Journal:

Article Title: CTLA-4 regulates allergen response by modulating GATA-3 protein level per cell

doi: 10.1111/j.1365-2567.2007.02537.x

Figure Lengend Snippet: Effects of in vitro CTLA-4 blockade on GATA-3 protein level/cell and frequency of IL-4-producing cells. Naive CD4+ T cells purified from the spleens of normal mice were induced to differentiate with anti-CD3 mAb, a recombinant form of CD80-Fc, IL-4 and anti-IFN-γ mAb for 3 days. Anti-CTLA-4 mAb was added in soluble form at increasing concentrations (0·1–10 μg/ml). (a) Mean fluorescence intensity for GATA-3 staining, (b) percentage of IL-4+ cells.

Article Snippet: 18 CD4 cell culture For in vitro experiments, splenic naive CD4 cells were purified from non-immunized BALB/c mice by immuno-magnetic cell sorting as previously described 19 and cultured in 24-well plates precoated with anti-CD3ε mAb (clone 145-2C11; 10 μg/ml) and/or recombinant mouse B7-1(CD80)/Fc chimeric protein (R & D Systems, cat. 740-B1; 0·33 μg/ml).

Techniques: In Vitro, Purification, Recombinant, Fluorescence, Staining

Journal: Current protocols

Article Title: Redefining Cell Culture Using a 3D Flipwell Co-culture System: A Mimetic for Gut Architecture and Dynamics In Vitro.

doi: 10.1002/cpz1.70107

Figure Lengend Snippet: Figure 15 THP-1 dramatic polarization toward an M1-like phenotype with Sepiapterin treatment in the 3D Flipwell coculture system.(A) Confocal images of THP-1 cells cocultured with Caco-2:HT- 29 in the 3D Flipwells and stained for pan macrophage marker CD68 (red) and CD80 (green). Pan macrophage CD68 marker is present in both the SEP treated sample and the untreated control. CD80 marker, typical of the M1 polarization phenotype, is more prominently present in the SEP treated sample with long pseudopodia extending outward typical of the M1 phenotype. (B) Con- focal images of THP-1 cells cocultured with Caco-2:HT-29 in the 3D Flipwell and stained for pan macrophage marker CD68 and CD163 to validate phenotypic polarization toward M1 phenotype and away from M2 phenotype. Scale bar = 25 μm.

Article Snippet: See the T erm s and C onditions (https://onlinelibrary.w iley.com s-and-conditions) on W iley O nline L ibrary for rules of use; O A articles are governed by the applicable C reative C om m ons L icense Anti-human MUC2 (996/1) (Thermo Fisher, cat. no. MA5-12345) Anti-human CK20 (Life Technologies, cat. no. PA5-82875) Anti-human CD80 (Cell Signaling Technologies, E3Q9V, cat. no. 15416) Anti-human CD163 (Abcam, cat. no. 156769) or anti-human CD68 (Novus Biologicals, cat. no. NB100-683) Secondary antibodies: Alexa Fluor 488 goat anti-rabbit IgG (Thermo Fisher, cat. no. A11008) for CD80 and CK20 Alexa Fluor 594 goat anti-mouse IgG (Thermo Fisher, cat. no. A11005) for MUC2 and CD68 or CD163 DAPI, VectaShield mounting medium (Vector Laboratories, cat. no. H-1000) 50-ml conical tubes (SpectraTube, Alkali Scientific cat. no. C2750) Plastic wrap Lab tape or shipping tape Tweezers 12-well tissue culture plate (Corning Costar, cat. no. 3513) 6-well dish Aluminum foil Scalpel Round cover slips, #1, 1 oz (Corning Float Glass, cat. no. 50143822, or Fisherbrand cat. no. 12-546-2 25CIR-2) Laboratory marker (VWR, cat. no 95042-566) Attofluor cell chamber coverslip holder (Thermo Fisher Scientific, cat. no. A7816) Confocal microscope and software [e.g., Leica Microsystems TCS SP5, multi-photon laser scanning, and Suite Advanced Fluorescence (LAS AF) software] Deep Petri dish bottoms, 100 × 25–mm deep polystyrene stackable Petri dish, sterile (USA Scientific, cat. no. 8609-0625) 10-, 200-, and 1000-μl pipet tips (Alkali Scientific PurePoint or equivalent) Beckman Coulter Allegra 6 centrifuge with a swing bucket rotor for 50-ml conical tubes Optional: ImmunoPen (Fisher Scientific, cat. no. 4021761EA) Wash, permeabilization, disassembly, and blocking steps 1.

Techniques: Staining, Marker, Control

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Co-targeting CD47 and VEGF elicited potent anti-tumor effects in gastric cancer

doi: 10.1007/s00262-024-03667-9

Figure Lengend Snippet: Targeting CD47 significantly inhibited the growth of gastric cancer in Hu-PDX models. A In the Hu-PDX1 model, tumor volume was measured twice a week and presented as mean ± SD. After treatment with SIRPα-Fc for 4 weeks, tumor weight was presented. ( n = 6 for control group and n = 8 for SIRPα-Fc group, * P < 0.05, ** P < 0.01) B Each line represented the tumor volume from an independent mouse in the Hu-PDX1 model. C In the Hu-PDX2 model, tumor volume was measured twice a week and presented as mean ± SD. After treatment with SIRPα-Fc for 4 weeks, tumor weight was presented. ( n = 6 per group, * P < 0.05, ** P < 0.01). D Each line represented the tumor volume from an independent mouse in the Hu-PDX2 model. E Representative photographs of immunohistochemical staining for CD80, CD163, and CD8 of tumor tissue sections and the number of CD80 + , CD163 + , and CD8 + cells in each group were normalized to the control group. The value of control was set to 1.0. ( * P < 0.05, ** P < 0.01)

Article Snippet: Anti-CD80 monoclonal antibody (66,406-1-Ig, Proteintech), anti-CD163 monoclonal antibody (GB13340, Servicebio), anti-CD8 monoclonal antibody (GB12068, Servicebio), anti-CD31 monoclonal antibody (GB113151, Servicebio), VEGFA Monoclonal antibody (19,003-1-AP, Proteintech), anti-CD47 monoclonal antibody (ab218810, Abcam), carboxyfluorescein diacetate succinimidyl ester (CFDA SE) (C0051, Beyotime), PerCP anti-CD68 (333,813, BioLegend), PE anti-CD11b (101,208, BioLegend), granulocyte–macrophage colony-stimulating factor (GM-CSF) (C003, novoprotein), FITC-labeled anti-CD47 (CC2C6, BioLegend), Human Lymphocyte separation medium (7,111,011, DAKEWE).

Techniques: Control, Immunohistochemical staining, Staining

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Co-targeting CD47 and VEGF elicited potent anti-tumor effects in gastric cancer

doi: 10.1007/s00262-024-03667-9

Figure Lengend Snippet: CD47 blockade combined with antiangiogenetic therapy elicited enhanced anti-tumor effect in Hu-PDX models of gastric cancer. A and B In the Hu-PDX1 model, tumor-bearing mice were treated with SIRPα-Fc and/or VEGFR1-Fc for 4 weeks, tumor volume and tumor weight were presented as mean ± SD. Each line represented the value of the tumor volume of a single mouse. C and D In the Hu-PDX2 model, tumor volume and tumor weight were presented after the same treatment in the Hu-PDX1 model. Each line represented the value of the tumor volume of a single mouse. ( n = 6 per group, * P < 0.05, ** P < 0.01). E Representative photographs of immunohistochemical staining for CD80, CD163, CD8, and CD31 of tumor tissue sections and the number of CD80 + , CD163 + , and CD8 + cells and the relative vessel density in each group were normalized to the control group. The value of control was set to 1.0. (* P < 0.05, ** P < 0.01)

Article Snippet: Anti-CD80 monoclonal antibody (66,406-1-Ig, Proteintech), anti-CD163 monoclonal antibody (GB13340, Servicebio), anti-CD8 monoclonal antibody (GB12068, Servicebio), anti-CD31 monoclonal antibody (GB113151, Servicebio), VEGFA Monoclonal antibody (19,003-1-AP, Proteintech), anti-CD47 monoclonal antibody (ab218810, Abcam), carboxyfluorescein diacetate succinimidyl ester (CFDA SE) (C0051, Beyotime), PerCP anti-CD68 (333,813, BioLegend), PE anti-CD11b (101,208, BioLegend), granulocyte–macrophage colony-stimulating factor (GM-CSF) (C003, novoprotein), FITC-labeled anti-CD47 (CC2C6, BioLegend), Human Lymphocyte separation medium (7,111,011, DAKEWE).

Techniques: Immunohistochemical staining, Staining, Control

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Co-targeting CD47 and VEGF elicited potent anti-tumor effects in gastric cancer

doi: 10.1007/s00262-024-03667-9

Figure Lengend Snippet: Bispecific fusion protein SIRPα-VEGFR1 elicited synergetic antitumor effect and prevented gastric cancer recurrence. A and B In the Hu-PDX1 model, tumor volume and tumor weight were measured and the data was presented as mean ± SD after treatment with SIRPα-Fc plus VEGFR1-Fc, and SIRPα-VEGFR1 for 4 weeks. Each line represented the value of the tumor volume of a single mouse. C and D In the Hu-PDX2 model, tumor volume and tumor weight were measured after the same treatment in the Hu-PDX1 model. E The number of CD80 + , CD163 + , and CD8 + cells and the relative vessel density in each group were normalized to the control group. The value of control was set to 1.0. (* P < 0.05, ** P < 0.01). F In the humanized tumor recurrence model, mice were treated with control, SIRPα-Fc + VEGFR1-Fc, SIRPα-VEGFR1 for 2 weeks, and tumor volume was measured. Each line represented the value of the tumor volume of a single mouse. G Survival curves for different treatment groups. (n = 6 per group, * P < 0.05, ** P < 0.01)

Article Snippet: Anti-CD80 monoclonal antibody (66,406-1-Ig, Proteintech), anti-CD163 monoclonal antibody (GB13340, Servicebio), anti-CD8 monoclonal antibody (GB12068, Servicebio), anti-CD31 monoclonal antibody (GB113151, Servicebio), VEGFA Monoclonal antibody (19,003-1-AP, Proteintech), anti-CD47 monoclonal antibody (ab218810, Abcam), carboxyfluorescein diacetate succinimidyl ester (CFDA SE) (C0051, Beyotime), PerCP anti-CD68 (333,813, BioLegend), PE anti-CD11b (101,208, BioLegend), granulocyte–macrophage colony-stimulating factor (GM-CSF) (C003, novoprotein), FITC-labeled anti-CD47 (CC2C6, BioLegend), Human Lymphocyte separation medium (7,111,011, DAKEWE).

Techniques: Control

Journal: Bioactive Materials

Article Title: A tantalum-containing zirconium-based metallic glass with superior endosseous implant relevant properties

doi: 10.1016/j.bioactmat.2024.04.014

Figure Lengend Snippet: Macrophage responses to different surface chemistry. A) ELISA results of IL-1β, TNF-α and IL-10. Ta0 and Ta3-A significantly suppressed the secretion of pro-inflammatory cytokines. B) qRT-PCR results of M1 and M2 activity related markers. Ta0 and Ta3-A significantly reduced the expression of pro-inflammatory markers. C) Heatmap of representative overlapped genes between Ta3-A vs cpTi and Ta0 vs cpTi comparisons, showing an overall decrease in pro-inflammatory genes. D) Western blot result of iNos, Cd80, Cd206 and β-actin. Data are presented as mean ± SE. * p < 0.05, ** p < 0.01 and *** p < 0.001.

Article Snippet: The membranes were blotted with 5 % non-fat dry milk (Sigma) for 1 h and then incubated overnight with CD80 Rabbit mAb (1:1000, Abclonal, Cat No: A23688), Mannose Receptor/CD206 Rabbit mAb (1:1000, Abclonal, Cat No: A21014), iNOS Polyclonal antibody (1:1000, Proteintech, Cat No: 22226-1-AP), BMP4 Polyclonal antibody (1:1000, Proteintech, Cat No: 12492-1-AP), SMAD9 Polyclonal antibody (1:1000, Proteintech, Cat No: 16397-1-AP), p -SMAD1/5/8 Rabbit mAb (1:1000, Cell Signaling, Cat No: 13820T), Alkaline Phosphate (ALPL) Rabbit mAb (1:1000, Abclonal, Cat No: A0514), Rabbit Anti-beta-Actin/HRP antibody (1:1000, Bioss, Cat No: bsm-52846R), followed by incubation with a goat anti-rabbit IgG secondary antibody HRP conjugated (1:1000, Beyotime, Cat No: A0208).

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

Journal: Molecular Biomedicine

Article Title: EZH2 coordinates inflammatory and pain-associated pathways across joint compartments in osteoarthritis

doi: 10.1186/s43556-026-00643-w

Figure Lengend Snippet: Proposed model illustrating the effects of EZH2 inhibition across multiple joint compartments in osteoarthritis. In OA fibroblast-like synoviocytes (FLS), increased EZH2 and H3K27me3 levels were associated with inflammatory and catabolic responses, which were reduced by pharmacological EZH2 inhibition with EPZ-6438. In human OA FLS, EPZ-6438 decreased inflammatory, catabolic and pain-associated mediators, including IL1B, MMP13, NGF and NTN1, reduced LDHA expression and increased autophagy-associated responses. Proteomic and ChIP-seq analyses further identified inflammatory, metabolic and axon guidance-related pathways, including inflammation-dependent EZH2 recruitment at the PAX6 locus. EZH2 inhibition also reduced inflammatory activation of macrophages and osteoclastogenesis in OA-derived bone marrow cells. Collectively, these findings support a broader role for EZH2 in inflammatory, catabolic and osteoimmune responses across multiple OA-relevant cellular compartments. The identification of PAX6, NTN1 and axon guidance-related pathways highlights neuro-associated molecular programs that may be relevant to pain-associated responses, although their direct contribution to pain or sensory nerve remodeling was not assessed in this study. The reduced structural joint damage and pain-related behaviors observed following EZH2 inhibition in the MIA-induced OA model are consistent with this multicellular framework

Article Snippet: The following antibodies were used for immune cell characterization: CD11b (REA592 VioBlue, 130–113–810, Miltenyi Biotec), F4/80 (REA126 PE-Vio770, 130–118–459, Miltenyi Biotec), CD80 (REA983 APC-Vio770, 130–116–463, Miltenyi Biotec), CD86 (REA1190 PE, 130–122–129, Miltenyi Biotec), MHC-II (REA813 FITC, 130–112–386, Miltenyi Biotec), CD38 (REA616 PerCP-Vio700, 130–109–260, Miltenyi Biotec), and iNOS (REA982 APC, 130–116–423, Miltenyi Biotec).

Techniques: Inhibition, Expressing, ChIP-sequencing, Activation Assay, Derivative Assay