Endotoxin (LPS) is a potent inducer of tumor necrosis factor-alpha (TNF-alpha) and manganese superoxide dismutase (MnSOD). Recent evidence suggests that LPS induction of TNF-alpha and MnSOD mRNAs is mediated through distinct intracellular signal transduction pathways. Membrane CD14 (mCD14) and Toll-like receptor-4 (TLR4) mediate LPS induction of TNF-alpha in macrophages. In the current study, we evaluated the role of mCD14 and TLR4 in LPS induction of MnSOD using peritoneal macrophages from CD14 knockout (CD14-KO) mice and mice with the Tlr4 gene point mutation (C3H/HeJ) or deletion (C57BL/10ScCr). We studied mCD14-dependent (1 and 10 ng/ml) and mCD14-independent (1,000 ng/ml) concentrations of LPS. Compared with control (BALB/c) macrophages, LPS at 1 and 10 ng/ml failed to induce TNF-alpha or MnSOD mRNA in CD14-KO macrophages. However, LPS at 1,000 ng/ml induced TNF-alpha and MnSOD mRNAs equally in macrophages from CD14-KO and control mice. LPS (1, 10, or 1,000 ng/ml) failed to induce TNF-alpha or MnSOD mRNA and failed to activate nuclear factor-kappaB in C3H/HeJ or C57BL/10ScCr macrophages. Measurements of TNF-alpha and MnSOD enzyme activity paralleled TNF-alpha and MnSOD mRNA levels. These data demonstrate that, like TNF-alpha, induction of MnSOD by LPS is mediated by mCD14 and TLR4 in murine macrophages.