Background: Endotoxin (ETX) is thought to be the primary inducer of proinflammatory mediator release associated with bacterial sepsis. Furthermore, a number of studies indicate that preexposure of animals to high doses of ETX produces macrophages (Møs) that are refractory to ex vivo stimulation with ETX. However, it is unknown if levels of ETX comparable to those typically encountered in sepsis induce a similar refractory state in Møs. Design: To assess this, peritoneal Møs (PMøs) were harvested from C3H/HeN mice (ETX sensitive) at 1 hour (early) or 24 hours (late) following cecal ligation and puncture (CLP) to induce polymicrobial sepsis, sham CLP, or laparotomy followed by peritoneal implantation of a minipump delivering either saline or ETX (0.025 μg/g of body weight, every 24 hours). Peritoneal Mø cultures were incubated with ETX, either 0 or 10 μg/mL, for 24 hours, and their ability to release interleukin-1, interleukin-6, and tumor necrosis factor was assessed by bioassay. Results: Chronic low-dose ETX with 0 μg of ETX media added produced early (at 1 hour) in vivo activation of PMø interleukin-1 release, which was comparable to that seen in mice subjected to CLP. However, unlike PMøs taken from CLP mice, PMøs from mice implanted with the ETX minipump at 1 or 24 hours showed no marked decline in their ability to respond to ETX (10 μg). Comparable changes were seen for interleukin-6 and tumor necrosis factor release. Conclusions: Bacterial component(s) other than ETX per se induces the sustained dysfunction in PMø capacity to produce proinflammatory cytokines during sepsis and/or peritonitis. Thus, agents directed against ETX alone may not be adequate in the treatment of polymicrobial sepsis. (Arch Surg. 1995;130:1178-1185)