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Clinical and Diagnostic Laboratory Immunology, Jan 1995, 18-24, Vol 2, No. 1
Copyright © 1995 by the American Society for Microbiology. All rights reserved.

Quantitative reverse transcription-PCR analysis of Legionella pneumophila-induced cytokine mRNA in different macrophage populations by high-performance liquid chromatography

Y Yamamoto, C Retzlaff, P He, TW Klein and H Friedman
Department of Medical Microbiology and Immunology, University of South Florida College of Medicine, Tampa 33612.

Cytokine production in macrophages infected by bacteria is critical for the course of infection. However, it is not known how infection of macrophages with opportunistic bacteria leads to cytokine production in different populations of cells. Since it is possible that cytokine genes may be differentially regulated by attachment rather than by active infection, the levels of various cytokine mRNAs were measured in alveolar macrophages (AMs), peritoneal resident macrophages (RMs), and peritoneally elicited macrophages (EMs) interacting with Legionella pneumophila by using cytochalasin D-treated macrophages and a newly developed quantitative reverse transcription-PCR procedure with high- performance liquid chromatographic analysis to determine cytokine mRNA formation. Increased levels of interleukin-1 beta (IL-1 beta), IL-6, tumor necrosis factor alpha, granulocyte-macrophage colony-stimulating factor, and macrophage inflammatory protein 2 mRNAs were quantitated in the macrophages responding to L. pneumophila attachment in vitro. Using this technique, we showed that the three different macrophage populations responded differently to bacterial attachment. We found that the levels of IL-6 and granulocyte-macrophage colony-stimulating factor mRNAs induced by the attachment of L. pneumophila to AMs were significantly lower than the levels in RMs but similar to the levels in EMs. Furthermore, the levels of MIP-2 mRNA in the AMs were found to be higher than those in the RMs, but similar levels were found in EMs. IL- 1 beta mRNA levels were higher in both AMs and RMs than in EMs, but tumor necrosis factor alpha levels were not different among the three macrophage populations examined.(ABSTRACT TRUNCATED AT 250 WORDS)

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