EVIDENCE FOR LIPOPOLYSACCHARIDE AS THE PREDOMINANT PROINFLAMMATORY MEDIATOR IN SUPERNATANTS OF ANTIBIOTIC-TREATED BACTERIA

Lipopolysaccharide (LPS), purified from gram-negative bacteria, is wel l known to Induce proinflammatory responses in monocytes and macrophag es, and release of LPS from the microbial surface has been suggested t o be an important initiating event in the sepsis syndrome. However, nu merous studies have documented that a variety of constituents present in the outer cell membrane of gram-negative bacteria have the capacity to activate cells of the immune system. Given that the majority of im munotherapeutic approaches designed to intervene in gram-negative seps is have to date targeted the LPS molecule, it would be of value to ass ess the relative proinflammatory properties of LPS and other gram-nega tive structures. Experiments were therefore undertaken to assess-stimu lation of human monocytes by components released from Escherichia coli following bacteriolysis by the cell wall-active antibiotic ceftazidim e. As assessed by both induction of procoagulant activity and release of tumor necrosis factor, bacterial culture supernatants contain signi ficant proinflammatory activity. When culture supernatants are fractio nated via either velocity sedimentation in sucrose gradients or isopyc nic density gradient ultracentrifugation in cesium chloride, the predo minant monocyte-stimulating activity is identified in LPS-containing f ractions. Further, such activity can be readily abrogated by the addit ion of polymyxin B. These results provide support for the hypothesis t hat LPS may be responsible for the majority of the proinflammatory act ivity released from E. coti following bacteriolysis in vitro.