Maturation State Of Dendritic Cells Research Articles

Single-Step Antigen Loading and Activation of Dendritic Cells by mRNA Electroporation for the Purpose of Therapeutic Vaccination in Melanoma Patients

A critical factor determining the effectiveness of currently used dendritic cell (DC)-based vaccines is the DC activation or maturation status. We have recently shown that the T-cell stimulatory capacity of DCs pulsed with tumor-antigen-derived peptides can be considerably increased by activating the DCs through electroporation with mRNA encoding CD40 ligand, CD70, and a constitutively active Toll-like receptor 4 (TriMix DCs). Here, we investigate whether TriMix DCs can be coelectroporated with whole tumor-antigen-encoding mRNA. The T-cell stimulatory capacity of TriMix DCs pulsed with the immunodominant MelanA-A2 peptide and that of TriMix DCs coelectroporated with MelanA mRNA were compared in vitro. TriMix DCs were also coelectroporated with mRNA encoding Mage-A3, Mage-C2, tyrosinase, or gp100. The capacity of these DCs to stimulate tumor-antigen-specific T cells in melanoma patients was investigated both in vitro before vaccination and after DC vaccination. Like peptide-pulsed TriMix DCs, TriMix DCs coelectroporated with MelanA mRNA are very potent in inducing MelanA-specific CD8(+) T cells in vitro. These T cells have an activated phenotype, show cytolytic capacity, and produce inflammatory cytokines in response to specific stimulation. TriMix DCs coelectroporated with tyrosinase are able to stimulate tyrosinase-specific CD8(+) T cells in vitro from the blood of nonvaccinated melanoma patients. Furthermore, TriMix DCs coelectroporated with Mage-A3, Mage-C2, or tyrosinase are able to induce antigen-specific CD8(+) T cells through therapeutic DC vaccination. TriMix DCs coelectroporated with whole tumor-antigen mRNA stimulate antigen-specific T cells in vitro and induce antigen-specific T-cell responses in melanoma patients through vaccination. Therefore, they represent a promising new approach for antitumor immunotherapy.

Dendritic Cell Adhesion to Cerebral Endothelium: Role of Endothelial Cell Adhesion Molecules and Their Ligands

Dendritic cells (DCs) have been increasingly implicated in the pathogenesis of neuroinflammation, and there is evidence that they are recruited to the brain across the blood-brain barrier. The molecular mechanisms mediating DC trafficking to the central nervous system are poorly understood. This study used an in vitro model of the human blood-brain barrier and monocyte-derived DCs to investigate the role of endothelial cell (EC) adhesion molecules and their ligands in the adhesion of immature and mature DCs to cerebral microvascular ECs. Adhesion of DCs to resting brain ECs was minimal, but activation of ECs with tumor necrosis factor significantly upregulated adhesion. Immature DCs adhered to activated ECs more avidly than mature DCs; this correlated with differences in the expression of adhesion molecule ligands between the mature and immature DCs. Blocking studies indicated that adhesion to cytokine-activated blood-brain barrier endothelium is mediated by intercellular adhesion molecule (ICAM)-1, ICAM-2, platelet-EC adhesion molecule (PECAM)-1, vascular cell adhesion molecule 1, CD18, and DC-specific ICAM-3-grabbing nonintegrin (DC-SIGN) for immature DCs and ICAM-1, CD18, DC-SIGN, and PECAM-1 for mature DCs. These results suggest that DC adhesion to cerebral ECs depends on the maturation state of DCs and the activation state of the endothelium, and that it is regulated by specific receptor-ligand interactions. This study thus further highlights the active role of human brain microvascular ECs in neuroinflammation.

Dendritic Cell Preparation for Immunotherapeutic Interventions

Much effort has been made over the last decade to use dendritic cells (DCs) in vaccines to induce specific antitumor immune responses. However, the great hope provided by in vitro and in vivo preclinical investigations was not translated to the clinic in terms of clinical efficacy. Thus, one of the challenges resides in optimizing DC-based therapy to give maximum clinical efficacy while using manufacturing processes that enable quality control and scale-up of consistent products. In this article, we review DC biology and the DC-based clinical trials performed to date and focus on the DC maturation status compatible with the goals of cancer immunotherapy. We also highlight the different approaches used in these clinical studies, such as the DC types or subtypes used and their preparation. Finally, we discuss the immunological and clinical outcomes in treated patients, with emphasis on the strategies that could be used to improve DC-based vaccination.

Post-apoptotic tumors are more palatable to dendritic cells and enhance their antigen cross-presentation activity

Critical issues for cytotoxic lymphocyte (CTL) cross-priming are (a) the maturation state of dendritic cells (DC), (b) the source of the tumor-associated antigens (TAA) and (c) the context in which they are delivered to DCs. Drug-induced apoptosis has recently been implicated in CTL cross-priming. However, since drug-treatment produces in vivo more tumor cells than the DC default apoptotic clearance program can cope with, they are expected to proceed to secondary necrosis and change their molecular pattern. Here we have addressed this issue on renal carcinoma cells (RCC) by using different apoptotic stimuli. UVC, but not γ-irradiation or anthracyclins, induced after 4 h treatment of the RCC cell line K1 a combination of apoptotic (phosphatydilserine and calreticulin plasma membrane mobilization) and necrotic (membrane incompetence) features. Heat shock protein (Hsp)-70 and chromatin-bound high mobility box 1 HMGB1 protein, typical of necrosis, were released during the further 20 h and thus made accessible to co-cultured monocyte-derived immature ( i) DC. UVC-treated, secondary necrotic RCC cell lines were cross-presented with higher efficiency by cytokine-matured ( m) DC than their early apoptotic (i.e. γ-irradiated) counterpart. Upstream events such as increased tumor uptake, activation of genes involved in the antigen-processing machinery, and increased expression of costimulatory and maturation molecules were also observed after loading iDC with secondary necrotic, but not apoptotic, tumor cells. These data offer a description of the molecular and immunogenic characteristics of post-apoptotic tumors which can be exploited to increase the efficiency of in vivo and ex vivo TAA delivery to the DC cross-presentation pathway.

CD40L+ CD4+ memory T cells migrate in a CD62P-dependent fashion into reactive lymph nodes and license dendritic cells for T cell priming

There is growing evidence that the maturation state of dendritic cells (DCs) is a critical parameter determining the balance between tolerance and immunity. We report that mouse CD4+ effector memory T (TEM) cells, but not naive or central memory T cells, constitutively expressed CD40L at levels sufficient to induce DC maturation in vitro and in vivo in the absence of antigenic stimulation. CD4+ TEM cells were excluded from resting lymph nodes but migrated in a CD62P-dependent fashion into reactive lymph nodes that were induced to express CD62P, in a transient or sustained fashion, on high endothelial venules. Trafficking of CD4+ TEM cells into chronic reactive lymph nodes maintained resident DCs in a mature state and promoted naive T cell responses and experimental autoimmune encephalomyelitis (EAE) to antigens administered in the absence of adjuvants. Antibodies to CD62P, which blocked CD4+ TEM cell migration into reactive lymph nodes, inhibited DC maturation, T cell priming, and induction of EAE. These results show that TEM cells can behave as endogenous adjuvants and suggest a mechanistic link between lymphocyte traffic in lymph nodes and induction of autoimmunity.

Generation of functionally mature dendritic cells from elutriated monocytes using polyinosinic : polycytidylic acid and soluble CD40 ligand for clinical application

Despite the increasing use of dendritic cell (DC) vaccination in clinical trials, optimal conditions for the generation of functionally mature DCs remain to be established. The current standard DC maturation protocol for clinical trials has been used as an inflammatory cytokine cocktail [tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-6 and prostaglandin E(2)], but this cocktail induced insufficient maturation of DCs derived from elutriated monocytes when cultured in X-VIVO 15. The aim of this study was to define effective combinations of stimulators for generating functionally mature DCs from elutriated monocytes under current good manufacturing practice conditions. We compared the functional capacity of DCs in response to all possible pairwise combinations of four different classes of stimuli: TNF-alpha, peptidoglycan, polyinosinic : polycytidylic acid [poly(I:C)] and soluble CD40 ligand (CD40L). Maturation status of DCs stimulated with combination of four stimuli was similar to that of the cytokine cocktail as assessed by the cell surface phenotype. However, only the combination of poly(I:C) + CD40L induced complete functional activation of the whole DC population, assessing IL-12p70 production, allostimulatory activity, migratory response to CCL19 and T helper 1-polarizing capacity. Thus, the protocol based on the combination of poly(I:C) and CD40L is more effective for the induction of clinical-grade DCs from elutriated monocytes than the standard cytokine cocktail.

Review of clinical studies on dendritic cell-based vaccination of patients with malignant melanoma: assessment of correlation between clinical response and vaccine parameters.

During the past years numerous clinical trials have been carried out to assess the ability of dendritic cell (DC) based immunotherapy to induce clinically relevant immune responses in patients with malignant diseases. A broad range of cancer types have been targeted including malignant melanoma which in the disseminated stage have a very poor prognosis and only limited treatment options with moderate effectiveness. Herein we describe the results of a focused search of recently published clinical studies on dendritic cell vaccination in melanoma and review different vaccine parameters which are frequently claimed to have a possible influence on clinical response. These parameters include performance status, type of antigen, DC maturation status, route of vaccine administration, use of adjuvant, and vaccine induced immune response. In total, 38 articles found through Medline search, have been included for analysis covering a total of 626 patients with malignant melanoma treated with DC based therapy. Clinical response (CR, PR and SD) were found to be significantly correlated with the use of peptide antigens (p = 0.03), the use of any helper antigen/adjuvant (p = 0.002), and induction of antigen specific T cells (p = 0.0004). No significant correlations between objective response (CR and PR) and the tested parameters were found. However, a few non-significant trends were demonstrated; these included an association between objective response and use of immature DCs (p = 0.08), use of adjuvant (p = 0.09), and use of autologous antigen preparation (p = 0.12). The categorisation of SD in the response group is debatable. Nevertheless, when the SD group were analysed separately we found that SD was significantly associated with use of peptide antigens (p = 0.0004), use of adjuvant (p = 0.01), and induction of antigen specific T cells (p = 0.0003). No specific route of vaccine administration showed superiority. Important lessons can be learned from previous studies, interpretation of these findings should, however, be done with reservation for the many minor deviations in the different treatment schedules among the published studies, which were not considered in order to be able to process and group the data.

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.

Toll‐like receptor‐4 and mitogen‐activated protein kinase signal system are involved in activation of dendritic cells in patients with acute coronary syndrome

Atherosclerosis is an inflammatory disease in which dendritic cells have been suggested to play an essential role. The underlying signalling mechanisms are unknown thus far. The family of Toll-like receptors (TLRs) initiates innate immune responses, and Toll-like receptor-4 (TLR4) has been considered to be an important player in the initiation and progression of atherosclerotic disease. The highly conserved mitogen-activated protein kinase (MAPK) family is one of the major kinase families that regulate cells by transducing extracellular into cellular responses. Three important members of this family are the extracellular signal-regulated kinase (ERK), p38, and c-Jun N-terminal kinase (JNK). The aim of the study was to investigate the expression of TLR4 and MAPK families on dendritic cells (DC) in patients with coronary arteriosclerosis disease. We have examined the expression of TLR4 protein and mRNA by flow cytometry and real-time quantitative reverse transcription polymerase chain reaction (RT-PCR). In addition, the expression of MAPK family proteins have been determined by Western blot analysis. We examined the expression level of CD80 to value the maturation state of DC. We compared the levels of cytokines in DC in response to lipopolysaccharide (LPS). The results showed that the expression of TLR4 and MAPK families are increased in the patients with acute coronary syndrome (ACS), compared with it in the patients with stable angina and controls. DC in ACS are activated evaluated by its mature marker and cytokine secreting responding to LPS. We suggest that TLR4 and MAPK families maybe involved in activation of circulating DC of ACS patients.

Human myoblasts modulate the function of antigen-presenting cells

Muscle biopsy specimens of myositis patients were analyzed for the presence of dendritic cells (DC) and macrophages (MPh) by immunohistochemistry. The interaction of DC and myoblasts (MB) was studied by coculture and effects on DC phenotype and function were assessed by flow cytometry and T-cell proliferation assays. Effects of MB-lysates on the phagocytic capacity of MPh were analyzed in bead-incorporation assays. Myositis specimens revealed a tendency towards more immature DC. MB modulated the maturation state of DC and DC recovered from MB-coculture had an inhibitory effect on T-cell proliferation. MB-lysates strongly stimulated MPh phagocytosis. Hypothetically, MB might modulate APC, counterbalancing immune-mediated damage.

Programmed death 1 ligand signaling regulates the generation of adaptive Foxp3 + CD4 + regulatory T cells

Although mature dendritic cells (DCs) are potent initiators of adaptive immune response, immature steady-state DCs contribute to immune tolerance. In this study, we show that ex vivo splenic DCs are capable of inducing conversion of naïve CD4(+) T cells to adaptive Foxp3(+)CD4(+) regulatory T cells (aTreg) in the presence of TGF-beta. In particular, when compared with splenic CD8alpha(-) DCs, the CD8alpha(+) DC subset were superior in inducing higher frequencies of conversion. This was not attributable to the difference in basal level of costimulation, because deficiency of CD40 or CD80/86 signaling did not diminish the differential induction of Foxp3. Conversion was regulated by DC maturation status. Further insights into the molecular mechanisms of conversion were gained by analyzing the contribution of several costimulatory and coinhibitory receptors. Costimulatory signals through GITR suppressed conversion, whereas coinhibitory signaling via programmed death 1 ligand (PD-L1) but not PD-L2 was required for conversion. Ex vivo PD-L1(-/-) DCs failed to support Foxp3 induction in the presence of TGF-beta. In vivo blocking PD-L1 signaling abolished conversion in a tumor-induced aTreg conversion model. Collectively, this study highlights the cellular and molecular parameters that might be exploited to control the de novo generation of aTregs and peripheral tolerance.

Melanoma and Lymphoma Rejection Associated With Eosinophil Infiltration Upon Intratumoral Injection of Dendritic and NK/LAK Cells

Dendritic cells (DCs) are promising tools for tumor immunotherapy. Their efficacy in the tumor environment increases when tumor cells die as a consequence of chemo/radiotherapy or when local stimuli promoting DC maturation and function are available. Dying tumor cells could represent a source of tumor antigens, which DCs cross-present to tumor-specific T cells. The outcome of cross presentation is in turn determined by the maturation state of DCs. Natural killer (NK)/lymphokine-activated killer (LAK) cells injected into growing tumors could both provide a source of dying cells for cross-presentation and deliver stimuli for DC maturation. Here, we report that NK/LAK cells recognized and killed in vivo major histocompatibility complex class I(low) highly tumorigenic, nonimmunogenic B16F1 melanoma cells when injected into exponentially growing neoplastic lesions. The simultaneous injection of immature DCs was required to heal animals. Similar results were obtained injecting NK/LAK cells and DC into growing Raucher leukaemia virus induced cell line lymphomas. Cured mice failed to reject other implantable tumors, and developed a specific cytotoxic response against the original neoplasm; moreover, they developed a long-lasting memory, and were protected against further challenges with living tumor cells only when both cell populations were introduced. The response associated to the preferential recruitment within tumors of eosinophils. The simultaneous injection in solid tumors of DCs and NK/LAK cells represents an attractive approach for antineoplastic immunotherapeutic strategies.

Differential Control of T Regulatory Cell Proliferation and Suppressive Activity by Mature Plasmacytoid versus Conventional Spleen Dendritic Cells

Anergy and suppression are cardinal features of CD4(+)CD25(+)Foxp3(+) T cells (T regulatory cells (Treg)) which have been shown to be tightly controlled by the maturation state of dendritic cells (DC). However, whether lymphoid organ DC subsets exhibit different capacities to control Treg is unclear. In this study, we have analyzed, in the rat, the role of splenic CD4(+) and CD4(-) conventional DC and plasmacytoid DC (pDC) in allogeneic Treg proliferation and suppression in vitro. As expected, in the absence of exogenous IL-2, Treg did not expand in response to immature DC. Upon TLR-induced maturation, all DC became potent stimulators of CD4(+)CD25(-) T cells, whereas only TLR7- or TLR9-matured pDC induced strong proliferation of CD4(+)CD25(+)Foxp3(+) T cells in the absence of exogenous IL-2. This capacity of pDC to reverse Treg anergy required cell contact and was partially CD86 dependent and IL-2 independent. In suppression assays, Treg strongly suppressed proliferation and IL-2 and IFN-gamma production by CD4(+)CD25(-) T cells induced by mature CD4(+) and CD4(-) DC. In contrast, upon stimulation by mature pDC, proliferating Treg suppressed IL-2 production by CD25(-) cells but not their proliferation or IFN-gamma production. Taken together, these results suggest that anergy and the suppressive function of Treg are differentially controlled by DC subsets.

MyD88 and IFN-αβ differentially control maturation of bystander but not Salmonella-associated dendritic cells or CD11cintCD11b+cells during infection

The interface between dendritic cells (DCs) and T cells is critical to elicit effective immunity against pathogens. The maturation state of DCs determines the quality of the interaction and governs the type of response. DCs can be matured directly through activating Toll-like receptors (TLRs) or indirectly by cytokines. We explore the role of the TLR adaptor MyD88 on DC maturation during Salmonella infection. Using Salmonella expressing GFP, we also examine the phenotype and function of bacteria-associated DCs matured in the absence of bacteria-mediated TLR signalling. MyD88 was required for upregulation of CD80 on DCs during infection, whereas CD86 and CD40 were upregulated independently of MyD88, although requiring a higher bacterial burden in the MLN. MyD88-independent upregulation was mediated by IFN-alphabeta produced during infection. In infected MyD88(-/-)IFN-alphabetaR(-/-) mice, which lack most bacteria-driven TLR signalling, indirect DC maturation was abolished. In contrast, DCs containing Salmonella upregulated co-stimulatory molecules independently of MyD88 and IFN-alphabeta, revealing a pathway of phenotypic maturation active in infected DCs. However, despite high co-stimulatory molecule expression, Salmonella-containing DCs from MyD88(-/-) or MyD88(-/-)IFN-alphabetaR(-/-) mice had a compromised capacity to activate T cells. Thus, bacterial stimulation of TLRs influences DC function at multiple levels that modulates their capacity to direct antibacterial immunity.

Thrombotic Occlusion of an Aortic Pseudoaneurysm

The mechanisms leading to drug allergy in predisposed patients, especially those related to T-cell-mediated drug hypersensitivity, are not well understood. A key event in allergic reactions to drugs is the maturation process undergone by dendritic cells (DCs). Although amoxicillin (AX) has been reported to interact and maturate DCs from patients with AX-induced delayed-type hypersensitivity, the cell signaling pathways related to AX-mediated DC maturation have not been elucidated. We sought to determine the role of the MAPK and NF-κΒ pathways on AX-induced DC maturation and functional status. For that purpose, in monocyte-derived-DCs from AX-delayed allergic patients and tolerant subjects, we analyzed the activation pattern of p38MAPK, JNK, and ERK signaling and the NF-κB, maturation markers as well as endocytosis and allostimulatory capacities driven by AX-stimulated-DCs. Our data reveal that AX induces an increase in the phosphorylation levels of the three MAPKsand activated NF-κB in DCs from allergic patients. Moreover, the inhibition of these pathways prevents the up-regulation of surface molecules induced by AX. Additionally, we observed that the allostimulatory capacity and the endocytosis down-regulation in AX-stimulated-DCs from allergic patients depend on JNK and NF-κB activities. Taken together, our data shed light for the first time on the main signaling pathways involved in DC maturation from AX-delayed allergic patient.

Distribution and clinical significance of blood dendritic cells in children with juvenile idiopathic arthritis

Background:A role for dendritic cells (DC) in the development of adult rheumatoid arthritis has been suggested. To date, this problem has been poorly explored in juvenile idiopathic arthritis (JIA).Objective:To analyse...

The cross-talk between dendritic and regulatory T cells: good or evil?

Immune responses against pathogens require fine regulation to avoid excessive inflammation, which could be harmful to the host. Moreover, the immune system must be tolerant to nonpathogenic antigens to prevent allergy, autoimmunity, and transplant rejection. There is accumulating evidence that interactions between dendritic cells (DC) and regulatory T (Treg) cells play a crucial role in the balance between immune response and tolerance. Communications between these cells are complex, bidirectional, and mediated by soluble or cell surface molecules. The maturation status of DC, which may be influenced by different microenvironmental factors, is considered as an important checkpoint for the induction of peripheral tolerance through modifications of the activation status of T cells. Moreover, several lines of experimental evidence suggest that different subsets or the functional status of DC are also involved in the promotion of Treg cell differentiation. A better knowledge of the regulatory mechanisms of the immune response induced or inhibited by DC via their interactions with Treg cells could be relevant for the development of new, immunotherapeutic approaches.

Combined treatment with intratumoral injection of dendritic cells and topical application of imiquimod for murine melanoma

The maturation state of dendritic cells is one of the factors that affect their capacity to induce antigen-specific cytotoxic T lymphocytes. Topical cutaneous application of imiquimod can induce the maturation and migration of cutaneous dendritic cells. To evaluate the synergistic effect of topical application of imiquimod plus intratumoral injection of syngeneic bone marrow-derived dendritic cells in the treatment of melanoma. For the B16F10 melanoma model, naive C57BL/6 mice were inoculated intradermally with 2x10(3) B16F10 melanoma cells in the right upper flank. Four groups (untreated control, dendritic cells alone, imiquimod alone and imiquimod plus dendritic cells) were included in the animal study, with five mice in each group. Tumour size was measured every 2 weeks, and histochemical and immunohistochemical staining carried out. ELISpot and PKH assays were performed to assess immune activity. Combined treatment of topical application of imiquimod and intratumoral injection of dendritic cells led to significant tumour regression, in contrast to partial eradication of the tumours with imiquimod or dendritic cells alone. These findings suggest that combination therapy with topical application of imiquimod and intratumoral administration of dendritic cells is a potent strategy for the treatment of melanoma.

Density of DC-LAMP+ mature dendritic cells in combination with activated T lymphocytes infiltrating primary cutaneous melanoma is a strong independent prognostic factor

As the most potent antigen presenting cells, dendritic cells (DCs) play key roles in the immune response against tumors. Their density in the tumor tissue has been associated with prognosis in patients with various cancers. However, few studies have been aimed at the presence and maturation state of DCs in cutaneous melanoma, with regard to their potential clinical correlates. In this study, the density of DCs expressing CD1a and the maturation marker DC-LAMP was determined by immunohistochemistry in primary tumor samples from 82 patients with cutaneous malignant melanoma. Intratumoral and peritumoral cell densities were analyzed in relation to tumor thickness and the subsequent development of metastases, as well as to patients' survival. CD1a(+) DCs were found both infiltrating melanoma cell nests and in the surrounding stroma, while DC-LAMP(+) mature DCs were generally confined to the peritumoral areas, associated with lymphocytic infiltrates. DC density values significantly correlated with the number of activated (CD25(+) or OX40(+)) T lymphocytes (p < 0.001). The degree of infiltration by CD1a(+) and DC-LAMP(+) DCs showed strong inverse correlation with the thickness of melanomas (p < 0.001). High peritumoral density of mature DCs was associated with significantly longer survival (p = 0.0195), while density of CD1a(+) cells had a prognostic impact of borderline significance (p = 0.0610). Moreover, combination of high peritumoral CD1a(+) or DC-LAMP(+) cell density with high number of CD25(+) or OX40(+) lymphocytes identified patient subgroups with more favorable survival compared to other subgroups. A multivariate survival analysis involving DC and activated T-cell densities alone and in combinations, as well as traditional prognostic factors, identified high DC-LAMP(+) cell/high OX40(+) cell density and Breslow index as independent predictors of good prognosis. These results suggest that the presence of CD1a(+) DCs primarily depends on the thickness of melanomas, without direct relationship with the patients' survival. On the other hand, the density of mature DCs, especially in association with that of activated T cells, proved of prognostic importance, suggesting that these parameters could be considered as signs of a functional immune response associated with better outcome of the disease.

Differential susceptibility of bone marrow-derived dendritic cells and macrophages to productive infection with Listeria monocytogenes

Dendritic cells (DC) are required for the immune response against Listeria monocytogenes and are permissive for infection in vivo and in vitro. However, it is unclear if DC provide a desirable intracellular niche for bacterial growth. To address this issue, we have compared the behaviour of L. monocytogenes in murine bone marrow-derived DC and macrophages (BMM). Similar to BMM, bacteria escaped to the cytosol in DC, replicated, and spread to adjacent cells. However, DC infection was less robust in terms of intracellular doubling time and total increase in bacterial numbers. Immunofluorescence analysis using a strain of L. monocytogenes that expresses green fluorescent protein upon bacterial entry into the cytosol suggested that a subpopulation of DC restricted bacteria to vacuoles, a finding that was confirmed by electron microscopy. In unstimulated DC cultures, L. monocytogenes replicated preferentially in phenotypically immature cells. Furthermore, DC that were induced to mature prior to infection were poor hosts for bacterial growth. We conclude that DC provide a suboptimal niche for L. monocytogenes growth, and this is at least in part a function of the DC maturation state. Therefore, the generation of an effective T cell response may be a net effect of both productive and non-productive infection of DC.