Lack of LytA counteracts PSGL-1 deficiency.

<p>(A) Phagocytosis of FAM-SE-labeled pneumococcal strains (D39, D39 <i>lytA</i>) using HL-60 cells exposed to IgG isotype control (open bar) or exposed to KPL-1 (striped bar). (B) Example of flow cytometry histograms showing phagocytosis by HL-60 cells exposed to IgG isotype control (red) or KPL-1 (green). Error bars represent the SDs and asterisks indicate statistical significance after incubation with KPL-1 compared to the exposure to IgG isotype control. **<i>P</i> <0.01 for the comparison between D39 <i>vs</i> D39 <i>lytA</i> in HL-60 cells exposed to KPL-1. (C) Bacterial levels recovered at 24 h from BALF and lungs of <i>PSGL-1</i>^−/− or wild-type mice after pneumonia infection with the pneumococcal D39 <i>lytA</i> mutant. (D). Bacterial levels recovered at 4 h and 24 h from blood of <i>PSGL-1</i>^−/− or wild-type mice after intraperitoneal infection with the pneumococcal D39 <i>lytA</i> strain.</p

Impact of PSGL-1 deficiency in the inflammatory response after pneumococcal infection.

<p>Cytokine levels (pg/ml) in wild-type mice (black bars) and <i>PSGL-1</i>^−/− mice (open bars) at 24 h after intranasal inoculation of <i>S</i>. <i>pneumoniae</i> D39 strain. (A) Cytokines determined in BALF. (B) Cytokines measured in sera. Error bars represent the SDs and asterisks indicate statistical significance of cytokine levels of <i>PSGL-1</i>^−/− mice compared to wild-type mice.</p

Contribution of PSGL-1 to cellular migration in lungs and blood of infected mice with <i>S</i>. <i>pneumoniae</i>.

<p>(A) Neutrophils, macrophages, CD4⁺T and CD8⁺T cells in the lungs of wild-type (black bars) or <i>PSGL-1</i>^−/− mice (open bars) after pneumonia infection. (B) Proportion of neutrophils, macrophages, CD4⁺T and CD8⁺T cells in the blood of wild-type (black bars) or <i>PSGL-1</i>^−/− mice (open bars) after intravenous infection. (C) Representative lung histological sections from infected mice that show granulocytes infiltrates in <i>PSGL-1</i>^−/− mice visualized by Ly6G staining. T cell infiltrates were observed by CD3 staining. Original magnification × 10. Scale bars represent 200 μm.</p

PSGL-1 on HL-60 cells and murine neutrophils is required for phagocytosis of <i>S</i>. <i>pneumoniae</i>.

<p>(A) Phagocytosis mediated by PSGL-1 of FAM-SE-labeled pneumococcal clinical isolates belonging to different serotypes using a flow cytometry assay. HL-60 cells were incubated with an IgG isotype negative control that does not block PSGL-1 (black bars), whereas cells exposed to the KPL-1 antibody had the PSGL-1 blocked (open bars). (B) Examples of flow cytometry histograms of pneumococcal binding to cells incubated with KPL-1 (red) or IgG control (green). (C) Bacterial survival indicated as CFU/ml of recovered bacteria from HL-60 cells previously exposed (open bars) or not (black bars) to KPL-1. (D−E) Phagocytosis of FAM-SE-labeled D39 strain by neutrophils isolated from the spleen of wild-type (black bar and green histogram) or <i>PSGL-1</i>^–/–mice (open bar and red histogram). The blue line in the histogram shows the pattern of non-infected cells. Error bars represent the SDs and asterisks indicate statistical significance after incubation with KPL-1 compared to the exposure to IgG isotype control or between OP using neutrophils from wild-type mice <i>vs</i> those from <i>PSGL-1</i>^–/–mice. Error bars represent the SDs and asterisks indicate statistical significance after incubation with KPL-1 compared to the exposure to IgG isotype control.</p

PSGL-1 contributes to the uptake of <i>S</i>. <i>pneumoniae</i>.

<p>Kinetics of pneumococcal phagocytosis of <i>S</i>. <i>pneumoniae</i> D39 (serotype 2) strain expressing GFP mediated by PSGL-1. Images are visualized at different times by live imaging confocal microscopy and environmental control (A) Phagocytosis of D39 strain using HL-60 cells with the PSGL-1 active. (B) Phagocytosis of D39 strain by HL-60 cells with the PSGL-1 blocked after incubation with KPL-1.</p

Contribution of PSGL-1 to host defense against invasive pneumococcal disease.

<p>(A) Bacterial levels recovered at 6 h and 24 h from BALF, lung and blood of <i>PSGL-1</i>^−/− mice (open bars) and wild-type mice (black bars) after pneumonia infection with the pneumococcal D39 strain. (B) Bacterial levels recovered from blood of <i>PSGL-1</i>^−/− mice (open bars) and wild-type mice (black bars) after sepsis caused by D39 strain. (C) Survival rates of <i>PSGL-1</i>^−/− mice (solid line) and wild-type mice (dotted line) after pneumococcal sepsis produced by D39 <i>S</i>. <i>pneumoniae</i> strain. (D) Bacterial levels recovered from blood of <i>PSGL-1</i>^−/− mice (open bars) and wild-type mice (black bars) after intravenous infection with D39 strain. (E-F) Bacterial counts in blood and survival of <i>PSGL-1</i>^−/− mice (open bars and solid line) and wild-type mice (black bars and dotted line) infected by the intraperitoneal route (sepsis) with TIGR4 strain. Error bars represent the SDs and asterisks indicate statistical significance of bacterial levels of <i>PSGL-1</i>^−/− mice compared to wild-type mice. For differences in survival between wild-type mice and <i>PSGL1</i>^−/− mice a long-rank test was used (<i>P</i> <0.01 for D39 strain and <i>P</i> <0.05 for TIGR4 strain).</p

DataSheet_1_Age-dependent nasal immune responses in non-hospitalized bronchiolitis children.pdf

Bronchiolitis in children is associated with significant rates of morbidity and mortality. Many studies have been performed using samples from hospitalized bronchiolitis patients, but little is known about the immunological responses from infants suffering from mild/moderate bronchiolitis that do not require hospitalization. We have studied a collection of nasal lavage fluid (NLF) samples from outpatient bronchiolitis children as a novel strategy to unravel local humoral and cellular responses, which are not fully characterized. The children were age-stratified in three groups, two of them (GI under 2-months, GII between 2-4 months) presenting a first episode of bronchiolitis, and GIII (between 4 months and 2 years) with recurrent respiratory infections. Here we show that elevated levels of pro-inflammatory cytokines (IL1β, IL6, TNFα, IL18, IL23), regulatory cytokines (IL10, IL17A) and IFNγ were found in the three bronchiolitis cohorts. However, little or no change was observed for IL33 and MCP1, at difference to previous results from bronchiolitis hospitalized patients. Furthermore, our results show a tendency to IL1β, IL6, IL18 and TNFα increased levels in children with mild pattern of symptom severity and in those in which non RSV respiratory virus were detected compared to RSV+ samples. By contrast, no such differences were found based on gender distribution. Bronchiolitis NLFs contained more IgM, IgG1, IgG3 IgG4 and IgA than NLF from their age-matched healthy controls. NLF from bronchiolitis children predominantly contained neutrophils, and also low frequency of monocytes and few CD4+ and CD8+ T cells. NLF from infants older than 4-months contained more intermediate monocytes and B cell subsets, including naïve and memory cells. BCR repertoire analysis of NLF samples showed a biased VH1 usage in IgM repertoires, with low levels of somatic hypermutation. Strikingly, algorithmic studies of the mutation profiles, denoted antigenic selection on IgA-NLF repertoires. Our results support the use of NLF samples to analyze immune responses and may have therapeutic implications.</p

Psychologické a sociální aspekty umírání a smrti

<p>Data are expressed as mean ± SEM. *p<0.05 vs. CONTROL; ^#p<0.05 vs. ALDO</p

(A) Representative images showing Sirius red staining, Masson trichrome and Hematoxilin/Eosin staining in control+PBS liposome (Control+Lipo), aldosterone+PBS liposome (Aldo+Lipo), control+clodronate liposome (Control+Clodr), or aldosterone+clodronate liposome (Aldo+Clodr) treated animals. Kidney collagen content (B), quantitative analyses of Col I and Fibronectin (FN) protein (C-D) and mRNA expression (E) in treated rats.

<p>Data are expressed as mean ± SEM. *p<0.05 vs Control+Lipo; #p<0.05 vs. Aldo+Lipo.</p

Kidney collagen content by Sirius red (A) and Masson trichrome and representative images of collagen content by Hematoxilin/Eosin (B) in control (CONTROL), spironolactone treated animals (SPIRO), aldosterone+salt-treated animals (ALDO) and aldosterone+salt plus spironolactone treated animals (ALDO+SPIRO).

<p>Data are expressed as mean ± SEM. *p<0.05 vs. CONTROL; ^#p<0.05 vs. ALDO</p