Journal: Cell systems

Article Title: Multiplex Enhancer Interference Reveals Collaborative Control of Gene Regulation by Estrogen Receptor Alpha Bound Enhancers

doi: 10.1016/j.cels.2017.08.011

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: DATA AND SOFTWARE AVAILABILITY Software: See Key Resources Table. table ft1 table-wrap mode="anchored" t5 caption a7 REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies H3K27ac rabbit polyclonal Active Motif Cat# 39133; RRID:AB_2561016 H3K9me3 rabbit polyclonal Abcam Cat# ab8898; RRID:AB_306848 FLAG mouse monoclonal Sigma-Aldrich Cat# F1804; RRID:AB_262044 RNA pol II 4H8 mouse monoclonal Abcam Cat# ab5408; RRID:AB_304868 ER alpha rabbit polyclonal Santa Cruz Cat#sc-544; RRID:AB_631469 Bacterial and Virus Strains Biological Samples Chemicals, Peptides, and Recombinant Proteins β-estradiol Sigma-Aldrich Cat# E2758 Trichostatin A Cayman Chemicals Cat# 89730; CAS 58880-19-6 Critical Commercial Assays ZR 96-well Quick-RNA Zymo Research Cat# R1053 KAPA Stranded mRNA-seq KAPA Biosystems Cat# KK8420 Power SYBR® Green RNA-to-CT™ 1-Step Applied Biosystems Cat# 4389986 Deposited Data Raw and analyzed ChIP-seq data This study {"type":"entrez-geo","attrs":{"text":"GSE99906","term_id":"99906"}} GSE99906 Raw and analyzed RNA-seq data This study {"type":"entrez-geo","attrs":{"text":"GSE99906","term_id":"99906"}} GSE99906 Experimental Models: Cell Lines Human: Ishikawa cells ECACC ECACC Cat# 99040201, RRID:CVCL_2529 Experimental Models: Organisms/Strains Ishikawa SID4x-dCas9-KRAB This study Oligonucleotides gBlocks, see Table S1 Primers for constructing and verifying dCas9 fusions and guide RNAs, see Table S2 Guide RNA targets, see Table S3 qPCR primers, see Table S4 Primers for sequencing ERBS deletions, see Table S5 Recombinant DNA dCas9-VP160 Cheng et al., 2013 Addgene 48227 pGL3-U6-PGK-Puro Shen et al., 2014 Addgene 51133 gRNA_cloningVector Mali et al., 2013 Addgene 41824 Software and Algorithms Bowtie Langmead et al., 2009 DOI: 10.1186/gb-2009-10-3-r25 MACS2 Zhang et al., 2008 DOI: 10.1186/gb-2008-9-9-r137 DESeq2 Love et al., 2014 DOI: 10.1186/s13059-014-0550-8 HISAT Kim et al., 2015 DOI: 10.1038/nmeth.3317 HTSeq Anders et al., 2015 DOI: 10.1093/bioinformatics/btu638 Bedtools Quinlan and Hall, 2010 DOI: 10.1093/bioinformatics/btq033 Prism 6 http://www.graphpad.com/scientific-software/prism/ Other Open in a separate window KEY RESOURCES TABLE Data produced in this study are available from GEO ( {"type":"entrez-geo","attrs":{"text":"GSE99906","term_id":"99906"}} GSE99906 ).

Techniques: Recombinant, SYBR Green Assay, Sequencing, Software

Journal: Cell Death & Disease

Article Title: USP12 promotes breast cancer angiogenesis by maintaining midkine stability

doi: 10.1038/s41419-021-04102-y

Figure Lengend Snippet: A The relative expression of VEGF-A and VEGF-C in the supernatant of MDA-MB-231 cells overexpressing USP12 was detected by ELISA. B The relative expression of VEGF-A and VEGF-C in the supernatant of MDA-MB-231 cells with USP12 knockdown was detected by ELISA. C , D Migration of HUVECs treated with supernatant from MDA-MB-231 cells with USP12 overexpression ( C ) and USP12 knockdown ( D ). Representative images are shown in the left panel. The quantitative results are shown in the right panel. E , F . Tube formation assay of HUVECs treated with supernatant from MDA-MB-231 cells with USP12 overexpression ( E ) and USP12 knockdown ( F ). Representative images are shown in the left panel. The quantitative results are shown in the right panel. The experiments were repeated three times. ** p < 0.01, *** p < 0.001; mean ± SD.

Article Snippet: Matrigel matrix (356234, Corning), the human VEGF-A ELISA kit (EKO588-96, Boster) and the human VEGF-C ELISA KIT (EKO539-96, Boster) were also used.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Migration, Over Expression, Tube Formation Assay

Journal: Frontiers in Pharmacology

Article Title: MiR-19a-3p Suppresses M1 Macrophage Polarization by Inhibiting STAT1/IRF1 Pathway

doi: 10.3389/fphar.2021.614044

Figure Lengend Snippet: List of primers used in the study.

Article Snippet: TNF-α, Chemokine (C-X-C motif) ligand 9 (CXCL9), CXCL10 in the supernatants were detected with mouse TNF-α (70-EK282/3-96), CXCL9 (70-EK2143/2-24), CXCL10 (70-EK268/2-96) ELISA Kit (MultiSciences, Hangzhou, China).

Techniques:

Journal: Frontiers in Pharmacology

Article Title: MiR-19a-3p Suppresses M1 Macrophage Polarization by Inhibiting STAT1/IRF1 Pathway

doi: 10.3389/fphar.2021.614044

Figure Lengend Snippet: STAT1 was up-regulated in M1 macrophages. (A) CXCL9, CXCL10, iNOS, and TNF-α were determined by qRT-PCR in control cells (RAW264.7 cells) and RAW264.7 cells treated with LPS and IFN-γ (data were pooled from three independent experiments). (B) CXCL9, CXCL10, TNF-α were exhibited by ELISA in RAW264.7 cells treated with LPS and IFN-γ and control cells (data were pooled from three independent experiments). (C) STAT1, iNOS and p-STAT1 protein levels were measured in control cells and LPS plus IFN-γ induced RAW264.7 cells by western blot (a representative blot, from three independent experiments). (D) qRT-PCR analysis of STAT1 expression in control cells and LPS plus IFN-γ induced RAW264.7 cells (data were pooled from three independent experiments). Statistical significance was calculated using unpaired Student’s t-test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: TNF-α, Chemokine (C-X-C motif) ligand 9 (CXCL9), CXCL10 in the supernatants were detected with mouse TNF-α (70-EK282/3-96), CXCL9 (70-EK2143/2-24), CXCL10 (70-EK268/2-96) ELISA Kit (MultiSciences, Hangzhou, China).

Techniques: Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot, Expressing

Journal: Frontiers in Pharmacology

Article Title: MiR-19a-3p Suppresses M1 Macrophage Polarization by Inhibiting STAT1/IRF1 Pathway

doi: 10.3389/fphar.2021.614044

Figure Lengend Snippet: Overexpressed miR-19a-3p suppresses RAW264.7 derived M1 polarization and down-regulated miR-19a-3p improves RAW264.7 derived M1 polarization. RAW264.7 cells transfected with miR-19a-3p mimics/NC or miR-19a-3p inhibitor/iNC for 24 h, then stimulated with LPS plus IFN-γ for 24 h. (A,B) miR-19a-3p was determined using qRT-PCR (data were pooled from three independent experiments). (C,D) TNF-α, CXCL9, CXCL10, iNOS were analyzed by qRT-PCR (data were pooled from three independent experiments). (E,F) TNF-α, CXCL9, CXCL10 were exhibited by ELISA (data were pooled from three independent experiments). (G,H) Western blot analysis the protein level of iNOS (a representative blot, from three independent experiments). NC: miR-19a-3p mimics negative control, iNC: miR-19a-3p inhibitor negative control. Statistical significance was calculated using unpaired Student’s t-test or ANOVA followed by Bonferroni test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: TNF-α, Chemokine (C-X-C motif) ligand 9 (CXCL9), CXCL10 in the supernatants were detected with mouse TNF-α (70-EK282/3-96), CXCL9 (70-EK2143/2-24), CXCL10 (70-EK268/2-96) ELISA Kit (MultiSciences, Hangzhou, China).

Techniques: Derivative Assay, Transfection, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot, Negative Control

Journal: Frontiers in Pharmacology

Article Title: MiR-19a-3p Suppresses M1 Macrophage Polarization by Inhibiting STAT1/IRF1 Pathway

doi: 10.3389/fphar.2021.614044

Figure Lengend Snippet: Overexpressed STAT1 reverses the inhibitory effect of miR-19a-3p in M1 phenotype polarization in RAW264.7 cells. RAW264.7 cells were co-transfected with miR-19a-3p mimics, STAT1 plasmid or NC, vector for 24 h, followed with LPS plus IFN-γ treatment for 24 h. (A,B) STAT1 and IRF1 mRNA levels were detected by qRT-PCR (data were pooled from three independent experiments). (C) p-STAT1, STAT1 and IRF1 protein levels were assessed by western blot (a representative blot, from three independent experiments). (D) TNF-α, CXCL9, and iNOS were detected by qRT-PCR (data were pooled from three independent experiments). NC: miR-19a-3p mimics negative control. Statistical significance was calculated using ANOVA followed by Bonferroni test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: TNF-α, Chemokine (C-X-C motif) ligand 9 (CXCL9), CXCL10 in the supernatants were detected with mouse TNF-α (70-EK282/3-96), CXCL9 (70-EK2143/2-24), CXCL10 (70-EK268/2-96) ELISA Kit (MultiSciences, Hangzhou, China).

Techniques: Transfection, Plasmid Preparation, Quantitative RT-PCR, Western Blot, Negative Control

Journal: Frontiers in Pharmacology

Article Title: MiR-19a-3p Suppresses M1 Macrophage Polarization by Inhibiting STAT1/IRF1 Pathway

doi: 10.3389/fphar.2021.614044

Figure Lengend Snippet: M1 macrophage increased obviously in the LPS challenged sepsis model. Male 5C7BL/6 J mice were challenged with 20 mg/kg LPS or saline by intraperitoneal injection for 24 h. (A) H&E staining of lung tissues treated with LPS and control mice (a representative experiment, from three independent experiments) (Scale bar, 50 μm, 200×). (B,C) miR-19a-3p, TNF-α, CXCL9, and CXCL10 in the peritoneal macrophages of mice treated with LPS and control mice were determined using qRT-PCR (data were pooled from three independent experiments with six mice per group). (D) TNF-α, CXCL9, and CXCL10 in serum of mice treated with LPS and control mice were measured by ELISA (data were pooled from three independent experiments with six mice per group). (E) STAT1 and IRF1 were detected in the peritoneal macrophages of mice treated with LPS and control mice (data were pooled from three independent experiments with six mice per group). (F) IRF1 and STAT1 were measured by western blot in the peritoneal macrophages of mice treated with LPS and control mice (a representative blot, from three independent experiment with three mice per group). Statistical significance was calculated using unpaired Student’s t-test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: TNF-α, Chemokine (C-X-C motif) ligand 9 (CXCL9), CXCL10 in the supernatants were detected with mouse TNF-α (70-EK282/3-96), CXCL9 (70-EK2143/2-24), CXCL10 (70-EK268/2-96) ELISA Kit (MultiSciences, Hangzhou, China).

Techniques: Injection, Staining, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Frontiers in Pharmacology

Article Title: MiR-19a-3p Suppresses M1 Macrophage Polarization by Inhibiting STAT1/IRF1 Pathway

doi: 10.3389/fphar.2021.614044

Figure Lengend Snippet: Overexpressed miR-19a-3p reduced the inflammation of lung tissues and inhibited M1 polarization in LPS-treated mice. Male 5C7BL/6 J mice were treated with agomiR-19a-3p or agomiR NC for 1 h, followed by the 20 mg/kg LPS challenge by intraperitoneal injection. (A) H&E staining of lung tissues of mice treated with agomiR-19a-3p or agomiR NC followed by challenging LPS (a representative experiment, from three independent experiments). (Scale bar, 50 μm, 200×). (B,C) miR-19a-3p, TNF-α, CXCL9, and CXCL10 in the peritoneal macrophages of mice using qRT-PCR (data were pooled from three independent experiments with ten mice per group). (D) TNF-α, CXCL9, and CXCL10 in serum of mice were measured by ELISA (data were pooled from three independent experiments with ten mice per group). (E) STAT1 and IRF1 were detected in the peritoneal macrophages of mice (data were pooled from three independent experiments with 10 mice per group). (F) IRF1 and STAT1 were measured by western blot in the peritoneal macrophages of mice (a representative blot, from three independent experiment with six mice per group). Statistical significance was calculated using unpaired Student’s t-test. ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: TNF-α, Chemokine (C-X-C motif) ligand 9 (CXCL9), CXCL10 in the supernatants were detected with mouse TNF-α (70-EK282/3-96), CXCL9 (70-EK2143/2-24), CXCL10 (70-EK268/2-96) ELISA Kit (MultiSciences, Hangzhou, China).

Techniques: Injection, Staining, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot