Ascofuranone prevents ER stress-induced insulin resistance via activation of AMP-activated protein kinase in L6 myotube cells.

The current study presents that ascofuranone isolated from a phytopathogenic fungus, Ascochyta viciae, has antitumor activity against various transplantable tumors and a considerable hypolipidemic activity. AMP-activated protein kinase (AMPK) plays a critical role in cellular glucose and lipid homeostasis. We found that ascofuranone improves ER stress-induced insulin resistance by activating AMPK through the LKB1 pathway. In L6 myotube cells, ascofuranone treatment increased the phosphorylation of the Thr-172 residue of the AMPK alpha subunit and the Ser-79 subunit of acetyl-CoA carboxylase (ACC) and cellular glucose uptake. Ascofuranone-induced phosphorylation of AMPK and ACC was not increased in A549 cells lacking LKB1. Interestingly, ascofuranone treatment also improved insulin signaling impaired by ER stress in L6 myotube cells. These effects were all reversed by pretreatment with Compound C, an AMPK inhibitor or with adenoviral-mediated dominant-negative AMPK alpha 2. Taken together, these results indicated that ascofuranone-mediated enhancement of glucose uptake and reduction of impaired insulin sensitivity in L6 cells is predominantly accomplished by activating AMPK, thereby mediating beneficial effects in type 2 diabetes and insulin resistance.

White button mushroom (Agaricus bisporus) lowers blood glucose and cholesterol levels in diabetic and hypercholesterolemic rats.

Agaricus bisporus (white button mushroom; WBM) contains high levels of dietary fibers and antioxidants including vitamin C, D, and B(12); folates; and polyphenols that may provide beneficial effects on cardiovascular and diabetic diseases. The objective of this study was to examine the hypothesis that intake of the fruiting bodies of WBM regulates anticholesterolemic and antiglycemic responses in rats fed a hypercholesterolemic diet (0.5% cholesterol; 14% fat) and rats with type 2 diabetes induced by injection of streptozotocin (STZ) (50 mg/kg body weight), respectively. The STZ-induced diabetic male Sprague-Dawley rats fed the Agaricus bisporus powder (ABP; 200 mg/kg of body weight) for 3 weeks had significantly reduced plasma glucose and triglyceride (TG) concentrations (24.7% and 39.1%, respectively), liver enzyme activities, alanine aminotransferase and aspartate aminotransferase (11.7% and 15.7%, respectively), and liver weight gain (P < .05). In hypercholesterolemic rats, oral feeding of ABP for 4 weeks resulted in a significant decrease in plasma total cholesterol (TC) and low-density lipoprotein (LDL) (22.8% and 33.1%, respectively) (P < .05). A similar significant decrease in hepatic cholesterol and TG concentrations was observed (36.2% and 20.8%, respectively) (P < .05). Decrease in TC, LDL, and TG concentrations was accompanied by a significant increase in plasma high-density lipoprotein concentrations. It was concluded that A bisporus mushroom had both hypoglycemic and hypolipidemic activity in rats.

Optimal Culture Conditions for Mycelial Growth and Exo-polymer Production of Ganoderma applanatum.

The effect of fermentation parameters and medium composition on the simultaneous mycelial growth and exo-polymer production from submerged cultures of Ganoderma applanatum was investigated in shake-flask cultures. The optimum initial pH for mycelial growth and exo-polymer production was 5.0 and 6.0, respectively. The optimum temperature was 25℃ and the optimum inoculum content was 3.0% (v/v). The optimal carbon and nitrogen sources were glucose and corn steep powder, respectively. After 12 days fermentation under these conditions, the highest mycelial growth was 18.0 g/l and the highest exo-polymer production was 3.9 g/l.

Antitumor Effects and Immunomodulating Activities of Phellinus linteus Extract in a CT-26 Cell-Injected Colon Cancer Mouse Model.

The antitumor effects of Phellinus linteus extract (Keumsa Linteusan) were investigated in a CT-26 cell-injected colon cancer mouse model. When administered orally (250~1,000 mg/kg body weight), Keumsa Linteusan significantly inhibited the growth of solid colon cancer. The highest dose was highly effective, reducing tumor formation by 26% compared with the control group. The anticomplementary activity of Keumsa Linteusan increased in a dose-dependent manner. Lysosomal enzyme activity of macrophages was increased by 2-fold (100 µg/ml) compared with the control group. Keumsa Linteusan can be regarded as a potent enhancer of the innate immune response, and can be considered as a very promising candidate for antitumor action.

Immunomodulating activities of water-soluble exopolysaccharides obtained from submerged culture of Lentinus lepideus.

Immunomodulating activities of water-soluble exopolysaccharides (LL-EX) obtained from submerged mycelial culture of Lentinus lepideus were studied and their effectiveness was compared with lipopolysaccharide (LPS). The influence of the LL-EX on macrophage cellular lysosomal enzyme activity was to stimulate up to 267%, 392%, and 464% at the level of 10, 50, and 100 microg/ml, respectively. When the LL-EX was further fractionated into LL-Fr.I and Fr.II by Sepharose CL-6B gel chromatography, the cellular lysosomal enzyme activity of LL-Fr.II (2.1- fold) was higher than Fr.I (1.2-fold). Moreover, both LL-Fr.I and Fr.II stimulated the cytokines IL-1beta, TNF-alpha, and IL-6 in macrophages. In mixed lymphocyte reaction, LL-Fr.I and Fr.II enhanced the splenocyte proliferation up to 1.2-fold and 1.4-fold (50 microg/ml), respectively, stimulating only T lymphocytes. The fractions of LL-EX not show any direct toxicity against human gastric adenocarcinoma cell (AGS). The molecular masses of LL-Fr.I and Fr.II were estimated to be about 1,986 kDa and 21 kDa, respectively. The total sugar and protein contents of the two fractions were 84.97% and 69.88% and 15.03% and 30.12%, respectively. The sugar and amino acid compositions of the LL-Fr.I and Fr.II were also analyzed in detail.

Ganoderma applanatum: a promising mushroom for antitumor and immunomodulating activity.

The antitumor effect of exo-biopolymer (EXP) produced by Ganoderma applanatum was investigated using sarcoma-180 bearing mice. EXP, when administered (10-80 mg/kg body weight: BW) intraperitoneally, significantly inhibited the growth of solid tumor and increased the natural killer (NK) cell activity. A dose of 40 mg/kg BW was found to be highly effective, as it reduced the tumor formation by 39.7%, and increased the NK cell activity of splenocytes by 51.6% compared with the control group. The complement activity of EXP was increased in accordance with an increase in concentration. The phosphatase activity of macrophages was increased by 0.7-fold (200 microg/mL) compared with the control group. This EXP contained 58.9% carbohydrate and 17.1% protein. The major sugar of EXP was composed of mannose and glucose, while the protein mainly consisted of serine, glycine and aspartic acid.

Isodihydrocapsiate stimulates plasma glucose uptake by activation of AMP-activated protein kinase.

AMP-activated protein kinase (AMPK) is an energy-sensing enzyme that is implicated as a key factor in controlling whole body homeostasis, including fatty acid oxidation and glucose uptake. We report that a synthetic structural isomer of dihydrocapsiate, isodihydrocapsiate (8-methylnonanoic acid 3-hydroxy-4-methoxy benzyl ester) improves type 2 diabetes by activating AMPK through the LKB1 pathway. In L6 myotube cells, phosphorylation of AMPK and acetyl-CoA carboxylase (ACC) and glucose uptake were significantly increased, whereas these effects were attenuated by an AMPK inhibitor, compound C. In addition, increased phosphorylation of AMPK and ACC by isodihydrocapsiate was significantly reduced by radicicol, an LKB1 destabilizer, suggesting that increased glucose uptake in L6 cells with isodihydrocapsiate treatment is predominantly accomplished by a LKB1-mediated AMPK activation pathway. Oral administration of isodihydrocapsiate to diabetic (db/db) mice reduced blood glucose levels by 40% after a 4-week treatment period. Our results support the development of isodihydrocapsiate as a potential therapeutic agent to target AMPK in type 2 diabetes.

Anti-tumor Effects of Exo- and Endo-biopolymers Produced from Submerged Cultures of Three Different Mushrooms.

The anti-tumor effects of exo- (EX) and endo-biopolymers (EN) produced from submerged mycelial cultures of Ganoderma applanatum (GA), Collybia confluens (CC), and Pleurotus eryngii (PE) were studied using Sarcoma 180 bearing mice. Solid tumor growth was inhibited most effectively when 40 mg/kg body weight (BW) of GA-EX or PE-EN was administered to the intraperitoneal (i.p.) cavity of BALB/c mice. The spleen and liver indexes were increased in mice following i.p. administration of GA-EX and PE-EN fractions. GA-EX and PE-EN reduced the tumor formation by 30.7% and 29.4%, respectively. GA-EX and PE-EN increased the natural killer (NK) cell activity of splenocytes by 41.3% and 28.9%, respectively.

Hypoglycemic Effects of Exo-biopolymers Produced by Five Different Medicinal Mushrooms in STZ-induced Diabetic Rats.

Hypoglycemic effects of exo-biopolymers (EBP) produced by submerged mycelial cultures of Coriolus versicolor, Cordyceps sinensis, Paecilomyces japonica, Armillariella mellea, and Fomes fomentarius were investigated in streptozotocin (STZ)-induced diabetic rats. The rats from each experimental group were orally administered with EBPs (100 mg/kg BW) daily for 2 weeks. Though the hypoglycemic effect was achieved in all the cases, however, C. versicolor EBP proved as the most potent one. The administration of the C. versicolor EBP substantially reduced (29.9%) the plasma glucose level as compared to the saline administered group (control). It also reduced the plasma total cholesterol (TC), triglyceride (TG), aspartate aminotransferase (AST) and, alanine aminotransferase (ALT) levels by 9.22, 23.83, 16.93, and 27.31%, respectively. The sugar and amino acid compositions of this EBP were also analyzed in detail.

Isolation and characterization of biopolymers extracted from the bark of Acanthopanax sessiliflorus and their anticomplement activity.

The crude biopolymer (AS-S1) and endo-biopolymer (AS-S2) were isolated from the dry stem bark of Acanthopanax sessiliflorus and tested for anti complement activity. The two potent anticomplement biopolymers, AS-1 and AS-2-Fr.I, were isolated by the combination of ion-exchange chromatography and gel filtration methods from the endo-biopolymers (AS-S2). The anticomplement activity of AS-1 (MW 12 kDa) and AS-2-Fr.I (MW 180 kDa) were found to be 84.4% and 100.0%, respectively, at the concentration of 25 microg/ml. Activated pathway of the complement system occurred in both classical and alternative pathways, as evidenced by crossed immunoelectrophoresis (CIEP), where a major pathway was detected to be the classical one. It was found that the anticomplement activities of the periodate oxidized were decreased significantly, but those of pronase digested biopolymers of AS-1 and AS-2-Fr.I were decreased very little. The AS-1 contained 2,4,6-tri-O-methyl-D-glucitol, 2,3,6-tri-O-methyl-D-glucitol, and 2,3,6-tri-O-methyl-D-galacitol, which indicated that AS-1 contained a (1 --> 3), (1 --> 4)-linked glucopyranosyl residue and a (1 --> 4)-linked galactosyl residue. AS-2-Fr.I contained mainly 2,4-di-O-methyl-D-mannitol and 2,3,4-tri-O-methyl-D-galacitol, which contained (1 --> 3), (1 --> 6) linked mannosyl and (1 --> 6) linked galactosyl residues.

Hypoglycemic effects of Ganoderma applanatum and Collybia confluens exo-polymers in streptozotocin-induced diabetic rats.

The hypoglycemic effects of Ganoderma applanatum exo-polymer (GAE) and Collybia confluens exo-polymer (CCE) produced by submerged mycelial cultures in streptozotocin (STZ)-induced diabetic rats were investigated. Hypoglycemic effects were achieved in both the GAE- and CCE-treated groups by administration at a level of 100 mg/kg body weight (BW) daily for 3 weeks. The administration of GAE and CCE substantially reduced the plasma glucose levels by as much as 22.0% and 25.9%, respectively, when compared with the control group. The GAE and CCE also lowered the plasma total cholesterol and triglyceride levels by 20.3% and 22.5%, and by 22.7% and 25.5%, respectively. Furthermore, the activity of alanine transaminase (ALT) and aspartate transaminase (AST) was decreased by 23.2% and 20.7% in the GAE-treated group, and it was also reduced by 28.7% and 23.6% in the CCE-treated group. The results strongly demonstrate the potential of GAE and CCE in combating diabetes in experimental animals.

Chemical characteristics and immuno-modulating activities of exo-biopolymers produced by Grifola frondosa during submerged fermentation process.

The immuno-modulating activities and chemical characteristics of exo-biopolymer (EX-GF) produced by a submerged mycelial culture of Grifola frondosa were studied. The EX-GF was fractionated into EX-GF-Fr.I, II, and III by Sephadex G-100 gel chromatography. Anti-complementary activity of EX-GF-Fr.III was highest (71.1%) among them, and its activation system occurred through both classical and alternative pathways, where the classical pathway found to be major one. Lysosomal enzyme activity and nitric oxide production ability of macrophage were also found to be mediated by EX-GF-Fr.III. The molecular weight of the EX-GF-Fr.I, II, and III was estimated to be about 163, 40, and 2.8 kDa, respectively. Total sugar and protein contents of the three fractions were 80.3, 61.9 and 89.3%, and 17.3, 35.2, and 10.7%, respectively. The sugar and amino acid compositions of the EX-GF-Fr.I, II, and III were also analyzed in detail.

Hypolipidemic Effects of Biopolymers Extracted from Culture Broth, Mycelia, and Fruiting Bodies of Auricularia auricula-judae in Dietary-induced Hyperlipidemic Rats.

Hypolipidemic effect of biopolymers extracted from culture broth (CP), mycelia (MP), and fruiting bodies (FP) of Auricularia auricula-judae was investigated in dietary-induced hyperlipidemic rats. The experimental animals were administrated (100 mg/kg body weight) with different biopolymers, daily for 4 weeks. Hypolipidemic effects were achieved in all the experimental groups, however, FP was proved to be the most potent one. The administration of the FP reduced the plasma triglyceride, total cholesterol, low-density lipoprotein cholesterol, and atherogenic index by 24.3, 28.5, 36.4, and 40.9%, respectively, while increased the high-density lipoprotein cholesterol level (9.0%), when compared to the saline (control) administered group.

Anti-complementary Activities of Exo- and Endo-biopolymer Produced by Submerged Mycelial Culture of Eight Different Mushrooms.

The Elfvingia applanata (EA), Hericium erinaceum (HE),Grifola frondosa (GF), Pholiota nameko (PN), Pleurotus eryngii (PE), Trametes suaveolens (TS), Fomes fomentarius (FF), and Inonotus obliquus (IO) could produce the endo- (EN) and exo-biopolymer (EX) in submerged culture. The highest anti-complementary activity of the EN was exhibited by PN (49.1%), followed by HE (38.6%), TS (37.0%),and FF (33.0%),whereas the high activity of the EX was found with GF (59.8%),followed by HE (36.3%),TS (30.8%),and IO (28.8%). The EN of P. nameko (EN-PN) and EX of G. frondosa (EX-GF) were found to contain 78.6% and 41.2% carbohydrates, while 21.4% and 58.8% protein, respectively. The sugar and amino acid compositions of EN-PN and EX-GF were also analyzed in detail.

Macrophage-stimulating activity of polysaccharides extracted from fruiting bodies of Coriolus versicolor (Turkey Tail Mushroom).

The macrophage-stimulating effect of polysaccharides extracted from Coriolus versicolor (Turkey Tail mushroom) was investigated, and their effectiveness was compared with that of lipopolysaccharide (LPS). The purified polysaccharide (CV-S2-Fr.I) of C. versicolor obtained by Sepharose CL-6B gel chromatography stimulated macrophage lysosomal enzyme activity by 250% at a concentration of 100 microg/mL, which was higher than that of LPS at the same concentration. When CV-S2-Fr.I was used in combination with interferon-gamma, there was a marked cooperative induction of nitric oxide production. However, CV-S2-Fr.I had no effect on nitric oxide production by itself. The proportion of C3-positive macrophages in the CV-S2-Fr.I group increased by 7.2-fold compared with the control group.

Chemical characteristics and immuno-stimulating properties of biopolymers extracted from Acanthopanax sessiliflorus.

During our search for macrophage stimulating compounds from medicinal plants, we isolated biopolymers from Acanthopanax sessiliflorus. Isolated fraction AS-5 showed maximum potential, and stimulated lysosonal enzymatic activity by 230% at 300 microg/ml. The nitric oxide (NO) producing ability of AS-5 100 microg/ml was 58 microM when treated with interferon-gamma and lipopolysaccharide 20 micro/ml. The lymphocyte proliferating effects of isolated biopolymer fractions were also investigated. Highest lymphoproliferative activity (a 2.8-fold enhancement compared to salines treated group was exhibited by AS-3 at 200 micro/ml followed by AS-5 and AS-6. The AS-3 fraction stimulated only T-lymphocytes and had little or no effect on B-lymphocyte proliferation. Partially methylated alditol acetates were prepared to elucidate the glycosyl linkage-compositions of the AS-3 and AS-5 biopolymers, and were analyzed by GC-MS. The AS-3 and AS-5 biopolymer fractions were found to contain 2,3,4-tri-O-methyl-D-glucitol, 2,3,4-tri-O-methyl-D-galacitol 3,4,6-tri-O-methyl-galacitol, 2-O-methyl-arabinitol and 2,4,6-tri-O-methyl-D-glucitol, 2,3,6-tri-O-methyl-D-galacitol linkages, respectively.

Production of an anti-complement exo-polymer produced by Auricularia auricula-judae in submerged culture.

Exo-polymer (EP) was produced at 1.2 g l(-1) in submerged culture of Auricular auricula-judae. Crude EP (AJ-0) has 70% anti-complementary activity (inhibition of total complementary hemolysis 50%; ITCH50). The activating pathway of the complement system occurred through both the classical and alternative pathways, though the major pathway was the classical one. Fractionation of AJ-0 using Sepharose CL-6B gel chromatography gave three major fractions (AJ-Fr-I, II and III) of which the first was the most active. The mycelial growth and EP production of A. auricula-judae were optimal at pH 6, 25 degrees C and pH 5, 25 degrees C, respectively.

Hypolipidemic effect of an Exo-biopolymer produced from a submerged mycelial culture of Hericium erinaceus.

The hypolipidemic effect of an exo-biopolymer produced from a submerged mycelial culture of Hericium erinaceus was investigated in dietary-induced hyperlipidemic rats. Hypolipidemic effects were proportionally increased with the increasing concentration of the exo-biopolymer for oral administration. The exo-biopolymer, at the dose of 200 mg/kg body weight, substantially reduced the plasma total cholesterol (32.9%), LDL cholesterol (45.4%), triglyceride (34.3%), phospholipid (18.9%), atherogenic index (58.7%), and hepatic HMG-CoA reductase activity (20.2%). It increased the plasma HDL cholesterol level (31.1%) as compared to the control group. The molecular mass of this exo-biopolymer measured by HPLC was under 40 kDa. Total sugar and protein contents were 91.2 and 8.8%, respectively. The sugar and amino acid compositions of the exo-biopolymer were analyzed in detail.

Hypoglycemic effect of a Lentinus edodes exo-polymer produced from a submerged mycelial culture.

The hypoglycemic effect of an exo-polymer produced from a submerged mycelial culture of Lentinus edodes was investigated in streptozotocin-induced diabetic rats. The administration of the exo-polymer (200 mg/kg BW) reduced the plasma glucose level by as much as 21.5%, and increased plasma insulin by 22.1% as compared to the control group. It also lowered the plasma total cholesterol and triglyceride levels by 25.1 and 44.5%, respectively. Gel chromatography of the exo-polymer revealed a single peak which is likely to have been a glycoprotein with a molecular weight of 52 kDa and was found to contain 83.5% carbohydrate and 16.5% protein. The Sugar and amino acid compositions of the exo-polymer were analyzed in detail.