma. Exosome can carry circular RNAs (circRNAs) to regulate drug resistance via sponging microRNAs (miRNAs). miRNAs can control mRNA expression by regulate the interaction with 3’UTR and methylation. Nanog homeobox (NANOG) is an important biomarker for TMZ resistance. Hitherto, it is unknown about the role of exosomal hsa_circ_0072083 (circ_0072083) in TMZ resistance in glioma, and whether it is associated with NANOG via regulating miRNA sponge and methylation. Methods TMZ-resistant (n = 36) and sensitive (n = 33) patients were recruited. The sensitive cells and constructed resistant cells were cultured and exposed to TMZ. circ_0072083, miR-1252-5p, AlkB homolog H5 (ALKBH5) and NANOG levels were examined via quantitative reverse transcription polymerase chain reaction and western blot. The half maximal inhibitory concentration (IC50) of TMZ, cell proliferation, apoptosis, migration and invasion were analyzed via Cell Counting Kit-8, colony formation, flow cytometry, wound healing and transwell assays. The in vivo function was assessed using xenograft model. The N6-methyladenosine (m6A) level was analyzed via methylated RNA immunoprecipitation (MeRIP). Target relationship was investigated via dual-luciferase reporter assay and RNA immunoprecipitation. Warburg effect was investigated via lactate production, glucose uptake and key enzymes expression. Exosome was isolated and confirmed via transmission electron microscopy and specific protein expression. Results circ_0072083 expression was increased in TMZ-resistant glioma tissues and cells. circ_0072083 knockdown restrained the resistance of resistant cells via decreasing IC50 of TMZ, proliferation, migration, invasion and xenograft tumor growth and increasing apoptosis. circ_0072083 silence reduced NANOG expression via blocking ALKBH5-mediated demethylation. circ_0072083 could regulate NANOG and ALKBH5 via targeting miR-1252-5p to control TMZ resistance. Warburg effect promoted the release of exosomal circ_0072083 in resistant cells. Exosomal circ_0072083 from resistant cells increased the resistance of sensitive cells to TMZ in vitro and xenograft model. Exosomal circ_0072083 level was enhanced in resistant patients, and it had a diagnostic value and indicated a lower overall survival in glioma. Conclusion Exosomal circ_0072083 promoted TMZ resistance via increasing NANOG via regulating miR-1252-5p-mediated degradation and demethylation in glioma.

scholarly journals Circ_0003266 sponges miR-503-5p to suppress colorectal cancer progression via regulating PDCD4 expression

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Caihong Wen ◽  
Xiaoqing Feng ◽  
Honggang Yuan ◽  
Yong Gong ◽  
Guangsheng Wang
Keyword(s):  
Colorectal Cancer ◽  
Cell Proliferation ◽  
Cell Lines ◽  
Cancer Progression ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Circular Rnas ◽  
Pdcd4 Expression ◽  
Novel Target

Abstract Background Circular RNAs (circRNAs) feature prominently in tumor progression. However, the biological function and molecular mechanism of circ_0003266 in colorectal cancer (CRC) require further investigation. Methods Circ_0003266 expression in 46 pairs CRC tissues / adjacent tissues, and CRC cell lines was detected by quantitative real-time polymerase chain reaction (qRT-PCR); after circ_0003266 was overexpressed or knocked down in CRC cells, cell proliferation, apoptosis, migration, and invasion were evaluated by the cell counting kit-8 (CCK-8), flow cytometry, and Transwell assays, respectively; the interaction among circ_0003266, miR-503-5p, and programmed cell death 4 (PDCD4) was confirmed using bioinformatics analysis and dual-luciferase reporter assay; PDCD4 protein expression in CRC cells was quantified using Western blot. Results Circ_0003266 was significantly lowly expressed in CRC tissues and cell lines. Circ_0003266 overexpression markedly repressed CRC cell proliferation, migration, and invasion, and accelerated the cell apoptosis, but its overexpression promoted the malignant phenotypes of CRC cells. PDCD4 was a direct target of miR-503-5p and circ_0003266 promoted PDCD4 expression by competitively sponging miR-503-5p. Conclusion Circ_0003266 suppresses the CRC progression via sponging miR-503-5p and regulating PDCD4 expressions, which suggests that circ_0003266 may serve as a novel target for the treatment of CRC.

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