Mariana de Oliveira Mauro

Evaluation of the antimutagenic and anticarcinogenic effects of inulin in vivo

The incidence of colorectal cancer is growing worldwide. The characterization of compounds present in the human diet that can prevent the occurrence of colorectal tumors is vital. The oligosaccharide inulin is such a compound. The aim of this study was to evaluate the antigenotoxic, antimutagenic and anticarcinogenic effects of inulin in vivo. Our study is based on 3 assays that are widely used to evaluate chemoprevention (comet assay, micronucleus assay, and aberrant crypt focus assay) and tests 4 protocols of treatment with inulin (pre-treatment, simultaneous, post-treatment, and pre + continuous). Experiments were carried out in Swiss male mice of reproductive age. In order to induce DNA damage, we used the pro-carcinogenic agent 1,2-dimethylhydrazine. Inulin was administered orally at a concentration of 50 mg/kg body weight following the protocols mentioned above. Inulin was not administered to the control groups. Our data from the micronucleus assay reveal antimutagenic effects of inulin in all protocols. The percentage of inulin-induced damage reduction ranged from 47.25 to 141.75% across protocols. These data suggest that inulin could act through desmutagenic and bio-antimutagenic mechanisms. The anticarcinogenic activity (aberrant crypt focus assay) of inulin was observed in all protocols and the percentages of damage reduction ranged from 55.78 to 87.56% across protocols. Further tests, including human trials, will be necessary before this functional food can be proven to be effective in the prevention and treatment of colon cancer.

Evaluation of the antimutagenic activity and mode of action of the fructooligosaccharide inulin in the meristematic cells of Allium cepa culture.

This study evaluated the mutagenicity and antimutagenicity of inulin in a chromosomal aberration assay in cultures of the meristematic cells of Allium cepa. The treatments evaluated were as follows: negative control--seed germination in distilled water; positive control--aqueous solution of methyl methanesulfonate (10 μg/mL MMS); mutagenicity--aqueous solutions of inulin (0.015, 0.15, and 1.50 μg/mL); and antimutagenicity--associations between MMS and the different inulin concentrations. The antimutagenicity protocols established were pre-treatment, simultaneous simple, simultaneous with pre-incubation, and post-treatment. The damage reduction percentage (DR%) was 43.56, 27.77, and 55.92% for the pre-treatment; -31.11, 18.51, and 7.03% for the simultaneous simple; 30.43, 19.12, and 21.11% for the simultaneous with pre-incubation; and 64.07, 42.96, and 53.70% for the post-treatment. The results indicated that the most effective treatment for inhibiting damages caused by MMS was the post-treatment, which was followed by the pre-treatment, suggesting activity by bioantimutagenesis and desmutagenesis. The Allium cepa assay was demonstrated to be a good screening test for this type of activity because it is easy to perform, has a low cost, and shows DR% that is comparable to that reported studies that evaluated the prevention of DNA damage in mammals by inulin.

A new synthetic resorcinolic lipid 3-Heptyl-3,4,6-trimethoxy-3H-isobenzofuran-1-one: Evaluation of toxicology and ability to potentiate the mutagenic and apoptotic effects of cyclophosphamide

Resorcinolic lipids have important biological actions, including anti-carcinogenic activity. Therefore, we evaluated the mutagenic, genotoxic, immunomodulatory and apoptotic potential and the biochemical and histopathological changes caused by the synthetic resorcinolic lipid 3-Heptyl-3,4,6-trimethoxy-3H-isobenzofuran-1-one, (AMS35AA; 10, 20 and 40 mg/kg) alone or in combination with cyclophosphamide (100 mg/kg) in Swiss mice. The results indicated that AMS35AA is not genotoxic or mutagenic and does not alter liver or kidney histology. However, the compound does cause an increase (p < 0.05) in the levels of glutamic-oxaloacetic transaminase and creatinine and in splenic phagocytosis and liver and kidney apoptosis. When combined with cyclophosphamide, AMS35AA caused increased (p < 0.05) mutagenic damage (although the compound had anti-genotoxic activity), splenic phagocytosis, neutropenia and glutamic-oxaloacetic transaminase and creatinine levels (even in the absence of histological damage) and induced liver and kidney apoptosis. We conclude that this resorcinolic lipid may be an important chemotherapy adjuvant that can potentiate mutagenic damage and increase apoptosis caused by cyclophosphamide without causing adverse effects. In addition, the immunomodulatory activity of the compound should be noted, which counters reductions in lymphocyte number, a primary side effect of cyclophosphamide in cancer therapy.

Gestational exposure to Byrsonima verbascifolia: teratogenicity, mutagenicity and immunomodulation evaluation in female Swiss mice.

ETHNOPHARMACOLOGICAL RELEVANCE Byrsonima verbascifolia is used in folk medicine to treat diarrhea, intestinal infections, chronic wounds, Chagas disease, inflammation and as a diuretic. However there is no investigation regarding the Byrsonima verbascifolia hydrometanolic extract (BVHME) used during gestation. MATERIALS AND METHODS The pregnant females were randomly divided into 5 groups. Control group received saline plus DMSO (1%) in a volume of 0.1 mL/10 g (b.w.), via gavage, for at least 15 days prior to mating and throughout the gestational period. The Pre-treatment group received the BVHME, via gavage, at a dose of 50 mg/kg (b.w.) for at least 15 days prior to mating and up to the appearance of the vaginal plug. The Organogenesis group received the BVHME at a dose of 50 mg/kg (b.w.), via gavage, on the 5-15th gestational day. The Gestational group received the BVHME at a dose of 50 mg/kg (b.w.), via gavage, throughout the gestational period (from the 1st to the 18th day of pregnancy). The Pre+Gestational group received the BVHME at a dose of 50mg/kg (b.w.), via gavage, for at least 15 days prior to mating and up to throughout the gestational period. The clinical signals of maternal and fetuses toxicity were evaluated, as the mutagenicity and immunomodulation tests were performed. RESULTS AND CONCLUSIONS The present investigation shows, for the first time, that the use of Byrsonima verbascifolia extract in pregnant Swiss mice, did not alter the female reproductive function, mutagenicity or immunostimulation as well as not interfere with embryofetal development at least in our experimental conditions.

Gochnatia polymorpha ssp. floccosa: bioprospecting of an anti-inflammatory phytotherapy for use during pregnancy.

ETHNOPHARMACOLOGICAL RELEVANCE Gochnatia polymorpha ssp. floccosa is used in folk medicine to treat inflammation and infections. Non-steroidal anti-inflammatory drugs (NSAIDs) are the most commonly consumed medications during pregnancy in women with inflammatory diseases. However, the relationship between the use of NSAIDs and the risk of miscarriage and birth defects and/or benefits is not fully understood. Thus, an investigation regarding the use of Gochnatia polymorpha during gestation is of relevance for developing safe anti-inflammatory drugs for use during pregnancy. MATERIALS AND METHODS The pregnant females were randomly divided into 5 groups. Control group received a hydroalcoholic solution (1.2%), via gavage, for at least 15 days prior to mating and throughout the gestational period. The pre-treatment group received Gochnatia polymorpha ethanol extract (GPEE), via gavage, at a dose of 100mg/kg body weight (b.w.) for at least 15 days prior to mating and up to the appearance of the vaginal plug. The organogenesis group received GPEE at a dose of 100mg/kg (b.w.), via gavage, on the 5-15th gestacional day. The pregnancy group received GPEE at a dose of 100mg/kg (b.w.), via gavage, throughout the gestational period (from the 1st to the 18th day of pregnancy). The pre+pregnancy group received GPEE at a dose of 100mg/kg (b.w.), via gavage, for at least 15 days prior to mating and throughout the entire gestational period. The clinical signals of maternal toxicity and teratogenesis were evaluated. Additional assays to evaluate chronic inflammation, antigenotoxicity and immunomodolatory activity were performed. RESULTS AND CONCLUSIONS The results indicated that GPEE does not interfere with reproductive performance or embryo-fetal development but does correlate with reduced weight and fetal length. The extract was not teratogenic or mutagenic or an immunomodulator. However, GPEE did exhibit effective anti-inflammatory activity. Based on this study, it can be inferred that GPEE is an important, safe anti-inflammatory agent for use during pregnancy according to the experimental design we utilized, which opens up possibilities for the bioprospecting of a new anti-inflammatory phytotherapy for use during pregnancy.

Pre-treatment with glutamine reduces genetic damage due to cancer treatment with cisplatin

Cisplatin is an effective antineoplastic drug. However, it provokes considerable collateral effects, including genotoxic and clastogenic activity. It has been reported that a diet rich in glutamine can help inhibit such collateral effects. We evaluated this activity in 40 Swiss mice, distributed into eight experimental groups: G1 - Control group (PBS 0.1 mL/10 g body weight); G2 - cisplatin group (cisplatin 6 mg/kg intraperitoneally); G3, G4, G5 - glutamine groups (glutamine at 150, 300, and 600 mg/kg, respectively; orally); G6, G7, G8 - Pre-treatment groups (glutamine at 150, 300, and 600 mg/kg, respectively; orally and cisplatin 6 mg/kg intraperitonially). For the micronucleus assay, samples of blood were collected (before the first use of the drugs at T0, then 24 (T1) and 48 (T2) hours after the first administration). For the comet assay, blood samples were collected only at T2. The damage reduction percentages for the micronucleus assay were 90.0, 47.3, and 37.3% at T1 and 46.0, 38.6, and 34.7% at T2, for G6, G7, and G8 groups, respectively. For the comet assay, the damage reduction percentages were 113.0, 117.4, and 115.0% for G6, G7, and G8, respectively. We conclude that glutamine is able to prevent genotoxic and clastogenic damages caused by cisplatin.

Analysis of the anti-inflammatory and chemopreventive potential and description of the antimutagenic mode of action of the Annona crassiflora methanolic extract

Abstract Context: Annona crassiflora Mart. (Annonaceae) is a medicinal plant that is widely used in folk medicine, which leads to its investigation as a potential source of new pharmacological principles. Objective: This study describes the anti-inflammatory, antiallodynic, and antimutagenic/chemopreventive activities of the leaves A. crassiflora methanolic extract. Its antimutagenic mode of action was analyzed in a plant or animal experimental model. Materials and methods: Total flavonoids were quantified by spectrophotometry at 415 nm and its composition was analyzed by 1H NMR spectra. Animals received orally, 30, 100, and 300 mg/kg of extract in both tests, carrageenan-induced paw edema and myeloperoxidase activity. Animals were treated with 100 and 300 mg/kg, in all the analyzed tests, pleural cell migration and protein exudation, carrageenan-induced cell migration into the pouch, induction of joint inflammation and carrageenan-induced allodynia response in the mouse paw. To evaluate the antimutagenic/chemopreventive activity through the Allium cepa test, we used 5, 10, and 15 mg/L of extract, and for the micronucleus test in the peripheral blood, we used the dose of 15 mg/kg. Results: The fractionation of the ethyl acetate (EA) fraction, resulting from the partition of the methanol extract of the A. crassiflora, afforded through chromatographic methods resulted in the isolation of kaempferol 3-O-β-glucoside and kaempferol 3-O-β-diglucoside. Oral treatment with 100 and 300 mg/kg of extract significantly inhibited the carrageenan-induced edema formation, with inhibitions of 53 ± 7% and 47 ± 10%; in MPO activity, the observed inhibitions were 60 ± 7% for 100 mg/kg treatment and 63 ± 7% for 300 mg/kg. The ACME reduced significantly the total leukocytes (an inhibition of 78 ± 9% with 100 mg/kg and 90 ± 7% with 300 mg/kg) and protein levels (approximately 100% inhibition with both doses) in the pleurisy model. In carrageenan-induced leukocyte migration into the pouch, the extract inhibited leukocyte migration only when administered 300 mg/kg per dose (the reduction was 43 ± 5%). Pretreatment with extract failed to reduce the zymosan-induced edema formation and did not inhibit the carrageenan-induced mechanical allodynia. Damage reduction in Allium cepa tested with different concentrations (5, 10, and 15 mg/L) was 66.17, 75.75, and 69.19% for the pre-treatment; 72.72, 33.33, and 22.22% for the simple simultaneous treatment; 100.50, 93.93, and 102.52% for the simultaneous treatment with pre-incubation; 89.39, 79.79, and 84.34%; for the post-treatment, and 86.36, 81.31, and 93.43% for the continuous treatment. The antimutagenic evaluation in the micronucleous test showed a damage reduction of 75.00 and 64.58% for the pre-treatment and simultaneous protocols, respectively. The post-treatment protocol increased the cyclophosphamide effects in 45.83%. Conclusion: These results suggest that this medicinal plant has chemopreventive and anti-inflammatory therapeutic potential.

Resistant starch: a functional food that prevents DNA damage and chemical carcinogenesis.

Resistant starch is formed from starch and its degradation products and is not digested or absorbed in the intestine; thus, it is characterized as a fiber. Because fiber intake is associated with the prevention of DNA damage and cancer, the potential antigenotoxic, antimutagenic, and anticarcinogenic capabilities of resistant starch from green banana flour were evaluated. Animals were treated with 1,2-dimethylhydrazine and their diet was supplemented with 10% green banana flour according to the following resistant starch protocols: pretreatment, simultaneous treatment, post-treatment, and pre + continuous treatment. The results demonstrated that resistant starch is not genotoxic, mutagenic, or carcinogenic. The results suggest that resistant starch acts through desmutagenesis and bio-antimutagenesis, as well as by reducing aberrant crypt foci, thereby improving disease prognosis. These findings imply that green banana flour has therapeutic properties that should be explored for human dietary applications.

Compounds used to produce cloned animals are genotoxic and mutagenic in mammalian assays in vitro and in vivo.

The compounds 6-dimethylaminopurine and cycloheximide promote the successful production of cloned mammals and have been used in the development of embryos produced by somatic cell nuclear transfer. This study investigated the effects of 6-dimethylaminopurine and cycloheximide in vitro, using the thiazolyl blue tetrazolium bromide colorimetric assay to assess cytotoxicity, the trypan blue exclusion assay to assess cell viability, the comet assay to assess genotoxicity, and the micronucleus test with cytokinesis block to test mutagenicity. In addition, the comet assay and the micronucleus test were also performed on peripheral blood cells of 54 male Swiss mice, 35 g each, to assess the effects of the compounds in vivo. The results indicated that both 6-dimethylaminopurine and cycloheximide, at the concentrations and doses tested, were cytotoxic in vitro and genotoxic and mutagenic in vitro and in vivo, altered the nuclear division index in vitro, but did not diminish cell viability in vitro. Considering that alterations in DNA play important roles in mutagenesis, carcinogenesis, and morphofunctional teratogenesis and reduce embryonic viability, this study indicated that 6-dimethylaminopurine and cycloheximide utilized in the process of mammalian cloning may be responsible for the low embryo viability commonly seen in nuclear transfer after implantation in utero.

Cardanol: toxicogenetic assessment and its effects when combined with cyclophosphamide

Abstract Cardanol is an effective antioxidant and is a compound with antimutagenic and antitumoral activity. Here, we evaluated the genotoxic and mutagenic potential of saturated side chain cardanol and its effects in combination with cyclophosphamide in preventing DNA damage, apoptosis, and immunomodulation. Swiss mice were treated with cardanol (2.5, 5 and 10 mg/kg) alone or in combination with cyclophosphamide (100 mg/kg). The results showed that cardanol is an effective chemopreventive compound, with damage reduction percentages that ranged from 18.9 to 31.76% in the comet assay and from 45 to 97% in the micronucleus assay. Moreover, cardanol has the ability to reduce the frequency of apoptosis induced by cyclophosphamide. The compound did not show immunomodulatory activity. A final interpretation of the data showed that, despite its chemoprotective capacity, cardanol has a tendency to induce DNA damage. Hence, caution is needed if this compound is used as a chemopreventive agent. Also, this compound is likely not suitable as an adjuvant in chemotherapy treatments that use cyclophosphamide.