Expression Of ILT4 Research Articles

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.

HLA-G and HLA-E in patients with juvenile idiopathic arthritis

To investigate the expression and release of HLA-G and HLA-E in JIA. Soluble (s)HLA-G and HLA-E were measured in sera from 58 JIA patients and 54 healthy donors. Surface expression of HLA-G, HLA-E and immunoglobulin-like transcript (ILT)2 and ILT4, two receptors for HLA-G, was assessed on T, B cells and monocytes from peripheral blood (PB) and SF of 12 JIA patients and from PB of 12 controls. Serum sHLA-G concentration was significantly lower in patients than in controls. Both sHLA-G and sHLA-E were detected in SF and sHLA-E concentration in SF was higher in extended oligoarticular/polyarticular than in limited oligoarticular JIA. Patients compared with controls showed: (i) down-regulation of HLA-E and ILT2 expression on T cells; (ii) up-regulation of HLA-E expression on B cells and monocytes; and (iii) down-regulation of ILT4 expression on monocytes. Comparing JIA patients' SF and PB we found: (i) up-regulation of HLA-E and ILT2 expression in T and B cells and monocytes; and (ii) down-regulation of ILT4 expression in monocytes. ILT4 was up-regulated in monocytes from oligoarticular extended/polyarticular compared with oligoarticular limited JIA. A lower concentration of sHLA-G in sera may predispose to JIA, as observed for other autoimmune diseases. sHLA-E concentration in SF correlate with the number of affected joints. Higher ILT2 expression on SF cell populations compared with PB may be related to high sHLA-G concentration in SF. Higher HLA-E expression in SF than in PB cell populations may protect them from NK cytolysis.

Early markers for protective mechanisms during rush venom immunotherapy

Allergen-specific venom immunotherapy (VIT) represents the only rational-based option to treat allergic sensitizations against bee and wasp venom. So far, there is not much knowledge about early induction of protective and tolerogenic pathways during VIT. To identify the earliest markers for protective mechanisms against allergic reactions in the peripheral blood during the build-up phase of VIT. PBMC and monocytes were isolated, and serum samples were taken before and during a five day build-up phase from 65 hymenoptera venom allergic patients. Expression level of tolerogenic markers was analyzed on mRNA and protein level. Serum levels of different soluble tolerogenic factors were measured. We observed significantly enhanced tryptophan degradation, elevated ILT4 expression of monocytes as well as IL-10 production of CD3(+) T cells only a few hours after the first injection on day 1, followed by increased IL-10 serum levels, monocyte apoptosis and elevated intracellular cAMP levels of monocytes on day 3 combined with a higher ILT3 protein expression and IL-10 secretion of monocytes on day 5. From these data, we conclude that tryptophan depletion, ILT3/4-mediated inhibition, higher IL-10 production as well as intracellular cAMP might contribute to early induction of protective mechanisms against allergic reactions during the build-up phase of VIT.

IL-10 enhances promoter activity of ILT4 gene and up-regulates its expression in THP-1 cells

This study examined the effect of IL-10 on immunoglobulin-like transcript (ILT4) expression of human monocytic leukemic cell line THP-1, especially the role of the ILT4 promoter activity. ILT4 promoter area was amplified by PCR, and was cloned into the eukaryotic expressing vector pGL3-Basic. The pGL3-ILTP obtained was tested by double endonuclease digestion and sequencing. Then, the recombinant plasmid was transfected into THP-1 cells by using lipofectamine. After culture with IL-10 for 12 h, the mRNA extracted from THP-1 cells was detected by RT-PCR and the protein was detected by FACS. The dual-luciferase reporter assay system was employed to detect the activity of ILT4 promoter with or without IL-10. The results showed that the activity of pGL3-ILTP was significantly increased and was more than ten times that of pGL3-Basic cells. After culture with IL-10 for 12 h, the expression of ILT4 protein and its mean fluorescence intensity (MFI) were increased. Moreover, the mRNA was remarkably higher than that of the control group. Dual-luciferase reporter assay revealed that ILT4 promoter was much more activated after being treated with IL-10. We were led to conclude that pGL3-ILTP containing ILT4 promoter was constructed successfully. The expression of ILT4 could be up-regulated by IL-10 both at the transcriptional and translational level. Furthermore, ILT4 promoter could be much more active after addition of IL-10. This study suggests that IL-10 up-regulates ILT4 expression on monocytes via increasing ILT4 gene promoter activity, which may have implication for inducing transplantation tolerance in clinical practice.

Applications of monitoring human leukocyte antigen G and its inhibitory receptors in post-renal transplantation

to study the feasibility of human leucocyte antigen-G (HLA-G) as a post-transplantation prognostic biomarker and discuss the correlation of its receptor expression and the mechanisms. a total of 215 recipients in our centre from February 2006 to June 2008 were divided into stable kidney function group (n = 173) and acute rejection group (n = 42). The soluble human leucocyte antigen-G5 (sHLA-G5) level in peripheral plasma was detected by ELISA. And the HLA-G receptor ILT-2, KIR2DL4 on T, B, NK lymphocytes were analyzed by flow cytometry (FCM). The sHLA-G5 cutoff level by ROC curve was employed to predict the events of acute post-transplantation rejection. And regression analysis was used to determine the association of sHLA-G5 with acute rejection. an optimal cutoff value of 139.0 microg/L could be defined for sHLA-G5 (sensitivity: 63.6%, specificity: 82.1%, AUC: 0.780). Binary regression analysis showed that sHLA-G5 played an independent role on acute rejection (P = 0.019, OR = 0.039, 95%CI: 2.091 - 5.661). The rate of HLA-G receptor ILT-2 on CD4(+)T cell, CD8(+)T cell and B cell in acute rejection group was statistically lower than that in stable kidney function group (21% ± 7% vs 52% ± 17%, 23% ± 6% vs 39% ± 16%, 21% ± 7% vs 39% ± 16%, all P < 0.05). The expression of KIR2DL4 on NK cells in acute rejection group was statistically lower than that in stable kidney function group (31% ± 10%vs 57% ± 21%, P < 0.05). sHLA-G5 level may be predicted for acute rejection with a high sensitivity and specificity. The up-regulated expression of ILT-2 and KIR2DLT may contribute to immunology tolerance in peripheral circulation.

Asthma and allergies in farming environments – The PASTURE/FORALLVENT/EFRAIM-Project

Background: Cross-sectional studies in farming and rural environments (ALEX, PARSIFAL) revealed new hints about protective effects of contact to farm animals and the consumption of farm milk on asthma, atopy, and hay fever. Therefore, the prospective birth cohort study PASTURE among farming and non-farming populations was started in 2002 funded by the European Commission. The aim was to assess the role of indoor exposure to microbes and to investigate immunological and genetic mechanisms. The cohort was followed up by FORALLVENT (2006–2008) and EFRAIM (2008–2012). Material and Methods: In total, 1133 children (46.8% from farming families) were recruited in 5 European counties (Austria, Finland, France, Germany, and Switzerland). Yearly detailed questionnaire data and diary data (first year of living) were obtained asking about respiratory diseases, activities on farms, and life style. In blood samples from cord blood and at the age 1, 4.5, and 6 years the T-cell and B-cell response was assessed; genetic markers and mRNA were measured. Furthermore, breast milk, cow's milk, faeces, and diverse dust samples (mattress, indoor floor, cow's shed) were available in the biobank for different time points. Skin examinations and lung function measurement were performed. Questionnaire data and blood samples of siblings were additionally collected. Results from first cord blood analyses Exposure to farming environment and working activities on farms of mothers during pregnancy affected the cytokine- and IgE-expression. Specific IgE to allergens was associated with reduced IFN-gamma and with maternal immunity to Toxoplasma gondii and rubella virus. Maternal vitamin D intake during pregnancy increased gene expression of ILT3 and ILT4. Discussion: In future, the interdisciplinary project team expects a better understanding of the mechanisms of developing allergic immune response. New insights into the observed strong protective effects will allow to identify necessary steps for further development of preventive strategies for asthma and allergies in childhood.

Human NK cells display major phenotypic and functional changes over the life span

Aging is generally associated with an increased predisposition to infectious diseases and cancers, related in part to the development of immune senescence, a process that affects all cell compartments of the immune system. Although many studies have investigated the effects of age on natural killer (NK) cells, their conclusions remain controversial because the diverse health status of study subjects resulted in discordant findings. To clarify this situation, we conducted the first extensive phenotypic and functional analysis of NK cells from healthy subjects, comparing NK cells derived from newborn (cord blood), middle-aged (18-60 years), old (60-80 years), and very old (80-100 years) subjects. We found that NK cells in cord blood displayed specific features associated with immaturity, including poor expression of KIR and LIR-1/ILT-2 and high expression of both NKG2A and IFN-gamma. NK cells from older subjects, on the other hand, preserved their major phenotypic and functional characteristics, but with their mature features accentuated. These include a profound decline of the CD56(bright) subset, a specific increase in LIR-1/ILT-2, and a perfect recovering of NK-cell function following IL2-activation in very old subjects. We conclude that the preservation of NK cell features until very advanced age may contribute to longevity and successful aging.

Inhibition of HLA-G Expression Via RNAi Abolishes Resistance of Extravillous Trophoblast Cell Line TEV-1 to NK Lysis

Remodelling of uterine spiral arteries occurs in the first trimester of pregnancy and involves an expanded and activated population of maternal natural killer (NK) cells in the decidua and extravillous trophoblast cells. Invasive trophoblasts encounter maternal NK cells during their invasion into the uterine tissue, posing the problem of susceptibility to NK lysis. Studies in vitro and in vivo suggested that the expression of HLA-G by invasive extravillous trophoblasts might provide invulnerability to NK cells, while there is still lack of direct evidence of HLA-G knockdown effect on trophoblast/NK interaction. A study was conducted to investigate the effects of down-regulated HLA-G on extravillous trophoblasts. The short hairpin RNA (shRNA) vector targeting HLA-G was constructed and transfected into the human first-trimester extravillous trophoblast cell line TEV-1. Western blotting and reverse transcription polymerase chain reaction (RT-PCR) revealed that in HLA-G shRNA transfected cells, the expression of HLA-G was significantly decreased. HLA-G expression was also visualised by confocal imaging. The HLA phenotype of TEV-1 cells and inhibitory receptors expression in NK cells were analysed by flow cytometry. A comparison between HLA-G knockdown and non-knockdown cells showed a significant difference in the HLA expression profile without altering HLA-C and HLA-E. Both primary NK cells and NK-92 cell line exhibited potent cytotoxicity against HLA-G knockdown cells via standard 4-h 51Cr release assays. Expression of ILT2, ILT4 and KIR2DL4 in NK cells was unchanged after 4h of co-culture, while KIR2DL4 expression increased after 48h. We conclude that HLA-G contributes to trophoblast/NK interaction, acting as a key regulator of NK cytolysis in this human extravillous trophoblast cell model. In addition, TEV-1 cells share common HLA phenotype characters with extravillous trophoblast cells, and thus might be used as a good cell model. HLA-C expression in trophoblasts is not correlated with HLA-G translation and HLA-C alone was sufficient to boost HLA-E surface expression. In addition, RNA interference could be employed as a feasible and effective method to study HLA-G functions.

Maternal vitamin D intake during pregnancy increases gene expression of ILT3 and ILT4 in cord blood

Recent studies indicate that prenatal vitamin D intake may protect against the development of atopic diseases in young children. Vitamin D has been shown to induce tolerogenic antigen-presenting cells such as dendritic cells. Whether the allergy-protective potential of prenatal vitamin D is mediated through such mechanisms is, however, unknown. To evaluate the association between prenatal vitamin D supplementation and tolerogenic antigen-presenting cells in cord blood (CB) as determined by mRNA measurement of immunoglobulin-like transcripts (ILT)3 and ILT4. A prospective multi-centre birth cohort was established in rural areas of five European countries. Information on maternal exposures including vitamin D intake was collected by questionnaires during pregnancy. The gene expression of ILT3 and ILT4 was analysed by real-time PCR in the CB of 927 children. Maternal vitamin D supplementation was assessed in Finland and France (n=349). Maternal vitamin D supplementation during pregnancy was associated with an increase in the gene expression of ILT3 (P=0.012) and ILT4 (P<0.001). This association remained significant for ILT4 (P=0.020) and showed a positive trend for the gene expression of ILT3 (P=0.059) after multivariate analysis controlling for various confounders. Vitamin D supplementation during pregnancy may increase the mRNA levels of ILT3 and ILT4 in CB. This finding may point towards an early induction of tolerogenic immune responses by maternal vitamin D intake.

Multiple Myeloma Derived Factors Generate Tolerogenic Dendritic Cells by Activation of STAT3: A Potential Target for Immunotherapy.

Abstract 3031Poster Board II-1007Immunotherapy is a promising therapy for lymphoproliferative malignancies including multiple myeloma (MM). While many phase I and II clinical trials has been conducted using dendritic cells (DC) pulsed with cancer-specific antigen as immunotherapy, none of these studies have shown a significant clinical impact on treatment of patients with cancer, including in MM. The failure of DC-based vaccine in MM has been associated with impaired DC function due to the immunosuppressive environment caused by the cancer cells. This effect may be in part mediated through hyper-activation of STAT3. Using MM as a model, we investigated the effect of MM-derived factors on the function of DC and evaluated the effect on STAT3 hyper-activation in the latter.We generated DC from monocytes of normal donors using IL-4 and GM-CSF for 48 hours, following maturation using IL-1β, TNF-αa, PGE2 for another 24 hours. MM-derived factors (MM-DF) were obtained by co-culture of the human MM cell line U266 cells with in-vitro generated osteoclasts for 48 hours, and the supernatant was filtered. The filtered supernatant containing MM-DF was used in a ratio of 1:1 with fresh RPMI-1640 in the generation of DC in-vitro. The phenotype of immature and maturated DC was evaluated by flow-cytometry for surface staining of CD40, CD80, CD83, CD86, ILT3, and MHC-II. Lysates of DC were prepared for Western blot to evaluate for phospo (p)-STAT3 and ILT3 expression. The ability of generated DC (in the presence or absence of MM-DF) to stimulate T cells was assessed using 3H-thymidine incorporation.MM-DF did not affect the generation of DC as assessed by CD86 and MHC-II expression, but significantly impaired the maturation of DC with failure of up-regulation of CD40 (64% ±19.3% of control; P=0.075), CD80 (41% ±20.5% of control; P=0.019), and CD83 (35% ±18.0% of control; P=0.0077) in 3 independent experiments. In addition, the surface expression of ILT3 on the generated DC was significantly increased in DC generated in presence of MM-DF (155% increase in mean fluorescence intensity compare to control), furthermore, western blot of DC lysates showed 2.5 fold-increase in the expression of ILT3 in DC generated in presence of MM-DF compare to control. The failure of maturation of DC with higher expression of ILT3 indicates the generation of tolerogenic DC (Transpl Immunol 2003;11:245). The expression of phoso-STAT3αa (pY705αa or the active form of pSTAT3) was also significantly increased in DC generated in the presence of MM-DF compare to control (1.6-2.3 fold-increase), furthermore, higher expression of phoso-STAT3β (negative form of pSTAT3) was found in DC generated in the absence of MM-DF (Figure 1). Finally the function of generated DC in the presence of MM-DF was impaired compare to control (40-60% decrease in ability to stimulate allogeneic T cell) as assessed by 3H-Thymidine incorporaton Assay.Our results demonstrate a significant effect of MM-DF on the maturation of DC rendering them tolerogenic, and this is associated with hyper-activation of STAT3. RNA silencing experiments to knock down STAT3 and pharmacological inhibition experiments are in progress to assess the ability to abrogate the effect of MM-DC on DC maturation and improve function. This provides a potential therapeutic target in immunotherapy trials. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

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.

Induction of ILT expression on nonprofessional antigen presenting cells: Clinical applications

Achieving tolerance toward solid organ allografts remains challenging. Nonprofessional antigen-presenting cells, mainly endothelial cells, representing the first barrier between the donor organ and the recipient immune system represent a promising platform to induce tolerance towards allografts without compromising the recipient's immune system. Expression of ILT3 and ILT4, two members of the immunoglobulin-like transcript (ILT) family, has been demonstrated in endothelial cells in vitro and in vivo resulting in modulated endothelium-dependent T-cell activation. We here review mechanisms and effects of ILT3/ILT4 upregulation on endothelial cells and their potential significance in solid organ transplantation.

Rapamycin downregulates the inhibitory receptors ILT2, ILT3, ILT4 on human dendritic cells and yet induces T cell hyporesponsiveness independent of FoxP3 induction

Dendritic cells (DC) mediate potent alloimmune responses through the positive costimulatory molecules CD40, CD80 and CD86 while the negative costimulatory molecules, immunoglobulin-like transcript 2 (ILT2), ILT3 and ILT4 have been associated with tolerogenic DC function. Due to the pivotal role played by DC in immunity the effect of the immunosuppressive agent rapamycin (RAPA) on the expression profile of both positive and negative costimulatory molecules during human DC differentiation and maturation was investigated. During monocyte differentiation to DC an increase in the mean fluorescence intensity (MFI) for CD40, CD80, CD83 and CD86 was observed in association with a concomitant downregulation of ILT2, ILT3, ILT4 and HLA-G expression. While DC differentiation in the presence of RAPA (10 nM) showed a reduction in the MFI for CD40, CD80 and CD86 expressing cells, treatment after differentiation had no effect on the expression of these costimulatory molecules. The inhibitory receptors were also downregulated by RAPA only when added during differentiation. In comparison to RAPA, Cyclosporin A (CsA) had relatively minor effects on DC phenotype whereas IFN-α showed induction of CD80, CD86 and HLA-G when added prior to differentiation. Functionally, RAPA-treated DC used as stimulators in a DC–T cell mixed lymphocyte reaction (MLR) showed 40% inhibition of T cell proliferation relative to untreated DC ( P = 0.001) whereas CsA-treated DC showed no difference, and IFN-α-treated DC stimulated T cell proliferation. Nevertheless, the induction of T cell hyporesponsiveness by coculture of RAPA-treated DC with T cells was not associated with the generation of increased numbers of FoxP3 positive T regulatory cells. In conclusion, although RAPA downregulated ILT2, ILT3 and ILT4 expression in DC, the inhibition of T cell proliferation by RAPA-treated DC is predominantly due to the reduction of CD40, CD80 and CD86 expression rather than the propensity to generate FoxP3 expressing regulatory cells.

Induction of immune tolerance through an IL‐10 dependent mechanism allows Entamoeba histolytica successful cololization in the colon

Entamoeba histolytica infects 10% of the world's population, which can lead to amoebic dysentery and/or liver abscess. The majority of infected individuals are completely asymptomatic. We hypothesized that E. histolytica is capable of inducing immune tolerance in antigen presenting cells thus promoting parasite survival in the colon in the absence of an overt inflammatory or immune response. In support of this, we show that dendritic cells (DC's) and macrophages (mφ) exposed to amoeba secreted proteins can induce a significantly up‐regulation of IL‐10 mRNA expression and increase phosphorylation of ILT2. Furthermore, low concentrations (10μg/mL) of amoebic proteins caused rapid 3–8‐fold up‐regulation of ILT2 inhibitory receptor on CD4+ T cells in the presence of mφ or DCs 36 h following exposure. However, there was no significant up‐regulation of ILT2 if T cells were deficient in IL‐10. Moreover, amoebic proteins caused a significantly increased in apoptosis in IL‐10 deficient T cells after 48 or 72 hr as compared to wild type controls. We conclude that E. histolytica is capable of inducing IL‐10 production in antigen presenting cells, which is required for up‐regulating ILT2 on CD4+ T cells and inhibition of T cells apoptosis. This work was supported by NSERC. JMR is a recepient of the QE II Graduate Scholarship from the Province of Alberta and CAG/CIHR studentship.

CD8 +FOXP3 +T Cells From Renal Transplant Recipients in Quiescence Induce Immunoglobulin-like transcripts-3 and -4 on Dendritic Cells From Their Respective Donors

In previous studies, CD8 +FOXP3 + T cells have been shown to upgrade the inhibitory receptors on dendritic cells (DC) in heart transplantation patients. The inhibitory receptor were immunoglobulin-like transcripts (ILT)-3 and ILT-4, which were highly expressed on tolerogenic dendritic cells. Our study focused on the CD8 +FOXP3 + T cells from allogeneic renal transplant recipients, seeking to dissert their function in inducing ILT-3– and ILT-4–expressing dendritic cells. We analyzed 11 quiescent renal transplant recipients and their respective donors. Reverse transcriptase polymerase chain reaction analysis showed that the levels of Foxp3 mRNA in CD8 + T cells after renal transplantation were significantly higher than those in CD8 + T cells before transplantation. We further investigated the capacity of allospecific CD8 +FOXP3 + T cells to induce upregulation of the inhibitory receptors ILT-3 and ILT-4 on DC. Data from FACS showed that CD8 +FOXP3 + T cells induced greater expression of ILT-3 and ILT-4 on DC in an allospecific fashion. This study confirmed that Foxp3 gene expression was enhanced among quiescent renal transplant patients, suggesting that CD8 +FOXP3 + T cells play an important role in unresponsiveness related to upregulated ILT-3 and ILT-4 receptors on DC.

21-OR: Expression of inhibitory receptors ILT3 and ILT4 on neoplastic B cells from patients with high-risk chronic lymphocytic leukemia: Implications for allogeneic stem cell transplantation

21-OR: Expression of inhibitory receptors ILT3 and ILT4 on neoplastic B cells from patients with high-risk chronic lymphocytic leukemia: Implications for allogeneic stem cell transplantation

Expression of inhibitory receptor ILT3 on neoplastic B cells is associated with lymphoid tissue involvement in chronic lymphocytic leukemia

T cell responses against leukemia-associated antigens have been reported in chronic lymphocytic leukemia (CLL). However, the relentless accumulation of CLL B cells in some patients indicates that anti-tumor immune responses are inefficient. Inhibitory receptors from the Ig-like transcript (ILT) family, such as ILT3 and ILT4, are crucial to the tolerogenic activity of antigen presenting cells. In this study, we examined the expression of ILT3 on CD5+ B cells obtained from 47 patients with CLL. Using flow cytometry and RT-PCR, we found that B CLL cells from 23 of 47 patients expressed ILT3 protein and mature ILT3 mRNA. ILT3 protein and mRNA were not found in normal B cells obtained from donors without CLL. Expression of ILT4 in normal and B CLL cells showed a pattern similar to ILT3. The frequency of ILT3 positive CLL B cells was higher in patients with lymphoid tissue involvement, suggesting that ILT3 may have prognostic value in CLL. Our findings indicate that expression of ILT3 and ILT4 on CLL B cells represents a phenotypic abnormality that may play a role in tolerization of tumor-specific T cells.

Analysis of expression and function of the inhibitory receptor ILT2 (CD85j/LILRB1/LIR-1) in peripheral blood mononuclear cells from patients with systemic lupus erythematosus (SLE)

The aim of this work was to study the expression and function of the inhibitory receptor ILT2/CD85j in peripheral blood mononuclear cells (PBMC) from patients with systemic lupus erythematosus (SLE). We studied 23 SLE patients as well as 17 patients with rheumatoid arthritis, 10 with fibromyalgia, and 23 healthy individuals. We found a variable level of expression of ILT2 in the PBMC from both SLE patients and controls, with no significant differences among them. However, when the expression of this receptor was assessed in cell subsets, significantly lower levels were detected in CD19+ lymphocytes from SLE patients compared with healthy controls. Functional assays performed in unfractionated PBMC, showed a significant diminished inhibitory activity of ILT2 in CD4+ and CD8+ cell subsets from SLE patients compared to either rheumatoid arthritis or fibromyalgia patients, and healthy individuals. Our results show that the PBMC from some patients with SLE show a defective expression of ILT2, and that most of them exhibit a poor function of this inhibitory receptor.

Placental expression of HLA-G, and its correlation with ILT2 expression on surface of CD4+ T lymphocytes and NK cells in preeclamptic women

Placental expression of HLA-G, and its correlation with ILT2 expression on surface of CD4+ T lymphocytes and NK cells in preeclamptic women

Aspirin-Treated Human DCs Up-Regulate ILT-3 and Induce Hyporesponsiveness and Regulatory Activity in Responder T Cells

Mature dendritic cells (mDCs) are potent antigen presenting cells, but immature DCs (iDCs) have been shown to have reduced antigen stimulatory capacity. Different strategies have been investigated to augment the tolerogenic capacity of dendritic cells (DCs). We demonstrate that in aspirin-treated human DCs, there is reduced expression of CD1a, HLA-DR and CD86, up-regulation of ILT-3 expression and marginal increases in PDL-1. Aspirin-treated DCs are partially resistant to phenotypic changes following maturational stimuli, such as lipopolysaccharide (LPS) or TNFalpha, IL-1alpha and PGE2. Aspirin-treated DCs demonstrate normal endocytic function, but have a reduced ability to stimulate allogeneic T cells, which is comparable to iDCs. Furthermore, they induce hyporesponsiveness and regulatory activity in responder naïve and memory T cells; for naïve T cells this is achieved more quickly and efficiently than with iDCs. We investigated the mechanism of this regulatory activity and found that both cell-cell contact and inhibitory cytokine activity are involved, although no one cytokine predominates in importance. Blocking ILT-3 or IL-12 does not diminish the capacity of these DCs to induce regulation or Foxp3 expression on the regulatory T cells. Results demonstrate that aspirin-treated DCs display tolerogenic potential, which is of interest in their therapeutic potential in reducing chronic allograft rejection.