Anti-genotoxic effect of aqueous extracts of sun mushroom (Agaricus blazei Murill lineage 99/26) in mammalian cells in vitro

The sun mushroom is the popular name for the Agaricus blazei Murill fungus, a mushroom native to south-eastern Brazil, which has been frequently used in popular medicine mainly in the form of tea to treat various ailments (stress, diabetes, etc.). In the present study, the genotoxic and/or anti-genotoxic effects ofA. blazei on mammalian cells in culture was assessed by checking the increase or reduction of micronucleus (MN) frequency and comets. The sun mushroom (lineage 99/26) was used as aqueous extracts prepared (2.5%) at three different temperatures (60, 25 and 4°C). The in vitro micronucleus (MN) test in binucleated cells and comet assay were used in V79 cells cultivated in HAM-F10+DMEM medium (1:1), supplemented with 10% of fetal bovine serum. The experiments were divided into four treatment types: 1. Negative control; 2. Positive control with MMS; 3. Treatments with the three forms of extracts (60, 25 and 4°C); and 4. Treatments with the extracts in different associations (simultaneous, pre-treatment, post-treatment and simultaneous after pre-incubation for 1 h) with MMS. None of the A. blazei extracts show genotoxic activity. In the comet assay no protecting effect was found. The results obtained in the MN test showed that the three forms of extracts used had protective activity, suggesting that the compound or active ingredients of A. blazei are always present in these extracts. The greater protective efficiency of the simultaneous treatment and simultaneous treatment with pre-incubation mixture with MMS suggests that the extracts have an antimutagenic action of the desmutagenic type. © 2002 Elsevier Science Ltd. All rights reserved.

Screening for In Vitro and In Vivo Antitumor Activities of the Mushroom Agaricus Blazei

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Mechanism of anticlastogenicity of Agaricus blazei Murill mushroom organic extracts in wild type CHO (K-1) and repair deficient (xrs5) cells by chromosome aberration and sister chromatid exchange assays

Agaricus blazei Murill is a medicinal mushroom native to Brazil. The present work assessed the clastogenic and anticlastogenic potential of organic extracts (ethanol and chloroform/methanol) from the lineage AB97/11 in chinese hamster CHO-K-1 (wild type) and CHO-xrs5 (repair deficient) cells using the chromosome aberration (CA) and sister chromatid exchange (SCE) assays. In these experimental conditions were observed: (a) anticlastogenic effect at concentrations of 0.06 and 0.09% of the EtOH extract and at the 0.03 and 0.06% concentrations of the C/MetOH extract in CHO-K-1; (b) absence of protector effect on CHO-xrs5 cells; and (c) absence of protector effect in the SCE assay. These results indicate that organic extracts of A. blazei lineage AB97/11 present bio-antimutagenic type protective activity. (C) 2003 Elsevier B.V. B.V. All rights reserved.

Chemoprevention of preneoplastic liver foci development by dietary mushroom Agaricus blazei Murrill in the rat

The chemopreventive potential of an Agaricus blazei (Ab) Murrill mushroom meal was investigated in a medium-term rat liver carcinogenesis assay. Male Wistar rats initiated for hepatocarcinogenesis with diethylnitrosamine (DEN, 200 mg/kg i.p.) were fed during a 6-week period with the dry powdered mushroom strains Ab 29 or 26, each one with opened (OB) or closed basidiocarp (CB), mixed at 10% level in a basal diet. All experimental animals and controls were subjected to partial hepatectomy at week 3 and killed at week 8. Chemopreventive activity of the mushroom meal was observed for the Ab 29 (OB and CB) and Ab 26 (CB) strains in terms of the number of putative preneoplastic altered foci of hepatocytes which express either the enzyme glutathione S-transferase, placental form (GST-P+) or the transforming growth factor-alpha, and for the Ab 29 (OB) and Ab 26 (CB) strains on the size of GST-P-divided by foci. This was associated with inhibition of foci cell proliferation in the animals fed the Ab 29 (013) and Ab 26 (CB) strains. The results suggest that the protective influence of the Ab meal against the DEN potential for rat liver carcinogenicity depends on both the strain and period of mushroom harvest. (C) 2003 Elsevier Ltd. All rights reserved.

Antimutagenic effects of the mushroom Agaricus blazei Murrill extracts on V79 cells

Agaricus blazei Murrill, a native mushroom in Brazil, has been widely consumed in different parts of the world due to its medicinal power. Its anticarcinogenic activity has been shown in experimental animals, and antimutagenic activity has been demonstrated only in Salmonella. In this work, the multagenic and antimutagenic activities of mushroom teas of strains AB96/07, AB96/09 and AB97/11 were evaluated in Chinese hamster V79 cells, using the comet assay and the micronucleus test. The cells were treated with three different concentrations (0.05, 0.1 and 0.15) of teas prepared from a 2.5% aqueous solution, under three different temperatures: (1) room (20-25 degreesC); (2) ice-cold (2-8 degreesC); and (3) warm (60 degreesC). The teas were applied in co-, pre- and post-treatments in combination with the mutagen methyl methanesulfonate (MMS; 1.6 x 10(-4) and 4 x 10(-4) M). The duration of the treatment was 1 h in the comet assay and 2 h in the micronucleus test. The results showed that the mushroom was not mutagenic itself. Nevertheless, the mushroom is an efficient antimutagen against the induction of micronuclei by MMS in all concentrations and preparations tested. The observed reductions in the frequencies of micronuclei ranged from 61.5 (room temperature 0.1% tea in post-treatment) to 110.3% (co-treatment with warm and ice-cold 0.15% tea). In the comet assay, the antimutagenic activity was detected only when the cells were pre-treated with the following teas: warm 0.1 and 0.15%, room temperature 0.05% and ice-cold 0.1%. The results indicate that the mushroom A. blazei extracts are antimutagenic when tested in V79 cells. (C) 2001 Elsevier B.V. B.V. All rights reserved.

beta-Glucan extracted from the medicinal mushroom Agaricus blazei prevents the genotoxic effects of benzo[a]pyrene in the human hepatoma cell line HepG2

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Effects of extracts from Brazilian sun-mushroom (Agaricus blazei) on the NK activity and lymphoproliferative responsiveness of Ehrlich tumor-bearing mice

Agaricus blazei Murrill, is an edible and medicinal mushroom which is popularly consumed due to its antitumoral properties. The immunomodulatory effects of methanol (METH), dichloromethane (DM) and n-hexane (HEX) extracts of this mushroom were evaluated in Ehrlich tumor-bearing mice. Subcutaneous inoculation of Ehrlich tumor cells inhibited the natural killer (NK) activity of spleen cells (specific lysis = 6.18 +/- 2.56%) compared with normal mice (17.59 +/- 7.77%). Treatment of tumor-bearing mice with the extracts for 10 days restored the natural killer activity against Yac-1 target cells and the best results were observed by treatment with the HEX extract (21.48 +/- 15.26%). Treatment of the animals with the HEX extract for 10 days was also able to stimulate the mitogen-induced lymphoproliferative activity of spleen cells. Thirty days after the treatment, all groups presented low proliferative activity. Specific antibody production was observed to be higher in the groups treated with the DM or METH extract 30 days after the treatment. Analysis of the 3 extracts by gas chromatography mass spectrum (GCMS) and magnetic nuclear resonance (MNR) showed that the HEX extract contains mainly sugar and fatty acids and that the METH extract also contains sugar and possibly amino acids. (C) 2004 Elsevier Ltd. All rights reserved.

Genotoxic and antigenotoxic effects of organic extracts of mushroom Agaricus blazei Murrill on V79 cells

Agaricus blazei Murrill, popularly known as the sun mushroom, is a native mushroom in SP, Brazil, that has been widely used in the treatment of cancer and many other pathologies in different parts of the world. A water-soluble protein-polysaccharide complex (1 → 6)β-D-glucan has been isolated from its fruiting body that showed immune-modulation activity. From organic extracts, linoleic acid has been isolated and determined to be the main substance with antimutagenic activity. Using both the micronucleus (MN) and comet (single cell microgel electrophoresis) assays, this study determined the genotoxic and antigenotoxic potential of A. blazei (AB) obtained from commercial sources or the following strains: a) strains AB 97/29 (young and sporulated phases); b) a mixture taken from AB 96/07, AB 96/09 and AB 97/ 11 strains; and c) commercial mushrooms from Londrina, PR and Piedade, SP, designated as AB PR and AB SP, respectively. The extracts from these mushrooms were isolated in chloroform:methanol (3:1) and used in vitro at three different concentrations. V79 cells (Chinese hamster lung cells) were exposed to the extracts under pre-, simultaneous and post-treatment conditions, combined with methyl methanesulfonate (MMS). Under the circumstances of this study, these organic extracts did not show any genotoxic or mutagenic effects, but did protect cells against the induction of micronuclei by MMS. Copyright by the Brazilian Society of Genetics.

Anticlastogenic effect of aqueous extracts of Agaricus blazei on CHO-k1 cells, studying different developmental phases of the mushroom

The Agaricus blazei Murill (ABM) mushroom, known as the sun mushroom, is native to Brazil and has become known for its medicinal properties. This study examined the anticlastogenic effect of Agaricus blazei in Chinese hamster ovary cells, CHO-k1, by means of a chromosome aberration test using methyl methanesulphonate (MMS, 10(-4)M) as the DNA damage inducing agent. Two mushroom lines were used, ABM 99/26 and ABM 97/11, and the latter was used in the young (Y) and sporulating (S) developmental phases. The cells were treated for 12 h with MMS alone or combined with aqueous extracts of A. blazei at a final concentration of 0.15%, which were prepared at three different temperatures: (a) hot (60 degreesC), (b) room temperature (25 degreesC) and (c) chilled (4 degreesC). Mushroom extracts showed a marked anticlastogenic effect against DNA damage, as evidenced by a decrease in the number of cells with breaks, regardless of the line used, or the developmental stage or the temperature at which the extract was prepared. Generally, the extracts were more effective in reducing the isochromatid type breaks. The data obtained suggest that extracts of A. blazei mushroom are anticlastogenic under the conditions tested, mainly during the G1 and S stages of the cell cycle, where chromosome breaks of the isochromatid type are produced by the MMS agent. (C) 2003 Elsevier Ltd. All rights reserved.

Non-mutagenic or genotoxic effects of medicinal aqueous extracts from the Agaricus blazei mushroom in V79 cells

Agaricus blazei Murill is a mushroom largely consumed due to its medicinal properties. Effects of aqueous extract from its lineage AB97/11 in 2 fruiting body development stages (closed and opened pileus) were evaluated on chinese hamster V79 cells using cytokinesis blocking micronucleus (CBMN) and comet assays. The cells were treated at 0.15% concentration of aqueous extract prepared at different temperatures: ice-cold (4°C), room temperature (21°C) and warm (60°C). The extracts were applied in mutagenicity and antimutagenicity protocols (simultaneous, pre-incubation and continuous). The results showed that the aqueous extracts of Agaricus blazei lineage AB97/11 obtained at the 3 temperatures and both development stages did not present mutagenic or antimutagenic effect in V79 cells either in CBMN or comet assay.

Variation of the antimutagenicity effects of water extracts of Agaricus blazei Murrill in vitro

Agaricus blazei Murill, popularly known as Sun Mushroom or Himematsutake, is native to Brazil. Nowadays, this mushroom has been target of great scientific interest due to its medical power and because it has shown antitumoral and immune modulatory properties. This work evaluated the mutagenic and antimutagenic potential from aqueous extracts prepared in different temperatures (4 degreesC, 25 degreesC and 60 degreesC) from the lineage AB 97/29 in two basidiocarp phases (young and sporulated) and from A. blazei commercialized in Londrina- PR - Brazil, named here as AB PR, and in Piedade- SP- Brazil, named as AB SP. Both micronucleus (MN) as comet assays were used. Chinese hamster lung V79 cells were treated in three antimutagenic experimental protocols: pre-, post- and simultaneous treatments, with the aqueous extracts of the A. blazei Murill and methyl methanesulfonate (MMS). The results suggested that under these circumstances of treatment, aqueous extracts of the A. blazei in both assays did not show any genotoxic potential. However, by the MN test, an antigenotoxic effect was shown against mutagenicity inducted by MMS for aqueous extracts at 60 degreesC of mushroom commercialized in Piedade- SP, in pre-, post- and simultaneous treatments and for AB PR only when used in pre-treatment. on the other hand, with comet assay, the results showed no protective effect in any case. The numbers indicated that different results can be get from A. blazei teas, and that not all of them seemed to be an efficient antimutagen against the induction of micronuclei by MMS. (C) 2003 Elsevier Ltd. All rights reserved.

Agaricus blazei (Himematsutake) does not alter the development of rat diethylnitrosamine-initiated hepatic preneoplastic foci

The modifying potential of crude extracts of the mushroom Agaricus blazei Murrill (Himematsutake) on the development and growth of glutathione S-transferase placental form (GST-P)-positive liver foci (liver preneoplastic lesion) was investigated in adult male Wistar rats. Six groups of animals were used. Groups 2 to 5 were given a single i.p. injection of 200 mg/kg b.w. of diethylnitrosamine (DEN) and groups 1 and 6 were treated with saline at the beginning of the experiment. After 2 weeks, animals of groups 3 to 6 were orally treated with three dose levels of aqueous extracts of the mushroom A. blazei (1.2, 5.6, 11.5, and 11.5 mg/ml of dry weight of solids) for 6 weeks. All animals were subjected to two-thirds partial hepatectomy at week 3 and sacrificed at week 8. Two hours before sacrifice, ten animals of each group were administered a single i.p injection of 100 mg/kg of bromodeoxyuridine (BrdU). Apoptotic bodies and BrdU-positive hepatocyte nuclei were quantified in liver sections stained for hematoxylin and eosin (H&E) (eosinophilic foci) and simultaneously stained for GST-P expression (GST-P-positive foci), respectively. The 6-week treatment with A. blazei did not alter the development (number and size) of GST-P-positive foci and did not affect the growth kinetics of liver normal parenchyma or foci in DEN-initiated animals. Our results indicate that the treatment with aqueous extracts of the mushroom A. blazei during the post-initiation stage of rat liver carcinogenesis does not exert any protective effect against the development of GST-P-positive foci induced by DEN. (Cancer Sci 2003; 94: 188-192).

Influence of aqueous extract of Agaricus blazei on rat liver toxicity induced by different doses of diethylnitrosamine

The modifying potential of prior administration of an aqueous extract of the mushroom Agaricus blazei Murrill (Agaricaceae) (Ab) on hepatotoxicity induced by different doses of diethylnitrosamine (DEN) in male Wistar rats was evaluated. During 2 weeks, animals of groups G3 (Ab+DEN50), G5 (Ab+DEN100), G7 (Ab+DEN200), and G8 (Ab-treated) were treated with the A. blazei through drinking water. After this period, groups G2 (DEN50), G3 (Ab+DEN50), G4 (DEN100) G5 (Ab+DEN100), G6 (DEN200), and G7 (Ab+DEN200) were given a single i.p. injection of 50, 100 and 200 mg/kg of DEN, respectively, while groups G1 (nontreated) and G8 (Ab-treated) were treated with 0.9% NaCl only. All animals were killed 48 h after DEN or NaCl treatments. The hepatocyte replication rate was estimated by the index of the proliferating cell nuclear antigen (PCNA) positive hepatocytes and the appearance of putative preneoplastic hepatocytes through expression of the enzyme glutathione S-transferase placental form (GSTP). After DEN-treatment, ALT levels, PCNA labeling index, and the number of GST-P positive hepatocytes were lower in rats that received A. blazei treatment and were exposed to 100 mg/kg of DEN. Our findings suggest that previous treatment with A. blazei exerts a hepatoprotective effect on both liver toxicity and hepatocarcinogenesis process induced by a moderately toxic dose of DEN. (C) 2002 Elsevier B.V. Ireland Ltd. All rights reserved.

Lack of chemoprevention of dietary Agaricus blazei against rat colonic aberrant crypt foci

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Evaluation of the antigenotoxicity of polysaccharides and beta-glucans from Agaricus blazei, a model study with the single cell gel electrophoresis/Hep G2 assay

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)