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A Method for Generating Pulmonary Neutrophilia Using Aerosolized Lipopolysaccharide
Published on: December 15, 2014
Neutrophil modulation of the pulmonary chemokine response to lipopolysaccharide
Michael C Holmes1, Ping Zhang, Steve Nelson
1Department of Physiology, Louisiana State University Health Sciences Center, New Orleans, Louisiana 70112, USA.
Abstract:
When cells within the intrapulmonary compartment are exposed to pathogens or their products such as lipopolysaccharide, they produce CXC chemokines in order to attract circulating neutrophils into the lower respiratory tract. Previous studies have shown that as neutrophils (PMNs) enter the lung, bronchoalveolar lavage (BAL) chemokine levels are decreased. In this study, we determined the intrapulmonary and systemic responses to two important rat chemokines, cytokine-induced neutrophil chemoattractant (CINC) and macrophage inflammatory protein-2 (MIP-2), to intratracheal (i.t.) LPS (100 microg in 0.5 mL of phosphate-buffered saline) under neutropenic (cyclophosphamide [CPA]) and neutrophilic (G-CSF) conditions. By 4 h after i.t. LPS, CPA pretreatment decreased PMN recruitment 83% and G-CSF increased PMN recruitment 91% compared with recruitment into the lung in vehicle-pretreated rats (42.7 +/- 19.3 million PMNs). Neutropenic rats had increased CINC and MIP-2 concentrations in BAL fluid 4 h after i.t. LPS when compared with levels seen in vehicle controls (P < 0.05). In vitro LPS-stimulated chemokine production by alveolar macrophages obtained from CPA- and vehicle-pretreated animals did not differ. The increase in BAL fluid chemokine levels in neutropenic rats corresponded to increased chemotaxis of neutrophils to BAL fluid from CPA-pretreated rats as compared with the chemotaxis response of PMN to BAL fluid from vehicle-pretreated rats. In contrast, G-CSF enhancement of neutrophil recruitment decreased chemotactic activity of BAL fluid collected 4 h after i.t. LPS. These data show that as neutrophils are recruited into the lung, they alter chemokine levels, which most likely serves to down-regulate the inflammatory response.
Insights
Neutrophils entering the lungs decrease inflammatory chemokine levels, suggesting a self-regulating mechanism. This study investigated cytokine-induced neutrophil chemoattractant (CINC) and macrophage inflammatory protein-2 (MIP-2) responses in rats.
Area of Science:
- Immunology
- Respiratory Medicine
- Cell Biology
Background:
- Intrapulmonary cells produce CXC chemokines to recruit neutrophils to the lower respiratory tract upon pathogen exposure.
- Previous research indicates a decrease in bronchoalveolar lavage (BAL) chemokine levels as neutrophils enter the lung.
Purpose of the Study:
- To investigate the intrapulmonary and systemic responses of rat chemokines, cytokine-induced neutrophil chemoattractant (CINC) and macrophage inflammatory protein-2 (MIP-2).
- To determine the effect of neutropenic (cyclophosphamide [CPA]) and neutrophilic (G-CSF) conditions on chemokine levels and neutrophil recruitment following intratracheal lipopolysaccharide (LPS) instillation.
Main Methods:
- Rats were pretreated with CPA (neutropenic) or G-CSF (neutrophilic) or vehicle before intratracheal LPS administration.
- Neutrophil counts in BAL fluid were quantified.
- Chemokine (CINC, MIP-2) concentrations in BAL fluid were measured.
- In vitro chemokine production by alveolar macrophages was assessed.
- Neutrophil chemotaxis towards BAL fluid was evaluated.
Main Results:
- CPA pretreatment significantly decreased neutrophil recruitment by 83%, while G-CSF increased it by 91% compared to controls.
- Neutropenic rats exhibited higher CINC and MIP-2 levels in BAL fluid post-LPS.
- In vitro, LPS-stimulated chemokine production by alveolar macrophages did not differ between CPA- and vehicle-treated groups.
- Increased BAL fluid chemokine levels in neutropenic rats correlated with enhanced neutrophil chemotaxis.
- G-CSF treatment led to decreased BAL fluid chemotactic activity.
Conclusions:
- Neutrophil recruitment into the lung alters local chemokine levels.
- These alterations in chemokine levels likely serve to down-regulate the inflammatory response.
- The findings highlight a feedback mechanism in pulmonary inflammation involving neutrophils and chemokines.
