Journal: Cellular immunology

Article Title: TSLP signaling blocking alleviates E-cadherin dysfunction of airway epithelium in a HDM-induced asthma model.

doi: 10.1016/j.cellimm.2017.02.003

Figure Lengend Snippet: Figure 1. HDM increases the expression of TSLP through PI3K/Akt signaling pathway in 16HBE cells (A) Western blot analysis of TSLP protein levels in 16HBE stimulated with HDM for the indicated times. (B) Western blot analysis of TSLP protein levels in 16HBE stimulated with HDM for the indicated concentrations. (C) The 16-HBE cells were treated with HDM with or without 50 uM LY294002, The expression of phosphorylated Akt and TSLP were measured by Western Blot Analysis. Data are presented as the mean ± SD. n=3. *P<0.05 vs. the control group. #P <0.05, vs. the HDM group

Article Snippet: HDM was purchased from ALK-Abello A/S (Denmark), methacholine was obtained from Sigma-Aldrich, anti-TSLP mAb was purchased from R&D Systems (MAB555; USA), and recombinant human TSLP was obtained from R&D systems.

Techniques: Expressing, Western Blot, Control

Journal: Cancers

Article Title: Non-Expressed Donor KIR3DL1 Alleles May Represent a Risk Factor for Relapse after T-Replete Haploidentical Hematopoietic Stem Cell Transplantation

doi: 10.3390/cancers15102754

Figure Lengend Snippet: Intracellular localization of KIR3DL1*019. Flow cytometry analysis of Jurkat cell line after stable transfection with KIR3DL1*002, *004, *019, *019 L86S , *019 S182P , and *019 L86S+S182P chimeric constructs containing eGFP. Only cells positive for eGFP fluorescence were examined. ( a ) Specific Z27 3.7 anti-KIR3DL1 mAb (blue peak; Beckman Coulter) with isotype MOPC-21 IgG control mAb (green peaks; Sony) were used in extracellular staining; ( c ) specific 177,407 anti-KIR3DL1 mAb (blue peak; R&D Systems) with isotype 11,711 IgG control (green peaks; R&D Systems) were used in intracellular staining. Fluorescent microscopy of Jurkat cells stably transfected with KIR3DL1*002, *004, *019, *019 L86S , *019 S182P , and *019 L86S+S182P chimeric constructs containing eGFP. F41-1E3 anti-HLA class I mAb (EFS Nantes) was used in ( b ) extracellular and ( d ) intracellular transfectant staining. Magnification 100×. Specific Z27 3.7 (Beckman Coulter) and 177,407 (R&D Systems) anti-KIR3DL1 mAbs were used in extracellular and intracellular staining, respectively. Merged images show co-localization of KIR3DL1-eGFP, HLA class I delimiting plasma membrane and KIR3DL1 in extracellular and intracellular staining.

Article Snippet: Intracellular labeling was performed on cells permeabilized with BD Cytofix/Cytoperm solution (Becton Dickinson, Franklin Lakes, NJ, USA) using the AF647-conjugated anti-KIR3DL1 mAb (clone 177407; R&D Systems), the corresponding IgG1 isotype control (clone 11711; R&D Systems), the AF555-conjugated anti-HLA class I mAb (clone F41-1E3), and the corresponding AF555-IgG1 isotype control (clone MOPC-21).

Techniques: Flow Cytometry, Stable Transfection, Construct, Fluorescence, Control, Staining, Microscopy, Transfection, Clinical Proteomics, Membrane

Journal: PLoS ONE

Article Title: IL-23 Induces Atopic Dermatitis-Like Inflammation Instead of Psoriasis-Like Inflammation in CCR2-Deficient Mice

doi: 10.1371/journal.pone.0058196

Figure Lengend Snippet: On day 12, ( A ) mRNA of indicated cytokines was measured by real-time RT-PCR from ears of WT PBS-injected or WT and CCR2 −/− IL-23-injected mice. Average of 11 mice per genotype in 3 experiments. *p<0.01. ( B ) Intracellular cytokine staining for IL-4 on gated CD3 + CD4 + T cells following stimulation with PMA and ionomycin was performed and day 12 and analyzed by flow cytometry. Number of IL-4 + CD3 + CD4 + T cells in draining lymph node (left panel) or IL-23-injected ear (right panel). ( C ) TSLP was measured by tissue ELISA of ears isolated from PBS-injected or IL-23-injected WT and CCR2 −/− mice. Average of 3 mice per genotype. *p<0.005 CCR2 KO IL-23 vs. all other groups. ( D ) Representative immunofluorescence staining of ear skin isolated on day 12 from IL-23-injected WT C57Bl/6 or CCR2 −/− mice. Sections were stained with Ig isotype control antibody or anti-TSLP (x600, original magnification). Data are reflective of 3 mice per genotype.

Article Snippet: Sections were then washed twice in PBS 0.1% Tween and stained overnight with 1 μg of an affinity purified polyclonal antibody to mouse TSLP (R & D Systems, BAF555) diluted in PBS 0.1% saponin at 4 ° C. After washing with PBS 0.1% Tween on ice, sections were stained with 1 μg of AlexaFluor 488-conjugated donkey anti-goat IgG (Invitrogen) for 1 hour at room temperature.

Techniques: Quantitative RT-PCR, Injection, Staining, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Isolation, Immunofluorescence, Control

Journal: bioRxiv

Article Title: Distinct roles for thymic stromal lymphopoietin (TSLP) and IL-33 in experimental eosinophilic esophagitis

doi: 10.1101/2025.02.25.640192

Figure Lengend Snippet: Experimental EoE was induced in wild type (WT) mice using oxazolone (OXA). Starting on day 19, the mice received two intraperitoneal injections per week of isotype control antibodies or anti-TSLP (clones M702 or clone 28F12, A). On day 36, the mice were euthanized, and esophageal tissues were fixed, paraffin embedded, and slides were generated. The slides were stained with anti-major basic protein (MBP, B) and esophageal eosinophils were quantified (C). Epithelial cell proliferation was determined using anti-Ki67 staining (D, E). H&E-stained slides (G) were analysed for lamina propria (F) as well as epithelial (H) thickness. Esophageal vascularization was determined using anti-CD31 staining (I, J). Single cell suspensions of esophageal tissue were obtained from oxazolone-challenged isotype control or anti-TSLP (M702)-treated mice using enzymatic digestion and the levels of eosinophils and CD4 + T cells determined by flow cytometry (K, L). Representative photomicrographs of anti-MBP (B), anti-Ki-67 (D), H&E (G) and anti-CD31 (I) are shown. Data are presented as mean ± SEM and are representative of n=3 experiments conducted with 10-12 mice per group, ns-nonsignificant, *-p<0.05, **-p<0.01, ***-p<0.001, ****-p<0.0001 .

Article Snippet: To substantiate these findings, TSLP was neutralized using an additional clone of anti-TSLP (BioXcell, clone 28F12).

Techniques: Control, Clone Assay, Generated, Staining, Flow Cytometry

Journal: bioRxiv

Article Title: Distinct roles for thymic stromal lymphopoietin (TSLP) and IL-33 in experimental eosinophilic esophagitis

doi: 10.1101/2025.02.25.640192

Figure Lengend Snippet: Experimental EoE was induced in wild type (WT) mice. Starting on day 19, the mice received two intraperitoneal injections per week of isotype control antibodies or anti-TSLP (clones M702 or clone 28F12). On day 36, the mice were euthanized, and esophageal tissue was obtained and subjected to bulk RNA sequencing. PCA plot of vehicle- and oxazolone (OXA)-challenged mice treated with either isotype control or anti-TSLP antibodies (clones M702 and/or 28F12) is shown (A). Heat plot analysis of all differentially expressed genes (absolute fold change > 2, p<0.05) (B) is presented. Analysis of the top differentially expressed secreted factors, cytokine receptors, adhesion molecules and extracellular matrix components is shown (C-D). Gene Set Enrichment Analysis (GSEA) and STRING analysis were performed on the DEGs, and comparison of OXA-challenged vehicle-treated mice vs. OXA-challenged anti-TSLP treated mice was performed (E-H). The top up and down regulated pathways, which were regulated by TSLP are presented (E-F). In C, D and E, each lane represents a different mouse.

Article Snippet: To substantiate these findings, TSLP was neutralized using an additional clone of anti-TSLP (BioXcell, clone 28F12).

Techniques: Control, Clone Assay, RNA Sequencing, Comparison