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Human alveolar macrophages synthesize endothelins by thrombin
Y Kobayashi1, Y Sakamoto, M Shibasaki
1Second Department of Internal Medicine, Saitama Medical School, Irumagun, Japan.
Abstract:
Alveolar macrophages (AM) play pivotal roles in the defense mechanism and the regulation of inflammatory processes in the airways. Macrophages express receptors for thrombin on their surfaces, and thrombin induces the chemotaxis and the proliferation of macrophages. Thrombin acts on vascular endothelial cells to synthesize endothelin (ET)-1. AM have been known to express prepro ET-1 mRNA. Thus, we hypothesized that thrombin stimulates AM so as to synthesize ET. Surgically resected human lungs were irrigated by saline to remove intravascular blood, then saline was instilled into the bronchus, and the fluids were recovered. AM were separated by Percoll density centrifugation (density, 1.060 g/ml). AM were resuspended in culture medium without FCS in the presence or absence of thrombin. ET was synthesized by thrombin in a concentration-dependent manner, and the amounts of ET synthesized by thrombin (10 U/ml) were equivalent to those by LPS (10 microg/ml). Dexamethasone (10(-6)-10(-10) M), IL-4 (100 U/ml), and TGF-beta (10 ng/ml) significantly suppressed the ET synthesis by thrombin (p < 0.05). In contrast, 1,25-dihydroxyvitamin D3 (10(-8)-10(-10) M) enhanced the ET synthesis up to approximately 300%. The analysis using high pressure liquid chromatography revealed that AM-derived ET consists of ET-1, ET-2, and ET-3. Major constituents were ET-2 and ET-1, and the ratio of ET-2/ET-1 was 1.7 +/- 0.4 (mean +/- SE). These results indicate that thrombin is a potent agonist for AM to synthesize ET.
Insights
Thrombin stimulates alveolar macrophages (AM) to produce endothelin (ET). This finding reveals a new pathway for regulating airway inflammation and ET synthesis in the lungs.
Area of Science:
- Pulmonary immunology
- Cellular signaling
- Molecular biology
Background:
- Alveolar macrophages (AM) are key immune cells in the airways, regulating defense and inflammation.
- Thrombin, a protease, influences macrophage behavior and stimulates endothelin (ET) production in endothelial cells.
- AM express prepro ET-1 mRNA, suggesting a potential role in ET synthesis.
Purpose of the Study:
- To investigate whether thrombin stimulates alveolar macrophages (AM) to synthesize endothelin (ET).
- To characterize the types and regulation of ET produced by AM in response to thrombin.
Main Methods:
- Human lungs were lavaged to isolate AM.
- AM were cultured with or without thrombin.
- Endothelin (ET) synthesis was measured.
- The effects of dexamethasone, IL-4, TGF-beta, and 1,25-dihydroxyvitamin D3 on ET synthesis were assessed.
- High-performance liquid chromatography (HPLC) was used to analyze ET components.
Main Results:
- Thrombin induced concentration-dependent ET synthesis by AM, comparable to LPS stimulation.
- Dexamethasone, IL-4, and TGF-beta suppressed thrombin-induced ET synthesis.
- 1,25-dihydroxyvitamin D3 significantly enhanced ET synthesis.
- AM-derived ET comprised ET-1, ET-2, and ET-3, with ET-2 and ET-1 as major components.
Conclusions:
- Thrombin acts as a potent agonist, stimulating alveolar macrophages (AM) to synthesize endothelin (ET).
- This pathway highlights a novel mechanism linking coagulation and airway inflammation.
- The findings suggest AM play a significant role in local ET production within the lungs.