Heme oxygenase-1 (HO-1) is an intracellular enzyme that degrades heme and inhibits immune responses and inflammation in vivo. In most cell types, HO-1 is inducible by inflammatory stimuli and oxidative stress. Here we demonstrate that human monocyte-derived immature dendritic cells (iDCs) and several but not all freshly isolated rat splenic DC subsets and rat bone marrow-derived iDCs, spontaneously express HO-1. HO-1 expression drastically decreases during human and rat DC maturation induced in vitro. In IntroductionHeme oxygenases (HOs) are the rate-limiting intracellular enzymes that degrade heme to biliverdin, free divalent iron, and CO (for a review, see Otterbein and Choi 1 ). Three distinct HO enzymes have been identified: HO-1, HO-2, and HO-3. 1 HO-1 is a stress responsive gene whose expression is induced by a variety of stimuli including heme, heavy metals, inflammatory cytokines, and nitric oxide. 1 HO-1 is known for its cytoprotective effect against oxidative injuries and inflammation. 1 Induction of HO-1 expression by pharmacologic activators or gene transfer has had therapeutic effects in a variety of conditions or disorders involving the immune system, including transplantation and inflammatory disorders. [2][3][4][5][6][7][8] Biliverdin and its metabolite, bilirubin, are known for their antioxidant 9 and immunosuppressive effects. 10 HO-1 and CO have been shown to inhibit lipopolysaccharide (LPS)-induced expression of proinflammatory cytokines and to increase LPS-induced expression of interleukin 10 (IL-10) in macrophages. 11,12 Moreover, IL-10 induces HO-1 expression in macrophages. [13][14][15] We previously reported that overexpression of HO-1, obtained with an HO-1-encoding adenovirus in rats having heart transplants, results in long-term allograft survival associated with an inhibition of cellular allogeneic immune responses, which could be mediated by adenoviral transduction of dendritic cells (DCs). 6 DCs play a central role in the induction of immunity and tolerance (for a review, see Steinman et al 16 ). In the absence of inflammation, immature DCs (iDCs) located in peripheral tissues specialize in taking up innocuous and cell-associated self antigens.They continuously capture antigens and migrate to draining lymph nodes where they can induce tolerance. 16 In the presence of danger signals, DCs undergo maturation, a process involving upregulation of surface major histocompatibility complex (MHC) class II and costimulatory molecules, secretion of proinflammatory and anti-inflammatory cytokines, and the acquired ability to stimulate differentiation of naive T cells into effector cells.Our working hypothesis was that DCs can express HO-1, which can regulate DC functions. In this study, we demonstrate that human and rat iDCs express HO-1 and that HO-1 expression is down-regulated by maturation stimuli. Our results also demonstrate that induction of HO-1 expression renders DCs refractory to LPS-induced maturation, but preserves IL-10 secretion, suggesting that HO-1 may be used to regulate DC f...

Medullary thymic epithelial cells (mTECs) are specialized for inducing central immunological tolerance to self-antigens. To accomplish this, mTECs must adopt a mature phenotype characterized by expression of the autoimmune regulator Aire, which activates the transcription of numerous genes encoding tissue-restricted self-antigens. The mechanisms that control mature Aire(+) mTEC development in the postnatal thymus remain poorly understood. We demonstrate here that, although either CD4(+) or CD8(+) thymocytes are sufficient to sustain formation of a well-defined medulla, expansion of the mature mTEC population requires autoantigen-specific interactions between positively selected CD4(+) thymocytes bearing autoreactive T cell receptor (TCR) and mTECs displaying cognate self-peptide-MHC class II complexes. These interactions also involve the engagement of CD40 on mTECs by CD40L induced on the positively selected CD4(+) thymocytes. This antigen-specific TCR-MHC class II-mediated crosstalk between CD4(+) thymocytes and mTECs defines a unique checkpoint in thymic stromal development that is pivotal for generating a mature mTEC population competent for ensuring central T cell tolerance.

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To investigate the role of Aire in thymic selection, we examined the cellular requirements for generation of ovalbumin (OVA)-specific CD4 and CD8 T cells in mice expressing OVA under the control of the rat insulin promoter. Aire deficiency reduced the number of mature singlepositive OVA-specific CD4 ؉ or CD8 ؉ T cells in the thymus, independent of OVA expression. Importantly, it also contributed in 2 ways to OVA-dependent negative selection depending on the Tcell type. Aire-dependent negative selection of OVA-specific CD8 T cells correlated with Aire-regulated expression of OVA. By contrast, for OVA-specific CD4 T cells, Aire affected tolerance induction by a mechanism that operated independent of the level of OVA expression, controlling access of antigen presenting cells to medullary thymic epithelial cell (mTEC)-expressed OVA. This study supports the view that one mechanism by which Aire controls thymic negative selection is by regulating the indirect presentation of mTEC-derived antigens by thymic dendritic cells. It also indicates that mTECs can mediate tolerance by direct presentation of Aire-regulated antigens to both CD4 and CD8 T cells. (Blood. 2011;118(9): 2462-2472) IntroductionFor many years investigators noted the ectopic expression of tissue-specific antigens (TSAs) in the thymus of normal and transgenic mice 1,2 and other species. 3,4 Although this observation was a fascinating one, the true impact of thymic expression of TSAs was not fully appreciated until the generation and analysis of Aire-deficient mice. [5][6][7] These studies were prompted by the discovery of a recessive autoimmune disease in humans referred to as autoimmune polyglandular syndrome type I or autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy. 8,9 This disease is associated with circulating tissue-specific autoantibodies that contribute to the destruction of target organs, mainly endocrine glands. [10][11][12] The first symptoms that typically appear during early childhood include chronic mucocutaneous candidiasis, hypoparathyroidism, and primary adrenocortical failure. In adulthood, patients develop endocrine autoimmune diseases, such as gonadal atrophy, type 1 diabetes, hypothyroidism, and hepatitis. In addition, several ectodermic diseases may arise. 10 This disease was mapped to the AIRE locus, where several mutations have now been reported. 13 The generation of Airedeficient mice revealed 2 important observations: first, that many TSAs shown to have ectopic thymic expression were regulated by Aire and, second, that mice deficient in Aire were prone to autoimmune disease. 14 The initial report by Anderson et al 5 suggested that Aire controlled self-tolerance by enabling ectopic expression of TSAs in the medullary thymic epithelial cells (mTECs), although not all TSA genes appeared to be controlled by Aire. The ability of Aire to control thymic tolerance by regulating expression of TSAs was formally demonstrated by use of the RIP-HEL transgenic system, where the model antigen hen egg lysozyme (HEL) was expressed...