Extraction and characterization of polysaccharide from fermented mycelia of Coriolus versicolor and its efficacy for treating nonalcoholic fatty liver disease.

Coriolus versicolor, a popular traditional Chinese medicinal herb, is widely used in China to treat spleen and liver diseases; however, the beneficial effects of C. versicolor polysaccharides (CVPs) on nonalcoholic fatty liver disease (NAFLD) remain elusive. Herein we isolated and purified a novel CVP (molecular weight, 17,478 Da) from fermented mycelium powder. This CVP was composed of mannose, galacturonic acid, glucose, galactose, xylose, and fucose at a molar ratio of 22:1:8:15:10:3. Methylation, gas chromatography-mass spectrometry, and nuclear magnetic resonance analyses indicated that the CVP backbone consisted of →1)-ß-D-Man-(6,4→1)-α-D-Gal-(3→1)-α-D-Man-(4→1)-α-D-Gal-(6→, with branches of →1)-α-D-Glc-(6→1)-α-D-Man-(4,3→1)-ß-D-Xyl-(2→1)-ß-D-Glc on the O-6 position of →1)-ß-D-Man-(6,4→ of the main chain. The secondary branches linked to the O-4 position of →1)-α-D-Man-(4,3→ with the chain of →1)-α-D-Fuc-(4→1)-α-D-Man. Further, CVP treatment alleviated the symptoms of NAFLD in an HFD-induced mice model. CVP altered gut microbiota, predominantly suppressing microbes associated with bile acids both in the serum and cecal contents. In vitro data showed that CVP reduced HFD-induced hyperlipidemia via farnesoid X receptor. Our results improve our understanding of the mechanisms underlying the cholesterol- and lipid-lowering effects of CVP and indicate that CVP is a promising candidate for NAFLD therapy.

Synthesis and structural characterization of xylan acetate ester and its antinephritic effects in rats with experimental chronic kidney disease.

Acetic acid has been shown to be effective in chronic kidney disease (CKD). However, it is a low-molecular-weight compound that allows it to be absorbed in the upper digestive tract so that it cannot function in colon. To overcome these deficiencies, an acetate-releasing xylan derivative, xylan acetate ester (XylA), was synthesized and selected in this study for its potential in the treatment of CKD. IR, NMR and HPGPC were used to characterize the structure of XylA and its antinephritic effects was evaluated in vivo. The results showed that acetate was successfully grafted onto the C-2 and C-3 positions of xylan and with a molecular weight at 69157 Da. XylA treatments could relieve the symptoms of CKD in an adenine-induced chronic renal failure (CRF) model and an adriamycin-induced focal segmental glomerulosclerosis (FSGS) model in SD rats. Further study indicated that XylA could upregulate the short-chain fatty acids (SCFAs) in vitro and vivo. Nevertheless, the relative abundance of Phascolarctobacterium in colon was increased after XylA treatment. XylA could upregulate G-protein-coupled receptor 41 (GPR41) expression, inhibit glomerular cell apoptosis and promoting proliferation. Our study expands the application of xylan and provides a new idea for the treatment of CKD with acetic acid.

Heparosan-based self-assembled nanocarrier for zinc(II) phthalocyanine for use in photodynamic cancer therapy.

Zinc(II) phthalocyanine (ZnPc) is a promising photosensitizer in photodynamic therapy (PDT) for melanoma treatment. However, the poor solubility of ZnPc limits its application. To overcome this limitation, heparosan (HP)-based nanoparticles were prepared by anchoring the l-lysine-linked α-linolenic acid branch to the carboxylic acid group to produce amphiphilic conjugates named heparosan with an l-lysine-linked α-linolenic acid branch (HLA). HLA conjugates could self-assemble into spherical nanoparticles in aqueous media and encapsulate ZnPc to form HLA-ZnPc nanoparticles. The cellular uptake of ZnPc could be improved by HLA carriers. These nanoparticles presented excellent photodynamic-mediated toxicity against mouse melanoma cells (B16) by markedly upregulating the intracellular reactive oxygen species (ROS) levels while showing no cytotoxicity to either B16 or normal cells (L02 and HK-2 cells) in the dark. Furthermore, HLA-ZnPc displayed excellent stability in both powder and Roswell Park Memorial Institute (RPMI) 1640 medium, indicating its promise for application in drug delivery and PDT. These results revealed a strategy for HP-based enhancement of ZnPc in PDT efficacy.

Novel amikacin resistance genes identified from human gut microbiota by functional metagenomics.

AIMS: The aim of this study was to evaluate the diversity and potential for horizontal transfer of amikacin resistance genes from the human gut. METHODS AND RESULTS: A library of human faecal microbiota was constructed and subjected to functional screening for amikacin resistance. In total, five amikacin resistance genes that conferred relatively high amikacin resistance, with minimum inhibitory concentrations (MICs) ranging from 64 to >512, were identified from the library, including a novel aminoglycoside acetyltransferase gene and a 16S rRNA methyltransferase (MTase) gene, labelled aac (6')-Iao and rmtI, respectively. AAC(6')-Iao showed the highest identity of 48% to AAC(6')-Ian from a clinical isolate Serratia marcescens, whereas RmtI shared the closest amino acid identity of 32% with ArmA from Klebsiella pneumonia. The MICs of these five subclones to six commonly used aminoglycosides were determined. Susceptibility analysis indicated that RmtI was associated with high resistance phenotype to 4,6-disubstituted 2-DOS aminoglycosides, whereas AAC(6')-Iao conferred resistance to amikacin and kanamycin. In addition, kinetic parameters of AAC(6')-Iao were determined, suggesting a strong catalytic effect on amikacin and kanamycin. CONCLUSIONS: Antibiotic resistance genes with low identity to known sequences can be uncovered by functional metagenomics. In addition, the diversity and prevalence of amikacin resistance genes merit further investigation in extended habitats, especially the 16S rRNA MTase gene that might have been underestimated in previous cognition. SIGNIFICANCE AND IMPACT OF STUDY: Two novel amikacin resistance genes were identified in this study, including a 16S rRNA methyltransferase gene rmtI and an aminoglycoside acetyltransferase gene aac(6')-Iao. This work would contribute to the in-depth study of the diversity and horizontal transfer potential of amikacin resistance genes in the microbiome of the human gut.

Preparation and characterization of 2-deacetyl-3-O-sulfo-heparosan and its antitumor effects via the fibroblast growth factor receptor pathway.

Heparosan, with a linear chain of disaccharide repeating units of â†’ 4) ß-D-glucuronic acid (GlcA) (1 â†’ 4)-α-D-N-acetylglucosamine (GlcNAc) (1→, is a potential starting chemical for heparin synthesis. However, the chemoenzymatic synthesis of single-site sulfated heparosan and its antitumor activity have not been studied. In this study, 2-deacetyl-3-O-sulfo-heparosan (DSH) was prepared successively by the N-deacetylation chemical reaction and enzymatic modification of human 3-O-sulfotransferase-1 (3-OST-1). Structural characterization of DSH was shown the success of the sulfation with the sulfation degree of 0.87. High performance gel permeation chromatography (HPGPC) analysis revealed that DSH had only one symmetrical sharp peak with a molecular weight of 9.6334 × 104 Da. Biological function studies showed that DSH could inhibit tumor cell (A549, HepG2 and HCT116) viability and induce the apoptosis of A549 cells. Further in vitro mechanistic studies showed that DSH may induce apoptosis via the JNK signaling pathway, and the upstream signal of this process may be fibroblast growth factor receptors. These results indicated that DSH could be developed as one of a potential chemical for tumor treatment.

Isolation, structural characterization of polysaccharide from Cephalosporium sinensis mycelia and its anti-nephritic effects in adenine-induced CKD rats.

In this study, a new polysaccharide (CSMP, Mw = 16,685 Da) was isolated and purified from Cephalosporium sinensis mycelia. Monosaccharide composition analysis indicated that CSMP consists of mannose, glucose and galactose. A detailed structural analysis revealed that CSMP has a backbone consisting of →2,6)-ß-D-Manp-(1→ and →3,6)-ß-D-Manp-(1→, as well as two branched chains including of α-D-Manp-(1→6)-α-D-Glcp-(1→ and α-D-Glcp-(1→4)-α-D-Glcp-(1→3)-ß-D-Galp-(1→2)-ß-D-Manp-(1→ attached to C6 of →2,6)-ß-D-Manp-(1→ and →3,6)-ß-D-Manp-(1→. Orally administrated CSMP showed renal protection function in adenine-induced chronic kidney disease (CKD) rats. Further analysis demonstrated that CSMP increased relative abundance of the genera Lactobacillus group, Clostridium coccoides group and Bifidobacterium, and decreased Echerichia subgroup. CSMP increased acetate, propionate and butyrate levels both in colon and cecum. The mechanisms behind these effects could be related to the down-regulation nuclear factor kappa-B (NF-κB) level by up-regulating expression of G protein-coupled receptor 41 (GPR41) and improvement regulatory T cells (Tregs) ratio by inhibiting histone deacetylase (HDAC) activity. These results indicated that CSMP could be developed as one of the potential drugs in the treatment of CKD.

An orally administered butyrate-releasing xylan derivative reduces inflammation in dextran sulphate sodium-induced murine colitis.

Butyrate has been shown to be effective in ulcerative colitis (UC). However, its oral administration is rare due to its rancid odour and unpleasant taste. In this study, the effect of a butyrate-releasing polysaccharide derivative, xylan butyrate ester (XylB), was evaluated in a dextran sodium sulphate (DSS)-induced UC model in C57BL/6 mice. Linear xylan was extracted from corn cobs. The C-2 and C-3 positions of the linear xylan were esterified with butyrate, forming XylB. The protective and therapeutic effects of XylB against UC were determined in a DSS-induced mouse model. The results showed that XylB treatments reversed the imbalance between pro- and anti-inflammatory cytokines. Moreover, XylB rebalanced the gut microbiota that interfered with DSS treatment and significantly decreased the relative abundance of the genera Oscillibacter, Ruminococcaceae UCG-009, Erysipelatoclostridium, and Defluviitaleaceae UCG-01. XylB increased butyrate content in the colon, upregulated G-protein coupled receptor 109A protein expression, inhibited histone deacetylase (HDAC) activity, and exerted anti-inflammatory activity through autophagy pathway activation and nuclear factor-κB (NF-κB) inhibition. XylB reduces inflammatory intestinal damage in mice, suggesting that it would be a potential drug for the treatment of UC and could be used to overcome the limitations of the oral administration of sodium butyrate.

Structural characterization and anti-inflammatory effect in hepatocytes of a galactoglucan from Antrodia camphorata mycelium.

The galactoglucan ACP2 was isolated from cultured Antrodia camphorata mycelium through anion-exchange column chromatography and Sephadex G-100 chromatography and shown to exhibit hepatoprotective function in L02 cells. Based on monosaccharide composition analysis, ACP2 was mainly composed of glucose, galactose, and 6-deoxyglucose in a molar ratio of 5 : 2 : 1. The average molecular weight of ACP2 was 1.93 × 104 Da. The primary structure of ACP2 was elucidated with Fourier-transform infrared spectroscopy, gas chromatography-mass spectrometry, and nuclear magnetic resonance spectroscopy. The results indicated the following composition: →6)-linked-ß-d-Galp-(1→, →6)-linked-α-d-Glcp-(1→, →3)-linked-α-d-Glcp-(1→, and →2,4)-linked-ß-d-Glcp-(1→, with terminal 6-deoxy-α-d-Glcp and α-d-Glcp. ACP2 alleviated lipopolysaccharide-induced hepatocyte inflammation by down-regulating the expressions of COX-2, IL-1ß, TNF-α and IL-6. The decreased expressions of TLR4, MyD88, NF-κB, and phosphorylated p38 in ACP2-treated L02 cells indicated that ACP2 might ameliorate inflammation through the TLR4 and p38/NF-κB signaling pathways.

Isolation, purification, structural characterization and immunostimulatory activity of water-soluble polysaccharides from Lepidium meyenii.

A water-soluble polysaccharide LMP-1 was isolated and purified by ion-exchange chromatography from maca (Lepidium meyenii Walp.). LMP-1 has a molecular weight of 1.01 × 104 Da, and is composed of glucose and arabinose with a molar ratio of 7.03:1.08. Methylation and the 1D and 2D NMR spectroscopy of LMP-1 revealed that it is mainly composed of →4)-α-D-Glcp-(1→, →6)-α-D-Glcp-(1→, →3)-α-D-Glcp-(1→, and ß-D-Araf-(1→, with branching at O-6 of →4,6)-α-D-Glcp-(1 â†’ . LMP-1 showed up-regulation of Toll-like receptor 4 (TLR4) and Toll-like receptor 2 (TLR2). The upstream proteins of Toll-like receptors (TLRs) (CD14 and MD2) and mRNA level of IL-1ß also increased. Increased transcription factor nuclear factor-kappa B (NF-κB) p65 was found in the nuclei and cytoplasm in LMP-1-treated RAW264.7 macrophages. These results indicated that LMP-1 activated RAW264.7 macrophages and elicited immunostimulatory activities via the TLRs/NF-κB signalling pathway.

ß-d-glucan from Antrodia Camphorata ameliorates LPS-induced inflammation and ROS production in human hepatocytes.

A water soluble Antrodia Camphorata polysaccharide was composed of ß-d-glucan, coded as ACP1. ACP1 was derived from cultured Antrodia Camphorata powder, which was extracted, separated and purified by DEAE-52 column and Sephadex G-100 chromatography. The molecular weight of ACP1 was 1.72×104Da, and ACP1 consisted of glucose and galactose in the mole ratio of 2.21:1. Structural identification by FTIR, GC-MS and NMR, inferred that ACP1 consisted of →1,3)-linked-ß-d-Glcp-(6→, →3)-linked-ß-d-Glcp-(1→, and →6)-linked-α-d-Galp-(1→, with terminal ß-d-Glcp and ß-d-Galp. The anti-inflammation and anti-oxidation effects of ACP1 were examined in LPS-treated L02 cells in this study. The results demonstrated that ACP1 not only attenuated ROS production, but also decreased expressions of TNF-α and IL-1ß. Furthermore, ACP1 reduced the proteins expression of NOX1, NOX2 and NOX4, as well as phosphorylation of ERK, p38 and Akt. The results suggested ACP1 was very likely to alleviate oxidation and inflammation via the suppression of NADPH oxidase activation and inhibition of ERK, p38 and Akt signaling pathways.

Effects of cultured Cordyceps mycelia polysaccharide A on tumor neurosis factor-α induced hepatocyte injury with mitochondrial abnormality.

Cordyceps sinensis mycelia polysaccharide A (CPS-A), was isolated from cultured Cordyceps mycelia by 65% alcohol extraction and ion-exchange column chromatography. The molecular weight of CPS-A was 1.2×104Da and the backbone was mainly composed of (1→2)-linked ß-d-mannopyranose, (1→2,4)-linked ß-d-mannopyranose and (1→4)-linked α-d-glucopyranose with terminal ß-d-mannopyranose and α-d-glucopyranose residues. CPS-A played a protective role against TNF-α induced mitochondria injury in L02 cells via up-regulation of mitofusin 2, peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), and membrane potential. CPS-A also played a protective role against TNF-α induced L02 cells apoptosis via up-regulation of Bcl-2 and down-regulation of Bid, Bax, cleaved caspase-3, cleaved caspase-9 and ROS production. Moreover, CPS-A attenuated both the normal expression and overexpression of TNF-α receptor 1 (TNFR1) induced by TNF-α administration. In conclusion, CPS-A was involved in TNF-α induced mitochondria abnormality via TNFR1/ROS/Mfn2 pathway.