Journal: bioRxiv

Article Title: The synergistic effect of circRNA methylation promotes pulmonary fibrosis

doi: 10.1101/2021.08.05.455186

Figure Lengend Snippet: m 6 A modification of circRNAs in silicosis. A Arraystar m 6 A-circRNA epitranscriptomic microarray flow. B The heat map shows the difference in the level of m 6 A-modified circRNAs, and the red and blue strips indicate increased and decreased m 6 A levels, respectively. C The volcanic map shows the difference in the level of m 6 A-modified circRNAs. D Heat map showing the difference in the quantity of m 6 A-modified circRNAs. The red and blue strips indicate circRNAs with upregulated and downregulated expression, respectively. E The volcano map shows the difference in the quantity of m 6 A-modified circRNAs.

Article Snippet: First, an Arraystar m 6 A-circRNA Epitranscriptomic microarray was used to investigate the differences in m 6 A modification in circRNAs in the lung tissues of mice in the silicosis and control groups ( ).

Techniques: Modification, Microarray, Expressing

Journal: bioRxiv

Article Title: The synergistic effect of circRNA methylation promotes pulmonary fibrosis

doi: 10.1101/2021.08.05.455186

Figure Lengend Snippet: Preliminary screening of circRNAs. A The total RNA m 6 A level in the lung tissues of mice was detected using the EpiQuik m 6 A RNA Methylation Quantification Kit. B The EpiQuik m 6 A RNA Methylation Quantification Kit detects total RNA m 6 A levels in human pulmonary fibroblasts treated with SiO 2 for different periods. C Screening process of circRNAs; the basic expression levels of 10 circRNAs in human pulmonary fibroblasts were detected by PCR. D Analysis of three circRNA m 6 A sites. The data are presented as means±SD. *P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: First, an Arraystar m 6 A-circRNA Epitranscriptomic microarray was used to investigate the differences in m 6 A modification in circRNAs in the lung tissues of mice in the silicosis and control groups ( ).

Techniques: Methylation, Expressing

Journal: bioRxiv

Article Title: The synergistic effect of circRNA methylation promotes pulmonary fibrosis

doi: 10.1101/2021.08.05.455186

Figure Lengend Snippet: METTL3 expression is elevated in pulmonary fibrosis. A-B WB and q-PCR were used to detect changes in the five representative m 6 A modification enzymes in pulmonary fibroblasts. C The expression of METTL3 was detected by cellular immunofluorescence. Scale=30 μm. D The expression of METTL3 in the lung tissues of mice at 7 and 28 days after modeling was detected by immunofluorescence. Scale=50 μm. E The expression of METTL3 in the lung tissues of healthy individuals and silicosis patients was detected by immunofluorescence. Scale=50 μm. The data are presented as means±SD. *P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: First, an Arraystar m 6 A-circRNA Epitranscriptomic microarray was used to investigate the differences in m 6 A modification in circRNAs in the lung tissues of mice in the silicosis and control groups ( ).

Techniques: Expressing, Modification, Immunofluorescence

Journal: bioRxiv

Article Title: The synergistic effect of circRNA methylation promotes pulmonary fibrosis

doi: 10.1101/2021.08.05.455186

Figure Lengend Snippet: METTL3 mediates RNA m 6 A modification and promotes the activation, migration and activity of human pulmonary fibroblasts. A The EpiQuik m 6 A RNA Methylation Quantification Kit was used to detect total RNA m 6 A levels in human pulmonary fibroblasts after METTL3 knockdown. B-E WB was used to detect the effects of METTL3 knockdown on the human pulmonary fibroblast activation markers FN1, Collagen 1 and α-SMA. F-G The wound healing assay was used to detect the effect of METTL3 knockdown on the migration of human pulmonary fibroblasts. Scale=325 μm. H The CCK-8 assay was used to detect the effect of METTL3 knockdown on the activity of human pulmonary fibroblasts. I-L WB was used to detect the effects of the SAH inhibitor on the fibroblast activation markers FN1, Collagen 1 and α-SMA. M The CCK-8 assay was used to detect the effect of the SAH inhibitor on the viability of human pulmonary fibroblasts. The data are presented as means±SD. *P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: First, an Arraystar m 6 A-circRNA Epitranscriptomic microarray was used to investigate the differences in m 6 A modification in circRNAs in the lung tissues of mice in the silicosis and control groups ( ).

Techniques: Modification, Activation Assay, Migration, Activity Assay, Methylation, Wound Healing Assay, CCK-8 Assay

Journal: bioRxiv

Article Title: The synergistic effect of circRNA methylation promotes pulmonary fibrosis

doi: 10.1101/2021.08.05.455186

Figure Lengend Snippet: METTL3-mediated circRNA m 6 A modification promotes the activation of human pulmonary fibroblasts. A In situ hybridization detection of hsa_circ_0000672 and hsa_circ_0005654 colocalization with METTL3. Scale=20 μm. B RIP test of the interaction of METTL3 with hsa_circ_0000672 and hsa_circ_0005654. C Changes in circRNA m 6 A modification after METTL3 knockdown were detected by me-RIP. D PCR was used to detect the changes in hsa_circ_0000672 and hsa_circ_0005654 expression after knocking down METTL3. E Hsa_circ_0000672 and hsa_circ_0005654 with the m 6 A site mutated or wild-type overexpression plasmids were constructed. F-G The overexpression efficiency of wild-type and m 6 A site-mutated overexpression plasmids in human pulmonary fibroblasts was detected by PCR. H-K WB was conducted to detect the effect of the wild-type overexpression plasmid on the human pulmonary fibroblast activation markers FN1, Collagen 1 and α-SMA. I-O WB was conducted to detect the effect of the m 6 A site mutated overexpression plasmids on the human pulmonary fibroblast activation markers FN1, Collagen 1 and α-SMA. The data are presented as means±SD, *P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: First, an Arraystar m 6 A-circRNA Epitranscriptomic microarray was used to investigate the differences in m 6 A modification in circRNAs in the lung tissues of mice in the silicosis and control groups ( ).

Techniques: Modification, Activation Assay, In Situ Hybridization, Expressing, Over Expression, Construct, Plasmid Preparation

Journal: bioRxiv

Article Title: The synergistic effect of circRNA methylation promotes pulmonary fibrosis

doi: 10.1101/2021.08.05.455186

Figure Lengend Snippet: Schematic diagram showing the mechanisms through which silica induced m 6 A modification of hsa_circ_0000672 and hsa_circ_0005654, followed by activation of fibroblast.

Article Snippet: First, an Arraystar m 6 A-circRNA Epitranscriptomic microarray was used to investigate the differences in m 6 A modification in circRNAs in the lung tissues of mice in the silicosis and control groups ( ).

Techniques: Modification, Activation Assay