Binding Specificity of a New Artificial ß-Glucan Recognition Protein and Its Application to ß-Glucan Detection in Mushroom Extracts.

ß-1,3-D-glucan (BG) activates innate immunity and enhances immune responses. Fungi, such as mushrooms, produce a relatively large amount of BG, the structure and molecular weight of which varies depending on the species of fungi. This study was conducted to develop a detection probe for quantifying or detecting BG from fungi using BG-binding proteins. The binding properties of a new ß-glucan recognition protein (BGRP) against various BGs were compared. With reference to the amino acid sequences of BGRP in insects, an artificial BGRP (supBGRP) was designed with higher production efficiency using gene recombination technology. SupBGRP was produced in Escherichia coli with high efficiency, and its reactivity with BG from fungi was the highest among the BG-binding proteins examined. SupBGRP exhibited high reactivity with 1,6-branched BG and will be useful for the quantification and detection of fungal BG.

Open-Label Study of the Influence of Food Containing the Royal Sun Mushroom, Agaricus brasiliensis KA21 (Agaricomycetes), on ß-Glucan-Specific Antibody Production in Healthy Human Volunteers.

The edible mushroom Agaricus brasiliensis contains a large amount ß-glucan, which is mainly composed of a ß-1,6-glucan structure. In this study, we investigated the effect of A. brasiliensis strain KA21 on the anti-ß-glucan antibody titer in healthy humans and the role of antibodies as an immunomodulator. Twenty-two healthy volunteers were fed the dried fruiting body of A. brasiliensis (900 or 1500 mg/day) for 12 weeks. The anti-ß-glucan antibody titer in the serum was determined by enzyme-linked immunosorbent assay. Immunoglobulin G (IgG) against ß-glucan was significantly upregulated after intake of A. brasiliensis. Murine experiments demonstrated improvement of anti-ß-glucan antibody production after intraperitoneal injection of Agaricus-derived ß-glucan. To understand the role of antibody against ß-glucan in exclusion of pathogenic fungi, we examined the interaction between HL-60 cells and antibody-treated heat-killed Candida albicans. Flow cytometry analysis indicated the upregulation of Candida-positive HL-60 cells after treatment with human IgG, whereas the competitive assay demonstrated that the main epitope of Candida-reacted IgG was the ß-1,6-glucan structure. Binding between HL-60 and IgG-opsonized C. albicans was suppressed by anti-Fcγ receptor 1 (FcγRI) neutralizing antibody. Finally, using FcγRI-expressed cells with the nuclear factor of activated T-cell reporter assay, we demonstrated that higher titers of anti-ß-glucan IgG can induce stronger Fc receptor-mediated cell activation through the formation of an antibody-ß-glucan complex. In conclusion, oral ingestion of A. brasiliensis KA21 promotes anti-ß-glucan antibody production and may contribute to preventing fungal infection through the activation of immune cells by forming antibody-ß-glucan complexes via an FcγR-dependent pathway.

Outdoor-Cultivated Royal Sun Medicinal Mushroom Agaricus brasiliensis KA21 (Agaricomycetes) Reduces Anticancer Medicine Side Effects.

We investigated whether outdoor-cultivated Agaricus brasiliensis (KA21) could reduce the side effects caused by the anticancer medicine 5-fluorouracil (5-FU). The adverse effects of 5-FU were analyzed in mice by orally administering the drug every day for 5 days. Leukopenia, diarrhea, body weight loss, anorexia, kidney injury, gastrointestinal tract injury, and hair loss were evaluated as side effects. We determined whether these side effects were reduced by the intake of outdoor-cultivated A. brasiliensis. The side effects were reduced in mice receiving the outdoor-cultivated A. brasiliensis but not in those receiving the indoor-cultivated A. brasiliensis. These results suggest that outdoor-cultivated A. brasiliensis is beneficial in reducing the side effects of the anticancer medicine and might, therefore, be useful in improving the quality of life of patients with cancer. Furthermore, because antioxidants have been reported to contribute to the suppression of the side effects of anticancer medicines, the antioxidant activities of different mushrooms were determined. The antioxidant activity of the outdoor-cultivated A. brasiliensis was the highest among all the tested mushrooms. These findings make it clear that the cultivation conditions of A. brasiliensis are important in suppressing the side effects of anticancer medicines. Strong antioxidant activity might be one of the mechanisms through which this pharmacological activity is mediated.

Immunoreactivity of the Cold Water Extract of Royal Sun Culinary-Medicinal Mushroom, Agaricus brasiliensis Strain KA21 (Agaricomycetes), Assessed by Immunoglobulin Preparations for Intravenous Injection.

The royal sun medicinal mushroom, Agaricus brasiliensis, is a health food material that helps to improve quality of life. A. brasiliensis has long been used as a tea by extraction with cold and hot water. Our group has been investigating the immunopharmacological activities of the A. brasiliensis KA21 strain, which is cultivated outdoors. We prepared cold water (AgCWE) and hot water (AgHWE) extracts of this strain. AgCWE contained a larger proportion of proteins, including enzymes, and showed a brownish color during the extraction process. By contrast, chemical and immunochemical analyses revealed that AgHWE contained large amounts of ß-1,3-/1,6-glucans. In an attempt to elucidate the immunochemical characteristics of AgCWE, reactivities to immunoglobulin (Ig) preparations for intravenous injection were analyzed and compared with standard materials. To characterize brownish high-molecular weight components, standard phenol compounds such as caffeic acid (CA), trans-ferulic acid (FA), and coumaric acid (CouA) were polymerized to brownish polymerized polyphenols (PPPs) (i.e., polymerized CA, polymerized FA, and polymerized CouA) by laccase or peroxidase. The results obtained revealed that intravenous Ig reacted with all PPPs and PPPs cross-reacted with AgCWE and AgHWE. The isotype of the anti-PPP antibody was found to be IgG1, in contrast to that of the ß-glucan antibody, which was mainly IgG2. These results strongly suggest that A. brasiliensis extracts contain immunoreactive components against various classes of Igs.

Open-Label Study of the Influence of Food Containing the Royal Sun Mushroom, Agaricus brasiliensis KA21 (Higher Basidiomycetes), on the Quality of Life of Healthy Human Volunteers.

We conducted an open-label study in which food containing Agaricus brasiliensis KA21 was consumed continuously for 12 weeks. A questionnaire for subjective evaluation of the efficacy of this food (hereafter, subjective evaluation questionnaire) revealed significant improvements compared with before its intake; there were improvements in the scores of the amounts of hair loss and gray hair, fatigue and general malaise, eye strain, shoulder stiffness, coldness of extremities, difficulty staying awake during the day, and ease of getting out of bed. These findings suggest that intake of food containing A. brasiliensis KA21 results in the above-mentioned subjectively evaluated improvements, and the possibility that A. brasiliensis KA21 improves the body's immunity. Moreover, no issues regarding the safety of the test food were found.

Differences in antioxidant activities of outdoor- and indoor-cultivated Agaricus brasiliensis, and protective effects against carbon tetrachloride-induced acute hepatic injury in mice.

BACKGROUND: Agaricus brasiliensis (A. brasiliensis) is a medicinal mushroom that exerts various pharmacological actions. We previously demonstrated that different cultivation conditions altered the activity of the polyphenol-related enzymes from this mushroom. However, the influence of cultivation conditions on the antioxidant activity of the fruiting bodies remains unclear. Therefore, in this study we compared the antioxidative effects of fruiting bodies of A. brasiliensis cultivated outdoors and indoors. In addition, we assessed whether different cultivation methods affected the hepatoprotective effects against CCl4-induced liver injury. METHODS: We assessed the antioxidative effects of mushrooms cultivated in open-air or indoors using the DPPH radical-scavenging assay. Furthermore, we prepared experimental feeds containing outdoor- or indoor-cultivated A. brasiliensis. Acute liver injury was induced by CCl4 injection in mice that consumed feed containing outdoor- or indoor-cultivated A. brasiliensis. The hepatoprotective effects of these mushrooms were then evaluated by monitoring the reduction in the circulating levels of alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase. The significance of the differences between the means was assessed using Student's t-test. Finally, histopathological analysis of liver was performed. RESULTS: In the DPPH assay, the antioxidant activity of outdoor-cultivated A. brasiliensis was higher than that of indoor-cultivated mushroom. Moreover, in the mouse model of CCl4-induced hepatitis, the oral administration of outdoor-cultivated A. brasiliensis reduced liver damage significantly, but indoor-cultivated mushrooms failed to inhibit hepatitis. The hepatoprotective effects of outdoor-cultivated A. brasiliensis were observed even when ingestion commenced only 1 day before CCl4 injection, and these effects were not affected by excessive heat treatment. CONCLUSIONS: Outdoor cultivation significantly enhanced the antioxidative activity of A. brasiliensis fruiting bodies. In addition, outdoor-cultivated A. brasiliensis was more effective at protecting against CCl4-induced liver injury in mice than mushrooms grown in a greenhouse.