Journal: Frontiers in Plant Science

Article Title: Melatonin Alleviates Low-Temperature Stress via ABI5-Mediated Signals During Seed Germination in Rice ( Oryza sativa L.)

doi: 10.3389/fpls.2021.727596

Figure Lengend Snippet: Effects of exogenous MT treatment on seed germination-associated physiological attributes under LT stress. Physical efficiency, soluble sugar contents, soluble protein contents, phospholipid contents, respiratory rate, and membrane permeability under LT stress (constant 20°C for 7 days) without (LT20) or with (LT20 + MT) MT treatment. Different letters denote significant variations between the treatments, and the average values were measured by Tukey’s HSD test at p < 0.05. Data were represented as mean ± SE of six biological replicates.

Article Snippet: Phospholipid contents were determined by using the Phospholipid Assay Kit (ab234050, Colorimetric, Abcam, Shanghai, China) according to the instructions of the manufacturer.

Techniques: Membrane, Permeability

Journal: The Journal of International Medical Research

Article Title: The therapeutic potential of irisin in alleviating acute lung injury via inflammation and ferroptosis modulation

doi: 10.1177/03000605251340338

Figure Lengend Snippet: Western blotting analysis on the effects of irisin on the expression level of ferroptosis-associated proteins. (a) Representative western blotting images for the expression of ACSL4, COX-2, GPX4, p-AMPK, and t-AMPK in lung tissues. GAPDH was selected as the loading control protein. (b) Quantification analysis of the related bands of ACSL4, COX-2, GPX4, p-AMPK, and t-AMPK in lung tissues. Lung tissues were harvested on day 3 post-CLP. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post-hoc test. n = 6 per group. Data are presented as means ± SEM. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.

Article Snippet: Subsequently, the membranes were incubated with rabbit anti-AMPK (MCE, Cat# HY- P80541 ), anti-p-AMPK (MCE, HY- P80452 ), GPX4 (MCE, HY- P80450 ), ACSL4 (Santa Cruz Biotechnology, Dallas, Texas, USA; Cat# sc-365230), COX-2 (Abcam, Cambridge, United Kingdom; Cat# ab283574), and GAPDH (MCE, HY-P80137) antibodies at 4°C overnight.

Techniques: Western Blot, Expressing, Control

Journal: Indian Journal of Ophthalmology

Article Title: Ranitidine protects Müller cells against ferroptosis in diabetic retinopathy by regulating the AKT1 / GSK3β pathway

doi: 10.4103/IJO.IJO_1522_25

Figure Lengend Snippet: Ra influences the AKT1 / GSK3β signaling pathway under high-glucose conditions. a and b. Western blot analysis assessed AKT1 and p- AKT1 protein levels in HG-treated rMC-1 cells. a and c. Western blot analysis assessed GSK3β and p- GSK3β protein levels in HG-treated rMC-1 cells. d-f. Effects of Ra on GPX4 and xCT expression in HG-treated rMC-1 cells assessed by Western blotting ( n = 3, *P < 0.05, ns: no significance, compared to HG group). Ra: Ranitidine, HG: High-glucose, rMC-1: Mouse retinal Müller cells, GPX4: Glutathione peroxidase 4, xCT: Cystine/glutamate transporter

Article Snippet: After being closed with 5% bovine serum albumin blocking solution for 2 h at room temperature, the membranes were incubated with primary antibodies against GPX4 (Med Chem Express, USA) (1:1000), xCT (Med Chem Express, USA) (1:1000), GSK3β (Med Chem Express, USA) (1:1000), p- GSK3β (Med Chem Express, USA) (1:1000), AKT1 (Zenbio, USA) (1:1000), or p- AKT1 (Med Chem Express, USA) (1:1000) proteins at 4°C overnight.

Techniques: Western Blot, Expressing

Journal: Indian Journal of Ophthalmology

Article Title: Ranitidine protects Müller cells against ferroptosis in diabetic retinopathy by regulating the AKT1 / GSK3β pathway

doi: 10.4103/IJO.IJO_1522_25

Figure Lengend Snippet: Ra inhibits HG-induced ferroptosis in rMC-1 cells, depending on GSK3β . a-c. The effects of Ra on the expression levels of AKT1 , P- AKT1 , GSK3β , and p- GSK3β in HG-cultured rMC-1 cells were assessed by Western blotting. d-f. The effects of Ra on the expression levels of GPX4 and xCT in HG-cultured rMC-1 cells were assessed by Western blotting ( n = 3, **P < 0.01, ***P < 0.001, ns: no significance, compared to HG group; # P < 0.05, ### P < 0.001, ns: no significance, compared to HG + Ra group). Ra: Ranitidine, HG: High-glucose, rMC-1: Mouse retinal Müller cells, GPX4: Glutathione peroxidase 4, xCT: Cystine/glutamate transporter

Article Snippet: After being closed with 5% bovine serum albumin blocking solution for 2 h at room temperature, the membranes were incubated with primary antibodies against GPX4 (Med Chem Express, USA) (1:1000), xCT (Med Chem Express, USA) (1:1000), GSK3β (Med Chem Express, USA) (1:1000), p- GSK3β (Med Chem Express, USA) (1:1000), AKT1 (Zenbio, USA) (1:1000), or p- AKT1 (Med Chem Express, USA) (1:1000) proteins at 4°C overnight.

Techniques: Expressing, Cell Culture, Western Blot

Journal: Frontiers in Pharmacology

Article Title: Empagliflozin attenuates dexamethasone-induced non-alcoholic steatohepatitis by regulation of ferroptosis, inflammation and autophagy

doi: 10.3389/fphar.2026.1741926

Figure Lengend Snippet: Impact of EMPA on DEXA-induced changes in hepatic oxidant/antioxidant biomarkers. (A) MDA, (B) 4HNE, (C) GSH, (D) GPX4, (E) Nrf2. Data were expressed as means ± SEM (n = 4). Mean values were compared using one-way ANOVA followed by post hoc Tukey’s multiple comparison test. a significance vs. CONTROL group, b significance vs. DEXA group, c significance vs. EMPA10/DEXA group. DEXA: Dexamethasone, EMPA: Empagliflozin, 4HNE: 4-Hydroxy-2-nonenal, MDA: Malondialdehyde, GSH: Glutathione, GPX4: Glutathione Peroxidase-4, Nrf2: Nuclear Factor Erythroid 2–Related Factor 2.

Article Snippet: Hepatic levels of reduced glutathione (GSH) and glutathione peroxidase-4 (GPX4) were estimated using ELISA kits from CUSABIO (Houston, United States, cat no. CSB-E12144r, CSB-EL009869RA, respectively) according to the manufacturers’ instructions, while, hepatic levels of nuclear factor erythroid 2-related factor 2 (Nrf2) was estimated using BT LAB ELISA kit (Shanghai, China, Cat no. E1083Ra).

Techniques: Comparison, Control

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: Wild-type (WT) and Plscr1 -/- mice were exposed to sublethal (300 pfu, B, C, and F–H ) or lethal (900 pfu, D and E ) influenza A virus (IAV) (WSN) infection. ( A ) Scheme of experiment. ( B ) Whole lungs of WT mice were analyzed for Plscr1 RNA by qRT-PCR. ( C, D ) Mean relative weight of mice post-sublethal or lethal infection. ( E ) Survival rate of mice post-lethal IAV infection. ( F ) Viral RNA load in the lungs was assessed by quantifying M gene by qRT-PCR. ( G ) Infectious viral titer in the lungs was assessed by plaque assays. ( H ) Representative staining for H1N1 in lungs. The scale bars represent 1 mm. Quantification was performed using ImageJ. Data are expressed as mean ± SEM of n=30 mice/group for weight loss post-sublethal infection and n=8 mice/group for weight loss and survival rate post-lethal infection. For the rest analysis, n=5–10 mice/group. All data were pooled from three independent experiments and described biological replicates. Log-rank (Mantel-Cox) test was used to compare survival rates. Ordinary two-way ANOVA tests were used to compare weight losses. * p <0.05, ** p <0.01, *** p <0.001, **** P <0.0001. dpi, days post-infection. CTCF, Corrected Total Cell Fluorescence.

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Virus, Infection, Quantitative RT-PCR, Staining, Fluorescence

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: Wild-type (WT) and Plscr1 -/- mice were exposed to sublethal (300 pfu) influenza A virus (IAV) (WSN) infection. ( A ) Total Bronchoalveolar lavage (BAL) leukocyte numbers. ( B ) Differential cell counts in BAL. ( C ) Representative lung sections stained with Hematoxylin and Eosin (H&E). Scale bars represent 3 mm (main) and 200 μm (inlays). ( D ) Whole lungs were analyzed for Ifna , Ifnb , Ifng, and Ifnl RNA by qRT-PCR. ( E ) Tnf-α and Ifn-λ concentrations in BAL by ELISA. Data are expressed as mean ± SEM of n=3–14 mice/group. All data were pooled from three independent experiments and described biological replicates. * p <0.05, ** p <0.01. dpi, days post-infection.

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Virus, Infection, Staining, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: ( A–E ) Wild-type (WT) and Plscr1 -/- mice were exposed to sublethal (300 pfu) IAV (WSN) infection. ( A ) Heatmap of interferons and their receptors in whole lungs by RNA-seq. ( B ) Whole lungs were analyzed for Ifnlr1 by qRT-PCR. ( C ) Heatmap of differential expressions of all interferon-stimulated genes (ISGs) in whole lungs by RNA-seq. Gene expressions were compared between groups within each row and color-labeled from row minimum (blue) to row maximum (red). ( D ) Localization of Ifn-λr1+ cells in the lungs of IAV-infected WT mice at 7 dpi. Sections stained for Ifn-λr1 (red), Foxj1, uteroglobin, or Sftpc (green), and DAPI (blue) are shown. Scale bars represent 50 μm (main) and 20 μm (inlays). ( E ) Representative staining for Ifn-λr1 in airways or alveoli of IAV-infected WT and Plscr1 -/- mice at 7dpi. Scale bars represent 50 μm. Quantifications were performed using ImageJ. ( F, G ) Calu-3 cells were analyzed for PLSCR1 ( F ) and IFNLR1 ( G ) RNA by qRT-PCR after recombinant IFN-λ and/or α-IFN-λR1 antibody treatment. Data are presented as fold change compared to non-treated group. ( H, I ) Chromatin-Immunoprecipitation of PLSCR1 and IFNLR1 promoter in Calu-3 cells followed by standard PCR ( H ) and real-time quantitative PCR ( I ). Data are expressed as mean ± SEM of n=4–12 mice or wells/group. For transcriptomic analysis, 9 mice from each PBS-treated group and 4 mice from each IAV-infected group were pooled together. All data were pooled from three independent experiments and described biological replicates. * p <0.05, ** p <0.01, *** p <0.001. dpi, days post-infection. CTCF, Corrected Total Cell Fluorescence. Scale bars represent 50 μm. Figure 3—source data 1. PDF file containing original gel for , indicating the relevant treatments. Figure 3—source data 2. Original gel corresponding to .

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Infection, RNA Sequencing, Quantitative RT-PCR, Labeling, Staining, Recombinant, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Fluorescence

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: Wild-type (WT) and Plscr1-/- mice were intranasally given 2.5 μg/g of body weight of poly(I:C) (HMW) constitutively for 6 days and sacrificed on day 7. ( A ) Scheme of experiment. ( B ) Total Bronchoalveolar lavage (BAL) leukocyte numbers. ( C ) Differential cell counts in BAL. ( D, F, G ) Whole lungs were analyzed for Ifna, Ifnb, Ifng, Ifnl ( D ); Plscr1 ( F ); and Ifnlr1 ( G ) RNA by qRT-PCR. ( E ) Representative lung sections stained with Hematoxylin and Eosin (H&E). Scale bars represent 3 mm (main) and 200 μm (inlays). Data are expressed as mean ± SEM of n=5–12 mice/group. All data were pooled from three independent experiments and described biological replicates. ns, not significant, *p<0.05, ***p<0.001.

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Quantitative RT-PCR, Staining

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: ( A ) Co-Immunoprecipitation of Plscr1 and Ifn-λr1 in whole mouse lungs followed by western blot. ( B ) Proximity ligation assay of Ifn-λr1 and Plscr1 in the lungs of wild-type (WT) mice infected or uninfected with IAV. Scale bars represent 50 μm. Quantifications were performed using ImageJ. ( C ) Colocalization of IFN-λR1 (green) and PLSCR1 (red) on Calu-3 cell membranes infected or uninfected with IAV in a non-permeabilized staining. Scale bars represent 10 μm. Data are expressed as mean ± SEM of n=6–7 lungs/group. All data were pooled from three independent experiments and described biological replicates. * p <0.05, ** p <0.01. PLA, Proximity Ligation Assay. CTCF, Corrected Total Cell Fluorescence. Figure 4—source data 1. PDF file containing original membrane for , indicating the relevant bands and treatments. The exposure time was adjusted to visualize Plscr1 (top) or Ifn-λr1 (bottom). Figure 4—source data 2. Original membrane corresponding to .

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Immunoprecipitation, Western Blot, Proximity Ligation Assay, Infection, Staining, Fluorescence, Membrane

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: PLSCR1 plasmids on PLV-EF1a-IRES-Hygro backbone were packaged into GFP-expressing lentivirus. PLSCR1 -/- A549 cells were transduced using lentivirus. After a 10 day hygromycin selection, cells were analyzed using flow cytometry. ( A ) Gating strategy for live, GFP+ and PLSCR1+A549 cells. ( B ) Surface, cytoplasm, and nuclear expression of PLSCR1. Data are presented as fold change compared to WT -PLSCR1 -/- A549 cells. ( C ) Lentiviral transduction efficiency.

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Expressing, Selection, Flow Cytometry, Transduction

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: PLSCR1 -/- A549 cells were transduced with mutated PLSCR1 plasmids using lentivirus and infected with influenza A virus (IA)V (PR8) for 24 hr at 1 MOI ( A–E ) or 10 MOI ( F ). ( A ) IFNLR1 RNA by qRT-PCR. ( B ) IFN-λR1 proteins by western blotting. ( C ) Proximity ligation assay of IFN-λR1 and PLSCR1. Scale bars represent 20 μm. Quantifications were performed using ImageJ. ( D ) Viral RNA load was assessed by quantifying M gene by qRT-PCR. ( E ) Infectious viral titer was assessed by plaque assays. ( F ) Cells were stained with crystal violet. Cell viability was quantified using ImageJ. Data are expressed as mean ± SEM of n=4–13 wells/group. All data were pooled from three independent experiments and described biological replicates. ns, not significant, * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001, ***** p <0.00001. CTCF, Corrected Total Cell Fluorescence. Figure 5—source data 1. PDF file containing original membrane for , indicating the relevant bands and treatments. The membrane was cut just under 50 kDa marker after transfer. The top part was incubated with α-Ifn-λr1 antibody and the bottom part was incubated with α-β-actin antibody. They were then incubated with corresponding secondary antibodies separately. The exposure time was adjusted to visualize Ifn-λr1 (top) or β-actin (middle). A colorimetric photo was taken to visualize the molecular weight markers (bottom). Lanes 2–5 were from an unrelated experiment. Figure 5—source data 2. Original membrane corresponding to .

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Transduction, Infection, Virus, Quantitative RT-PCR, Western Blot, Proximity Ligation Assay, Staining, Fluorescence, Membrane, Marker, Incubation, Molecular Weight

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: WIld-type (WT), Plscr1 -/- , Ifnlr1 -/- and Plscr1 -/- ;Ifnlr1 -/- mice were exposed to sublethal (300 pfu) influenza A virus (IAV) (WSN) infection and sacrificed at 3 dpi. ( A ) Representative immunofluorescent staining for DAPI and Ifn-λr1 in lungs. Scale bars represent 100 μm. ( B ) Mean relative weight of mice. ( C ) Total Bronchoalveolar lavage (BAL) leukocyte numbers. ( D ) Neutrophil percentages in BAL. ( E ) Infectious viral titer in the lungs was assessed by plaque assays. Data are expressed as mean ± SEM of n=3–12 mice/group. All data were pooled from three independent experiments and described biological replicates. ns, not significant, * p <0.05, ** p <0.01, *** p <0.001.

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Virus, Infection, Staining

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: Wild-type (WT) mice were exposed to 2500 EID50 IAV (PR8) infection. Lungs were used for single-cell RNA sequencing analysis at 0, 1, 3, 6, and 21 dpi. ( A ) Two-dimensional UMAP representation of individual cells obtained from different timepoints. ( B ) Violin plot of aggregated Plscr1 expressions in all epithelial cell clusters. Red dots represent mean expression levels. ( C ) Violin plot of time-dependent Plscr1 expressions in both ciliated epithelial cell clusters. ( D ) Gene Ontology (GO) analysis for upregulated pathways in Ciliated Epithelial Cells-1. ( E ) Heatmap of the most differentially expressed genes of Ciliated Epithelial Cells-1 at different timepoints. ( F ) Violin plot of aggregated Plscr1 expressions in all immune cell clusters. Red dots represent mean expression levels. ( G ) Violin plot of time-dependent Plscr1 expressions in alveolar macrophage cluster. ( H ) Violin plot of time-dependent Plscr1 expressions in neutrophil cluster.

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Infection, RNA Sequencing, Expressing

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: Plscr1 floxStop and Plscr1 floxStop ;Foxj1-Cre + mice were exposed to sublethal (300 pfu) influenza A virus (IAV) (WSN) infection and sacrificed at 3 dpi. ( A ) Schematic representation of the experimental design of ciliated epithelial cell conditional Plscr1 KI mice. ( B ) Validation of Plscr1 overexpression in lungs of Plscr1 floxStop ;Foxj1-Cre + mice by qRT-PCR. ( C ) Representative immunofluorescent staining for Plscr1, Ifn-λr1, and Foxj1 in lungs. Scale bars represent 50 μm (main) and 10 μm (inlays).( D ) Mean relative weight of mice. ( E ) Viral RNA load in the lungs was assessed by quantifying M gene by qRT-PCR. ( F ) Infectious viral titer in the lungs was assessed by plaque assays. ( G ) Total Bronchoalveolar lavage (BAL) leukocyte numbers. ( H ) Neutrophil percentages in BAL. ( I ) Whole lungs were analyzed for Ifnlr1 RNA by qRT-PCR and Ifn-λr1 protein by western blot. ( J ) Whole lungs were analyzed for Ifna , Ifnb , Ifng , and Ifnl RNA by qRT-PCR. ( K ) Model depicting proposed mechanism of PLSCR1-regulated IFN-λ signaling. Data are expressed as mean ± SEM of n=3–10 mice/group. All data were pooled from three independent experiments and described biological replicates. ns, not significant, * p <0.05, ** p <0.01, *** p <0.001. dpi, days post-infection. Figure 8—source data 1. PDF file containing original membrane for , indicating the relevant bands and treatments. The membrane was cut just under 50 kDa marker after transfer. The top part was incubated with α-Ifn-λr1 antibody and the bottom part was incubated with α-β-actin antibody. They were then incubated with corresponding secondary antibodies separately. The exposure time was adjusted to visualize Ifn-λr1 (top) or β-actin (middle). A colorimetric photo was taken to visualize the molecular weight markers (bottom). Lanes 2 and 3 were from an unrelated experiment. Figure 8—source data 2. Original membrane corresponding to .

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Virus, Infection, Biomarker Discovery, Over Expression, Quantitative RT-PCR, Staining, Western Blot, Membrane, Marker, Incubation, Molecular Weight

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: Plscr1 floxStop and Plscr1 floxStop ;Lyz2-Cre + mice were exposed to sublethal (300 pfu) influenza A virus (IAV) (WSN) infection. ( A ) Validation of Plscr1 overexpression in lungs of Plscr1 floxStop ;Lyz2-Cre + mice by qRT-PCR. ( B ) Mean relative weight of mice. ( C ) Total Bronchoalveolar lavage (BAL) leukocyte numbers. ( D ) Differential cell counts in BAL. ( E ) Viral RNA load in the lungs was assessed by quantifying M gene by qRT-PCR. ( F ) Representative lung sections stained with Hematoxylin and Eosin (H&E). Scale bars represent 3 mm (main) and 200 μm (inlays). ( G–I ) Whole lungs were analyzed for Ifna , Ifnb , Ifng , Ifnl ( G ); Ifnlr1 ( H ); and Plscr1 ( I ) RNA by qRT-PCR. Data are expressed as mean ± SEM of n=15–16 mice/group for weight loss. For the rest analysis, n=3–7 mice/group. All data were pooled from three independent experiments and described biological replicates. ns, not significant, ** p <0.01. dpi, days post-infection.

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Virus, Infection, Biomarker Discovery, Over Expression, Quantitative RT-PCR, Staining

Journal: eLife

Article Title: Phospholipid scramblase 1 (PLSCR1) regulates interferon-lambda receptor 1 (IFN-λR1) and IFN-λ signaling in influenza A virus (IAV) infection

doi: 10.7554/eLife.104359

Figure Lengend Snippet: Influenza infection is usually detected by intracellular pattern recognition receptors (PRRs) such as TLR 3 and 7, RIG-I, and MDA5. These PRRs activate the expression of IFNL in early infection stage through IRF-3 and NK-κB-controlled transcriptions. IFN-λ secreted by the infected cells interacts with IL-10R2 and IFN-λR1 on neighboring cell surfaces, which results in activation of expression of various IFN-stimulated genes, including PLSCR1 . In ciliated airway epithelial cells, PLSCR1 can further enhance the transcription of IFNLR1 by directly binding to its promoter region as a transcriptional factor, or interact with IFN-λR1 on the cell membrane.

Article Snippet: They were stained overnight at 4 °C with mouse anti-human PLSCR1 monoclonal antibody (1:50 dilution, R&D Systems, cat #MAB8137, clone #875327) and rabbit anti-human IFN-λR1 polyclonal antibody (1:100 dilution, Invitrogen, cat #PA5-98608, lot #ZB4228832A).

Techniques: Infection, Expressing, Activation Assay, Binding Assay, Membrane