Journal: Cell Death & Disease

Article Title: ANGPTL2 inhibits macrophage pyroptosis and alleviates rheumatoid arthritis progression by regulating mitophagy via IGFBP5

doi: 10.1038/s41419-026-08537-z

Figure Lengend Snippet: A Representative macroscopic images showing paw swelling in WT and Angptl2 –/– CIA mice. B Assessment of arthritis progression, including body weight changes, paw thickness, and clinical arthritis scores over time. C Representative 3D μCT reconstruction images of ankle joints. Quantification of talus bone volume (mm³) ( D ) and the ratio of bone volume to tissue volume (BV/TV, %) ( E ) in ankle joints. F Representative H&E-stained sections of ankle joints showing inflammation and structural damage. G Representative TRAP-stained sections of ankle joints showing osteoclast distribution and activity. H Representative ALP-stained sections of ankle joints indicating osteoblast activity and bone formation in periarticular regions. I Dual immunofluorescence (IF) staining of ankle joint sections using anti-F4/80 and antibodies against NLRP3, Caspase-1, GSDMD, or IL-1β in CIA mice. The average IF signals for total NLRP3, Caspase-1, GSDMD, and IL-1β expression, and co-localization with F4/80 + macrophages were determined. T: tibia; TAL: talus; ST: synovial tissue. J Western blot analysis showing protein levels of ANGPTL2, NLRP3, Caspase-1, GSDMD, and IL-1β in the joints of CIA mice. * p < 0.05, ** p < 0.01, *** p <0.001, **** p <0.0001 in the indicated groups.

Article Snippet: The membranes were treated with 5% non-fat milk in TBST for an hour at room temperature and then incubated overnight at 4 °C with the following primary antibodies: ANGPTL2 (Proteintech, Wuhan.

Techniques: Staining, Activity Assay, Immunofluorescence, Expressing, Western Blot

Journal: Cell Death & Disease

Article Title: ANGPTL2 inhibits macrophage pyroptosis and alleviates rheumatoid arthritis progression by regulating mitophagy via IGFBP5

doi: 10.1038/s41419-026-08537-z

Figure Lengend Snippet: A qPCR analysis of Angptl2 , Nlrp3 , Caspase-1 , Gsdmd , and Il-1β expression in RAW264.7 cells stimulated with LPS alone or LPS + ATP. B ELISA quantification of IL-1β secretion in culture supernatants from RAW264.7 cells stimulated with LPS alone or LPS + ATP. C Time-course qPCR analysis of Angptl2 , Nlrp3 , Caspase-1 , Gsdmd , and Il-1β mRNA expression in RAW264.7 cells primed with LPS for 0, 3, 6, 12, and 24 h, followed by 1-h ATP stimulation prior to RNA extraction. D Corresponding time-course Western blot analysis of ANGPTL2, NLRP3, GSDMD, Caspase-1, and IL-1β expression in RAW264.7 cells under identical conditions. E qPCR analysis of Angptl2 , Nlrp3 , Caspase-1 , Gsdmd , and Il-1β expression in BMDMs stimulated with LPS alone or LPS + ATP. F ELISA quantification of IL-1β secretion in culture supernatants from BMDMs stimulated with LPS alone or LPS + ATP. G Time-course qPCR analysis of Angptl2 , Nlrp3 , Caspase-1 , Gsdmd , and Il-1β mRNA expression in BMDMs primed with LPS for 0, 3, 6, 12, and 24 h, followed by 1-h ATP stimulation prior to RNA extraction. H Corresponding time-course Western blot analysis of ANGPTL2, NLRP3, GSDMD, Caspase-1, and IL-1β expression in BMDMs under identical conditions. * p < 0.05, ** p < 0.01, *** p <0.001, **** p <0.0001 in the indicated groups.

Article Snippet: The membranes were treated with 5% non-fat milk in TBST for an hour at room temperature and then incubated overnight at 4 °C with the following primary antibodies: ANGPTL2 (Proteintech, Wuhan.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, RNA Extraction, Western Blot

Journal: Cell Death & Disease

Article Title: ANGPTL2 inhibits macrophage pyroptosis and alleviates rheumatoid arthritis progression by regulating mitophagy via IGFBP5

doi: 10.1038/s41419-026-08537-z

Figure Lengend Snippet: qPCR analysis of Angptl2 , Nlrp3 , Caspase-1 , Gsdmd , and Il-1β mRNA expression in RAW264.7 cells ( A ) and BMDMs ( B ) following si Angptl2 or siNC transfection, with or without LPS + ATP stimulation. Western blot analysis of ANGPTL2, NLRP3, GSDMD, Caspase-1, and IL-1β protein expression in RAW264.7 cells ( C ) and BMDMs ( D ) under identical conditions. ELISA measurement of IL-1β levels in culture supernatants of RAW264.7 cells ( E ) and BMDMs ( F ). Cell death assessment using LDH release assay in RAW264.7 cells ( G ) and BMDMs ( H ) under identical conditions. Representative TEM images showing morphological features of pyroptotic cell death in RAW264.7 cells ( I ) and BMDMs ( J ). Representative flow cytometry plots showing Annexin V and PI staining of RAW264.7 cells ( K ) and BMDMs ( L ) under identical conditions. Quantification of Annexin V + PI + populations in RAW264.7 cells ( M ) and BMDMs ( N ). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 in the indicated groups.

Article Snippet: The membranes were treated with 5% non-fat milk in TBST for an hour at room temperature and then incubated overnight at 4 °C with the following primary antibodies: ANGPTL2 (Proteintech, Wuhan.

Techniques: Expressing, Transfection, Western Blot, Enzyme-linked Immunosorbent Assay, Lactate Dehydrogenase Assay, Flow Cytometry, Staining

Journal: Cell Death & Disease

Article Title: ANGPTL2 inhibits macrophage pyroptosis and alleviates rheumatoid arthritis progression by regulating mitophagy via IGFBP5

doi: 10.1038/s41419-026-08537-z

Figure Lengend Snippet: A Volcano plot of DEGs comparing Angptl2 –/– and NC BMDMs following LPS + ATP stimulation. Genes with significant upregulation (yellow) or downregulation (blue) are highlighted based on thresholds of adjusted p < 0.05 and |log2(fold change)| ≥2. Non-significant genes are shown in gray. B Heatmap showing hierarchical clustering of DEGs between Angptl2 –/– and NC BMDMs following LPS + ATP stimulation. Each column represents an individual sample, and each row represents a gene. Color gradient indicates Z-score normalized expression levels (red: upregulated, green: downregulated). C GSEA plot showing enrichment of the gene set “negative regulation of autophagy of mitochondrion” between groups. D GO enrichment dot plot for DEGs. Enriched terms are classified by Biological Process (BP), Cellular Component (CC), and Molecular Function (MF). Dot size reflects gene count, color indicates adjusted p-value, and X-axis denotes gene ratio. E KEGG pathway enrichment map for DEGs in KO-LPS vs. NC-LPS. Dot size reflects gene count, color indicates adjusted p-value, and X-axis denotes gene ratio.

Article Snippet: The membranes were treated with 5% non-fat milk in TBST for an hour at room temperature and then incubated overnight at 4 °C with the following primary antibodies: ANGPTL2 (Proteintech, Wuhan.

Techniques: Expressing

Journal: Cell Death & Disease

Article Title: ANGPTL2 inhibits macrophage pyroptosis and alleviates rheumatoid arthritis progression by regulating mitophagy via IGFBP5

doi: 10.1038/s41419-026-08537-z

Figure Lengend Snippet: A Western blot analysis of mitophagy-related proteins (PINK1, PARKIN, LC3B, and p62) in joint tissues from WT and Angptl2 –/– mice under basal and CIA-induced conditions. B Western blot analysis of mitophagy markers in RAW264.7 cells and BMDMs transfected with si Angptl2 or siNC, with or without LPS + ATP stimulation. Flow cytometry analysis of mitochondrial membrane potential using JC-1 staining in RAW264.7 cells ( C ) and BMDMs ( D ). The ratio of red (aggregates) to green (monomers) fluorescence (R/G) was calculated to assess mitochondrial integrity. Intracellular ROS levels measured by flow cytometry in RAW264.7 cells ( E ) and BMDMs ( F ) under the indicated conditions. G Flow cytometry analysis of mito-Keima-loaded RAW264.7 cells to assess mitophagic flux under different conditions. H Representative confocal microscopy images showing colocalization of LC3B (red) and mitochondria (MitoTracker Green) in BMDMs. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 in the indicated groups.

Article Snippet: The membranes were treated with 5% non-fat milk in TBST for an hour at room temperature and then incubated overnight at 4 °C with the following primary antibodies: ANGPTL2 (Proteintech, Wuhan.

Techniques: Western Blot, Transfection, Flow Cytometry, Membrane, Staining, Fluorescence, Confocal Microscopy

Journal: Cell Death & Disease

Article Title: ANGPTL2 inhibits macrophage pyroptosis and alleviates rheumatoid arthritis progression by regulating mitophagy via IGFBP5

doi: 10.1038/s41419-026-08537-z

Figure Lengend Snippet: A Western blot analysis of ANGPTL2 expression in BMDMs following treatment with recombinant mouse ANGPTL2 protein (rMmANGPTL2). B qPCR analysis of Nlrp3 , Caspase-1 , Gsdmd , and Il-1β mRNA expression in BMDMs stimulated with LPS + ATP and treated with increasing concentrations of rMmANGPTL2 (0, 250, 500, 750, 1000 ng/mL). C Flow cytometry analysis of mitochondrial membrane potential using JC-1 staining in BMDMs under NC, LPS + ATP, or LPS + ATP + rMmANGPTL2 treatment conditions. D Flow cytometry analysis of intracellular ROS levels in BMDMs under NC, LPS + ATP, or LPS + ATP + rMmANGPTL2 treatment conditions. qPCR ( E ) and Western blot ( F ) analysis of mitophagy-related markers in BMDMs following LPS + ATP stimulation with or without rMmANGPTL2 and the autophagy inhibitor 3-MA. qPCR ( G ) and Western blot ( H ) analysis of pyroptosis-associated markers in BMDMs under the same treatment conditions. I Flow cytometry analysis of mito-Keima-loaded RAW264.7 cells to assess mitophagic flux under the same treatment conditions. J Representative confocal microscopy images showing LC3B and mitochondria colocalization in BMDMs under the same treatment conditions. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 in the indicated groups.

Article Snippet: The membranes were treated with 5% non-fat milk in TBST for an hour at room temperature and then incubated overnight at 4 °C with the following primary antibodies: ANGPTL2 (Proteintech, Wuhan.

Techniques: Western Blot, Expressing, Recombinant, Flow Cytometry, Membrane, Staining, Confocal Microscopy

Journal: Cell Death & Disease

Article Title: ANGPTL2 inhibits macrophage pyroptosis and alleviates rheumatoid arthritis progression by regulating mitophagy via IGFBP5

doi: 10.1038/s41419-026-08537-z

Figure Lengend Snippet: A Heatmap showing the expression correlation between ANGPTL2 and insulin-like growth factor (IGF) family members, including IGF1, IGF2, and IGFBP proteins. qPCR ( B ) and Western blot ( C ) analysis of IGFBP5 expression in BMDMs following Igfbp5 knockdown and LPS + ATP stimulation. D Western blot analysis of mitophagy-related proteins (PINK1, PARKIN, LC3B, p62) and NLRP3 in BMDMs after Igfbp5 knockdown with LPS + ATP treatment. qPCR ( E ) and Western blot ( F ) analysis of IGFBP5 expression in BMDMs following Angptl2 knockdown and LPS + ATP stimulation. qPCR ( G ) and Western blot ( H ) analysis of IGFBP5 expression in BMDMs treated with rMmANGPTL2 under LPS + ATP stimulation. I – J Co-immunoprecipitation (Co-IP) assay to evaluate protein-protein interaction between ANGPTL2 and IGFBP5 in RAW264.7 cells. K 3D model of structured ANGPTL2 interfaces with IGFBP5 predicted by HDOCK. L Western blot analysis of mitophagy-related proteins in BMDMs treated with LPS + ATP and rMmANGPTL2, with or without Igfbp5 knockdown. * p < 0.05, ** p < 0.01, *** p <0.001, **** p <0.0001 in the indicated groups.

Article Snippet: The membranes were treated with 5% non-fat milk in TBST for an hour at room temperature and then incubated overnight at 4 °C with the following primary antibodies: ANGPTL2 (Proteintech, Wuhan.

Techniques: Expressing, Western Blot, Knockdown, Co-Immunoprecipitation Assay

Journal: Cell Death & Disease

Article Title: ANGPTL2 inhibits macrophage pyroptosis and alleviates rheumatoid arthritis progression by regulating mitophagy via IGFBP5

doi: 10.1038/s41419-026-08537-z

Figure Lengend Snippet: A Representative images of mouse paws from control, CIA, and AAV- Angptl2 -treated groups showing macroscopic joint swelling. B Assessment of arthritis progression, including body weight changes, paw thickness, and clinical arthritis scores over time. C Representative 3D μCT reconstruction images of ankle joints. D Representative H&E-stained sections of ankle joints illustrating synovial inflammation and tissue architecture. E Representative TRAP-stained sections showing osteoclast distribution and activity. F Representative ALP-stained sections indicating osteoblast activity and bone formation. G Representative dual immunofluorescence images of ankle joint sections stained for F4/80 and either NLRP3, GSDMD, Caspase-1, or IL-1β. The average IF signals for total NLRP3, GSDMD, Caspase-1, and IL-1β expression, and co-localization with F4/80 + macrophages were determined. T: tibia; TAL: talus; ST: Synovial Tissue. H Western blot analysis of ANGPTL2 protein levels in joint tissues from each group. I Western blot analysis of pyroptosis and inflammasome-related proteins (NLRP3, GSDMD, Caspase-1, IL-1β) in joint tissues. J Western blot analysis of mitophagy-related proteins (PINK1, PARKIN, LC3B, p62) in joint tissues. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 in the indicated groups.

Article Snippet: The membranes were treated with 5% non-fat milk in TBST for an hour at room temperature and then incubated overnight at 4 °C with the following primary antibodies: ANGPTL2 (Proteintech, Wuhan.

Techniques: Control, Staining, Activity Assay, Immunofluorescence, Expressing, Western Blot

Journal: PLoS ONE

Article Title: Angiopoietin Like Protein 2 (ANGPTL2) Promotes Adipose Tissue Macrophage and T lymphocyte Accumulation and Leads to Insulin Resistance

doi: 10.1371/journal.pone.0131176

Figure Lengend Snippet: A, Western blot analysis of FLAG-tagged ANGPTL2 in the liver (upper panel: lean mice, lower panel: db/db mice). B, Body weight (n = 7–8 mice / group). C, Plasma ANGPTL2 levels 2 week after adenovirus injection (n = 6–8). D, Fasting glucose levels at 2 weeks (n = 7–8 mice / group). E, Glucose tolerance test at 2 weeks after treatment (Left: Lean mice, Right: db/db mice, n = 7–8 mice / group, in Experiment 1). F, Insulin tolerance test 2 weeks after treatment in db/db mice (n = 7–8 mice / group, in Experiment 2). G, Quantitative RT-PCR of mRNAs encoding gluconeogenesis related genes in the liver (Left: Lean mice, Right: db/db mice, n = 8). Data are mean ± SEM, **: P<0.01, *: P<0.05 compared with LacZ group.

Article Snippet: Briefly, C-terminal FLAG ANGPTL2 expression vector was obtained from Origene (Myc-DDK-tagged-Human angiopoietin-like 2, RC201917, Rockville, MD).

Techniques: Western Blot, Injection, Quantitative RT-PCR

Journal: PLoS ONE

Article Title: Angiopoietin Like Protein 2 (ANGPTL2) Promotes Adipose Tissue Macrophage and T lymphocyte Accumulation and Leads to Insulin Resistance

doi: 10.1371/journal.pone.0131176

Figure Lengend Snippet: A, Western blotting analysis of FLAG fusion ANGPTL2 protein with anti-FLAG and anti-human Angptl2 antibody (Left: with 2-mercaptoethanol (2-ME), Right: without 2-ME). B, TNF-α concentration in rANGPTL2-treated THP-1 conditioned media (72 hr treatment, n = 3). C, Quantitative RT-PCR of mRNAs encoding Angptl2 and pro-inflammatory related genes in THP-1 cells (24 hr treatment, n = 3). D, Quantitative RT-PCR of mRNAs encoding M2 macrophage markers in THP-1 cells (24 hr treatment, n = 3). Data are mean ± SEM, **: P<0.01, *: P<0.05 compared with BSA group.

Article Snippet: Briefly, C-terminal FLAG ANGPTL2 expression vector was obtained from Origene (Myc-DDK-tagged-Human angiopoietin-like 2, RC201917, Rockville, MD).

Techniques: Western Blot, Concentration Assay, Quantitative RT-PCR

Journal: PLoS ONE

Article Title: Angiopoietin Like Protein 2 (ANGPTL2) Promotes Adipose Tissue Macrophage and T lymphocyte Accumulation and Leads to Insulin Resistance

doi: 10.1371/journal.pone.0131176

Figure Lengend Snippet: A, Representative images of the liver (Left: LacZ, Right: Angptl2, db/db mice). B, Liver triglyceride levels (Left: Lean mice, Right: db/db mice, n = 7–8 animals per group). C, Representative images of oil red O staining (Left: LacZ, Right: Angptl2, db/db mice). D, Oil red O staining area (n = 7–8, db/db mice). E, Quantitative RT-PCR of mRNAs encoding genes related to fatty acid metabolism in the liver of lean mice (n = 8) F, Quantitative RT-PCR of mRNAs encoding genes related to fatty acid metabolism in the liver of db/db mice (n = 7–8). Data are mean ± SEM, **: P<0.01, *: P<0.05 compared with LacZ group.

Article Snippet: Briefly, C-terminal FLAG ANGPTL2 expression vector was obtained from Origene (Myc-DDK-tagged-Human angiopoietin-like 2, RC201917, Rockville, MD).

Techniques: Staining, Quantitative RT-PCR

Journal: PloS one

Article Title: Krüppel like factor 4 promoter undergoes active demethylation during monocyte/macrophage differentiation.

doi: 10.1371/journal.pone.0093362

Figure Lengend Snippet: Figure 1. PU.1 binds specifically to KLF4 promoter during differentiation of myeloid progenitors to macrophages. A) PU/ER(T) cells were grown in presence of 95% Ethanol or 100 nM Tamoxifen for time periods as indicated. Cytosolic and nuclear protein fractions were prepared and translocation of PU.1 from cytosol to nucleus was detected by immunoblotting with anti PU.1 rabbit polyclonal antibodies. Purity of cytoplasmic and nuclear fractions was analyzed by immunoblotting with a-Tubulin and HDAC2 proteins. B) Nuclear extracts were prepared from differentiating bone marrow derived macrophages on day 1 and 7 and in vitro binding to biotin labeled 76 bp oligonucleotide probe representing 2118/2113 PU.1 binding element in the KLF4 promoter was determined by EMSA as described in methods. C) PU/ER(T) cells were treated with Ethanol or 100 nM Tamoxifen for 1, 4 or 24 h and nuclear extracts were prepared. An equal amount of nuclear protein extract from Ethanol and Tamoxifen treatment was analyzed for DNA binding activity using biotin labeled 76 bp oligonucleotide probe representing 2118/2113 PU.1 binding element in the KLF4 promoter. Specificity of PU.1-KLF4 DNA complex in experiment B & C was determined by the ability of the PU.1-KLF4 DNA complex to form supershift with anti PU.1 antibody. D) Specificity of PU.1-KLF4 DNA complex was determined by including 5, 15, 25 and 100 picomole of excess unlabelled KLF4 promoter oligo in the DNA binding reaction. E) In vivo binding of PU.1 to KLF4 promoter was determined by Chromatin-immuno precipitation in differentiating BMDM on day 1, 2 and 7 and in F) PU/ER(T) cells treated with Tamoxifen for 1 and 24 h. Error bars represent standard deviation and * is used where ever p value is #0.05 and ** is used when p#0.005. doi:10.1371/journal.pone.0093362.g001

Article Snippet: Myc tagged AICDA and FLAG tagged human KLF4 expression vectors were purchased from Origene (RC202949) and SABiosciences (DAM603), respectively.

Techniques: Translocation Assay, Western Blot, Derivative Assay, In Vitro, Binding Assay, Labeling, Activity Assay, In Vivo, Chromatin Immunoprecipitation, Standard Deviation

Journal: PloS one

Article Title: Krüppel like factor 4 promoter undergoes active demethylation during monocyte/macrophage differentiation.

doi: 10.1371/journal.pone.0093362

Figure Lengend Snippet: Figure 4. PU.1 transcriptionally regulates KLF4 expression that is sensitive to promoter methylation. A) PU/ER(T) cells were electroporated with 1.6 kb pGL3-KLF4 vector, after 16 h of electroporation treated with ethanol or 100 nM Tamoxifen for 24 h and analyzed for Luciferase reporter activity B) HEK293 cells were transfected with empty pGL3, 1.6 kb KLF4-pGL3, pCMV-PU.1 or pCMV control vectors separately or in combination as indicated in figure, and after 36 h cell extracts were prepared in Promega cell lysis buffer and analyzed for luciferase activity. C) HEK293 cells were transfected with empty pGL3, 1.6 kb mutant pGL3-KLF4, pCMV-PU.1 or pCMV control vectors separately or in combination as indicated in figure, and after 36 h cell extracts were prepared in Promega cell lysis buffer and analyzed for luciferase activity. D) PU/ER(T) cells were treated with Tamoxifen or ethanol for indicated time periods and expression of KLF4 was determined by immunoblotting. E) PU/ER(T) cells were electroporated with either control pCMV or KLF4 over expression vector and grown in IMDM for 72 h and analyzed for surface expression of F4/80. F) The biotin labeled KLF4 promoter oligo was methylated using M.SssI in presence of S-Adenosyl Methionine and used as a probe to determine the DNA binding activity in the Ethanol and Tamoxifen treated nuclear extracts as described in figure 1. G) Empty pGL3 vector, pGL3-KLF4 Luciferase vector were in vitro methylated using M.SssI in presence of S-Adenosyl Methionine and electroporated along with control pCMV or pCMV-PU.1 expression vector. After 36 h, cell extracts were prepared in Promega cell lysis buffer and analyzed for luciferase activity. H) Empty pcpgf-basic, pcpgf- KLF4 were in vitro methylated using M.sssI in presence of S-Adenosyl Methionine and electroporated along with control pCMV or pCMV-PU.1 expression vector in to HEK293 cells and grown in HEK293Blue detection medium for 36 h. Relative promoter activity was compared by mSEAP reporter activity as measured by the absorbance of the HEK293 Blue growth medium at 630 nm. For figures 4 B, C, G, H reporter gene activities of pGL3-KLF4/pGL3-mutant KLF4/pcpg-KLF4 were not significantly different when co-transfected with the pCMV control plasmid. Figures 4 A to 4 C and 4 G, H means were compared and * is used where ever p value is #0.05 and ** is used when p#0.005. Error bars represent standard deviation. doi:10.1371/journal.pone.0093362.g004

Article Snippet: Myc tagged AICDA and FLAG tagged human KLF4 expression vectors were purchased from Origene (RC202949) and SABiosciences (DAM603), respectively.

Techniques: Expressing, Methylation, Plasmid Preparation, Electroporation, Luciferase, Activity Assay, Transfection, Control, Lysis, Mutagenesis, Western Blot, Over Expression, Labeling, Binding Assay, In Vitro, Standard Deviation

Journal: PloS one

Article Title: Krüppel like factor 4 promoter undergoes active demethylation during monocyte/macrophage differentiation.

doi: 10.1371/journal.pone.0093362

Figure Lengend Snippet: Figure 6. AICDA is essential for KLF4 promoter demethylation. A) The relative methylation index of the proximal M1 CpG region in KLF4 promoter was compared on 1st and 7th day of differentiating BMDM from wild type and GADD45a knock out macrophages as described in Figure 3 C&D. B) Knockdown of AICDA protein in AICDA-shRNA or control-shRNA electroporated PU/ER(T) cells. C) Methylation index of the proximal KLF4 promoter M1 CpG region was compared in AICDA shRNA or control shRNA electroporated PU/ER(T) cells as described in Figure 3C&D. D) Expression of F4/80 was analyzed in control shRNA or AICDA-shRNA electroporated PU/ER(T) cells treated with or without tamoxifen for 72 h. E) Morphological changes in ethanol/tamoxifen treated PU/ER(T) cells were analyzed in AICDA depleted cells in comparison with control cells. Representative histograms of flow cytometry, western blots and cell staining were presented. Figure 6A and C, * is used where ever p value is #0.05 and ** is used when p#0.005. Error bars represent standard deviation. doi:10.1371/journal.pone.0093362.g006

Article Snippet: Myc tagged AICDA and FLAG tagged human KLF4 expression vectors were purchased from Origene (RC202949) and SABiosciences (DAM603), respectively.

Techniques: Methylation, Knock-Out, Knockdown, shRNA, Control, Expressing, Comparison, Flow Cytometry, Western Blot, Staining, Standard Deviation