Human Immature Dendritic Cells Research Articles

Killed Whole-Cell Oral Cholera Vaccine Induces CCL20 Secretion by Human Intestinal Epithelial Cells in the Presence of the Short-Chain Fatty Acid, Butyrate

Short-chain fatty acids (SCFAs), such as acetate, butyrate, and propionate, modulate immune responses in the gut. However, the effect of SCFAs on mucosal vaccine-induced immune cell migration is poorly understood. Here, we investigated whether SCFAs modulate chemokine expression induced by the killed whole-cell oral cholera vaccine, Shanchol™, in human intestinal epithelial cells. Shanchol™ induced expression of CCL2, CCL5, CCL20, and CXCL10 at the mRNA level, but not at the protein level. Interestingly, CCL20 secretion was substantially increased by co-stimulation with Shanchol™ and butyrate, while neither acetate nor propionate showed such effect. Enhanced CCL20 secretion was associated with GPR109A activation, and histone deacetylase (HDAC) inhibition. In addition, co-treatment with Shanchol™ and butyrate synergistically increased the secretion of adenosine triphosphate (ATP). Moreover, CCL20 secretion was decreased by inhibiting the extracellular ATP receptor P2X7. However, neither inflammasomes nor caspases were involved in CCL20 production. The culture supernatant of cells treated with Shanchol™ and butyrate augmented human immature dendritic cell migration. Collectively, these results suggest that butyrate enhances Shanchol™-induced CCL20 production in human intestinal epithelial cells via HDAC inhibition and ATP-P2X7 signaling by activating GPR109A. These effects potentially enhance the mucosal immune responses in the gut induced by this oral cholera vaccine.

A practical approach to assess inhalation toxicity of metal oxide nanoparticles in vitro.

Exposure of humans to metal oxide nanoparticles (NPs) occurs mainly via air, and inhaled metal oxide NPs may generate inflammation. The aim of this study was to investigate the proinflammatory potential of six metal oxide NPs (CeO2 , Mn2 O3 , CuO, ZnO, Co3 O4 and WO3 ; 27-108μg ml-1 ) using human primary 3-dimensional airway epithelium (MucilAir™) and dendritic cell (DC) models. Metal oxide NPs were mainly aggregated/agglomerated in the cell media, as determined by dynamic light scattering, scanning electron microscopy and differential centrifugal sedimentation. WO3 and ZnO were highly soluble, both with and without respiratory mucus. Proinflammatory signalling by the epithelium was evaluated after a 24hour exposure by increased interleukin-6 and -8 and monocyte chemoattractant protein 1 cytokine release, which occurred only for CuO. Moreover, maturation of immature human DCs, which play a key role in the lung immune system, were evaluated by expression of surface markers HLA-DR, CD80, CD83 and CD86 after a 48hour exposure. Only Mn2 O3 consistently upregulated DC maturation markers. Furthermore, by addition of medium from metal oxide NP-exposed 3-dimensional airway cultures to metal oxide NP-exposed DC cultures, the interplay between lung epithelium and DCs was studied. Such an interplay was again only observed for Mn2 O3 and in one of five DC donors. Our results show that, even when using dosages that represent very high in vivo exposure levels, up to 27hours of constant human airway exposure, metal oxide NPs cause minimal proinflammatory effects and that epithelial cells not necessarily interfere with DC maturation upon metal oxide NP exposure. The present approach exemplifies a relevant translation towards human safety assessment.

Combining R-DOTAP and a particulate antigen delivery platform to trigger dendritic cell activation: Formulation development and in-vitro interaction studies

The utilization of the cationic lipid R-DOTAP as immune cell stimulant (e.g. its stimulating effects on immature dendritic cells) and correspondingly as possible adjuvant for vaccination is well known. Likewise, it is described in literature that solid polymer particles loaded with antigens can be size-tailored in a manner to be suitable for phagocytosis by antigen presenting cells. The effects of DOTAP-microparticle combinations, however, are not well understood. This study aimed therefore to explore the potential of R-DOTAP stabilized microparticles (MP) to act as a carrier platform for antigens e.g. for cancer vaccination. It was investigated whether or not a combination of R-DOTAP and PLGA leads to a boosted adjuvant effect in dendritic cell maturation.For proper comparison, neutral and negatively charged MPs of comparable sizes were developed. Toxicity, uptake, routing and maturation of the MP platform was assessed in-vitro on human immature dendritic cells (iDCs).Interestingly, none of the tested placebo formulations (without antigen) was capable to induce DC maturation when compared to LPS as positive control. This is in contrast to experiments previously reported in literature, where R-DOTAP (e.g. in liposomal form) triggered iDC maturation even without antigen. Possible reasons and further approaches are discussed in the paper.

Artificial human antigen-presenting cells are superior to dendritic cells at inducing cytotoxic T-cell responses.

Peptide recognition through the MHC class I molecule by cytotoxic T lymphocytes (CTLs) leads to the killing of cancer cells. A potential challenge for T-cell immunotherapy is that dendritic cells (DCs) are exposed to the MHC class I-peptide complex for an insufficient amount of time. To improve tumour antigen presentation to T cells and thereby initiate a more effective T-cell response, we generated artificial antigen-presenting cells (aAPCs) by incubating human immature DCs (imDCs) with poly(lactic-co-glycolic) acid nanoparticles (PLGA-NPs) encapsulating tumour antigenic peptides, followed by maturation with lipopolysaccharide. Tumour antigen-specific CTLs were then induced using either peptide-loaded mature DCs (mDCs) or aAPCs, and their activities were analysed using both ELISpot and cytotoxicity assays. We found that the aAPCs induced significantly stronger tumour antigen-specific CTL responses than the controls, which included both mDCs and aAPCs loaded with empty nanoparticles. Moreover, frozen CTLs that were generated by exposure to aAPCs retained the capability to eradicate HLA-A2-positive tumour antigen-bearing cancer cells. These results indicated that aAPCs are superior to DCs when inducing the CTL response because the former are capable of continuously presenting tumour antigens to T cells in a sustained manner. The development of aAPCs with PLGA-NPs encapsulating tumour antigenic peptides is a promising approach for the generation of effective CTL responses in vitro and warrants further assessments in clinical trials.

Abstract 4606: Preclinical characterization of a novel fully human IgG1 anti-PD-L1 mAb CK-301

Abstract Antibodies targeting Programmed Death-1 (PD-1), or its ligand, PD-L1, have demonstrated remarkable efficacy in subsets of cancer patients, with inhibition of the interaction between PD-1 on T-cells and PD-L1 on tumor cells leading to the recovery of anti-tumor immune response and immune-mediated eradication of tumors. However, not all patients respond to existing PD-1 and PD-L1 targeting agents and relapses to therapy still occur. Therefore, there exists a need to identify additional therapeutics and approaches to engage the immune system to enhance the efficacy of current anticancer therapies. Using phage and yeast display approaches, we have discovered and optimized a novel, fully human, PD-L1 specific IgG1 antibody, CK-301, which exhibits subnanomolar binding affinity for PD-L1. CK-301 blocks binding of PD-L1 to both PD-1 and B7-1 in enzyme-linked immunosorbent assays (ELISA) and cell-based competition assays. Using an assay measuring inhibition of a nuclear factor of activated T-cells (NFAT) reporter caused by PD-L1 binding to PD-1, we demonstrate that CK-301 completely reverses reporter inhibition at concentration of less than 1 µg/ml, IC50 of the dose response curve is 80ng/ml. CK-301 enhances IFN-gamma secretion in allogeneic mixed lymphocyte reaction (MLR) using primary human T-cells and immature dendritic cells. CK-301 can also trigger antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) mediated killing of PD-L1+ cell lines, including lymphoma cells. CK-301 has similar subnanomolar affinity for cynomolgus monkey PD-L1 as for human PD-L1, hence we chose Macaca fascicularis for pre-clinical toxicology and safety pharmacology studies. Single-dose administration of CK-301 to monkeys up to the highest tested dose of 100 mg/kg was shown to be safe and demonstrated linear dose-dependent pharmacokinetic (PK) properties over the dose range from 1 to 100 mg/kg with a half-life of 15 days at 100 mg/kg. A first-in-human Phase 1 study of CK-301 is planned to commence in mid-2017. Citation Format: Leonid Gorelik, George Avgerinos, Yune Kunes, Wayne A. Marasco. Preclinical characterization of a novel fully human IgG1 anti-PD-L1 mAb CK-301 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4606. doi:10.1158/1538-7445.AM2017-4606

Immature human DCs efficiently translocate endocytosed antigens into the cytosol for proteasomal processing

Cross-presentation of endocytosed antigen is essential for induction of CD8 effector T cell responses and a hallmark of dendritic cells (DCs). The mode of antigen processing in this context is controversial and some models imply translocation of the antigen from the endosomes into the cytosol. To test this hypothesis we made use of the pro-apoptotic properties of cytochrome c when in the cytosol, and confirmed that it indeed triggered apoptosis of human immature DCs but only at high concentrations. Proteasome inhibitors reduced the required concentration of cytochrome c thousand-fold, indicating that protein translocated into the cytosol is rapidly degraded by proteasomes. Mature DCs were also susceptible to cytochrome c-triggered apoptosis at high concentrations but proteasome inhibitors did not increase their sensitivity. Other cross-presenting cells such as B cells and monocytes were not sensitive to cytochrome c at all, indicating that they do not shuttle internalized antigen into the cytosol. Thus, processing of internalized antigens seems to follow different pathways depending on cell type and, in case of DCs, maturation state. Immature DCs appear to have a unique capacity to shuttle external antigen into the cytosol for proteasomal processing, which could explain their efficiency in antigen cross-presentation.

CD40L-Dependent Pathway Is Active at Various Stages of Rheumatoid Arthritis Disease Progression.

The inflammatory CD40-CD40L pathway is implicated in various autoimmune diseases, but the activity status of this pathway in various stages of rheumatoid arthritis (RA) progression is unknown. In this study, we used gene signatures of CD40L stimulation derived from human immature dendritic cells and naive B cells to assess the expression of CD40-downstream genes in synovial tissues from anti-citrullinated protein Ab-positive arthralgia, undifferentiated arthritis (UA), early RA, and established RA cohorts in comparison with healthy donors. Interestingly, the expression of CD40LG and active full-length CD40 was increased in the disease tissues, whereas that of a dominant-negative CD40 isoform was decreased. Gene set variation analysis revealed that CD40L-responsive genes in immature dendritic cells and naive B cells were significantly enriched in synovial tissues from UA, early RA, and established RA patients. Additionally, CD40L-induced naive B cell genes were also significantly enriched in synovial tissues from arthralgia patients. In our efforts to characterize downstream mediators of CD40L signaling, we have identified GPR120 and KDM6B as novel components of the pathway. In conclusion, our data suggest that therapeutic CD40-CD40L blocking agents may prove efficacious not only in early and established RA, but also in inhibiting the progression of the disease from arthralgia or UA to RA.

Cytotoxicity and Intracellular Uptake of Fluorescent BSA-AuNCs from Human Monocyte-Derived Immature Dendritic Cells

Cytotoxicity and Intracellular Uptake of Fluorescent BSA-AuNCs from Human Monocyte-Derived Immature Dendritic Cells

Beyond attachment: Roles of DC-SIGN in dengue virus infection.

Dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN), a C-type lectin expressed on the plasma membrane by human immature dendritic cells, is a receptor for numerous viruses including Ebola, SARS and dengue. A controversial question has been whether DC-SIGN functions as a complete receptor for both binding and internalization of dengue virus (DENV) or whether it is solely a cell surface attachment factor, requiring either hand-off to another receptor or a co-receptor for internalization. To examine this question, we used 4 cell types: human immature dendritic cells and NIH3T3 cells expressing either wild-type DC-SIGN or 2 internalization-deficient DC-SIGN mutants, in which either the 3 cytoplasmic internalization motifs are silenced by alanine substitutions or the cytoplasmic region is truncated. Using confocal and super-resolution imaging and high content single particle tracking, we investigated DENV binding, DC-SIGN surface transport, endocytosis, as well as cell infectivity. DC-SIGN was found colocalized with DENV inside cells suggesting hand-off at the plasma membrane to another receptor did not occur. Moreover, all 3 DC-SIGN molecules on NIH3T3 cells supported cell infection. These results imply the involvement of a co-receptor because cells expressing the internalization-deficient mutants could still be infected.

Microvesicles released from Giardia intestinalis disturb host-pathogen response in vitro

Giardia intestinalis (G.I), is an anaerobic protozoan and the aetiological agent of giardiasis, a diarrhoea present worldwide and associated with poverty. G.I has a simple life cycle alternating between cyst and trophozoite. Cysts are transmitted orally to the stomach and transform to trophozoites in the intestine by a multifactorial process. Recently, microvesicles (MVs) have been found to be released from a wide range of eukaryotic cells. We have observed a release of MVs during the life cycle of G.I., identifying MVs from active trophozoites and from trophozoites differentiating to the cyst form. The aim of the current work was to investigate the role of MVs from G.I in the pathogenesis of giardiasis. MVs from log phase were able to increase the attachment of G. intestinalis trophozoites to Caco-2 cells. Moreover, MVs from G. intestinalis could be captured by human immature dendritic cells, resulting in increased activation and allostimulation of human dendritic cells. Lipid rafts participate in the MV biogenesis and in the attachment to Caco-2 cells. Nevertheless, proteomic analysis from two types of MVs has shown slight differences at the protein levels. An understanding of biogenesis and content of MVs derived from trophozoites might have important implications in the pathogenesis of the disease.

HIV Interferes with Mycobacterium tuberculosis Antigen Presentation in Human Dendritic Cells

HIV coinfection is the most prominent risk factor for progression of Mycobacterium tuberculosis (Mtb) infection into active tuberculosis (TB) disease. The mechanisms behind the increased transition from latent to active TB in coinfected individuals have not been well elucidated at the cellular level. We hypothesized that HIV infection contributes to Mtb pathogenesis by interfering with the dendritic cell (DC)-mediated immune control. Mtb-antigen processing and presentation are key events in the immune response against TB. Human immature DCs coinfected with HIV/Mtb had decreased expression of human leukocyte antigen antigen D related and the costimulatory molecules CD40, CD80, and CD86. In addition, Mtb-infected DCs triggered a significant release of the proinflammatory cytokines IL-6, IL-1β, and tumor necrosis factor-α, whereas coinfected DCs did not. To assess the DC antigen presentation capacity, we measured interferon-γ from co-cultures of DCs and autologous Mtb antigen-specific CD4+ T cells. Interferon-γ release was significantly reduced when purified protein derivative- and Ag85B-specific CD4+ T cells had been activated with coinfected DCs compared to Mtb-infected DCs, and this effect was attributed to Mtb antigen processing rather than peptide-major histocompatibility complex class II loading. Evaluating autophagy as a measure of vesicular processing and maturation further revealed that HIV efficiently blocks initiation of this pathway during coinfection. Overall, our results demonstrate that HIV impairs Mtb antigen presentation in DCs, thereby suppressing an important cell linking innate and adaptive immune response in TB.

Cap analogs modified with 1,2-dithiodiphosphate moiety protect mRNA from decapping and enhance its translational potential.

Along with a growing interest in mRNA-based gene therapies, efforts are increasingly focused on reaching the full translational potential of mRNA, as a major obstacle for in vivo applications is sufficient expression of exogenously delivered mRNA. One method to overcome this limitation is chemically modifying the 7-methylguanosine cap at the 5′ end of mRNA (m7Gppp-RNA). We report a novel class of cap analogs designed as reagents for mRNA modification. The analogs carry a 1,2-dithiodiphosphate moiety at various positions along a tri- or tetraphosphate bridge, and thus are termed 2S analogs. These 2S analogs have high affinities for translation initiation factor 4E, and some exhibit remarkable resistance against the SpDcp1/2 decapping complex when introduced into RNA. mRNAs capped with 2S analogs combining these two features exhibit high translation efficiency in cultured human immature dendritic cells. These properties demonstrate that 2S analogs are potentially beneficial for mRNA-based therapies such as anti-cancer immunization.

New roles for CD14 and IL-β linking inflammatory dendritic cells to IL-17 production in memory CD4+ T cells.

Interleukin (IL)-1β has proven to be crucial in the differentiation of human and mouse Th17 cells. Although it has become evident that IL-1β has potent IL-17-inducing effects on CD4+ T cells directly, it has not yet been explored whether IL-1β can also prime dendritic cells (DCs) for a Th17 instruction program. Here, we show that human immature DCs exposed to IL-1β promote IL-17 production in human memory CD4+ T cells. IL-1β-primed DCs express high levels of CD14 that mediate IL-17 production through direct interaction with T cells. Moreover, culturing human CD4+CD45RO+ memory T cells with soluble CD14 is sufficient for the upregulation of retinoic acid-related orphan receptor-γ thymus and IL-17 production. In addition, in a human in situ model using tissue-resident skin DCs, upregulation of CD14 expression induced by IL-1β on skin residents DCs promotes IL-17 production in memory T cells; strongly suggesting the in vivo relevance of this mechanism. Our findings uncover new roles for IL-1β and CD14, and may therefore have important consequences for the development of new therapies for Th17-mediated autoimmune diseases and bacterial and fungal pathogenic infections.

Distinct expression of interferon-induced protein with tetratricopeptide repeats (IFIT) 1/2/3 and other antiviral genes between subsets of dendritic cells induced by dengue virus 2 infection.

Dengue virus (DENV) infection is an emerging public health hazard threatening inhabitants of the tropics and sub-tropics. Dendritic cells (DCs) are one of the major targets of DENV and the initiators of the innate immune response against the virus. However, current invitro research on the DENV-DC interaction is hampered by the low availability of exvivo DCs and donor variation. In the current study, we attempted to develop a novel invitro DC model using immature DCs derived from the myeloid leukaemia cell line MUTZ-3 (IMDCs) to investigate the DENV-DC interaction. The IMDCs morphologically and phenotypically resembled human immature monocyte-derived dendritic cells (IMMoDCs). However, the permissiveness of IMDCs to DENV2 was lower than that of IMMoDCs. RT-PCR arrays showed that a group of type I interferon (IFN) -inducible genes, especially IFIT1, IFITM1, and IFI27, were significantly up-regulated in IMMoDCs but not in IMDCs after DENV2 infection. Further investigation revealed that IFIT genes were spontaneously expressed at both transcriptional and protein levels in the naive IMDCs but not in the naive IMMoDCs. It is possible that the poor permissiveness of IMDCs to DENV2 was a result of the high basal levels of IFIT proteins. We conclude that the IMDC model, although less permissive to DENV2, is a useful platform for studying the suppression mechanism of DENV2 and we expand the knowledge of cellular factors that modulate DENV2 infection in the human body.

Class 3 semaphorins induce F-actin reorganization in human dendritic cells: Role in cell migration.

Class 3 semaphorins (Semas) are soluble proteins that are well recognized for their role in guiding axonal migration during neuronal development. In the immune system, Sema3A has been shown to influence murine dendritic cell (DC) migration by signaling through a neuropilin (NRP)-1/plexin-A1 coreceptor axis. Potential roles for class 3 Semas in human DCs have yet to be described. We tested the hypothesis that Sema3A, -3C, and -3F, each with a unique NRP-1 and/or NRP-2 binding specificity, influence human DC migration. In this report, we find that although NRP-1 and NRP-2 are expressed in human immature DCs (imDCs), NRP-2 expression increases as cells mature further, whereas expression of NRP-1 declines dramatically. Elevated levels of RNA encoding plexin-A1 and -A3 are present in both imDCs and mature DC (mDCs), supporting the relevance of Sema/NRP/plexin signaling pathways in these cells. Sema3A, -3C, and -3F bind to human DCs, with Sema3F binding predominantly through NRP-2. The binding of these Semas leads to reorganization of actin filaments at the plasma membrane and increased transwell migration in the absence or presence of chemokine CCL19. Microfluidic chamber assays failed to demonstrate consistent changes in speed of Sema3C-treated DCs, suggesting increased cell deformability as a possible explanation for enhanced transwell migration. Although monocytes express RNA encoding Sema3A, -3C, and -3F, only RNA encoding Sema3C increases robustly during DC differentiation. These data suggest that Sema3A, -3C, and -3F, likely with coreceptors NRP-1, NRP-2, and plexin-A1 and/or -A3, promote migration and possibly other activities of human DCs during innate and adaptive immune responses.

Exogenous Thyropin from p41 Invariant Chain Diminishes Cysteine Protease Activity and Affects IL-12 Secretion during Maturation of Human Dendritic Cells.

Dendritic cells (DC) play a pivotal role as antigen presenting cells (APC) and their maturation is crucial for effectively eliciting an antigen-specific immune response. The p41 splice variant of MHC class II-associated chaperone, called invariant chain p41 Ii, contains an amino acid sequence, the p41 fragment, which is a thyropin-type inhibitor of proteolytic enzymes. The effects of exogenous p41 fragment and related thyropin inhibitors acting on human immune cells have not been reported yet. In this study we demonstrate that exogenous p41 fragment can enter the endocytic pathway of targeted human immature DC. Internalized p41 fragment has contributed to the total amount of the immunogold labelled p41 Ii-specific epitope, as quantified by transmission electron microscopy, in particular in late endocytic compartments with multivesicular morphology where antigen processing and binding to MHC II take place. In cell lysates of treated immature DC, diminished enzymatic activity of cysteine proteases has been confirmed. Internalized exogenous p41 fragment did not affect the perinuclear clustering of acidic cathepsin S-positive vesicles typical of mature DC. p41 fragment is shown to interfere with the nuclear translocation of NF-κB p65 subunit in LPS-stimulated DC. p41 fragment is also shown to reduce the secretion of interleukin-12 (IL-12/p70) during the subsequent maturation of treated DC. The inhibition of proteolytic activity of lysosomal cysteine proteases in immature DC and the diminished capability of DC to produce IL-12 upon their subsequent maturation support the immunomodulatory potential of the examined thyropin from p41 Ii.

Immature human dendritic cells enhance their migration through KCa3.1 channel activation

Migration capacity is essential for dendritic cells (DCs) to present antigen to T cells for the induction of immune response. The DC migration is supposed to be a calcium-dependent process, while not fully understood. Here, we report a role of the KCa3.1/IK1/SK4 channels in the migration capacity of both immature (iDC) and mature (mDC) human CD14+-derived DCs. KCa3.1 channels were shown to control the membrane potential of human DC and the Ca2+ entry, which is directly related to migration capacities. The expression of migration marker such as CCR5 and CCR7 was modified in both types of DCs by TRAM-34 (100nM). But, only the migration of iDC was decreased by use of both TRAM-34 and KCa3.1 siRNA. Confocal analyses showed a close localization of CCR5 with KCa3.1 in the steady state of iDC. Finally, the implication of KCa3.1 seems to be limited to the migration capacities as T cell activation of DCs appeared unchanged. Altogether, these results demonstrated that KCa3.1 channels have a pro-migratory effect on iDC migration. Our findings suggest that KCa3.1 in human iDC play a major role in their migration and constitute an attractive target for the cell therapy optimization.

DC-SIGN Mediated Dengue Virus Entry into Cells

DC-SIGN (a single-pass, C-type lectin, transmembrane protein) is a receptor for dengue virus (DENV) and variety of other pathogens. It exists in small clusters on the plasma membranes of dendritic and other immune cells. Using NIH3T3 cells that express full length DC-SIGN or a mutant form lacking the cytoplasmic tail as well as primary human immature dendritic cells, we investigated DENV binding, surface transport, endocytosis and infection. We employed confocal microscopy, super-resolution imaging and large-scale single particle tracking. After binding, the virus/receptor complexes co-migrate to clathrin coated pits where the complexes are endocytosed followed by release of the viral genome for replication. Suprisingly, dengue binding, even to a few DC-SIGN clusters, induces a global cellular response in which both loaded and unloaded DC-SIGN clusters exhibit dramatically increased lateral mobility, possibly to enhance encounters with coated pits. In addition, a small but significant fraction of DC-SIGN clusters in the plasma membrane undergo rapid, microtubule-based directed transport towards the cell center at velocities which can exceed 1μm/s. Since the measured velocities are increased after dengue binding, this activity may be required to bring captured pathogens from the leading margins back to perinuclear zone for processing by dendritic cells. A controversy exists as to whether DC-SIGN functions simply as an attachment factor for dengue viruses or whether it plays additional roles in viral entry. The absence of the DC-SIGN cytoplasmic region reduced both dengue binding and endocytosis. However, infection still occurred, albeit reduced, suggesting that DC-SIGN must be able to act in concert with a co-receptor containing cell entry motifs. Supported by NIH GM41402.

Actin Dynamics Regulates Dendritic Cell-Mediated Transfer of HIV-1 to T Cells

Whereas human dendritic cells (DCs) are largely resistant to productive infection with HIV-1, they have a unique ability to take up the virus and transmit it efficiently to T lymphocytes through a process of trans-infection or trans-enhancement. To elucidate the molecular and cell biological mechanism for trans-enhancement, we performed an shRNA screen of several hundred genes involved in organelle and membrane trafficking in immature human monocyte-derived dendritic cells (MDDCs). We identified TSPAN7 and DNM2, which control actin nucleation and stabilization, as having important and distinct roles in limiting HIV-1 endocytosis and in maintaining virus particles on dendrites, which is required for efficient transfer to T lymphocytes. Further characterization of this process may provide insights not only into the role of DCs in transmission and dissemination of HIV-1 but also more broadly into mechanisms controlling capture and internalization of pathogens.

Generation of regulatory dendritic cells after treatment with paeoniflorin.

Regulatory dendritic cells are a potential therapeutic tool for assessing a variety of immune overreaction diseases. Paeoniflorin, a bioactive glucoside extracted from the Chinese herb white paeony root, has been shown to be effective at inhibiting the maturation and immunostimulatory function of murine bone marrow-derived dendritic cells. However, whether paeoniflorin can program conventional dendritic cells toward regulatory dendritic cells and the underlying mechanism remain unknown. Here, our study demonstrates that paeoniflorin can induce the production of regulatory dendritic cells from human peripheral blood monocyte-derived immature dendritic cells in the absence or presence of lipopolysaccharide (LPS) but not from mature dendritic cells, thereby demonstrating the potential of paeoniflorin as a specific immunosuppressive drug with fewer complications and side effects. These regulatory dendritic cells treated with paeoniflorin exhibited high CD11b/c and low CD80, CD86 and CD40 expression levels as well as enhanced abilities to capture antigen and promote the proliferation of CD4(+)CD25(+) T cells and reduced abilities to migrate and promote the proliferation of CD4(+) T cells, which is associated with the upregulation of endogenous transforming growth factor (TGF)-β-mediated indoleamine 2,3-dioxygenase (IDO) expression. Collectively, paeoniflorin could program immature dendritic cells (imDCs) and imDCs stimulated with LPS toward a regulatory DC fate by upregulating the endogenous TGF-β-mediated IDO expression level, thereby demonstrating its potential as a specific immunosuppressive drug.