Identification and biological properties of bacterial endotoxin

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Biological and biochemical prpperties of bacterial endotoxin (lipopolysaccharide, LPS) were examined.1. LPS-responsiveness of a CD14-negative cell line.LPS-responsiveness of non-macrophage ST2 cells derived from murine bone marrow stroma was examined. ST2 cells did not express CD14 mRNA.The cells as well as macrophages expressed IL-6 mRNA in response to LPS (at a minimal does of 0.1 ng/ml), but did neither TNF nor nitric oxide. Taxol, an LPS agonist, induced IL-6 mRNA expression by ST2 cells. Phodobacter sphaeroides lipid A (RsDPLA), an LPS antagonist, inhibited both LPS-and taxol-induced IL-6 mRNA expression.2. LPS-responsiveness and LPS-binding properties of ST2 cells transfected with murine CD14 cDNA.To examine whether other LPS receptor molecules than CD14 are required for CD14-dependent cell activation in macrophages, we prepared LPS-responsive ST2 and LPS-non-responsive COS cells transfected with murine CD14 cDNA expression vectors.3. CD14-independent LPS-binding in LPS-responsive cells.To clarify whether CD14-negative ST2 cells have LPS-binding sites on the cells, binding studies of radiolabeled LPS to the cells were examined. ^<125>I-LPS showed time-and dose-saturable specific binding to ST2 cells, suggesting that there exists a CD14-independent binding sites on ST2 cells.Collectively, these results strongly suggests a CD14-independent signal transduction pathway for IL-6 mRNA-expression in response to LPS.