R-spondin 2 mediates neutrophil egress into the alveolar space through increased lung permeability.

Abstract Objective R-spondin 2 (RSPO2) is required for lung morphogenesis, activates Wnt signaling, and is upregulated in idiopathic lung fibrosis. Our objective was to investigate whether RSPO2 is similarly important in homeostasis of the adult lung. While investigating the characteristics of bronchoalveolar lavage in RSPO2-deficient (RSPO2-/-) mice, we observed unexpected changes in neutrophil homeostasis and vascular permeability when compared to control (RSPO2+/+) mice at baseline. Here we quantify these observations to explore how tonic RSPO2 expression impacts lung homeostasis. Results Quantitative PCR (qPCR) analysis demonstrated significantly elevated myeloperoxidase (MPO) expression in bronchoalveolar lavage fluid (BALF) cells from RSPO2-/- mice. Likewise, immunocytochemical (ICC) analysis demonstrated significantly more MPO+ cells in BALF from RSPO2-/- mice compared to controls, confirming the increase of infiltrated neutrophils. We then assessed lung permeability/barrier disruption via Fluorescein Isothiocyanate (FITC)-dextran instillation and found a significantly higher dextran concentration in the plasma of RSPO2-/- mice compared to identically treated RSPO2+/+ mice. These data demonstrate that RSPO2 may be crucial for blood-gas barrier integrity and can limit neutrophil migration from circulation into alveolar spaces associated with increased lung permeability and/or barrier disruption. This study indicates that additional research is needed to evaluate RSPO2 in scenarios characterized by pulmonary edema or neutrophilia.

Download Full-text

R-spondin 2 mediates neutrophil egress into the alveolar space through increased lung permeability.

10.21203/rs.2.18790/v1 ◽

2019 ◽

Author(s):

Sergio Randell Jackson ◽

Maria Fernanda De Mello Costa ◽

Christopher Frances Pastore ◽

Gan Zhao ◽

Aaron I. Weiner ◽

...

Keyword(s):

Neutrophil Migration ◽

Fluorescein Isothiocyanate ◽

Lavage Fluid ◽

Permeability Barrier ◽

Cell Content ◽

Barrier Disruption ◽

Lung Permeability ◽

Balf Cell ◽

Lung Vascular Permeability ◽

Experimental Injury

Abstract Objective: R-spondin 2 (RSPO2) is required for proper lung morphogenesis. Our objective was to investigate whether RSPO2 is similarly important in homeostasis of the adult lung. Unexpectedly, we observed changes in neutrophil migration and lung vascular permeability in RSPO2-deficient (RSPO2-/-) mice compared to RSPO2 control (RSPO2+/+) mice, independent of experimental injury/challenge. Here we use multiple methods to quantify these observations to further understand how tonic RSPO2 expression regulates lung homeostasis. Results: Quantitative PCR (qPCR) analysis demonstrated significantly higher myeloperoxidase (MPO) expression in bronchoalveolar lavage fluid (BALF) cell content from RSPO2-/- mice compared to RSPO2+/+ mice. Immunocytochemical (ICC) analysis likewise demonstrated significantly more MPO+ cells in BALF from RSPO2-/- mice compared to RSPO2+/+ mice, confirming the increase of infiltrated neutrophils. We then assessed lung permeability/barrier disruption via Fluorescein isothiocyanate (FITC)-dextran instillation and found a significantly higher dextran concentration in the plasma of RSPO2-/- mice compared to identically treated RSPO2+/+ mice. These data demonstrate that RSPO2 may be crucial for lung barrier integrity and can facilitate an increase in neutrophil migration from circulation into alveolar spaces due to increased lung permeability/barrier disruption. Our studies suggest additional research is needed to evaluate RSPO2 in scenarios exhibiting either pulmonary edema or neutrophilia.

Download Full-text

Initial recruitment of neutrophils to alveolar structures in acute lung injury

Journal of Applied Physiology ◽

10.1152/jappl.1991.70.4.1575 ◽

1991 ◽

Vol 70 (4) ◽

pp. 1575-1585 ◽

Cited By ~ 14

Author(s):

G. C. Kindt ◽

J. E. Gadek ◽

J. E. Weiland

Keyword(s):

Respiratory Distress Syndrome ◽

Respiratory Distress ◽

Bronchoalveolar Lavage ◽

Bacterial Pneumonia ◽

Bronchoalveolar Lavage Fluid ◽

Distress Syndrome ◽

Neutrophil Migration ◽

Lavage Fluid ◽

Neutrophil Influx ◽

Lung Lymph

The adult respiratory distress syndrome and bacterial pneumonia are both characterized by an influx of neutrophils into the lung. The neutrophil has been implicated as having a “pathological” role in adult respiratory distress syndrome, in contrast to its role in bacterial pneumonia. We hypothesized that processes resulting in neutrophil recruitment to the lung are distinct, depending on whether the inflammatory stimulus arises in the intravascular or the alveolar compartment of the lung. Anesthetized sheep with lung lymph fistulas were utilized to access the three compartments of the lung relevant to studies of transpulmonary neutrophil migration. Serum, lung lymph, and bronchoalveolar lavage fluid were studied for neutrophil influx and chemotactic activity before and after administration of endotoxin by either an intravascular or inhaled alveolar route. Both groups developed significant neutrophil influx into the lymph and bronchoalveolar lavage fluid by 3 h postendotoxin. Those animals receiving intravascular endotoxin developed chemotactic gradients opposing neutrophil migration into the lung in contrast to animals receiving alveolar endotoxin, suggesting that neutrophil influx into the lung occurs by random migration.

Download Full-text

Immunomodulating Effects of HMR 3004 on Pulmonary Inflammation Caused by Heat-Killed Streptococcus pneumoniae in Mice

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.42.12.3309 ◽

1998 ◽

Vol 42 (12) ◽

pp. 3309-3312 ◽

Cited By ~ 17

Author(s):

Michel Duong ◽

Marie Simard ◽

Yves Bergeron ◽

Nathalie Ouellet ◽

Mélanie Côté-Richer ◽

...

Keyword(s):

Nitric Oxide ◽

Streptococcus Pneumoniae ◽

Bronchoalveolar Lavage ◽

Bacterial Pneumonia ◽

Bronchoalveolar Lavage Fluid ◽

Pulmonary Inflammation ◽

Antimicrobial Properties ◽

Fluorescein Isothiocyanate ◽

Lavage Fluid ◽

Lung Edema

ABSTRACT We investigated the influence of HMR 3004, a new ketolide antibiotic, on the pulmonary inflammation induced by heat-killed fluorescein isothiocyanate-labeled Streptococcus pneumoniae. HMR 3004 downregulated (P < 0.05) the pneumococcus-induced release of interleukin-6 (IL-6), IL-1β, and nitric oxide in bronchoalveolar lavage fluid. The drug limited (P < 0.05) neutrophil recruitment to lung tissues and alveoli but did not interfere with phagocytosis. HMR 3004 totally abrogated lung edema. By reducing inflammation in addition to possessing antimicrobial properties, HMR 3004 may participate in improving the outcome of bacterial pneumonia.

Download Full-text

The effect of induced sputum and bronchoalveolar lavage fluid from patients with chronic obstructive pulmonary disease on neutrophil migration in vitro

Medicina ◽

10.3390/medicina46050044 ◽

2010 ◽

Vol 46 (5) ◽

pp. 315 ◽

Cited By ~ 4

Author(s):

Agnė Babušytė ◽

Jolanta Jeroch ◽

Rimantas Stakauskas ◽

Kristina Stravinskaitė ◽

Kęstutis Malakauskas ◽

...

Keyword(s):

Bronchoalveolar Lavage ◽

Bronchoalveolar Lavage Fluid ◽

Neutrophil Migration ◽

Lavage Fluid ◽

Interleukin 8 ◽

Induced Sputum ◽

Chronic Obstructive ◽

Healthy Individuals ◽

Copd Patients

Objective. The aim of study was to investigate a chemotactic effect of induced sputum and bronchoalveolar lavage fluid on blood neutrophils in patients with chronic obstructive pulmonary disease (COPD) and healthy individuals. Material and methods. Forty-three smokers with COPD, 19 ex-smokers with COPD, 13 healthy smokers, and 17 healthy nonsmokers were recruited to the study. Neutrophils were isolated from peripheral blood of study individuals. For the same experimental conditions, pooled induced sputum and bronchoalveolar lavage fluid of 20 COPD patients were used. Neutrophil chemotaxis in vitro was performed in cell-transmigration chamber. Substances tested for chemoattraction (interleukin-8, induced sputum, bronchoalveolar lavage fluid directly or in addition to interleukin-8) were added to lower wells. Upper wells were filled with 2.5×106/mL of neutrophil culture and incubated for 2 hours. Migration was analyzed by flow cytometry. Results. Interleukin-8 (10–100 ng/mL) induced a dose-dependant neutrophil migration in all the groups. Only 100 ng/L of interleukin-8 induced more intensive chemotaxis of neutrophils from COPD smokers as compared to ex-smokers (P<0.05). Such difference between healthy individuals was obtained using 30 ng/mL of interleukin-8 (P<0.05). Induced sputum/interleukin-8 (10–100 ng/mL), as well as induced sputum directly, induced neutrophil migration (P<0.05). Chemotaxis of neutrophils isolated from COPD patients and healthy nonsmokers did not depend on additional interleukin-8 concentration. Bronchoalveolar lavage fluid/interleukin-8 (30–100 ng/mL) induced more intensive migration of neutrophils from COPD patients than bronchoalveolar lavage fluid (P<0.05) alone. Conclusions. Migration of neutrophils isolated from patients with COPD was more intensive compared to healthy individuals. Induced sputum and bronchoalveolar lavage fluid directly and with addition of interleukin-8 stimulated chemotaxis, and it was higher in neutrophils from COPD patients. Migration of neutrophils did not depend on smoking status.

Download Full-text