Evaluation of the genotoxic and anti-genotoxic activities of Silybin in human hepatoma cells (HepG2)

Silybin (SB), a constituent of the medicinal plant Silybum marianum, is reported to be a potent hepatoprotective agent, but little is currently known regarding its genotoxicity, mutagenicity and potential chemopreventive properties. In this study, we evaluated the ability of SB to induce DNA migration and micronuclei (MN) formation in human hepatoma cells (HepG2). Also, possible preventive effects of SB on MN formation induced by three different mutagens, bleomycin (BLEO), benzo[a] pyrene (B[alpha] P) and aflatoxin B(1) (AFB(1)), were studied. To clarify the possible mechanism of SB antimutagenicity, three treatment protocols were applied: pretreatment, in which SB was added before the application of the mutagens; simultaneous treatment, in which SB was added during treatment and post-treatment, in which SB was added after the application of the mutagens. At concentrations up to 100 mu M, SB was non-genotoxic, while at a concentration of 200 mu M, SB induced DNA migration, generated oxidized DNA bases, reduced cell viability, decreased the replicative index of the cells and induced oxidative stress. It is noteworthy that SB was able to reduce the genotoxic effect induced by B[alpha] P, BLEO and AFB1 in pretreatment and simultaneous treatments but had no significant effect on DNA damage induction in post-treatment. Taken together, our findings indicate that SB presents anti-genotoxic activity in vitro, which suggests potential use as a chemopreventive agent.

In vitro Antitumour Activity of Orsellinates

Lichen phenolic compounds exhibit antioxidant, antimicrobial, antiproliferative. and cytotoxic activities. The purpose of this study was to evaluate the anticancer activity of lecanoric acid, a secondary metabolite of the lichen Parmotrema tinctorum, and its derivatives, orsellinates, obtained by structural modification. A cytotoxicity assay was carried out hi vitro with sulforhodamine B (SRB) using HEp-2 larynx carcinoma, MCF7 breast carcinoma, 786-0 kidney carcinoma, and B16-F10 murine melanoma cell lines, in addition to a normal (Vero) cell line in order to calculate the selectivity index of the compounds. n-Butyl orsellinate was the most active compound, with IC(50) Values (the concentration that inhibits 50% of growth) ranging from 7.2 to 14.0 mu g/ml, against all the cell lines tested. The compound was more active (IC(50), = 11.4 mu g/mL) against B16-F10 cells than was cisplatin (12.5 mu g/mL). Conversely, lecanoric acid and methyl orsellinate were less active against all cell lines, having an IC(50) value higher than 50 mu g/mL. Ethyl orsellinate was more active against HEp-2 than against MCF7, 786-0, or B16-F10 cells. The same pattern was observed for n-propyl and n-butyl orsellinates. n-Pentyl orsellinate was less active than n-propyl or n-butyl orsellinates against HEp-2 cells. The orsellinate activity increased with chain elongation (from methyl to n-butyl), a likely consequence of an increase in lipophilicity. The results revealed that the structural modification of lecanoric acid increases the cytotoxic activity of the derivatives tested.

Effect of Annatto on Micronuclei Induction by Direct and Indirect Mutagens in HepG2 Cells

Annatto (AN), a natural food colorant rich in carotenoids, has been reported as being an effective antioxidant, but little is known about its potential chemopreventive properties. In this Study, we evaluated the ability of AN to protect human hepatoma cells (HepG2) from micronucleus (MN) induction against three different mutagens: benzo(a)pyrene (B(a)P), doxorubicin (DXR), and methyl methanesulfonate (MMS). In an attempt to clarify the possible mechanism of anti mutagenicity of AN, three protocols of treatment were applied (pretreatment; simultaneous treatment, and post-treatment with AN following treatment with the mutagens). Also, cells exposed only to AN were assayed for cytotoxicity and mutagenicity. A dosage up to 10 mu g/ml of AN was devoid of mutagenic activity. Protective effects were seen on micronuclei induced by B(a)P and DXR using pre and simultaneous treatment, but AN had no significant effect on MN induction by MMS in any of the protocols. Our results also show that exposure of cells to concentrations of AN higher than 10 mu g/ml decreased cell viability. Taken together, our findings indicate that AN presents antimutagenic activity in vitro, but its protective effect is dependent on the mutagen and on type of treatment suggesting its potential use as a chemopreventive agent. Environ. Mol. Mutagen. 50:808-814, 2009. (C) 2009 Wiley-Liss, Inc.

Bioactive Chemical Constituents and Comparative Antimicrobial Activity of Callus Culture and Adult Plant Extracts from Alternanthera tenella

Crude extracts of a callus culture (two culture media) and adult plants (two collections) from Alternanthera tenella Colla (Amaranthaceae) were evaluated for their antibacterial and antifungal activity, in order to investigate the maintenance of antimicrobial activity of the extracts obtained from plants in vivo and in vitro. The antibacterial and antifungal activity was determined against thirty strains of microorganisms including Gram-positive and Gram-negative bacteria, yeasts and dermatophytes. Ethanolic and hexanic extracts of adult plants collected during the same period of the years 1997 and 2002 [Ribeirao Preto (SP), collections 1 and 2] and obtained from plant cell callus culture in two different hormonal media (AtT43 and AtT11) inhibited the growth of bacteria, yeasts and dermatophytes with inhibition halos between 6 and 20 mm. For the crude extracts of adult plants bioassay-guided fractionation, purification, and isolation were performed by chromatographic methods, and the structures of the isolated compounds were established by analysis of chemical and spectral evidences (UV, IR, NMR and ES-MS). Steroids, saponins and flavonoids (aglycones and C-glycosides) were isolated. The minimum inhibitory concentration (MIC) of the isolated compounds varied from 50 to 500 mu g/mL.

The Protective Role of Lychnophora ericoides Mart. (Brazilian Arnica) in 1,2-Dimethylhydrazine-Induced Experimental Colon Carcinogenesis

Aberrant crypt foci (ACF) and colon rectal mucosal epithelial cell proliferation have been shown to be increased in patients with colon cancer and have been largely used for early detection of factors that influence colorectal carcinogenesis in rats. Fifty male Wistar rats were randomly divided into 5 groups. The groups G1 to G4 were given 4 injections of the carcinogen 1,2-dimethylhydrazine (DMH). The G2 group received Lychnophora ericoides (LE) extracts for 6 wk. The groups G3 and G4 received LE for 4 wk and 2 wk, respectively, at the postinitiation and initiation phases of colonic carcinogenesis. The group G5 was the control. Forty-two days after the first injections of DMH for the neoplasic induction, we observed a statistically significant decrease in the number of aberrant crypt foci (ACF) and an attenuation of the increase in cell proliferation induced by DMH in all the LE-treated groups. Thus, we concluded that Lychnophora ericoides extracts were effective against the development of cancer. These data suggest that LE has a protective influence on the process of colon carcinogenesis, suppressing both the initiation and the promotion of colonic carcinogenesis.

Cytotoxic and mutagenic evaluation of extracts from plant species of the Miconia genus and their influence on doxorubicin-induced mutagenicity: An in vitro analysis

The Miconia genus, a plant widely used for medicine, occurs in tropical America and its extracts and isolated compounds have demonstrated antibiotic, antitumoral, analgesic and antimalarial activities. However, no study concerning its genotoxicity has been conducted and it is necessary to determine its potential mutagenic effects to develop products and chemicals from these extracts. This study assessed the cytotoxicity, mutagenicity and the protective effects of methanolic extracts from Miconia species on Chinese hamster lung fibroblast cell cultures (V79). The cytotoxicity was evaluated using a clonogenic assay. Cultures exposed to the extract of Miconia albicans up to a concentration of 30 mu g/mL, M. cabucu up to 40 mu g/mL, M. albicans up to 40 mu g/mL and M. stenostachya up to 60 mu g/mL exhibited a cytotoxic effect on the cells. The clonogenic assay used three non-cytotoxic concentrations (5, 10 and 20 mu g/mL) to evaluate mutagenicity and antimutagenicity of the extracts. Cultures were treated with these three extract concentrations (mutagenicity test) or the extract associated with doxorubicin (DXR) (antimutagenicity test) in three protocols (pre-, simultaneous and post-treatments). Distilled water and DXR were used as negative and positive controls, respectively. In the micronucleus (MN) test, a significant reduction was observed in MN frequency in cultures treated with DXR and extracts compared to those receiving only DXR; a significant reduction was also observed for the presence of mutagenicity in all treatments. This study confirmed the safe use of Miconia extracts at the concentrations tested and reinforced the therapeutic properties previously described for Miconia species by showing their protective effects on doxorubicin-induced mutagenicity. (C) 2010 Elsevier GmbH. All rights reserved.

Association Among Microalbuminuria and Oxidative Stress Biomarkers in Patients With Type 2 Diabetes

Aim: Hyperglycemia in diabetes mellitus (DM) may be one of the most important factors responsible for the development of oxidative stress, which promotes the main complications in DM patients. Therefore, this study evaluated if the hyperglycemia could be related to oxidative stress biomarkers, lipid profile, and renal function in type 2 diabetes patients without clinic complications. Methods: Plasmatic malondialdehyde (MDA), serum protein carbonyl (PCO), serum creatinine levels, microalbuminuria, glycated hemoglobin, and lipid profile were analyzed in 37 type 2 diabetic patients and 25 subjects with no diabetes. Results: Serum creatinine levels were within the reference values, but microalbuminuria presented increased levels in all the patients compared with controls (P G 0.05) and above of the reference values. The MDA, PCO, low- density lipoprotein, and triglyceride levels showed positive correlation with microalbuminuria levels. Moreover, glycated hemoglobin presented positive correlation with MDA, PCO, and microalbuminuria levels. Conclusions: The hyperglycemia could be responsible for the increase of the microalbuminuria levels and for the oxidation process in lipids and proteins in DM patients. Therefore, we suggested that the microvascular lesion is a direct consequence from hyperglycemia and an indirect one from the increased oxidative stress. Malondialdehyde and protein carbonyl levels could be suggested as additional biochemical evaluation to verify tissue damage in type 2 DM patients.

Anticlastogenic and antigenotoxic effects of selenomethionine on doxorubicin-induced damage in vitro in human lymphocytes

The use of antioxidants during chemotherapy has been shown to reduce or prevent the undesirable effects experienced by healthy cells. Micronutrient selenium is well known for its antioxidant properties; however, selenium exhibits a bimodal nature in that both its beneficial and toxic properties lie within a limited and narrow dose range. The present study investigated the possible protective effects of selenomethionine (SM) on the cytotoxicity, genotoxicity and clastogenicity of the chemotherapic doxorubicin (DXR), a key chemotherapic used in cancer treatment. Human peripheral lymphocytes were treated in vitro with varying concentrations of SM (0.25 mu M, 0.5 mu M, 1.0 mu M and 2.0 mu M), tested in combination with DXR (0.15 mu g/mL). SM alone was not cytotoxic and when combined with DXR treatment, reduced the DNA damage index significantly, the frequency of chromosomal aberrations, the number of aberrant metaphases and the frequency of apoptotic cells. The mechanism of chemoprotection of SM may be related to its antioxidant properties as well as its ability to interfere with DNA repair pathways. Therefore this study showed that SM is effective in reducing the genetic damage induced by the antitumoral agent DXR. (C) 2007 Elsevier Ltd. All rights reserved.