Cytotoxic, genotoxic, and oxidative stress-inducing effect of an l-amino acid oxidase isolated from Bothrops jararacussu venom in a co-culture model of HepG2 and HUVEC cells.

Hepatocellular carcinoma incidence rates have increased worldwide, which encouraged the development of new chemotherapeutic drugs. l-Amino acid oxidases from snake venoms are cytotoxic towards human tumor cells in in vitro monoculture systems, which do not simulate the tumor microenvironment. We examined the antitumor potential of BjussuLAAO-II, an l-amino acid oxidase from Bothrops jararacussu venom, in hepatocarcinoma cells (HepG2) in monoculture and co-culture with human umbilical vein endothelial cells (HUVEC) in vitro. All the concentrations tested (0.25-5.00 µg/mL) were cytotoxic (MTT and clonogenic survival assays) towards HepG2 and HUVEC cells in monoculture, and increased oxidative stress by 2',7'-dichlorofluorescin diacetate fluorescence assay. Only 1.00 and 5.00 µg/mL exerted these effects in HepG2 cells co-cultured with HUVEC cells, and were genotoxic (comet assay) to HUVEC cells in monoculture. BjussuLAAO-II at 5.00 µg/mL induced DNA, but not chromosomal damage (micronucleus assay) in HepG2 cells in mono- and co-culture. The cytotoxicity and genotoxicity was more pronounced in monoculture, indicating that the tumor microenvironment influences the cellular response. BjussuLAAO-II caused cell death and DNA damage in HepG2 cells in vitro by inducing oxidative stress. Therefore, BjussuLAAO-II is a promising molecule for the development of new antitumor drugs.

Laurus nobilis (laurel) aqueous leaf extract's toxicological and anti-tumor activities in HPV16-transgenic mice.

Cancers induced by human papillomavirus (HPV) infection remain a significant public health threat, fueling the study of new therapies. Laurel (Laurus nobilis) compounds and extracts recently showed in vitro activity against HPV-transformed cell lines. This work aims to evaluate the in vivo efficacy and hepatic toxicity of a laurel extract in a transgenic mouse model of HPV16-induced cancer. The extract was administered in drinking water (20 mg per animal per day) for three consecutive weeks, using four experimental groups (n = 10) (group I: HPV16-/- without treatment, group II: treated HPV16-/-, group III: HPV16+/- without treatment and group IV: treated HPV16+/-). Following the treatment period, animals were sacrificed and skin samples were used to classify skin lesions histologically. Toxicological parameters included hematological and biochemical blood markers, splenic and hepatic histology and hepatic oxidative stress. The extract did not prevent the progression of HPV16-induced cutaneous lesions in this model. The treated wild-type animals showed mild hepatitis, while transgenic animals suffered weight loss. However, there were no changes concerning hematological, biochemical and hepatic oxidative stress markers.

The toxin BjussuLAAO-II induces oxidative stress and DNA damage, upregulates the inflammatory cytokine genes TNF and IL6, and downregulates the apoptotic-related genes BAX, BCL2 and RELA in human Caco-2 cells.

Colorectal carcinoma is one of the most common cancers in adults. As chemotherapy, the first-choice treatment for colorectal carcinoma, is often infeasible due to acquired tumor resistance and several adverse effects, it is important to discover and explore new molecules with better therapeutic action. Snake venom toxins have shown promising results with high cytotoxicity against tumor cells, but their mechanisms of action remain unclear. Here we examined how BjussuLAAO-II, an L-amino acid oxidase isolated from Bothrops jararacussu snake venom, exerts cytotoxicity towards colorectal adenocarcinoma human cells (Caco-2) and human umbilical vein endothelial cell line (HUVEC). A 24-h treatment with BjussuLAAO-II at 0.25 - 5.00 µg/mL diminished cell viability by decreasing (i) mitochondrial activity, assessed by reduction of 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide and resazurin; (ii) the activity of acid phosphatases; and (iii) lysosomal function, assessed by neutral red uptake. BjussuLAAO-II also increased intracellular levels of reactive oxygen species and DNA damage, as assessed by fluorescence and the comet assay, respectively. BjussuLAAO-II altered the expression of cell proliferation-related genes, as determined by RT-qPCR: it elevated the expression of the inflammatory cytokine genes TNF and IL6, and lowered the expression of the apoptotic-related genes BAX, BCL2, and RELA. Therefore, BjussuLAAO-II induces Caco-2 cells death by acting on multiple intracellular targets, providing important data for further studies to assess whether these effects are seen in both tumor and normal cells, with the aim of selecting this drug for possible therapeutic purposes in the future.

Protective effects of the exopolysaccharide Lasiodiplodan against DNA damage and inflammation induced by doxorubicin in rats: Cytogenetic and gene expression assays.

The lasiodiplodan (LS) is a ß-(1→6)-d-glucan produced by the fungus Lasiodiplodia theobromae and some of the biological activities of LS were reported as hypoglycemic, anticoagulant, anti-proliferative and anticancer action; however, its effects on DNA instability and modulation of gene expression are still unclear. Aims of study were investigate the genotoxic effects of lasiodiplodan, and its protective activity against DNA damage induced by doxorubicin (DXR) and its impact on the expression of genes associated with DNA damage and inflammatory response pathways. Therefore, Wistar rats were treated (15 days) orally with LS (5.0; 10 and 20mg/kg bw) alone and in combination with DXR (15mg/kg bw; administrated intraperitoneally on 14th day) as well as their respective controls: distilled water and DXR. Monitoring of DNA damage was assessed by comet and micronucleus (MN) assays and gene expression was evaluated by PCR-Arrays. Treatments with LS alone did not induce disturbances on DNA; when LS was given in combination with DXR, comet and MN formations were reduced to those found in the respective controls. Moreover, LS was able to reduce the disturbances on gene expressions induced by DXR treatment, since the animals that receive LS associated with DXR showed no alteration in the expression of genes related to DNA damage response. Also, DXR induced several up- and down-regulation of several genes associated to inflammatory process, while the animals that received LS+DXR had their gene expression patterns similar to those found in the control group. In conclusion, our results showed that LS did not induce disturbances on DNA stability and significantly reduce the DNA damage and inflammation caused by DXR exposure. In addition, we give further information concerning the molecular mechanisms associated to LS protective effects which seems to be a promising nutraceutical with chemopreventive potential.

Chemopreventive effect and lack of genotoxicity and mutagenicity of the exopolysaccharide botryosphaeran on human lymphocytes.

Carbohydrate biopolymers of fungal-origin are an important natural resource in the search for new bioagents with therapeutic and nutraceutical potential. In this study the mutagenic, genotoxic, antigenotoxic and antioxidant properties of the fungal exopolysaccharide botryosphaeran, a (1→3)(1→6)-ß-D-glucan, from Botryosphaeria rhodina MAMB-05, was evaluated. The mutagenicity was assessed at five concentrations in Salmonella typhimurium by the Ames test. Normal and tumor (Jurkat cells) human T lymphocyte cultures were used to evaluate the genotoxicity and antigenotoxicity (Comet assay) of botryosphaeran alone and in combination with the mutagen methyl methanesulfonate (MMS). The ability of botryosphaeran to reduce the production of reactive oxygen and nitrogen species (RONS) generated by hydrogen peroxide was assessed using the CM-H2DCFDA probe in lymphocyte cultures under different treatment times. None of the evaluated botryosphaeran concentrations were mutagenic in bacteria, nor induced genotoxicity in normal and tumor lymphocytes. Botryosphaeran protected lymphocyte DNA against damage caused by MMS under simultaneous treatment and post-treatment conditions. However, botryosphaeran was not able to reduce the RONS generated by H2O2. Besides the absence of genotoxicity, botryosphaeran exerted a protective effect on human lymphocytes against genotoxic damage caused by MMS. These results are important in the validation of botryosphaeran as a therapeutic agent targeting health promotion.

Euthanasia using gaseous agents in laboratory rodents.

Several questions have been raised in recent years about the euthanasia of laboratory rodents. Euthanasia using inhaled agents is considered to be a suitable aesthetic method for use with a large number of animals simultaneously. Nevertheless, its aversive potential has been criticized in terms of animal welfare. The data available regarding the use of carbon dioxide (CO2), inhaled anaesthetics (such as isoflurane, sevoflurane, halothane and enflurane), as well as carbon monoxide and inert gases are discussed throughout this review. Euthanasia of fetuses and neonates is also addressed. A table listing currently available information to ease access to data regarding euthanasia techniques using gaseous agents in laboratory rodents was compiled. Regarding better animal welfare, there is currently insufficient evidence to advocate banning or replacing CO2 in the euthanasia of rodents; however, there are hints that alternative gases are more humane. The exposure to a volatile anaesthetic gas before loss of consciousness has been proposed by some scientific studies to minimize distress; however, the impact of such a measure is not clear. Areas of inconsistency within the euthanasia literature have been highlighted recently and stem from insufficient knowledge, especially regarding the advantages of the administration of isoflurane or sevoflurane over CO2, or other methods, before loss of consciousness. Alternative methods to minimize distress may include the development of techniques aimed at inducing death in the home cage of animals. Scientific outcomes have to be considered before choosing the most suitable euthanasia method to obtain the best results and accomplish the 3Rs (replacement, reduction and refinement).

Microparticles Containing Curcumin Solid Dispersion: Stability, Bioavailability and Anti-Inflammatory Activity.

This work aimed at improving the solubility of curcumin by the preparation of spray-dried ternary solid dispersions containing Gelucire®50/13-Aerosil® and quantifying the resulting in vivo oral bioavailability and anti-inflammatory activity. The solid dispersion containing 40% of curcumin was characterised by calorimetry, infrared spectroscopy and X-ray powder diffraction. The solubility and dissolution rate of curcumin in aqueous HCl or phosphate buffer improved up to 3600- and 7.3-fold, respectively. Accelerated stability test demonstrated that the solid dispersion was stable for 9 months. The pharmacokinetic study showed a 5.5-fold increase in curcumin in rat blood plasma when compared to unprocessed curcumin. The solid dispersion also provided enhanced anti-inflammatory activity in rat paw oedema. Finally, the solid dispersion proposed here is a promising way to enhance curcumin bioavailability at an industrial pharmaceutical perspective, since its preparation applies the spray drying, which is an easy to scale up technique. The findings herein stimulate further in vivo evaluations and clinical tests as a cancer and Alzheimer chemoprevention agent.

Effects of lutein and chlorophyll b on GSH depletion and DNA damage induced by cisplatin in vivo.

Recent studies have proposed the use of low concentrations of phytochemicals and combinations of phytochemicals in chemoprevention to reduce cytotoxicity and simulate normal ingestion through diet. The purpose of the present study was to evaluate whether the DNA damage, chromosome instability, and oxidative stress induced by cisplatin (cDDP) are modulated by a combination of the natural pigments lutein (LT) and chlorophyll b (CLb). The protective effects observed for synergism between phytochemicals have not been completely investigated. The comet assay and micronucleus test were performed and the catalase activities and glutathione (GSH) concentrations were measured in the peripheral blood, bone marrow, liver, and kidney cells of mice. The comet assay and micronucleus test results revealed that the pigments LT and CLb were not genotoxic or mutagenic and that the pigments presented antigenotoxic and antimutagenic effects in the different cell types evaluated. This protective effect is likely related to antioxidant properties in peripheral blood cells through the prevention of cDDP-induced GSH depletion. Altogether our results show that the combination of LT and CLb, which are both usually present in the same foods, such as leafy green vegetables, can be used safely.

Loss of heterozygosity in the fragile histidine triad (FHIT) locus and expression analysis of FHIT protein in patients with breast disorders.

PURPOSE OF INVESTIGATION: The fragile histidine triad (FHIT) gene is a tumor suppressor frequently inactivated in various types of tumors. The authors evaluated the occurrence of loss of heterozygosity (LOH) in the FHIT locus and FHIT protein changes in breast tissue. MATERIALS AND METHODS: Blood and breast tissue samples were obtained from 35 women with mammary disorders. The occurrence of LOH in FHIT locus was assayed by polymerase chain reaction (PCR), and the results obtained from blood and breast tissues from each patient were compared. FHIT protein expression was evaluated by immunohistochemistry. RESULTS: LOH in the FHIT gene occurred in 48.6% (17/35) of patients with mammary disorder. Among patients with malignant breast disorders, 59.1% (13/22) presented LOH in the FHIT gene in comparison with patients with benign breast lumps, in which the LOH was observed in 30.8% (4/13) of women, suggesting that changes in this gene occur prior to the process of mammary carcinogenesis. The changes in the locus of the FHIT gene occur with greater frequency in the coded region of the gene, principally near exons 5 and 8, where the FRA3B site and the histidine triad respectively are found. Changes in FHIT did not modify protein expression. The association between menopause and LOH in the FHIT gene was evident. CONCLUSIONS: LOH in the FHIT gene may be related to menopause in women with breast disorders.

The anaesthetic combination of ketamine/midazolam does not alter the acquisition of spatial and motor tasks in adult mice.

The ketamine/midazolam association of a dissociative with a sedative agent is used for the induction and maintenance of anaesthesia in laboratory animals. Anaesthesia may interfere with research results through side-effects on the nervous system, such as memory impairment. It is known that ketamine and midazolam affect cognition; however, their effects have not been clarified when used in a context of balanced anaesthesia. Thus, this study evaluated the effects of ketamine/midazolam on the acquisition of motor and of a spatial memory task in adult mice. Twenty-eight C57BL/6 adult male mice were divided into three groups: untreated control, treated with ketamine/midazolam (75 mg/kg / 10 mg/kg) and treated with midazolam (10 mg/kg) groups. Respiratory rate, heart rate and systolic pressure were measured every 5 min in the animals treated with ketamine/midazolam, as this was the only group that exhibited loss of the righting reflex. One day after treatment, animals were tested in the open field, rotarod and radial arm maze. There were no differences between treatments regarding open-field activity, rotarod performance or number of working and reference memory errors in the radial arm maze task. In conclusion, the learning process of spatial and motor tasks was not disrupted by the anaesthetic combination of ketamine/midazolam. These results suggest its safe use in adult mice in projects where acquisition of a spatial and motor task is necessary.

Apoptotic neurodegeneration and spatial memory are not affected by sedative and anaesthetics doses of ketamine/medetomidine combinations in adult mice.

BACKGROUND: Ketamine is increasingly popular in clinical practice and its combination with α(2)-agonists can provide good anaesthetic stability. Little is known about the effects of this combination in the brain. Therefore, we investigated the effects of different concentrations of ketamine combined with medetomidine on cognition and its potential apoptotic neurodegenerative effect in adult mice. METHODS: Seventy-eight C57BL/6 adult mice were divided into six different groups (saline solution, 1 mg kg(-1) medetomidine, 25 mg kg(-1) ketamine+1 mg kg(-1) medetomidine, 75 mg kg(-1) ketamine+1 mg kg(-1) medetomidine, 25 mg kg(-1) ketamine, and 75 mg kg(-1) ketamine). Eight animals per group were tested in the T-maze, vertical pole, and open-field test. Five animals per group were used for histopathological [haematoxylin and eosin (HE) staining] and immunohistochemical analyses [caspase-3 activation and expression of neurotrophin brain-derived neurotrophic factor (BDNF)]. Cells showing clear HE staining and positive immunoreactions for caspase-3 and BDNF in the retrosplenial cortex, visual cortex, pyramidal cell layer of the cornu Ammonis 1 and cornu Ammonis 3 areas of the hippocampus, and in the granular layer of the dentate gyrus were counted. RESULTS: There were no differences between groups regarding the number of dead cells and cells showing positive immunoreactions in the different areas of the brain studied. Similarly, no differences were detected in the number of trials to complete the T-maze task. Nevertheless, α(2)-agonist decreased hyperlocomotion caused by ketamine in the open field. CONCLUSIONS: Neither apoptotic neurodegeneration nor alterations in spatial memory were observed with different concentrations of ketamine combined with medetomidine in adult mice.

Performance of electroencephalogram-derived parameters in prediction of depth of anaesthesia in a rabbit model.

BACKGROUND: The index of consciousness (IoC), the permutation entropy (PE), and the approximate entropy are recent EEG-derived indices of anaesthetic depth. In this study, a rabbit model under fentanyl and isoflurane anaesthesia was used to compare the performance of these indices and also the classic median and spectral edge frequency 95%. METHODS: EEG recordings were obtained from six rabbits. Animals received fentanyl for premedication, followed by induction with propofol and maintenance with isoflurane. Anaesthetic depth was evaluated according to a clinical scale from 1 (awake) to 4 (surgical anaesthesia). Animals were submitted to surgical implantation of a small device in the lumbar muscles. A correction factor for the EEG suppression ratio was applied to the spectral parameters and to the PE. The correlation of the indices with the clinical scale of anaesthesia was analysed using prediction probability. Repeated-measures analysis of variance or its non-parametric equivalent was used to analyse the indices values at the study times and to compare their variability. RESULTS: The IoC showed the best mean prediction probability value [0.94 (0.01)] followed by burst suppression-corrected PE [0.91(0.03)]. Both parameters also showed less variability than the others. CONCLUSIONS: The IoC and PE are promising indices for anaesthetic depth monitoring. The PE might benefit from the application of a burst suppression correction at deeper stages of anaesthesia. The rabbit is useful as a translational research animal model for the validation of clinical indices.

Genotoxic effects of aluminum, iron and manganese in human cells and experimental systems: a review of the literature.

There is considerable evidence indicating an increase in neurodegenerative disorders in industrialized countries. The clinical symptoms and the possible mutagenic effects produced by acute poisoning and by chronic exposure to metals are of major interest. This study is a review of the data found concerning the genotoxic potential of three metals: aluminum (Al), iron (Fe) and manganese (Mn), with emphasis on their action on human cells.

Rumen biohydrogenation-derived fatty acids in milk fat from grazing dairy cows supplemented with rapeseed, sunflower, or linseed oils.

The effects of supplementation with rapeseed, sunflower, and linseed oils (0.5 kg/d; good sources of oleic, linoleic, and linolenic acids, respectively) on milk responses and milk fat fatty acid (FA) profile, with special emphasis on rumen-derived biohydrogenation intermediates (BI), were evaluated in a replicated 4 x 4 Latin square study using 16 grazing dairy cows. The dietary treatments were 1) control diet: 20-h access to grazing pasture supplemented with 5 kg/d of corn-based concentrate mixture (96% corn; CC); 2) RO diet: 20-h access to grazing supplemented with 4.5 kg/d of CC and 0.5 kg of rapeseed oil; 3) SO diet: 20-h access to grazing supplemented with 4.5 kg/d of CC and 0.5 kg of sunflower oil; and 4) LO diet: 20-h access to grazing supplemented with 4.5 kg/d of CC and 0.5 kg of linseed oil. Milk fatty acids were converted to methyl esters and analyzed by gas-liquid chromatography and silver-ion HPLC. Dietary treatments had no effect on milk production or on milk protein content and milk protein production. Supplementation with rapeseed and sunflower oils lowered milk fat content and milk fat production, but linseed oil had no effect. Inclusion of dietary vegetable oils promoted lower concentrations of short-chain (including 4:0) and medium-chain FA (including odd- and branched-chain FA) and 18:3n-3, and higher concentrations of C(18) FA (including stearic and oleic acids). The BI concentration was higher with the dietary inclusion of vegetable oils, although the magnitude of the concentration and its pattern differed between oils. The RO treatment resulted in moderate increases in BI, including trans 18:1 isomers and 18:2 trans-7,cis-9, but failed to increase 18:1 trans-11 and 18:2 cis-9,trans-11. Sunflower oil supplementation resulted in the highest concentrations of the 18:1 trans-10, 18:1 cis-12, and 18:2 trans-10,trans-12 isomers. Concentrations of 18:1 trans-11 and 18:2 cis-9,trans-11 were higher than with the control and RO treatments but were similar to the LO treatment. Concentration of BI in milk fat was maximal with LO, having the highest concentrations of some 18:1 isomers (i.e., trans-13/14, trans-15, cis-15, cis-16), most of the nonconjugated 18:2 isomers (i.e., trans-11,trans-15, trans-11,cis-15, cis-9,cis-15, and cis-12,cis-15), and conjugated 18:2 isomers (i.e., trans-11,cis-13, cis-12,trans-14, trans-11,trans-13, trans-12,trans-14, and trans-9,trans-11), and all conjugated 18:3 isomers. The LO treatment induced the highest amount and diversity of BI without decreasing milk fat concentration, as the RO and SO treatments had, suggesting that the BI associated with 18:3n-3 intake may not be the major contributors to inhibition of mammary milk fat synthesis.

Correlation between clinical signs of depth of anaesthesia and cerebral state index responses in dogs during induction of anaesthesia with propofol.

The cerebral state index (CSI) is used for monitoring EEG and depth of anaesthesia. The objective of this study was to analyse the correlation between ocular reflexes, CSI and estimated propofol plasma concentrations (PropCP) in dogs during induction of anaesthesia with propofol. Fourteen dogs were premedicated with acepromazine 0.05 mg kg(-1) IM. Anaesthesia was induced with a 200 ml h(-1) propofol 1% constant infusion rate until loss of corneal reflex using RugLoop II software with Beths' pharmacokinetic model to estimate PropCp. Palpebral reflex (PR) and the corneal reflex (CR) were tested every 30s and classified as present (+) or absent (-), and eyeball position was registered as rotated ventromedialy (ERV) or centred (EC). Heart rate (HR), mean arterial pressure (MAP) and CSI values were analyzed from baseline before the beginning of propofol infusion (T0) until loss of CR; CSI and PropCp, CSI and anaesthetic planes, and PropCp and anaesthetic planes were compared using correlation analysis. PropCp reached 7.65+/-2.1 microg ml(-1) at the end of the study. CSI values at T0 were 89.2+/-3.8. Based on the observation of ocular reflexes and eyeball position, it was possible to define five anaesthetic planes: A (superficial) to E (deep), being A (PR+/CR+/EC), B (PR+/ERV/CR+), C (PR-/ERV/CR+), D (PR-/EC/CR+) and E (PR-/EC/CR-). There was a significant correlation between PropCp and the anaesthetic planes (R=0,861; P<0.01). No significant correlation was observed between CSI and the anaesthetic planes or between CSI and PropCp. MAP decreased significantly from T0 until loss of corneal reflex (from 98+/-14 mm Hg to 82+/-12 mm Hg); HR did not change significantly (from 101+/-30 bpm to 113+/-16 bpm). The CSI monitoring was not consistent with the clinical observations observed in the different stages of depth anaesthesia. This could limit the use of CSI for monitoring depth of anaesthesia with propofol.

Intraperitoneal anaesthesia with propofol, medetomidine and fentanyl in mice.

Fast recoveries are essential when looking for a safe anaesthetic protocol to use on mice. Propofol is a short-acting anaesthetic agent, which provides a smooth, fast recovery. A recent study carried out in our laboratory showed that the intraperitoneal (i.p.) administration of propofol combined with a fast-acting opioid does not provide a sufficiently stable anaesthesia. In this experiment, we hypothesized that the additional application of medetomidine would increase muscle relaxation and analgesia. Fifty-four male CD1 mice, divided into six groups of five and three groups of eight, were used to test nine different combinations of propofol, medetomidine and fentanyl administered i.p. and reversed with atipamezole 30 min after induction. These combinations were composed in the following manner: propofol 75 mg/kg, medetomidine 1 and 2 mg/kg and fentanyl 0.1, 0.15 and 0.2 mg/kg. The depth of anaesthesia, loss of righting reflex, loss of pedal withdrawal reflex, pulse rate and respiratory rate were recorded along with the duration and quality of the recovery. The combination of propofol and medetomidine provided a predictable induction, hypnosis and muscle relaxation, but surgical anaesthesia (loss of pedal withdrawal reflex) was not achieved. The addition of fentanyl increased analgesia leading to surgical anaesthesia. We concluded that a combination of 75/1/0.2 mg/kg of propofol, medetomidine and fentanyl, respectively, is a safe, easy and reversible technique for i.p. anaesthesia in mice, providing a surgical window of 15 min and restraint for 30 min with a fast recovery.

Genotoxic effect of Physalis angulata L. (Solanaceae) extract on human lymphocytes treated in vitro

Physalis angulata L (Solanaceae) is a medicinal plant from North of Brazil, whose different extracts and infusions are commonly used in the popular medicine for the treatment of malaria, asthma, hepatitis, dermatitis and rheumatism. However, the genotoxic effects of P. angulata on human cells is not well known. The main purpose of the present study was to evaluate the in vitro genotoxic effects of aqueous extract of P. angulata using the comet assay and the micronucleus assay in human lymphocytes provided from 6 healthy donors. Treatments with P. angulata extracts were performed in vitro in order to access the extent of DNA damage. The comet assay has shown that treatments with P. angulata at 0.5, 1.0, 2.0, 3.0 and 6.0 microg/mL in culture medium were genotoxic. Lymphocytes treated with P. angulata at the concentrations of 3.0 and 6.0 microg/mL in culture medium showed a statistically significant increase in the frequency of micronucleus (p<0.05), however, the cytokinesis blocked proliferation index (CBPI) was not decreased after P. angulata treatment. In conclusion, the present work demonstrated the genotoxic effects of P. angulata extract on human lymphocytes in vitro.

Genotoxic effect of Physalis angulata L. (Solanaceae) extract on human lymphocytes treated in vitro

Physalis angulata L (Solanaceae) is a medicinal plant from North of Brazil, whose different extracts and infusions are commonly used in the popular medicine for the treatment of malaria, asthma, hepatitis, dermatitis and rheumatism. However, the genotoxic effects of P. angulata on human cells is not well known. The main purpose of the present study was to evaluate the in vitro genotoxic effects of aqueous extract of P. angulata using the comet assay and the micronucleus assay in human lymphocytes provided from 6 healthy donors. Treatments with P. angulata extracts were performed in vitro in order to access the extent of DNA damage. The comet assay has shown that treatments with P. angulata at 0.5, 1.0, 2.0, 3.0 and 6.0 microg/mL in culture medium were genotoxic. Lymphocytes treated with P. angulata at the concentrations of 3.0 and 6.0 microg/mL in culture medium showed a statistically significant increase in the frequency of micronucleus (p<0.05), however, the cytokinesis blocked proliferation index (CBPI) was not decreased after P. angulata treatment. In conclusion, the present work demonstrated the genotoxic effects of P. angulata extract on human lymphocytes in vitro.(AU)

Intraperitoneal propofol and propofol fentanyl, sufentanil and remifentanil combinations for mouse anaesthesia.

The combination of propofol and a rapid-acting opioid, such as fentanyl, sufentanil or remifentanil, is a relatively safe, total intravenous anaesthesia technique, commonly used in humans and which has been investigated in laboratory animals. The objective of this study was to evaluate these combinations for anaesthesia of mice by the intraperitoneal (i.p.) route. Sixty-seven mice, divided into groups of four, were used to test 28 combinations of propofol alone and propofol with fentanyl, sufentanil or remifentanil administered i.p. The dose ranges of drugs studied were propofol 50-200 mg/kg, fentanyl 0.2-0.4 mg/kg, sufentanil 0.05-0.1 mg/kg and remifentanil 0.2-1.0 mg/kg. The loss of righting reflex (RR) and the loss of pedal withdrawal reflex (PWR) were recorded along with the duration and quality of recovery. The results obtained in these studies were unpredictable. The same dose combinations of propofol and opioids were associated with different responses in different individuals. Higher doses did not induce loss of RR and PWR in all animals and were associated with high mortality rates. An adequate hypnotic level was only observed with higher doses of propofol. The synergistic effect of propofol and the opioids was not sufficient to allow surgical procedures. Animals that reached PWR loss showed tail rigidity, shaking limbs and scratched their heads with their forefeet. Higher opioid doses induced respiratory depression and higher death rates. The inconsistency between and within groups may be associated with the i.p. route. The results reported here show that the i.p. route is not appropriate for mouse anaesthesia using propofol alone or in combination with fentanyl, sufentanil or remifentanil.