Mengyao Yu

Characterisation and immunostimulatory activity of an α-(1→6)-d-glucan from the cultured Armillariella tabescens mycelia

IPS-B2, an intracellular polysaccharide with anti-tumor effect, was isolated from cultured Armillariella tabescens mycelia by hot water extraction, anion-exchange and gel chromatography. Based on the results of paper chromatography (PC), gas chromatography (GC), infra-red (IR) spectroscopy and (13)C NMR, IPS-B2 was characterized as an α-(1→6)-d-glucan with a molecular weight of 49.5kDa. The effects of IPS-B2 on murine peritoneal macrophages were further investigated. The results demonstrated that IPS-B2 induced nitric oxide (NO) and cytokines (TNF-α, IL-1β and IL-6) production in macrophages. The outcomes of real-time reverse transcription-polymerase chain reaction (RT-PCR) proved that the transcribing level of inducible NO synthase (iNOS), TNF-α, IL-1β and IL-6 mRNA in the peritoneal macrophages have been augmented by IPS-B2. These data suggest that the anti-tumor activity of the polysaccharide from A. tabescens may due to activation of macrophage.

Molecular Diversity of Auricularia polytricha Revealed by Inter-Simple Sequence Repeat and Sequence-Related Amplified Polymorphism Markers

Due to unsatisfying attempts to fingerprint Auricularia polytricha, two different molecular maker systems—Inter-Simple Sequence Repeats (ISSR) and Sequence-related amplified polymorphism (SRAP)—were established and tested to quantify molecular diversity among 19 strains of this fungus. A total of 202 (99.0%) and 459 (95.9%) polymorphic bands were detected by 13 ISSR primers and 14 SRAP primer combinations, respectively. By parsimony method, a phylogenetic tree was constructed based on each analysis; the two trees show that 19 A. polytricha strains were distributed into five or four groups. These results demonstrated that both methods were suitable for discriminating among strains of A. polytricha, and the novel SRAP markers are more efficient and preferable. The result also indicated the high level of genetic diversity of A. polytricha and their relationship between each other. These findings would benefit future research in A. polytricha, especially in breeding and medicine development. It also gives a useful method for fingerprinting of other fungi.

Antioxidant and immunomodulatory activities of a polysaccharide from Flammulina velutipes

OBJECTIVE To evaluate the antioxidant and immunomodulatory activities of a unique polysaccharide from the medicinal fungus Flammulina velutipes in vitro. METHODS Using water extraction and alcohol precipitation, crude polysaccharides were obtained. After purification by DEAE-cellulose 52 ion exchange chromatography and Sephacryl S-300 HR gel filtration chromatography, High performance liquid chromatography equipped with evaporative light-scattering detector, Infrared radiation and Nuclear magnetic resonance were used to evaluate the structure of the polysaccharide. Its immunomodulatory activity was measured by examining the production of nitric oxide (NO) and cytokine secretion, and via lymphocyte proliferation experiments. Its effects on the scavenging activities of hydroxyl radical, superoxide anion and reducing power were also measured. RESULTS A water-soluble polysaccharide, Flammulina velutipes polysaccharide I-A (FVP I-A), was obtained with a molecular mass of 8.14 x 10(4) Da determined by high performance gel permeation chromatography. An in vitro antioxidant assay indicated that FVP I-A could scavenge hydroxyl radical, superoxide anion and possessed reducing power and could largely promote NO production and augment the interleukin-1β, interleukin-6, and tumor necrosis factor-α secretion by RAW264.7 macrophages (P < 0.05). Moreover, FVP I-A could promote lymphocyte proliferation (P < 0.05), and synergistically enhance the augmentation of the proliferation of mouse lymphocytes by concanavalin A and lipopolysaccharides (P < 0.01, P < 0.05). CONCLUSION The FVP I-A obtained from Flammulina velutipes possessed antioxidant activity and could enhance non-specific and specific immune responses in vitro.

Dehydropachymic acid decreases bafilomycin A1 induced β-Amyloid accumulation in PC12 cells

ETHNOPHARMACOLOGICAL RELEVANCE Fuling, the sclerotium of Poria cocos, was frequently used in traditional Chinese medicine (TCM) formulae for Alzheimers disease (AD) intervention over the past 10 centuries. And its extracts exhibited significant effects in both cellular and animal models of AD in previous studies. However, its mechanisms on prevention and treatment of AD have not been well elucidated yet. AIM OF THE STUDY To investigate the effect and corresponding mechanisms of dehydropachymic acid, which is one of the major triterpenes in P. cocos, on the clearance of β-amyloid accumulation in bafilomycin A1 induced PC12 cells. MATERIALS AND METHODS MTT assay was used to examine the DPA effect on the viability of PC12 cells stable transfected with pCB6-APP (PC12-APP). PC12-APP cells were treated with DPA at the concentration of 6.25, 12.5, 25μg/mL for 4h, and then co-treated with 50nmol/L bafilomycin A1 for 48h except the controls. The Aβ1-42 content in culture medium was determined by ELISA. The intracellular amount of APP, Aβ1-42, LC3, cathepsin D was measured by Western blotting and normalized to GAPDH loading control. The PC12 cells stable transfected with pSelect-LC3-GFP (PC12-LC3-GFP) was used in the fluorescence microscopy estimation of autophagosomes accumulation. The internal pH in lysosome was detected by LysoTracker Red staining. RESULTS DPA had no significant effect on the cell viability but could significantly decrease Aβ1-42 content in culture medium and eliminate the intracellular accumulation of APP and Aβ1-42 in bafilomycin A1 induced PC12-APP cells. Furthermore, DPA lowered the LC3-II/LC3-I ratio and reduced the GFP-labeled LC3 puncta which were elevated by bafilomycin A1. And the increase in internal pH of lysosome and decrease in mCatD amount in Bafilomycin A1 induced PC12-APP cells were restored by DPA treatment. These results indicated that DPA could restore the lysosomal acidification and recover the autophgic flux which is impaired by bafilomycin A1. CONCLUSIONS DPA could effectively clear the accumulation of Aβ1-42 in bafilomycin A1 impaired PC12 cells through restoring the lysosomal acidification and recovering the autophgic flux. And these results highlight its therapeutic potential for AD treatment.

Isolation of a wild Morchella spp. strain and the effects of its extract on ethanol-induced gastric mucosal lesions in rats.

A Morchella spp. strain was isolated from a wild morel mushroom, and the effects of its mycelia extract on the ethanol-induced gastric mucosal lesions of rats were investigated in vivo. Sequence analysis of internal transcribed spacer suggested that this Morchella spp. strain (strain No. M1) was clustered together with M. conica in the phylogenetic tree. The superoxide dismutase (SOD) activity increased significantly compared to the control. However, the malondialdehyde (MDA) level and myeloperoxidase (MPO) activity decreased significantly compared to the control. These results indicated that M1 is one member of M. conica and the protective effects of M1 extract against the ethanol-induced gastric lesions may be related to the increased SOD activity and decreased MDA level and MPO activity in rats.