Journal:

Article Title: Interleukin-3, but not granulocyte-macrophage colony-stimulating factor and interleukin-5, inhibits apoptosis of human basophils through phosphatidylinositol 3-kinase: requirement of NF-?B-dependent and -independent pathways

doi: 10.1046/j.1365-2567.2002.01517.x

Figure Lengend Snippet: Interleukin-3 (IL-3) induces CD69 expression on human basophils and eosinophils. Flow cytometry analysis data are shown of CD69 expression in basophils and eosinophils after 18 hr of culture in medium alone or in medium containing 300 pm IL-3. The figure shows representative histograms of three independent experiments. The percentage of positive cells is indicated.

Article Snippet: Specific mouse anti-human CD69 (MCA 736) and isotype-matched control antibody [mouse immunoglobulin G2b (IgG2b) negative control, MCA 691] were obtained from Serotec (Toronto, Ontario, Canada).

Techniques: Expressing, Flow Cytometry

Journal:

Article Title: Interleukin-3, but not granulocyte-macrophage colony-stimulating factor and interleukin-5, inhibits apoptosis of human basophils through phosphatidylinositol 3-kinase: requirement of NF-?B-dependent and -independent pathways

doi: 10.1046/j.1365-2567.2002.01517.x

Figure Lengend Snippet: The phosphatidylinositol 3-kinase (PI3-K) inhibitor LY294002 (LY) blocks interleukin-3 (IL-3)-induced cell survival and CD69 expression in human basophils. (a) LY inhibits the IL-3-induced basophil survival. Freshly isolated human basophils were cultured in medium alone or with 300 pm IL-3, or in the presence or absence of LY294002 (25 µm or 50 µm), or 50 µm PD98059. At 48 hr, basophils were collected and stained with annexin-V and propidium iodide (prop. iod.), and analysed by flow cytometry. Annexin-V fluorescein isothiocyanate (FITC) and propidium iodide negativity indicates cell viability (% cell survival). *P < 0·05 when compared with IL-3 alone. (b) LY inhibits CD69 expression on basophils. The experimental condition was the same as in (a), except that the cells were cultured for 18 hr and then stained with anti-CD69 or isotype-control antibody (IgG2b). A representative histogram is shown of human basophils stained with anti-CD69 or control antibody after 18 hr of culture with IL-3 alone and IL-3 plus 50 µm LY. Values in parenthesis indicate the mean fluorescence intensity. The figure is representative of three independent experiments.

Article Snippet: Specific mouse anti-human CD69 (MCA 736) and isotype-matched control antibody [mouse immunoglobulin G2b (IgG2b) negative control, MCA 691] were obtained from Serotec (Toronto, Ontario, Canada).

Techniques: Expressing, Isolation, Cell Culture, Staining, Flow Cytometry, Control, Fluorescence

Journal:

Article Title: Interleukin-3, but not granulocyte-macrophage colony-stimulating factor and interleukin-5, inhibits apoptosis of human basophils through phosphatidylinositol 3-kinase: requirement of NF-?B-dependent and -independent pathways

doi: 10.1046/j.1365-2567.2002.01517.x

Figure Lengend Snippet: Lactacystin (LC) inhibits interleukin-3 (IL-3)-induced cell survival and CD69 surface expression in human basophils. (a) LC inhibits IL-3-induced cell survival. Freshly isolated human basophils were cultured in medium alone, or in medium containing IL-3 or IL-3 + LC, for 48 hr. Basophils were then collected and stained with annexin-V and propidium iodide, and analysed by using flow cytometry. Annexin-V fluorescein isothiocyanate (FITC) and propidium iodide (prop. iod.) negativity indicates cell viability (% cell survival). These data presented here are representative of three experiments. (b) LC inhibits IL-3-induced cell-surface expression of CD69. A representative histogram is shown of human basophils stained with anti-CD69 or control antibody after 18 hr of culture with IL-3 alone or IL-3 + 10 µm LC. The experimental conditions were the same as in (a), except that the cells were cultured for 18 hr and then stained with anti-CD69 or isotype-control antibody (IgG2b). Values in parenthesis indicate the mean fluorescence intensity. The figure is representative of three independent experiments.

Article Snippet: Specific mouse anti-human CD69 (MCA 736) and isotype-matched control antibody [mouse immunoglobulin G2b (IgG2b) negative control, MCA 691] were obtained from Serotec (Toronto, Ontario, Canada).

Techniques: Expressing, Isolation, Cell Culture, Staining, Flow Cytometry, Control, Fluorescence

Journal:

Article Title: Interleukin-3, but not granulocyte-macrophage colony-stimulating factor and interleukin-5, inhibits apoptosis of human basophils through phosphatidylinositol 3-kinase: requirement of NF-?B-dependent and -independent pathways

doi: 10.1046/j.1365-2567.2002.01517.x

Figure Lengend Snippet: A schema presenting our hypothesis that interleukin-3 (IL-3) inactivates phosphatidylinositol 3-kinase (PI3-K), initiating signalling cascades that lead to the generation of survival genes through both an NF-κB pathway, as well as through an alternate mechanism. In summary, we demonstrated that the in vitro spontaneous rate of apoptosis of human basophils is higher than that of eosinophils, and that IL-3 inhibits basophil apoptosis as well as up-regulating basophil CD69 surface expression via a PI3-K-dependent mechanism(s) that entails new RNA and protein synthesis, partially via NF-κB signalling. As basophils are active participants in allergic reactions, and IL-3 is one of the most abundant proinflammatory cytokines in the secretions from allergic tissue, IL-3-mediated inhibition of basophil apoptosis may exacerbate the inflammation associated with allergic disorders.

Article Snippet: Specific mouse anti-human CD69 (MCA 736) and isotype-matched control antibody [mouse immunoglobulin G2b (IgG2b) negative control, MCA 691] were obtained from Serotec (Toronto, Ontario, Canada).

Techniques: In Vitro, Expressing, Inhibition

Journal: Frontiers in Immunology

Article Title: Proline metabolism reprogramming of trained macrophages induced by early respiratory infection combined with allergen sensitization contributes to development of allergic asthma in childhood of mice

doi: 10.3389/fimmu.2022.977235

Figure Lengend Snippet: Lung CD11b + trained macrophages increased after PVM-OVA early sensitization and mediated asthma in childhood mice. (A) The lung CD11b + cells in the memory phase after sensitization by flow cytometry. (B) Lung CD11b + macrophages morphologic evaluation. (C) Timeline of mouse adoptive transfer experiment. (1) PBS transfer group, (2) PBS-OVA transfer group, (3) PVM transfer Group, (4) PVM-OVA transfer group. (D) RI and lung Cdyn in mice after adoptive transfer of the lung CD11b + macrophages following OVA challenge. PVM-OVA transfer group compared with PBS transfer group, *, with PBS-OVA transfer group, #, with PVM transfer group, <. (E) Relative expression of MCP-1 and IL-5 in lung tissue following OVA challenge by Real-time PCR. Each experimental group contained three biology repeats. Data are presented as means standard deviations of 5-6 mice per group. (F) H&E staining of lung tissue slices after adoptive transfer the lung CD11b + macrophages (100X) and inflammation scores following OVA challenge. (G) CD11b + CD69 + macrophages or CD11b + TLR4 + macrophages in lungs by flow cytometry. (H) TLR4 expression of the sorted CD11b+ macrophages after LPS or OVA stimulation by flow cytometry. (I) Relative mRNA expression of chemokines MCP-1 or GRO-α of the sorted CD11b + macrophages after LPS or OVA stimulation by real-time PCR. *P<0.05. ** P<0.01, ***P<0.001, **** P<0.001. ## means PVM-OVA transfer group compared with PBS-OVA transfer group (P<0.01). The symbol “■” but not “<” means PVM-OVA transfer group compared with PVM transfer group (P<0.05).

Article Snippet: Lung single cells were incubated with fluorescein labeled antibodies, anti-mouse CD11b-PE, CD69-FITC, TLR4-APC, CD49a-PE-vio770, Gr-1-FITC (MACS, Miltenyi, Germany).

Techniques: Flow Cytometry, Adoptive Transfer Assay, Expressing, Real-time Polymerase Chain Reaction, Staining