Intestinal macrophages are known to display profound inflammatory anergy in response to lipopolysacchraide (LPS). To study the mechanisms of unresponsiveness of intestinal macrophages to LPS, we compared the mRNA expression of molecules associated with signal transduction of intestinal macrophages with those of other tissue macrophages. Also cellular localization of CD14 protein was examined. Intestinal, alveolar and peritoneal macrophages were isolated from rats or mice. The expression of mRNA was assessed by real-time PCR, and cellular localization of CD14 protein was examined by flow cytometry. Cellular responses to LPS were examined by production of TNF and NO. The expression of CD14 mRNA in intestinal macrophages was lower than for peritoneal macrophages but higher than for alveolar macrophages. The mRNA expression of other molecules corresponding to intracellular signal transduction in intestinal macrophages was similar with alveolar and peritoneal macrophages. Despite the presence of CD14 mRNA, proteins of CD14 were not detected on cell surfaces of intestinal macrophages, and induction of TNF or NO responding to LPS were not detected. Flow cytometric analysis demonstrated that CD14 protein was not expressed on the cell surface but was expressed inside intestinal macrophages. The unresponsiveness of intestinal macrophages after LPS exposure is considered to be largely attributed to the lack of CD14 protein on their cell surfaces. However, CD14 protein was expressed inside of the cells, suggesting that post-transcriptional regulation rather than transcriptional suppression may play a dominant role in determining the phenotype of the intestinal macrophages.