Journal: Cellular Microbiology

Article Title: Influenza A virus-mediated priming enhances cytokine secretion by human dendritic cells infected with Streptococcus pneumoniae

doi: 10.1111/cmi.12122

Figure Lengend Snippet: Preceding influenza infection leads to elevated levels of secreted IL-12p70 and IL-6 after co-infection with SP. MDDCs were infected with IAV for 4 h before infection with SP. The cells were further incubated for 18 h before the concentration of IL-12p70 (A), IL-6 (B), TNFα (C), IL-1β (D), IFN-β (E) and IL-8 (F) in the supernatants was measured by ELISA. Each dot in a graph represents the cytokine concentration from cells of one donor. The graphs show mean ± SEM of seven (A, C), three (B), five (D, E) or four (F) independent experiments with different donors. Statistical analysis was performed using paired Student’s t -test. (* P < 0.05, ** P < 0.01).

Article Snippet: Cells were pelleted and medium was replaced with R10 medium containing antibodies against human IFN-α (No. 31101-1, PBL Interferon source), IFN-β (No. 31410-1, PBL Interferon source) and IFNAR (MMHAR-2, PBL Interferon source) at concentrations of 5000 U ml −1 , 2000 U ml −1 and 20 μg ml −1 respectively.

Techniques: Infection, Incubation, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: Cellular Microbiology

Article Title: Influenza A virus-mediated priming enhances cytokine secretion by human dendritic cells infected with Streptococcus pneumoniae

doi: 10.1111/cmi.12122

Figure Lengend Snippet: Type I IFNs are responsible for the priming of DCs to produce IL-12p70. MDDCs were incubated with different concentrations of recombinant human IFN-α 4 h prior to infection with SP. The cells were further incubated for 18 h before the concentration of IL-12p70 in the supernatants was measured by ELISA. MDDCs were sequentially infected with IAV and SP in the presence of neutralizing antibodies against IFN-α, IFN-β and IFNAR. After 22 h incubation the cells were lysed and total RNA was isolated and assayed for the presence of IL-12p35 and IL-12p40 mRNA. Ct values were normalized against γ-actin and the relative induction of the genes was calculated using the ΔΔCt method. Values represent mean ± SEM three independent experiments with different donors (A) or one representative donor of 3 (B, C). Statistical analysis was performed using paired Student’s t -test. (* P < 0.05).

Article Snippet: Cells were pelleted and medium was replaced with R10 medium containing antibodies against human IFN-α (No. 31101-1, PBL Interferon source), IFN-β (No. 31410-1, PBL Interferon source) and IFNAR (MMHAR-2, PBL Interferon source) at concentrations of 5000 U ml −1 , 2000 U ml −1 and 20 μg ml −1 respectively.

Techniques: Incubation, Recombinant, Infection, Concentration Assay, Enzyme-linked Immunosorbent Assay, Isolation

Journal: The EMBO Journal

Article Title: Function of HNRNPC in breast cancer cells by controlling the dsRNA‐induced interferon response

doi: 10.15252/embj.201899017

Figure Lengend Snippet: Concentrations of IFNβ, measured by ELISA, in the culturing media of MCF7 and T47D cells 48 hours after siRNA transfections. siNC: non‐targeting siRNA as a negative control, siHN‐1: siRNA sequence 1 for HNRNPC, siHN‐2: siRNA sequence 2 for HNRNPC, siLMNA: siRNA for LMNA as another negative control. Each sample has three replicates. Data represent mean ± SD. The normal MCF7 (left) and T47D (right) cells were cultured in the media collected from the corresponding cells 48 h after siRNA‐mediated gene knock‐down. Expressions of the ISGs in these normal cells were measured with qPCR. Each sample has three replicates. Data represent mean ± SD. Different doses of the IFNβ antibody were added to the media of the siRNA‐transfected MCF7 (left) or T47D (right) cells right before the media were transferred to the wild‐type cells. The wild‐type MCF7 or T47D cells were then cultured in these media for 48 hours, and the expressions of ISGs were measured by qPCR. Each sample has three replicates. Data represent mean ± SD. Growth curves of the MCF7 (left) and T47D (right) cells cultured in the media collected from the siRNA‐transfected cells. Each sample has three replicates. Data represent mean ± SD.

Article Snippet: Ruxolitinib (2 mM) (Invivogen), 0–20 U/ml of IFNAR2 antibody (PBL Interferon Source 21385‐1), 0–500 U/ml of IFNβ antibody (PBL Interferon Source 31401‐1) were added when changing the medium 6 h after siRNA transfection.

Techniques: Enzyme-linked Immunosorbent Assay, Transfection, Negative Control, Sequencing, Cell Culture, Knockdown

Journal: The EMBO Journal

Article Title: Function of HNRNPC in breast cancer cells by controlling the dsRNA‐induced interferon response

doi: 10.15252/embj.201899017

Figure Lengend Snippet: A–C In the siRNA‐transfected MCF7 (left) or T47D cells (right), the interferon signaling pathway was blocked at different stages by means of IFNβ neutralization (A), IFNAR2 neutralization (B), or JAK‐STAT inhibition (C). The expressions of the ISGs were measured by qPCR. Each sample has three replicates. Data represent mean ± SD. D, E Growth curves of the MCF7 (D) or T47D cells (E) upon HNRNPC knock‐down but with the interferon response blocked with the IFNβ antibody. Each sample has three replicates. Data represent mean ± SD. F Growth curves of the MCF7 (left) or T47D (right) cells upon HNRNPC knock‐down and JAK‐STAT inhibition with ruxolitinib (5 μM). siNC: non‐targeting siRNA as a negative control, siLMNA: siRNA for LMNA as another negative control, siHN‐1: siRNA sequence 1 for HNRNPC, siHN‐2: siRNA sequence 2 for HNRNPC. Each sample has three replicates. Data represent mean ± SD.

Article Snippet: Ruxolitinib (2 mM) (Invivogen), 0–20 U/ml of IFNAR2 antibody (PBL Interferon Source 21385‐1), 0–500 U/ml of IFNβ antibody (PBL Interferon Source 31401‐1) were added when changing the medium 6 h after siRNA transfection.

Techniques: Transfection, Neutralization, Inhibition, Knockdown, Negative Control, Sequencing