Dagmar von Bubnoff

Systemic application of CpG-rich DNA suppresses adaptive T cell immunity via induction of IDO

CpG‐rich oligonucleotides (CpG‐ODN) bind to Toll‐like receptor 9 (TLR9) and are used as powerful adjuvants for vaccination. Here we report that CpG‐ODN not only act as immune stimulatory agents but can also induce strong immune suppression depending on the anatomical location of application. In agreement with the adjuvant effect, subcutaneous application of antigen plus CpG‐ODN resulted in antigen‐specific T cell activation in local lymph nodes. In contrast, systemic application of CpG‐ODN resulted in suppression of T cell expansion and CTL activity in the spleen. The suppressive effect was mediated by indoleamine 2,3‐dioxygenase (IDO) as indicated by the observation that CpG‐ODN induced IDO in the spleen and that T cell suppression could be abrogated by 1‐methyl‐tryptophan (1‐MT), an inhibitor of IDO. No expression of IDO was observed in lymph nodes after injection of CpG‐ODN, explaining why suppression was restricted to the spleen. Studies with a set of knockout mice demonstrated that the CpG‐ODN‐induced immune suppression is dependent on TLR9 stimulation and independent of type I and type II interferons. The present study shows that for the use of CpG‐ODN as an adjuvant in vaccines, the route of application is crucial and needs to be considered. In addition, the results indicate that down‐modulation of immune responses by CpG‐ODN may be possible in certain pathological conditions.

FcεRI Induces the Tryptophan Degradation Pathway Involved in Regulating T Cell Responses

FcεRI is suspected to play a pivotal role in the pathophysiology of atopic disorders such as atopic dermatitis. In search for genes differentially regulated by FcεRI on APCs, a differential cDNA bank of receptor-stimulated and unstimulated monocytes was established. By means of suppression subtractive hybridization, we identified kynurenine 3-monooxygenase and subsequently indoleamine 2,3-dioxygenase (IDO) to be overexpressed in FcεRI-activated monocytes. IDO is the rate-limiting enzyme in the catabolism of the essential amino acid tryptophan. We show that cross-linking of FcεRI on monocytes results in low tryptophan concentrations associated with impaired T cell stimulatory capacity. Importantly, T cell suppression could be prevented by the addition of tryptophan or inhibition of IDO. Moreover, stimulation of T cells by FcεRI-activated monocytes was increased compared with T cell stimulation by nonactivated monocytes if exogenous supply of tryptophan was available. We speculate that the expression of IDO by FcεRI+ APCs in vivo allows these cells to regulate T cell responses in atopic disorders by inhibiting or stimulating T cell proliferation, depending on the metabolic environment.

Tryptophan Deprivation Induces Inhibitory Receptors ILT3 and ILT4 on Dendritic Cells Favoring the Induction of Human CD4+CD25+ Foxp3+ T Regulatory Cells

Tryptophan catabolism through IDO activity can cause nonresponsiveness and tolerance acting on T cells. Given the crucial importance of dendritic cells (DCs) in the initiation of a T cell response, surprisingly little is known about the impact of IDO activity and tryptophan deprivation on DCs themselves. In the present study, we show that human DCs differentiated under low-tryptophan conditions acquire strong tolerogenic capacity. This effect is associated with a markedly decreased Ag uptake as well as the down-regulation of costimulatory molecules (CD40, CD80). In contrast, the inhibitory receptors ILT3 and ILT4 are significantly increased. Functionally, tryptophan-deprived DCs show a reduced capacity to stimulate T cells, which can be restored by blockade of ILT3. Moreover, ILT3highILT4high DCs lead to the induction of CD4+CD25+ Foxp3+ T regulatory cells with suppressive activity from CD4+CD25− T cells. The generation of ILT3highILT4high DCs with tolerogenic properties by tryptophan deprivation is linked to a stress response pathway mediated by the GCN2 kinase. These results demonstrate that tryptophan degradation establishes a regulatory microenvironment for DCs, enabling these cells to induce T regulatory cells. The impact of IDO thus extends beyond local immune suppression to a systemic control of the immune response.

Natural killer cells in atopic and autoimmune diseases of the skin

Natural killer (NK) cells are best known for their ability to recognize and kill tumor cells and virally infected cells and for their ability to produce large amounts of some cytokines, such as IFN-gamma. Recent research has substantially expanded our view on the function of NK cells in the immune system in health and disease. In addition to the better-studied functions in cancer and autoimmunity, contributions from NK cells to allergies and various skin diseases have emerged. We briefly recount the traditional NK cell functions before focusing on their roles in atopic dermatitis, psoriasis, alopecia areata, and pemphigus vulgaris. Although this field is still developing, strong data are available that indicate NK cell involvement. In patients with allergic diseases, the production of T(H)2 cytokines by NK cells contributes to the known immune deviation. In patients with psoriasis, their pathophysiologic role seems to be especially the production of IFN-gamma. NK cell overactivation can be found in patients with alopecia areata and pemphigus vulgaris. Many details are still unclear; however, we believe that there is solid evidence that NK cells actively participate in a number of diseases that have not been traditionally linked to this type of lymphocyte.

Identification of IDO-Positive and IDO-Negative Human Dendritic Cells after Activation by Various Proinflammatory Stimuli

Dendritic cells (DCs) can induce tolerance or immunity. We identified and characterized an IDO-expressing and an IDO-negative human DC population after stimulation by various proinflammatory stimuli. IDO expression was strongly dependent on the maturation status of the cells (CD83-positive cells only). The two DC subpopulations remained IDO positive and IDO negative, respectively, over a time period of at least 48 h. IDO enzyme activity of human DCs was highest during stimulation by strongly maturation-inducing TLR ligands such as highly purified LPS (TLR4 ligand) or polyriboinosinic-polyribocytidilic acid (TLR3 ligand); factors of the adaptive immune system such as IFN-γ, a mixture of cytokines, and IFN-α had lesser stimulatory capacity for IDO induction and activity. After stimulation with CD40L, IDO-positive DCs expressed significantly increased levels of B7 family molecules such as CD40, CD80, CD86, ICOS ligand, as well as PD-L1 (B7-H1) and PD-L2 (B7-DC) compared with the IDO-negative DC subset. At the same time, the inhibitory receptors Ig-like transcripts 3 and 4 were significantly downregulated on IDO-positive cells. Functionally, IDO-positive DCs produced significantly more IL-1β and IL-15 and less IL-10 and IL-6 than the IDO-negative subset after CD40L stimulation. These results show that IDO expression is associated with a distinctive phenotype and functional capacity in mature DCs. It seems likely that the IDO-positive DC subset possesses a regulatory function and might skew a T cell response toward tolerance.

Indoleamine 2,3-dioxygenase–expressing antigen-presenting cells and peripheral T-cell tolerance: Another piece to the atopic puzzle?

There is growing evidence that dendritic cells, the major antigen-presenting cells and T-cell activators, have a broad effect on peripheral T-cell tolerance and regulation of immunity. Very recently, a new feature of regulatory antigen-presenting cells was observed. Certain dendritic cells, monocytes, and macrophages express the enzyme indoleamine 2,3-dioxygenase, and thus because of enhanced degradation of the essential amino acid tryptophan, they modulate T-cell activity in specific local tissue environments. In this review we discuss the various and apparently disparate effects of indoleamine 2,3-dioxygenase induction in cells of the immune system. We place current knowledge about this mechanism in the context of atopy. We introduce the hypothesis that tryptophan degradation might add to the ability to control and downregulate allergen-specific T-cell responses in atopic individuals.

Indoleamine 2,3-dioxygenase–expressing myeloid dendritic cells and macrophages in infectious and noninfectious cutaneous granulomas

BACKGROUND The enzyme indoleamine 2,3-dioxygenase (IDO) degrades the essential amino acid tryptophan, and this degradation is an immunosuppressive mechanism that is mainly used by antigen-presenting cells. IDO-expressing dendritic cells and macrophages have previously been identified as components of lymph node granulomas after Listeria monocytogenes infection. In this study we undertook an analysis of IDO expression in granulomas of infectious and noninfectious origin in the human skin. METHODS Lesional skin biopsy specimens (n = 22) from different granulomatous skin disorders (lupus vulgaris, sarcoidosis, granuloma annulare, leprosy) were analyzed. Immunohistochemistry was performed to identify and locate the enzyme IDO within the inflammatory granulomatous infiltrate (IDO, CD11c, CD68, S100, CD3, Foxp3). Two-color immunofluorescence of IDO in combination with multiple markers was applied to characterize the IDO-expressing cells. RESULTS Cutaneous granulomas of different origin strongly express IDO, mainly in the center and in the ring wall of the granulomas. We demonstrate that in infectious, but also in noninfectious human cutaneous granulomas the large myeloid CD11c(+)S100(+)CD68(-) dendritic cells and the CD68(+) macrophages express IDO. LIMITATIONS This study was limited by the lack of details about the exact stage or maturity of granuloma formation in the specimens investigated. CONCLUSION These findings reveal that IDO expression in myeloid dendritic cells and macrophages is part of an integrated response of granuloma formation, which may be a unifying feature of granulomatous reactions in the skin.

Kinetics of Gene Induction After FcεRI Ligation of Atopic Monocytes Identified by Suppression Subtractive Hybridization

The high-affinity receptor for IgE, FcεRI, on APCs plays an important role in the initiation and chronicity of inflammatory atopic diseases. To understand the molecular regulation of FcεRI-mediated processes, differentially expressed genes are of great interest to be identified. Suppression subtractive cDNA hybridization has been used to identify genes induced after FcεRI stimulation on atopic monocytes. Overexpression of the identified genes was determined by semiquantitative RT-PCR analysis of transcripts from the tester (stimulated) and driver (unstimulated) monocytes. Results were confirmed and kinetics of the transcripts established using blood cells from additional atopics at 4 and 24 h of FcεRI induction. The following sequences were identified: monocyte chemoattractant protein 1, macrophage-inflammatory protein 1β, IL-6, βA subunit of inhibin/activin, IFN-stimulated gene of 54 kDa, IL-1R antagonist, and kynurenine 3-monooxygenase. Chemokines are highly expressed during the early and late phase after FcεRI cross-linking, whereas proinflammatory and differentiation stimuli rapidly decline after an initial overexpression. Kynurenine 3-monooxygenase, an enzyme involved in the degradation of the amino acid tryptophan, is significantly up-regulated during the late phase after 24 h of FcεRI induction. These results demonstrate that the analysis of the profile of gene induction following activation of FcεRI on atopic monocytes may reveal how these cells might participate in the regulation of atopic disorders.

Transcriptional profiling identifies an interferon‐associated host immune response in invasive squamous cell carcinoma of the skin

Squamous cell carcinoma (SCC) and basal cell carcinoma (BCC) represent the 2 most common types of nonmelanoma skin cancer. Both derive from keratinocytes but show a distinct biological behavior. Here we present transcriptional profiling data of a large cohort of tumor patients (SCC, n = 42; BCC, n = 114). Differentially expressed genes reflect known features of SCC and BCC including the typical cytokeratin pattern as well as upregulation of characteristic cell proliferation genes. Additionally, we found increased expression of interferon (IFN)‐regulated genes (including IFI27, IFI30, Mx1, IRF1 and CXCL9) in SCC, and to a lower extent in BCC. The expression of IFN‐regulated genes correlated with the extent of the lesional immune‐cell infiltrate. Immunohistological examinations confirmed the expression of IFN‐regulated genes in association with a CXCR3+ cytotoxic inflammatory infiltrate on the protein level. Of note, a small subset of SCC samples with low expression of IFN‐regulated genes included most organ transplant recipients receiving immunosuppressive medication. Collectively, our findings support the concept that IFN‐associated host responses play an important role in tumor immunosurveillance in the skin.

Primary immunodeficiency in combination with transverse upper limb defect and anal atresia in a 34‐year‐old patient with Jacobsen syndrome

We describe a 34‐year‐old male patient with Jacobsen syndrome associated with a broad spectrum of anomalies and an increased susceptibility to infections. Features commonly seen in Jacobsen syndrome were short stature, mental retardation, congenital heart disease, cryptorchidism, strabismus, distal hypospadia glandis, and mild thrombocytopenia. Chromosome analysis disclosed a mosaic 46,XY,del(11)(q24.1)/46,XY karyotype with a very low percentage of normal cells. In addition, transverse upper limb defect, imperforate anus, and hearing impairment were noted. Cellular anomalies include functional impairment and deficiency of T‐helper cells, and a low serum immunoglobulin M (IgM)‐level. The presence of a transverse limb defect and primary immunodeficiency has not been reported previously in Jacobsen syndrome.