control empty vectors plv-egfp  (Genechem)

Journal: Journal of Leukocyte Biology

Article Title: IL-4Rα on dendritic cells in neonates and Th2 immunopathology in respiratory syncytial virus infection

doi: 10.1189/jlb.4A1216-536R

Figure Lengend Snippet: (A) Flow cytometric histogram graphs show a representative example of IL-4Rα expression on DC subsets. (B) IL-4Rα MFI on CD11b+ mDCs. (C) IL-4Rα MFI on CD103+ mDCs. (D) IL-4Rα MFI on pDCs. Shaded histograms represent FMO controls. Data are representative of 3 independent experiments with 4–5 mice/group. *P < 0.05.

Article Snippet: BMDCs were then harvested and infected with third-generation lentiviral vectors (pLV-IL-4Rα-eGFP or control pLV-eGFP) (Cyagen Biosciences, Santa Clara, CA, USA) at a multiplicity of infection 20 in Opti-MEM media (Thermo Fisher Scientific) and 5 µg/ml of polybrene (Cyagen Biosciences) for 3 h. These cells were cultured in DC medium for 4 d. Transduced cells were then infected with RSV at a multiplicity of infection 50 overnight, and CD11b + mDCs that were positive for eGFP were sorted with a SH800 cell sorter (Sony Biotechnology, San Jose, CA, USA).

Techniques: Expressing

Journal: Journal of Leukocyte Biology

Article Title: IL-4Rα on dendritic cells in neonates and Th2 immunopathology in respiratory syncytial virus infection

doi: 10.1189/jlb.4A1216-536R

Figure Lengend Snippet: IL-4Rα−/−DC or IL-4Rα−/loxDC mice were infected at 5 d old and reinfected with RSV 4 wk later (IL-4Rα−/−DCRR and IL-4Rα−/loxDCRR, respectively). Control mice were sham infected (IL-4Rα−/−DCsham and IL-4Rα−/loxDCsham). Pulmonary Th profile was measured via flow cytometry at 6 d after reinfection, and virus gene expression was calculated via real-time RT-PCR at 4 d after primary infection. (A) Percentage (upper panel) and numbers (middle panel) of pulmonary CD4+ T cells expressing IFN-γ, IL-4, or both IFN-γ and IL-4. The bottom panel displays representative flow plots. (B) Relative expression of the RSV NS1 gene in the lung at 4 d after primary infection. Data are representative of 3 independent experiments with 4–5 mice/group. *P < 0.05.

Article Snippet: BMDCs were then harvested and infected with third-generation lentiviral vectors (pLV-IL-4Rα-eGFP or control pLV-eGFP) (Cyagen Biosciences, Santa Clara, CA, USA) at a multiplicity of infection 20 in Opti-MEM media (Thermo Fisher Scientific) and 5 µg/ml of polybrene (Cyagen Biosciences) for 3 h. These cells were cultured in DC medium for 4 d. Transduced cells were then infected with RSV at a multiplicity of infection 50 overnight, and CD11b + mDCs that were positive for eGFP were sorted with a SH800 cell sorter (Sony Biotechnology, San Jose, CA, USA).

Techniques: Infection, Flow Cytometry, Expressing, Quantitative RT-PCR

Journal: Journal of Leukocyte Biology

Article Title: IL-4Rα on dendritic cells in neonates and Th2 immunopathology in respiratory syncytial virus infection

doi: 10.1189/jlb.4A1216-536R

Figure Lengend Snippet: Deletion of IL-4Rα on CD11c + cells attenuated Th2-biased cytokine responses in the lung upon RSV reinfection

Article Snippet: BMDCs were then harvested and infected with third-generation lentiviral vectors (pLV-IL-4Rα-eGFP or control pLV-eGFP) (Cyagen Biosciences, Santa Clara, CA, USA) at a multiplicity of infection 20 in Opti-MEM media (Thermo Fisher Scientific) and 5 µg/ml of polybrene (Cyagen Biosciences) for 3 h. These cells were cultured in DC medium for 4 d. Transduced cells were then infected with RSV at a multiplicity of infection 50 overnight, and CD11b + mDCs that were positive for eGFP were sorted with a SH800 cell sorter (Sony Biotechnology, San Jose, CA, USA).

Techniques:

Journal: Journal of Leukocyte Biology

Article Title: IL-4Rα on dendritic cells in neonates and Th2 immunopathology in respiratory syncytial virus infection

doi: 10.1189/jlb.4A1216-536R

Figure Lengend Snippet: IL-4Rα−/−DC or IL-4Rα−/loxDC mice were infected and reinfected with RSV, as previously described. BALF cellularity, airways resistance in response to methacholine (MeCh) challenge, and histology were performed at 6 d after reinfection. (A) BALF cellularity. Macs, macrophages; Lymph, lymphocytes; Neutro, neutrophils; Eos, eosinophils. (B) Airway resistance. (C) H&E-stained lung sections and inflammation score. Arrowheads point to cellular infiltrates. (D) PAS-stained lung sections and PAS score. Arrowheads point to mucus. Scale bar, 400 μm in the upper panels. The bottom panels are images taken under higher magnification of the selected area in the upper panel (inset box). Scale bar, 100 μm in the bottom panels. Data are representative of 2 independent experiments with 4–5 mice/group. *P < 0.05.

Article Snippet: BMDCs were then harvested and infected with third-generation lentiviral vectors (pLV-IL-4Rα-eGFP or control pLV-eGFP) (Cyagen Biosciences, Santa Clara, CA, USA) at a multiplicity of infection 20 in Opti-MEM media (Thermo Fisher Scientific) and 5 µg/ml of polybrene (Cyagen Biosciences) for 3 h. These cells were cultured in DC medium for 4 d. Transduced cells were then infected with RSV at a multiplicity of infection 50 overnight, and CD11b + mDCs that were positive for eGFP were sorted with a SH800 cell sorter (Sony Biotechnology, San Jose, CA, USA).

Techniques: Infection, Staining

Journal: Journal of Leukocyte Biology

Article Title: IL-4Rα on dendritic cells in neonates and Th2 immunopathology in respiratory syncytial virus infection

doi: 10.1189/jlb.4A1216-536R

Figure Lengend Snippet: Neonatal IL-4Rα−/−DC and IL-4Rα−/loxDC were infected with RSV. CD11b+ mDCs were isolated from those mice at 6 d after infection and cultured with CD4+ T cells purified from the spleens of naïve, adult BALB/c mice. Th profile was measured after 72 h coculture using flow cytometry. (A) Percentage of pulmonary CD4+ T cells expressing IFN-γ. (B) Percentage of pulmonary CD4+ T cells expressing IL-4. (C) Ratio of pulmonary CD4+ T cells expressing IL-4 and IFN-γ. Data are representative of 2 independent experiments with 3 samples/group. *P < 0.05.

Article Snippet: BMDCs were then harvested and infected with third-generation lentiviral vectors (pLV-IL-4Rα-eGFP or control pLV-eGFP) (Cyagen Biosciences, Santa Clara, CA, USA) at a multiplicity of infection 20 in Opti-MEM media (Thermo Fisher Scientific) and 5 µg/ml of polybrene (Cyagen Biosciences) for 3 h. These cells were cultured in DC medium for 4 d. Transduced cells were then infected with RSV at a multiplicity of infection 50 overnight, and CD11b + mDCs that were positive for eGFP were sorted with a SH800 cell sorter (Sony Biotechnology, San Jose, CA, USA).

Techniques: Infection, Isolation, Cell Culture, Purification, Flow Cytometry, Expressing

Journal: Journal of Leukocyte Biology

Article Title: IL-4Rα on dendritic cells in neonates and Th2 immunopathology in respiratory syncytial virus infection

doi: 10.1189/jlb.4A1216-536R

Figure Lengend Snippet: Neonatal or adult wild-type mice and neonatal IL-4Rα−/−DC mice were infected with RSV. Maturation markers (CD80 and CD86) and MHC-II were measured on lung CD11b+ mDCs via flow cytometry. (A) MFI of CD80 on CD11b+ mDCs. (B) MFI of CD86 on CD11b+ mDCs. (C) MFI of MHCII on CD11b+ mDCs. Data are representative of 3 independent experiments with 4–5 mice/group. *P < 0.05.

Article Snippet: BMDCs were then harvested and infected with third-generation lentiviral vectors (pLV-IL-4Rα-eGFP or control pLV-eGFP) (Cyagen Biosciences, Santa Clara, CA, USA) at a multiplicity of infection 20 in Opti-MEM media (Thermo Fisher Scientific) and 5 µg/ml of polybrene (Cyagen Biosciences) for 3 h. These cells were cultured in DC medium for 4 d. Transduced cells were then infected with RSV at a multiplicity of infection 50 overnight, and CD11b + mDCs that were positive for eGFP were sorted with a SH800 cell sorter (Sony Biotechnology, San Jose, CA, USA).

Techniques: Infection, Flow Cytometry

Journal: Journal of Leukocyte Biology

Article Title: IL-4Rα on dendritic cells in neonates and Th2 immunopathology in respiratory syncytial virus infection

doi: 10.1189/jlb.4A1216-536R

Figure Lengend Snippet: BMDCs were isolated from adult mice, transduced with pLV-IL-4Rα-eGFP or pLV-eGFP control, and infected with RSV. CD11b+ mDCs that were positive for eGFP were sorted and adoptively transferred into the lungs of adult mice. Those mice were then infected with RSV, and 6 d later, pulmonary Th subsets were quantified. (A) Percentage of CD4+ T cells expressing IFN-γ or IL-4 or both IFN-γ and IL-4. (B) Number of CD4+ T cells expressing IFN- γ, IL-4, or both IFN-γ and IL-4. Data are representative of 2 independent experiments with 3–4 mice/group. *P < 0.05.

Article Snippet: BMDCs were then harvested and infected with third-generation lentiviral vectors (pLV-IL-4Rα-eGFP or control pLV-eGFP) (Cyagen Biosciences, Santa Clara, CA, USA) at a multiplicity of infection 20 in Opti-MEM media (Thermo Fisher Scientific) and 5 µg/ml of polybrene (Cyagen Biosciences) for 3 h. These cells were cultured in DC medium for 4 d. Transduced cells were then infected with RSV at a multiplicity of infection 50 overnight, and CD11b + mDCs that were positive for eGFP were sorted with a SH800 cell sorter (Sony Biotechnology, San Jose, CA, USA).

Techniques: Isolation, Transduction, Infection, Expressing