Retroviral transduction of acute myeloid leukaemia-derived dendritic cells with OX40 ligand augments their antigen presenting activity

Recent studies have shown that human myeloid leukaemia cells can differentiate into dendritic cell (DC)-like cells (leukaemia-DCs) when cultured with a combination of cytokines. In the present study, we examined whether the transduction of leukaemia-DCs with OX40 ligand (OX40L), a member of the tumour necrosis factor (TNF) family, resulted in augmentation of their antigen presenting activity. Bicistronic retroviral vectors expressing both human OX40L and enhanced green fluorescent protein (EGFP) or EGFP alone were generated and used for transduction. Fresh leukaemic cells from five patients with acute myeloid leukaemia (AML) were isolated and retrovirally transduced with OX40L during the culture with a combination of cytokines from stem cell factor, fms-like tyrosine kinase (Flt)-3 ligand, granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin-4 (IL-4) and TNF-alpha. After 7 d, the majority of cells showed DC-like morphology, and expressed higher levels of CD80, CD86 and HLA-DR than fresh leukaemic cells. The transduction efficiency was 8.5-27.2%. Leukaemia-DCs transduced with OX40L elicited higher proliferative response of allogeneic CD4+ T cells than fresh leukaemic cells, non-transduced, or mock-transduced leukaemia-DCs. Co-culture of allogeneic CD4+ T cells with OX40L-transduced leukaemia-DCs was superior in the generation of interferon (IFN)-gamma producing CD4+ T cells and in production of IFN-gamma. Furthermore, OX40L-transduced leukaemia-DCs could elicit significant proliferative response of human leucocyte antigen-matched T cells from the donor in allogeneic stem cell transplantation. These results indicate that retroviral transduction of leukaemia-DCs with OX40L augments their antigen presenting cell activity and thus renders them more suitable for tumour vaccines or ex vivo stimulation of leukaemia-specific T cells.     

Murine liver plasmacytoid dendritic cells become potent immunostimulatory cells after Flt-3 ligand expansion

The liver has unique immunological properties. Although dendritic cells (DCs) are central mediators of immune regulation, little is known about liver DCs. Plasmacytoid DCs (pDCs) are a recently identified subtype of murine liver DC. We sought to define the function of freshly isolated murine liver pDCs. We found that normal liver pDCs were weak in stimulating T cells, yet they possessed a proinflammatory cytokine profile with high tumor necrosis factor-alpha and low IL-10 secretion. To facilitate the investigation of murine liver pDCs, we expanded them in vivo with fms-like tyrosine kinase 3 ligand (Flt3L). After Toll-like receptor-9 ligation, expanded liver pDCs secreted high levels of IFN-alpha and were able to stimulate NK cells, NKT cells, and antigen-specific CD8+ T cells in vitro. In addition, Flt3L expansion alone generated pDCs capable of activating antigen-specific CD8+ T cells in vivo. Conclusion: Unstimulated liver pDCs exist in a latent state with the potential to become potent activators of the innate and adaptive immune systems through their interactions with other immune effectors. Our findings have implications for understanding the role of the liver in tolerance and immunity.     

OX40 ligand shuts down IL-10-producing regulatory T cells

IL-10-producing CD4(+) type 1 regulatory T (Tr1) cells play a critical role in the maintenance of peripheral tolerance. Although immunosuppressive drugs, cytokines, costimulatory molecules, and immature dendritic cells are implicated in the induction of Tr1 cells, the signals that negatively regulate the generation and function of Tr1 cells have been elusive. We report that OX40 ligand (OX40L) completely inhibited the generation of IL-10-producing Tr1 cells from na?ve and memory CD4(+) T cells induced by the immunosuppressive drugs dexamethasone and vitamin D3. This unique function of OX40L was not shared by two costimulatory TNF family members, GITR ligand and 4-1BB ligand. OX40L strongly inhibited the generation of IL-10-producing Tr1 cells induced by two physiologic stimuli, the inducible costimulatory ligand and immature dendritic cells. In addition, OX40L strongly inhibited IL-10 production and suppressive function of differentiated IL-10-producing Tr1 cells. These two novel functions of OX40L shed light on the mechanism by which OX40/OX40L regulates immunity and tolerance.     

Introduction of OX40 ligand into lymphoma cells elicits anti-lymphoma immunity in vivo

OBJECTIVE: OX40, a member of the TNF receptor superfamily, and its ligand (OX40L) play crucial roles in induction and maintenance of integrated T cell immune response. Engagement of OX40L delivers a costimulatory signal to T cells. In this study, we investigated whether inoculation of OX40L-transfected EL4, a murine T cell lymphoma cell line, could induce anti-lymphoma immunity in mice. MATERIALS AND METHODS: Female C57BL/6 mice were inoculated with 1 x 10(5) cells of parental EL4, OX40L-transfected EL4 (EL4-OX40L), or mock control vector-transfected EL4 (EL4-mock), and then the tumor size, overall survival, CTL activity of spleen cells, and the immunohistochemistry were compared. RESULTS: While both parental EL4 and EL4-mock grew rapidly, EL4-OX40L was rejected or grew slower than parental EL4 or EL4-mock. Pretreatment of mice with either anti-CD4 or anti-CD8 mAb accelerated the growth of EL4-OX40L, suggesting that both CD4+ and CD8+ T cells were involved in anti-lymphoma immunity. The immunohistochemical study revealed the infiltration of CD8+ T cells into the tumor of EL4-OX40L. In vitro CTL assay demonstrated that spleen cells of mice that had rejected EL4-OX40L had significant cytotoxic activity against parental EL4. CONCLUSION: The gene transfer of OX40L into lymphoma cells is an eligible and efficient modality to induce anti-lymphoma immunity.     

Leukotriene C4 induces migration of human monocyte-derived dendritic cells without loss of immunostimulatory function

Generation of human monocyte-derived dendritic cells (DCs) for cancer vaccination involves ex vivo maturation in the presence of proinflammatory cytokines and prostaglandin E(2) (PGE(2)). Although the inclusion of PGE(2) during maturation is imperative for the induction of DC migration, PGE(2) has unfavorable effects on the immunostimulatory capacity of these cells. Like PGE(2), leukotrienes (LTs) are potent mediators of DC migration. We therefore sought to characterize the migratory and immunologic properties of DCs that matured in the presence of LTB(4), LTC(4), LTD(4), and PGE(2). Here, we demonstrate that DCs matured in the presence of LTC(4), but not LTB(4) or LTD(4), are superior to PGE(2)-matured DCs in stimulating CD4(+) T-cell responses and in inducing antigen-specific cytotoxic T lymphocytes (CTLs) in vitro without concomitant induction or recruitment of regulatory T cells (Tregs). LTC(4)-matured DCs migrate efficiently through layers of extracellular matrix and secrete higher levels of immunostimulatory IL-12p70 while producing reduced levels of immune-inhibitory IL-10, IL12p40, indoleamine-2,3-dioxidase, and TIMP-1 (tissue inhibitor of matrix metalloproteinases). Intracellular calcium mobilization and receptor antagonist studies reveal that, in contrast to LTD(4), LTC(4) did not signal through CysLTR(1) in DCs. Collectively, our data suggest that LTC(4) represents a promising candidate to replace PGE(2) in DC maturation protocols for cancer vaccination.     

Correlation of OX40 ligand on B cells with serum total IgE and IL-4 levels by CD4+ T cells in allergic rhinitis

Introduction and objectives

Allergic rhinitis (AR) is a classic Th2-mediated disease, with important contributions to the pathology of interleukins 4, 5, and 13. The co-stimulatory molecule of OX40 and its ligand interaction participate in the immune response by regulation of Th1/Th2 cells balance. Considering the paucity of information on the relation between OX40 ligand (OX40L) and AR, this study aimed to examine its expression on B lymphocytes.

Expansion of HIV-specific CD4+ and CD8+ T cells by dendritic cells transfected with mRNA encoding cytoplasm- or lysosome-targeted Nef

Transfection with synthetic mRNA is a safe and efficient method of delivering antigens to dendritic cells for immunotherapy. Targeting antigens to the lysosome can sometimes enhance the CD4+ T-cell response. We transfected antigen-presenting cells (APCs) with mRNA encoding Gag-p24 and cytoplasmic, lysosomal, and secreted forms of Nef. Antigen-specific cytotoxic T cells were able to lyse the majority of transfected targets, indicating that transfection was efficient. Transfection of APCs with a Nef construct bearing lysosomal targeting signals produced rapid and prolonged antigen presentation to CD4+ and CD8+ T cells. Polyclonal CD4+ and CD8+ T-cell lines recognizing multiple distinct epitopes were expanded by coculture of transfected dendritic cells with peripheral blood mononuclear cells from viremic and aviremic HIV-infected subjects. Importantly, lysosome-targeted antigen drove a significantly greater expansion of Nef-specific CD4+ T cells than cytoplasmic antigen. The frequency of recognition of CD8 but not CD4 epitopes by mRNA-expanded T cells was inversely proportional to sequence entropy and was similar to ex vivo responses from a large chronic cohort. Thus human dendritic cells transfected with mRNA encoding lysosome-targeted HIV antigen can expand a broad, polyclonal repertoire of antiviral T cells, offering a promising approach to HIV immunotherapy.     

Increase in the immunostimulatory effect of dendritic cells by pulsing with serum derived from pancreatic and colorectal cancer patients

Both we and others have observed a relative resistance of solid tumor cells to immunological effector cells in vitro, which may be one reason for the clinical phenomenon of resistance of patients with pancreatic carcinoma or other solid tumors to immunological therapeutic approaches. Dendritic cells (DC) are professional antigen-presenting cells which can process and present tumor-associated antigens such as CA 19-9. Here we tested DC pulsed with serum containing CA 19-9 for their capacity to stimulate immunological effector cells against pancreatic carcinoma cells. Coculture of immunological effector cells with DC led to a significant increase in cytotoxic activity as measured by a lactic dehydrogenase release assay. Most interestingly, cytotoxic activity against tumor cells was further increased using DC pulsed with patient-derived CA 19-9 containing serum. Similar results have been obtained using either autologous or allogeneic serum from patients with pancreas carcinoma. The effect of serum on the cytotoxicity of effector cells increased in a dose-dependent manner. Interestingly, heat inactivation led to a significant loss of immunostimulatory capacity of the serum. Cytotoxicity was partially inhibited by using an antibody directed against CA 19-9 on the surface of the target cells. Best results were obtained when adding CA 19-9 protein to CA 19-9 containing serum for pulsing of DC. In conclusion, DC pulsed with CA 19-9 containing serum increased the cytotoxic activity of immunological effector cells against pancreatic cancer cells. DC pulsed with CA 19-9 containing serum with or without additional exogenous CA 19-9 protein may have an impact on immunotherapeutic protocols for patients with CA 19-9 secreting tumors.     

Isolation of renin-producing human cells by transfection with three simian virus 40 mutants.

A human juxtaglomerular cell (JGC) tumor was used for the immortalization of renin-secreting cells. The transfection of primary JGC with three different simian virus 40 (SV40) mutants resulted in the continuous production of renin-secreting cells. The most efficient renin-producing cells (producing about 400 pg of renin per 24 hr per ml of culture medium) were those transfected with the PAS SV40 mutant. The renin production was stable and the cell cultures have been maintained for greater than 1 year. Two types of cells were cultured together and could not be separated: round and birefringent cells, which exhibited features of mast cells, and elongated cells containing myofilaments and secretory granules. Immunocytochemical staining showed the presence of renin in this latter cell type. The renin produced by the transfected cells was not stored within the cells but was released rapidly into the medium. More than 95% of the renin produced was prorenin, which, after activation, had characteristics similar to those of pure human standard renin as to its enzymatic, immunologic, and biochemical properties, except that it was less glycosylated. These stable JGC tumoral cell lines provide a unique system for studying human renin biosynthesis and its regulation in vitro.     

IL-33 Promotes the Induction and Maintenance of Th2 Immune Responses by Enhancing the Function of OX40 Ligand

Background: In Th2 immune responses, TSLP is a key player by induction of OX40-ligand (OX40L) on dendritic cells (DCs), which is the trigger to induce Th2 cell-mediated allergic cascade. Thus, TSLP-DC-OX40L axis might be the principal pathway in the inflammatory cascades in atopic dermatitis and asthma. IL-33, which is produced by epithelial cells, has been implicated in the Th2 immune responses and pathogenesis of the allergic disorders. However, the role of IL-33 in the Th2-polarizing TSLP-DC-OX40L axis still remains largely elusive. We focused on the ability of IL-33 to promote OX40L-mediated Th2 responses.Methods: Purified human naïve or memory CD4 + T cells were stimulated with recombinant OX40L or TSLP- treated DCs (TSLP-DCs) in the presence of IL-33, and the cytokine production by the primed T cells was examined. We also performed immunohistochemical analyses for the expression of IL-33 in specimens of lymph node and skin from the patients with atopic dermatitis.Results: IL-33 remarkably enhanced TSLP-DCs-driven or OX40L-driven Th2 responses from naïve T cells and the Th2 functional attributes of CRTH2 + CD4 + Th2 memory cells by the increased production of IL-5, IL-9, and IL-13. In addition, IL-33 was expressed in the nuclei of epithelial cells in not only skin lesion but also lymph nodes of the patient with atopic dermatitis, suggesting a specialized role in adaptive T cell-priming phase.Conclusions: IL-33 works as a positive regulator of TSLP-DC-OX40L axis that initiates and maintains the Th2 cell-mediated inflammatory responses, and therefore, it would be a new therapeutic target for the treatment of allergic disorders.     

Expression of lymphocyte-endothelial receptor-ligand pairs, alpha4beta7/MAdCAM-1 and OX40/OX40 ligand in the colon and jejunum of patients with inflammatory bowel disease

BACKGROUND: The interaction between leucocytes and vascular endothelial cells is essential for leucocyte migration into inflammatory sites. AIMS: To study the local expression of the pairs of complementary molecules, alpha4beta7/mucosal addressin cell adhesion molecule (MAdCAM-1) and OX40/OX40 ligand in the lamina propria of the colon and jejunum of patients with inflammatory bowel disease. METHODS: Ten patients with active ulcerative colitis (UC), nine with active Crohn's disease (CD), and seven irritable bowel syndrome (IBS) controls were submitted to endoscopic and peroral jejunal biopsies. Specimens were immunostained by indirect alkaline phosphatase using antibodies against CD3, intercellular adhesion molecule (ICAM) 1, alpha4beta7, MAdCAM-1, and OX40. An OX40-mouse-IgG fusion protein was used to detect OX40 ligand on frozen sections. Immunohistological analysis was carried out by optical microscopy using a computer assisted image analyser. RESULTS: Colonic lamina propria of patients with CD and UC showed increased density of CD3+, alpha4beta7+, and OX40+ cells compared with IBS controls. ICAM-1, MAdCAM-1, and OX40 ligand positive vessels were also increased compared with IBS controls. No significant difference was found in the density of any of these cells in the jejunal mucosa of patients compared with IBS controls. CONCLUSIONS: The expression of MAdCAM-1 and OX40 ligand on gut endothelial and OX40+ cells is increased in sites of mucosal inflammation in patients with inflammatory bowel disease. No evidence was found for increased lamina propria T cells or increased vascular adhesion molecule expression in the proximal intestine of patients with distal inflammatory bowel disease.     

The upregulated expression of OX40/OX40L and their promotion of T cells proliferation in the murine model of asthma

Objective

To investigate whether the expression of OX40/OX40 ligand (OX40L) was upregulated in a murine model of asthma and their significance in the pathogenesis of asthma.

Methods

After an ovalbumin-sensitized/challenged murine model of asthma was established, the expressions of OX40, OX40L in peripheral blood mononuclear cells (PBMCs) and bronchoalveolar lavage fluid (BALF) cell pellets were measured. Then T cell proliferation was analyzed by cell counting kit-8 (CCK8), and the protein levels of OX40 and OX40L in the lungs were determined by immunohistochemistry. The concentrations of IL-4 and IFN-γ in BALF and T cell culture supernatant were evaluated by ELISA.

Results

The percentages of CD4⁺OX40⁺, CD19⁺OX40L⁺, F4/80⁺OX40L⁺ in PBMCs and BALF cell pellets were higher in asthma group than in control group (all P<0.01). The proliferation capacity of T cells in asthma group was higher than that in control group (P<0.05). In asthma group, stimulation of OX40 by anti-OX40 mAb obviously promoted T cell proliferation and secretion of IL-4 and IFN-γ. Immunohistochemistry assay showed that OX40 and OX40L protein levels were higher in asthma group than those in control group (all P<0.05).

Conclusions

The expressions of OX40 and OX40L were upregulated in the murine asthmatic model. The upregulation of OX40/OX40L signals could induce the proliferation and cytokines secretion of T cells in asthmatic mice, indicating that OX40/OX40L signal was involved in the pathogenesis of asthma.     

OX40/OX40L分子在炎症性肠病免疫病理机制中的作用

OX40/OX40L是一对重要的协同刺激分子,它们能增强CD 4~ T细胞的扩增、成熟和产生效应功能,还可使T细胞免于凋亡,并促进记忆性T细胞产生。因此,OX40/OX40L在机体的免疫应答和多种疾病中起重要作用。目前的研究表明免疫异常是炎症性肠病发病的重要因素,此文就OX40/OX40L的生物学功能及其在IBD中的作用作一综述。     

OX40 ligand fusion protein delivered simultaneously with the BCG vaccine provides superior protection against murine Mycobacterium tuberculosis infection

Mycobacterium tuberculosis infection claims approximately 2 million lives per year, and improved efficacy of the BCG vaccine remains a World Health Organization priority. Successful vaccination against M. tuberculosis requires the induction and maintenance of T cells. Targeting molecules that promote T-cell survival may therefore provide an alternative strategy to classic adjuvants. We show that the interaction between T-cell-expressed OX40 and OX40L on antigen-presenting cells is critical for effective immunity to BCG. However, because OX40L is lost rapidly from antigen-presenting cells following BCG vaccination, maintenance of OX40-expressing vaccine-activated T cells may not be optimal. Delivering an OX40L:Ig fusion protein simultaneously with BCG provided superior immunity to intravenous and aerosol M. tuberculosis challenge even 6 months after vaccination, an effect that depends on natural killer 1.1(+) cells. Attenuated vaccines may therefore lack sufficient innate stimulation to maintain vaccine-specific T cells, which can be replaced by reagents binding inducible T-cell costimulators.     

Efficient stimulation of HIV-1-specific T cells using dendritic cells electroporated with mRNA encoding autologous HIV-1 Gag and Env proteins

Infection with human immunodeficiency virus type 1 (HIV-1) is characterized by dysfunction of HIV-1-specific T cells. To control the virus, antigen-loaded dendritic cells (DCs) might be useful to boost and broaden HIV-specific T-cell responses. In the present study, monocyte-derived DCs from nontreated HIV-1-seropositive patients were electroporated with codon-optimized ("humanized") mRNA encoding consensus HxB-2 (hHXB-2) Gag protein. These DCs elicited a strong HIV-1 Gag-specific interferon-gamma (IFN-gamma) response by an HLA-A2-restricted CD8+ T-cell line. Moreover, hHXB-2 gag mRNA-electroporated DCs also triggered IFN-gamma secretion by autologous peripheral blood mononuclear cells (PBMCs), CD4+ T cells, and CD8+ T cells from all patients tested. Next, a novel strategy was developed using autologous virus sequences. Significant specific IFN-gamma T-cell responses were induced in all patients tested by DCs electroporated with patients' autologous polymerase chain reaction (PCR)-amplified and in vitro-transcribed proviral and plasma viral mRNA encoding either Gag or Env. The stimulatory effect was seen on PBMCs, CD8+ T cells, and CD4+ T cells, demonstrating both major histocompatibility complex (MHC) class I and MHC class II antigen presentation. Moreover, a significant interleukin-2 (IL-2) T-cell response was induced by DCs electroporated with hHxB-2 or proviral gag mRNA. These findings open a major perspective for the development of patient-specific immunotherapy for HIV-1 disease.     

Altered peptide ligand vaccination with Flt3 ligand expanded dendritic cells for tumor immunotherapy

Most tumor-associated antigens represent self-proteins and as a result are poorly immunogenic due to immune tolerance. Here we show that tolerance to carcinoembryonic antigen (CEA), which is overexpressed by the majority of lethal malignancies, can be reversed by immunization with a CEA-derived peptide. This peptide was altered to make it a more potent T cell antigen and loaded onto dendritic cells (DCs) for delivery as a cellular vaccine. Although DCs are rare in the blood, we found that treatment of advanced cancer patients with Flt3 ligand, a hematopoietic growth factor, expanded DCs 20-fold in vivo. Immunization with these antigen-loaded DCs induced CD8 cytotoxic T lymphocytes that recognized tumor cells expressing endogenous CEA. Staining with peptide-MHC tetramers demonstrated the expansion of CD8 T cells that recognize both the native and altered epitopes and possess an effector cytotoxic T lymphocyte phenotype (CD45RA(+)CD27(-)CCR7(-)). After vaccination, two of 12 patients experienced dramatic tumor regression, one patient had a mixed response, and two had stable disease. Clinical response correlated with the expansion of CD8 tetramer(+) T cells, confirming the role of CD8 T cells in this treatment strategy.     

rAAV-AFP转染人外周血单核细胞来源树突状细胞增强免疫刺激功能

目的:探讨表达甲胎蛋白抗原的重组腺相关病毒rAAV-AFP转染对人外周血单核细胞来源树突状细胞(DC)免疫刺激功能的影响．方法:用rAAV-AFP转染新分离的DC;苔盼蓝拒染法检测每天的活细胞比率;流式细胞仪检测DC表面分子CD80,CD86,CD83,CD40, CD1a,HLA-DR及AFP的表达;并用3H-TdR掺入法检测转染前后DC刺激自体淋巴细胞增殖能力;MTT法检测DC诱导T细胞的杀伤活性．结果:成熟DC 77．7％表达AFP蛋白;转染对活细胞百分率和成熟DC表型无影响,与未转染组无显著差异(P>0．05);转染后DC仍保持较强的刺激自体淋巴细胞增殖的能力,与未转染组也无显著性差异(P>0．05),并且可诱导出特异性杀伤,效应细胞和靶细胞为80:1,40:1,20:1 时,与未转染组相比均有显著差异(35．5±5．5 vs 20．6±4．7;28．7±3．6 vs 15.3±2．5;16．2±2．8 vs 9．6±1．8;均P<0．01)．结论:rAAV可负载AFP基因在DC中表达, rAAV-AFP转染DC对其功能无明显影响,免疫功能更强．     

共刺激分子OX40-OX40L相互作用对ApoE-/-小鼠淋巴细胞活化T细胞核因子C1表达的影响

目的 探讨共刺激分子OX40-OX40L相互作用对载脂蛋白E基因敲除(ApoE-/-)小鼠淋巴细胞中活化T细胞核因子C1(NFATc1)表达的影响.方法 采用未经治疗的ApoE-/-小鼠颈动脉粥样斑块模型,取其脾脏淋巴细胞,以不同浓度刺激型OX40抗体或抑制型OX40L抗体体外特异性刺激或阻断OX40-OX40L相互作用,不同时间点收获细胞,分别应用实时荧光定量PCR和流式细胞技术检测小鼠淋巴细胞NFATc1 mRNA和蛋白表达.结果 (1)抗OX40抗体特异性刺激OX40-OX40L轴后,小鼠淋巴细胞NFATc1 mRNA及蛋白表达明显上调,anti-OX40浓度为20 μg/ml时刺激作用最强,刺激时间24 h最佳(P<0.01).(2)抗OX40L抗体特异性阻断OX40-OX40L轴能明显抑制NFATc1 mRNA及蛋白表达,anti-OX40L为20 μg/ml时抑制作用最强,抑制时间24 h最佳(P<0.001).结论 OX40-OX40L相互作用能调控ApoE-/-小鼠淋巴细胞NFATc1的表达.

Abstract：

Objective To investigate the effect of OX40/OX40L interaction on the nuclear factor of activated T cells c1(NFATc1) in ApoE-/- mice.Methods Lymphocytes were prepared from mouse spleens after Collar-treated Surgery, then incubated with a range of agonistic anti-OX40 mAbs and inhibitory anti-OX40L mAb to stimulate or inhibit OX40-OX40L interaction in vitro. The expression of NFATc1 mRNA and protein in lymphocytes of ApoE-/- mice was measured by Real Time PCR and flow cytometry, respectively.Results (1)After stimulating OX40-OX40L signal pathway, the expression of NFATc1 mRNA and protein in leukocytes of ApoE-/- mice was significantly increased, with maximal effect occurring at 20 μg/ml anti-OX40 mAb- stimulated, and peaked at 24 h at any concentration(P<0.01). (2)Anti-OX40L mAb significantly suppressed the expression of NFATc1 in leukocytes of ApoE-/- mice, with maximal effect occurring at 20 μg/ml anti-OX40L mAb, and peaked at 24 h(P<0.001).ConclusionsOX40-OX40L interaction can regulate the mRNA and protein expressions of NFATc1 in lymphocytes of ApoE-/- mice.     

Functional defects of dendritic cells in patients with CD40 deficiency

We have recently identified 2 patients with a rare autosomal recessive form of hyper IgM disease, known as HIGM3, caused by mutations in the CD40 gene. These patients had opportunistic infections observed on X-linked hyper IgM syndrome (HIGM), suggesting that the CD40-CD40 ligand interaction is important for promoting T-cell-mediated immunity. To evaluate whether innate immunity signals may substitute CD154 for inducing the maturation of dendritic cells (DCs), we analyzed monocyte-derived DCs in these patients. Monocyte-derived DCs of HIGM3 subjects on ex vivo stimulation with tumor necrosis factor-alpha (TNF-alpha) or lipopolysaccharide (LPS) combined with interferon-gamma (IFN-gamma) normally express all the markers of mature DCs, such as CD83 and DC-LAMP. However, cell surface levels of HLA-DR in mature DCs are reduced, as is costimulatory activity of these cells for allogeneic naive T cells. In addition, CD40-deficient DCs secrete lower amounts of interleukin-12 (IL-12) but larger quantities of IL-10 than control subjects. Finally, analysis of circulating plasmacytoid DCs demonstrates a normal percentage of this subset in CD40-deficient cells, but IFN-alpha secretion in response to herpes simplex virus 1 (HSV-1) infection is severely reduced in patients. These observations suggest that the severe impairment of DC maturation may contribute to the defect of T-cell-mediated immunity observed in HIGM3 patients.