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Macrophages participate in physiologic and pathologic processes through elaboration of distinct activation programs. Studies with macrophage cell systems have revealed much concerning the importance of this pleiotropic cell; however, these studies are inherently limited by three factors: heterogeneity of the target cell population, poor capacity to elaborate various activation programs, and lack of a genetically tractable model system for loss- and gain-of-function studies. Although definitive, hematopoietic lineages can be isolated from embryonic stem (ES) cells, these isolation procedures are inefficient and time-consuming and require elaborate cell-sorting protocols. We therefore examined whether myeloid precursors, capable of differentiating into macrophages, could be conditionally expanded in vitro. Here, we report methods for selective isolation and immortalization of ES cell-derived myeloid precursors by estrogen-regulated HoxA9 protein. Using this new macrophage differentiation system, an unlimited number of custom-designed macrophages with defined functional characteristics can be generated from any targeted ES cell. In combination with knockout or small interfering RNA knockdown technologies, this macrophage differentiation system provides a powerful tool for high throughput analysis of regulatory mechanisms controlling macrophage activation in health and disease.

Complex interplay between T helper (Th) cells and macrophages contributes to the formation and progression of atherosclerotic plaques. While Th1 cytokines promote inflammatory activation of lesion macrophages, Th2 cytokines attenuate macrophage-mediated inflammation and enhance their repair functions. In spite of its biologic importance, the biochemical and molecular basis of how Th2 cytokines promote maturation of anti-inflammatory macrophages is not understood. We show here that in response to interleukin-4 (IL-4), signal transducer and activator of transcription 6 (STAT6) and PPARgamma-coactivator-1beta (PGC-1beta) induce macrophage programs for fatty acid oxidation and mitochondrial biogenesis. Transgenic expression of PGC-1beta primes macrophages for alternative activation and strongly inhibits proinflammatory cytokine production, whereas inhibition of oxidative metabolism or RNAi-mediated knockdown of PGC-1beta attenuates this immune response. These data elucidate a molecular pathway that directly links mitochondrial oxidative metabolism to the anti-inflammatory program of macrophage activation, suggesting a potential role for metabolic therapies in treating atherogenic inflammation.