Ig-like Transcript Research Articles

Ig-like Transcript 7, but Not Bone Marrow Stromal Cell Antigen 2 (Also Known as HM1.24, Tetherin, or CD317), Modulates Plasmacytoid Dendritic Cell Function in Primary Human Blood Leukocytes

The Ig-like transcript (ILT) 7 is a surface molecule selectively expressed by human plasmacytoid dendritic cells (pDCs). ILT7 cross-linking suppresses pDC activation and type I IFN (IFN-I) secretion following TLR7/9 engagement. The bone marrow stromal cell Ag 2 (BST2, aka HM1.24, tetherin, or CD317) is expressed by different cell types upon exposure to IFN-I and is a natural ligand for ILT7. In this study, we show that ILT7 expression decreased spontaneously in pDCs upon in vitro culture, which correlates with pDC differentiation measured as increased side scatter properties and CCR7 expression. TLR7/9 ligands, as well as HIV, induced BST2 upregulation on all tested cell types except T cells, which required TCR stimulation to respond to TLR9L-induced IFN-I. IFN-γ, IL-4, IL-10, and TNF-α had only marginal effects on BST2 expression in blood leukocytes compared with TLR9L. Preincubation with ILT7 cross-linking Ab inhibited IFN-I production in PBMCs treated with TLR7/9L or HIV, whereas BST2 blockade did not affect IFN-I responses even when BST2 upregulation was further boosted with TCR agonists or immunoregulatory cytokines. Our data indicate that BST2-mediated ILT7 cross-linking may act as a homeostatic regulatory mechanism on immature circulating pDC, rather than a negative feedback for activated mature pDCs that have downregulated ILT7.

Immunoglobulin-like transcript receptors on human dermal CD14+dendritic cells act as a CD8-antagonist to control cytotoxic T cell priming

Human Langerhans cells (LCs) are highly efficient at priming cytolytic CD8(+) T cells compared with dermal CD14(+) dendritic cells (DCs). Here we show that dermal CD14(+) DCs instead prime a fraction of naïve CD8(+) T cells into cells sharing the properties of type 2 cytokine-secreting CD8(+) T cells (TC2). Differential expression of the CD8-antagonist receptors on dermal CD14(+) DCs, the Ig-like transcript (ILT) inhibitory receptors, explains the difference between the two types of DCs. Inhibition of CD8 function on LCs inhibited cytotoxic T lymphocytes (CTLs) and enhanced TC2 generation. In addition, blocking ILT2 or ILT4 on dermal CD14(+) DCs enhanced the generation of CTLs and inhibited TC2 cytokine production. Lastly, addition of soluble ILT2 and ILT4 receptors inhibited CTL priming by LCs. Thus, ILT receptor expression explains the polarization of CD8(+) T-cell responses by LCs vs. dermal CD14(+) DCs.

A novel role of CD1c in regulating CD1d-mediated NKT cell recognition by competitive binding to Ig-like transcript 4.

Humans express four MHC-like CD1 molecules CD1a, b, c and d that are capable of presenting a wide variety of self or foreign lipid antigens to T cells. Much progress has been made in elucidating the function of CD1d-restricted NKT cells in both innate and adaptive immune responses. However, knowledge of the other CD1 molecules is less well defined in terms of lipid presentation and immune regulation. We have previously shown that immunoglobulin-like transcript 4 (ILT4) binds to CD1d and inhibits its recognition by NKT cells. In this study, we show that CD1c can also interact specifically with ILT4 with a higher affinity than that of CD1d. Furthermore, changes in CD1c expression seem to modulate CD1d function; up-regulation of CD1c enhances NKT recognition of CD1d and down-regulation reduces CD1d recognition. We propose that CD1c can act as a sink for the inhibitory receptor ILT4: when CD1c is up-regulated, ILT4 is recruited to CD1c, thus reducing the inhibitory effect of ILT4 on CD1d recognition. Consequently, CD1c could be a potential target for modulating NKT activity. NKT, CD1d, CD1c, ILT4, antigen presentation.

Downregulation of Inflammatory MicroRNAs by Ig-like Transcript 3 Is Essential for the Differentiation of Human CD8+ T Suppressor Cells

We have investigated the mechanism underlying the immunoregulatory function of membrane Ig-like transcript 3 (ILT3) and soluble ILT3Fc. microRNA (miRNA) expression profile identified genes that were downregulated in ILT3-induced human CD8(+) T suppressor cells (Ts) while upregulated in T cells primed in the absence of ILT3. We found that miR-21, miR-30b, and miR-155 target the 3'-untranslated region of genes whose expression was strongly increased in ILT3Fc-induced Ts, such as dual specificity phosphatase 10, B cell CLL/lymphoma 6, and suppressor of cytokine signaling 1, respectively. Transfection of miRNA mimics or inhibitors and site-specific mutagenesis of their 3'-untranslated region binding sites indicated that B cell CLL/lymphoma 6, dual specificity phosphatase 10, and suppressor of cytokine signaling 1 are direct targets of miR-30b, miR-21, and miR-155. Primed CD8(+) T cells transfected with miR-21&30b, miR-21&155, or miR-21&30b&155 inhibitors displayed suppressor activity when added to autologous CD3-triggered CD4 T cells. Luciferase reporter assays of miR-21 and miR-155 indicated that their transcription is highly dependent on AP-1. Analysis of activated T cells showed that ILT3Fc inhibited the translocation to the nucleus of the AP-1 subunits, FOSB and c-FOS, and the phosphorylation of ZAP70 and phospholipase C-γ 1. In conclusion, ILT3Fc inhibits T cell activation and induces the generation of Ts targeting multiple inflammatory miRNA pathways.

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.

LILRA3 Binds Both Classical and Non-Classical HLA Class I Molecules but with Reduced Affinities Compared to LILRB1/LILRB2: Structural Evidence

Structurally, Group 1 LILR (Leukocyte Immunogloblin (Ig)-Like Receptor, also known as Ig-like transcripts, ILT; Leukocyte Ig-like receptor, LIR; and CD85) members are very similar in terms of the HLAIs (human leukocyte antigen class I molecules) binding region and were hypothesized that they all bind to HLAIs. As one of the Group 1 LILRs, LILRA3 is the only secretory LILR and may greatly control the inhibitory immune response induced by LILRB1, LILRB2, and other HLA-binding LILR molecules like LILRA1. Nevertheless, little was known about the binding of LILRA3 to HLAIs. In this report, we present the crystal structure of the LILRA3 domain 1 (D1) and evaluate the D1 and D1D2 (domain 1 and domain 2) binding to classical and non-classical HLAIs using BIAcore® surface plasmon resonance analysis (SPR). We found that LILRA3 binds both classical HLA-A*0201 and non-classical HLA-G1 but with reduced affinities compared to either LILRB1 or LILRB2. The polymorphic amino acids and the LILRA3 D1 structure support this notion.

Identification of a New Phenotype of Tolerogenic Human Dendritic Cells Induced by Fungal Proteases from Aspergillus oryzae

We characterized a new pathway to induce tolerogenic dendritic cells (DCs) following treatment of human monocyte-derived DCs with proteases from the fungus Aspergillus oryzae (ASP). ASP-treated DCs (ASP-DCs) exhibit a CD80(-)CD83(-)CD86(-)Ig-like transcript (ILT)2(-)ILT3(-)ILT4(+) phenotype, do not secrete cytokines or chemokines, and express tolerogenic markers such as glucocorticoid-induced leucine zipper, NO synthetase-2, retinaldehyde dehydrogenase-1 or retinaldehyde dehydrogenase-2. When cocultured with naive CD4(+) T cells, ASP-DCs induce an anergic state that can be reversed by IL-2. Generated T cells mediate a suppressive activity in third-party experiments that is not mediated by soluble factors. A comparison between dexamethasone-treated DCs used as a reference for regulatory T cell-inducing DCs and ASP-DCs reveals two distinct phenotypes. In contrast to dexamethasone, ASP treatment induces glucocorticoid-induced leucine zipper independently of glucocorticoid receptor engagement and leads to NF-κB p65 degradation. Abrogation of protease activities in ASP using specific inhibitors reveals that aspartic acid-containing proteases are key inducers of regulatory genes, whereas serine, cysteine, and metalloproteases contribute to NF-κB p65 degradation. Collectively, those features correspond to a previously unreported anergizing phenotype for human DCs. Such regulatory mechanisms may allow fungi to downregulate host immune responses and provide clues for new approaches to treat proinflammatory disorders.

Proper Regrafting of Ig-Like Transcript 2 after Trogocytosis Allows a Functional Cell–Cell Transfer of Sensitivity

The acquisition by T cells of exogenous ligands originally expressed by APC has been already described. However, reports essentially focused on the outward signaling of acquired ligands and their effects on surroundings cells. We investigated the function of transferred receptors (not ligands) on the T cells that acquired them (not on cells they interact with). We show that inhibitory Ig-like transcript 2 receptors efficiently transfer from monocytes to autologous T cells by trogocytosis and integrate within the plasma membrane of the acquirer T cells. Furthermore, the acquired receptors can access compatible signaling machinery within acquirer T cells and use it to signal and alter the functions of their new host cells. These data are a formal demonstration that a transferred molecule may send signals to its new host cell. We also provide evidence that sensitivity to modulatory molecules can be acquired from other cells and introduce the notion of intercellular transfer of sensitivities.

Elevated HLA-G Expression On Myeloid Dendritic Cells Correlates With Regulatory T Cell Foxp3 Expression In Operationally Tolerant Pediatric Liver Transplant Recipients

Introduction: Human leukocyte antigen (HLA)-G is a non-classical HLA class I molecule expressed as membrane-bound and soluble isoforms. Interaction of HLA-G with its receptor, Ig-like transcript (ILT)4 on professional antigen (Ag)-presenting dendritic cells (DC) down-regulates their T cell costimulatory ability. We examined the expression of HLA-G and ILT4, together with other immune regulatory molecules, on circulating DC subsets in three groups of clinically stable, pediatric liver recipients: operationally tolerant (TOL: 26); prospective immunosuppressive drug weaning (PW: 28) and maintenance immunosuppression (MI: 24), and in healthy controls (HC: 28). Methods: PBMC were stained with a lineage monoclonal antibody (mAb) cocktail, andwith mAbs, directed against blood DC Ag (BDCA)-1 (CD1; monocytoid [m]DC),and BDCA-2 (CD303; plasmacytoid [p]DC), as well as with mAbs against HLA-G and ILT4 (CD85d) or CMRF-44, inducible costimulatory ligand (ICOSL) or glucocorticoid-induced tumor necrosis factor-related protein ligand (GITRL), then analyzed by flow cytometry. Serum HLA-G levels were determined by ELISA. Results: Both circulating mDC and pDC in HC expressed surface HLA-G and ILT4. The incidence of mDC expressing HLA-G (but not ILT4) was significantly higher than that of pDC, although fluorescence intensity (MFI) was lower for HLA-G and ILT4 on mDC, compared with pDC. In TOL patients, mDC but not pDC expressed higher HLA-G (both% positive cells and MFI) than in PW or MI patients or HC. By contrast, ILT4, CMRF-44, GITRL or ICOSL expression on either pDC or mDC did not differ significantly between the groups. In TOL patients, the incidence of regulatory T cells (Treg) and the intensity of Foxp3 expression by Treg were significantly higher than in MI or HC groups. In addition, the intensity of HLA-G expression on circulating mDC correlated significantly with that of Foxp3 in the TOL group. The incidence of patients with circulating HLA-G levels>100pg/ml was greatest in the TOL group. Conclusion: Higher HLA-G expression on circulating mDC,- the predominant DC subset, in TOL pediatric liver transplant recipients compared with MI or HC, suggests a possible role of HLA-G in immune regulation. This may be correlated with enhanced Foxp3 expression by host Treg. The findings also suggest the potential of HLA-G expression on mDC as a biomarker of operational pediatric liver transplant tolerance.

BCL6 Is Required for Differentiation of Ig-Like Transcript 3-Fc–Induced CD8+ T Suppressor Cells

Ig-like transcript 3 (ILT3) is an inhibitory receptor expressed by tolerogenic dendritic cells. When human CD8(+) T cells are allostimulated in the presence of recombinant ILT3-Fc protein, they differentiate into antigenic specific T suppressor (Ts) cells that inhibit CD4 and CD8 T cell effector function both in vitro and in vivo. ILT3-Fc-induced CD8(+) Ts cells express high amounts of BCL6 that are crucial to their function. Knockdown of BCL6 from unprimed human T cells prevents their differentiation into Ts cells, whereas ex vivo overexpression of BCL6 converts CD8(+) T cells into Ts cells. NOD/SCID mice transplanted with human pancreatic islets and humanized by injection of human PBMCs tolerate the graft and develop BCL6(high) CD8(+) Ts cells when treated with ILT3-Fc before or after the onset of rejection. This indicates that ILT3-Fc acts through BCL6 and is a potent immunosuppressive agent for reversing the onset of allo- or possibly autoimmune attacks against pancreatic islets.

Granulocyte‐colony stimulating factor drives the in vitro differentiation of human dendritic cells that induce anergy in naïve T cells

G-CSF is a modulator of T-cell and DC functions. Previous reports show that monocytes from G-CSF-treated (post-G) healthy donors differentiate into tolerogenic DC in vitro in the presence of autologous serum, containing high levels of IL-10 and IFN-α, and in turn induce type 1 Treg (Tr1) cells. However, the direct effect of G-CSF on DC differentiation was not investigated. Here, we show that monocytes differentiated in the presence of exogenous G-CSF (G-DC) remain CD14+CD1a−, but acquire a DC-like morphology, express CD83 and CD86 and low levels of the tolerogenic markers Ig-like transcript (ILT)4 and HLA-G. G-DC spontaneously produce IL-10 and, upon stimulation, low levels of IL-12. G-DC display low stimulatory capacity and induce anergy in naïve T cells, but do not confer suppressive function. Therefore, in vitro differentiation of monocyte-derived DC in the presence of G-CSF can replicate some but not all features of post-G DC. These findings indicate that the tolerogenic properties of G-CSF do not exclusively reside in its direct effect on DC, which in turn induce T-cell anergy, but also in its ability to generate a tolerogenic milieu in vivo, which is necessary for Tr1 cell induction and cannot be replicated in vitro.

Differentiation of type 1 T regulatory cells (Tr1) by tolerogenic DC-10 requires the IL-10–dependent ILT4/HLA-G pathway

AbstractType 1 T regulatory (Tr1) cells suppress immune responses in vivo and in vitro and play a key role in maintaining tolerance to self- and non–self-antigens. Interleukin-10 (IL-10) is the crucial driving factor for Tr1 cell differentiation, but the molecular mechanisms underlying this induction remain unknown. We identified and characterized a subset of IL-10–producing human dendritic cells (DCs), termed DC-10, which are present in vivo and can be induced in vitro in the presence of IL-10. DC-10 are CD14+, CD16+, CD11c+, CD11b+, HLA-DR+, CD83+, CD1a−, CD1c−, express the Ig-like transcripts (ILTs) ILT2, ILT3, ILT4, and HLA-G antigen, display high levels of CD40 and CD86, and up-regulate CD80 after differentiation in vitro. DC-10 isolated from peripheral blood or generated in vitro are potent inducers of antigen-specific IL-10–producing Tr1 cells. Induction of Tr1 cells by DC-10 is IL-10–dependent and requires the ILT4/HLA-G signaling pathway. Our data indicate that DC-10 represents a novel subset of tolerogenic DCs, which secrete high levels of IL-10, express ILT4 and HLA-G, and have the specific function to induce Tr1 cells.

The red jungle fowl leukocyte receptor complex contains a large, highly diverse number of chicken immunoglobulin-like receptor (CHIR) genes

The chicken Ig-like receptor (CHIR) gene family is located on microchromosome 31, the orthologous region to the mammalian leukocyte receptor complex. CHIR are equally related to the mammalian killer Ig-like receptors and leukocyte Ig-like transcripts, but they occur in a much higher number and diversity. The chicken microchromosome 31 has been neglected in the genome sequence analysis. Here, we provide a first analysis of this region. For this purpose bacterial artificial chromosome (BAC) sequences originating from a single inbred red jungle fowl that served as basis for the chicken genome project were screened for the presence of CHIR sequences and eight BACs were identified as major CHIR containing regions. Since the sequences of these BACs that were available in the database were not complete, sequence gaps were further closed by novel data from the chicken genome project. The entire sequence was aligned into 26 contigs covering 875 kbp that contained 84 functional CHIR and 46 CHIR pseudogenes that were hampered by different reasons such as premature stop codons. The 84 functional CHIR were further categorized into 35 activating (CHIRA), 26 inhibitory (CHIRB) and 23 bifunctional (CHIRAB) genes. A detailed comparison of the annotated sequence taking also into account the previously published CHIR BAC sequence originating from an Lohman selected leghorn chicken revealed that the CHIR locus seems to be a very active region with a high degree of gene reorganization that resembles a constant birth and death evolution. The present report provides a framework for the future completion of the CHIR locus. It further suggests that the entire microchromosome 31 may resemble a locus of extraordinary genomic diversity that is beneficial for the development of a large CHIR repertoire, but that has therefore lost all other genes, where such a diversification would be fatal.

Tyrosine nitration in the human leucocyte antigen‐G‐binding domain of the Ig‐like transcript 2 protein

Ig-like transcript 2 (ILT2) is a suppressive receptor that participates in the control of the autoimmune reactivity. This action is usually carried out in a proinflammatory microenvironment where there is a high production of free radicals and NO. However, little is known regarding whether these conditions modify the protein or affect its suppressive functions. The present study aimed to investigate the suppressive response of the ILT2 receptor under oxidative stress. To address this topic, we treated the ILT2-expressing natural killer cell line, NKL, with the NO donor N-(4-[1-(3-aminopropyl)-2-hydroxy-2-nitrosohydrazino]butyl)propane-1,3-diamine (DETA-NO). We observed that DETA-NO caused ILT2 protein nitration. MS analysis of the chimeric recombinant human ILT2-Fc protein after treatment with the peroxynitrite donor 3-(morpholinosydnonimine hydrochloride) (SIN-1) showed the nitration of Tyr35, Tyr76 and Tyr99, which are involved in human leucocyte antigen-G binding. This modification is selective because other Tyr residues were not modified by SIN-1. Recombinant human ILT2-Fc treated with SIN-1 bound a significantly higher quantity of human leucocyte antigen-G than untreated recombinant human ILT2-Fc. DETA-NO did not modify ILT2 mRNA expression or protein expression at the cell surface. Preincubation of NKL cells with DETA-NO decreased the cytotoxic lysis of K562-human leucocyte antigen-G1 cells compared to untreated NKL cells (P < 0.05) but increased cytotoxicity against K562-pcDNA cells (P < 0.05). Intracellular tyrosine phosphorylation produced after human leucocyte antigen-G binding was not affected by DETA-NO cell pretreatment. These results support the hypothesis that the ILT2human leucocyte antigen-G interaction should have a central role in tolerance under oxidative stress conditions when other tolerogenic mechanisms are inhibited.

Ig-Like Transcript 3 Regulates Expression of Proinflammatory Cytokines and Migration of Activated T Cells

Ig-like transcript 3 (ILT3), an inhibitory receptor expressed by APC is involved in functional shaping of T cell responses toward a tolerant state. We have previously demonstrated that membrane (m) and soluble (s) ILT3 induce allogeneic tolerance to human islet cells in humanized NOD/SCID mice. Recombinant sILT3 induces the differentiation of CD8(+) T suppressor cells both in vivo and in vitro. To better understand the molecular mechanisms by which ILT3 suppresses immune responses, we have generated ILT3 knockdown (ILT3KD) dendritic cells (DC) and analyzed the phenotypic and functional characteristics of these cells. In this study, we report that silencing of ILT3 expression in DC (ILT3KD DC) increases TLR responsiveness to their specific ligands as reflected in increased synthesis and secretion of proinflammatory cytokines such as IL-1alpha, IL-1beta, and IL-6 and type I IFN. ILT3KD-DC also secretes more CXCL10 and CXCL11 chemokines in response to TLR ligation, thus accelerating T cell migration in diffusion chamber experiments. ILT3KD-DC elicit increased T cell proliferation and synthesis of proinflammatory cytokines IFN-gamma and IL-17A both in MLC and in culture with autologous DC pulsed with CMV protein. ILT3 signaling results in inhibition of NF-kappaB and, to a lesser extent, MAPK p38 pathways in DC. Our results suggest that ILT3 plays a critical role in the control of inflammation.

Ig-like transcript 4 inhibits lipid antigen presentation through direct CD1d interaction.

NKT cells recognize lipid Ags presented by CD1d molecules and play an important role in the regulation of innate and adaptive immune responses. In this study, we report the identification of a membrane-associated protein, Ig-like transcript 4 (ILT4), as a novel human CD1d receptor that inhibits CD1d-mediated immune responses. We found that native CD1d tetramer generated by mammalian cells was able to specifically bind human monocytes in the peripheral blood, and this binding was at least partly mediated by monocyte-expressed ILT4. The interaction between ILT4 and CD1d involves the two N-terminal domains of ILT4 and the Ag-binding groove of CD1d (alpha1/alpha2 domain). This interaction has been identified on the cell surface as well as in the endosomal and lysosomal compartments. Functional analysis showed that ILT4 could block the loading of lipid Ags such as alpha-GalCer, and consequently inhibited NKT recognition. The interaction between ILT4 and CD1d may provide new insights into the regulation of NKT-mediated immunity.

Expression of Ig-Like Transcript 4 Inhibitory Receptor in Human Non-small Cell Lung Cancer

Human Ig-like transcript 4 (ILT-4) is a member of the inhibitory receptor family for immune function. Little is known about the expression levels of ILT-4 in tumor cells. We have studied the expression levels of ILT-4 both in vitro in cancer cell lines and in vivo in tumor tissues from 70 patients with non-small cell lung cancer (NSCLC) by reverse transcriptase-polymerase chain reaction, fluorescence-activated cell sorting, and immunohistochemical analysis. Three cancer cell lines (H1299, A549, and U1810) express ILT-4 messenger RNA, and only two cell lines (H1299 and A549) express ILT-4 protein on the cell surface. Approximately 37.1% of 70 tumor tissue samples express ILT-4, which is localized in the cell membrane and cytoplasm. In addition, tumor cells and stromal and plasma cells also express ILT-4. The number of infiltrating lymphoid cells in the tumor tissues that express B7-H3 was much lower than those that did not, but there is no significant correlation between ILT-4 expression and disease progression including nodal metastasis. These findings suggest that ILT-4 is frequently expressed in both tumor and stromal cells of NSCLC, and it might play an important role in regulation of the host immune system.

Expression of the CD85j (leukocyte Ig‐like receptor 1, Ig‐like transcript 2) receptor for class I major histocompatibility complex molecules in idiopathic inflammatory myopathies

Expression of class I major histocompatibility complex (MHC) molecules on the surface of muscle cells is a biologic feature of idiopathic inflammatory myopathies (IIM). Class I MHC-transgenic mouse models support a causative role for class I MHC expression by muscle cells in the pathogenesis of IIM. The muscle lesions are characterized by leukocyte infiltration. We undertook this study to analyze the expression in muscle lesions of various class I MHC-specific receptors on leukocytes and natural killer (NK) cells. We generated a panel of cell transfectants to control the immunofluorescence analysis of class I MHC receptor expression. We then analyzed the expression of CD158 (killer cell Ig-like receptors [KIRs]) and CD85j (leukocyte Ig-like receptor 1, Ig-like transcript 2) on muscle sections prepared from 14 patients with IIM (5 with dermatomyositis [DM], 5 with polymyositis [PM], and 4 with sporadic inclusion body myositis [IBM]). We could not detect the presence of NK cells in inflammatory lesions. However, the class I MHC receptor CD85j, but no KIRs, was expressed by inflammatory cells infiltrating muscle lesions in IIM. CD85j is expressed in PM and sporadic IBM at the sites of partial invasion and in DM in perivascular inflammation, paving the way for dissecting the role of CD85j in the pathogenesis of inflammatory myopathies.

Maturation of dendritic cells depends on proteolytic cleavage by cathepsin X

The maturation status of dendritic cells (DCs) is crucial for effective antigen presentation and initiation of the primary immune response. Maturation stimuli cause the adhesion of immature DCs to the extracellular matrix, which is accompanied by recruitment of the CD11b/CD18 [macrophage antigen-1 (Mac-1)] integrin receptor, cytoskeleton reorganization, and podosome formation. Cathepsin X, a cysteine protease expressed in DCs and other APCs, is involved in Mac-1 activation. We have shown that during maturation, cathepsin X translocates to the plasma membrane of maturing DCs, enabling Mac-1 activation and consequently, cell adhesion. In mature DCs, cathepsin X redistributes from the membrane to the perinuclear region, which coincides with the de-adhesion of DCs, formation of cell clusters, and acquisition of the mature phenotype. Inhibition of cathepsin X activity during DC differentiation and maturation resulted in an altered phenotype and function of mature DCs. It reduced surface expression of costimulatory molecules, increased expression of inhibitory Ig-like transcripts 3 and 4 (ILT3 and ILT4), almost completely abolished cytokine production, diminished migration, and reduced the capacity of DCs to stimulate T lymphocytes. These results stress the importance of cathepsin X in regulating DC adhesion, a crucial event for their maturation and T cell activation.

Modulation of dendritic cell differentiation by HLA-G and ILT4 requires the IL-6—STAT3 signaling pathway

The expression of Ig-like transcript (ILT) inhibitory receptors is a characteristic of tolerogenic dendritic cells (DCs). However, the mechanisms of modulation of DCs via ILT receptors remain poorly defined. HLA-G is a preferential ligand for several ILTs. Recently, we demonstrated that triggering of ILT4 by HLA-G1 inhibits maturation of human monocyte-derived conventional DCs and murine DCs from ILT4 transgenic mice, resulting in diminished expression of MHC class II molecules, CD80 and CD86 costimulatory molecules, and prolongation of skin allograft survival. Different isoforms of HLA-G have diverse effects on the efficiency to induce ILT-mediated signaling. In this work, we show that HLA-G1 tetrameric complex and HLA-G5 dimer, but not HLA-G5 monomer, induce strong ILT-mediated signaling. We determined that the arrest of maturation of ILT4-positive DCs by HLA-G ligands involves the IL-6 signaling pathway and STAT3 activation. Ligation of ILT4 with HLA-G on DCs results in recruitment of SHP-1 and SHP-2 protein tyrosine phosphatases. We propose a model where SHP-2 and the IL-6-STAT3 signaling pathway play critical roles in the modulation of DC differentiation by ILT4 and HLA-G.