Immunotherapy against angiotensin II receptor ameliorated insulin resistance in a leptin receptor-dependent manner.

The angiotensin II type 1 receptor (AT1R) signaling pathway is reported to modulate glucose metabolism. Targeting AT1R, our group invented ATRQß-001 vaccine, a novel immunotherapeutic strategy to block the activation of AT1R. Here, we evaluated the therapeutic efficacy of ATRQß-001 vaccine in insulin resistance, and investigated the mechanism. Our results showed that ATRQß-001 vaccine and specific monoclonal antibody against epitope ATR-001 (McAb-ATR) decreased fasting serum insulin concentration and improved glucose and insulin tolerance in ob/ob mice. These beneficial effects were verified in high-fat diet-induced obese mice. McAb-ATR activated insulin signaling in skeletal muscle and insulin-resistant C2C12 myotubes without affecting liver or white adipose tissue of ob/ob mice. Mechanistically, the favorable impact of McAb-ATR on insulin resistance was abolished in db/db mice and in C2C12 myotubes with leptin receptor knockdown. AT1R knockdown also eradicated the effects of McAb-ATR in C2C12 myotubes. Furthermore, McAb-ATR treatment was able to activate the leptin receptor-mediated JAK2/STAT3 signaling in skeletal muscle of ob/ob mice and C2C12 myotubes. Additionally, angiotensin II downregulated the leptin signaling in skeletal muscle of ob/ob and diet-induced obese mice. We demonstrated that ATRQß-001 vaccine and McAb-ATR improved whole-body insulin resistance and regulated glucose metabolism in skeletal muscle in a leptin receptor-dependent manner. Our data suggest that immunotherapy targeting AT1R is a novel strategy for treating insulin resistance.

Immunolocalization of leptin and leptin receptor in colorectal mucosa of ulcerative colitis, Crohn's disease and control subjects with no inflammatory bowel disease.

The expression of leptin and leptin receptor (Ob-R) has been partially elucidated in colon of patients with inflammatory bowel diseases (IBDs), even though leptin is involved in angiogenesis and inflammation. We previously reported overexpression of GLUT5 fructose transporter, in aberrant clusters of lymphatic vessels in lamina propria of IBD and controls. Here, we examine leptin and Ob-R expression in the same biopsies. Specimens were obtained from patients with ulcerative colitis (UC), Crohn's disease (CD) and controls who underwent screening for colorectal cancer, follow-up after polypectomy or with a history of lower gastrointestinal symptoms. Immunohistochemistry revealed leptin in apical and basolateral membranes of short epithelial portions, Ob-R on the apical pole of epithelial cells. Leptin and Ob-R were also identified in structures and cells scattered in the lamina propria. In UC, a significant correlation between leptin and Ob-R in the lamina propria was found in all inflamed samples, beyond non-inflamed samples of the proximal tract, while in CD, it was found in inflamed distal samples. Most of the leptin and Ob-R positive areas in the lamina propria were also GLUT5 immunoreactive in inflamed and non-inflamed mucosa. A significant correlation of leptin or Ob-R expression with GLUT5 was observed in the inflamed distal samples from UC. Our findings suggest that there are different sites of leptin and Ob-R expression in large intestine and those in lamina propria do not reflect the status of mucosal inflammation. The co-localization of leptin and/or Ob-R with GLUT5 may indicate concomitance effects in colorectal lamina propria areas.

Correlation between leptin and IFN-γ involved in granulosa cell apoptosis in PCOS.

OBJECTIVES: Our study aimed to explore the relationship between leptin and IFN-γ in PCOS patients, and confirmed the effect of leptin-induced IFN-γ on granulosa cells furtherly. METHODS: 29 patients with PCOS and 36 healthy controls were enrolled. Leptin level and the proportion of Th1 cells were detected and association between them were analyzed. Meanwhile, peripheral blood mononuclear cells (PBMCs) isolated from PCOS patients were treated with leptin and then the proportion of Th1 was analyzed. Besides that, the apoptotic level of KGN cells was monitored after IFN-γ treatment. RESULTS: In the circulation of PCOS patients, leptin level dramatically increased compared with controls. And, this was associated with upregulated Th1 cells proportion and IFN-γ level. In vitro, Th1 cells proportion increased after leptin treated PBMCs from PCOS patients. Furthermore, for KGN cells, the percentage of live cells decreased and later apoptosis cells increased after IFN-γ treatment. CONCLUSIONS: Our results indicated that leptin takes part in process of PCOS via inducing expression of IFN-γ. Our findings highlight the importance of the connection between leptin and inflammation in PCOS and provide new insights therapeutic strategy for this disease.

The Role of the Adipokine Leptin in Immune Cell Function in Health and Disease.

Leptin is a critical mediator of the immune response to changes in overall nutrition. Leptin is produced by adipocytes in proportion to adipose tissue mass and is therefore increased in obesity. Despite having a well-described role in regulating systemic metabolism and appetite, leptin displays pleiotropic actions, and it is now clear that leptin has a key role in influencing immune cell function. Indeed, many immune cells have been shown to respond to leptin directly via the leptin receptor, resulting in a largely pro-inflammatory phenotype. Understanding the role of adipose-tissue derived mediators in inflammation is critical to determining the pathophysiology of multiple obesity-associated diseases, such as type 2 diabetes, autoimmune disease, and infection. This review, therefore, focuses on the latest data regarding the role of leptin in modulating inflammation.

Alternatively activated macrophage-derived secretome stimulates ovarian cancer spheroid spreading through a JAK2/STAT3 pathway.

High-grade serous ovarian cancer (HGSOC) metastasizes when tumor spheroids detach from the primary tumor and re-attach throughout the peritoneal cavity. Once the cancer cells have implanted in these new sites, the development of metastatic lesions is dependent on the disaggregation of cancer cells from the spheroids and subsequent expansion across the collagenous extracellular matrix (ECM). As HGSOC progresses an increase in alternatively activated macrophages (AAMs) in the surrounding ascites fluid has been observed and AAMs have been shown to enhance tumor invasion and growth in a wide range of cancers. We hypothesized that soluble factors from AAMs in the peritoneal microenvironment promote the disaggregation of ovarian cancer spheroids across the underlying ECM. We determined that co-culture with AAMs significantly increased HGSOC spheroid spreading across a collagen matrix. Multivariate modeling identified AAM-derived factors that correlated with enhanced spread of HGSOC spheroids and experimental validation showed that each individual cell line responded to a distinct AAM-derived factor (FLT3L, leptin, or HB-EGF). Despite this ligand-level heterogeneity, we determined that the AAM-derived factors utilized a common signaling pathway to induce spheroid spreading: JAK2/STAT3 activation followed by MMP-9 mediated spreading. Furthermore, immunostaining demonstrated that FLT3, LEPR, EGFR, and pSTAT3 were upregulated in metastases in HGSOC patients, with substantial patient-to-patient heterogeneity. These results suggest that inhibiting individual soluble factors will not inhibit AAM-induced effects across a broad group of patients; instead, the downstream JAK2/STAT3/MMP-9 pathway should be examined as potential therapeutic targets to slow metastasis in ovarian cancer.

Evidence for leptin receptor immunoreactivity in the gastrointestinal tract and gastric leptin regulation in the rainbow trout (Oncorhynchus mykiss).

In this study, evidence for leptin receptor (LR) and gastric leptin immunoreactivity along the digestive tract of the rainbow trout (Oncorhynchus mykiss), is reported. Besides this, the regulation of gastric leptin and its transcript by fatty acids was analyzed in vitro. LR was detected mainly in the cells of the stomach gastric glands and in the brush border of the epithelium of the anterior, middle and distal intestine. In the stomach LR was co-distributed with leptin. The regulation of gastric leptin and its transcript by fatty acids was analyzed by in vitro incubations. Rabbit polyclonal antibodies anti rainbow trout leptin were developed and employed to detect leptin concentration in the stomach and in the incubation medium. Stomach slices were incubated with butyric (4:0), oleic (18:1n-9), α-linolenic (18:3n-3) and arachidonic fatty acids (20:4n-6). All fatty acids caused an increase in the protein in both the stomach and culture medium, while leptin transcript was not modified. Overall, the results confirm the gastric leptin release upon nutritional modulation.

C-Reactive Protein (CRP) and Leptin Receptor in Obesity: Binding of Monomeric CRP to Leptin Receptor.

While leptin deficiency or dysfunction leads to morbid obesity, obese subjects are characterized paradoxically by hyperleptinemia indicating lack of response to leptin. C-reactive protein (CRP) has been suggested to be a key plasma protein that could bind to leptin. To examine whether CRP interferes with leptin action, mediated through its cell surface receptor, docking studies of CRP with the extracellular domain of the leptin receptor were done employing bioinformatics tools. Monomeric CRP docked with better Z-rank score and more non-bond interactions than pentameric CRP at the CRH2-FNIII domain proximal to the cell membrane, distinct from the leptin-docking site. Interaction of CRP with leptin receptor was validated by solid phase binding assay and co-immunoprecipitation of CRP and soluble leptin receptor (sOb R) from human plasma. Analysis of the serum levels of leptin, CRP, and sOb R by ELISA showed that CRP levels were significantly elevated (p < 0.0001) in non-morbid obese subjects (n = 42) compared to lean subjects (n = 32) and correlated positively with body mass index (BMI) (r = 0.74, p < 0.0001) and leptin (r = 0.8, p < 0.0001); levels of sOb R were significantly low in obese subjects (p < 0.001) and showed a negative correlation with BMI (r = -0.26, p < 0.05) and leptin (r = -0.23, p < 0.05) indicating a minimal role for sOb R in sequestering leptin.

Ablation of the leptin receptor in myeloid cells impairs pulmonary clearance of Streptococcus pneumoniae and alveolar macrophage bactericidal function.

Leptin is a pleiotropic hormone produced by white adipose tissue that regulates appetite and many physiological functions, including the immune response to infection. Genetic leptin deficiency in humans and mice impairs host defenses against respiratory tract infections. Since leptin deficiency is associated with obesity and other metabolic abnormalities, we generated mice that lack the leptin receptor (LepRb) in cells of the myeloid linage (LysM-LepRb-KO) to evaluate its impact in lean metabolically normal mice in a murine model of pneumococcal pneumonia. We observed higher lung and spleen bacterial burdens in LysM-LepRb-KO mice following an intratracheal challenge with Streptococcus pneumoniae. Although numbers of leukocytes recovered from bronchoalveolar lavage fluid did not differ between groups, we did observe higher levels of pulmonary IL-13 and TNFα in LysM-LepRb-KO mice 48 h post infection. Phagocytosis and killing of ingested S. pneumoniae were also impaired in alveolar macrophages (AMs) from LysM-LepRb-KO mice in vitro and were associated with reduced LTB4 and enhanced PGE2 synthesis in vitro. Pretreatment of AMs with LTB4 and the cyclooxygenase inhibitor, indomethacin, restored phagocytosis but not bacterial killing in vitro. These results confirm our previous observations in leptin-deficient ( ob/ob) and fasted mice and demonstrate that decreased leptin action, as opposed to metabolic irregularities associated with obesity or starvation, is responsible for the defective host defense against pneumococcal pneumonia. They also provide novel targets for therapeutic intervention in humans with bacterial pneumonia.

E74-Like Factor (ELF3) and Leptin, a Novel Loop Between Obesity and Inflammation Perpetuating a Pro-Catabolic State in Cartilage.

BACKGROUND/AIMS: The E74-like factor 3 (ELF3) is an inflammatory mediator that participates in cartilage destruction in osteoarthritis. Leptin and other adipokines negatively impact articular cartilage, triggering catabolic and inflammatory responses in chondrocytes. Here, we investigated whether leptin induces ELF3 expression in chondrocytes and the signaling pathway involved in this process. METHODS: We determined mRNA and protein levels of ELF3 by RT-qPCR and Western blotting using cultured human primary chondrocytes and the human T/C-28a2 chondrocyte cell line. Further, we measured luciferase activities of different reporter constructs, and we assessed the contribution of leptin to the induction of ELF3 mRNA by knocking down hLEPR gene expression using siRNA technology. RESULTS: Leptin synergizes with IL-1ß in inducing ELF3 expression in chondrocytes. We also found that PI3K, p38, and JAK2 signaling pathways are at play in the leptin-driven induction of ELF3. Moreover, we confirm the participation of NFΚB in the leptin/IL-1ß synergistic induction of ELF3. CONCLUSION: Here we show, for the first time, the regulation of ELF3 expression by leptin, suggesting that this transcription factor likely mediates the inflammatory responses triggered by leptin in articular chondrocytes.

Siglec-1 and -2 as potential biomarkers in autoimmune disease.

Autoimmune diseases (ADs) are currently treated with anti-inflammatory and immunosuppressive drugs, aimed at reducing symptoms of disease in order to improve quality of life for patients. However, for a significant number of patients these therapies are ineffective, leading to an increased risk of irreversible damage and eventual disability in certain cases. Growing evidence has implicated glycosylated proteins and their cognate receptors in modulation of the autoimmune response. This review will summarize these findings with particular focus on sialic acid-binding immunoglobulin-like lectin (Siglec)-1 and Siglec-2 involvement in AD. Fluctuations in these glycosylation-dependent pathways could act as sentinels of disease activity or drug responses. If validated, protein modification and cellular response markers could help clinicians achieve remission earlier.

Human breast milk and adipokines--A potential role for the soluble leptin receptor (sOb-R) in the regulation of infant energy intake and development.

Concentrations of different adipokines in human breast milk are thought to be able to affect energy intake of the infant. Leptin is a hormone synthesized by adipose tissue and the human placenta and favors satiety. The availability of leptin in breast milk is influenced by epithelial cells of the mammary gland that are known to be able to produce leptin, as well as leptin from maternal circulation that is transported to the breast milk, and which can thus in turn reach neonatal blood after absorption. Research so far as mainly focused on leptin concentrations in breast milk. However, evidence suggests that in addition to leptin concentrations levels of the so-called soluble leptin receptor (sOb-R), the main high-affinity binding protein for leptin in humans, are necessary in order to calculate the free leptin index (FLI) and to assess function of the leptin axis. FLI is calculated from the ratio of leptin to the sOb-R, and serves as the main parameter for assessing function of the leptin axis throughout maturation and development. Here we propose that assessing sOb-R levels in addition to leptin concentrations in breast milk could serve as a valuable tool to investigate effects of the leptin axis in breast milk because sOb-R concentrations can impact available leptin levels, and which in turn can have significant implications for infant energy intake and related development.

Leptin Regulation of Immune Responses.

Leptin is a regulatory hormone with multiple roles in the immune system. We favor the concept that leptin signaling 'licenses' various immune cells to engage in immune responses and/or to differentiate. Leptin is an inflammatory molecule that is capable of activating both adaptive and innate immunity. It can also 'enhance' immune functions, including inflammatory cytokine production in macrophages, granulocyte chemotaxis, and increased Th17 proliferation. Leptin can also 'inhibit' cells; CD4(+) T cells are inhibited from differentiating into regulatory T cells in the presence of elevated leptin, while NK cells can exhibit impaired cytotoxicity under the same circumstances. Consequently, understanding the effect of leptin signaling is important to appreciate various aspects of immune dysregulation observed in malnutrition, obesity, and autoimmunity.

Leptin receptor signaling in T cells is required for Th17 differentiation.

The hormone leptin plays a key role in energy homeostasis, and the absence of either leptin or its receptor (LepR) leads to severe obesity and metabolic disorders. To avoid indirect effects and to address the cell-intrinsic role of leptin signaling in the immune system, we conditionally targeted LepR in T cells. In contrast with pleiotropic immune disorders reported in obese mice with leptin or LepR deficiency, we found that LepR deficiency in CD4(+) T cells resulted in a selective defect in both autoimmune and protective Th17 responses. Reduced capacity for differentiation toward a Th17 phenotype by lepr-deficient T cells was attributed to reduced activation of the STAT3 and its downstream targets. This study establishes cell-intrinsic roles for LepR signaling in the immune system and suggests that leptin signaling during T cell differentiation plays a crucial role in T cell peripheral effector function.

Immunohistochemical evidence of leptin and its receptor in the carpal glands of domestic pigs and wild boar.

BACKGROUND: In recent years, the protein hormone leptin has been the subject of numerous studies designed to clarify and interpret its functional significance; it has been speculated that this goes well beyond the control of appetite and energy metabolism. In particular, the presence of leptin and its receptor has been observed in various glands anatomically and functionally related to the reproductive apparatus. This has led to the hypothesis that leptin may act directly in the functional control of these glands and, in general, the control of reproductive function. HYPOTHESIS/OBJECTIVES: The presence and distribution of leptin and its receptor in the carpal glands of domestic pigs and wild boar are examined, using immunohistochemical techniques. ANIMALS: Tissue samples were collected from five domestic pigs and five wild boar, following slaughter. RESULTS: The presence of leptin and its receptor was demonstrated in the glands, localized in the dark cells of the glandular secretory epithelium. In addition, no difference was observed between wild boar and domestic pigs. CONCLUSIONS AND CLINICAL IMPORTANCE: We hypothesize that leptin may be produced by the carpal gland and may act on the gland's secretory epithelial cells with an autocrine/paracrine mechanism, thus affecting the secretory activity of the gland itself.

Creating leptin-like biofunctions by active immunization against chicken leptin receptor in growing chickens.

In this study, immunization against chicken leptin receptor (cLEPR) extracellular domain (ECD) was applied to investigate leptin regulation and LEPR biofunction in growing chicken pullets. A recombinant protein (cLEPR ECD) based on the cLEPR complemenary DNA sequence corresponding to the 582nd to 796th amino acid residues of cLEPR mature peptide was prepared and used as antigen. Immunization against cLEPR ECD in growing chickens increased anti-cLEPR ECD antibody titers in blood, enhanced proportions of phosphorylated janus kinase 2 (JAK2) and served as signal transducer and activator of transcription 3 (STAT3) protein in liver tissue. Chicken live weight gain and abdominal fat mass were significantly decreased (P < 0.05), but feed intake was stimulated by cLEPR ECD immunization (P < 0.05). The treatment also upregulated the gene expression levels of lepR, AMP-activated protein kinase (AMPK), acetyl CoA carboxylase-2 (ACC2), and uncoupling protein 3 (UCP3) in liver, abdominal fat, and breast muscle (P < 0.05) but decreased fasn expression levels (P < 0.01). Apart from that of lepR, the expression of appetite-regulating genes, such as orexigenic genes, agouti-related peptide (AgRP) and neuropeptide Y (NPY), were upregulated (P < 0.01), whereas the anorexigenic gene proopiomelanocortin (POMC) was downregulated in the hypothalamic tissue of cLEPR-immunized pullets (P < 0.01). Blood concentrations of metabolic molecules, such as glucose, triglycerides, and very-low-density lipoprotein, were significantly decreased in cLEPR-immunized pullets but those of cholesterol, high-density lipoprotein, and low-density lipoprotein increased. These results demonstrate that antibodies to membrane proximal cLEPR ECD enhance cLEPR signal transduction, which stimulates metabolism and reduces fat deposition in chickens.

Screening of synthetic phage display scFv libraries yields competitive ligands of human leptin receptor.

Initially considered the main endogenous anorexigenic factor, fat-derived leptin turned out to be a markedly pleiotropic hormone, influencing diverse physiological processes. Moreover, hyperleptinemia in obese individuals has been linked to the onset or progression of serious disorders, such as cancer, autoimmune diseases, and atherosclerosis, and antagonizing peripheral leptin's signalization has been shown to improve these conditions. To develop an antibody-based leptin antagonist we have devised a tailored panning procedure and screened two phage display libraries of single chain variable antibody fragments (scFvs) against recombinant leptin receptor. One of the scFvs was expressed in Escherichia coli and its interaction with leptin receptor was characterized in more detail. It was found to recognize a discontinuous epitope and to compete with leptin for receptor binding with IC50 and Kd values in the nanomolar range. The reported scFv represents a lead for development of leptin antagonists that may ultimately find use in therapy of various hyperleptinemia-related disorders.

Hepatitis C virus core protein stimulates fibrogenesis in hepatic stellate cells involving the obese receptor.

Hepatitis C virus core protein (HCVcp), which is secreted by infected cells, is reported as an immunomodulator in immune cells. However, the effects of HCVcp on hepatic stellate cells (HSCs), the key cells in liver fibrosis, still remain unclear. In this study, we investigated the effects of HCVcp on obese receptor (ObR) related downstream signaling pathways and fibrogenic gene expression in HSCs. LX-2, a human HSC line, was incubated with HCVcp. Inhibitors and short interfering RNAs were used to interrogate the mechanisms of HCVcp action on HSCs. HCVcp (20-100 ng/ml) concentration-dependently stimulated α-smooth muscle actin (α-SMA) protein expression and mRNA expression of α-SMA, procollagen α2(I) and TGF-ß1 genes, with a plateau of 220% of controls at 100 ng/ml. HCVcp induced mRNA and protein expression of ObR. Blocking of Ob-Rb with a neutralizing antibody inhibited phosphorylation of signal transducer and activator of transcription 3 (STAT3) and AMPKα stimulated by HCVcp. Furthermore, knockdown of Ob-Rb down-regulated HCVcp-induced STAT3, AKT, and AMPKα phosphorylation, and reversed HCVcp-suppressed mRNA expression of matrix metalloproteinase (MMP)-1, peroxisome proliferator-activated receptor (PPAR)γ and sterol regulatory element binding protein-1c (SREBP-1c) genes. AMPKα signaling blockade reversed HCVcp-suppressed SREBP-1c mRNA expression. HCVcp stimulated reactive oxygen species formation and gp91(phox) (a component of NADPH oxidase) protein expression, together with AKT phosphorylation, leading to suppression of PPARγ and SREBP-1c genes. Our results provide a new finding that HCVcp induced ObR-dependent Janus Kinase (JAK) 2-STAT3, AMPKα, and AKT signaling pathways and modulated downstream fibrogenetic gene expression in HSCs.

Leptin exacerbates collagen-induced arthritis via enhancement of Th17 cell response.

OBJECTIVE: To determine the role of leptin in modulating Th17 cell response and joint inflammation in mice with collagen-induced arthritis (CIA). METHODS: Leptin receptor expression on T cells was examined by polymerase chain reaction (PCR) analysis, immunofluorescence microscopy, and flow cytometry. Effects of leptin on Th17 cell differentiation and proliferation were evaluated by quantitative PCR, carboxyfluorescein diacetate succinimidyl ester proliferation assay, and flow cytometry. Dynamic changes in leptin concentrations in the joint tissue and synovial fluid of mice with CIA were determined by immunohistochemistry analysis and enzyme-linked immunosorbent assay (ELISA). Arthritis symptoms and joint pathology in mice with CIA were assessed after injection of leptin into the knee joint. Th1 and Th17 cell populations in the spleen, draining lymph nodes, and joint tissue were analyzed by flow cytometry and enzyme-linked immunospot assay. Interleukin-17 messenger RNA and protein levels in the joint tissue were measured by PCR analysis and ELISA. RESULTS: In culture, leptin treatment significantly increased Th17 cell generation from naive CD4+ T cells. During CIA development, markedly elevated levels of leptin were detected in the joint tissue and synovial fluid. Moreover, injection of leptin into the knee joint of collagen-immunized mice resulted in an early onset of arthritis and substantially increased the severity of clinical symptoms, accompanied by more pronounced synovial hyperplasia and joint damage. Further examination by immunofluorescence microscopy confirmed the presence of significantly increased numbers of Th17 cells in the joint tissue and draining lymph nodes of leptin-treated mice with CIA. CONCLUSION: The results of this study identify a previously undescribed function of leptin in enhancing Th17 cell response and exacerbating joint inflammation in mice with CIA.

Leptin and mucosal immunity.

Enhanced susceptibility to infection has long been recognized in children with congenital deficiency of leptin or its receptor. Studies in mice have demonstrated that leptin deficiency affects both the innate and acquired immune systems. Here, we review recent studies that demonstrate the impact on immunity of a common non-synonomous polymorphism of the leptin receptor. In a Bangladesh cohort of children, the presence of two copies of the ancestral Q223 allele was significantly associated with resistance to amebiasis. Children and mice with at least one copy of the leptin receptor 223R mutation were more susceptible to amebic colitis. Leptin signaling in the intestinal epithelium and downstream STAT3 (signal transducer and activator of transcription 3) and SHP2 (Src homology phosphatase 2) signaling were required for protection in the murine model of amebic colitis. Murine models have also implicated leptin in protection from other infections, including Mycobacterium tuberculosis, Klebsiella pneumoniae, and Streptococcus pneumoniae. Thus, the role of leptin signaling in infectious disease and specifically leptin-mediated protection of the intestinal epithelium will be the focus of this review.

Specific glycan elements determine differential binding of individual egg glycoproteins of the human parasite Schistosoma mansoni by host C-type lectin receptors.

During infection with the blood fluke Schistosoma mansoni, glycan motifs present on glycoproteins of the parasite's eggs mediate immunomodulatory effects on the host. The recognition of these glycan motifs is primarily mediated by C-type lectin receptors on dendritic cells and other cells of the immune system. However, it is not yet known which individual glycoproteins interact with the different C-type lectin receptors, and which structural components are involved. Here we investigated the structural basis of the binding of two abundant egg antigens, kappa-5 and IPSE/a1, by the C-type lectin receptor dendritic cell-specific ICAM3-grabbing non-integrin, macrophage galactose-type lectin and mannose receptor. In the natural soluble form, the secretory egg glycoprotein IPSE/a1 interacts with dendritic cells mainly via mannose receptors. Surprisingly, in plate-based assays mannose receptors preferentially bound to mannose conjugates, while in cell-based assays, IPSE/a1 is bound via the fucosylated Galb1-4(Fuca1-3)GlcNAc (LeX) motif on diantennary N-glycans. Kappa-5, in contrast, is bound by dendritic cells viaall three C-type lectin receptors studied and for a minor part also via other, non-C-type lectin receptors.Kappa-5 interacts with macrophage galactose-type lectins via the GalNAcb1-4GlcNAc antenna present on its triantennary N-glycans, as well as the GalNAcb1-4(Fuca1-3) GlcNAc antennae present on a minor N-glycan subset. Dendritic cell-specific ICAM3-grabbing non-integrin binding of kappa-5 was mediated via the GalNAcb1-4(Fuca1-3)GlcNAc antennae, whereas binding of mannose receptors may involve either GalNAcb1-4(Fuca1-3)GlcNAc antennae or the fucosylated and xylosylated chitobiose core. This study provides a molecular and structural basis for future studies of the interaction between C-type lectin receptors and other soluble egg antigen glycoproteins and their effects on the host immune response.