Search Results (17)

Flammulina filiformis is one of the widely produced edible fungi worldwide. It is rich in γ-aminobutyric acid (GABA), a non-protein amino acid with important physiological functions in humans. To investigate the functions of key genes in the GABA metabolic pathway of F. filiformis, we isolated the monokaryon Fv-HL23-1 from the factory-cultivated F. filiformis strain Fv-HL23 and then sequenced and assembled the genome using the PacBio Sequel and Illumina NovaSeq sequencing platforms. The results showed that the genome comprised 140 scaffolds with a total length of 40.96 Mb, a GC content of 49.62%, an N50 of 917,125 bp, and 14,256 protein-coding genes. Phylogenetic analysis based on the whole genome revealed a close evolutionary relationship of Fv-HL23-1 with Armillaria mellea, Lentinula edodes, and Schizophyllum commune. A total of 589 carbohydrate-active enzymes were identified in the genome of Fv-HL23-1, suggesting its strong lignocellulose degradation ability, and 108 CYP450 gene family members were identified, suggesting important functions such as resistance to stress, secondary metabolite synthesis, and growth and development. The F. filiformis proteins glutamate decarboxylase 1 (Ff-GAD1) and glutamate decarboxylase 2 (Ff-GAD2), which may be responsible for GABA synthesis, were identified by protein alignment. Molecular docking analysis showed that Ff-GAD2 may have better catalytic activity than Ff-GAD1. To verify the function of Ff-gad2, its heterologous expression in the mycelia of the mononuclear Hypsizigus marmoreus was analyzed. Compared with wild type, the GABA content of mycelia was increased by 85.40–283.90%, the growth rate was increased by 9.39 ± 2.35%, and the fresh weight was increased by 18.44 ± 7.57%. Ff-GAD2 may play a catalytic role in GABA synthesis. In addition, the expression of the full-length Ff-gad2 gene was increased by 7.96 ± 1.39 times compared with the exon expression level in H. marmoreus mycelia, suggesting that the intron may contribute to the heterologous expression of Ff-GAD2. Based on whole-genome sequencing, we analyzed the enzyme system related to the important life activities of F. filiformis, focusing on the function of Ff-GAD, a key enzyme in the GABA synthesis pathway. The results lay a foundation for elucidating the GABA metabolism pathway of edible fungi and developing targeted breeding strategies for GABA-producing edible fungi. Full article

In the industrial production of Lentinula edodes and Pleurotus eryngii, slow growth of the mother seed and insufficient hyphal vitality can significantly affect the cultivation process. To shorten the growth period on traditional PDA medium, two strains of L. edodes and P. eryngii were cultured with different proportions of P. tuber-regium and Wolfiporia hoelen sclerotium powders added into the medium to investigate the effect on the mycelial growth. Compared to the PDA, the addition of sclerotia powder significantly enhanced the growth of mycelia, with an optimal addition ratio of 2%. Transcriptome sequencing was performed after culturing L. edodes and P. eryngii on PDA, PDA with 2% P. tuber-regium sclerotium powder, and PDA with 2% W. hoelen sclerotium powder. GO enrichment analysis of the differentially expressed genes (DEGs) of L. edodes and P. eryngii strains cultured in the sclerotia powder media showed significant changes in oxidoreductase and glucosidase activities. Changes were observed in KEGG annotation for carbohydrate metabolism, glycolysis, pyruvate metabolism, and other energy metabolic pathways. Moreover, carbohydrate-active enzyme (CAZyme) family genes were predominantly upregulated. The increase in the activity of CAZyme and oxidoreductases promotes the degradation of nutrients in the sclerotia into small-molecule substances, which explains why the sclerotia powder culture medium promotes mycelial growth. Full article

Spent mushroom substrates (SMSs) and mushroom basal bodies (MBBs) are significant by-products because of their nutrient content even after harvesting. This study aimed to evaluate the effect of these two by-products, derived from Agaricus bisporus (Ab) and Cyclocybe cylindracea (Cc) cultivation, as potential growth and biochemical composition add-value enhancers of edible mushroom mycelia such as Pleurotus ostreatus, C. cylindracea, and Lentinula edodes. Fungal growth substrates enriched with SMS and MBB extracts significantly affected the growth of mushroom mycelia. In particular, on P. ostreatus, the MBBs Ab and Cc extracts determined an increase in mycelial weight by 89.5%. Also, by-products influenced mushrooms’ mycelial texture, which appeared more floccose and abundant in growth. FT-IR analysis showed that L. edodes mycelium, grown on MBB substrates, showed the highest increase in bands associated with proteins and chitin. Results demonstrated that mushroom by-products enhance mycelial growth and confer an enrichment of compounds that could increase mycelial resistance to pathogens and make a nutraceutical improvement. Full article

A standardized extract of cultured Lentinula edodes mycelia (ECLM, AHCC^®) has been shown to have beneficial effects on organ metabolism. ECLM has been indicated to have liver protective properties by suppressing inflammatory responses. The pathogenesis of hepatic ischemia-reperfusion injury is thought to involve the induction of inflammatory mediators. However, whether ECLM affects inflammatory mediators caused by warm hepatic ischemia-reperfusion injury and partial hepatectomy (HIRI+PH) has not been clarified. In this study, we evaluated the protective effects of ECLM against liver damage caused by HIRI+PH. Rats were fed a normal diet (HIRI+PH) or a normal diet with 2% ECLM (HIRI+PH and ECLM) for ten days, then the liver and duodenal ligament were clamped and subjected to 15 min of hepatic ischemia. After 70% hepatectomy, the inflow occlusion was released, and liver and blood samples were collected at 3, 6, and 24 h. The effect of ECLM on mortality induced by 30 min of ischemia and hepatectomy was evaluated. The results showed that ECLM attenuated pathological liver damage, including apoptosis, in the rats treated with HIRI+PH, and decreased serum aminotransferase activity; ECLM decreased mRNA levels of the inflammation-related genes inducible nitric oxide synthase and C-X-C motif chemokine ligand 1, and increased mRNA levels of interleukin 10, an anti-inflammatory cytokine; ECLM increased hepatocyte growth factor mRNA levels and Ki-67 labeled nuclei in the liver at 24 h; ECLM significantly reduced HIRI+PH-induced mortality. In conclusion, ECLM may prevent HIRI+PH-induced liver injury in part by suppressing various inflammatory responses and promoting liver regeneration. Full article

Puberty is a critical developmental period of life characterized by marked physiological changes, including changes in the immune system and gut microbiota development. Exposure to inflammation induced by immune stressors during puberty has been found to stimulate central inflammation and lead to immune disturbance at distant sites from the gut; however, its enduring effects on gut immunity are not well explored. Therefore, in this study, we used a pubertal lipopolysaccharides (LPS)-induced inflammation mouse model to mimic pubertal exposure to inflammation and dysbiosis. We hypothesized that pubertal LPS-induced inflammation may cause long-term dysfunction in gut immunity by enduring dysregulation of inflammatory signaling and epigenetic changes, while prebiotic/probiotic intake may mitigate the gut immune system deregulation later in life. To this end, four-week-old female Balb/c mice were fed prebiotics/probiotics and exposed to LPS in the pubertal window. To better decipher the acute and enduring immunoprotective effects of biotic intake, we addressed the effect of treatment on interleukin (IL)-17 signaling related-cytokines and pathways. In addition, the effect of treatment on gut microbiota and epigenetic alterations, including changes in microRNA (miRNA) expression and DNA methylation, were studied. Our results revealed a significant dysregulation in selected cytokines, proteins, and miRNAs involved in key signaling pathways related to IL-17 production and function, including IL-17A and F, IL-6, IL-1β, transforming growth factor-β (TGF-β), signal transducer and activator of transcription-3 (STAT3), p-STAT3, forkhead box O1 (FOXO1), and miR-145 in the small intestine of adult mice challenged with LPS during puberty. In contrast, dietary interventions mitigated the lasting adverse effects of LPS on gut immune function, partly through epigenetic mechanisms. A DNA methylation analysis demonstrated that enduring changes in gut immunity in adult mice might be linked to differentially methylated genes, including Lpb, Rorc, Runx1, Il17ra, Rac1, Ccl5, and Il10, involved in Th17 cell differentiation and IL-17 production and signaling. In addition, prebiotic administration prevented LPS-induced changes in the gut microbiota in pubertal mice. Together, these results indicate that following a healthy diet rich in prebiotics and probiotics is an optimal strategy for programming immune system function in the critical developmental windows of life and controlling inflammation later in life. Full article

Mycelial cultures of Lentinula edodes, an edible and medicinal mushroom, have been used in our previous research to obtain selenium-containing immunomodulatory preparations. Our current attempts to obtain a new preparation containing both selenium and zinc, two micronutrients necessary for the functioning of the immune system, extended our interest in the simultaneous accumulation of these elements by mycelia growing in media enriched with selenite and zinc(II) ions. Subsequently, we have studied the effects of new L. edodes mycelium water extracts with different concentrations of selenium and zinc on the activation of T cell fraction in human peripheral blood mononuclear cells (PBMCs). Flow cytometry analysis was used to measure the expression of activation markers on human CD4+ and CD8+ T cells stimulated by anti-CD3 and anti-CD3/CD28 antibodies (Abs). It was demonstrated that statistically significant changes were observed for PD-1 and CD25 antigens on CD8+ T cells. The selenium and zinc content in the examined preparations modified the immunomodulatory activity of mycelial polysaccharides; however, the mechanisms of action of various active ingredients in the mycelial extracts seem to be different. Full article

The coronavirus disease 2019 (COVID-19) pandemic has significantly impacted global public health safety and the economy. Multiple antiviral drugs have been developed, and some have received regulatory approval and/or authorization. The use of nutraceuticals can be beneficial for preventing and treating COVID-19 complications. AHCC is a standardized, cultured extract of an edible mushroom Lentinula edodes of the Basidiomycete family of fungi that is enriched in acylated α-1,4-glucans. Here, we evaluated the effects of the oral administration of AHCC on the host response to SARS-CoV-2 infection in two murine models, K18-hACE2 transgenic mice and immunocompetent BALB/c mice. Oral administration of AHCC every other day for one week before and one day post SARS-CoV-2 infection in both strains of mice decreased the viral load and attenuated inflammation in the lungs. AHCC treatment also significantly reduced SARS-CoV-2-induced lethality in the K18-hACE2 mice. AHCC administration enhanced the expansion of γδ T cells in the spleen and lungs before and after viral infection and promoted T helper 1-prone mucosal and systemic T cell responses in both models. In AHCC-fed BALB/c mice, SARS-CoV-2 specific IgG responses were also enhanced. In summary, AHCC supplementation enhances host resistance against mild and severe COVID-19 infection primarily via the promotion of innate and adaptive T cell immune responses in mice. Full article

Mycelial ageing is associated with ROS and autophagy in Lentinula edodes. However, the underlying cellular and molecular mechanisms between ROS and autophagy remain obscure. This study induced autophagy in L. edodes mycelia through exogenous H₂O₂ treatment. Results showed that 100 μM H₂O₂ treatment for 24 h significantly inhibited mycelial growth. H₂O₂ caused the depolarisation of MMP and accumulation of TUNEL-positive nuclei, which was similar to the ageing phenotype of L. edodes mycelia. Transcriptome analysis showed that differentially expressed genes were enriched in the mitophagic, autophagic, and MAPK pathways. LeAtg8 and LeHog1 were selected as hub genes. RNA and protein levels of LeATG8 increased in the H₂O₂-treated mycelia. Using fluorescent labelling, we observed for the first time the classic ring structure of autophagosomes in a mushroom, while 3D imaging suggested that these autophagosomes surrounded the nuclei to degrade them at specific growth stages. Phospho-LeHOG1 protein can translocate from the cytoplasm to the nucleus to regulate mycelial cells, resisting ROS-induced oxidative stress. Furthermore, LeATG8 expression was suppressed when LeHOG1 phosphorylation was inhibited. These results suggest that the LeATG8-dependent autophagy in L. edodes mycelial is closely associated with the activity or even phosphorylation of LeHOG1. Full article

The spawn of Lentinula edodes and other basidiomycete fungi tend to age with long-term culture. This causes heavy yield losses if aging spawn is used for propagation. In this study, we cultivated dikaryotic L. edodes mycelia in plates for 60 days to produce intrinsic aging phenotypes. We found that intracellular reactive oxygen species levels increased in contrast to mitochondrial depolarization and also observed greater DNA fragmentation with longer culture time. Transcriptome analysis of mycelia at different growth stages revealed pronounced expression differences between short- and long-term cultures. In particular, “phenylalanine, tyrosine, and tryptophan biosynthesis”, “mitophagy and autophagy”, “MAPK signaling pathway”, and “ABC transporter” were among the enriched terms in the mycelial aging process. Weighted correlation network analysis identified LeAtg8, LeHog1, LePbs2, and LemTOR as key genes during aging. Western blotting confirmed that LeATG8 and phosphorylated LeHOG1 protein levels were significantly upregulated in aging mycelia. Our combined analytical approach provides insights into the mechanisms that regulate mycelial aging, indicating that autophagy/mitophagy plays a major role in counteracting the effects of age on mycelial growth development. Full article

It has been demonstrated that lignin was efficiently degraded by Lentinula edodes (L. edodes). However, the process of lignin degradation and utilization by L. edodes has not been discussed in detail. Therefore, the effects of lignin on L. edodes mycelium growth, chemical compositions, and phenolic profiles were investigated herein. It has been revealed that 0.10% lignin acted as the most effective concentration to accelerate mycelia growth, which yielded the highest biomass of 5.32 ± 0.07 g/L. Furthermore, a 0.10% concentration of lignin promoted the accumulation of phenolic compounds, especially protocatechuic acid, with peak value of 48.5 ± 1.2 μg/g. In contrast, the higher concentration of lignin (0.20%) exerted an inhibitory effect on the growth of L. edodes. Overall, the application of lignin at the optimal concentration of 0.10% could not only enhance the mycelial growth but also accumulate the phenolic acids and raise the nutritional and medical values of L. edodes. Full article

We reported the development of an effective cancer treatment using a multidisciplinary treatment, including photodynamic therapy (PDT) with indocyanine green (ICG) liposomes and a combination of Lentinula edodes mycelia (LEM) and hydrogen gas inhalation therapy. ICG liposomes were prepared by adding 5 mg of ICG to 50 mL liposomes. Later, 25 mL of ICG liposomes were diluted with 250 mL of 5% glucose solution and administered intravenously to the patient. We selected the multi-laser delivery system (MLDS), a laser irradiator for performing PDT. Further, the patients received a combination of LEM and hydrogen gas inhalation therapy throughout the treatment. We reported two cases of PDT therapy, one with middle intrathoracic esophagus carcinoma and the other with hypopharyngeal cancer. In the first case, the MLDS laser was directly attached to the endoscope and directed to the cancer area with wavelengths of 810 nm. After the treatment, a biopsy demonstrated no tumor recurrence. In the second case, the patient was treated with endovascular PDT using ICG liposomes and MLDS fiber optics. Later, tumor shrinkage was demonstrated after the first round and disappeared after six months. In conclusion, the present findings suggest that the effect of PDT using ICG liposomes with LEM and hydrogen gas may eradicate cancer without burdening patients by enhancing tumor immunity. Full article

The AHCC standardized extract of cultured Lentinula edodes mycelia, and the standardized extract of Asparagus officinalis stem, trademarked as ETAS, are well known supplements with immunomodulatory and anticancer potential. Several reports have described their therapeutic effects, including antioxidant and anticancer activity and improvement of immune response. In this study we aimed at investigating the effects of a combination of AHCC and ETAS on colorectal cancer cells and biopsies from healthy donors to assess the possible use in patients with colorectal cancer. Our results showed that the combination of AHCC and ETAS was synergistic in inducing a significant decrease in cancer cell growth, compared with single agents. Moreover, the combined treatment induced a significant increase in apoptosis, sparing colonocytes from healthy donors, and was able to induce a strong reduction in migration potential, accompanied by a significant modulation of proteins involved in invasiveness. Finally, combined treatment was able to significantly downregulate LGR5 and Notch1 in SW620 cancer stem cell (CSC) colonospheres. Overall, these findings support the potential therapeutic benefits of the AHCC and ETAS combinatorial treatment for patients with colorectal cancer. Full article

The probiotic Bifidus BB536 (BB536), which contains Bifidobacterium longum, has been shown to have enhanced probiotic effects when given together with a standardized extract of cultured Lentinula edodes mycelia (AHCC^®, Amino Up Co. Ltd., Sapporo, Japan). BB536 and AHCC^® may modulate T cell and dendritic cell (DC) phenotypes, and cytokine profiles to favour anti-inflammatory responses following antibiotic ingestion. We tested the hypothesis that orally administered BB536 and/or AHCC^®, results in modulation of immune effector cells with polarisation towards anti-inflammatory responses following antibiotic usage. Forty healthy male volunteers divided into 4 equal groups were randomised to receive either placebo, BB536, AHCC^® or a combination for 12 days in a double-blind manner. After 7 days volunteers also received 250 mg azithromycin for 5 days. Cytokine profiles from purified CD3+ T cells stimulated with PDB-ionomycin were assessed. CD4+ CD25+ forkhead box P3 (Foxp3) expression and peripheral blood DC subsets were assessed prior to treatment and subsequently at 7 and 13 days. There was no difference in cytokine secretion from stimulated CD3+ T cells between treatment groups. Compared with baseline, Foxp3 expression (0.45 ± 0.1 vs. 1.3 ± 0.4; p = 0.002) and interferon-gamma/interleukin-4 (IFN-γ/IL-4) ratios were increased post-treatment in volunteers receiving BB536 (p = 0.031), although differences between groups were not significant. For volunteers receiving combination BB536 and AHCC^®, there was an increase in myeloid dendritic cells (mDC) compared with plasmacytoid DC (pDC) counts (80% vs. 61%; p = 0.006) at post treatment time points. mDC2 phenotypes were more prevalent, compared with baseline, following combination treatment (0.16% vs. 0.05%; p = 0.002). Oral intake of AHCC^® and BB536 may modulate T regulatory and DC phenotypes to favour anti-inflammatory responses following antibiotic usage. Full article

Invasive pulmonary aspergillosis represents one of the most serious fungal infections among immunocompromised patients. In this study, we aimed to analyze the in vivo efficacy of prophylactic oral amphotericin B (AMB) encapsulated in modified chitosan-nanoparticles (Nanomerics’ Molecular Envelope Technology (MET)) supplemented with a standardized extract of cultured Lentinula edodes mycelia (AHCC^®) in a murine model of pulmonary aspergillosis. We determined fungal burden and survival of mice and additionally, we carried out a cytokine analysis in an attempt to understand the immunomodulation of the extract. Our results evidenced equivalent efficacy between orally administered AMB-MET and the intravenous liposomal AMB marketed formulation. Addition of the AHCC^® supplement significantly improved efficacy in terms of burden reduction and survival increase of both oral and intravenous AMB therapies compared to the untreated control group. Moreover, a protective effect of the extract was observed in terms of weight loss. Regarding the cytokine profiles, the Th1 immune response was stimulated in treated animals when compared to the control group. This response was marked by an enhancement in the MCP-1, GM-CSF, VEGF, RANTES and IL-17 levels and a decrease in the IL-6, a biomarker related to the severity of the infection. Full article

Lentinula edodes is the most consumed mushroom in Asia due to its nutritional and medicinal values, and the optimal reference gene is crucial for normalization of its gene expression analysis. Here, the expression stability of 18 candidate reference genes (CRGs) in L. edodes was analyzed by three statistical algorithms (geNorm, NormFinder and BestKeeper) under different stresses (heat, cadmium excess and Trichoderma atroviride infection), different substrates (straw, sawdust and corn stalk) and different development stages (mycelia, primordia and fruit bodies). Among the 18 CRGs, 28S, Actin and α-tub exhibited the highest expression stability in L. edodes under all conditions, while GPD, SPRYP and MSF showed the least stable expression. The best reference gene in different conditions was different. The pairwise variation values showed that two genes would be sufficient for accurate normalization under different conditions of L. edodes. This study will contribute to more accurate estimation of the gene relative expression levels under different conditions using the optimal reference gene in qRT-PCR (quantitative reverse transcription polymerase chain reaction) analysis. Full article