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Complement receptor expression on neutrophils at an inflammatory site, the Pseudomonas-infected lung in cystic
M Berger1, R U Sorensen, M F Tosi
1Department of Pediatrics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
Abstract:
Activation of human neutrophils (PMN) is accompanied by rapid upregulation of CR1, the C3b receptor, and CR3, the iC3b receptor, which also serves as the PMN's major adherence protein. This is necessary for migration and phagocytosis, but the extent of expression of these proteins on PMN at inflammatory sites has not been determined. We used monoclonal antibodies and flow cytometry to assess CR1 and CR3 expression on PMN in bronchoalveolar lavage (BAL) fluid of cystic fibrosis (CF) patients chronically infected with pseudomonas and in sterile joint fluid of arthritis patients. Resting peripheral blood PMN from these patients and normals expressed similar low levels of CR1 and CR3, and the patients' PMN increased CR1 and CR3 expression normally when stimulated in vitro. CR3 expression on CF BAL PMN was 90 +/- 12% of that on the same patient's blood cells stimulated in vitro with FMLP. In contrast, CR1 expression on BAL PMN was only 27 +/- 8% of that on stimulated blood cells. Similar results were obtained for joint PMN. This pattern could be reproduced in vitro by treating FMLP-stimulated blood cells with BAL supernatants or with pseudomonas or PMN elastase. The serine protease inhibitors, PMSF and alpha 1-antitrypsin prevented the lavage supernatant from reducing CR1 expression, while metalloprotease inhibitors had no effect. Treatment of PMN with elastase in vitro decrease their ability to kill opsonized Pseudomonas aeruginosa. These results suggest that PMN at inflammatory sites have maximally upregulated expression of their complement receptors, but that CR1 is then cleaved by proteolysis in situ. Although not related to the basic defect in CF, this may interfere with efficient phagocytosis and contribute to the CF patient's inability to eradicate chronic lung infection.
Insights
Neutrophils (PMN) at inflammatory sites upregulate complement receptors CR1 and CR3. However, CR1 is cleaved by proteolysis, potentially impairing phagocytosis and chronic infection clearance in cystic fibrosis patients.
Area of Science:
- Immunology
- Cell Biology
- Respiratory Medicine
Background:
- Neutrophils (PMN) express complement receptors CR1 (C3b receptor) and CR3 (iC3b receptor) crucial for migration and phagocytosis.
- The expression levels of CR1 and CR3 on PMN at inflammatory sites remain undetermined.
- Chronic Pseudomonas aeruginosa infection in cystic fibrosis (CF) patients highlights the need to understand PMN function in inflammatory environments.
Purpose of the Study:
- To determine the expression levels of CR1 and CR3 on PMN in bronchoalveolar lavage (BAL) fluid of CF patients and joint fluid of arthritis patients.
- To investigate the mechanisms underlying altered complement receptor expression at inflammatory sites.
- To assess the functional consequences of altered CR1 expression on PMN's ability to combat Pseudomonas aeruginosa.
Main Methods:
- Monoclonal antibodies and flow cytometry were used to quantify CR1 and CR3 expression on PMN.
- PMN were isolated from peripheral blood, BAL fluid of CF patients, and joint fluid of arthritis patients.
- In vitro experiments involved stimulating PMN with FMLP and treating them with lavage supernatants or purified proteases.
Main Results:
- PMN from inflammatory sites (BAL fluid, joint fluid) showed maximal CR3 upregulation but significantly reduced CR1 expression compared to stimulated blood PMN.
- In vitro treatment of stimulated PMN with BAL supernatants or Pseudomonas/PMN elastase mimicked the reduction in CR1 expression.
- Protease inhibitors (PMSF, alpha 1-antitrypsin) blocked the reduction of CR1 expression by lavage supernatants, implicating serine proteases.
- Treatment of PMN with elastase in vitro impaired their ability to kill opsonized Pseudomonas aeruginosa.
Conclusions:
- PMN at inflammatory sites exhibit maximal CR3 expression, but CR1 is cleaved by proteolysis in situ.
- This proteolytic cleavage of CR1 may hinder efficient phagocytosis of pathogens.
- While not the primary defect in CF, impaired CR1 function could contribute to the difficulty in clearing chronic lung infections.
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