Ikaros limits basophil development by suppressing C/EBP-α expression.

The Ikaros gene (Ikzf1) encodes a family of zinc-finger transcription factors implicated in hematopoietic cell differentiation. Here we show that Ikaros suppresses the development of basophils, which are proinflammatory cells of the myeloid lineage. In the absence of extrinsic basophil-inducing signals, Ikaros(-/-) (Ik(-/-)) mice exhibit increases in basophil numbers in blood and bone marrow and in their direct precursors in bone marrow and the spleen, as well as decreased numbers of intestinal mast cells. In vitro culture of Ik(-/-) bone marrow under mast cell differentiation conditions also results in predominance of basophils. Basophil expansion is associated with an increase in basophil progenitors, increased expression of Cebpa and decreased expression of mast cell-specifying genes Hes1 and microphthalmia-associated transcription factor (Mitf). Ikaros directly associates with regulatory sites within Cebpa and Hes1 and regulates the acquisition of permissive H3K4 tri-methylation marks at the Cebpa locus and reduces H3K4 tri-methylation at the Hes1 promoter. Ikaros blockade in cultured cells or transfer of Ik(-/-) bone marrow into irradiated Ik(+/+) recipients also results in increased basophils confirming a cell-intrinsic effect of Ikaros on basophil development. We conclude that Ikaros is a suppressor of basophil differentiation under steady-state conditions and that it acts by regulating permissive chromatin modifications of Cebpa.

A cell permeable peptide inhibitor of NFAT inhibits macrophage cytokine expression and ameliorates experimental colitis.

Nuclear factor of activated T cells (NFAT) plays a critical role in the development and function of immune and non-immune cells. Although NFAT is a central transcriptional regulator of T cell cytokines, its role in macrophage specific gene expression is less defined. Previous work from our group demonstrated that NFAT regulates Il12b gene expression in macrophages. Here, we further investigate NFAT function in murine macrophages and determined the effects of a cell permeable NFAT inhibitor peptide 11R-VIVIT on experimental colitis in mice. Treatment of bone marrow derived macrophages (BMDMs) with tacrolimus or 11R-VIVIT significantly inhibited LPS and LPS plus IFN-γ induced IL-12 p40 mRNA and protein expression. IL-12 p70 and IL-23 secretion were also decreased. NFAT nuclear translocation and binding to the IL-12 p40 promoter was reduced by NFAT inhibition. Experiments in BMDMs from IL-10 deficient (Il10(-/-)) mice demonstrate that inhibition of IL-12 expression by 11R-VIVIT was independent of IL-10 expression. To test its therapeutic potential, 11R-VIVIT was administered systemically to Il10(-/-) mice with piroxicam-induced colitis. 11R-VIVIT treated mice demonstrated significant improvement in colitis compared to mice treated with an inactive peptide. Moreover, decreased spontaneous secretion of IL-12 p40 and TNF in supernatants from colon explant cultures was demonstrated. In summary, NFAT, widely recognized for its role in T cell biology, also regulates important innate inflammatory pathways in macrophages. Selective blocking of NFAT via a cell permeable inhibitory peptide is a promising therapeutic strategy for the treatment of inflammatory bowel diseases.

Altered macrophage function contributes to colitis in mice defective in the phosphoinositide-3 kinase subunit p110δ.

BACKGROUND & AIMS: Innate immune responses are crucial for host defense against pathogens but need to be tightly regulated to prevent chronic inflammation. Initial characterization of mice with a targeted inactivating mutation in the p110δ subunit of phosphoinositide 3-kinase (PI3K p110δ(D910A/D910A)) revealed defects in B- and T-cell signaling and chronic colitis. Here, we further characterize features of inflammatory bowel diseases in these mice and investigate underlying innate immune defects. METHODS: Colons and macrophages from PI3K p110δ(D910A/D910A) mice were evaluated for colonic inflammation and innate immune dysfunction. Colonic p110δ messenger RNA expression was examined in interleukin (IL)-10(-/-) and wild-type germ-free mice during transition to a conventional microbiota. To assess polygenic impact on development of colitis, p110δ(D910A/D910A) mice were backcrossed to IL-10(-/-) mice. RESULTS: A mild spontaneous colitis was shown in PI3K p110δ(D910A/D910A) mice at 8 weeks, with inflammation increasing with age. An inflammatory mucosal and systemic cytokine profile was characterized by expression of IL-12/23. In PI3K p110δ(D910A/D910A) macrophages, augmented toll-like receptor signaling and defective bactericidal activity were observed. Consistent with an important homeostatic role for PI3K p110δ, wild-type mice raised in a germ-free environment markedly up-regulated colonic PI3K p110δ expression with the introduction of the enteric microbiota; however, colitis-prone IL-10(-/-) mice did not. Moreover, PI3K p110δ(D910A/D910A) mice crossed to IL-10(-/-) mice developed severe colitis at an early age. CONCLUSIONS: This study describes a novel model of experimental colitis that highlights the importance of PI3K p110δ in maintaining mucosal homeostasis and could provide insight into the pathogenesis of human inflammatory bowel disease.

Mast cells: multifaceted immune cells with diverse roles in health and disease.

Mast cells were discovered more than 100 years ago and until recently, have been considered renegades of the host with the sole purpose of perpetuating allergy. The discovery of mast cell-deficient mice that could be reconstituted with mast cells (the so called "mast cell knock-in" mice) has allowed the study of the in vivo functions of mast cells and revealed several new facets of these cells. It is now evident that mast cells have a much broader impact on many physiological and pathologic processes. Mast cells, particularly through their dynamic interaction with the nervous system, have been implicated in wound healing, tissue remodeling, and homeostasis. Perhaps the most progress has been made in our understanding of the role of mast cells in immunity outside the realm of allergy, and host defense. Mast cells play critical roles in both innate and adaptive immunity, including immune tolerance. Greater insight into mast cell biology has prompted studies probing the additional consequences of mast cell dysfunction, which reveal a central role for mast cells in the pathogenesis of autoimmune disorders, cardiovascular disorders, and cancer. Here, we review recent developments in the study of mast cells, which present a complex picture of mast cell functions.

Carbon monoxide ameliorates chronic murine colitis through a heme oxygenase 1-dependent pathway.

Heme oxygenase (HO)-1 and its metabolic product carbon monoxide (CO) play regulatory roles in acute inflammatory states. In this study, we demonstrate that CO administration is effective as a therapeutic modality in mice with established chronic colitis. CO administration ameliorates chronic intestinal inflammation in a T helper (Th)1-mediated model of murine colitis, interleukin (IL)-10-deficient (IL-10(-/-)) mice. In Th1-mediated inflammation, CO abrogates the synergistic effect of interferon (IFN)-gamma on lipopolysaccharide-induced IL-12 p40 in murine macrophages and alters IFN-gamma signaling by inhibiting a member of the IFN regulatory factor (IRF) family of transcription factors, IRF-8. A specific signaling pathway, not previously identified, is delineated that involves an obligatory role for HO-1 induction in the protection afforded by CO. Moreover, CO antagonizes the inhibitory effect of IFN-gamma on HO-1 expression in macrophages. In macrophages and in Th1-mediated colitis, pharmacologic induction of HO-1 recapitulates the immunosuppressive effects of CO. In conclusion, this study begins to elucidate potential etiologic and therapeutic implications of CO and the HO-1 pathway in chronic inflammatory bowel diseases.