Journal: Scientific reports

Article Title: Messenger RNA delivery of a cartilage-anabolic transcription factor as a disease-modifying strategy for osteoarthritis treatment.

doi: 10.1038/srep18743

Figure Lengend Snippet: Figure 2. Histological analysis of mouse OA knee joints exposed to intra-articular injections of PEG- PAsp(DET) polyplex nanomicelles carrying RUNX1-FL or EGFP mRNA. (a) Images of representative sections stained with safranin-O after one month of intra-articular injection of 1 μ g RUNX1-FL or EGFP mRNA-loaded PEG-PAsp(DET) polyplex nanomicelles. Scale bars, 100 μ m. (b) Immunostaining on the serial sections of (a) using an anti-RUNX1 antibody. Brown spots indicate positive staining. Counter staining was performed with methyl green. Scale bars, 100 μ m. (c) Scoring of OA status with the OARSI scoring system and the osteophyte scoring system. The score of normal cartilage is 0. Data are expressed as the mean ± SD (N = 10).

Article Snippet: After being blocked in 6% milk/Tris buffered saline with 0.1% Tween (TBST), the membranes were incubated with a horseradish peroxidase (HRP)-conjugated anti-Flag M2 mouse monoclonal antibody (1:1000, A8592; Sigma-Aldrich) or an anti-Runx1 rabbit antibody (1:100, sc28679; Santa Cruz Biotechnology) overnight.

Techniques: Staining, Injection, Immunostaining

Journal: Scientific reports

Article Title: Messenger RNA delivery of a cartilage-anabolic transcription factor as a disease-modifying strategy for osteoarthritis treatment.

doi: 10.1038/srep18743

Figure Lengend Snippet: Figure 3. Histological analysis of mouse OA knee joints exposed to intra-articular injection of PEG- PAsp(TET) polyplex nanomicelles carrying RUNX1-FL or EGFP mRNA. (a) Images of representative sections stained with safranin-O after one month of intra-articular injection of 1 μ g RUNX1-FL or EGFP mRNA-loaded PEG-PAsp(TET) polyplex nanomicelles. Scale bars, 100 μ m. (b) Immunostaining of tibias using an anti-RUNX1 antibody on representative sections in (a). Brown spots indicate positive staining. Counter staining was performed with methyl green. Images of medial and lateral sites are shown. Medial sites were supposed to be more afflicted by the OA induction than lateral sites. Inset boxes indicate the magnified region shown below each image. Scale bars, 100 μ m. (c) Immunostaining of tibias using anti-FLAG and anti-GFP antibodies on representative sections in (a). Brown spots indicate positive staining. Counter staining was performed with methyl green. Scale bars, 100 μ m. (d) Scoring of OA status with both the OARSI scoring system and the osteophyte scoring system. The score of normal cartilage is 0. Data are expressed as the mean ± SD (N = 10). *P < 0.05 versus the EGFP mRNA-injected group.

Article Snippet: After being blocked in 6% milk/Tris buffered saline with 0.1% Tween (TBST), the membranes were incubated with a horseradish peroxidase (HRP)-conjugated anti-Flag M2 mouse monoclonal antibody (1:1000, A8592; Sigma-Aldrich) or an anti-Runx1 rabbit antibody (1:100, sc28679; Santa Cruz Biotechnology) overnight.

Techniques: Injection, Staining, Immunostaining

Journal: Scientific reports

Article Title: Messenger RNA delivery of a cartilage-anabolic transcription factor as a disease-modifying strategy for osteoarthritis treatment.

doi: 10.1038/srep18743

Figure Lengend Snippet: Figure 4. Immunohistochemical analyses of mouse OA knee joints exposed to intra-articular injection of PEG-PAsp(TET) polyplex nanomicelles carrying RUNX1-FL or EGFP mRNA. (a) Immunostaining using anti-Sox9, anti-type II collagen, and anti-PCNA antibodies on sections after one month of intra-articular injection of 1 μ g RUNX1-FL or EGFP mRNA-loaded PEG-PAsp(TET) polyplex nanomicelles. Brown spots indicate positive staining. Counter staining was performed with methyl green. Inset boxes indicate the magnified region shown on the right of each image. Scale bars, 100 μ m. Quantitative data of stained cells are shown below each image. Data are expressed as the mean ± SD (N = 3). *P < 0.05 versus the EGFP mRNA- injected group. (b) Immunostaining using anti-type X collagen, anti-MMP13, and anti- IL-1ß antibodies on sections after one-month intra-articular injection of 1 μ g RUNX1-FL or EGFP mRNA-loaded PEG-PAsp(TET)

Article Snippet: After being blocked in 6% milk/Tris buffered saline with 0.1% Tween (TBST), the membranes were incubated with a horseradish peroxidase (HRP)-conjugated anti-Flag M2 mouse monoclonal antibody (1:1000, A8592; Sigma-Aldrich) or an anti-Runx1 rabbit antibody (1:100, sc28679; Santa Cruz Biotechnology) overnight.

Techniques: Immunohistochemical staining, Injection, Immunostaining, Staining

Journal: Cell

Article Title: Cooperative binding of transcription factors orchestrates reprogramming

doi: 10.1016/j.cell.2016.12.016

Figure Lengend Snippet: Experimental Model and Subject Details

Article Snippet: Rabbit polyclonal anti-Runx1 , Novus Biologicals , Cat#NBP1-61277.

Techniques: Recombinant, Sample Prep, DNA Library Preparation, Isolation, Sequencing, Luciferase, Software

Journal:

Article Title: The E2A-HLF Oncoprotein Activates Groucho -Related Genes and Suppresses Runx1

doi: 10.1128/MCB.21.17.5935-5945.2001

Figure Lengend Snippet: Downregulation of expression of the RUNX1 gene induced by E2A-HLF. Northern blot analysis was performed on poly(A)+ RNA (1 μg per lane) prepared from FL5.12 cells expressing the indicated cDNAs under the control of a zinc-regulated metallothionein promoter. Cells were cultured in the presence or absence of zinc (100 μM) and IL-3 as described in the legend to Fig. ​Fig.1.1. The blot was hybridized with the RUNX1 or Gapdh cDNA probe as indicated. Mobilities of the 18S and 28S rRNAs are shown.

Article Snippet: A rabbit polyclonal antibody against Runx1 (AML1/RHD; Ab-2) was obtained from Calbiochem, and a goat polyclonal antibody against actin (C-11) was obtained from Santa Cruz Biotechnology.

Techniques: Expressing, Northern Blot, Cell Culture

Journal:

Article Title: The E2A-HLF Oncoprotein Activates Groucho -Related Genes and Suppresses Runx1

doi: 10.1128/MCB.21.17.5935-5945.2001

Figure Lengend Snippet: Time course of RUNX1 and Grg6 mRNA expression in E2A-HLF-transfected FL5.12 cells. (A) Total RNA was isolated at different time points after addition of zinc (100 μM) and hybridized with RUNX1, Grg6, and Gapdh antisense mRNA probes. GAPDH was used as an internal control. The protected RNA fragments (solid arrowheads) were separated by an 8% sequencing gel. Open arrowheads, RNA probes. (B) Immunoblot analysis of Runx1 in E2A-HLF-transfected FL5.12 cells. Extracts were prepared from cells stably transfected with either the pMT-CB6+ vector or the metallothionein promoter-regulated E2A-HLF construct after the cells were cultured in IL-3-containing medium for the indicated times in hours after the addition of 100 μM ZnSO4. The blot was immunoblotted with Runx1 rabbit antisera, HLF(C) rabbit antisera, and actin goat antisera (C11), respectively.

Article Snippet: A rabbit polyclonal antibody against Runx1 (AML1/RHD; Ab-2) was obtained from Calbiochem, and a goat polyclonal antibody against actin (C-11) was obtained from Santa Cruz Biotechnology.

Techniques: Expressing, Transfection, Isolation, Sequencing, Western Blot, Stable Transfection, Plasmid Preparation, Construct, Cell Culture

Journal:

Article Title: The E2A-HLF Oncoprotein Activates Groucho -Related Genes and Suppresses Runx1

doi: 10.1128/MCB.21.17.5935-5945.2001

Figure Lengend Snippet: Expression of RUNX1 and Grg6 in FL5.12 cells stably transfected with plasmids expressing E2A-HLF, TLE1, TLE2, and RDA3. Total RNA was isolated from cells with or without a 24-h incubation with zinc (100 μM) and was hybridized with RUNX1, Grg6, and Gapdh antisense RNA probes. Solid arrowheads, protected RNA fragments; open arrowheads, RNA probes.

Article Snippet: A rabbit polyclonal antibody against Runx1 (AML1/RHD; Ab-2) was obtained from Calbiochem, and a goat polyclonal antibody against actin (C-11) was obtained from Santa Cruz Biotechnology.

Techniques: Expressing, Stable Transfection, Transfection, Isolation, Incubation

Journal: Cell reports

Article Title: Secreted phosphoprotein 1 slows neurodegeneration and rescues visual function in mouse models of aging and glaucoma

doi: 10.1016/j.celrep.2022.111880

Figure Lengend Snippet:

Article Snippet: Rabbit anti-Runx1 , LSBio , Cat# LS-B13948.

Techniques: Plasmid Preparation, Virus, Recombinant, Negative Control, Phospho-proteomics, Produced, Software

Journal: Frontiers in Molecular Biosciences

Article Title: RUNX1 Upregulates CENPE to Promote Leukemic Cell Proliferation

doi: 10.3389/fmolb.2021.692880

Figure Lengend Snippet: RUNX1 in leukemia cells regulates cell cycle-associated biological process. (A) Workflow for choosing differential expression genes and the RUNX1 occupied genes. Heat map showed the RNA-seq differentially expressed genes (DEGs) (FDR p < 0.1 and >2 -fold change) between THP-1 cells and CD34 + cells. RUNX1 occupied genes was defined as genes contained RUNX1 binding around ±1 kb relative to TSS. (B) Workflow for choosing THP-1 specific RUNX1 potential target genes. For THP-1 specific RUNX1 potential target genes, we overlapped DEGs with THP-1 specific RUNX1 occupied genes. (C) Gene Ontology (GO) analysis of 1784 differentially expressed genes that contain RUNX1 binding site but without MLL-AF9 occupied in promoter regions (±1 kb relative to TSS) for THP-1 cells. Black font indicates the biological pathways that are related to cell growth; gray font indicates other biological pathways. (D) RUNX1 peaks located in the proximal vicinity of TSSs of CENPE. (E) qPCR confirmed FANCD2 and CENPE mRNA expression of THP-1 cells and CD34 + cells with or without RUNX1 shRNA. (one-way ANOVA, * p < 0.05, ** p < 0.01,*** p < 0.001; error bars, median ± SD).

Article Snippet: Thereafter, the membrane was probed using antibodies against RUNX1 (Cell Signaling, 2527S, 1:500).

Techniques: Quantitative Proteomics, RNA Sequencing, Binding Assay, Expressing, shRNA

Journal: Frontiers in Molecular Biosciences

Article Title: RUNX1 Upregulates CENPE to Promote Leukemic Cell Proliferation

doi: 10.3389/fmolb.2021.692880

Figure Lengend Snippet: Cell number phenotype caused by RUNX1 overexpression (OE) and knockdown (KD). (A) Scheme of lentivirus transduction experiments. RUNX1 OE and KD virus were transducted into THP-1 and CD34 + cells. After FC sorting, GFP + cells were transferred into fresh medium and culture in vitro for further experiment. (B) qRT-PCR confirmed the relative RUNX1 mRNA expression of THP-1 and CD34 + cells with empty vector, RUNX1 and RUNX1 shRNA in vitro . ALL groups were compared with Ctrl. All reactions were normalized against GAPDH. (C) THP-1 and CD34 + cells were transfected with the RUNX1 and RUNX1 shRNA virus, followed by efficiency check at 60 h post-transduction before growth curves were monitored daily. The cell number was normalized to Day 0 before plotting. Blue trace is the empty vector control. Red, green, and purple traces are cells transfected with individual RUNX1, RUNX1 sh1 and sh2, respectively. (one-way ANOVA, * p < 0.05, ** p < 0.01,*** p < 0.001; error bars, median ± SD).

Article Snippet: Thereafter, the membrane was probed using antibodies against RUNX1 (Cell Signaling, 2527S, 1:500).

Techniques: Over Expression, Knockdown, Transduction, Virus, In Vitro, Quantitative RT-PCR, Expressing, Plasmid Preparation, shRNA, Transfection, Control

Journal: Frontiers in Molecular Biosciences

Article Title: RUNX1 Upregulates CENPE to Promote Leukemic Cell Proliferation

doi: 10.3389/fmolb.2021.692880

Figure Lengend Snippet: RUNX1 knockdown may affect leukemia cell differentiation ability. (A) Cell morphology change and cell aggregation in cell culture at day 5. (B) Colony formation of human THP-1 cells and CD34 + cells after transduction of empty vector, RUNX1 and shRUNX1. The colony morphology could be observed at (B) and the whole colony number and subtype calculated at (C) , colony number (upper) and colony size (lower) were respectively analyzed at (D) . (one-way ANOVA, * p < 0.05, ** p < 0.01,*** p < 0.001; error bars, median ± SD).

Article Snippet: Thereafter, the membrane was probed using antibodies against RUNX1 (Cell Signaling, 2527S, 1:500).

Techniques: Knockdown, Cell Differentiation, Cell Culture, Transduction, Plasmid Preparation

Journal: Frontiers in Molecular Biosciences

Article Title: RUNX1 Upregulates CENPE to Promote Leukemic Cell Proliferation

doi: 10.3389/fmolb.2021.692880

Figure Lengend Snippet: RUNX1 promotes cell proliferation and inhibits cell apoptosis in leukemia cells. (A) FC analysis of cell proliferation in THP-1 cells. Flow plots (left) and histograms (right) show the proliferation ratio of THP-1 cells transducted with control, OE (RUNX1), and KD (RUNX1 shRNA). (B) . FC analysis of cell apoptosis in THP-1 cells. Flow plots (left) and histograms (right) show the apoptosis (blue and red box) and active cell (black box) ratio. The cell proliferation in (C) and apoptosis in (D) in CD34 + cells were similar described with (A,B) . (one-way ANOVA, * p < 0.05, ** p < 0.01,*** p < 0.001; error bars, median ± SD).

Article Snippet: Thereafter, the membrane was probed using antibodies against RUNX1 (Cell Signaling, 2527S, 1:500).

Techniques: Control, shRNA

Journal: Frontiers in Molecular Biosciences

Article Title: RUNX1 Upregulates CENPE to Promote Leukemic Cell Proliferation

doi: 10.3389/fmolb.2021.692880

Figure Lengend Snippet: RUNX1 directly regulates CENPE can rescue cell proliferation. (A) qRT-PCR confirmed the relative mRNA expression (CENPE, PIWI4, ZMPSTE24, XPO1, CLSPN and HMMR) of THP-1 cells with empty vector, RUNX1 and RUNX1 shRNA in vitro . (B) Growth curve of THP-1 cells that rescued with CENPE. Growth curve of THP-1 cells that contained different groups. Blue trace is the empty vector control. Orange and green are cells transfected with individual CENPE and shRUNX1, respectively. The purple trace is transfected with CENPE in RUNX1 knockdown of THP-1 cells. (C) FC analysis of cell proliferation in THP-1 cells treated with Control, shRUNX1, and shRUNX1+ CENPE. (one-way ANOVA, * p < 0.05, ** p < 0.01,*** p < 0.001; error bars, median ± SD). (D) The colony subtypes of THP-1 cells after transduction of CENPE (labeled as CENPE OE) and CENPE + shRUNX1 (labeled as CENPE + sh2) were observed under the microscope [control (labeled as Ctrl) and shRUNX1 (labeled as sh2) conditions are shown in , but not shown here to avoid redundancy]. shows images for all conditions in triplicate.

Article Snippet: Thereafter, the membrane was probed using antibodies against RUNX1 (Cell Signaling, 2527S, 1:500).

Techniques: Quantitative RT-PCR, Expressing, Plasmid Preparation, shRNA, In Vitro, Control, Transfection, Knockdown, Transduction, Labeling, Microscopy