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As dendritic cells (DCs) are among the first cells to encounter antigens, these cells trigger both innate and T cell responses, and are the most potent antigen-presenting cells. Brucella spp., which is an intracellular facultative and stealthy pathogen, is able to evade the bactericidal activities of professional phagocytes. Several studies have demonstrated that Brucella can survive and replicate intracellularly, thereby provoking impaired maturation of DCs. Therefore, the interaction between DCs and Brucella becomes an interesting model to study the immune response. In this review, we first will describe the most common techniques for DCs differentiation in vitro as well as general features of brucellosis. Then, the interaction of DCs and Brucella, including pathogen recognition, molecular mechanisms of bacterial pathogenesis, and intracellular trafficking of Brucella to subvert innate response, will be reviewed. Finally, we will debate diversity in immunological DC response and the controversial role of DC activation against Brucella infection.

Dengue is a worldwide expanding threat caused by dengue virus (DENV) infection. To date, no specific treatment or effective vaccine is available. Antibodies produced by plasma cells (PCs) might be involved concomitantly in protection and severe dengue immunopathology. Although a massive appearance of PCs has been reported during acute DENV infection in humans, this response has been poorly characterized. Here, we show the dynamic of PC generation in immune-competent mice cutaneously inoculated with DENV compared with two control experimental groups: mice inoculated with inactivated DENV or with PBS. We found that PC numbers increased significantly in the skin-draining lymph node (DLN), peaking at day 10 and abruptly decreasing by day 14 after DENV inoculation. Class-switched IgG+ PCs appeared from day 7 and dominated the response, while in contrast, the frequency of IgM+ PCs decreased from day 7 onwards. Even though the kinetic of the response was similar between DENV- and iDENV-inoculated mice, the intensity of the response was significantly different. Interestingly, we demonstrated a similar PC response to virus antigens (E and prM) by ELISPOT. In situ characterization showed that PCs were distributed in the medullary cords and in close proximity to germinal centers (GCs), suggesting both an extrafollicular and a GC origin. Proliferating PCs (Ki-67+) were found as early as 3-day postinoculation, and in-depth analysis showed that these PCs were in active phases of cell cycle during the kinetic. Finally, we found a progressive appearance of high-affinity neutralizing DENV-specific IgG further supporting GC involvement. Of note, these antibodies seem to be highly cross-reactive, as a large proportion recognizes Zika virus (ZIKV). The strong PC response to skin-inoculated DENV in this work resembles the findings already described in humans. We consider that this study contributes to the understanding of the in vivo biology of the humoral immune response to DENV in an immunocompetent murine model.

Dengue virus infection (DENV-2) is transmitted by infected mosquitoes via the skin, where many dermal and epidermal cells are potentially susceptible to infection. Most of the cells in an area of infection will establish an antiviral microenvironment to control viral replication. Although cumulative studies report permissive DENV-2 infection in dendritic cells, keratinocytes, and fibroblasts, among other cells also infected, little information is available regarding cell-to-cell crosstalk and the effect of this on the outcome of the infection. Therefore, our study focused on understanding the contribution of fibroblast and dendritic cell crosstalk to the control or promotion of dengue. Our results suggest that dendritic cells promote an antiviral state over fibroblasts by enhancing the production of type I interferon, but not proinflammatory cytokines. Infected and non-infected fibroblasts promoted partial dendritic cell maturation, and the fibroblast-matured cells were less permissive to infection and showed enhanced type I interferon production. We also observed that the soluble mediators produced by non-infected or Poly (I:C) transfected fibroblasts induced allogenic T cell proliferation, but mediators produced by DENV-2 infected fibroblasts inhibited this phenomenon. Additionally, the effects of fibroblast soluble mediators on CD14+ monocytes were analyzed to assess whether they affected the differentiation of monocyte derived dendritic cells (moDC). Our data showed that mediators produced by infected fibroblasts induced variable levels of monocyte differentiation into dendritic cells, even in the presence of recombinant GM-CSF and IL-4. Cells with dendritic cell-like morphology appeared in the culture; however, flow cytometry analysis showed that the mediators did not fully downregulate CD14 nor did they upregulate CD1a. Our data revealed that fibroblast-dendritic cell crosstalk promoted an antiviral response mediated manly by type I interferons over fibroblasts. Furthermore, the maturation of dendritic cells and T cell proliferation were promoted, which was inhibited by DENV-2-induced mediators. Together, our results suggest that activation of the adaptive immune response is influenced by the crosstalk of skin resident cells and the intensity of innate immune responses established in the microenvironment of the infected skin.

Gaining knowledge of the neoplastic side of the three main cells—B cells, Follicular Helper T (Tfh) cells, and follicular dendritic cells (FDCs) —involved in the germinal center (GC) reaction can shed light toward further understanding the microuniverse that is the GC, opening the possibility of better treatments. This paper gives a review of the more complex underlying mechanisms involved in the malignant transformations that take place in the GC. Whilst our understanding of the biology of the GC-related B cell lymphomas has increased—this is not reviewed in detail here—the dark side involving neoplasms of Tfh cells and FDCs are poorly studied, in great part, due to their low incidence. The aggressive behavior of Tfh lymphomas and the metastatic potential of FDCs sarcomas make them clinically relevant, merit further attention and are the main focus of this review. Tfh cells and FDCs malignancies can often be misdiagnosed. The better understanding of these entities linked to their molecular and genetic characterization will lead to prediction of high-risk patients, better diagnosis, prognosis, and treatments based on molecular profiles.

Background Co-circulation of dengue virus (DENV) and chikungunya virus (CHIKV) is increasing worldwide but information on the viral dynamics and immune response to DENV-CHIKV co-infection, particularly in young infants, is scant. Methods Blood samples were collected from 24 patients, aged 2 months to 82 years, during a CHIKV outbreak in Mexico. DENV and CHIKV were identified by RT-PCR; ELISA was used to detect IgM and IgG antibodies. CHIKV PCR products were cloned, sequenced and subjected to BLAST analysis. To address serological findings, HMEC-1 and Vero cells were inoculated with DENV-1, DENV-2 and CHIKV alone and in combination (DENV-2-CHIKV and DENV-1-CHIKV); viral titers were measured at 24, 48 and 72 h. Results Nine patients (38%) presented co-infection, of who eight were children. None of the patients presented severe illness. Sequence analysis showed that the circulating CHIKV virus belonged to the Asian lineage. Seroconversion to both viruses was only observed in the four patients five years or older, while the five infants under two years of age only seroconverted to CHIKV. Viral titers in the CHIKV mono-infected cells were greater than in the DENV-1 and DENV-2 mono-infected cells. Furthermore, we observed significantly increased CHIKV progeny and reduction of DENV progeny in the co-infected cells. Conclusions In our population, DENV-CHIKV co-infection was not associated with increased clinical severity. Our in vitro assay findings strongly suggest that the lack of DENV IgG conversion in the co-infected infants is due to suppression of DENV replication by the Asian lineage CHIKV. The presence of maternal antibody and immature immune responses in the young infants may also play a role. Electronic supplementary material The online version of this article (10.1186/s13071-018-2942-1) contains supplementary material, which is available to authorized users.

Dengue is a global health problem without current specific treatment nor safe vaccines available. While severe dengue is related to pre-existing non-neutralizing dengue virus (DENV) antibodies, the role of T cells in protection or pathology is unclear. Using cutaneous DENV infection in immunocompetent mice we previously showed the generation of PNA+ germinal centers (GCs), now we assessed the activation and proliferation of B and T cells in draining lymph nodes (DLNs). We found a drastic remodelling of DLN compartments from 7 to 14 days post-infection (dpi) with greatly enlarged B cell follicles, occupying almost half of the DLN area compared to ~24% in naive conditions. Enormous clusters of proliferating (Ki-67+) cells inside B follicles were found 14 dpi, representing ~33% of B cells in DLNs but only ~2% in non-infected mice. Inside GCs, we noticed an important recruitment of tingle body macrophages removing apoptotic cells. In contrast, the percentage of paracortex area and total T cells decreased by 14–16 dpi, compared to controls. Scattered randomly distributed Ki-67+ T cells were found, similar to non-infected mice. CD69 expression by CD4+ and CD8+ T cells was minor, while it was remarkable in B cells, representing 1764.7% of change from basal levels 3 dpi. The apparent lack of T cell responses cannot be attributed to apoptosis since no significant differences were observed compared to non-infected mice. This study shows massive B cell activation and proliferation in DLNs upon DENV infection. In contrast, we found very poor, almost absent CD4+ and CD8+ T cell responses.

Lymph nodes (LNs) have evolved to maximize antigen (Ag) collection and presentation as well as lymphocyte proliferation and differentiation - processes that are spatially regulated by stromal cell subsets, including fibroblastic reticular cells (FRCs) and follicular dendritic cells (FDCs). Here we showed that naive neonatal mice have poorly organized LNs with few B and T cells and undetectable FDCs, whereas adult LNs have numerous B cells and large FDC networks. Interestingly, immunization on the day of birth accelerated B cell accumulation and T cell recruitment into follicles as well as FDC maturation and FRC organization in neonatal LNs. However, compared with adults the formation of germinal centers was both delayed and reduced following immunization of neonatal mice. Although immunized neonates poorly expressed activation-induced cytidine deaminase (AID), they were able to produce Ag-specific IgGs, but with lower titres than adults. Interestingly, the Ag-specific IgM response in neonates was similar to that in adults. These results suggest that despite an accelerated structural maturation of LNs in neonates following vaccination, the B cell response is still delayed and reduced in its ability to isotype switch most likely due to poor AID expression. Of note, naive pups born to Ag-immunized mothers had high titres of Ag-specific IgGs from day 0 (at birth). These transferred antibodies confirm a mother-derived coverage to neonates for Ags to which mothers (and most likely neonates) are exposed, thus protecting the neonates while they produce their own antibodies. Finally, the type of Ag used in this study and the results obtained also indicate that T cell help would be operating at this stage of life. Thus, neonatal immune system might not be intrinsically immature but rather evolutionary adapted to cope with Ags at birth.

Dengue virus (DENV) is one of the most significant human viral pathogens transmitted by mosquitoes and can cause from an asymptomatic disease to mild undifferentiated fever, classical dengue, and severe dengue. Neutralizing memory antibody (Ab) responses are one of the most important mechanisms that counteract reinfections and are therefore the main aim of vaccination. However, it has also been proposed that in dengue, some of these class-switched (IgG) memory Abs might worsen the disease. Although these memory Abs derive from B cells by T-cell-dependent processes, we know rather little about the (acute, chronic, or memory) B cell responses and the complex cellular mechanisms generating these Abs during DENV infections. This review aims to provide an updated and comprehensive perspective of the B cell responses during DENV infection, starting since the very early events such as the cutaneous DENV entrance and the arrival into draining lymph nodes, to the putative B cell activation, proliferation, and germinal centers (GCs) formation (the source of affinity-matured class-switched memory Abs), till the outcome of GC reactions such as the generation of plasmablasts, Ab-secreting plasma cells, and memory B cells. We discuss topics very poorly explored such as the possibility of B cell infection by DENV or even activation-induced B cell death. The current information about the nature of the Ab responses to DENV is also illustrated.

Evasion or subversion of host immune responses have been shown for a variety of microorganisms, and this might be the case for Trichophyton rubrum, the most common pathogenic fungus causing chronic dermatophytosis in humans. Keratinocytes, the main epidermal cells, have important roles as a first defense against microbial challenges in local immune reactions. Epidermal keratinocytes express several Toll-like receptors and produce host defense peptides, cytokines and chemokines in response to various stimuli. We analyzed the expression of Toll-Like receptor TLR2, TLR4, TLR6, and Human Beta Defensin (HBD)-1, HBD-2, Interleukin IL-1b and IL-8 production, when exposing primary keratinocyte cultures to T. rubrum. We observed changes in size and granularity of keratinocytes stimulated with either whole conidia or conidial homogenates compared to other treatments. Intact conidia decreased keratinocytes’ TLR2 and TLR6 expression without affecting that of TLR4, while conidial homogenates increased the expression of these three receptors. Interestingly, whole conidia decreased HBD-1 and HBD-2 production, whereas conidial homogenate increased it. No changes were observed in IL-1b and IL-8 production after stimulation with conidia or conidial homogenate. CONCLUSIONS. Our results suggest that: 1) Keratinocytes can recognize and respond to cell wall components of T. rubrum; 2) Viable intact conidia inhibit TLR-2 and TLR6 expression and decrease HBD-1 and HBD-2 production; 3) Conidial homogenate from T. rubrum increases the expression of TLR2, TLR4 and TLR6 and induces HBD-1 and HBD-2 production; 4) Therefore, innate immune functions of keratinocytes as the first level of local skin immunity are apparently manipulated by T. rubrum, likely to ensure its establishment, persistence and survival.

This study used an in vivo mouse model to analyze the response of dendritic cells (DCs) in Peyer's patches (PPs) within the first 48 h of infection with the wild-type murine rotavirus EDIM (EDIM wt ). After the infection, the absolute number of DCs was increased by 2-fold in the PPs without a modification of their relative percentage of the total cell number. Also, the DCs from PPs of infected mice showed a time-dependent migration to the subepithelial dome (SED) and an increase of the surface activation markers CD40, CD80, and CD86. This response was more evident at 48 h postinfection (p.i.) and depended on viral replication, since DCs from PPs of mice inoculated with UV-treated virus did not show this phenotype. As a result of the activation, the DCs showed an increase in the expression of mRNA for the proinflammatory cytokines interleukin-12/23p40 (IL-12/23p40), tumor necrosis factor alpha (TNF-α), and beta interferon (IFN-β), as well as for the regulatory cytokine IL-10. These results suggest that, a short time after rotavirus infection, the DCs from PPs play a critical role in controlling the infection and, at the same time, avoiding an excessive inflammatory immune response.

Dengue virus (DENV) has four serotypes, which can cause from asymptomatic disease to severe dengue. Heterologous secondary infections have been associated to a greater risk of potentially fatal dengue due to non-neutralizing memory antibodies (Abs), which facilitate the infection, such as anti-precursor membrane (prM) Abs, among other mechanisms. Usually, class-switched memory Abs are generated mainly through germinal centers (GCs). However, the cellular events underlying these Ab responses to DENV, especially during repeated/secondary infections, have been poorly studied. We wanted to know whether there is involvement of GC reactions during cutaneous DENV infection and whether there is any sort of preferential Ab responses to defined viral proteins. Intradermal DENV inoculation at a relatively low dose efficiently infects immune-competent BALB/c mice, inducing higher quantities of DENV-specific GC B cells and larger GCs than the control conditions. Interestingly, GCs exhibited as much prM as envelope (E) and non-structural 3 viral proteins in situ. Intriguingly, despite the much larger abundance of E protein than of prM protein in the virions, infected animals showed similar amounts of circulating Abs and Ag-specific GC B cells both for prM and for E proteins, even significantly higher for prM. To the best of our knowledge, there are no reports of the GC responses during DENV infection. This relatively stronger anti-prM response could be triggered by DENV to preferentially promote Abs against certain viral proteins, which might favor infections by facilitating DENV invasion of host cells. It is thus conceivably that DENV might have evolved to induce this kind of Ab responses.

Dendritic cells (DC) are often arranged in planar layers in tissues with high antigenic exposure, such as skin and mucosae. Providing an en face view, this arrangement optimizes in situ analysis regarding morphology (even of individual dendrites), topographic distribution (regular/clustered) and quantification. The few reports on human genital DC usually utilize single markers and conventional sections, restricting immunolabelling only to cell parts sectioned by the cut. To better assess DC in situ, we labelled epithelial sheets, prepared from fresh cervix biopsies, with antibodies to major histocompatibility complex (MHC)-CII, CD1a and Langerin, revealing (with each of these markers) a dense DC network in a planar-like, regular distribution. Using the hybrid capture system to detect the high-risk mucotropic human papilloma virus (HPV) group, 16 positive and five negative women were studied and the results were compared between these groups. DC frequency per area was substantially reduced (to approximately 50% for the three markers) in samples from all HPV-infected patients compared with samples from controls. Unlike HPV(-) samples, Langerin(+) DC in HPV(+) cervix exhibited a highly accentuated dendritic appearance. We believe this to be the first study using these three DC-restricted markers (Langerin, CD1a and MHC-CII) in cervical epithelial sheets from high-risk HPV(+) donors and also the first study to demonstrate the morphological and quantitative changes triggered by high-risk HPV infection. Cervical DC reduction in early, premalignant high-risk HPV infection might represent viral subversion strategies interfering with efficient antigen handling by the immune system's peripheral sentinels, the DC, perhaps hampering appropriate recruitment and subsequent development of effector (cytotoxic) T cells.

Dengue virus (DV) infections are serious causes of morbidity and mortality worldwide, and adaptive (secondary) immune response appears to influence the severity of this arboviral disease (Morens 1994; Halstead & O'Rourke 1977; Rigau-Perez et al. 1998). There are an estimated 50–100 million cases of dengue fever and 250,000-500,000 cases of dengue haemorrhagic fever (DHF) annually in the world (WHO 1997). The dengue shock syndrome (DSS), the most severe and potentially fatal form of the disease, especially in developing countries, is less frequent. In the model proposed here, we have chosen to examine DV2, because it is the prevalent serotype (Gomez-Dantes et al. 2004). The high seroprevalence together with the co-circulation of multiple serotypes indicates that Mexico could be in risk of an important DHF outbreak (Gomez-Dantes et al. 2004). The exact pathogenic mechanisms following DV infection are not well understood, especially in the severe forms of the infection (DHF and DSS). The immune response to DV seems at least partially involved in the process (Halstead 1988; Avirutnan et al. 1998; Diamond et al. 2000). Several theories have been advanced including facilitation of DV infection by non-neutralizing antibodies (Halstead & O'Rourke 1977; Halstead et al. 1977; Morens 1994), the potential differential virulence of each infecting serotype (Leitmeyer et al. 1999; Kawaguchi et al. 2003), CD4-CD8 T-cell cross-reactivity and host factors such as a particular, but yet undefined susceptibility (Klenerman & Zinkernagel 1998; Mongkolsapaya et al. 2003). While there is an excellent research on dengue about the molecular structure and epidemiology of this virus, the basic mechanisms of the immunopathology still remain obscure (Haywood 1994; Kuhn et al. 2002). For instance, until very recently (Johnston et al. 2000; Wu et al. 2000), it was unknown whether DV was able to replicate in cutaneous cells. Likewise, there are two other features largely unexplored regarding DV infection: (i) the very early stages of the infection, because, by necessity, patients are examined when disease is almost over, when the patients are under recovery or when they have entered into DSS and (ii) which type of responses, if any, occur cutaneously, in situ, following DV inoculation when mosquitoes feed. We wanted to develop an in situ approach that would enable us to study the early immune pathology of the cellular events regarding the infectious process and the first reactions of the peripheral immune system, the dendritic cells (DCs). DCs constitute a system of highly dynamic sentinel cells, whose most studied element in peripheral non-lymphoid tissues is the epidermal Langerhans cells (LCs). It is now known that LCs perform multiple tasks including antigen capturing and processing, antigen ferrying to regional lymphoid tissues, and ultimately, cognate antigen presentation to naive lymphocytes, once in secondary lymphoid organs (Flores-Romo 2001). Under the influence of a variety of stimuli such as cytokines, antigens or microbial products, LCs become activated, start to migrate into the dermis and concomitantly to up-regulate antigen-presenting molecules and to express co-stimulatory and maturation markers (Cumberbatch & Kimber 1992; Cumberbatch et al. 1997). Nevertheless, despite that LCs as sensors of the external antigenic world are the most exposed cells of the immune system during mosquito feeding, and the initial virus inoculation through skin, the interactions of these cells with DV and the outcome of such potential interplay regarding both the virus and the LC have received little attention in situ (Wu et al. 2000). We attempted to address some of these issues by establishing a model with fresh, healthy, non-cadaveric human skin explants exposed to DV, to analyse whether local infection and viral replication are feasible experimentally, to assess the most early phases of the initiation of the immune responses in situ and the immediate local repercussions of viral inoculation upon local antigen-presenting cells (APCs), particularly on the DCs, the sentinel posts of the immune system in the skin.

SummaryAtherosclerosis is a complex disease involved in major fatal events such as myocardial infarction and stroke. It is the result of interactions between metabolic, dietetic and environmental risk factors acting on a genetic background that could result in endothelial susceptibility. Our aim was to determine the patterns of expression of adhesion molecules and whether phosphatidylserine is translocated to the cell surface of human umbilical vein endothelial cells (HUVECs) isolated from healthy newborns born to parents with a strong family history of myocardial infarction under TNF-α or oxLDL stimulated conditions. Compared to control HUVECs, experimental cords showed: (a) a four-fold increase in VCAM-1 expression under basal conditions, which showed no change after stimulation with the pro-atherogenic factors; (b) a two-fold increase in basal P-selectin expression that reached a 10-fold increase with any of the pro-atherogenic factors; (c) a basal ICAM-1 expression similar to P-selectin that was not modified by the pro-atherogenic molecules; (d) a similar PECAM-1 expression. Unexpectedly, phospathidylserine expression in experimental cord HUVECs was significantly increased (211 817 versus 3354 TFU) but was not associated to apoptotic death as the percentage of dead cells induced by TNF-α treatment was very low (0·55 versus 9·87% in control HUVECs). The latter result was corroborated by TUNEL staining. T cell adherence to HUVECs was highly up-regulated in the genetically predisposed samples. The analysis of nonpooled HUVECs, from newborns to family predisposed myocardial-infarction individuals, might represent a useful strategy to identify phenotypical and functional alterations, and hopefully, to take early preventive actions.

Pulmonary tuberculosis, an infection caused by Mycobacterium tuberculosis has been declared a global health emergency by the World Health Organization (WHO). Tuberculosis is a re-emergent expanding disease, responsible for a great mortality and morbidity throughout the world. It has been estimated that one-third of the world's population is currently infected with M. tuberculosis, with 8 million new cases per year. Nevertheless, only 5–10% will develop the chronic pulmonary disease (Dye et al. 1999; WHO 2000). Most people will be able to control the infection through efficient immune responses, especially the cellular arm of immunity. However, the bacilli might not be eliminated completely and may persist dormant for years in a latent state, a phenomenon not well understood yet (Flynn & Ernest 2000; Hernandez-Pando et al. 2000; Manabe & Bishai 2000). M. tuberculosis spreads from a person with pulmonary tuberculosis through aerosols, entering the body via the respiratory tract, reaching the lungs where phagocytic cells become infected, especially the alveolar macrophages (AMs). Apparently, M. tuberculosis can survive within AM for long-lasting periods, causing either a latent infection or subsequently developing the pulmonary disease (Wiegeshaus et al. 1989; Andersen 1997). This bacillus has evolved several mechanisms to evade the host immune responses, including the inhibition of antimicrobial activity of macrophages (Schluger & Rom 1998). It is known that the first line of defence against M. tuberculosis is the AM, which interacts with the bacilli through different surface molecules like complement, mannose and Toll-like receptors (Arthur & Dannenberg 1991; Schluger & Rom 1998). Despite the fact that dendritic cells (DCs) are present throughout the respiratory tract, from the upper nasal mucosa to parenchymal lung tissue, little is known about lung DCs in vivo during tuberculosis, either in the naturally occurring disease or in experimental models of infection. Lung DCs are found within pleura and the alveolar septal walls; additionally they can be found, although in very small numbers, in the alveolar spaces (Holt & Schon–Hegrad 1987; McWilliam et al. 1995; Holt 2000). In vitro assays have shown that monocyte-derived DCs and macrophages internalize the mycobacteria, resulting in DC maturation and activation with a differential production of cytokines. Interleukin-12 (IL-12) was almost exclusively produced by DCs, while macrophages produced IL-10 and the pro-inflammatory cytokines tumour necrosis factor-α (TNF-α), IL-1 and IL-6 (Mohagheghpour et al. 2000; Giacomini et al. 2001). M. tuberculosis grew equally well within nonactivated DCs and macrophages; however, activation with IFN-γ and LPS inhibited the growth of this intracellular microorganism. While macrophages can kill the intracellular bacteria, DCs seem unable to do so and, apparently, only restrict the growth of this bacillus (Bodnar et al. 2001). Experiments conducted in vivo with the M. bovis (BCG) showed that, following intravenous administration, bacilli were detected in similar percentage in splenic macrophages and in splenic DCs. After 2 weeks, BCG bacilli were still viable without apparent growth within DCs (Jiao et al. 2001). In this study, using four different markers in situ, we describe the main lung antigen-presenting cells (APCs) in the susceptible BALB/c mice during experimental pulmonary tuberculosis, which is induced by intratracheal inoculation of the virulent strain of M. tuberculosis H37Rv. Pulmonary infection with M. tuberculosis induced an increase of parenchymal lung APC; these cells displayed differential tissue distribution and morphological features during the infection and, apparently, also a differential capacity to interact with bacilli in vivo.

CD40, a glycoprotein expressed on B lymphocytes plays an important role in B cell development, growth and differentiation. The ligand for the CD40 is a 39-kDa glycoprotein (CD154) expressed on the surface of activated T lymphocytes and is essential for thymus-dependent humoral immunity. The expression of CD154 is tightly regulated and its transient expression reduces the chances of potentially deleterious bystander activation of B cells. Stimulation through CD40 has been studied in vitro by using antibodies against CD40, by membranes of activated T cells or lately, by CD154 transfected cells. In this work we have evaluated the outcome of CD40-CD40 ligand interaction in vitro and in vivo by using CD154-transfected L929 cells. In vitro assays showed that CD154-L929 cells can induce on B cells: IL-4-dependent proliferation, up-regulation of CD23, CD54 and class II molecules and can also rescue WEHI-231 B cell lymphoma from anti-IgM-induced apoptosis. Interestingly, in vivo assays revealed that when CD154-L929 cells were inoculated into the spleen, mice developed a strong but transient production of anti-erythrocyte autoantibodies. Through B lymphocyte activation with CD154-transfected L929 cells both in vitro and in vivo, our data reveal that enforced and prolonged expression of CD40 ligand overcomes the tightly regulated mechanisms of B cell activation, triggering the production of autoantibodies. This system might be used to evaluate the early steps of an autoimmune response and the role of CD40-CD154 in the induction of primary responses in vivo.

Whereas CD40–CD40 ligand interactions are important for various dendritic cell (DC) functions in vitro, their in vivo relevance is unknown. We analyzed the DC status of CD40 ligand −/− mice using a contact hypersensitivity (CHS) model system that enables multiple functions of DCs to be assessed in vivo. Immunohistochemistry of skin sections revealed no differences in terms of numbers and morphology of dendritic epidermal Langerhans cells (LCs) in unsensitized CD40 ligand −/− mice as compared with wild-type C57BL/6 mice. However, after contact sensitization of CD40 ligand −/− mice, LCs failed to migrate out of the skin and substantially fewer DCs accumulated in draining lymph nodes (DLNs). Furthermore, very few antigen-bearing DCs could be detected in the paracortical region of lymph nodes draining sensitized skin. This defect in DC migration after hapten sensitization was associated with defective CHS responses and decreased cutaneous tumor necrosis factor (TNF)-α production and was corrected by injecting recombinant TNF-α or an agonistic anti-CD40 monoclonal antibody. Thus, CD40–CD40 ligand interactions in vivo regulate the migration of antigen-bearing DCs from the skin to DLNs via TNF-α production and play a vital role in the initiation of acquired T cell–mediated immunity.

Human CD34+ multilineage progenitor cells (CD34HPC) from cord blood and bone marrow express CD40, a member of the tumor necrosis factor–receptor family present on various hematopoietic and nonhematopoietic cells. As hyper-IgM patients with mutated CD40 ligand (CD40L) exhibit neutropenia, no B cell memory, and altered T cell functions leading to severe infections, we investigated the potential role of CD40 on CD34HPC development. CD40activated cord blood CD34HPC were found to proliferate and differentiate independently of granulocyte/macrophage colony-stimulating factor, into a cell population with prominent dendritic cell (DC) attributes including priming of allogeneic naive T cells. DC generated via the CD40 pathway displayed strong major histocompatibility complex class II DR but lacked detectable CD1a and CD40 expression. These features were shared by a dendritic population identified in situ in tonsillar T cell areas. Taken together, the present data demonstrate that CD40 is functional on CD34HPC and its cross-linking by CD40L+ cells results in the generation of DC that may prime immune reactions during antigen-driven responses to pathogenic invasion, thus providing a link between hematopoiesis, innate, and adaptive immunity.

Analysis of the cDNA encoding murine interleukin (IL) 17 (cytotoxic T lymphocyte associated antigen 8) predicted a secreted protein sharing 57% amino acid identity with the protein predicted from ORF13, an open reading frame of Herpesvirus saimiri. Here we report on the cloning of human IL-17 (hIL-17), the human counterpart of murine IL-17. hIL-17 is a glycoprotein of 155 amino acids secreted as an homodimer by activated memory CD4+ T cells. Although devoid of direct effects on cells of hematopoietic origin, hIL-17 and the product of its viral counterpart, ORF13, stimulate epithelial, endothelial, and fibroblastic cells to secrete cytokines such as IL-6, IL-8, and granulocyte-colony-stimulating factor, as well as prostaglandin E2. Furthermore, when cultured in the presence of hIL-17, fibroblasts could sustain the proliferation of CD34+ hematopoietic progenitors and their preferential maturation into neutrophils. These observations suggest that hIL-17 may constitute (a) an early initiator of the T cell-dependent inflammmatory reaction; and (b) an element of the cytokine network that bridges the immune system to hematopoiesis.

Tuberculosis (TB), one of the major world health problems, is a chronic infection caused by members of the Mycobacterium tuberculosis complex (MTC). In 2009, tuberculosis (TB) caused 1.7 million deaths and 9.4 million new cases. Although recent efforts to improve TB prevention, diagnosis and treatment have contributed to a 35% decrease in the death rate, the emergence of mycobacterial strains with highly virulent phenotypes combined with pandemic HIV infections has added new challenges to control TB.

The scavenger receptor family comprises transmembrane proteins involved in the recognition of polyanionic ligands. Several studies have established that members of this family are involved both in immunity and in developmental processes. In Drosophila melanogaster, one of the best characterized scavenger receptors is Croquemort, which participates in the recognition of apoptotic cells in the embryo. Although comparative genomic studies have revealed the presence of four orthologs of this receptor in the malaria vector Anopheles gambiae, little is known about their function. We have investigated the expression pattern of the four Croquemort orthologs during the mosquito life cycle. Croquemort transcripts SCRBQ2 and SCRBQ4 are expressed at all the developmental stages, while expression of Croquemort transcripts SCRBQ1 and SCRBQ3 is more restricted. We have also investigated the expression of the four Croquemort orthologs in the different organs of the adult female. Croquemort transcript SCRBQ2 is highly expressed in the A. gambiae female midgut. SCRBQ2 midgut gene expression was up-regulated after a non-infected or a Plasmodium berghei-infected blood meal, compared to its expression in midguts of sugar-fed females. Interestingly, knockdown of SCRBQ2 expression by dsRNA injection resulted in a 62.5% inhibition of oocyst formation, suggesting that SCRBQ2 plays a role in Plasmodium-mosquito interactions.

Identifying the properties of a molecule involved in the efficient activation of the innate and adaptive immune responses that lead to long-lasting immunity is crucial for vaccine and adjuvant development. Here we show that the papaya mosaic virus (PapMV) is recognized by the immune system as a pathogen-associated molecular pattern (PAMP) and as an antigen in mice (Pamptigen). A single immunization of PapMV without added adjuvant efficiently induced both cellular and specific long-lasting antibody responses. PapMV also efficiently activated innate immune responses, as shown by the induction of lipid raft aggregation, secretion of pro-inflammatory cytokines, up-regulation of co-stimulatory molecules on dendritic cells and macrophages, and long-lasting adjuvant effects upon the specific antibody responses to model antigens. PapMV mixed with Salmonella enterica serovar Typhi (S. typhi) outer membrane protein C increased its protective capacity against challenge with S. typhi, revealing the intrinsic adjuvant properties of PapMV in the induction of immunity. Antigen-presenting cells loaded with PapMV efficiently induced antibody responses in vivo, which may link the innate and adaptive responses observed. PapMV recognition as a Pamptigen might be translated into long-lasting antibody responses and protection observed. These properties could be used in the development of new vaccine platforms.

"Los mamíferos producen lactoferrina, una glicoproteína monomérica no hémica de unión a hierro que es considerada componente esencial de la inmunidad innata del hospedero.El objetivo de esta revisión es presentar los principales efectos inmunorreguladores de la lactoferrina heteróloga, administrada por vía oral o parenteral en animales de experimentación sanos o infectados con bacterias patógenas. La lactoferrina bovina y la humana estimulan o inhiben diversos componentes humorales y celulares de la inmunidad implicados en la prevención y/o resolución de infecciones y de la inflamación asociada a estas últimas. El efecto modulador de la lactoferrina heteróloga tiene implicaciones prácticas por sus posibles aplicaciones terapéuticas, profilácticas y adyuvantes. La comprensión integral de la influencia de la lactoferrina sobre la respuesta inmunitaria y sus limitantes son de fundamental relevancia para su aplicación racional en el control de diversas enfermedades causadas por bacterias patógenas."

Dendritic cells (DCs) are located at body surfaces such as the skin, respiratory and genital tracts, and intestine. To further analyze intestinal DCs, we adapted an epidermal sheet separation technique and obtained two intestinal layers, facing the lumen and serosa. Unexpectedly, immunolabeling of the layer toward the serosa revealed a regular, dense, planar network of cells with prominent dendritic morphology within the external muscular layer and with increasing frequency along the length of the intestine. Direct examination of the serosal-disposed layers showed a significant fraction of the DCs to express DEC-205/CD205, CD11c, Langerin/CD207, Fcγ receptor/CD16/32, CD14, and low levels of activation markers, CD25, CD80, CD86, and CD95. By more sensitive FACS analyses, cells from this layer contained two CD11c+populations of CD45+CD205+, CD19-leukocytes, MHC II+and MHC II-. When ovalbumin conjugated to an anti-DEC-205 antibody was injected into mice, the conjugate targeted to these DCs, which upon isolation were able to stimulate ovalbumin-specific, CD4+and CD8+T cell antigen receptor-transgenic T cells.In vivo, these DCs responded to two microbial stimuli, systemic LPS and oral live bacteria, by up-regulating CD80, CD86, DEC-205, and Langerin within 12 h. This network of DCs thus represents a previously unrecognized antigen-presenting cell system in the intestine.

SummaryIt has been shown recently that different genotypes of Mycobacterium tuberculosis induce distinct immune responses in the host, as reflected by variations in cytokine and iNOS expression. Because these molecules are probably regulated by multiple factors in vivo this complex phenomenon was partially analysed by assessing cytokine and iNOS expression by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) in an in vitro model of bone marrow-derived macrophages infected with three different M. tuberculosis genotypes: Canetti, H37 Rv and Beijing. Although the three genotypes induced production of iNOS and the different cytokines tested at 24 h post-infection, macrophages infected with the Beijing isolate expressed the highest levels of mRNA for iNOS, interleukin (IL)-1β, tumour necrosis factor (TNF)-α, IL-12 cytokines and lower levels of IL-10 compared with cells infected with other genotypes. This expression pattern has been associated with infection control, but during infection in vivo with the Beijing genotype it is lost upon progression to chronic phase. The failure to control infection is likely to be influenced by cytokines produced by other cell types and bacterial molecules expressed during the course of disease. Results presented in this work show that each genotype has the ability to induce different levels of cytokine expression that could be related to its pathogenesis during infection.

La respuesta inmune desarrollada durante la infección por Entamoeba histolytica no ha sido completamente determinada. La inmunidad mediada por células se ha asociado a la resolución de la amibiasis, principalmente por neutrófilos y macrófagos. La generación de mediadores de oxígeno por macrófagos es uno de los principales mecanismos de ataque contra este parásito. Otras células del sistema inmune, tales como los eosinófilos se han relacionado con la limitación de la infección. La generación de citocinas y quimiocinas por parte de macrófagos, neutrófilos, linfocitos y células epiteliales no sólo dirige la migración de las células que están encargadas de destruir al parásito, sino que también orquesta la respuesta inmune que llevará a la efectiva resolución de la infección. Hasta el momento, no está clara la respuesta inmune que efectivamente ataca a este parásito y que contribuye a resolver la amibiasis, esto es debido en parte a la variabilidad de los modelos usados, por lo que el objetivo de esta revisión es concluir cuál es el papel de la respuesta inmune celular desarrollada durante la amibiasis que contribuye a controlar la infección.

SUMMARY Despite tuberculosis resurgence and extensive dendritic cell (DC) research, there are no in vivo studies evaluating DC within regional lymphoid tissue during airways infection with virulent Mycobacterium tuberculosis (Mtb) H37Rv. Using DC-specific antibodies, immunocytochemistry, flow cytometry and Ziehl–Neelsen (ZN) for bacilli staining, we searched for Mtb and DC changes within mediastinal lymph nodes, after intratracheal (ITT) inoculation of virulent Mtb. ZN and immunocytochemistry in frozen and paraffin sections of mediastinal lymph nodes identified Mtb until day 14 after ITT inoculation, associated with CD11c + and Dec205 + DC. Analysing CD11c, MHC-CII, and Dec205 combinations by flow cytometry in MLN suspensions revealed that CD11c + /MHC-CII + and CD11c + /Dec205 + DC did not increase until day 14, peaked on day 21, and sharply declined by day 28. No changes were seen in control, salineinoculated animals. The costimulatory molecules evaluated in CD11c + DCs followed a similar trend; the CD80 increase was negligible, slightly surpassed by CD40. CD86 increased earlier and the three markers peaked at day 21, declining by day 28. While antigen-specific proliferation was not evident for MLN CD4 + T cells at 2 weeks postinfection, delayed-type hypersensitivity responses upon ITT inoculation revealed that, as early as day 3 and 7, both the priming and peripheral systemic immune responses were clearly established, persisting until days 14–21. While airways infection with virulent Mtb triggers an early, systemic peripheral response maintained for three weeks, this seems dissociated from regional events within mediastinal lymph nodes, such as antigen-specific T-cell reactivity and a delay in the influx and local activation of DC.

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.

Dendritic cells (DC) are cells of the hematopoletic system specialized in capturing antigens and initiating T cell-mediated immune responses. We show here that human DC generated in vitro by culturing CD34+ cord blood progenitor cells in granulocyte macrophage colony stimulating factor plus tumor necrosis factor-alpha express the Fas antigen (APO-1, CD95) and undergo apoptosis upon triggering of Fas by mAb. However, only a proportion of the cells die in response to Fas ligation, an observation that may be related to the virtual absence of the bcl-2 protein in about half of the cells. Ligation of DC CD40 by culture on CD40L-transfected fibroblastic cells up-regulates the expression of bcl-2 and, concomitantly, renders DC virtually resistant to Fas-induced apoptosis. Parallel experiments with mature, interdigitating dendritic cells (IDC) isolated from tonsils revealed that IDC express Fas but do not enter into apoptosis following Fas ligation, a finding that may be explained by their high levels of bcl-2. Thus, upon encountering antigen-specific T cells, DC become resistant to Fas-induced apoptosis, as a consequence of CD40 ligation and possibly by mechanisms associated to the up-regulation of bcl-2 protein expression.

Background When mosquitoes infected with DENV are feeding, the proboscis must traverse the epidermis several times (“probing”) before reaching a blood vessel in the dermis. During this process, the salivary glands release the virus, which is likely to interact first with cells of the various epidermal and dermal layers, cells which could be physiologically relevant to DENV infection and replication in humans. However, important questions are whether more abundant non-hematopoietic cells such as fibroblasts become infected, and whether they play any role in antiviral innate immunity in the very early stages of infection, or even if they might be used by DENV as primary replication cells. Methodology/Principal Findings Fibroblasts freshly released from healthy skin and infected 12 hours after their isolation show a positive signal for DENV. In addition, when primary skin fibroblast cultures were established and subsequently infected, we showed DENV-2 antigen-positive intracellular signal at 24 hours and 48 hours post-infection. Moreover, the fibroblasts showed productive infection in a conventional plaque assay. The skin fibroblasts infected with DENV-2 underwent potent signaling through both TLR3 and RIG- 1, but not Mda5, triggering up-regulation of IFNβ, TNFα, defensin 5 (HB5) and β defensin 2 (HβD2). In addition, DENV infected fibroblasts showed increased nuclear translocation of interferon (IFN) regulatory factor 3 (IRF3), but not interferon regulatory factor 7 (IRF7), when compared with mock-infected fibroblasts. Conclusions/Significance In this work, we demonstrated the high susceptibility to DENV infection by primary fibroblasts from normal human skin, both in situ and in vitro. Our results suggest that these cells may contribute to the pro-inflammatory and anti-viral microenvironment in the early stages of interaction with DENV-2. Furthermore, the data suggest that fibroblast may also be used as a primary site of DENV replication and provide viral particles that may contribute to subsequent viral dissemination., Author Summary In this work, we demonstrate that that both human whole skin and freshly isolated skin fibroblasts are productively infected with Dengue virus (DENV). In addition, primary skin fibroblast cultures were established and subsequently infected with DENV-2; we showed in these cells the presence of the viral antigen NS3, and we found productive viral infection by a conventional plaque assay. Of note, the infectivity rate was almost the same in all the primary cultures analyzed from different donors. The skin fibroblasts infected with DENV-2 underwent signaling through both TLR3 and RIG-1, but not Mda5, triggering up-regulation of IFNβ, TNFα, defensin 5 (HB5) and β defensin 2 (HβD2). In addition, DENV infected fibroblasts showed increased nuclear translocation of interferon (IFN) regulatory factor 3 (IRF3), but not interferon regulatory factor 7 IRF7, when compared with mock-infected fibroblasts. Our data suggest that fibroblasts might even participate producing mediators involved in innate immunity that activate and contribute to the orchestration of the local innate responses. This work is the first evaluating primary skin fibroblast cultures obtained from different humans, assessing both their susceptibility to DENV infection as well as their ability to produce molecules crucial for innate immunity.

Immunoglobulin E (IgE) is central to the induction of allergic diseases through its binding to the high-affinity receptor (Fc epsilon R1) on mast cells and basophils. Crosslinking by allergens of the bound IgE leads to the release of various inflammatory mediators. IgE production by B cells requires a physical interaction with T cells, involving a number of surface adhesion molecules, as well as the soluble factors interleukin-4 (IL-4) and IL-13 (ref. 5) produced by T cells, basophils and mast cells. Here we report that, in the presence of IL-4, mast and basophilic cell lines can provide the cell contact signals that are required for IgE synthesis. The human cell lines HMC-1 (mast) and KU812 (basophilic) both express the ligand for CD40 (CD40L) which is shown to be responsible for the IgE production. Moreover, freshly isolated purified human lung mast cells and blood basophils are also shown to express CD40L and to induce IgE production. This evidence suggests that mast cells and basophils may therefore play a key role in allergy not only by producing inflammatory mediators, but also by directly regulating IgE production independently of T cells.