Innate phase production of IFN-γ by memory and effector T cells expressing early activation marker CD69 during infection with Cryptococcus deneoformans in the lungs.

Cryptococcus deneoformans is a yeast-type fungus that causes fatal meningoencephalitis in immunocompromised patients and evades phagocytic cell elimination through an escape mechanism. Memory T (Tm) cells play a central role in preventing the reactivation of this fungal pathogen. Among these cells, tissue-resident memory T (TRM) cells quickly respond to locally invaded pathogens. This study analyzes the kinetics of effector T (Teff) cells and Tm cells in the lungs after cryptococcal infection. Emphasis is placed on the kinetics and cytokine expression of TRM cells in the early phase of infection. CD4+ Tm cells exhibited a rapid increase by day 3, peaked at day 7, and then either maintained their levels or exhibited a slight decrease until day 56. In contrast, CD8+ Tm cells reached their peak on day 3 and thereafter decreased up to day 56 post-infection. These Tm cells were predominantly composed of CD69+ TRM cells and CD69+ CD103+ TRM cells. Disruption of the CARD9 gene resulted in reduced accumulation of these TRM cells and diminished interferon (IFN) -γ expression in TRM cells. TRM cells were derived from T cells with T cell receptors non-specific to ovalbumin in OT-II mice during cryptococcal infection. In addition, TRM cells exhibited varied behavior in different tissues. These results underscore the importance of T cells, which produce IFN-γ in the lungs during the early stage of infection, in providing early protection against cryptococcal infection through CARD9 signaling.

Structure of the human Bre1 complex bound to the nucleosome.

Histone H2B monoubiquitination (at Lys120 in humans) regulates transcription elongation and DNA repair. In humans, H2B monoubiquitination is catalyzed by the heterodimeric Bre1 complex composed of Bre1A/RNF20 and Bre1B/RNF40. The Bre1 proteins generally function as tumor suppressors, while in certain cancers, they facilitate cancer cell proliferation. To obtain structural insights of H2BK120 ubiquitination and its regulation, we report the cryo-electron microscopy structure of the human Bre1 complex bound to the nucleosome. The two RING domains of Bre1A and Bre1B recognize the acidic patch and the nucleosomal DNA phosphates around SHL 6.0-6.5, which are ideally located to recruit the E2 enzyme and ubiquitin for H2BK120-specific ubiquitination. Mutational experiments suggest that the two RING domains bind in two orientations and that ubiquitination occurs when Bre1A binds to the acidic patch. Our results provide insights into the H2BK120-specific ubiquitination by the Bre1 proteins and suggest that H2B monoubiquitination can be regulated by nuclesomal DNA flexibility.

Continuous Electrocardiographic Monitoring for 24 Hours Before Death in Patients with Terminal Cancer.

BACKGROUND: Purposeless physiological monitoring at the end-of-life is not recommended. However, studies on how families feel regarding the death of patients with terminal cancer without continuous electrocardiographic monitoring (CEM) are lacking. OBJECTIVES: To explore the impact on the quality of care and the feelings and psychological distress experienced by families when CEM is not used during the 24 hours preceding a patient's death. METHODS: In this multicenter cross-sectional, self-report questionnaires were distributed to 1087 bereaved families at Japanese specialized palliative care units in 2018. RESULTS: Out of 671 responses, 394 valid responses were analyzed. Families of nonmonitored patients (NM-group) accounted for 79.2%, while those with bedside electrocardiogram monitoring (MB-group) and remote nurse station monitoring (MC-group) comprised 11.9% and 8.9%, respectively. In the NM-group, 85.5% expressed satisfaction without CEM, which was more than 10% lower than other groups. While 14% in the NM-group desired patient monitoring, families who received adequate explanations about CEM had lower proportions compared to the MB-group (P = .021). Univariate analyses showed no significant differences in evaluations of the quality of care and families' psychological distress (mean scores of Overall Care Satisfaction, Care Evaluation Scale, Good Death Inventory, Brief Grief Questionnaires) across all groups. CONCLUSION: While the majority of NM-group were satisfied with their patient's care without CEM, the proportion of dissatisfied families was higher than in other groups. Although not using CEM is not a major hindrance to end-of-life care for patients with terminal cancer, providing sufficient explanations may be important for satisfactory care.

CARD9-mediated macrophage responses and collagen fiber capsule formation caused by textured-type breast implants.

BACKGROUND: An increasing number of women are undergoing breast implantation for cosmetic purposes and for reconstructive purposes after breast excision. The surface morphology of the breast implant is one of the key factors associated with the induction of capsule contraction. The effect of surface morphology on the inflammatory response following implant insertion remains unclear, however. This study conducted comparative analyses to determine the effect of the textured and smooth surface morphology of silicone sheets. METHODS: Each type of silicone sheet was inserted into the subcutaneous pocket below the panniculus carnosus in C57BL/6 mice and mice with genetic disruption of CARD9, Dectin-1, Dectin-2, or Mincle. We also analyzed the collagen fiber capsule thickness, histological findings, and macrophage inflammatory response, including TGF-ß synthesis. RESULTS: We found that textured surface morphology contributed to the formation of collagen fiber capsules and the accumulation of fibroblasts and myofibroblasts, and was accompanied by the accumulation of TGF-ß-expressing macrophages and foreign-body giant cells. CARD9 deficiency attenuated collagen fiber capsule formation, macrophage responses, and TGF-ß synthesis, although the responsible C-type lectin receptors (CLRs) remain to be clarified. CONCLUSIONS: These results suggest that CARD9 may have a strong impact on silicone sheet insertion through the regulation of macrophage responses.

Cutaneous wound healing promoted by topical administration of heat-killed Lactobacillus plantarum KB131 and possible contribution of CARD9-mediated signaling.

Optimal conditions for wound healing require a smooth transition from the early stage of inflammation to proliferation, and during this time alternatively activated (M2) macrophages play a central role. Recently, heat-killed lactic acid bacteria (LAB), such as Lactobacillus plantarum (L. plantarum) have been reported as possible modulators affecting the immune responses in wound healing. However, how signaling molecules regulate this process after the administration of heat-killed LAB remains unclear. In this study, we examined the effect of heat-killed L. plantarum KB131 (KB131) administration on wound healing and the contribution of CARD9, which is an essential signaling adaptor molecule for NF-kB activation upon triggering through C-type lectin receptors, in the effects of this bacterium. We analyzed wound closure, histological findings, and inflammatory responses. We found that administration of KB131 accelerated wound closure, re-epithelialization, granulation area, CD31-positive vessels, and α-SMA-positive myofibroblast accumulated area, as well as the local infiltration of leukocytes. In particular, M2 macrophages were increased, in parallel with CCL5 synthesis. The acceleration of wound healing responses by KB131 was canceled in CARD9-knockout mice. These results indicate that the topical administration of KB131 accelerates wound healing, accompanying increased M2 macrophages, which suggests that CARD9 may be involved in these responses.

Mouse Model of Latent Cryptococcal Infection and Reactivation.

AbstractCryptococcus neoformans is an opportunistic fungal pathogen that frequently causes fatal meningoencephalitis in patients with impaired immune responses. This fungus, an intracellularly growing microbe, evades host immunity, leading to a latent infection (latent C. neoformans infection: LCNI), and cryptococcal disease is developed by its reactivation when host immunity is suppressed. Elucidation of the pathophysiology of LCNI is difficult due to the lack of mouse models. Here we show the established methods for LCNI and reactivation.

Structural basis of transcription regulation by CNC family transcription factor, Nrf2.

Several basic leucine zipper (bZIP) transcription factors have accessory motifs in their DNA-binding domains, such as the CNC motif of CNC family or the EHR motif of small Maf (sMaf) proteins. CNC family proteins heterodimerize with sMaf proteins to recognize CNC-sMaf binding DNA elements (CsMBEs) in competition with sMaf homodimers, but the functional role of the CNC motif remains elusive. In this study, we report the crystal structures of Nrf2/NFE2L2, a CNC family protein regulating anti-stress transcriptional responses, in a complex with MafG and CsMBE. The CNC motif restricts the conformations of crucial Arg residues in the basic region, which form extensive contact with the DNA backbone phosphates. Accordingly, the Nrf2-MafG heterodimer has approximately a 200-fold stronger affinity for CsMBE than canonical bZIP proteins, such as AP-1 proteins. The high DNA affinity of the CNC-sMaf heterodimer may allow it to compete with the sMaf homodimer on target genes without being perturbed by other low-affinity bZIP proteins with similar sequence specificity.

PAMPs and Host Immune Response in Cryptococcal Infection.

Cryptococcus spp. are yeast-type opportunistic fungal pathogens with thick polysaccharide capsules that infect the lungs via airborne routes and frequently cause fatal meningoencephalitis. The cellular immune mechanism plays a central role in controlling cryptococcal infection and is critically regulated by Th1-Th2 immune balance. Pathogens that have invaded the host are recognized by innate immune cells, and appropriate immune responses are initiated. Pathogen-associated molecular patterns (PAMPs) are recognized by macrophages and dendritic cells via pattern recognition receptors (PRRs), which trigger the inflammatory responses as the first line of host defense. Some PRRs, such as Toll-like receptors (TLRs), NOD-like receptors (NLRs), and C-type lectin receptors (CLRs), are involved in the recognition of cryptococcal components, such as glucuronoxylomannan (GXM), mannoproteins (MPs), and nucleic acids. However, some cryptococcal cell components suppress the host immune response. This review will highlight the cryptococcal components involved in host immune responses. Future research is expected to promote the understanding of the mechanism of host immune response to Cryptococcus, which will lead to the development of new vaccines and therapies for cryptococcal infection.

Practical Validation of United States Centers for Disease Control and Prevention Assays for the Detection of Human Respiratory Syncytial Virus in Pediatric Inpatients in Japan.

The World Health Organization initiated a global surveillance system for respiratory syncytial virus (RSV) in 2015, and the pilot surveillance is ongoing. The real-time RT-PCR RSV assays (Pan-RSV and duplex assays) developed by the United States Centers for Disease Control and Prevention are applied as the standard assays. To introduce these as standard assays in Japan, their practicality was evaluated using 2261 specimens obtained from pediatric inpatients in Japan, which were collected from 2018 to 2021. Although the Pan-RSV and duplex assays had similar analytical sensitivities, they yielded 630 (27.9%) and 786 (34.8%) RSV-positive specimens, respectively (p < 0.001). Although sequencing analysis showed mismatches in the reverse primer used in the Pan-RSV assay, these mismatches did not affect its analytical sensitivity. The analysis of read numbers of RSV isolates from air−liquid interface culture of human bronchial/tracheal epithelial cells showed that the duplex assay had a greater number of reads than did the Pan-RSV assay. Therefore, the duplex assay has superior detection performance compared with the Pan-RSV assay, but the two assays have similar analytical sensitivities.

Suitability of NIID-MDCK cells as a substrate for cell-based influenza vaccine development from the perspective of adventitious virus susceptibility.

The practical use of cell-based seasonal influenza vaccines is currently being considered in Japan. From the perspective of adventitious virus contamination, we assessed the suitability of NIID-MDCK cells (NIID-MDCK-Cs) as a safe substrate for the isolation of influenza viruses from clinical specimens. We first established a sensitive multiplex real-time PCR system to screen for 27 respiratory viruses and used it on 34 virus samples that were isolated by passaging influenza-positive clinical specimens in NIID-MDCK-Cs. Incidentally, the limit of detection (LOD) of the system was 100 or fewer genome copies per reaction. In addition to influenza viruses, human enterovirus 68 (HEV-D68) genomes were detected in two samples after two or three passages in NIID-MDCK-Cs. To further investigate the susceptibility of NIID-MDCK-Cs to adventitious viruses, eight common respiratory viruses were subjected to passages in NIID-MDCK-Cs. The genome copy numbers of seven viruses other than parainfluenza 3 decreased below the LOD by passage 4. By passaging in NIID-MDCK-Cs, the genome numbers of the input HEV-D68, 1 × 108 copies, declined to 102 at passage 3 and to under the LOD at passage 4, whereas those of the other six viruses were under the LOD by passage 3. These results implied that during the process of isolating influenza viruses with NIID-MDCK-Cs, contaminating viruses other than parainfluenza 3 can be efficiently removed by passages in NIID-MDCK-Cs. NIID-MDCK-Cs could be a safe substrate for isolating influenza viruses that can be used to develop cell-based influenza vaccine candidate viruses.

Variation in Thermal Stability among Respiratory Syncytial Virus Clinical Isolates under Non-Freezing Conditions.

Virus isolates are not only useful for diagnosing infections, e.g., respiratory syncytial virus (RSV), but can also facilitate many aspects of practical viral studies such as analyses of antigenicity and the action mechanisms of antivirals, among others. We have been isolating RSV from clinical specimens from patients with respiratory symptoms every year since our first isolation of RSV in 1964, and isolation rates have varied considerably over the years. As collected clinical specimens are conventionally stored in a refrigerator from collection to inoculation into cells, we hypothesized that certain storage conditions or associated factors might account for these differences. Hence, we evaluated the thermal stability of a total of 64 viruses isolated from 1998 to 2018 upon storage at 4 °C and 20 °C for a defined duration. Interestingly, and contrary to our current understanding, 22 strains (34%) showed a greater loss of viability upon short-term storage at 4 °C than at 20 °C. Thirty-seven strains (57%) showed an almost equal loss, and only five strains (8%) were more stable at 4 °C than at 20 °C. This finding warrants reconsideration of the temperature for the temporary storage of clinical samples for RSV isolation.

Observational study of rebiopsy in EGFR-TKI-resistant patients with EGFR mutation-positive advanced NSCLC.

The identification of acquired resistance mutations has been essential in non-small-cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR) active mutations. Rebiopsy plays a pivotal role in selecting the optimal treatment for patients who develop resistance to initial EGFR-tyrosine kinase inhibitors (EGFR-TKIs). This multicenter, observational study was conducted to investigate the details of rebiopsy in Japanese clinical practice. The primary endpoints were the implementation rate of rebiopsy and the concordance rate for T790M mutation detection between histological and cytological specimens using the cobas EGFR Mutation Test, version 2. One hundred ninety-four patients with EGFR-mutant NSCLC were enrolled, and 120 patients developed acquired resistance to EGFR-TKIs. The median age was 68 years (range 20-87), and 52.5% of the patients were women. Rebiopsy was performed in 109 patients, and the implementation rate of rebiopsy was 90.8%. The success rates of rebiopsy in the total, histology, cytology and liquid biopsy populations were 67.9%, 81.3%, 66.7% and 43.8%, respectively. The positive percent agreement and the negative percent agreement in the detection of the T790M mutation between the histological and cytological specimens were both 90.9%. Obtaining histological or cytological tissue samples at rebiopsy may contribute to improving the detection rate of the T790M mutation (trial registration number: UMIN000026019).

PD-1 blockade therapy augments the antitumor effects of lymphodepletion and adoptive T cell transfer.

Lymphodepleting cytotoxic regimens enhance the antitumor effects of adoptively transferred effector and naïve T cells. Although the mechanisms of antitumor immunity augmentation by lymphodepletion have been intensively investigated, the effects of lymphodepletion followed by T cell transfer on immune checkpoints in the tumor microenvironment remain unclear. The current study demonstrated that the expression of immune checkpoint molecules on transferred donor CD4+ and CD8+ T cells was significantly decreased in lymphodepleted tumor-bearing mice. In contrast, lymphodepletion did not reduce immune checkpoint molecule levels on recipient CD4+ and CD8+ T cells. Administration of anti-PD-1 antibodies after lymphodepletion and adoptive transfer of T cells significantly inhibited tumor progression. Further analysis revealed that transfer of both donor CD4+ and CD8+ T cells was responsible for the antitumor effects of a combination therapy consisting of lymphodepletion, T cell transfer and anti-PD-1 treatment. Our findings indicate that a possible mechanism underlying the antitumor effects of lymphodepletion followed by T cell transfer is the prevention of donor T cell exhaustion and dysfunction. PD-1 blockade may reinvigorate exhausted recipient T cells and augment the antitumor effects of lymphodepletion and adoptive T cell transfer.

Structural basis of the regulation of the normal and oncogenic methylation of nucleosomal histone H3 Lys36 by NSD2.

Dimethylated histone H3 Lys36 (H3K36me2) regulates gene expression, and aberrant H3K36me2 upregulation, resulting from either the overexpression or point mutation of the dimethyltransferase NSD2, is found in various cancers. Here we report the cryo-electron microscopy structure of NSD2 bound to the nucleosome. Nucleosomal DNA is partially unwrapped, facilitating NSD2 access to H3K36. NSD2 interacts with DNA and H2A along with H3. The NSD2 autoinhibitory loop changes its conformation upon nucleosome binding to accommodate H3 in its substrate-binding cleft. Kinetic analysis revealed that two oncogenic mutations, E1099K and T1150A, increase NSD2 catalytic turnover. Molecular dynamics simulations suggested that in both mutants, the autoinhibitory loop adopts an open state that can accommodate H3 more often than the wild-type. We propose that E1099K and T1150A destabilize the interactions that keep the autoinhibitory loop closed, thereby enhancing catalytic turnover. Our analyses guide the development of specific inhibitors of NSD2.

Topical Administration of Heat-Killed Enterococcus faecalis Strain KH2 Promotes Re-Epithelialization and Granulation Tissue Formation during Skin Wound-Healing.

Lactic acid bacteria (LAB) are known to have beneficial effects on immune responses when they are orally administered as bacterial products. Although the beneficial effects of LAB have been reported for the genera Lactobacillus and Lactococcus, little has been uncovered on the effects of the genus Enterococcus on skin wound-healing. In this study, we aimed to clarify the effect of heat-killed Enterococcus faecalis KH2 (heat-killed KH2) strain on the wound-healing process and to evaluate the therapeutic potential in chronic skin wounds. We analyzed percent wound closure, re-epithelialization, and granulation area, and cytokine and growth factor production. We found that heat-killed KH2 contributed to the acceleration of re-epithelialization and the formation of granulation tissue by inducing tumor necrosis factor-α, interleukin-6, basic fibroblast growth factor, transforming growth factor (TGF)-ß1, and vascular endothelial growth factor production. In addition, heat-killed KH2 also improved wound closure, which was accompanied by the increased production of TGF-ß1 in diabetic mice. Topical administration of heat-killed KH2 might have therapeutic potential for the treatment of chronic skin wounds in diabetes mellitus. In the present study, we concluded that heat-killed KH2 promoted skin wound-healing through the formation of granulation tissues and the production of inflammatory cytokines and growth factors.

Deficiency of lung-specific claudin-18 leads to aggravated infection with Cryptococcus deneoformans through dysregulation of the microenvironment in lungs.

Cryptococcus deneoformans is an opportunistic fungal pathogen that infects the lungs via airborne transmission and frequently causes fatal meningoencephalitis. Claudins (Cldns), a family of proteins with 27 members found in mammals, form the tight junctions within epithelial cell sheets. Cldn-4 and 18 are highly expressed in airway tissues, yet the roles of these claudins in respiratory infections have not been clarified. In the present study, we analyzed the roles of Cldn-4 and lung-specific Cldn-18 (luCldn-18) in host defense against C. deneoformans infection. luCldn-18-deficient mice exhibited increased susceptibility to pulmonary infection, while Cldn-4-deficient mice had normal fungal clearance. In luCldn-18-deficient mice, production of cytokines including IFN-γ was significantly decreased compared to wild-type mice, although infiltration of inflammatory cells including CD4+ T cells into the alveolar space was significantly increased. In addition, luCldn-18 deficiency led to high K+ ion concentrations in bronchoalveolar lavage fluids and also to alveolus acidification. The fungal replication was significantly enhanced both in acidic culture conditions and in the alveolar spaces of luCldn-18-deficient mice, compared with physiological pH conditions and those of wild-type mice, respectively. These results suggest that luCldn-18 may affect the clinical course of cryptococcal infection indirectly through dysregulation of the alveolar space microenvironment.

TMPRSS2 Activates Hemagglutinin-Esterase Glycoprotein of Influenza C Virus.

Influenza C virus (ICV) has only one kind of spike protein, the hemagglutinin-esterase (HE) glycoprotein. HE functions similarly to hemagglutinin (HA) and neuraminidase of the influenza A and B viruses (IAV and IBV, respectively). It has a monobasic site, which is cleaved by some host enzymes. The cleavage is essential to activating the virus, but the enzyme or enzymes in the respiratory tract have not been identified. This study investigated whether the host serine proteases, transmembrane protease serine S1 member 2 (TMPRSS2) and human airway trypsin-like protease (HAT), which reportedly cleave HA of IAV/IBV, are involved in HE cleavage. We established TMPRSS2- and HAT-expressing MDCK cells (MDCK-TMPRSS2 and MDCK-HAT). ICV showed multicycle replication with HE cleavage without trypsin in MDCK-TMPRSS2 cells as well as IAV did. The HE cleavage and multicycle replication did not appear in MDCK-HAT cells infected with ICV without trypsin, while HA cleavage and multistep growth of IAV appeared in the cells. Amino acid sequences of the HE cleavage site in 352 ICV strains were completely preserved. Camostat and nafamostat suppressed the growth of ICV and IAV in human nasal surface epithelial (HNE) cells. Therefore, this study revealed that, at least, TMPRSS2 is involved in HE cleavage and suggested that nafamostat could be a candidate for therapeutic drugs for ICV infection. IMPORTANCE Influenza C virus (ICV) is a pathogen that causes acute respiratory illness, mostly in children, but there are no anti-ICV drugs. ICV has only one kind of spike protein, the hemagglutinin-esterase (HE) glycoprotein on the virion surface, which possesses receptor-binding, receptor-destroying, and membrane fusion activities. The HE cleavage is essential for the virus to be activated, but the enzyme or enzymes in the respiratory tract have not been identified. This study revealed that transmembrane protease serine S1 member 2 (TMPRSS2), and not human airway trypsin-like protease (HAT), is involved in HE cleavage. This is a novel study on the host enzymes involved in HE cleavage, and the result suggests that the host enzymes, such as TMPRSS2, may be a target for therapeutic drugs of ICV infection.

Role of Dectin-2 in the Phagocytosis of Cryptococcus neoformans by Dendritic Cells.

The cell walls and capsules of Cryptococcus neoformans, a yeast-type fungal pathogen, are rich in polysaccharides. Dectin-2 is a C-type lectin receptor (CLR) that recognizes high-mannose polysaccharides. Previously, we demonstrated that Dectin-2 is involved in cytokine production by bone marrow-derived dendritic cells (BM-DCs) in response to stimulation with C. neoformans. In the present study, we analyzed the role of Dectin-2 in the phagocytosis of C. neoformans by BM-DCs. The engulfment of this fungus by BM-DCs was significantly decreased in mice lacking Dectin-2 (Dectin-2 knockout [Dectin-2KO]) or caspase recruitment domain-containing protein 9 (CARD9KO), a common adapter molecule that delivers signals triggered by CLRs, compared to wild-type (WT) mice. Phagocytosis was likewise inhibited, to a similar degree, by the inhibition of Syk, a signaling molecule involved in CLR-triggered activation. A PI3K inhibitor, in contrast, completely abrogated the phagocytosis of C. neoformans. Actin polymerization, i.e., conformational changes in cytoskeletons detected at sites of contact with C. neoformans, was also decreased in BM-DCs of Dectin-2KO and CARD9KO mice. Finally, the engulfment of C. neoformans by macrophages was significantly decreased in the lungs of Dectin-2KO mice compared to WT mice. These results suggest that Dectin-2 may play an important role in the actin polymerization and phagocytosis of C. neoformans by DCs, possibly through signaling via CARD9 and a signaling pathway mediated by Syk and PI3K.

Synthesis of Lignans Based on a Borate-mediated One-pot Sequential Suzuki-Miyaura Coupling of Cyclic Boranes.

Lignans are a group of polyphenolic phytochemicals that possess a large spectrum of chemical structures and biological activities. Here the syntheses of lignans - anwulignan, burseran, dehydroxycubebin, ruburisandrin B, and sesamin - are achieved based on a borate-mediated one-pot sequential Suzuki-Miyaura coupling of cis- and trans-fused bicyclic boranes, which were prepared by diastereoselective cyclic hydroboration of exo-cyclic diene with cyclopentyl- and thexylboranes, respectively. A one-pot sequential Suzuki-Miyaura coupling of each cyclic borate with various aryl bromides initiated by activation of the cyclic borane with the carbon nucleophile provided 2,3-dibenzylbutane derivatives with different aromatic substituents. Finally, the syntheses of naturally occurring lignans were accomplished in several steps from the products of Suzuki-Miyaura coupling.

Dectin-2-mediated initiation of immune responses caused by influenza virus hemagglutinin.

Antigen-presenting cells express pattern recognition receptors (PRRs), which sense pathogen-associated molecular patterns from microorganisms and lead to the induction of inflammatory responses. C-type lectin receptors (CLRs), the representative PRRs, bind to microbial polysaccharides, among which Dectin-2 and Mincle recognize mannose-containing polysaccharides. Because influenza virus (IFV) hemagglutinin (HA) is rich in mannose polysaccharides, Dectin-2 or Mincle may contribute to the recognition of HA. In this study, we addressed the possible involvement of Dectin-2 and Mincle in the viral recognition and the initiation of cytokine production. Interleukin (IL)-12p40 and IL-6 production by bone marrow-derived dendritic cells (BM-DCs) upon stimulation with HA was significantly reduced in Dectin-2 knockout (KO) mice compared to wild-type (WT) mice whereas there was no difference between WT mice and Mincle KO mice. BM-DCs that were treated with Syk inhibitor resulted in a significant reduction of cytokine production upon stimulation with HA. The treatment of BM-DCs with methyl-α-D-mannopyranoside (ManP) also led to a significant reduction in cytokine production by BM-DCs that were stimulated with HA, except for the A/H1N1pdm09 subtype. IL-12p40 and IL-6 synthesis by BM-DCs was completely diminished upon stimulation with HA treated with concanavalin A (ConA)-bound sepharose beads. Finally, GFP expression was detected in reporter cells that were transfected with the Dectin-2 gene, but not with the Mincle gene, when stimulated with HA derived from the A/H3N2 subtype. These data suggested that Dectin-2 may be a key molecule as the sensor for IFV to initiate the immune response and regulate the pathogenesis of IFV infection.