Journal: Journal of immunotoxicology

Article Title: The early growth response factor-1 contributes to interleukin-13 production by mast cells in response to stem cell factor stimulation.

doi: 10.1080/15476910802129612

Figure Lengend Snippet: FIG. 4. Egr-1 deficiency has no effects on c-Kit and IgE receptor expression or mast-cell viability. (A) Bone marrow cells from wild-type mice or Egr-1– deficient mice were cultured in conditioned media in vitro for 4 wk and exam- ined by flow cytometry for c-Kit and IgE receptor expression. For c-Kit anal- ysis, BMMCs were stained with FITC conjugated rat anti-mouse c-Kit mAb or FITC conjugated rat IgG2a isotypic control. For analysis of IgE receptor expression, BMMC were sensitized with IgE overnight and then stained with FITC-conjugated anti-IgE antibody (mouse IgG1). No difference in c-Kit or IgE receptor expression was observed between Egr-1+/+ and Egr-1−/−BMMCs. (B) BMMC from Egr-1-deficient mice or wild-type mice cultured in complete media containing WEHI-3B supernatants (a source of IL-3) were healthy and showed ≥96% viability. To induce mast cell death, WEHI-3B supernatant was removed from culture media for various days. Mast cell viability was examined by trypan blue exclusion assay. Data were expressed as mean ± SD (n = 3 independent experiments).

Article Snippet: Fluorescein isothiocyanate (FITC) anti-mouse CD117 monoclonal antibody (mAb) (CL8936F), FITC rat IgG2a (CLCR2A01) were purchased from Cedarlane Laboratories Limited (Ontario, Canada).

Techniques: Expressing, Cell Culture, In Vitro, Cytometry, Staining, Control, Trypan Blue Exclusion Assay

Journal: Frontiers in Cardiovascular Medicine

Article Title: Deficiency of Myeloid Pfkfb3 Protects Mice From Lung Edema and Cardiac Dysfunction in LPS-Induced Endotoxemia

doi: 10.3389/fcvm.2021.745810

Figure Lengend Snippet: Myeloid-specific Pfkfb3 deficiency attenuates LPS-induced inflammatory responses. (A) Representative images (left) and quantification (right) for immunohistochemical staining of the neutrophil marker Ly6G in lung sections of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 5). (B) Representative images (left) and quantification (right) for immunohistochemical staining of the macrophage marker Mac2 in lung sections of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 5). (C–E) qPCR analysis of the mRNA levels of Il1b (C) , Il6 (D) and Nos2 (E) in the lung of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 6). (F,G) ELISA analysis of Il1b (F) and Il6 (G) in serum of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 4). (H) NO levels in serum of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 3–4). All data are represented as mean ± SEM, ** P < 0.01 and *** P < 0.001 for Pfkfb3 WT vs. Pfkfb3 ΔMϕ (unpaired two-tailed Student's t test).

Article Snippet: After antigen retrieval with Antigen Unmasking Solution (H-3301, Vector Laboratories, Burlingame, CA, USA) at 98°C for 10 min, sections were blocked with avidin solution with 10% normal rabbit serum for 1 h at room temperature, and incubated in biotin blocking solution with primary antibodies against Mac2 (3 μg/mL, CL8942F, Cedarlane, Burlington, NC, USA), or Ly6G (3 μg/mL, 551459, BD biosciences, San Jose, CA, USA) at 4°C overnight.

Techniques: Immunohistochemical staining, Staining, Marker, Injection, Enzyme-linked Immunosorbent Assay, Two Tailed Test

Journal: PLoS ONE

Article Title: Syntaxin Binding Protein 1 Is Not Required for Allergic Inflammation via IgE-Mediated Mast Cell Activation

doi: 10.1371/journal.pone.0058560

Figure Lengend Snippet: A, Wild type, STXBP1 mutant, or heterozygous mice were identified by PCR using DNA from liver cells. B , After 4–5 weeks of culture, liver-derived mast cells (LMC) were examined by toluidine blue staining to determine mast cell purity. A representative photomicrograph is shown (original magnification = ×100). C , LMCs were sensitized with anti-TNP IgE, stained with FITC-conjugated anti-IgE or anti-c-kit antibodies, and examined by flow cytometry. D, STXBP1 +/+ and STXBP1 −/− LMCs were sensitized with anti-TNP IgE and stimulated with TNP-BSA (10 ng/ml) for various times. Whole cell lysates were analyzed by Western blotting for STXBP1 and Actin. E, Sec1/Munc18 (SM) family gene expression profiles were identified by RT-PCR using mRNA from wild type mouse bone marrow-derived mast cells (BMMC), and wild type (STXBP1 +/+ ) or STXBP1 −/− LMCs, with or without TNP-BSA stimulation. All members of SM family are expressed in wild type BMMCs and LMCs. Similar results were observed in 2 independent experiments.

Article Snippet: FITC-conjugated rat anti-mouse IgE (IgG1), FITC-rat IgG1 and FITC-conjugated rat anti-mouse CD117 (c-kit) were purchased from Cedarlane Laboratories (Burlington, ON, Canada).

Techniques: Mutagenesis, Derivative Assay, Staining, Flow Cytometry, Western Blot, Gene Expression, Reverse Transcription Polymerase Chain Reaction