[Fungal Beta-glucans: Structure and Effect on Host Immune Responses].

Fungi are eukaryotic microorganisms that show complex life cycles, including both anamorph and teleomorph stages. Beta-1,3-1,6-glucans (BGs) are major cell wall components in fungi. BGs are also found in a soluble form and are secreted by fungal cells. Studies of fungal BGs extensively expanded from 1960 to 1990 due to their applications in cancer immunotherapy. However, progress in this field slowed down due to the low efficacy of such therapies. In the early 21st century, the discovery of C-type lectin receptors significantly enhanced the molecular understanding of innate immunity. Moreover, pathogen-associated molecular patterns (PAMPs) and pattern recognition receptors (PRRs) were also discovered. Soon, dectin-1 was identified as the PRR of BGs, whereas BGs were established as PAMPs. Then, studies on fungal BGs focused on their participation in the development of deep-seated mycoses and on their role as a source of functional foods. Fungal BGs may have numerous and complex linkages, making it difficult to systematize them even at the primary structure level. Moreover, elucidating the structure of BGs is largely hindered by the multiplicity of genes involved in cell wall biosynthesis, including those for BGs, and by fungal diversity. The present review mainly focused on the characteristics of fungal BGs from the viewpoint of structure and immunological activities.

Existence of natural mouse IgG mAbs recognising epitopes shared by malondialdehyde acetaldehyde adducts and Porphyromonas gingivalis.

Natural Abs are produced by B lymphocytes in the absence of external Ag stimulation. They recognise self, altered self and foreign Ags, comprising an important first-line defence against invading pathogens and serving as innate recognition receptors for tissue homeostasis. Natural IgG Abs have been found in newborns and uninfected individuals. Yet, their physiological role remains unclear. Previously, no natural IgG Abs to oxidation-specific epitopes have been reported. Here, we show the cloning and characterisation of mouse IgG mAbs against malondialdehyde acetaldehyde (MAA)-modified low-density lipoprotein. Sequence analysis reveals high homology with germline genes, suggesting that they are natural. Further investigation shows that the MAA-specific natural IgG Abs cross-react with the major periodontal pathogen Porphyromonas gingivalis and recognise its principle virulence factors gingipain Kgp and long fimbriae. The study provides evidence that natural IgGs may play an important role in innate immune defence and in regulation of tissue homeostasis by recognising and removing invading pathogens and/or modified self-Ags, thus being involved in the development of periodontitis and atherosclerosis.

Differential Responses of Pattern Recognition Receptors to Outer Membrane Vesicles of Three Periodontal Pathogens.

Highly purified outer membrane vesicles (OMVs) of the periodontal pathogens, Porphyromonas gingivalis, Treponema denticola and Tannerella forsythia were produced using tangential flow ultrafiltration, ultracentrifugation and Optiprep density gradient separation. Cryo-TEM and light scattering showed OMVs to be single lipid-bilayers with modal diameters of 75 to 158 nm. Enumeration of OMVs by nanoparticle flow-cytometry at the same stage of late exponential culture indicated that P. gingivalis was the most prolific OMV producer. P. gingivalis OMVs induced strong TLR2 and TLR4-specific responses and moderate responses in TLR7, TLR8, TLR9, NOD1 and NOD2 expressing-HEK-Blue cells. Responses to T. forsythia OMVs were less than those of P. gingivalis and T. denticola OMVs induced only weak responses. Compositional analyses of OMVs from the three pathogens demonstrated differences in protein, fatty acids, lipopolysaccharide, peptidoglycan fragments and nucleic acids. Periodontal pathogen OMVs induced differential pattern recognition receptor responses that have implications for their role in chronic periodontitis.

Hagfish C1q: its unique binding property.

Hagfish C1q (HaC1q) was identified and characterized as a pattern-recognition molecule (PRM) in the hagfish complement system. The serum from hagfish, Eptatretus burgeri, was applied to a GlcNAc-agarose column and eluted sequentially with GlcNAc and EDTA. Four (31, 27, 26, and 19 kDa) and one (26 kDa) proteins were detected as bound molecules in the GlcNAc- and the EDTA-eluates, respectively. Among these, the 26 kDa protein from the EDTA eluate was found to be a homologue of mammalian C1q through cDNA analysis. HaC1q had an ability to bind to various microbes in a Ca(2+)-dependent manner and its target ligands on the microbes were lipopolysaccharide, lipoteichoic acid, and peptidoglycan. The binding of HaC1q to GlcNAc-agarose was not inhibited by an excess amount of monosaccharide such as GlcNAc. While HaC1q bound to Sepharose 6B with a matrix of GlcNAc-agarose (polymer of agarobiose), it did not bind to Sepharose 4B that contained lower concentration of agarobiose than Sepharose 6B. Therefore, the target of HaC1q on GlcNAc-agarose was concluded to be agarobiose and high density of the target moiety seemed to be required for the stable binding. This finding was in accordance with the known behavior of other lectins involved in the complement system. We have concluded that HaC1q recognizes agarobiose-like structures present on the surface of microbes and acts as a pattern-recognition molecule in the process for elimination of invading microbes.

Molecular cloning and expression analysis of mannose receptor C type 1 in grass carp (Ctenopharyngodon idella).

Mannose receptor C type 1 (MRC1) is a pattern-recognition receptor (PRR) which plays a significant role in immune responses. Much work on MRC1 has been done in mammals and birds while little in fish. In this study, we cloned and characterized MRC1 in grass carp (gcMR). The full-length gcMR contained 5291bp encoding a putative protein of 1432 amino acids. The predicted amino acid sequences showed that gcMR contained a signal peptide, a cysteine-rich (CR) domain, a fibronectin type II (FN II) domain, eight C-type lectin-like domains (CTLDs), a transmembrane domain and a short cytoplasmic domain. gcMR were constitutively expressed in different organs with the higher expression in spleen and head kidney. During embryonic development, gcMR transcript levels were highest at cleavage stage. The up-regulation expression of gcMR, IL-1ß and TNF-α in liver, spleen, head kidney and intestine after Aeromonas hydrophila infection indicating it involved in innate immune regulation during bacterial infections.

A novel C1q-domain-containing (C1qDC) protein from Mytilus coruscus with the transcriptional analysis against marine pathogens and heavy metals.

The C1q-domain-containing (C1qDC) proteins, which are involved in various processes of vertebrates, are important pattern recognition receptors in innate immunity of invertebrates. In present study, a novel C1qDC was identified from Mytilus coruscus (designated as McC1qDC), which was 917 bp in length encoding 236 amino acids with a typical signal peptide of 19 amino acid residues in N-terminus. Based on its conserved C1q domain and molecular architecture of 10 ß-strand jelly-roll folding topology structure, McC1qDC might be classified as a member of the C1q family. The mRNA transcript of McC1qDC was predominantly detectable in the hemocytes, and a less degree in gill, gonad and mantle, but trace in foot, adductor and digestive gland. Upon induction by Vibrio harveyi and Vibrio alginolyticus, McC1qDC expression was significantly up-regulated. Time-dependent mRNA expression of McC1qDC was found during copper and cadmium exposure for its heavy metal-binding domain. These results indicated that McC1qDC was a novel member of the C1qDC protein family as a pattern recognition receptor against pathogens, and might be developed as a potential indicator for monitoring heavy metals pollution.

Characterization of a pattern recognition molecule vitellogenin from carp (Cyprinus carpio).

Pattern recognition proteins function in innate immune responses by binding to molecules on the surface of invading pathogens and initiating host defense reactions. To explore the role of vitellogenin (Vg) in fish innate immunity, we purified Vg from Carp by gel filtration combined with diethylaminoethyl (DEAE) chromatography. The purified Vg was confirmed by MALDI-TOF mass spectrometry. Antibacterial activity analysis showed that Vg inhibited bacterial activity to Escherichia coli and Staphylococcus aureus in a dose-dependent manner. Vg bound to the surface of Gram-negative and Gram-positive bacteria. It also agglutinated E. coli and S. aureus and weakly to Saccharomyces cerevisiae. Vg showed a strong binding activity to lipopolysaccharides from Gram-negative bacteria. Vg-treated macrophage enhanced phagocytosis to E. coli and S. aureus. Vg also bind with macrophage function as opsonins to promote phagocytosis. The results suggest that Vg serves as a pattern recognition molecule and opsonins in antibacterial defense and as an effector in fish innate immunity.