Journal: ACS Applied Materials & Interfaces

Article Title: IL-4/Nanohydroxyapatite Codelivery in a Dual-Bionic Scaffold: Design and Immunomodulatory Endogenous Osteogenesis Mechanism

doi: 10.1021/acsami.5c23172

Figure Lengend Snippet: Schematic illustration of the bioactive PHS/SG@IL-4 dual-bionic scaffold: from bionic design to critical-sized calvarial regeneration. Inspired by the “soft-hard” integration structure of natural bone, we fabricated PCL/nHA composite filaments to construct gyroid bionic scaffolds through FDM 3D printing. Subsequently, SG hydrogel loaded with interleukin-4 (SG@IL-4) was infused into the scaffold pores using a vacuum-assisted technique. The resulting scaffold provides sustained release of IL-4 and bioactive nHA. These signals work synergistically to polarize macrophages toward the M2 phenotype and subsequently promote bone regeneration by modulating the PI3K-AKT and calcium signaling pathways.

Article Snippet: The SG/IL-4 hydrogel was prepared by homogeneously mixing the SG hydrogel precursor solution with recombinant mouse interleukin-4 (IL-4, Solarbio, China, cat. no. P00196) to achieve a final concentration of 1 μg/mL.

Techniques: Construct, Protein-Protein interactions

Journal: NPJ Parkinson's disease

Article Title: Altered expression of the immunoregulatory ligand-receptor pair CD200-CD200R1 in the brain of Parkinson's disease patients.

doi: 10.1038/s41531-022-00290-2

Figure Lengend Snippet: Fig. 2 CD200R1 expression in human microglia-like cells treated with inflammatory stimuli. mRNA expression of the four CD200R1 mRNA variants in microglia-like cells treated for 24 h with LPS (100 ng/mL) (a) or IL4 (50 ng/mL) (b) by qRT-PCR after gene-specific retrotranscription. GAPDH and RPS18 were used as reference genes. Individual points are represented and bars indicate the positions of the mean ± SEM of 3–4 independent experiments. *p < 0.05 and **p < 0.01; two-tailed paired Student’s t-test.

Article Snippet: Cells were treated with LPS from Escherichia coli (100 ng/mL; 026:B6; Sigma-Aldrich) or IL4 (50 ng/mL; recombinant mouse IL4 expressed in CHO cells, Creative BioMart, Shirley, NY, USA) and processed after 24 h. Control and treated cells were derived from the same donor.

Techniques: Expressing, Quantitative RT-PCR, Two Tailed Test

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: We treated cDCs at day 6-7 of culture with IL-4 or left them untreated for 24 h. We then stimulated the cDCs with CpG 1826 (10 ug/ml) or R848 (1 ug/ml) for 6 h and then analyzed by qPCR the expression of the IFN-responsive genes ISG15 and Mx-1. All of the conditions were normalized against the control (untreated DCs in medium only) in each experiment. Results are average of three independent experiments, performed with three independent bone marrow-derived cultures from 3 different mice; * p <0.05; ** p <0.01.

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Expressing, Derivative Assay

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: A. We analyzed the gene expression of IRF7 by qPCR after stimulation with CpG 1826 (10 ug/ml) or R848 (1 ug/ml) for 6 h in cDCs treated with IL-4 or left untreated for 24 h. All of the conditions were normalized against the control (untreated DCs in medium only) in each experiment. Results are the average of three independent experiments; * p <0.05; ** p <0.01. B. We analyzed the IRF7 protein expression by Western blotting in cDCs 8 h after stimulation with CpG or R848. Bar graph represents the ratio of integrated density values of IRF7 vs GAPDH (loading control). One representative blot is shown from testing five independent cultures.

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Expressing, Western Blot

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: A . We treated cDCs with IL-4 or left them untreated for 24 h. We then stimulated the cDCs with CpG or R848 for 24 h and then analyzed MHC class I expression. Histogram bars represent averages and standard errors (SE) of the median fluorescence intensity (MdFI) of seven experiments conducted with seven independent cDC cultures; *p<0.05,**p<0.01. B . Western Blot analyses of STAT2 and STAT1 expression in cDCs treated or not IL-4 for 24 h and then stimulated for 8 h with 10 ug/ml of CpG 1826 or 1 ug/ml of R848. Actin was used as loading control. We show a representative blot of three independent experiments. Bar graph represents the ratio of integrated density values of STAT1 or STAT2 vs Actin (loading control).

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Expressing, Fluorescence, Western Blot

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: A. We analyzed the gene expression of IL-6, TNFα, IL-12p35 and IL-12p40 by qPCR after stimulation with CpG or R848 for 6 h in cDCs treated or not with IL-4 for 24 h. Averages and SE of four independent cDC cultures are shown (* p <0.05; ** p <0.01, *** p <0.001). B. We measured by ELISA the levels of IL-6, TNFα and IL-12p70 in the supernatants of DC cultures treated or not with IL-4 for 24 h and then stimulated with R848 or CpG 1826 for 24 h. Averages and SE of four independent cultures are shown.

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Expressing, Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: A. We analyzed by qPCR the gene expression of the IFN-responsive genes IRF7, ISG15 and Mx-1 induced by 6 h stimulation with CpG or R848 in wild type (WT) (C57BL/6) and STAT6-KO cDCs treated or not for 24 h with IL-4. Results were normalized to WT control cells not treated with IL-4. B. We analyzed the MHC Class I expression on cDCs by flow cytometry. Results are shown as median fluorescence intensity (MdFI). Averages and SE of three independent BMDC cultures are shown. (* p<0.05,**p<0.01, ***p<0.0001). C. Western Blot analyses of STAT2 expression in cDCs treated or not for 24 h with IL-4 and harvested 8 h after stimulation with CpG or R848 in WT and STAT6-KO BMDCs. GAPDH was used as loading control. One blot representative of three independent experiments is shown. Numbers below the blot represent the percentage of the normalized integrated density values against GAPDH (loading control).

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Expressing, Flow Cytometry, Fluorescence, Western Blot

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: We measured by ELISA the levels of IL-6 and IL-12p70 in the supernatants of cDC cultures from wild type and STAT6-KO mice treated or not for 24 h with IL-4 and then stimulated with CpG or R848 for 6 h (IL-6) or 48 h (IL-12p70). Averages and SE of three independent cultures are shown (* p<0.05).

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: A. We analyzed by qPCR the gene expression of TLR7 and TLR9 in cDCs treated or not with IL-4 for 24 h. Graphs show averages and SE of six independent cultures. B. We analyzed the protein expression of TLR7 in the total lysates of cDC treated or not for 24 h with IL-4. C. We analyzed by immunofluorescence staining TLR7 expression in cDCs after 24 h treatment with IL-4. The nucleus was stained with DAPI and TLR7 was stained using a rabbit polyclonal anti-body to TLR7 and a secondary anti-rabbit IgG conjugated to Cy3. A minimum of five fields in each slide was read for each condition at a magnification of 40×. Picture representative of five independent cultures.

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Expressing, Immunofluorescence, Staining

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: A. We analyzed the gene expression of SOCS1, -2 and -3 in cDCs by qPCR after 24 h treatment with IL-4. Results were normalized against untreated cells. Graphs show averages and SE of 6 independent cultures. B. We analyzed the protein expression of SOCS2 in the total cDC lysates after 24 h treatment with IL-4. Results were normalized against GAPDH. C . We analyzed the protein expression of SOCS2 in wild-type (WT) and STAT6-KO cDCs that had been treated or not with IL-4 for 24 h and then stimulated with CpG or R848 for 8 h. Results were normalized against GAPDH. Blot representative of three sets of experiments.

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Expressing

Journal: PLoS ONE

Article Title: IL-4 Suppresses the Responses to TLR7 and TLR9 Stimulation and Increases the Permissiveness to Retroviral Infection of Murine Conventional Dendritic Cells

doi: 10.1371/journal.pone.0087668

Figure Lengend Snippet: We analyzed the expression of IFN-responsive genes induced by CpG or R848 stimulation in splenic DCs. We cultured the total splenocytes from C57BL/6 mice as 30–50 million cells in 3 ml each well in GM-CSF-complete medium and 2.5 ng/ml of IL-4 overnight and then stimulated with 50 ug CpG or 5 ug R848 for 4 h. Subsequently we sorted the CD11c+ DCs using Miltenyi magnetic bead separation. All the conditions were normalized against the control (GM-CSF medium only) in each experiment. Line graphs show the response calculated as fold change from no IL-4 control of three independent experiments (* p <0.05).

Article Snippet: Resting DC cultures at day 6 or 7 were treated for 24 h with recombinant mouse IL-4 at 2.5 ng/ml (BD Biosciences), a dose we have previously found efficient to affect cDCs .

Techniques: Expressing, Cell Culture