Preferential production of IL-12 by peritoneal macrophages activated by liposomes prepared from neoglycolipids containing oligomannose residues
Introduction
Antigen-presenting cells (APCs)1, especially macrophages and dendritic cells (DCs), play a central role in the innate immune response to a wide range of bacterial components [1], [2]. On activation by these components, APCs produce pro-inflammatory cytokines such as tumor necrosis factor (TNF)-α, interleukin (IL)-12, IL-6 and IL-1, express high levels of MHC class II and co-stimulatory molecules such as CD80 and CD86 on their cell surface, and mature to produce IL-12, which greatly potentiates IFN-γ-producing Th1-type T cell differentiation [3], [4]. Thus, the maturation and cytokine production profiles of APCs are critical in the link between innate and adaptive immunity.
APCs interact with pathogens using pattern recognition receptors (PRRs), which can recognize highly conserved molecular signatures: so-called pathogen-associated molecular patterns (PAMPs) [5]. Toll-like receptors (TLRs) are a major pattern recognition receptor family. TLRs respond to PAMPs, and the subsequent signaling events can lead to detection of microbial infections, activation of innate immune responses, subsequent activation and modulation of adaptive immunity, and consequent triggering of the antibacterial host defense response [1], [2]. In addition to TLRs, carbohydrate-binding receptors are another part of the larger PRR family and are expressed on a wide variety of cell types, including APCs. Carbohydrate-binding receptors participate in recognition of carbohydrate structures associated with a wide variety of microbes and microbial-derived products [6], [7], [8]. The macrophage mannose receptor (CD206) expressed on macrophages and DCs recognizes mannose, fucose and N-acetylglucosamine residues of glycoconjugates, and is a well-characterized endocytic receptor in uptake of bacteria, yeast, and other pathogenic microorganisms [9]. The β-glucan receptor, dectin-1, mediates phagocytosis of nonopsonic yeast by macrophages and acts in synergy with TLR-2 to augment pro-inflammatory cytokine responses [10], [11]. DC-specific ICAM-3 grabbing nonintegrin (DC-SIGN), a new dendritic cell-specific membrane lectin, binds to high-mannose oligosaccharides and is involved in uptake of mannose-exposed particles and microbes by DCs [12], [13], [14], [15], [16], and DC-SIGN-related proteins, SIGNR1 and SIGNR3, expressed on macrophages also recognize polysaccharides and microbes [17]. Thus, carbohydrate-binding receptors may play important roles in recognition of PAMPs in innate immunity.
Recently, we have reported that liposomes coated with synthesized neoglycolipids constructed from mannotriose and dipalmitoylphosphatidylethanolamine (Man3-DPPE) are preferentially and rapidly taken up by peritoneal macrophages, and that liposome-incorporating macrophages accumulate in extranodal lymphoid tissues following injection of Man3-DPPE-coated liposomes (hereafter referred to as oligomannose-coated liposomes, OMLs) into the peritoneal cavity [18], [19]. It has also been shown that intraperitoneal immunization of OMLs entrapped with soluble leishmanial antigen induces an antigen-specific Th1 immune response and protects against subsequent infection with Leishmania major in BALB/c mice [20], [21]. These findings indicate that specific incorporation of OMLs into peritoneal macrophages triggers the induction of Th1 immune response, but the underlying mechanism is unclear. Up-regulation of co-stimulatory molecules and MHC class II on APCs is crucial in determining the nature and extent of the immune response [22], [23], [24], and IL-12 secretion from APCs is critical for development of Th1 cells and initiation of a cell-mediated immune response [3]. Thus, the present study was conducted to investigate the activation of peritoneal macrophages by OMLs and the subsequent profile of pro-inflammatory cytokines. Our results indicate that OMLs exhibit a novel adjuvant activity that results in increased expression of MHC class II and co-stimulatory molecules and leads to preferential induction of IL-12 production by peritoneal macrophages.
Section snippets
Antibodies and reagents
Fluoroscein isothiocyanate (FITC)-, phycoerythrin (PE)-, or peridinin chlorophyll protein-cyanin 5.5 (PerCP-Cy5.5)-labeled antibodies directed against mouse CD3ε, CD11b, CD19, CD40, Gr.-1 and I-A/I-E class II molecules and an Fc-block (anti-mouse CD16/32) were purchased from BD PharMingen (San Diego, CA, USA). PE-labeled anti-F4/80 antibody was purchased from Caltag Laboratories (Burlingame, CA, USA). FITC-conjugated antibodies directed against mouse CD11c, mouse CD80, and CD86 were purchased
Intraperitoneal immunization of OMLs induces a Th1 response against encased antigen
Liposomes coated with Man3-DPPE were used as oligomannose-coated liposomes (OMLs) in all experiments, unless otherwise indicated. We used OMLs consisting of DPPC:cholesterol:Man3-DPPE (10:10:1) and with a particle size of 1 μm, since it has been shown that OMLs of this composition and size are preferentially taken up by peritoneal macrophages (PEMs), with subsequent induction of a Th1 immune response specific for an antigen entrapped in the OML in mice [21]. To confirm preferential uptake of
Discussion
Our results show that production of IL-12 in response to OMLs (Man3-DPPE- or Man5-DPPE-coated liposomes) is a characteristic of peritoneal macrophages, with subsequent maturation to antigen-presenting cells with enhanced expression of co-stimulatory molecules and MHC class II molecules, and induction of an antigen-specific Th1 immune response. Thus, OMLs possess novel adjuvant properties for induction of cell-mediated immunity. These responses are characteristic to OMLs, since liposomes coated
Acknowledgments
This work was supported by the Program for Promotion of Basic Research Activities for Innovative Biosciences (PROBRAIN), and in part by a grant for Hi-Tech Research from Tokai University, and by the Industrial Technology Research Grant Program from the New Energy and Industrial Technology Development Organization (NEDO) of Japan.
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2016, Immunology LettersCitation Excerpt :These characteristic abilities suggest that CLRs expressed on APCs are potentially useful for antigen delivery for vaccination, as well as immune modulation. We have generated liposomes coated with a synthetic neoglycolipid constructed from mannotriose and dipalmitoylphosphatidylethanolamine (Man3-DPPE) for targeting mannose-binding CLRs and have shown that vaccination with these Man3-DPPE-coated liposomes (oligomannose-coated liposomes; OMLs) with encased antigens can elicit antigen-specific CLTs and Th1 responses, with preferential production of IL-12 from APCs, up-regulation of co-stimulatory molecules and MHC class II on APCs, and presentation of antigenic peptides of the encased antigens on both MHC class I and class II molecules [11–13]. Therefore, OMLs could act as a novel adjuvant and antigen delivery vehicle for induction of antigen specific cellular immune responses.
SIGNR1-mediated phagocytosis, but not SIGNR1-mediated endocytosis or cell adhesion, suppresses LPS-induced secretion of IL-6 from murine macrophages
2015, CytokineCitation Excerpt :We also showed that SIGNR1 directly mediates the uptake of several NGL-coated liposomes, including OMLs, using non-phagocytic CHO cells transfected with SIGNR1 and murine macrophage-like RAW264.7 cells that stably expressed SIGNR (RAW-SIGNR1 cells) [13,14]. Since spontaneous secretion of IL-6 from peritoneal phagocytic cells was suppressed by OML uptake and certain levels of SIGNR1 are expressed murine peritoneal phagocytic cells [10,12], SIGNR1-mediated recognition of ligands may affect the production of IL-6 from phagocytic cells, as is the case for human DC-SIGN. In addition to mediating uptake of carbohydrate-decorated particles, SIGNR1 also mediates internalization of carbohydrate-bearing soluble polymers, such as ovalbumin and dextran, and cell adhesion to carbohydrate-coated solid phases [11,14].
Comparison of the carbohydrate preference of SIGNR1 as a phagocytic receptor with the preference as an adhesion molecule
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