Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: Domain organizations of LDLR and PCSK9 and structures of LDLR β-propeller domain and of PCSK9·LDLR complex. A, domain organizations of LDLR (top) and PCSK9 (bottom). LDLR is composed of the ligand binding domain (LBD), containing seven repeats (L1–L7) of ∼40 residues, the EGF precursor homology domain (EGFPH), consisting of EGFA, EGFB, six-bladed (1,–6) β-propeller, and EGFC domains, the O-linked sugar regions, and the transmembrane domain (TMD). Mature PCSK9 is formed after auto-cleavage between the prodomain (Pro) and the catalytic domain (Catalytic). The hinge domain (H) and the C-terminal histidine-rich domain (CHRD) are indicated. B, LDLR β-propeller domain with its six blades and their corresponding residues. The positions of Arg410 and G592 in the β-propeller domain of the LDLR are displayed. C, overall structure of the PCSK9·LDLR complex (PDB code 3M0C) shows two interfaces of interaction between the two proteins as follows: a major interface between catalytic domain of PCSK9 (green) and the EGFA domain of LDLR (turquoise) and a minor interface (boxed) between the prodomain of PCSK9 (red) and the LDLR β-propeller domain (blue). Box, details of the prodomain/β-propeller interface; yellow dotted line shows the van der Waals interaction between Leu108 (PCSK9) and Leu647 (LDLR-WT), whereas the blue dotted line depicts the “putative” ionic interaction between the GOF mutation L108R (PCSK9) and Glu626 (LDLR-WT).

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Ligand Binding Assay, Mutagenesis

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: Functional classification of LDLR loss of function mutations

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Functional Assay, Mutagenesis, Blocking Assay, Binding Assay

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: Evolution of LDLc concentration of patient III-3 following different therapies and family pedigree. A, LDLc concentrations following several lipid-lowering treatments. B, family pedigree. The prepositus is marked with an arrow. Round symbols, females. Square symbols, males. Black outlined symbols, family members for which data were available. Gray outlined symbols, family members for which no data were available. Symbols crossed through: deceased individuals. LDLR-R410S allele, dark gray shading. LDLR-G592E allele, light gray shading.

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Concentration Assay

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: Total and cell surface expression of wild-type LDLR and its two mutants, R410S and G592E. HEK293 cells were transfected with V5-tagged LDLR-WT (WT), LDLR-R410S (RS), LDLR-G592E (GE), or both mutants (RS/GE). A, WB analysis of LDLR expression. B, LDLR sensitivity to endoglycosidase H (Endo H). C, LDLR levels after 18 h in absence (−) or presence (+) of 2.5 μmol/liter MG132, a proteasome inhibitor. D, FACS analyses of cell surface LDLR expression. E, immunofluorescence microscopy of total (PERM) and cell surface (NON PERM) LDLR in HepG2 cells overexpressing WT, RS, GE, or RS/GE. Quantification of cell surface LDLR is shown, using 12 transfected cells (EGFP-positive)/condition/experiment. Non-transfected cells expressed ∼100-fold less LDLR than transfected ones. Scale bar, 15 μm. Data are representative of at least two independent experiments. Quantifications are averages ± S.D. *, p < 0.05; **, p < 0.01; ***, p < 0.001 (t test).

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Expressing, Transfection, Immunofluorescence, Microscopy

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: Effects of PCSK9 on LDLR degradation. A and B, HEK293 cells were co-transfected with V5-tagged LDLR-WT (WT) or LDLR-R410S (RS) and PCSK9-WT (PC9-WT), PCSK9-DY (PC9-DY) or empty vector (v), as control, and analyzed by WB using anti-human LDLR (A), or by FACS for cell surface LDLR expression (B). C and D, HEK293 cells overexpressing V5-tagged WT or RS were incubated for 7 h with conditioned media from HEK293 cells (input): no PCSK9 control media (Cnt) or PCSK9 media (∼1.8 μg/ml), PC9-WT or GOF PC9-DY, and analyzed by WB with anti-human LDLR (C) or by FACS for cell surface LDLR expression (D). E, mouse primary hepatocytes lacking both PCSK9 and LDLR were transfected with V5-tagged WT or RS and incubated for 18 h with conditioned media from HEK293 as described in C. Cell lysates were immunoblotted with anti-V5-HRP for LDLR. Data are representative of at least three independent experiments. Quantifications are averages ± S.D. *, p < 0.05; **, p < 0.01; ***, p < 0.001 (t test).

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Transfection, Plasmid Preparation, Control, Expressing, Incubation

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: PCSK9 displays similar binding to the cell surface LDLR-WT and LDLR-RS. A and B, HEK293 cells transfected with LDLR-WT (WT) or LDLR-R410S (RS) were incubated for 4 h at 4 °C with conditioned media from HEK293 cells: no PCSK9 control media (Cnt) or media containing PCSK9-WT. PCSK9 was non-tagged (PC9-WT) ∼1.3 μg/ml (A), or mCherry-tagged (PC9-mCherry), ∼1 μg/ml (B). PCSK9 binding to cell surface receptor was quantified by FACS (A) or fluorescence spectroscopy (B). Bars represent averages ± S.D. from three independent experiments. C, HEK293 cells overexpressing LDLR-WT (WT) or LDLR-RS (RS) were incubated with purified recombinant human PCSK9 for 4 h at 4 °C. The curve through the data is the fit to a rectangular hyperbola and is representative of two independent experiments. The average PCSK9 binding constants determined from the two experiments are 1.8 ± 0.8 μm (WT) and 1.3 ± 0.1 μm (RS).

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Binding Assay, Transfection, Incubation, Control, Cell Surface Receptor Assay, Fluorescence, Spectroscopy, Purification, Recombinant

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: GOF PCSK9-L108R in the prodomain rescues PCSK9-mediated degradation of LDLR-R410S via the extracellular pathway. HEK293 cells overexpressing LDLR-WT (WT) or LDLR-R410S (RS) were incubated for 18 h with conditioned media from HEK293 cells: no PCSK9 control-media (Cnt) or PCSK9-media (∼1.1 μg/ml), PCSK9-WT (PC9-WT), or GOF PCSK9-L108R (PC9-LR). A, total LDLR quantification by ELISA. B, FACS analyses of cell surface LDLR. Data are representative of four independent experiments. Bars are averages ± S.D. *, p < 0.05; **, p < 0.01 (t test).

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Incubation, Control, Enzyme-linked Immunosorbent Assay

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: Effect of loss of Arg410 on LDLR expression, functionality, and sensitivity to extracellular PCSK9-mediated degradation. A–D, HEK293 cells transfected with LDLR-WT (WT) or LDLR-R410 mutants (RS, RK, RE, RA) were analyzed for total LDLR expression by ELISA (A), cell surface LDLR expression by FACS (B), sensitivity of LDLR to endoglycosidase H, by WB (C), and cell surface LDLR expression, by FACS, following an 18-h incubation with conditioned media from HEK293 cells: no PCSK9 control media (Cnt), or PCSK9 media (∼1.1 μg/ml), PCSK9-WT (PC9-WT), or GOF PCSK9-DY (PC9-DY) (D). E and F, immunofluorescence microscopy in HepG2 cells transfected with WT or RS, RK, RE, RA mutants. E, total LDLR, under permeabilized conditions (PERM), and cell surface LDLR, under non-permeabilized conditions (NON PERM). F, DiI-LDL internalization after 4-h incubation with 5 μg/ml DiI-LDL at 37 °C. Quantifications in E and F were derived from analyses of 12 transfected cells (EGFP-positive)/condition/experiment. Scale bar, 15 μm. Bars in A, B, and F are averages ± S.D. of five independent experiments, and bars in C and D are averages ± S.D. of two independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001 (t test).

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Expressing, Transfection, Enzyme-linked Immunosorbent Assay, Incubation, Control, Immunofluorescence, Microscopy, Derivative Assay

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: Functionality of LDLR-WT and LDLR-R410S. A–C, HEK293 cells transfected with LDLR-WT (WT) or LDLR-R410S (RS) were evaluated. A, 125I-LDL binding to LDLR after a 1-h incubation at 4 °C with increasing concentrations of 125I-LDL (200 cpm/pg). B, time-dependent 125I-LDL internalization after continuous incubation with 2 ng of 125I-LDL at 37 °C. C, time-dependent 125I-LDL degradation after continuous incubation with 2 ng of 125I-LDL (4 ng/ml) at 37 °C. At each time point, the medium was precipitated with ice-cold TCA, and the TCA-soluble material was used as a measure of the degraded 125I-LDL. Data are representative of three independent experiments each performed in four biological replicates. Points are the averages ± S.D. within one experiment. D and E, HepG2 cells overexpressing WT or RS were incubated with DiI-LDL (5 μg/ml) and analyzed by confocal microscopy as follows: DiI-LDL binding to the cell surface after 4-h incubation at 4 °C (D) or DiI-LDL internalization after 4 h at 37 °C (E). Quantifications in D and E were derived from analyses of 12 transfected cells (EGFP-positive)/condition/experiment. Scale bar, 15 μm. Data are representative of at least two independent experiments. Bars are averages ± S.D. **, p < 0.01 (t test).

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Transfection, Binding Assay, Incubation, Confocal Microscopy, Derivative Assay

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: Residue Arg410 of the LDLR is important for LDL release in acidic early endosomes and for its delivery to late endosomes. A, HepG2 cells overexpressing LDLR-WT (WT) or LDLR-R410S (RS) were incubated with DiI-LDL (5 μg/ml) for 4 h at 37 °C and analyzed by immunofluorescence under permeabilized conditions. Upper panels, co-localization of DiI-LDL with markers for early endosomal compartment (EEA1) and quantification. Lower panels, co-localization of DiI-LDL with markers for late endosomal/lysosomal compartment (Lamp1) and quantification. Quantifications were derived from analyses of 12 transfected cells (EGFP-positive)/condition/experiment. Scale bar, 15 μm. B and C, HEK293 cells overexpressing WT or RS were incubated for 1 h at 4 °C with 15 μg/ml DiI-LDL at pH 7.4, washed, and switched to pH 5.3 for an additional hour at 4 °C. At each pH, the amount of fluorescence from DiI-LDL bound to each receptor was measured by FACS. B, flow cytometry plots for WT (left) and RS (right). C, quantification of DiI-LDL bound at each pH relative to pH 7.4. D, HEK293 cells overexpressing WT or RS were incubated with DiI-LDL (6 μg/ml) for 1 h at 4 °C, washed, and then switched to fresh serum-free DMEM at 37 °C for the indicated times. DiI-LDL released into media was measured and reported relative to DiI-LDL bound at time 0. Data are averages ± S.D. of at least three independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001 (t test).

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Residue, Incubation, Immunofluorescence, Derivative Assay, Transfection, Fluorescence, Flow Cytometry

Journal: The Journal of Biological Chemistry

Article Title: The Proprotein Convertase Subtilisin/Kexin Type 9-resistant R410S Low Density Lipoprotein Receptor Mutation

doi: 10.1074/jbc.M116.769430

Figure Lengend Snippet: At pH 5, the β-propeller mobility of LDLR-R410S is reduced compared with LDLR-WT, and for both receptors the structures at pH 5 are more mobile than the ones at pH 7. A, β-propeller configurations of LDLR-WT (WT) and LDLR-R410S (RS) at pH 7 (left) and pH 5 (right) were generated by molecular dynamics simulations starting with the LDLR-WT structure from PDB code 1N7D. The six blades (β1–β6) and flexible amino acid regions are indicated. The B-factor putty scale is shown as a measure of region flexibility (blue-to-red translated into rigid-to-flexible). B, plots of root mean squared fluctuations values (Å) for each β-propeller residue of LDLR-WT (blue trace) and LDLR-R410S (red trace) at pH 7 (top) and pH 5 (bottom) that were used to calculate the B-factors and generate the configurations in A.

Article Snippet: The supernatants corresponding to the non-denatured cell lysates were subjected to measurement of cell-bound PCSK9 levels (in-house ELISA) ( 60 ), total human LDLR protein levels (human LDLR DuoSet ELISA development kit, DY218; R&D Systems; see below), and total protein (Bio-Rad DC Protein assay).

Techniques: Generated, Residue

Journal: Mucosal immunology

Article Title: IL-11 acts as an alarmin-like pro-inflammatory mediator regulating mucosal responses during helminth infection

doi: 10.1016/j.mucimm.2026.01.005

Figure Lengend Snippet: (A) Published data by our group showing the total number of larvae found in the lungs in different days of infection summarizing the kinetics of Ascaris larval migration in the experimentally infected mice lungs . (B) Transcriptional profile of Ascaris -infected lungs of mice at day 8 post-infection (peak of larval burden in the lungs). Volcano plot shows the significant upregulated and downregulated genes, with the log2 fold change, in the lungs of Ascaris -infected mice over naïve lungs obtained from a nanostring analysis of 507 genes associated with inflammation. Dotted line indicates the p-value threshold in 0.05. Genes labelled in red are differentially expressed genes (DEG) associated with eosinophil activation pathway. Genes labelled in green are DEG associated with neutrophil activation pathway. Line graph showing the profile of cytokine/chemokine levels (mean ± SEM) in the lung tissue homogenate of C57/BL-6 mice during different time points of pulmonary larval migration, including IL-6 (C), IL-11 (D), G-CSF (E), CXCL-1 (F). Dotted line in each graph represents the baseline levels of each analyte in naïve lung homogenates. P values are represented in the graph in comparison to naïve lungs. *p < 0.05, **p < 0.01. *p < 0.001. Scatter plots showing the parasite burden in the lungs of C57BL6 mice infected with different loads of Ascaris eggs (G) and the respective IL-11 protein levels in the lungs of these mice at day 8p.i (H), followed by the Spearman correlation analysis between number of larvae trafficking in the lungs and the levels of IL-11 (pg/mL) (I). p and r values are indicated in the graphs. Differences between naïve and Ascaris -infected groups were considered statistically significant when P < 0.05. Data are representative of 2 independent experiments with n = 5 for each group per experiment.

Article Snippet: After incubation, mouse IL-11 levels were measured in the culture supernatant by using the Duoset ELISA Kit (R&D Systems).

Techniques: Infection, Migration, Activation Assay, Comparison

Journal: Mucosal immunology

Article Title: IL-11 acts as an alarmin-like pro-inflammatory mediator regulating mucosal responses during helminth infection

doi: 10.1016/j.mucimm.2026.01.005

Figure Lengend Snippet: Immunophenotypic analysis of CD45 + lung myeloid cells on the course of larval migration in the lungs of Ascaris -infected (n = 5) and naïve (n = 5) C57/BL6 mice. Representative flow cytometry dot plots showing the frequency of CD11b + Ly6G + neutrophils at day 8p.i (A), SiglecF + CD11ceosinophils at day 18p.i (B) and CD11b + F4/80 + macrophages at day 18p.i (B) in the lungs, and their associate scatter dot plots with their absolute numbers at day 0, 5, 8 and 18p.i. (D) Flow cytometry dot plot of Ascaris -infected lung cells showing the frequency of Lineage − CD11b + CD24 + neutrophils at day 8p.i of C57/BL6 mice treated with isotype control (n = 5) or anti-Ly6G antibodies (n = 5). IL-6 and IL-11 levels at day 8p.i in the neutrophil-depleted Ascaris -infected lung homogenates in comparison with the isotype control. Data is represented by the geometric mean, and the p-values are indicated in each graph. Differences between the groups were considered statistically significant when P < 0.05. Data are representative of 2 independent experiments with n = 5 for each group per experiment.

Article Snippet: After incubation, mouse IL-11 levels were measured in the culture supernatant by using the Duoset ELISA Kit (R&D Systems).

Techniques: Migration, Infection, Flow Cytometry, Control, Comparison

Journal: Mucosal immunology

Article Title: IL-11 acts as an alarmin-like pro-inflammatory mediator regulating mucosal responses during helminth infection

doi: 10.1016/j.mucimm.2026.01.005

Figure Lengend Snippet: (A) Using publicly available single-nucleus RNA-sequencing data from naïve lung cells , a total of 31 clusters of cells in the lung tissue of C57BL6 mice were identified based on their expression profile using Seurat and annotated fine cell types on the ImmGen mouse database using SingleR. (B) Expression level of IL-11 was investigated among the clusters. The frequency of IL-11 + lung cells deconvoluted by cell type among the four major subsets including epithelial, stromal, endothelial and immune cells (C). Confocal imaging analysis of naïve (panels a and b) and Ascaris -infected lungs (c and d) staining with anti-CD45 cells for immune cells (green), anti-EpCAM for epithelial cells (red), intracellular anti-IL-11 for IL-11 expression (yellow) and DAPI for nuclei (blue).

Article Snippet: After incubation, mouse IL-11 levels were measured in the culture supernatant by using the Duoset ELISA Kit (R&D Systems).

Techniques: RNA Sequencing, Expressing, Imaging, Infection, Staining

Journal: Mucosal immunology

Article Title: IL-11 acts as an alarmin-like pro-inflammatory mediator regulating mucosal responses during helminth infection

doi: 10.1016/j.mucimm.2026.01.005

Figure Lengend Snippet: Fresh-frozen lung tissue section of four naïve and four Ascaris -infected WT mice were hybridized with probes to Il11 and probes to either Epcam or Pecam (probe set 1) (A), or probes to Acta2 and Pdgfra (probe set 3 (B)]. Nuclei were counterstained with DAPI (blue). Scale bar = 20 μm. The frequency of IL-11 expressing Epcam + epithelial cells (C), Pecam + endothelial cells (D), Pdgfrα + fibroblasts (E), Acta2 + smooth muscle cells (F) and Acta2 + Pdgfrα + cells (myofibroblasts) (G) were calculated based in the total number of IL11 + DAPI + cells. Data is represented by the geometric mean, and the p-values are indicated in each graph. Differences between the groups were considered statistically significant when P < 0.05.

Article Snippet: After incubation, mouse IL-11 levels were measured in the culture supernatant by using the Duoset ELISA Kit (R&D Systems).

Techniques: Infection, Expressing

Journal: Mucosal immunology

Article Title: IL-11 acts as an alarmin-like pro-inflammatory mediator regulating mucosal responses during helminth infection

doi: 10.1016/j.mucimm.2026.01.005

Figure Lengend Snippet: Recombinant murine IL-11 (2 μg/day) or PBS were daily administered intranasally for 7 days in both naïve (n = 5) and Ascaris -infected (n = 5) C57/BL-6 mice. Parasite burden and cytokine/chemokine profiling were performed in the lungs at day 8p.i (A). IL-11 levels were measured in the lungs homogenates of all four groups (B). Heatmap analysis showing the transformed z-score data of cytokines (C), growth-factors (D) and chemokines (E) levels (pg/mL) in the lung’s homogenate of both naïve and Ascaris -infected C57/BL-6 mice that received either PBS or rIL-11. Scatter plots showing IL-6, CXCL-1, G-CSF and CXCL-5 in the lung homogenate of naïve mice (F) and Ascaris -infected mice (G) that received either PBS or rIL-11. (H) Representative histological analysis of lung section stained with H&E from Ascaris -infected mice at day 8p.i treated daily with PBS (a-c) or treated daily with rIL-11 (d-f). Parameters such as peribronchial (*) and perivascular cellular infiltration (#) were used to calculate the total pulmonary inflammation score (I). Septum thickness (arrows), Ascaris larva (arrowhead) and hemorrhagic areas (h) are also indicated in the panels a and d. Frequency of Ly6G + neutrophils by flow cytometry in all four groups of analysis (J). Data is represented by the geometric mean, and the p-values are indicated in each graph. Differences between the groups were considered statistically significant when P < 0.05. Data are representative of 2 independent experiments with n = 5 for each group per experiment.

Article Snippet: After incubation, mouse IL-11 levels were measured in the culture supernatant by using the Duoset ELISA Kit (R&D Systems).

Techniques: Recombinant, Infection, Transformation Assay, Staining, Flow Cytometry

Journal: Mucosal immunology

Article Title: IL-11 acts as an alarmin-like pro-inflammatory mediator regulating mucosal responses during helminth infection

doi: 10.1016/j.mucimm.2026.01.005

Figure Lengend Snippet: Both mouse bronchial EpCAM + epithelial cells (Cell Biologics, Inc) and mouse bronchial fibroblasts MM14.Lu (ATCC) were cultured at 1 × 10 5 cells/well and stimulated in the absence (media) or presence of 100 ng/mL of recombinant murine cytokines for 24 h at 37 °C in 5 % CO 2 . After incubation, IL-11 levels were measured in the culture supernatant (A). Both cell types were also stimulated in absence or in the presence of recombinant mouse TGF-b1 at different concentrations (1, 10, 100, 1000 ng/mL) in the same culture conditions as above, for IL-11 quantification in the culture supernatant (B). Finally, both mouse bronchial fibroblasts and epithelial cells were stimulated in the same conditions with recombinant mouse IL-11 in a dose–response manner (0, 0.2, 2, 20 and 200 ng/mL) for the quantification of CXCL-1 (C), IL-6 (D) and G-CSF (E) levels in the culture supernatant. Scatter plots represent the geometric mean of four replicates, and the p-values are indicated in each graph. Differences between the groups were considered statistically significant when P < 0.05. Data are representative of 2 independent in vitro experiments with 3–4 replicates each.

Article Snippet: After incubation, mouse IL-11 levels were measured in the culture supernatant by using the Duoset ELISA Kit (R&D Systems).

Techniques: Cell Culture, Recombinant, Incubation, In Vitro

Journal: Mucosal immunology

Article Title: IL-11 acts as an alarmin-like pro-inflammatory mediator regulating mucosal responses during helminth infection

doi: 10.1016/j.mucimm.2026.01.005

Figure Lengend Snippet: Ascaris larval migration from the pulmonary circulation into the lung parenchyma and airways induces mechanical and inflammatory injury to endothelial and epithelial barriers. This tissue damage triggers the rapid upregulation of IL-11 within the lung microenvironment. Based on our data, EpCAM + epithelial cells, PECAM + endothelial cells, and peribronchial stromal populations, including fibroblasts, myofibroblasts, pericytes and airway smooth muscle cells, represent the major sources of IL-11 in helminth-infected lungs. Fibroblasts, and likely differentiated myofibroblasts, respond to TGF-β signaling by inducing IL-11 expression, consistent with established profibrotic pathways. In contrast, the mechanisms by which larval stages directly stimulate epithelial, endothelial, and smooth muscle cells remain unclear. Here, we hypothesize that these structural cell populations act as early responders to tissue damage, rapidly upregulating IL-11 and positioning it as an alarmin-like mediator that amplifies downstream inflammatory responses during acute infection. IL-11 signaling within epithelial and stromal compartments promotes the production of neutrophil-associated mediators, including CXCL-1, G-CSF, and IL-6, leading to an early and robust recruitment of neutrophils from the circulation into lung tissue. Recruited neutrophils, themselves a major source of IL-6, further contribute to sustained pulmonary inflammation. Notably, IL-11-producing cell populations express high levels of IL-11Rα1, supporting a model in which IL-11 signaling is reinforced through autocrine and paracrine feedback loops within damaged lung niches during helminth infection. Figure created with BioRender.

Article Snippet: After incubation, mouse IL-11 levels were measured in the culture supernatant by using the Duoset ELISA Kit (R&D Systems).

Techniques: Migration, Infection, Expressing

Journal: Journal of Veterinary Pharmacology and Therapeutics

Article Title: Pharmacokinetics of the cytochrome P-450 substrates phenytoin, theophylline, and diazepam in healthy Greyhound dogs

doi: 10.1111/j.1365-2885.2011.01316.x

Figure Lengend Snippet: Phenytoin pharmacokinetic parameters in 6 healthy Greyhounds after IV administration of sodium phenytoin (12 mg/kg equivalent to 11 mg/kg phenytoin).

Article Snippet: Drug administration and sample collection Drugs were administered in a non-randomized design, theophylline as aminophylline, 10 mg/kg IV equivalent to 7.88 mg/kg theophylline, (aminophylline 25 mg/mL, Hospira, Inc, Lake Forest, Il, USA), then sodium phenytoin, 12 mg/kg equivalent to 11 mg/kg phenytoin IV (sodium phenytoin 50 mg/mL, Hospira, Inc, Lake Forest, Il, USA), and then diazepam, 0.5 mg/kg IV (5 mg/mL, Hospira, Inc, Lake Forest, Il, USA), with at least 3 weeks between each treatment.

Techniques:

Journal: Journal of Veterinary Pharmacology and Therapeutics

Article Title: Pharmacokinetics of the cytochrome P-450 substrates phenytoin, theophylline, and diazepam in healthy Greyhound dogs

doi: 10.1111/j.1365-2885.2011.01316.x

Figure Lengend Snippet: Plasma concentrations (mean and standard deviation) of phenytoin in 6 healthy Greyhounds after IV sodium phenytoin (12 mg/kg equivalent to 11 mg/kg phenytoin base).

Article Snippet: Drug administration and sample collection Drugs were administered in a non-randomized design, theophylline as aminophylline, 10 mg/kg IV equivalent to 7.88 mg/kg theophylline, (aminophylline 25 mg/mL, Hospira, Inc, Lake Forest, Il, USA), then sodium phenytoin, 12 mg/kg equivalent to 11 mg/kg phenytoin IV (sodium phenytoin 50 mg/mL, Hospira, Inc, Lake Forest, Il, USA), and then diazepam, 0.5 mg/kg IV (5 mg/mL, Hospira, Inc, Lake Forest, Il, USA), with at least 3 weeks between each treatment.

Techniques: Clinical Proteomics, Standard Deviation