Nomenclature of monocytes and dendritic cells in blood

The hallmark of immunity is antigen (Ag) recognition. From early life until death we face countless Ags; many are deleterious and come from the outside world. However, lymphocytes, the mediators of specific acquired immunity, are usually not exposed where foreign antigens are encountered. Thus a system must ferry these Ags from the periphery into inner tissues where lymphocytes comfortably reside or traffic. Clearly, translocating Ags from the periphery to the lymphoid niches is a pivotal function whereby the dendritic cell (DC) system initiates immune responses. DCs, however, are not passive Ag couriers; a dynamic process not fully understood is triggered by Ag deposition. Strategically poised at the boundaries between the inner and the outside world, in a way bridging innate and acquired immunity, DCs sample, trap, engulf, and digest Ags of very diverse origin, in what is called Ag processing. Antigenic fragments are then regurgitated, exposed at the cell surface through one of three molecular pathways [Major histocompatibility complex (MHC) CI and CII, and CD1)] responsible for an efficacious Ag presentation. Concomitantly, DC bear an arsenal of powerful costimulatory molecules [CD40; CD80; CD86; DC-specific/intercellular adhesion molecule type 3-grabbing, non-integrin (DC-SIGN)] and the potential to produce critical cytokines [chemokines, interleukin-12 (IL-12), etc.], thus ensuring the initiation and the fate of acquired immunity. The DC system is particularly efficient, not just translocating, but also transforming antigens sampled at the earlier local milieu into an immunologically accesible language, becoming an excellent reporter of the previous antigenic past, translating the conundrum posed by viruses, bacteria, proteins, etc., into T-cell receptor (TCR) ligands; a short-peptide vocabulary that T cells especially now understand. A highly dynamic process is thus triggered by encountering Ags; while these are carried and processed to be appropriately presented, DCs in turn experience a variety of changes: migratory, phenotypical, functional; all encompassed in the term maturation. Essentially, DC maturation means to change from an antigen-capturing to an antigen-presenting, T-cell-priming mode, a process whereby DCs convert antigens into efficacious immunogens, express the necessary cytokines and costimulatory molecules, thus appropriately initiating the specific, acquired clonal immunity.

BackgroundGiant Cell Arteritis (GCA) and Polymyalgia Rheumatica (PMR) are overlapping diseases occurring exclusively in people older than 50 years. Antigen-presenting cells (APCs), including monocytes and dendritic cells (DCs), are main...

Expansion of CD141int Dendritic Cells in Myelodysplastic Syndrome (MDS)

Evolution has provided two distinct and highly sophisticated defense mechanisms to human beings for survival in a hostile environment. The innate immune system is aimed to react rapidly (from within minutes to a few hours) and in a rather simple way with little variation to attacks of pathogens. In contrast, the acquired immune system provides a more adaptive and highly specific defense response to foreign structures. In addition, it has the unique ability to induce tolerance of self-structures. The mechanisms of acquired immunity involve several steps of recognition and reactions in which various different cell types are engaged. Among antigen-presenting cells (APC), dendritic cells (DC) fulfill a pivotal function by providing information about invading pathogens under optimal conditions to other partners (e.g., effector cells) of the immune system. Thus, after having been neglected for years, DC research is experiencing a revival due to the central role of these cells in the complex machinery of the adaptive immune response. Moreover, understanding the role DC play in pathophysiologic conditions may be a key step in developing treatment strategies for several disease entities. Since many different DC types have been identified during the last years, including follicular DC and thymic DC, the present review will focus on the "classical" DC as they have been described initially by Steinman and Cohn. The first member of the DC system was described more than 100 years ago by Paul Langerhans (1868) and was originally thought to be a type of cutaneous nerve cell. After it had then been considered for a time to be an immature melanocyte, Birbeck (1) described the unique ultrastructural feature of the Langerhans cell (LC), which was named after him. Birbeck granules (BG) are rod-shaped structures with a central, periodically striated lamella and, depending on the section viewed, are tennis-racket shaped. BG are found exclusively in LC from man and other mammals, but not in other DC. They are considered the primary marker of epidermal LC. Nowadays, LC are best recognized in the skin by their CD1a expression. In the 1970s, Steinman & Cohn first described the structure and function of DC from mouse spleen suspensions (2). Morphologically, DC are characterized by their numerous thin, elongate cytoplasmic processes, which give them a veil-like appearance. They exhibit features of metabolically active cells with scattered mitochondria; a recognizable Golgi apparatus; some lysosomes, phagolysosomes, and lipid droplets; and a well-developed endoplasmic reticulum. They have large and often indented nuclei with heterochromatin preferentially deposited at the nuclear membrane (3). DC have been found in virtually all types of epithelia (skin, mucous membranes, lung) and as interstitial DC in the heart and kidney as well as in other organs. In addition, various subtypes of DC were also discovered in blood and in the lymphatic system (4). These represent different stages of maturation and are connected by circulatory pathways. Beside their typical dendritic structure in tissue and in suspension, DC were initially characterized mainly by their high expression of major histocompatibility complex (MHC) class II HLA-DR and their high stimulatory activity toward allogeneic T cells. Although they ultimately act as highly specialized APC, DC have to undergo four main stages of differentiation and maturation before they fulfill their main function in the lymphoid organs. Since the first demonstration that epidermal LC are derived from bone-marrow cells by Katz et al. (5), many efforts have been made to characterize the precursor cells of DC and LC in bone marrow and blood (Fig. 1). Thus, the ontogenesis and the development of techniques for in vitro generation of DC have been the focus in this field of research, especially considering possible therapeutic implications (see below). Ontogenesis of dendritic cells (DC). Before being able to activate naive T cells (Tn) (primary immune response), DC must undergo profound maturation step which occurs during their migration to regional lymph nodes. In peripheral tissue, DC may also trigger secondary immune response when encountering memory T cells (Tm) in transit through tissue. Although it is well established that DC derive from bone-marrow CD34+ stem cells, two main strategies have been followed over the past years. First, in 1992, Caux et al. described a system that generates CD1a+ LC-like DC from CD34+ stem cells by supplementing granulocyte/macrophage colony stimulating factor (GM-CSF) and tumor-necrosis factor alpha (TNF-α) (6). The generation of LC/DC was optimized later by adding stem cell factor (SCF) and/or FLT-3 ligand, resulting in a higher yield of CD1a+ cells, with a typical dendritic structure, strong expression of MHC class II antigens, CD4, CD40, CD54, CD58, CD80, CD83, and CD86, and the presence of BG in 10–20% of the cells. Most importantly, these cells exhibit a potent capacity to stimulate the proliferation of naive T cells and to present soluble antigens to clones of CD4+ T cells. On the other hand, in 1994, Sallusto & Lanzavecchia (7) were able to generate CD1a+ DC corresponding to interstitial DC in their phenotype by culturing monocytes with GM-CSF and interleukin (IL)-4. CD14+ monocytes undergo maturation into CD1a+ DC, which, however, lack BG and are therefore not considered LC but are more similar to dermal DC since they express CD11b, CD68, and the coagulation factor XIIIa. Typically, after 7 days of culture with GM-CSF and IL-4, monocytes give rise to immature DC which need further stimulation with CD40 ligand, endotoxin, or TNF-α to reach the full maturation stage of highly stimulatory DC. However, if monocytes are cultured with macrophage colony stimulating factor (M-CSF) alone, they differentiate into macrophage-like cells (CD14+, CD1a−, CD83−) and synthesize IL-10 (8). While these DC are now classified as myeloid DC because they are known to derive from myeloid precursors (see below), more recently, a novel type of so-called lymphoid DC has been described. These lymphoid DC derive from CD4+/CD3−/CD11c− plasmocytoid cells from the blood and the tonsils (9, 10). These precursors do not differentiate into macrophages with GM-CSF or M-CSF. Lymphoid DC are dependent on IL-3, but not on GM-CSF, and are less active in phagocytosis. When localized in peripheral blood or in nonlymphoid tissue, DC are considered to be functionally immature. This refers to the fact that DC in tissues are highly specialized for capturing and processing foreign or autologous protein antigens or haptens. Uptake of high-molecular-weight antigens by DC may occur through macropinocytosis or more specifically through a number of membrane receptors such as FcγRII and FcεRI loaded with the adequate antibodies. DC also express membrane receptors bearing multiple lectin domains such as the mannose receptor and the DEC-205 molecule(11). These structures enable DC to internalize antigens by receptor-mediated endocytosis, a pathway which leads to antigen uptake into specialized compartments inside DC and allows efficient processing and subsequent loading of these antigens on MHC class II molecules. In contrast, uptake of low-molecular-weight haptens, e.g., DNCB or oxazolone (12, 13), mostly occurs via binding to surface glycoproteins and subsequent internalization. Experiments with MHC knockout mice suggest that presentation of such haptens is achieved through MHC class I molecules to CD8+ T cells rather than via MHC class II. A further characteristic of DC is the high stability of MHC class I or class II molecules on their cell surface, allowing them to be loaded for a long time with defined antigens. At this stage of maturation, DC are able to stimulate memory T cells trafficking through the tissue, initiating a secondary immune response at the site of contact with the captured antigen. However, since macrophages and other cells are as efficient as DC in this type of stimulatory activity, it is assumed that triggering a secondary immune response is not the primary task of DC under normal conditions. In recent years, it has become clear that the migration of many cell types including DC is tightly regulated by chemokines. The expression of chemokines at different anatomic sites and in different pathologic states in combination with the differential expression of chemokine receptors on cells during different maturation stages is the basis of a complex signaling network that orchestrates cell migration and cell interaction in the immune response (14). Specifically for DC, it has been shown that the chemokine receptor profile expressed on immature DC (CCR1, CCR2, CCR5, and CCR6) mainly recognizes chemokines that are released during inflammatory processes. This allows the accumulation of DC that are geared toward antigen uptake at sites of inflammation. Release of cytokines such as IL-1 and TNF-α further perpetuates this process by inducing immature DC to release even more inflammatory chemokines. Conversely, mature DC downregulate their receptors for inflammatory chemokines and express different chemokine receptors (CCR4, CCR7, CXCR4, SLC, and ELC). These allow them to receive signals which will attract them to the regional lymphatics and eventually to the T-cell-rich areas of the lymph node. Thus, after antigen uptake, tissue DC migrate to the regional lymph nodes. For example, LC seem to be able to migrate quite fast; i.e., several millimeters within 30 min (15). On their way to the lymph node, DC begin a profound metamorphosis, leading to significant changes in their structure and phenotype. In the afferent lymphatic vessels, DC have been described as so-called veiled cells and as interdigitating cells in the T-cell-rich paracortical zones of secondary lymphoid tissues. As DC mature, they lose their antigen uptake capacity and their function shifts toward antigen presentation. One of the hallmarks of this development is the upregulation of peptide-loaded MHC class II and costimulatory molecules (CD80, CD86) on the surface of these cells. In the meantime, DC rapidly downregulate and sometimes completely abolish the expression of Fc receptors. Migration and maturation of DC seem to be linked processes in vivo since factors such as lipopolysaccharides (LPS), TNF-α, and IL-1 induce both processes (16). In vitro, TNF-α has been shown to induce maturation of monocyte-derived DC, also leading to upregulation of CD80, CD86, CD83, and MHC class II. All these molecules are crucial for efficient antigen presentation to resting naive T cells. Priming of naive T cells is one of the crucial tasks that DC have to fulfill. To do so, DC and naive T cells have to colocalize in the paracortical zone of the lymph nodes. An interesting finding was the fact that naive T cells express chemokine receptors (e.g., CCR7) that allow them to receive the signals sent by mature DC which release ELC and DC-specific chemokines (DC-CK1). After having reached the T-cell area, a single DC can prime hundreds of naive T cells. In this process, peptides bound to MHC class II or MHC class I on DC are presented to T cells via the T-cell receptor complex (TCR). Recently, it became clear that, in addition to the signals received via the TCR, costimulatory signals are of key importance in initiating and directing a T-cell response. Interaction of the costimulatory molecules CD80 and CD86 with their counterparts on T cells, i.e., CD28 or CTLA-4, determines whether this stimulation will result in an antigen-specific proliferation of T cells or tolerance. Indeed, additional factors present at the site of DC–T-cell interaction such as IL-10 may modify CD80/CD28 signaling by blocking downstream events in signal transduction, thereby leading to antigen-specific tolerance (17). An important observation was that DC can release IL-12. This cytokine is involved in the induction of a Th1 T-cell response. Likewise, other cytokines such as IFN-γ may induce a Th1 response, whereas IL-4 has been shown to direct the T-cell response toward Th2. This capacity to influence the type of T-cell response may explain why some antigens induce an allergic reaction and others do not. It is interesting that the cytokines and factors released during T-cell priming also induce a different chemokine receptor repertoire on stimulated T cells. Whereas Th1 cells express CCR1, CCR2, CCR5, CXCR3, and CXCR5, Th2 cells are characterized by the expression of CCR2, CCR3, CCR4, and CXCR5 (14). The differential expression pattern may recruit these cells to specific types of inflammation (allergic vs nonallergic) and determine which other cell types may be involved in a particular inflammatory response. As far as allergic reactions are concerned, it is noteworthy that Th2 cells, eosinophils, and basophils share the expression of the chemokine receptor CCR3, whereas Th1 cells and monocytes, which can differentiate into DC, share CCR1 and CCR5. Once antigen presentation has been achieved, DC are not supposed to recirculate in peripheral blood or lymphatic vessels. Indeed, it is assumed that DC will be killed by T cells or will die by apoptosis on site (18, 19). As mentioned above, the primary task of DC is to inform the immune system about the invasion of foreign and potentially harmful proteins. Much interest has been focused over the last 25 years on the possible pathophysiologic role of DC in a variety of conditions, especially in allergic inflammatory diseases. Allergic contact dermatitis (ACD) is the archetype of cell-mediated hypersensitivity reactions in which DC play a pivotal role in the sensitization process. While the contact of irritant compounds on the skin leads to the secretion of TNF-α and GM-CSF by keratinocytes, low-molecular-weight haptens (e.g., nickel, DNCB, or oxazolone) stimulate the additional release of IL-1α, IP-10, and MIP-2. These chemokines activate DC and endothelial cells, leading to an accumulation of even more DC at the site of antigen contact. Moreover, application of hapten induces the release of IL-1β by epidermal LC and thereby promotes their egress from the epithelium. After the uptake of the antigen, DC process it while migrating to the regional lymph nodes where it will be presented to antigen-specific naive T cells. Little is known about the mechanisms which enable DC to be highly efficient in priming naive T cells. Another remarkable property of DC is their ability to present exogenous antigens on MHC class I and II molecules. This leads to the activation of both CD4+ and CD8+ hapten-specific T cells (20, 21). Whereas classical delayed-type hypersensitivity reactions are mediated by CD4+ effector cells, contact dermatitis is mediated by CD8+ effector cells (22-24). Other cytokines released during the sensitization process have been implicated in directing the type of immune response mounted by T cells. It has been shown that IL-10 converts LC/DC from potent inducers of a primary immune response to hapten-specific tolerizing cells. A significant decrease in mRNA signals for IL-1α, IL-1β, and TNF-α confirms the immunomodulatory role of this cytokine in contact hypersensitivity reactions (25, 26). On the other hand, IL-12 which is released by keratinocytes and by DC themselves (25, 27), is known as a strong inducer of the Th1 response. After a second contact with a contact allergen, antigen-specific memory T cells can be stimulated either by DC or by APC less potent than DC (e.g., macrophages or monocytes) and, due to their specific homing molecules, elicit an immune response at the appropriate anatomic site. Atopic diathesis is characterized by three major diseases, i.e., allergic rhinoconjunctivitis, allergic asthma, and and is with Thus, it is assumed that mechanisms e.g., secretion of IL-4 and are of crucial importance in diseases. specific may play a role in the of these conditions. Since of the react do not have direct to cells in the blood or in lymphoid tissue, and presentation to T cells must be by APC localized in tissues at the with the i.e., in the the and other Thus, as they the first of defense in these peripheral DC are considered the best for priming naive T cells toward In the of the which has research during the last it was resting T cells are into Th1 or Th2 cells during antigen presentation. While it became clear that IL-12 by DC is mainly for the to Th1 it was a of which cells may be the of IL-4, which shifts T-cell response to the Th2 et al. some that may be the signal which cells to the Th2 type recently, et al. have shown that myeloid DC are for T cells into Th1 to as while lymphoid DC direct T cells into Th2 in an way to as (Fig. Moreover, mechanisms are these DC and T cells. of the mechanisms of Th2 stimulation by lymphoid or other and/or costimulatory will modify understanding of has the immune system to an appropriate of immune types of dendritic cells and DC types seem to derive from different and are to Th1 and Th2 It has been that monocytes, and myeloid express the receptor for lymphoid this structure has not been The FcεRI on LC and several important from this receptor on effector cells of i.e., cells and Indeed, it is not expressed on these cells but to be regulated by signals of the inflammatory the cells. Thus, the FcεRI expression is on LC and a described inflammatory dendritic epidermal cell from skin of dermatitis However, the lack or the surface expression of the receptor complex is due to the expression of the which is for the surface expression of the structure, while the is present in a inside the cells the FcεRI on LC and as well as on monocytes, the This has in to LC and from normal LC receptor are not receptor is of a role of FcεRI in antigen by monocytes, and blood DC that have been by FcεRI receptor-mediated are into MHC class II compartments such as in which processing and loading of MHC class II molecules occur This in leads to an optimal antigen presentation to CD4+ T cells, as a for antigen In this one may about the role of DC in the of It is well that molecules and effector cells such as cells, or are the result of an efficient defense system. It has been that this system has been toward because of the lack of have been to the role of DC in the network of immunity and allergic As mentioned above, antigen uptake, and presentation are the main of DC. Among the of antigen which and receptor endocytosis, the last provides the efficient and specific This to be the for Indeed, the expression of high FcεRI on DC of several important First, DC their ability to react to by binding large of molecules with various This the of FcεRI by a defined at the cell the allow the of rather large which, under normal are not via the i.e., by of FcεRI on DC is followed by via receptor-mediated via and However, in to the receptor where and different compartments this for antigen uptake by DC, i.e., specifically via and may whether the foreign structure will be and to MHC class ultimately leading to a higher of specific peptides in the of surface MHC class II molecules. DC high receptor will full cell activation FcεRI inducing the and release of defined may to the subsequent antigen presentation. One may that DC with specific can the secondary immune response and further trigger the by and more antigen-specific Th2 cells. DC are the potent of naive T i.e., they are to a primary immune response. At first antigen uptake and subsequent presentation seem rather in the primary reaction since specific be present at the However, it be that complex structures captured via FcεRI on DC are by these cells in a way leading the and presentation of the T cell This then a primary reaction these antigens, thereby to the variety of the It is a that, as above, antigen uptake and FcεRI on DC to the and release of of directing T cells toward a defined phenotype and/or i.e., Th1 or Th2 cells. This in the of DC to be especially considering recent an important role of IL-12 and in T cells toward either Th2 or The role and function of APC in allergic disease high of both and DC were found in the and the of the of from DC of the BG a feature which them as LC. the represent LC at a different maturation stage or DC of a different to be The number of DC is in the and rapidly further the that higher are in the to with the antigen Indeed, it has been in after that the number of DC after antigen At the of the CD1a+ DC were in the and vessels, to the epithelium. In the second of these cells were found the of the As is little that DC are able to within the these changes are to in their and/or The pivotal role of DC for antigen processing is further by their with a of This with the in epithelia such as the normal human where the corresponding DC e.g., are with a of days or The interaction of DC with other cell types such as cells that can be identified in the to be The of is Although are this is a complex that is characterized by and Whereas the is characterized by release from cells, the is by an inflammatory in the the inflammation leads to to the is a allergic disease with a response and it has been that the of and other be due to a higher of Thus, pathogens that prime for a Th1 immune response. In addition, stimulation of the immune system with to for Th2 memory and thereby the type of immune response with The maturation of DC function in the is an important factor in the of the memory cell in the of this maturation process may be a key of the of Recently, it has been by et al. that different of DC may a direct over Th1 vs Th2 differentiation of naive T cells The first for the induction of an immune response to is that these molecules to cells. Although the a highly regulated and is in the to after In addition, induces cells to express GM-CSF, which DC to the site of antigen contact As far as antigen uptake by DC is concerned, the response within the tissue is the of MHC class II DC The class cells within the a within after antigen the DC their and to a more of veiled cells. DC within the is and in an in the of these cells from the to the lymph nodes. Another that may to an response of to may be that in the inflammatory process DC are from It is known that monocyte-derived DC from allergic in the expression of and the receptor for and even an upregulation of and into more potent cells than from normal Whereas DC are in priming the immune system to other APC may play a crucial role in the secondary immune response to In this they may to the of The major APC in the of the macrophages the and DC, the specific cells, type II cells, and, to a cells. The interaction of DC with other APC as well as with other effector cells of the immune system an active field of on DC in the skin the and release of which may a inflammatory as has been for cells. Thus, from a pathophysiologic of DC, and LC and DC in the have been to play a crucial role in dermatitis since they may represent the the and antigen-specific cells the skin This is by the observation that the presence of LC bearing molecules is a to by application of on the skin of may represent the of an delayed-type hypersensitivity reaction in and The of may be by cytokines derived from cells. The expression of and on endothelial cells is leading to the and invasion of other cells, such as macrophages or and by cytokines as well as have been shown to leading to an activation of and Th1 and cells. Thus, IL-12 may for the of the cytokine pattern and the from a Th2 to a Th1 response with the subsequent release of This cytokine is for the and of and determines the of the disease Indeed, the observation that IFN-γ mRNA in such was by a of IL-12 expression the of the Th2 to Th1 in the of As a DC have a crucial role in the of various especially in direct contact with the environment. pathophysiologic role in allergic contact as well as in other allergic diseases, is now well Moreover, they seem to have a central role in the and presentation of antigens. strategies have now been to DC in the of hypersensitivity reactions and, on the other hand, to these cells as a to Recently, have that the unique function of DC to In of their at tissues such as the skin and or DC be for therapeutic In the is known to the of LC/DC well as that of cells) and is in the treatment of inflammatory skin diseases. the capacity of DC to induce an immune response, the mechanisms are far from Indeed, DC seem to their expression of several functionally molecules such as HLA-DR or CD86, but they their stimulatory activity recently, it has been shown that a generation of i.e., and which, in to can be interesting with to DC They the expression of costimulatory molecules, the of distinct DC in inflammatory tissue and decrease the stimulatory activity of DC in vitro, as well as in after application of molecules with the binding of to receptor or compounds defined activation mechanisms by DC in represent in the of conditions. made in understanding the ontogenesis of DC and the techniques for their generation in vitro have to an and therapeutic in vitro DC may be either to hypersensitivity reactions in contrast, to the immune response in a as for A number of pathologic conditions are known to be by distinct of hypersensitivity Among diseases, and allergic are the DC with appropriate and by cytokines such as IL-10 or may be to and as well as T cells. have been because with LC in a of hapten-specific tolerance Another interesting is the of the immunomodulatory of such as This to the phenotype and the function of DC. It the expression of the costimulatory molecules CD86 and CD40, and the and release of IL-1 and but the of IL-10 Thus, may represent a and able to DC and to them from potent to The first therapeutic for the treatment of by with DC have been established Thus, DC may as for as the and of an immune response is regulated at the of DC. are to antigens or peptides to particular presentation class II vs class within DC. are and DC can induce primary and secondary immune of all years after the of the DC in the skin by Paul of the of these cells and especially the made in the last may be considered in the understanding of crucial pathophysiologic in diseases. Most importantly, this is about to of therapeutic and the of in vitro DC in has a in This was by the of the and by the

There are minimal data on the frequencies of monocyte subsets and dendritic cells (DCs) in children with Gaucher disease (GD), as nearly all previous studies have involved adult patients. Consequently, we aimed to describe the changes in these cell subpopulations in children with GD type 1 who were on regular enzyme replacement therapy (ERT). This case-control study included 25 children with GD1 and 20 healthy controls. All participants underwent investigations such as complete blood count and flow cytometric assessment of DC and monocyte frequencies and phenotype. We found that GD1 children had significantly reduced percentages of both types of DCs, i.e., plasmacytoid DCs and myeloid DCs, compared to the control group. There was also a significant reduction in absolute monocyte numbers and percentage of classical monocyte. Moreover, the GD1 children had higher frequencies of non-classical and intermediate monocytes than the control group. Our results so far indicate that, when compared to the control group, the GD1 children had significantly reduced total and classical monocyte, with significantly decreased frequencies for both types of DCs. These changes can contribute to immunological abnormalities in pediatric patients with GD1. Children with Gaucher disease type 1 (GD1) have significantly reduced total and classical monocyte frequencies, with decreasing percentages for both types of dendritic cells. GD1 children had significantly reduced frequencies of myeloid and plasmacytoid dendritic cells as compared to the controls. The GD1 children also had significant changes in monocyte subsets when compared to the controls. Our results show that monocytes and dendritic cells' significant changes could contribute to immunological abnormalities in pediatric patients with GD1.

Objectives: Type 2 diabetes patients show an increased susceptibility to suffer from infections of the skin, urinary tract, pulmonary system or other organs. The underlying cause for a weakened immune defense in type diabetic patients is yet unclear. Dendritic cells as potent antigen presenting cells play a key role in initiating a specific cellular immune defense. Here we investigate the percentage of peripheral blood dendritic cells (BDC) and their subsets, myeloid dendritic cells type 1 (MDC1) and plasmacytoid dendritic cells (PDC), on peripheral blood mononuclear cells (PBMC).

Dendritic cells (DCs) elicit potent anti-tumoral T-cell responses in vitro and in vivo. However, different types of DC have yet to be compared for their capacity to induce anti-tumor responses in vivo at different developmental stages. Herein, we correlated the efficiencies of different types of monocyte-derived DC as vaccines on the resulting anti-tumor immune responses in vivo. Immature and mature DCs were separately pulsed with a peptide derived from tyrosinase, MelanA/MART-1 or MAGE-1 and a recall antigen. Both DC populations were injected every 2 weeks in different lymph nodes of the same patient. Immune responses were monitored before, during and after vaccination. Mature DCs induced increased recall antigen-specific CD4(+) T-cell responses in 7/8 patients, while immature DCs did so in only 3/8. Expansion of peptide-specific IFN-gamma-producing CD8(+) T cells was observed in 5/7 patients vaccinated with mature DCs but in only 1/7 using immature DCs. However, these functional data did not correlate with the tetramer staining. Herein, immature DCs also showed expansion of peptide-specific T cells. In 2/4 patients vaccinated with mature DCs, we observed induction of peptide-specific cytotoxic T cells, as monitored by chromium-release assays, whereas immature DCs failed to induce peptide-specific cytotoxic T cells in the same patients. Instead, FCS-cultured immature DCs induced FCS-specific IgE responses in 1 patient. Our data demonstrate that this novel vaccination protocol is an efficient approach to compare different immunization strategies within the same patient. Thus, our data define FCS-free cultured mature DCs as superior inducers of T-cell responses in melanoma patients.

Dendritic cells (DC) have a central role in the initiation of adequate immune responses. They recognize pathogens by means of Toll-like receptors (TLR) and link innate to adaptive immune responses by releasing proinflammatory cytokines and inducing T cell proliferation. We conducted this study to evaluate the expression and function of TLR on human lung DC subsets and to study their T cell stimulatory capacity. TLR gene expression by human pulmonary DC was evaluated by RT-PCR, while protein expression was analyzed by flow cytometry. We investigated cytokine release by DC in response to different TLR ligands. T cell stimulatory capacity was evaluated by mixed leukocyte reactions of purified lung DC with allogeneic T cells. Myeloid dendritic cells type 1 (mDC1) and myeloid dendritic cells type 2 (mDC2) express mRNA transcripts for TLR1, TLR2, TLR3, TLR4, TLR6, and TLR8. Flow cytometric analysis demonstrated high TLR2 protein expression for mDC1 and moderate TLR4 expression for mDC2. mDC1 and mDC2 release proinflammatory cytokines (TNF-alpha, IL-1beta, IL-6, and IL-8) in response to TLR2 and TLR4 ligands. TLR3 ligands induce cytokine release in mDC1, but not in mDC2. Plasmacytoid DC (pDC) express TLR7 and TLR9 and release proinflammatory cytokines in response to imiquimod and IFN-alpha in response to CpG oligonucleotides. mDC1 are strong inducers of T cell proliferation, while pDC hardly induce any T cell proliferation. mDC2 have an intermediate T cell-stimulatory capacity. Our results show divergent roles for the different human lung DC subsets, both in innate and adaptive immune responses.

Myeloid blood CD11c+ dendritic cells and monocyte-derived dendritic cells differ in their ability to stimulate T lymphocytes

Human dendritic cells (DC) are highly professional antigen presenting cells for the priming of naive cytotoxic T cells. Gene transfer in DC would be a useful strategy to load DC with relevant de novo synthesized antigens for immunotherapeutical purposes. As a first step towards a DC-based gene therapy, we examined the efficiency of nonviral transfection in different types of cultured human dendritic cells with a humanized red-shifted green fluorescent protein reporter gene. Plasmid DNA transfection by electroporation or lipofection was used to transfect CD34+ progenitor cell-derived DC (PC-DC) and Langerhans' cells (PC-LC), as well as monocyte-derived DC (Mo-DC). While lipofection was unsuccessful in all types of DC, we obtained high-efficiency gene transfer by electroporation in PC-LC (16%) and PC-DC (12%). In contrast, electroporation was strikingly less efficient in Mo-DC (< or = 2%). The potent allostimulatory capacity of DC was still retained in electroporated PC-DC and PC-LC. In conclusion, electroporation of antigen expressing plasmid DNA is an efficient tool for nonviral gene transfer in PC-DC and PC-LC, but not in Mo-DC and could be useful for the development of DC-based tumor immunotherapy.

The current study aims to evaluate the effect of non-surgical periodontal treatment on the modulation of monocyte phenotype, in the presence or absence of diabetes. The identification, quantification, and phenotypic characterization of monocyte subtypes (classical, intermediate, and non-classical) were performed by flow cytometry, at baseline and 1month after the end of non-surgical periodontal treatment, in patients with periodontitis, associated or not with diabetes. There was an increase in non-classical monocytes after treatment and a reduction in intermediate monocytes, without differences for the classical subtype, regardless of the diabetes status. Furthermore, there was a reduction in intermediate monocytes and an increase in non-classical and classical monocytes after treatment in the diabetes group, while no significant differences were observed for classical, intermediate, and non-classical monocytes in the group without diabetes. Comparisons between the two groups showed significant differences for classical, intermediate, and non-classical monocytes at baseline; these differences were not found one month after treatment. Non-surgical periodontal treatment leads to modulation of monocytes to a less inflammatory phenotype, especially in individuals with diabetes. A better understanding of the role of these biomarkers in the periodontitis contex may constitute a new strategic target for a better treatment of patiens with diabetes associated to periodontitis. Brazilian Registry of Clinical Trials-RBR-35szwc. Jhefferson Miranda Alves and Danielle Borges Germano contributed equality to this study and should be considered first authors.

Despite their distinct etiology, several lines of evidence suggest that innate immunity plays a pivotal role in both juvenile idiopathic arthritis (JIA) and septic arthritis (SA) pathophysiology. Indeed, monocytes and dendritic cells (DC) are involved in the first line of defense against pathogens and play a critical role in initiating and orchestrating the immune response. The aim of this study was to compare the number and phenotype of monocytes and DCs in peripheral blood (PB) and synovial fluid (SF) from patients with JIA and SA to identify specific cell subsets and activation markers associated with pathophysiological mechanisms and that could be used as biomarkers to discriminate both diseases. The proportion of intermediate and non-classical monocytes in the SF and PB, respectively, were significantly higher in JIA than in SA patients. In contrast the proportion of classical monocytes and their absolute numbers were higher in the SF from SA compared with JIA patients. Higher expression of CD64 on non-classical monocyte was observed in PB from SA compared with JIA patients. In SF, higher expression of CD64 on classical and intermediate monocyte as well as higher CD163 expression on intermediate monocytes was observed in SA compared with JIA patients. Moreover, whereas the number of conventional (cDC), plasmacytoid (pDC) and inflammatory (infDC) DCs was comparable between groups in PB, the number of CD141+ cDCs and CD123+ pDCs in the SF was significantly higher in JIA than in SA patients. CD14+ infDCs represented the major DC subset in the SF of both groups with potent activation assessed by high expression of HLA-DR and CD86 and significant up-regulation of HLA-DR expression in SA compared with JIA patients. Finally, higher activation of SF DC subsets was monitored in SA compared with JIA with significant up-regulation of CD86 and PDL2 expression on several DC subsets. Our results show the differential accumulation and activation of innate immune cells between septic and inflammatory arthritis. They strongly indicate that the relative high numbers of CD141+ cDC and CD123+ pDCs in SF are specific for JIA while the over-activation of DC and monocyte subsets is specific for SA.

The Impact of Graft-Versus-Host Disease on Dendritic Cell Homeostasis and Their Potential Use As Biomarker to Predict the Severity of Chronic Graft-Versus-Host Disease

Background Intestinal dendritic cells (DCs) and macrophages govern the mechanisms of immune homeostasis having a role in inflammatory bowel disease (IBD) onset. However, the profile of their circulating precursors (DC and monocytes) in IBD has not been previously described in depth. Our aim was to characterise blood DC and monocyte subsets in healthy controls (HCs) and IBD patients in order to understand their potential implication in IBD pathogenesis. Methods 18 HC and 64 IBD patients were recruited. IBD patients were categorised into Crohn’s disease (CD) and ulcerative colitis (UC), either endoscopically active (aCD and aUC) or quiescent (qCD and qUC), based on the SES-CD or the Mayo index endoscopic subscore. Blood circulating type 1 conventional DC (cDC1), type 2 conventional DC (cDC2), plasmacytoid DC (pDC), classical monocytes, non-classical monocytes and intermediate monocytes were identified by flow cytometry and characterised for the expression of 18 homing and activation markers (β7, CCR1, CCR2, CCR3, CCR5, CCR6, CCR7, CCR9, CCRL1, CD40, CD86, CD137L, CD274 (PD-L1), CLA, CXCR1, CXCR3, ICOSL and HLA-DR). Association between markers and the presence, type or activity of IBD was tested by logistic regression. Discriminant canonical analysis was also performed to classify the patients on their own endoscopy category. Results All groups (HC, aCD, qCD, aUC and qUC) were separated from the others based on the discriminant canonical analyses of the 18 markers applied over all DC and monocytes subsets (Figure 1). Specifically, CCRL1, CCR3 and CCR5 expression on cDC1, CCRL1 on non-classical monocytes and CCR9 and b7 on classical monocytes were highly associated to IBD. CCR3 displayed an odds ratio (OR) of 2.29 along with its 95% confidential interval (CI) between 1.11 and 4.75, showing a strong association with activity in CD; whereas the other markers displayed an inverse association with IBD. Hence, expression of CCRL1 on cDC1 and non-classical monocytes from aUC showed an OR (95% CI) of 0.23 (0.08–0.66) and 0.52 (0.28–0.95), respectively. In the case of qUC, CCR5 on cDC1 and β7 on classical monocytes displayed an OR (95% CI) of 0.10 (0.01–0.83) and 0.56 (0.34–0.90), respectively. CCR9 was inversely associated to qCD with an OR (95% CI) of 0.64 (0.46–0.89) in the classical monocytes subset. Indeed, the same markers (excluding β7) were also associated with IBD when all DC and monocyte subsets were considered at the same time. Conclusion Differences on the expression of migration markers CCR3, CCR5, CCR9, b7 and decoy receptor CCRL1 on circulating DC and monocyte subsets from IBD groups suggest the presence of constitutive migratory differences underlying IBD pathogenesis in CD or UC and its condition (inflamed or non-inflamed).

Dendritic cells (DCs) as antigen presenting cells play an important role in the initiation of an autoimmune disease like type 1 diabetes. Although there is evidence from the NOD mouse model that the function and frequency of DCs is altered in type 1 diabetes, there is little data on dendritic cells in human type 1 diabetes. We investigated peripheral blood myeloid (mDC1 and mDC2) and plasmacytoid dendritic cells (pDCs) in 15 type 1 diabetes patients with recent onset (within the last 3 months) of type 1 diabetes as well as in 15 patients with long standing (more than 5 years) type 1 diabetes by flow cytometry. Both groups were compared to age matched controls. We observed a significantly reduced percentage of pDCs of peripheral blood mononuclear cells (PBMCs) in both recent onset (0.13 vs. 0.25%, p=0.01) and long standing type 1 diabetes patients (0.13 vs. 0.24%, p=0.01). The absolute counts of pDCs per ml of blood were also significantly lower in both recent onset (mean 9560 vs. 13524, p=0.048) and long-standing diabetes (mean 7869 vs. 12202; p=0.05). The percentage of mDC1 was significantly diminished in recent onset (0.21% vs. 0.30%, p=0.034), but not in long standing type 1 diabetes. Our study demonstrates a persisting reduction of peripheral plasmacytoid DCs in type 1 diabetes patients. Since pDCs are involved in the control of immune responses and inducing regulatory cells, a reduced number of pDCs may predispose to an autoimmune reaction in the pancreatic islets.