Targeting dendritic cells with biomaterials: developing the next generation of vaccines

Current vaccine and immunotherapy technology faces ongoing challenges in both efficacy and practicality: many chronic diseases cannot yet be addressed by vaccination, and several vaccines that do function well require multiple injections, which is a substantial limitation in various parts of the world. A possible key to developing the next generation of vaccines is the ability to deliver antigen to dendritic cells (DCs) more specifically and induce the subsequent activation of T-cell immunity. However, antigen delivery to, and activation of, DCs is a complex problem, involving antigen transport to DC-rich areas, DC binding and antigen uptake, and antigen processing and presentation. Addressing these challenges requires novel and multidisciplinary approaches, for example, the application of biomaterials to immunotechnology. Here, we review the latest advances in biomaterial drug vehicles, such as polymer microparticles and nanoparticles, and liposomes, that are being used to target DCs in new strategies for vaccination.     

Interaction of Neisseria meningitidis with human dendritic cells

Infection with Neisseria meningitidis serogroup B is responsible for fatal septicemia and meningococcal meningitis. The severity of disease directly correlates with the production of the proinflammatory cytokines tumor necrosis factor alpha (TNF-alpha), interleukin-1 (IL-1), IL-6, and IL-8. However, the source of these cytokines has not been clearly defined yet. Since bacterial infection involves the activation of dendritic cells (DCs), we analyzed the interaction of N. meningitidis with monocyte-derived DCs. Using N. meningitidis serogroup B wild-type and unencapsulated bacteria, we found that capsule expression significantly impaired neisserial adherence to DCs. In addition, phagocytic killing of the bacteria in the phagosome is reduced by at least 10- to 100-fold. However, all strains induced strong secretion of proinflammatory cytokines TNF-alpha, IL-6, and IL-8 by DCs (at least 1,000-fold at 20 h postinfection [p.i.]), with significantly increased cytokine levels being measurable by as early as 6 h p.i. Levels of IL-1beta, in contrast, were increased only 200- to 400-fold at 20 h p.i. with barely measurable induction at 6 h p.i. Moreover, comparable amounts of cytokines were induced by bacterium-free supernatants of Neisseria cultures containing neisserial lipooligosaccharide as the main factor. Our data suggest that activated DCs may be a significant source of high levels of proinflammatory cytokines in neisserial infection and thereby may contribute to the pathology of meningococcal disease.     

Interaction of large DNA viruses with dendritic cells

Dendritic cells (DC) with their unique capacity to prime naïve T cells are crucial in the induction of immunological responses, including anti-tumoral and anti-viral immunity. DC based immunotherapies are thus currently considered a particularly promising approach for cellular immunotherapy. The cloning of tumor associated antigens (TAAs) together with the possibility of manipulating viral genomes by biotechnological techniques has sparked the interest of using genetically modified viruses to transduce DC in order to achieve antigenic expression of TAA with the aim of inducing a protective immune response. An increasing number of modified viral vectors has been designed for gene therapy purposes and consecutively has been used for the ex vivo transduction of DC. It has been shown that viral vectors genetically engineered to express TAA or immune modifiers like cytokines or costimulatory molecules can lead to a high level of transgene expression. Furthermore, these studies have also revealed that viruses have developed several immune evasion mechanisms specifically targeting DC. Therefore, analysing the interactions of viruses with DC is crucial for the development of new viral vectors suitable for the transduction of DC. In this report we describe the interaction of two large DNA viruses, herpes simplex virus type1 (HSV-1) and vaccinia virus (VV), with DC generated from peripheral blood mononuclear cells.     

Interaction of Listeria monocytogenes with mouse dendritic cells.

In this study, the interaction of murine dendritic cells with Listeria monocytogenes was investigated. Dendritic cells are efficient antigen-presenting cells, play a key role in the immune response, and are capable of migrating over substantial distances between sites of infection and lymphoid tissues. L. monocytogenes EGD invaded dendritic cells, escaped from phagosomes into the cytoplasm, and there directed actin nucleation, polymerization, and polarization in a typical fashion, thereby achieving intracellular movement and cell-to-cell spread. The internalization process appears to be independent of the inl locus. Interestingly, an intact microtubular function was essential for efficient uptake, whereas in a previous report, microtubule disruption did not affect bacterial spread in Caco-2 cells. The results obtained also suggest that L. monocytogenes binds to glycosylated receptors of dendritic cells. Uptake of Listeria cells was mediated by a protein kinase-dependent transducing phosphorylation signal that induces the actin polymerization-dependent phagocytic process. To achieve efficient uptake, de novo protein synthesis of eukaryotic and prokaryotic cells is also required. Despite the killing of dendritic cells, wild-type bacteria were found to persist in small numbers in some cells for at least 24 h. When different isogenic mutants of the EGD strain were analyzed for their capability to interact with dendritic cells, it was observed that some virulence-attenuated mutants (i.e., prfA and delta hly) persisted in large numbers for even longer times. Invasion of dendritic cells by L. monocytogenes, which in turn could result in either cell death or persistent infection, might have an important role in the pathogenesis of listeriosis, leading to impaired immune responses with inefficient bacterial clearance and/or promoting bacterial spread.     

Interaction of Mycobacterium avium with human monocyte-derived dendritic cells

The mechanism by which mycobacteria elicit class I-restricted T-cell responses remains undefined because these organisms have been shown to reside exclusively within membrane-bound vesicles in macrophages (Mphi), their primary host cells. We studied the interaction of M. avium with dendritic cells (DC) because they are the most potent antigen-presenting cells and are abundant at M. avium infection sites. We observed that both DC and Mphi, generated from human peripheral blood monocytes by short-term culture, internalized M. avium. The onset of programmed cell death and the percentage of apoptotic cells in infected DC and Mphi were comparable. However, following infection, DC secreted significantly larger amounts of interleukin-12, but not interleukin-1beta, than infected autologous Mphi. Further analysis of infected cells showed that while phagosomes failed to acidify in both M. avium-infected DC and Mphi, bacilli grew more slowly in DC. Electron microscopy studies revealed that M. avium resided within endocytic vacuoles in both cell types. The vacuolar membrane surrounding some bacilli in approximately 10% of the vacuoles in DC possessed several breaks. The importance of this finding will have to be addressed in future studies.     

Interaction of human serum lipoproteins with biomaterials

Human serum was incubated with representative portions of polyvinyl chloride (PVC) blood storage bags and vascular prostheses. The in vitro interaction process with lipoprotein was followed by polyacrylamide gel electrophoresis (PAGE) using sudan black and nitroblue tetrazolium (NBT) in the prestaining procedure. Densitometric scan of all the lipoprotein bands in serum after incubation with PVC bag material when prestained with sudan black showed remarkable increase in intensity. However, in the same experiment when NBT was used for prestaining no increase in the intensity of any of the lipoprotein bands could be observed. Since sudan black is known to bind cholesterol specifically we suggest that a molecular unfolding occurs when lipoprotein interacts with PVC bag material. When similar experiments were carried out with vascular prosthesis there was conspicuous decrease in the intensity of the high-density lipoprotein (HDL) band especially when stained with NBT. This indicates preferential adhesion of HDL during interaction with vascular prosthesis.     

Interaction of porcine conventional dendritic cells with swine influenza virus

Swine influenza virus (SwIV) causes sub-acute or acute respiratory infections on swine farms and pigs can act as “mixing vessels” for new influenza strains. Knowledge of the immune response of SwIV in its natural host, pigs, is very limited. Dendritic cells (DCs) mediate the induction of immunity to pathogens, but their interaction with SwIV has not been fully characterized. Thus, porcine bone marrow derived DCs (poBMDCs) were exposed to a circulating strain of H3N2 SwIV in vitro. Infection of poBMDCs resulted in structures resembling influenza virus inside poBMDCs in vesicles and also free in cytoplasm. Viral progeny was undetectable in supernatant but limited replication was detected in the first 8 h after infection. However, viral particles from infected-poBMDCs were able to induce cytopathic effect in susceptible cells only when cell-to-cell interaction was favoured. The data generated in our studies reveal the particular interaction of H3N2 SwIV with conventional DCs.     

Interaction of Leishmania parasites with dendritic cells and its functional consequences

Interaction between dendritic cells (DC) and T cells is essential for the generation of cell mediated immunity and thus DC play a critical role in the initiation of immune responses against Leishmania parasites. Although macrophages (Mphi) are the major targets of all species of Leishmania, a number of studies demonstrated the infection of DC by Leishmania. DC specific intracellular adhesion molecule 3-grabbing nonintegrin (DC-SIGN), has been reported to be the receptor for Leishmania amastigotes. The functional consequences in DC after Leishmania infections appear to depend on species of Leishmania. Some species of Leishmania enhance the surface expression of co-stimulatory molecules and CD40-ligand-induced IL-12 production in DC. On the other hand other species down-regulate co-stimulatory molecules and inhibit IL-12 production. The intrinsic differences among Leishmania species with regard to alteration of cell surface molecules and IL-12 production in DC may contribute to the healing and non-healing forms of the disease.     

The role of integrins in the recognition and response of dendritic cells to biomaterials

Biomaterials have the potential to be utilized as immunostimulatory or immunosuppressive delivery agents for biologics. It is hypothesized that this is directed by the ability of a biomaterial to drive dendritic cells (DC) in situ toward an immunostimulatory or an immunosuppressive phenotype, respectively. However, the specific pattern recognition receptors (PRRs) that DCs use to recognize and respond to biomaterials are unknown. From among the many receptors that DCs use to recognize and respond to foreign entities, herein the focus is on integrins. A biomaterial that induces DC maturation, namely poly(lactic-co-glycolic) acid (PLGA), supported increased human monocyte-derived DC adhesion and up-regulation of integrin receptor gene expression, measured via RT-PCR, as compared to culture on tissue culture polystyrene (TCPS). This was not observed for a biomaterial that does not support DC maturation. Through antibody-blocking techniques, the adhesion to both TCPS and PLGA was found to be β(2) integrin dependent and β(1) independent. Significantly, inhibiting β(2)-mediated adhesion to biomaterials via blocking antibodies also lowered the level of maturation of DCs (CD86 expression). β(2) integrins (but not β(1)) were found localized in biomaterial-adherent DC podosomes and also were found in direct contact with the PLGA surface. Therefore, it appeared that β(2) integrin-mediated adhesion is involved in determining the state of DC maturation on the PLGA surface. DC adhesion to biomaterials may be engaged or avoided to manipulate an immune response to biological component delivered with a biomaterial carrier.     

Interaction of Brucella suis and Brucella abortus rough strains with human dendritic cells

Brucella is a facultative intracellular pathogen of various mammals and the etiological agent of brucellosis. We recently demonstrated that dendritic cells (DCs), which are critical components of adaptive immunity, are highly susceptible to Brucella infection. Furthermore, Brucella prevented the infected DCs from engaging in maturation processes and impaired their capacity to present antigen to naive T cells and to secrete interleukin-12 (IL-12). The lipopolysaccharide (LPS) phenotype is largely associated with the virulence of Brucella. Depending on whether they express the O-side chain of LPS or not, the bacteria display a smooth or rough phenotype. Rough Brucella mutants are attenuated and induce a potent protective T-cell-dependent immune response. Due to the essential role of DCs in the initiation of T-cell-dependent adaptive immune responses, it seemed pertinent to study the interaction between rough Brucella strains and human DCs. In the present paper, we report that, in contrast to smooth bacteria, infection of DCs with rough mutants of Brucella suis or Brucella abortus leads to both phenotypic and functional maturation of infected cells. Rough mutant-infected DCs then acquire the capacity to produce IL-12 and to stimulate naive CD4⁺ T lymphocytes. Experiments with rough and smooth purified LPS of Brucella supported the hypothesis of an indirect involvement of the O-side chain. These results provide new data concerning the role of LPS in Brucella virulence strategy and illuminate phenomena contributing to immune protection conferred by rough vaccine strains.     

Interaction between Borrelia burgdorferi and immature human dendritic cells

Antigen uptake and the following maturation of dendritic cells (DCs) are pivotal to the initiation of specific antimicrobial immune responses. DCs also play an important role in the recruitment and activation of the cells of the innate immune system. We have examined the interactions of DCs with Borrelia burgdorferi to find explanations for the difficulties the human immune system has in dealing with the bacterium. Phagocytosis of B. burgdorferi by immature DCs and the effect of the bacterium on the maturation and interleukin-8 (IL-8) secretion of DCs were studied. Borreliae were phagocytized and processed into fragments by DCs; narrow tube-like pseudopods and broad pseudopods were used for the engulfment. The immature DC population gained a heterogeneous appearance within 2 h of incubation with the borreliae. A 24 h coculture with borreliae induced maturation and IL-8 secretion in the DCs in a manner comparable with the effect of lipopolysaccharides. All strains studied, including a mutant strain lacking outer surface proteins A and B, were capable of inducing these responses. Thus, our results did not show any clear inadequacy concerning the way DCs are dealing with B. burgdorferi. However, further studies on the subject are required.     

Compatibility of embryonic stem cells with biomaterials

Periodontal bone defects and atrophy of the jaws in an aging population are of special concern. Tissue engineering using embryonic stem cells (ESCs) and biomaterials may offer new therapeutic options. The purpose of this study is to evaluate the compatibility of ESCs with biomaterials and the influence of biomaterials on the osteogenic gene expression profile.Therefore, ESCs are cultured with various biomaterials. The cytocompatibility of murine ESCs is measured regarding the proliferation of the cells on the materials by CyQUANT assay, the morphology by scanning electron microscopy, and the influence on the gene expression by real time PCR.The results show that insoluble collagenous bone matrix, followed by beta-tricalciumphosphate, is most suitable for bone tissue engineering regarding cell proliferation, and phenotype. The gene expression analysis indicates that biomaterials do influence the gene expression of ESCs.Our results provide new insight into the cytocompatibility of ESCs on different scaffolds.     

Interaction between dendritic cells and natural killer cells during pregnancy in mice

A complex regulation of innate and adaptive immune responses at the maternal fetal interface promotes tolerance of trophoblast cells carrying paternally derived antigens. Such regulatory functions involve uterine dendritic cells (uDC) and natural killer (uNK) cells. The existence of a NK and DC "cross talk" has been revealed in various experimental settings; its biological significance ranging from cooperative stimulation to cell lysis. Little is known about the presence or role of NK and DC cross talk at the maternal fetal interface. The present study shows that mouse NK and DC interactions are subject to modulation by trophoblast cells in vitro. This interaction promotes a tolerogenic microenvironment characterized by downregulation of the expression of activation markers on uNK cells and uDC and dominance of Th2 cytokines. NK and DC interactions would also influence uterine cell proliferation and this process would be strongly modulated by trophoblast-derived signals. Indeed; while low proliferation rates were observed upon regular coculture allowing direct contact between uterine cells and trophoblasts, incubation in a transwell culture system markedly increased uterine cell proliferation suggesting that soluble factors are key mediators in the molecular "dialog" between the mother and the conceptus during the establishment of mouse pregnancy. Our data further reveal that the regulatory functions of trophoblast cells associated with tolerance induction are impaired in high abortion murine matings. Interestingly, we observed that secretion of interleukin-12p70 by uDC is dramatically abrogated in the presence of uNK cells. Taken together, our results provide the first evidence that a delicate balance of interactions involving NK cells, DC, and trophoblasts at the mouse maternal fetal interface supports a successful pregnancy outcome.     

Differential functional effects of biomaterials on dendritic cell maturation

The immunological outcome of dendritic cell (DC) treatment with different biomaterials was assessed to demonstrate the range of DC phenotypes induced by biomaterials commonly used in combination products. Immature DCs (iDCs) were derived from human peripheral blood monocytes, and treated with different biomaterial films of alginate, agarose, chitosan, hyaluronic acid (HA), or 75:25 poly(lactic-co-glycolic acid) (PLGA) and a comprehensive battery of phenotypic functional outcomes was assessed. Different levels of functional changes in DC phenotype were observed depending on the type of biomaterial films used to treat the DCs. Treatment of DCs with PLGA or chitosan films supported DC maturation, with higher levels of DC allostimulatory capacity, pro-inflammatory cytokine release, and expression of CD80, CD86, CD83, HLA-DQ and CD44 compared with iDCs, and lower endocytic ability compared with iDCs. Alginate film induced pro-inflammatory cytokine release from DCs at levels higher than from iDCs. Dendritic cells treated with HA film expressed lower levels of CD40, CD80, CD86 and HLA-DR compared with iDCs. They also exhibited lower endocytic ability and CD44 expression than iDCs, possibly due to an insolubilized (cross-linked) form of high molecular weight HA. Interestingly, treatment of DCs with agarose film maintained the DC functional phenotype at levels similar to iDCs except for CD44 expression, which was lower than that of iDCs. Taken together, these results can provide selection criteria for biomaterials to be used in immunomodulating applications and can inform potential outcomes of biomaterials within combination products on associated immune responses as desired by the application.     

Macrophage and dendritic cell phenotypic diversity in the context of biomaterials

Macrophages (M?) and dendritic cells (DCs) are critical antigen presenting cells that play pivotal roles in host responses to biomaterial implants. Although M?s have been widely studied for their roles in the inflammatory responses against biomaterials, the roles that DCs play in the host responses toward implanted materials have only recently been explored. DCs are of significant research interest because of the emergence of a large number of combination products that cross-traditional medical device boundaries. These products combine biomaterials with biologics, including cells, nucleic acids, and/or proteins. The biomaterial component may evoke an inflammatory response, primarily mediated by neutrophils and M?s, whereas the biologic component may elicit an immunogenic immune response, initiated by DCs involving lymphocyte activation. Control of M? phenotypic balance from proinflammatory M1 to reparative M2 is a goal of investigators to optimize the host response to biomaterials. Similarly, control of DC phenotype from proinflammatory to toleragenic is of interest in vaccine delivery and tissue engineering/transplantation situations, respectively. This review discusses the interconnection between innate and adaptive immunity, the comparative and contrasting phenotypes and roles of M?s and DCs in immunity, their responses to biomaterials and the strategies to modulate their phenotype for applications in tissue engineering and vaccine delivery. Furthermore, the collaboration between and unique roles of DCs and M?s needs to be addressed in future studies to gain a more complete picture of host responses toward combination products.     

Artificial cells as bioreactive biomaterials

Artificial cells can have the same dimensions as biological cells. They can enclose and retain a large variety of bioreactants. Artificial cell membranes can have the required permeability characteristics to allow the rapid equilibration of external molecules to be acted on by the enclosed bioreactants. Polymer, protein, protein-lipid, polymer-lipid, lipid or other materials can form artificial cell membranes. It is possible to vary the membrane thickness and permeability characteristics over a wide range. Many different forms of bioreactive artificial cells are available. This article contains only a few examples. This includes its applications in (1) red blood cell substitutes; (2) immobilisation of enzymes, multienzymes with cofactor recycling, cell cultures and other biotechnological applications; (3) hemoperfusion in kidney failure, poisoning, removal of aluminum and iron; (4) use in liver failure; (5) other applications in microencapsulation.     

Interactions of acinar cells on biomaterials with various surface properties

The purpose of this study is to evaluate the interactions of rat parotid acinar cells on biomaterials with different surface properties. The biomaterials used in this study included polyvinyl alcohol (PVA), chitosan, poly (ethylene-co-vinyl alcohol) (EVAL), and polyvinylidene fluoride (PVDF). Cell morphology was observed by photomicroscope. Cell growth and differentiated characteristic function were separately assayed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) reduction activity and amylase activity. Results indicated that behaviors of acinar cells on materials might differ to a great extent depending on the surface hydrophilicity and morphology of the materials. On the relatively hydrophobic materials, the abilities of acinar cells to adhere and proliferate increased simultaneously. In addition, porous PVDF had higher cell growth compared with dense PVDF. Therefore, the hydrophobic PVDF with a porous structure was the best substrate for culturing acinar cells. According to our findings, a tubular PVDF scaffold with dense outer surface to prevent saliva leakage and with porous inner surface for the cell growth was proposed to serve as an artificial salivary gland for future use in the treatment of patients with salivary hypofunction.     

In vitro and in vivo interactions of cells with biomaterials

The biocompatibility of materials at an implant site involves a complex interaction of cells and tissues with the biomaterial. This cell-cell and cell-polymer interaction evokes the release of mediators such as chemotactic and growth factors that elicit and sustain inflammatory responses at the implant site. In this review, we summarize the interaction of cells with biomaterials in vitro and in vivo.