Loren C. Denlinger 1990-1998
University Hospitals and Clinics, Madison, WI
University of Michigan 1990
Medical Microbiology & Immunology PhD 1996
Richard Proctor, Paul Bertics
Adenine nucleotide regulation of murine endotoxicity
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The release of endotoxin (lipopolysaccharide, LPS) into the bloodstream can result in lethal septic shock. Macrophages express several classes of LPS receptors and are stimulated by LPS to produce multiple inflammatory mediators responsible for the immunotoxicity of LPS (e.g., tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO)). Recent evidence suggests that purinoreceptors, which bind extracellular adenine nucleotides, have a controlling influence on macrophage function. Therefore, the major hypothesis of my dissertation is that extracellular adenine nucleotides serve as an early inflammatory mediator, that regulates LPS-stimulated macrophage production of TNF-alpha and NO. In Chapters 2 and 3, I have demonstrated that selected purinoreceptor agonists synergize with toxic LPS preparations to induce the production of NO by murine RAW 264.7 macrophages, whereas a purinoreceptor antagonist, 2-MeS-ATP, inhibits the production of TNF-alpha and NO by LPS-treated macrophages. 2-MeS-ATP administration to mice reduces LPS-induced serum levels of TNF-alpha and partially (50%) blocks NO production by peritoneal macrophages cultured ex vivo. This purinoreceptor antagonist does not block all LPS-stimulated inflammatory mediators, however, 2-MeS-ATP treatment protects 95% of mice given a lethal dose of LPS, suggesting that extracellular adenine nucleotides can regulate LPS-stimulated events that are important to the pathophysiology of murine endotoxemia. Using nine endotoxins that varied in their capacities to induce murine lethality by over 160-fold, I have shown that the nuclear translocation of NF-kappaB and the production of TNF-alpha are stimulated equally by toxic and nontoxic endotoxins (Chapter 4). In contrast, LPS-induced NO production by RAW 264.7 cells predicts endotoxin lethality. Cotreatment with exogenous interferon-gamma normalizes macrophage production of NO induced by toxic and nontoxic endotoxins, and this cytokine induces signals distinct from those regulated by macrophage purinoreceptors. Adenine nucleotides are an early component of the cascade of inflammatory mediators released by macrophages, and I propose that there are events subsequent to adenine nucleotide release that are preferentially activated by toxic preparations of LPS. Future identification of LPS-induced, macrophage-generated effectors, that are produced subsequent to adenine nucleotide release, should provide information concerning the variance in toxicity of LPS preparations. |
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| Thesis Publications |
- Denlinger LC, Fisette PL, Sommer JA, Watters JJ, Prabhu U, Proctor RA, Bertics PJ. Cutting Edge: The nucleotide receptor P2X7 contains multiple protein- and lipid-interaction motifs including a potential binding site for bacterial lipopolysaccharide. J Immunol 167:1871-1876, 2001.
- Watters JJ, Sommer JA, Fisette PL, Pfeiffer ZA, Aga M, Prabhu U, Guerra AN, Denlinger LC, Bertics PJ. The P2X7 Nucleotide Receptor: Modulation of LPS-induced macrophage signaling and mediator production. Drug Dev Res 53:91-104, 2001.
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