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Endotoxin induces the expression of macrophage inflammatory protein 1 alpha mRNA by rat alveolar and bone
J W Christman1, T R Blackwell, H B Cowan
1Department of Veterans Affairs, Vanderbilt University, Nashville, Tennessee 37232-2650.
Abstract:
Macrophage inflammatory protein 1 alpha (MIP-1 alpha) is a newly described cytokine that is present in large amounts in the culture supernatant of an endotoxin-stimulated murine macrophage-like cell line (RAW 264.7). There is increasing information that suggests that this cytokine mediates acute neutrophilic inflammation, although the mechanism of mediation is unknown. Data examining the production and regulation of MIP-1 alpha by primary rat macrophages are lacking, and MIP-1 alpha has not been studied previously in an animal model of endotoxin-induced neutrophilic alveolitis. In this study, we performed Northern analysis of steady-state rat MIP-1 alpha mRNA using an oligonucleotide probe complementary to amino acids 4-13 of murine MIP-1 alpha. Our data demonstrate that rat alveolar and bone marrow-derived macrophages can be induced by in vitro endotoxin treatment to express a 1.1-kb MIP-1 alpha mRNA. Expression of the mRNA could be elicited by treatment with 0.1 to 10.0 micrograms/ml of endotoxin in vitro with peak steady-state levels detectable up to 9 h after adding endotoxin to the media. Alveolar macrophages recovered by whole lung lavage from endotoxin-treated rats expressed increased amounts of the mRNA homologous to MIP-1 alpha mRNA when treated in vitro with endotoxin. We also found that rat neutrophils could be induced by endotoxin in vitro to express the MIP-1 alpha mRNA. We were able to identify MIP-1 alpha in culture supernatant from endotoxin-stimulated rat alveolar and bone marrow-derived macrophages by immunoprecipitation with a specific goat anti-murine MIP-1 alpha.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
This study shows that endotoxin stimulates rat macrophages and neutrophils to produce Macrophage inflammatory protein 1 alpha (MIP-1 alpha) mRNA. This cytokine is involved in acute neutrophilic inflammation.
Area of Science:
- Immunology
- Cell Biology
- Molecular Biology
Background:
- Macrophage inflammatory protein 1 alpha (MIP-1 alpha) is a cytokine implicated in acute neutrophilic inflammation.
- The production and regulation of MIP-1 alpha by primary rat macrophages and its role in endotoxin-induced neutrophilic alveolitis are not well understood.
Purpose of the Study:
- To investigate the expression and regulation of MIP-1 alpha mRNA in primary rat macrophages and neutrophils following endotoxin stimulation.
- To examine the role of MIP-1 alpha in an animal model of endotoxin-induced neutrophilic alveolitis.
Main Methods:
- Northern analysis was used to detect MIP-1 alpha mRNA in rat alveolar and bone marrow-derived macrophages and neutrophils.
- Primary rat macrophages were stimulated in vitro with varying concentrations of endotoxin.
- Immunoprecipitation was employed to identify MIP-1 alpha in culture supernatants.
Main Results:
- Endotoxin treatment induced the expression of a 1.1-kb MIP-1 alpha mRNA in rat alveolar and bone marrow-derived macrophages in vitro.
- Peak mRNA levels were observed up to 9 hours after endotoxin exposure.
- Endotoxin also induced MIP-1 alpha mRNA expression in rat neutrophils.
- MIP-1 alpha was identified in the supernatant of stimulated macrophages.
Conclusions:
- Rat alveolar and bone marrow-derived macrophages, as well as neutrophils, can be induced by endotoxin to express MIP-1 alpha mRNA.
- These findings provide insights into the cellular sources and regulation of MIP-1 alpha during endotoxin-induced inflammation in rats.