Journal: Cell Death & Disease

Article Title: Circular RNA circUBE2J2 acts as the sponge of microRNA-370-5P to suppress hepatocellular carcinoma progression

doi: 10.1038/s41419-021-04269-4

Figure Lengend Snippet: A The cluster heat map showed the differentially expressed circRNAs in HCC tissues compared with those in adjacent nontumor tissues. Red color indicates high expression level, and green color indicates low expression level. B Volcano plots illustrate that among significantly different expressed circRNAs in HCC tissues relative to normal tissues. The green, red, and black points represent downregulated, upregulated, and no statistically significant difference circular RNAs (circRNAs), respectively. x -axis: log2 ratio of circRNA expression levels between normal and tumor tissues. y -axis: the P value (−log10 transformed) of circRNAs. C Upper panel: illustration of the annotated genomic region of UBE2J2 , the putative different RNA splicing forms, and scheme illustrating the production of circUBE2J2. Convergent (blue) and divergent (red) primers were designed to amplify the linear or back-splicing products. Lower panel: total RNA from HepG2 cells with or without RNase-R treatment were subjected to polymerase chain reaction (PCR). D qRT-PCR analysis of circUBE2J2 and mUBE2J2 RNA after treatment with Actinomycin D at the indicated time points in HCC cells. E qRT-PCR detection show the differential expression of circUBE2J2 in 75 paired HCC tissues. F CircUBE2J2 expression among 75 patients was analyzed for survival probability . G The expression of circUBE2J2 was analyzed by in situ hybridization on HCC tissue. The representative photomicrographs of circUBE2J2 level with high, equal, and low in HCC tumors, as compared with normal tissues, were shown. The altered level of circUBE2J2 between HCC and non-tumor tissues in 50 HCC patients were summarized ( n = number of cases). Data are presented as mean ± SD. * p < 0.05, ** p < 0.01; NS no significance.

Article Snippet: Five pairs of HCC samples (tumor tissues and matched non-tumor tissues) were analyzed by Arraystar Human circRNA Array.

Techniques: Expressing, Transformation Assay, Polymerase Chain Reaction, Quantitative RT-PCR, In Situ Hybridization

Journal: Cell Death & Disease

Article Title: Circular RNA circUBE2J2 acts as the sponge of microRNA-370-5P to suppress hepatocellular carcinoma progression

doi: 10.1038/s41419-021-04269-4

Figure Lengend Snippet: A RAP assays were performed using an biotin probe against circUBE2J2 on extracts from HepG2 cells. Relative expression levels of circRNA were evaluated by qRT-PCR. B The relative expression level of potential target miRNA of circUBE2J2 was assessed by qRT-PCR. C , D RIP experiments were performed using an antibody against AGO2 on extracts from HLF and HepG2 cells. E Schematic diagram of reporter gene structure. F A schematic drawing showing the putative binding sites. The mutant version of circUBE2J2 is presented. G , H Relative luciferase activity determined 48 h after transfecting HEK293T cells ( G ) and HepG2 cells ( H ) with miR-370-5P mimic/NC or circUBE2J2 WT/Mut. WT wild type, Mut, mutant-type. I The expression of circUBE2J2 in MHCC97H, HLF, and HepG2 cells after transfection with miR-370-5P mimcs or miR-370-5P inhibitor. J The expression of miR-370-5P in HepG2, HLF, and MHCC97H cells after transfection with circUBE2J2-OE or circUBE2J2 shRNA. K RAP assay were performed using an probe against miR370-5P on extracts from HepG2 cells. Relative expression levels of circRNA were evaluated by qRT-PCR. L FISH assay analysis for the co-localization of miR-370-5P and circUBE2J2 in HCC cells. M miR-370-5P co-localized with circUBE2J2 in HCC adjacent non-tumor and tumor tissues was detected by FISH. Data are from three independent experiments (mean ± SEM). The data are represented as the mean ± SD, n = 3. * p < 0.05; ** p < 0.01; NS no significance.

Article Snippet: Five pairs of HCC samples (tumor tissues and matched non-tumor tissues) were analyzed by Arraystar Human circRNA Array.

Techniques: Expressing, Quantitative RT-PCR, Binding Assay, Mutagenesis, Luciferase, Activity Assay, Transfection, shRNA

Journal: Molecular Cancer

Article Title: Circular RNA cTFRC acts as the sponge of MicroRNA-107 to promote bladder carcinoma progression

doi: 10.1186/s12943-019-0951-0

Figure Lengend Snippet: Up-regulated circRNAs in BC tumor tissues and its correlation with prognosis of patients. a cTFRC increased in BC tissues as compared to that in the matched nontumor tissues analyzed by circRNAs Arraystar Chip. b Schematic representation of the high expression level of cTFRC in 57 BC patients tissues compared with adjacent normal patients tissues by qPCR. c cTFRC upregulated in recurrent BC patients. d The high expression levels of cTFRC in BC patients with high grade. e Advanced T stage is associated with higher cTFRC levels. f The expression of cTFRC higher in patients with lymphatic metastasis. g Prognostic significance of cTFRC expression for BC patients was performed with cTFRC values by using the median value as the cutoff

Article Snippet: Fig. 1 Up-regulated circRNAs in BC tumor tissues and its correlation with prognosis of patients. a cTFRC increased in BC tissues as compared to that in the matched nontumor tissues analyzed by circRNAs Arraystar Chip. b Schematic representation of the high expression level of cTFRC in 57 BC patients tissues compared with adjacent normal patients tissues by qPCR. c cTFRC upregulated in recurrent BC patients. d The high expression levels of cTFRC in BC patients with high grade. e Advanced T stage is associated with higher cTFRC levels. f The expression of cTFRC higher in patients with lymphatic metastasis. g Prognostic significance of cTFRC expression for BC patients was performed with cTFRC values by using the median value as the cutoff

Techniques: Expressing

Journal: Molecular Therapy. Nucleic Acids

Article Title: The circular RNA Ataxia Telangiectasia Mutated regulates oxidative stress in smooth muscle cells in expanding abdominal aortic aneurysms

doi: 10.1016/j.omtn.2023.08.017

Figure Lengend Snippet: Circular RNAs are deregulated in human abdominal aortic aneurysm (A) Volcano plot depicting downregulated (51, blue) and upregulated (40, red). Circular RNAs (circRNAs) in human elective human abdominal aortic aneurysm (eAAA, n = 11) vs . control (CTRL, n = 6) aorta specimens, as resulted by array experiments. Log2 fold change and -log10 p value are plotted on the x and y axes, respectively. IDs of circRNAs meant for a first round of validation are highlighted. Statistics: unpaired t test; p values <0.05 are considered significant. (B) Pie chart illustrating the proportion of exonic (89.8%), intronic (5.7%), sense overlapping (3.4%), and antisense (1.1%) array-identified differentially expressed circRNAs. Absolute numbers are further indicated for each group. (C) Real-time quantitative PCR (qPCR) validation of hsa_circ_0005660 (c NFIX ), hsa_circ_0003641 (c ATM ), hsa_circ0042103 (c MYOCD ), hsa_circ003218 (c BMPR2 ), hsa_circ0004771 (c NRIP1 ), and hsa_circ0005615 (c NFATC3 ) differential expression in human eAAA (N = 8) and CTRL (N = 4) aortas. 2 –ddCT was calculated by normalizing on RPLPO . Data are represented as mean ± SEM. Statistics: unpaired t test; p values <0.05 are considered significant. NS, not significant; eAAA, elective AAA.

Article Snippet: The resulting labeled cDNA was then purified and 1 μg was fragmented, heated, and subsequently hybridized with an 8 × 15k commercially available array chip displaying 13,617 human circRNAs (Arraystar, no. AS-S-CR-H-V2.0) for 17 h at 65°C in an Agilent Hybridization Oven.

Techniques: Control, Biomarker Discovery, Real-time Polymerase Chain Reaction, Quantitative Proteomics

Journal: Frontiers in Oncology

Article Title: CircRNA_036186 mediates HNSCC progression by regulating 14-3-3ζ

doi: 10.3389/fonc.2024.1498139

Figure Lengend Snippet: Identification of circRNA_036186-miR-193b-3p-14-3-3ζ-regulatory axis. (A) The heatmap illustrates the expression profiles of 287 circRNAs, of which 146 exhibited increased expression and 141 exhibited decreased expression. The colouration gradually transitions from blue to red, indicating increased expression. (B) The five most highly ranked circRNAs and their target miRNAs, each corresponding to a node, are shown with two interacting genes based on base sequence pairing connected by a solid line. (C) The intersection results of the HNSCC occurrence and development group, the prognosis group, and the competing endogenous RNA networks in OncomiR. (D) The predicted target mRNAs from three databases—Wayne ’ s map, miR-193b-3pmicroT-CDS, Tarbase, and TargetScan—were taken as intersections. (E) A search of the Tarbase database revealed that miR-193b-3p has a base sequence that binds and interacts with the 14-3-3ζ target. (F, G) The results of the RT-PCR experiments indicated that circRNA_036186 and miR-193b-3p exhibited elevated expression in HNSCC tissues relative to paracancerous tissues in five pairs of HNSCC and paracancerous tissue samples. (H, I) The results of RT-PCR experiments indicated that the mRNA expression of circRNA_036186 and miR-193b-3p was markedly elevated in all three HNSCC cell lines relative to HOK. (n=3, * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001).

Article Snippet: We used the Arratstar Super RNA Labeling Kit to amplify the enriched circRNA, which was then transcribed into fluorescent cRNA.The labelled cRNA was hybridised to the Arraystar Human circRNA V2 chip.

Techniques: Expressing, Sequencing, Reverse Transcription Polymerase Chain Reaction

Journal: Frontiers in Oncology

Article Title: CircRNA_036186 mediates HNSCC progression by regulating 14-3-3ζ

doi: 10.3389/fonc.2024.1498139

Figure Lengend Snippet: Top 5 circRNA.

Article Snippet: We used the Arratstar Super RNA Labeling Kit to amplify the enriched circRNA, which was then transcribed into fluorescent cRNA.The labelled cRNA was hybridised to the Arraystar Human circRNA V2 chip.

Techniques:

Journal: Frontiers in Oncology

Article Title: CircRNA_036186 mediates HNSCC progression by regulating 14-3-3ζ

doi: 10.3389/fonc.2024.1498139

Figure Lengend Snippet: Effect of down-regulation of circRNA_0361863p and miR-193b-3p on 14-3-3ζ in HSC2. (A) The results of the RT-PCR experiment indicated that si-circ_0036186 had been successfully transfected into HSC2. (B) The results of the RT-PCR experiments indicated that miR-193b-3p had been successfully transfected into HSC2. (C) Reverse transcription polymerase chain reaction (RT-PCR) experiments were conducted to ascertain the impact of transfection on 14-3-3ζ mRNA expression. (D, E) Western blotting experiments were conducted to ascertain the impact of transfection on 14-3-3ζ protein expression. (n=3, * p <0.05, *** p <0.001, **** p <0.0001).

Article Snippet: We used the Arratstar Super RNA Labeling Kit to amplify the enriched circRNA, which was then transcribed into fluorescent cRNA.The labelled cRNA was hybridised to the Arraystar Human circRNA V2 chip.

Techniques: Reverse Transcription Polymerase Chain Reaction, Transfection, Reverse Transcription, Polymerase Chain Reaction, Expressing, Western Blot

Journal: Frontiers in Oncology

Article Title: CircRNA_036186 mediates HNSCC progression by regulating 14-3-3ζ

doi: 10.3389/fonc.2024.1498139

Figure Lengend Snippet: Effect of down-regulation of circRNA_0361863p and miR-193b-3p on the proliferation, migration, invasion, and scratch healing rate of HSC2 cells. (A–D) The Transwell assay assessed the impact of circRNA_0361863p and miR-193b-3p down-regulation on cell migration and invasion of HSC2. (E, F) The Scratch test assessed the scratch healing rate of HSC2 after the down-regulation of circRNA_0361863p and miR-193b-3p. (G) The cell activity of HSC2 was evaluated after the down-regulation of circRNA_0361863p and miR-193b-3p through CCK-8 experiments. (n=3, **** p <0.0001).

Article Snippet: We used the Arratstar Super RNA Labeling Kit to amplify the enriched circRNA, which was then transcribed into fluorescent cRNA.The labelled cRNA was hybridised to the Arraystar Human circRNA V2 chip.

Techniques: Migration, Transwell Assay, Activity Assay, CCK-8 Assay