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PubMed2002

DC-SIGN, a C-type lectin on dendritic cells that unveils many aspects of dendritic cell biology.

Dendritic cells (DC) are present in essentially every tissue where they operate at the interface of innate and acquired immunity by recognizing pathogens and presenting pathogen-derived peptides to T cells. It is becoming clear that not all C-type lectins on DC serve as antigen receptors recognizing pathogens through carbohydrate structures. The C-type lectin DC-SIGN is unique in that it regulates adhesion processes, such as DC trafficking and T-cell synapse formation, as well as antigen capture. Moreover, even though several C-type lectins have been shown to bind HIV-1, DC-SIGN does not only capture HIV-1 but also protects it in early endosomes allowing HIV-1 transport by DC to lymphoid tissues, where it enhances trans infection of T cells. Here we discuss the carbohydrate/protein recognition profile and other features of DC-SIGN that contribute to the potency of DC to control immunity.

Animals↗
PubMed2004

[Preparation of dendritic cell tumor-vaccine with dendritic cells and problems in its clinical uses--review].

Dendritic cell (DC), the most powerful antigen presenting cell, has been paid close attention to tumor immunotherapy. The tumor antigens were extracted by different ways to pulse DC, and then the mature DC were infused to patients to accelerate anti-tumor immuno responses. Its clinical use would provide a new therapeutic pathway other than surgical operation, irradiation, chemotherapy. In this paper, biological characteristics of DC, tumor-vaccine preparation and its clinical use were reviewed.

Cancer Vaccines↗
PubMedDate not supplied

Mature dendritic cells are superior to immature dendritic cells in expanding antigen-specific naive and memory CD8+ T cells.

BACKGROUND: For successful dendritic cell (DC)-based immunotherapy, it is critical to identify the most potent stage of human DCs, including immature DCs (imDCs) and mature DCs (mDCs). MATERIALS AND METHODS: imDCs were obtained by culturing monocytes in the presence of GM-CSF and IL-4 for 5- 7 days and imDCs were further cultured for 24-48 h in the presence of TNFalpha, IL-6, IL-1beta and PGE2 to obtain mDCs. Melan-A- and EBV (BRF1) peptides were used and the frequency of antigen-specific CD8+ T cells was assessed using appropriate tetramers. RESULTS: mDCs were potent antigen-presenting cells for the induction and proliferation of antigen-specific naive and memory CD8+ T cells and may overcome regulatory functions that suppress antigen-specific CD8+ T cells. CONCLUSION: Our findings that mDCs can efficiently expand antigen-specific naive and memory CD8 + T cells have important implications in the development of vaccination strategies and support the use of antigen-loaded mature DCs in human clinical trials

Antigen Presentation↗
PubMed2006

Nonviral transfection of human umbilical cord blood dendritic cells is feasible, but the yield of dendritic cells with transgene expression limits the application of this method in cancer immunotherapy.

Dendritic cells (DC) generated from human umbilical cord blood might replace patients' DC in attempts to elicit tumor-specific immune response in cancer patients. We studied the efficiency of transfection of human cord blood DC with plasmid DNA carrying the enhanced version of green fluorescent protein (EGFP) as a reporter gene, to test if nonviral gene transfer would be a method to load DC with protein antigens for immunotherapy purposes. Cord blood mononuclear cells were cultured in serum-free medium in the presence of granulocyte-monocyte colony stimulating factor (GM-CSF), stem cell factor (SCF) and Flt-3 ligand (FL), to generate DC from their precursors, and thereafter transfected by electroporation. Maturation of DC was induced by stimulation with GM-CSF, SCF, FL and phorbol myristate acetate (PMA). Transfected DC strongly expressed EGFP, but transfection efficiency of DC, defined as HLA-DR(+) cells lacking lineage-specific markers, did not exceed 2.5%. Expression of the reporter gene was also demonstrated in the DC generated from transfected, purified CD34(+) cord blood cells, by stimulation with GM-CSF, SCF, FL, and tumor necrosis factor alpha (TNF-alpha). Transfection of CD34(+) cells was very efficient, but proliferation of the transfected cells was much reduced as compared to the untransfected cells. Therefore, the yield of transgene-expressing DC was relatively low. In conclusion, nonviral transfection of cord blood DC proved feasible, but considering the requirements for immunotherapy in cancer patients, transfection of differentiated DC or generation of DC from transfected hematopoietic stem cells provide only a limited number of DC expressing the transgene.

Antigens, CD34↗
PubMed2006

Dendritic cell-tumor cell hybrids enhance the induction of cytotoxic T lymphocytes against murine colon cancer: a comparative analysis of antigen loading methods for the vaccination of immunotherapeutic dendritic cells.

Dendritic cells (DCs) have been used successfully for inducing effective anti-tumor immune responses in advanced cancer patients undergoing tumor-specific immunotherapy. Appropriate antigen pulsing is a crucial parameter for optimizing the efficacy of immunotherapy as well as anti-tumor protection therapy. Using a murine colon cancer model, we evaluated the anti-tumor efficacy of four different preparations of DC vaccines that contained either a whole tumor or its derivatives, including i) DCs pulsed with tumor lysate, ii) DCs pulsed with necrotic tumor cells, iii) DCs pulsed with apoptotic tumor cells, and iv) DC-tumor cell fusion hybrids. Our data show that DC-tumor cell fusion hybrids and DCs pulsed with irradiated apoptotic tumor cells were more potent than DCs with freeze-thawed necrotic tumor cells for the induction of protective anti-tumor responses. The vaccination of DCs pulsed with tumor lysate failed to elicit any anti-tumor effect. In animals administered with higher doses of a tumor-cell challenge, DC-tumor cell fusion hybrids elicited the most effective anti-tumor response. Among the preparations tested, mice immunized with DC-tumor cell fusion hybrids resulted in the greatest induction of cytotoxicity as measured by the cytotoxic T lymphocyte activity of both the splenocytes and the Thy1.2-positive T lymphocytes. Furthermore, the in vitro production of IFN-gamma polarized to the Th1 cytokine responses was highest in the splenocytes derived from mice vaccinated with DC-tumor cell fusion hybrids. Our results suggest that DC-tumor cell fusion hybrids are more potent inducers of protection against solid tumors, such as colon cancer, than other antigen-loading strategies using whole tumor cell materials.

Animals↗
PubMed1990

HIV-1 infection and virus production in follicular dendritic cells in lymph nodes. A case report, with analysis of isolated follicular dendritic cells.

Follicular dendritic cells (FDC) from axillary lymphoid tissue of a patient with acquired immune deficiency syndrome (AIDS) were analyzed for the presence of gag and env proteins and env mRNA of human immunodeficiency virus type-1 (HIV-1), both in a purified FDC suspension and on frozen sections. Isolated cells with morphologic and immunocytochemical features of FDC expressed HIV-1 core (gag) proteins p15, p17, p24, and envelope (env) protein gp41; in addition HIV-1 env mRNA was detected in some of these cells. This corresponded with intense expression of HIV-1 proteins by FDC in germinal centers in situ, and the presence of HIV-1 mRNA-positive cells in germinal follicles. These findings led us to conclude that FDC are infected and able to produce HIV-1. Such infection may contribute significantly to the destruction of the FDC network during the lymphadenopathy phase after HIV-1 infection.

Acquired Immunodeficiency Syndrome↗
PubMed1989

The role of dendritic cells as stimulators of minor lymphocyte-stimulating locus-specific T cell responses in the mouse. I. Differential capacity of dendritic cells to stimulate minor lymphocyte-stimulating locus-reactive T cell hybridomas and the primary anti-minor lymphocyte-stimulating locus mixed lymphocyte reaction.

The response of T cells to minor lymphocyte-stimulating locus (Mls) determinants remains poorly understood with respect to the antigenic determinants responsible for T cell stimulation and the types of APC capable of stimulating the response. In this report, we demonstrate that highly purified dendritic cells (DC) as well as B cells have the capacity to stimulate Mls-specific responses. Unseparated spleen cells, purified DC, resting B cells, and activated B cells were compared for their capacity to stimulate several Mls-reactive T cell hybridomas. Whereas the entire panel of Mls-reactive T cell hybridomas was stimulated strongly by unseparated spleen cells and activated B cells, the hybridomas responded only weakly to purified DC or resting B cells. Activation of resting B cells with either B cell stimulatory factor-1 (1 day pre-treatment) or LPS/dextran (2 or 3 day pre-treatment) greatly augmented their Mls-stimulatory capacity. In contrast, the Mls-stimulatory capacity of DC was not augmented by a 1-day pre-treatment with either B cell stimulatory factor-1 or supernatant from the DC-induced primary anti-Mls-MLR. In the primary anti-Mls-MLR, both purified DC and LPS/dextran-stimulated B blasts were found to elicit vigorous T cell proliferative responses. Much weaker responses were elicited by unseparated spleen cells. The stimulation of the primary anti-Mls-MLR by purified DC was further confirmed by producing Mls-specific T cell clones which were preferentially stimulated by DC. Autologous (Mlsb) DC were found to markedly enhance the primary anti-Mls-MLR response to small numbers of Mlsa B blasts. Thus, DC possess other "accessory cell" properties that augment the primary anti-Mls-MLR despite the predicted low level of Mls determinant expression on DC based on the results obtained with Mls-reactive hybridomas. Possible accessory cell properties of DC relevant to this phenomenon are discussed.

Animals↗
PubMed1995

Endotoxin-mediated dendritic cell release from the intestine. Characterization of released dendritic cells and TNF dependence.

Dendritic cells (DC) acquire Ag in peripheral tissues and transport it to lymph nodes where they efficiently activate resting T cells. We have shown that i.v. endotoxin causes increased release of intestinal DC into lymph. In this paper we further characterize the release of DC and the properties of the released cells. A total of 50 micrograms of endotoxin injected i.v. causes an increase in DC output within 6 h that peaks between 12 and 24 h, with a maximum output of 8 to 15 times normal. At the same time lymphocyte output is markedly decreased. The increased output of DC is followed by a decrease to subnormal levels. The stimulated release of DC is almost totally blocked by a monoclonal anti-TNF-alpha Ab. A second injection of TNF-alpha does not result in further DC release. DC are not released from lymph nodes into efferent lymph by endotoxin. DC collected from lymph after endotoxin treatment show increased expression of the p55 IL-2 receptor and the OX48 Ag but otherwise resemble normal lymph DC. In functional assays they show no significant differences from normal in their ability to stimulate a MLR or to present Ags to sensitized T cells. Immunocytochemistry with the use of MRC OX62 suggests that the DC are released into lymph from the lamina propria of the small intestine. The stimulated release of DC mediated by TNF-alpha may be important in regulating Ag presentation in lymph nodes draining inflammatory sites.

Animals↗
PubMed1993

Identification of a novel population of CD8 alpha+ beta- bone marrow-derived dendritic epidermal cells. This unique population is subsequently outnumbered by thymus-independent expansion of the CD8 alpha+ beta+ dendritic epidermal cells.

Maturation of some T cell populations has been suggested to occur in epithelial tissues. The CD8 molecule, which is expressed as heterodimers made of alpha/beta-chains on virtually all peripheral T cells, is preferentially expressed as homodimers made only of alpha-chains on a subset of CD8+ mouse gut intraepithelial lymphocytes, which arise from bone marrow (BM) precursors in a thymus-independent mechanism. This unique population of CD8 alpha+ beta- T cells, however, has not been identified in other lymphoid tissues in normal adult mice. Because our previous study demonstrated that reconstitution of thymectomized irradiated mice with T cell-depleted BM cells resulted in the appearance of CD8+, TCR-alpha beta+ dendritic epidermal cells (DEC) and suggested that a proportion of the BM-derived DEC may mature within the epidermis, we asked whether the unique phenotype of CD8 alpha+ beta- could be detected among the CD8+ BM-derived DEC. In this report for the first time we identified this unique population of CD8 alpha+ beta- cells among the DEC. Although this population comprised the predominant fraction of the BM-derived DEC in the early post-transfer period (2 to 3 mo), gradual shift from the CD8 alpha+ beta+ phenotypes occurred during the late post-transfer period (4 to 6 mo) independently of the presence of the thymus. Kinetic studies on the BM-derived DEC revealed that this phenotypic shift could be caused by the subsequent expansion of the CD8 alpha+ beta+ DEC. In contrast, the CD8 alpha+ beta- population was the major subset of the BM-derived intraepithelial lymphocytes throughout the entire observation period and the phenotypic shift with age was never observed. These results indicate that CD8 alpha+ beta- cells are preferentially detected on T cells that home to the epithelial tissues and that in the epidermis, but not in the gut epithelia, the subsequent expansion of the CD8 alpha+ beta+ DEC would dilute the high number of the CD8 alpha+ beta- cells.

Animals↗
PubMed1996

In vitro differentiation of CD34+ hematopoietic progenitor cells toward distinct dendritic cell subsets of the birbeck granule and MIIC-positive Langerhans cell and the interdigitating dendritic cell type.

We have demonstrated recently that Birbeck granule-positive Langerhans cells (LC) can be derived from CD34+ peripheral blood progenitor cells in the presence of a seven-cytokine cocktail (CC7-7). Here, we show that the sequential use of early-acting hematopoietic growth factors, stem cell factor, interleukin (IL)-3, and IL-6, followed on day 8 by differentiation in the two-factor combination IL-4 plus granulocytemacrophage colony-stimulating factor (GM-CSF) (CC4GM) is more efficient and allows the cells to be arrested in the LC stage for more than 1 week while continuous maturation occurs in CC7-7. Maturation of LC to interdigitating dendritic cells (DC) could specifically be induced within 60 hours by addition of tumor necrosis factor-alpha (20 ng/mL) or lipopolysaccharide (100 ng/mL). Using LC that had been enriched to greater than 90% CD1a+ cells by an immunoaffinity column, we were able to define clear-cut differences between LC and DC that corroborate data of the respective cells derived from epithelial borders (LC) or from lymph nodes (LN) and spleen (DC). Thus, molecules and functions involved in antigen (AG) uptake and processing were highly expressed in LC, while those involved in AG presentation were at maximum in DC. LC were CD1a+2 DR+2, CD23+, CD36+, CD80-, CD86-, and CD25-, while DC were CD1a+/- DR+3, CD23-, CD36-, CD80+, CD86+2, and CD25+, CD40 and CD32 were moderately expressed and nearly unchanged on maturation, in contrast to monocyte-derived DC. Macropinocytosis of fluorescein isothiocyanate-dextran was dominant in LC, as were multilamellar major histocompatibility complex (MHC) class II compartments (MIICs), which were detected by electron microscopy. The functional dichotomy of these cell types was finally supported by testing the AG-presenting cell function for tetanus toxoid to primed autologous T-cell lines, which was optimal when cells were loaded with AG as LC and subsequently induced to become DC.

Antigen Presentation↗
PubMed1996

Induction of nitric oxide synthase in mouse dendritic cells by IFN-gamma, endotoxin, and interaction with allogeneic T cells: nitric oxide production is associated with dendritic cell apoptosis.

Nitric oxide (NO) is an important effector molecule that is involved in immune regulation and host defense. In this study, highly purified NLDC 145+ (DEC-205+) MHC class II(bright) B7-2+ dendritic cells (DC) propagated from normal mouse bone marrow in response to granulocyte-macrophage CSF + IL-4 were induced to produce NO by IFN-gamma and LPS. NO production was inhibited by the nitric oxide synthase (NOS) inhibitor N(G)-monomethyl-L-arginine (NMMA). Nitrite also accumulated in mixed leukocyte culture supernatants as the result of coculture of DC with purified naive allogeneic T cells. Furthermore, NO production was induced by CD40 ligation. Suboptimal T cell proliferation observed at high relative concentrations of DC correlated with increased NO production and was mitigated by NMMA. Induction of mRNA for an inducible NOS (iNOS) in DC was confirmed by Northern blotting, whereas intracellular iNOS was visualized by two-color flow cytometry and by both immunofluorescent and immunogold labeling in a subpopulation of IFN-gamma + LPS-stimulated cells. Both endogenous NO production and exposure of unstimulated DC to the NO donor S-nitroso-N-acetyl-penicillamine (SNAP) resulted in DC apoptosis. Thus, although DC function initially as the most potent APCs for T cell activation, DC induced to synthesize NOS by IFN-gamma may inhibit (allogeneic) T cell proliferation: NO may suppress lymphocyte proliferation and also induce apoptosis of the most potent source of alloantigenic stimulation.

Animals↗
PubMed1996

Functional comparison of spleen dendritic cells and dendritic cells cultured in vitro from bone marrow precursors.

We have compared dendritic cells (DC) isolated from mouse spleen, or generated in vitro from bone marrow (BM) precursors cultured in granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4), for the ability to process and present soluble antigen and stimulate major histocompatibility complex (MHC) Class II-restricted T cells. DC from spleen or BM cultures were equally able to stimulate the in vitro proliferation of allogeneic T cells or of antigen-specific T-cell receptor (TCR)-transgenic T cells. Both DC populations also induced comparable levels of IL-2 secretion by a T-cell hybridoma. Therefore, splenic and BM-derived DC express comparable levels of (Antigen + MHC Class II) ligands and/or costimulatory molecules and have comparable ability to stimulate T-cell responses. When presentation of a native protein antigen, rather than peptide, was evaluated, BM-derived DC were at least 50 times better than splenic DC at stimulating the proliferation of TCR-transgenic T cells. The antigen processing ability of the two populations was similar only when splenic DC were used immediately ex vivo. Therefore, unlike spleen DC, BM-derived DC maintain the capacity to process protein antigen for MHC Class II presentation during in vitro culture. Due to these characteristics, BM-derived DC may represent a useful tool in immunotherapy studies, as they combine high T-cell stimulatory properties with the capacity to process and present native antigen.

Amino Acid Sequence↗
PubMed1997

Developmental pathways of dendritic cells in vivo: distinct function, phenotype, and localization of dendritic cell subsets in FLT3 ligand-treated mice.

We have recently shown that Flt3 ligand administration dramatically increases dendritic cell (DC) numbers in various mouse tissues. This has enabled the identification of distinct mature DC subpopulations. These have been designated: population C (CD11c(bright) CD11b(bright)), D (CD11c(bright) CD11b(dull)), and E (CD11c(bright) CD11b(negative)) This report demonstrates that the mature DC subsets (C, D, and E) from Flt3 ligand-treated mice differ with respect to phenotype, geographic localization, and function. The myeloid Ags CD11b, F4/80, and Ly-6C are predominantly expressed by population C, but not D or E. In addition, a subset of population C-type DC expresses 33D1 and CD4. In contrast, DC within population D and E selectively express the lymphoid-related DC markers CD8alpha, DEC 205, CD1d, as well as CD23, elevated levels of CD117 (c-kit), CD24 (HSA), CD13, and CD54. Immunohistology indicates that the different DC subsets reside in distinct microenvironments, with populations D and E residing in the T cell areas of the white pulp, while DC within population C localize in the marginal zones. These DC subpopulations showed different capacities to phagocytose FITC-zymosan and to secrete IL-12 upon stimulation with Staphylococcus aureus cowan I strain + IFN-gamma + granulocyte-macrophage-CSF. Population C-type DC were more phagocytic but secreted little inducible IL-12 while population D- and E-type DC showed poor phagocytic capacity and secreted considerably higher levels of IL-12. These results underscore the importance of viewing DC development in vivo, as an interplay between distinct lineages and a maturational dependence on specific microenvironmental signals.

Animals↗
PubMed1998

The role of tumor necrosis factor alpha in modulating the quantity of peripheral blood-derived, cytokine-driven human dendritic cells and its role in enhancing the quality of dendritic cell function in presenting soluble antigens to CD4+ T cells in vitro.

Because dendritic cells (DC) are critically involved in both initiating primary and boosting secondary host immune responses, attention has focused on the use of DC in vaccine strategies to enhance reactivity to tumor-associated antigens. We have reported previously the induction of major histocompatibility complex class II-specific T-cell responses after stimulation with tumor antigen-pulsed DC in vitro. The identification of in vitro conditions that would generate large numbers of DC with more potent antigen-presenting cell (APC) capacity would be an important step in the further development of clinical cancer vaccine approaches in humans. We have focused attention on identifying certain exogenous cytokines added to DC cultures that would lead to augmented human DC number and function. DC progenitors from peripheral blood mononuclear cells (PBMC) were enriched by adherence to plastic, and the adherent cells were then cultured in serum-free XVIVO-15 medium (SFM) for 7 days with added granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4). At day 7, cultures contained cells that displayed the typical phenotypic and morphologic characteristics of DC. Importantly, we have found that the further addition of tumor necrosis factor alpha (TNFalpha) at day 7 resulted in a twofold higher yield of DC compared with non-TNFalpha-containing DC cultures at day 14. Moreover, 14-day cultured DC generated in the presence of TNFalpha (when added at day 7) demonstrated marked enhancement in their capacity to stimulate a primary allogeneic mixed leukocyte reaction (8-fold increase in stimulation index [SI]) as well as to present soluble tetanus toxoid and candida albicans (10- to 100-fold increases in SI) to purified CD4+ T cells. These defined conditions allowed for significantly fewer DC and lower concentrations of soluble antigen to be used for the pulsing of DC to efficiently trigger specific T-cell proliferative responses in vitro. When compared with non-TNFalpha-supplemented cultures, these DC also displayed an increased surface expression of CD83 as well as the costimulatory molecules, CD80 and CD86. Removal of TNFalpha from the DC cultures after 2 or 4 days reduced its enhancing effect on DC yield, phenotype, and function. Thus, the continuous presence of TNFalpha over a 7-day period was necessary to achieve the maximum enhancing effect observed. Collectively, our findings point out the importance of exogenous TNFalpha added to cultures of cytokine-driven human DC under serum-free conditions, which resulted in an enhanced number and function of these APC. On the basis of these results, we plan to initiate clinical vaccine trials in patients that use tumor-pulsed DC generated under these defined conditions.

Antigen Presentation↗
PubMed1998

Neutralization of tumor necrosis factor activity shortly after the onset of dendritic cell hematopoiesis reveals a novel mechanism for the selective expansion of the CD14-dependent dendritic cell pathway.

The CD14-dependent and -independent dendritic cell (DC) pathways are instituted simultaneously when CD34(+) progenitor cells are treated with granulocyte-macrophage colony-stimulating factor (GM-CSF)/tumor necrosis factor (TNF) +/- stem cell factor (SCF) (GTS). If TNF activity is neutralized within 48 hours of cytokine exposure, DC development is halted and myelogranulocytic hematopoiesis takes place. In this study, we show that disruption of TNF activity at a later time point produced a distinct alteration within the DC system. Instead of downregulating DC development, treatment of GTS cultures with antibodies to TNF (anti-TNF) on day 3 provoked the selective expansion of the CD14-dependent (monocyte) DC pathway from progenitor cell populations lacking CD14 and CD1a. After an initial decrease in proliferation, anti-TNF produced a rebound in cell growth that yielded intermediate myeloid progenitors exhibiting CD14-dependent DC differentiation potential and CD14(+)CD1a+ DC precursors. Cultures enriched in CD14-dependent DCs were more potent stimulators of a mixed leukocyte reaction, compared with control GTS cultures containing both types of DCs. The intermediate progenitors expanded in the presence of anti-TNF were CD115(+)CD33(+)DR+, long-lived, and displayed clonogenic potential in methylcellulose. When exposed to the appropriate cytokine combinations, these cells yielded granulocytes, monocytes, and CD14-dependent DCs. Antigen-presenting function was acquired only when DC maturation was induced from these myelodendritic progenitors with GM-CSF + interleukin-4 or GTS. These studies show a novel mechanism by which TNF regulates the DC system, as well as providing a strategy for the amplification of the CD14-dependent DC pathway from immature progenitors. Although TNF is required to ensure the institution of DC hematopoiesis from CD34(+) progenitor cells, its activity on a later progenitor appears to limit the development of CD14-dependent DCs.

Antibodies, Monoclonal↗
PubMed1998

Prostaglandin E2 induces the final maturation of IL-12-deficient CD1a+CD83+ dendritic cells: the levels of IL-12 are determined during the final dendritic cell maturation and are resistant to further modulation.

Activation of immature dendritic cells (DC) in peripheral tissues induces their migration to lymph nodes and their maturation into CD83+ DC, which are able to prime naive T cells. The inflammatory cytokines IL-1beta and TNF-alpha induce mature DC, which can secrete IL-12 and promote the development of Th0/Th1-biased cells. DC maturation factors with a Th2-promoting function have not been described. Here we show that PGE2, although it does not induce final DC maturation by itself, synergizes with IL-1beta and TNF-alpha, and allows their effectiveness at 100-fold lower concentrations. While being phenotypically identical with the DC matured in the presence of high concentrations of IL-1beta and TNF-alpha alone, DC matured in the additional presence of PGE2 show impaired IL-12 production and bias naive Th cell development toward the Th2. The ability of DC to produce IL-12 is also suppressed by IL-10, which in contrast to PGE2, inhibits their maturation. The differences in the ability to produce IL-12, established during the final DC maturation, are stable after the removal of modulatory factors. Importantly, fully mature DC become unsusceptible to PGE2 and IL-10. This indicates that the levels of IL-12 production in vivo, in mature DC interacting with Th cells within the lymph nodes, are mainly predetermined at the stage of immature DC in peripheral tissues. These data imply that the character of pathogen-induced local inflammatory reaction can "instruct" local DC to initiate Th1 or Th2-biased responses.

Adjuvants, Immunologic↗
PubMed1998

Activation of cutaneous dendritic cells by CpG-containing oligodeoxynucleotides: a role for dendritic cells in the augmentation of Th1 responses by immunostimulatory DNA.

Genetic vaccination depends at least in part on the adjuvant properties of plasmids, properties that have been ascribed to unmethylated CpG dinucleotides in bacterial DNA. Because dendritic cells (DC) participate in the T cell priming that occurs during genetic vaccination, we reasoned that CpG-containing DNA might activate DC. Thus, we assessed the effects of CpG oligodeoxynucleotides (CpG ODN) on Langerhans cell (LC)-like murine fetal skin-derived DC (FSDDC) in vitro and on LC in vivo. Treatment with CpG ODN as well as LPS induced FSDDC maturation, manifested by decreased E-cadherin-mediated adhesion, up-regulation of MHC class II and costimulator molecule expression, and acquisition of enhanced accessory cell activity. In contrast to LPS, CpG ODN stimulated FSDDC to produce large amounts of IL-12 but only small amounts of IL-6 and TNF-alpha. Injection of CpG ODN into murine dermis also led to enhanced expression of MHC class II and CD86 Ag by LC in overlying epidermis and intracytoplasmic IL-12 accumulation in a subpopulation of activated LC. We conclude that immunostimulatory CpG ODN stimulate DC in vitro and in vivo. Bacterial DNA-based vaccines may preferentially elicit Th1-predominant immune responses because they activate and mobilize DC and induce them to produce large amounts of IL-12.

Adjuvants, Immunologic↗
PubMed1998

Dendritic cell-based vaccines in the setting of peripheral blood stem cell transplantation: CD34+ cell-depleted mobilized peripheral blood can serve as a source of potent dendritic cells.

We are investigating the use of tumor-pulsed dendritic cell (DC)-based vaccines in the treatment of patients with advanced cancer. In the current study, we evaluated the feasibility of obtaining both CD34+ hematopoietic stem/ progenitor cells (HSCs) and functional DCs from the same leukapheresis collection in adequate numbers for both peripheral blood stem cell transplantation (PBSCT) and immunization purposes, respectively. Leukapheresis collections of mobilized peripheral blood mononuclear cells (PBMCs) were obtained from normal donors receiving granulocyte colony-stimulating factor (G-CSF) (for allogeneic PBSCT) and from intermediate grade non-Hodgkin's lymphoma or multiple myeloma patients receiving cyclophosphamide plus G-CSF (for autologous PBSCT). High enrichment of CD34+ HSCs was obtained using an immunomagnetic bead cell separation device. After separation, the negative fraction of mobilized PBMCs from normal donors and cancer patients contained undetectable levels of CD34+ HSCs by flow cytometry. This fraction of cells was then subjected to plastic adherence, and the adherent cells were cultured for 7 days in GM-CSF (100 ng/ml) and interleukin 4 (50 ng/ml) followed by an additional 7 days in GM-CSF, interleukin 4, and tumor necrosis factor alpha (10 ng/ml) to generate DCs. Harvested DCs represented yields of 4.1+/-1.4 and 5.8+/-5.4% of the initial cells plated from the CD34+ cell-depleted mobilized PBMCs of normal donors and cancer patients, respectively, and displayed a high level expression of CD80, CD86, HLA-DR, and CD11c but not CD14. This phenotypic profile was similar to that of DCs derived from non-CD34+ cell-depleted mobilized PBMCs. DCs generated from CD34+ cell-depleted mobilized PBMCs elicited potent antitetanus as well as primary allogeneic T-cell proliferative responses in vitro, which were equivalent to DCs derived from non-CD34+ cell-depleted mobilized PBMCs. Collectively, these results demonstrate the feasibility of obtaining both DCs and CD34+ HSCs from the same leukapheresis collection from G-CSF-primed normal donors and cancer patients in sufficient numbers for the purpose of combined PBSCT and immunization strategies.

Antigens, CD34↗