Endotoxin tolerance in macrophages is a key regulatory mechanism to limit the innate immune response to infection or injury. Long considered a state of unresponsiveness to Toll-like receptor activation, tolerance is now recognized as a state of altered responsiveness to infection or injury. Endotoxin tolerance leads to a shift away from a pro-inflammatory response toward a response with key anti-inflammatory and pro-resolution features. Advances in our understanding of Toll-like receptor function have identified a number of molecular mechanisms that promote tolerance, but how these are integrated to achieve gene-specific regulation is an important outstanding question. The potential to harness the mechanisms of endotoxin tolerance to promote the resolution of chronic inflammation warrants the continued investigation of this fundamental feature of innate immunity. This review focuses on the endotoxin tolerant state, our understanding of the underlying molecular mechanisms, and the clinical significance of endotoxin tolerance.