Pattern of injury with a graded excitotoxic insult and ensuing chronic medial septal damage in the rat brain

Brain damage caused by an acute injury depends on the initial severity of the injury and the time elapsed after the injury. To determine whether these two variables activate common mechanisms, we compared the response of the rat medial septum to insult with a graded series of concentrations of a-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) with the time-course effects of a low dose of AMPA. For this purpose we conducted a dose-response study at concentrations of AMPA between 0.27 and 10.8 nmol to measure atrophy of the septal area, losses of cholinergic and GABAergic neurons, astroglial and microglial reactions, and calcification. Cholinergic neurons, whose loss paralleled the degree of septal atrophy produced by AMPA, are more sensitive than GABAergic neurons to the injury produced by AMPA. At doses of AMPA above 2.7 nmol, calcification and the degree of microglial reaction increased only in the GABAergic region of the septal area, whereas atrophy and neuronal loss reached a plateau. We chose the 2.7-nmol dose of AMPA to determine how these parameters were modified between 4 days and 6 months after injection. We found that atrophy and neuronal loss increased progressively through the 6-month study period, whereas astrogliosis ceased to be observed after 1 month, and calcium precipitates were never detected. We conclude that septal damage does not increase with the intensity of an excitotoxic insult. Rather, it progresses continuously after the insult. Because these two situations involve different mechanisms, short-term paradigms are inappropriate for interpreting the pathogenic mechanisms responsible for long-term neurodegenerative processes.

Nitric oxide sustains IL-1β expression in human dendritic cells enhancing their capacity to induce IL-17-producing T-cells.

The role played by lung dendritic cells (DCs) which are influenced by external antigens and by their redox state in controlling inflammation is unclear. We studied the role played by nitric oxide (NO) in DC maturation and function. Human DCs were stimulated with a long-acting NO donor, DPTA NONOate, prior to exposure to lipopolysaccharide (LPS). Dose-and time-dependent experiments were performed with DCs with the aim of measuring the release and gene expression of inflammatory cytokines capable of modifying T-cell differentiation, towardsTh1, Th2 and Th17 cells. NO changed the pattern of cytokine release by LPS-matured DCs, dependent on the concentration of NO, as well as on the timing of its addition to the cells during maturation. Addition of NO before LPS-induced maturation strongly inhibited the release of IL-12, while increasing the expression and release of IL-23, IL-1β and IL-6, which are all involved in Th17 polarization. Indeed, DCs treated with NO efficiently induced the release of IL-17 by T-cells through IL-1β. Our work highlights the important role that NO may play in sustaining inflammation during an infection through the preferential differentiation of the Th17 lineage.

Suitability of human and mammalian cells of different origin for the assessment of genotoxicity of metal and polymeric engineered nanoparticles.

Nanogenotoxicity is a crucial endpoint in safety testing of nanomaterials as it addresses potential mutagenicity, which has implications for risks of both genetic disease and carcinogenesis. Within the NanoTEST project, we investigated the genotoxic potential of well-characterised nanoparticles (NPs): titanium dioxide (TiO2) NPs of nominal size 20 nm, iron oxide (8 nm) both uncoated (U-Fe3O4) and oleic acid coated (OC-Fe3O4), rhodamine-labelled amorphous silica 25 (Fl-25 SiO2) and 50 nm (Fl-50 SiO) and polylactic glycolic acid polyethylene oxide polymeric NPs - as well as Endorem® as a negative control for detection of strand breaks and oxidised DNA lesions with the alkaline comet assay. Using primary cells and cell lines derived from blood (human lymphocytes and lymphoblastoid TK6 cells), vascular/central nervous system (human endothelial human cerebral endothelial cells), liver (rat hepatocytes and Kupffer cells), kidney (monkey Cos-1 and human HEK293 cells), lung (human bronchial 16HBE14o cells) and placenta (human BeWo b30), we were interested in which in vitro cell model is sufficient to detect positive (genotoxic) and negative (non-genotoxic) responses. All in vitro studies were harmonized, i.e. NPs from the same batch, and identical dispersion protocols (for TiO2 NPs, two dispersions were used), exposure time, concentration range, culture conditions and time-courses were used. The results from the statistical evaluation show that OC-Fe3O4 and TiO2 NPs are genotoxic in the experimental conditions used. When all NPs were included in the analysis, no differences were seen among cell lines - demonstrating the usefulness of the assay in all cells to identify genotoxic and non-genotoxic NPs. The TK6 cells, human lymphocytes, BeWo b30 and kidney cells seem to be the most reliable for detecting a dose-response.

Oncobiguanides: Paracelsus" law and nonconventional routes for administering diabetobiguanides for cancer treatment

The dose makes the poison, the common motto of toxicology first expressed by Paracelsus more than 400 years ago, may effectively serve to guide potential applications for metformin and related biguanides in oncology. While Paracelsus' law for the dose-response effect has been commonly exploited for the use of some anti-cancer drugs at lower doses in non-neoplastic diseases (e.g., methotrexate), the opposite scenario also holds true; in other words, higher doses of non-oncology drugs, such as anti-diabetic biguanides, might exert direct anti-neoplastic effects. Here, we propose that, as for any drug, there is a dose range for biguanides that is without any effect, one corresponding to"diabetobiguanides" with a pharmacological effect (e.g., insulin sensitization in type 2 diabetes, prevention of insulin-dependent carcinogenesis, indirect inhibition of insulin and growth factor-dependent cancer growth) but with minimal toxicity and another corresponding to 'oncobiguanides' with pharmacological (i.e., direct and strong anticancer activity against cancer cells) as well as toxic effects. Considering that biguanides demonstrate a better safety profile than most oncology drugs in current use, we should contemplate the possibility of administering biguanides through non-conventional routes (e.g., inhaled for carcinomas of the lung, topical for skin cancers, intravenous as an adjunctive therapy, rectal suppositories for rectal cancer) to unambiguously investigate the therapeutic value of high-dose transient biguanide exposure in cancer. Perhaps then, the oncobiguanides, as we call them here, could be viewed as a mechanistically different type of anti-cancer drugs employed at doses notably higher than those used chronically when functioning as diabetobiguanides

Correlation of anti-inflammatory activity with phenolic content in the leaves of syzygium cumini (l.) skeels (myrtaceae)

Phenolic contents of extracts of Syzygium cumini leaves, collected monthly over a one-year period, were quantitatively determined by the modified Folin-Ciocalteau method. Extracts and tannin-free fractions were assayed by their potential to inhibit mouse paw edema induced by C48/80. HPLC showed high molecular weight phenolic species and flavonoids in the active extracts and fractions. The highest total phenolic content corresponded to the most potent degree of inhibition and the flavonoids were supposed to be the main species responsible for the activity, given that the flavonoid-enriched ethyl-acetate fraction maintained its effect down to a dose of 0.01 mg/kg in a dose-response manner.

A toxicidade em ambientes aquáticos: discussão e métodos de avaliação

Aquatic toxicity tests are assays performed with different aquatic organisms of different ecological organization levels. Such tests are a source of information on the toxicity of a given substance or wastewater under controlled conditions, and they complement the physico-chemical analyses. Moreover, they allow one to evaluate the risks resulting from the presence of toxic substances in the environment. Algae, crustaceans, fishes and bacteria are frequently used in toxicity tests. In this work, we will present the main aspects related to the aquatic toxicity tests and a discussion of their applicability will also be presented.

Evaluation of resistance in a selected field strain of Haemonchus contortus to ivermectin and moxidectin using the Larval Migration on Agar Test

Haemonchus contortus is one of the most common and economically significant causes of disease in small ruminants worldwide, and the control programs of parasitic nematodes - including H. contortus - rely mostly on the use of anthelmintic drugs. The consequence of the use of this, as the sole sanitary strategy to avoid parasite infections, was the reduction of the efficacy of all chemotherapeutic products with a heavy selection for resistance. The widespread of anthelmintic resistance and the difficulty of its early diagnosis has been a major concern for the sustainable parasite management on farms. The objective of this research was to determine and compare the ivermectin (IVM) and moxidectin (MOX) effect in a selected field strain of H. contortus with a known resistance status, using the in vitro larval migration on agar test (LMAT). Third stage larvae of the selected isolate were obtained from faecal cultures of experimentally infected sheep and incubated in eleven increasing diluted concentrations of IVM and MOX (6, 12, 24, 48, 96, 192, 384, 768, 1536, 3072 and 6144µg/mL). The dose-response sigmoidal curves were obtained using the R² value of >0.90 and the lethal concentration (LC50) dose for the tested anthelmintic drugs using a four-parameter logistic model. The LC50 value for MOX was significantly lower than IVM (1.253µg/mL and 91.06µg/mL), identifying the H. contortus isolate as considerably less susceptible to IVM compared to MOX. Furthermore, the LMAT showed a high consistency (p<0.0001) and provided to be a useful diagnostic tool for monitoring the resistance status of IVM and MOX in H. contortus field isolate, as well as it may be used for official routine drug monitoring programs under the Ministry of Agriculture (MAPA) guidance.

Tolerance of eucalypt (Eucalyptus spp.) seedlings to imazapyr in nutritive solution

Imazapyr has presented excellent results in controlling coppices in stand reforms of eucalypt forests, despite differences in the efficacy levels. To find out whether these different responses are caused by the genetic variability of the cultivated materials, two experiments were carried out under greenhouse conditions with different imazapyr doses in a hydroponic system in plastic vases containing 2,500 mL solution. The clones IEF-1 (Eucalyptus grandis x Eucalyptus sp. hybrid), GE 463 (E. urophylla x E. grandis), and MN 445 (E. grandis x Eucalyptus sp. hybrid) were used in the first assay, and IEF-1, IEF2 (E. grandis x E. urophylla) x Eucalyptus sp. hybrid) and the clones 129 and 7182 (E. grandis x Eucalyptus sp. hybrids) in the second assay. Thirty days after transplanting the seedlings to a nutritive solution, imazapyr was applied to the solution at doses of 0.00, 0.05, 0.10, 0.20, 0.40, 0.80, 1.60 and 3.20 muL L-1. Clone GE 463 proved to be more tolerant to imazapyr than clones IEF-1 and MN 445 in the first assay; however, in the second, clone 7182 was the most tolerant. Thus, doses should also be differentiated when controlling coppices, according to the cultivated clone.

Control of glyphosate resistant hairy fleabane (Conyza bonariensis) with dicamba and 2,4-D

Auxyn type herbicides such as dicamba and 2,4-D are alternative herbicides that can be used to control glyphosate-resistant hairy fleabane. With the forthcoming possibility of releasing dicamba-resistant and 2,4-D-resistant crops, use of these growth regulator herbicides will likely be an alternative that can be applied to the control of glyphosate resistant hairy fleabane (Conyza bonariensis). The objective of this research was to model the efficacy, through dose-response curves, of glyphosate, 2,4-D, isolated dicamba and glyphosatedicamba combinations to control a brazilian hairy fleabane population resistant to glyphosate. The greenhouse dose-response studies were conducted as a completely randomized experimental design, and the rates used for dose response curve construction were 0, 120, 240, 480, 720 and 960 g a.i. ha-1 for 2,4-D, dicamba and the dicamba combination, with glyphosate at 540 g a.e. ha-1. The rates for glyphosate alone were 0, 180, 360, 540, 720 and 960 g a.e. ha-1. Herbicides were applied when the plants were in a vegetative stage with 10 to 12 leaves and height between 12 and 15 cm. Hairy fleabane had low sensitivity to glyphosate, with poor control even at the 960 g a.e. ha-1 rate. Dicamba and 2,4-D were effective in controlling the studied hairy fleabane. Hairy fleabane responds differently to 2,4-D and dicamba. The combination of glyphosate and dicamba was not antagonistic to hairy fleabane control, and glyphosate may cause an additive effect on the control, despite the population resistance.

Resistance to ACCase inhibitors in Eleusine indica from Brazil involves a target site mutation

Eleusine indica (goosegrass) is a diploid grass weed which has developed resistance to ACCase inhibitors during the last ten years due to the intensive and frequent use of sethoxydim to control grass weeds in soybean crops in Brazil. Plant dose-response assays confirmed the resistant behaviour of one biotype obtaining high resistance factor values: 143 (fenoxaprop), 126 (haloxyfop), 84 (sethoxydim) to 58 (fluazifop). ACCase in vitro assays indicated a target site resistance as the main cause of reduced susceptibility to ACCase inhibitors. PCR-generated fragments of the ACCase CT domain of the resistant and sensitive reference biotype were sequenced and compared. A point mutation was detected within the triplet of aspartate at the amino acid position 2078 (referred to EMBL accession no. AJ310767) and resulted in the triplet of glycine. These results constitute the first report on a target site mutation for a Brazilian herbicide resistant grass weed.

Limited occurrence of resistant radish (Raphanus sativus) to AHAS-inhibiting herbicides in Argentina

Radish has developed feral and weedy biotypes, which is a concern for agriculture around the world. In Argentina, it is one of the most widespread and troublesome crop weeds. In Brazil, this species has developed herbicide-resistance to acetohydroxyacid synthase (AHAS) inhibiting herbicides. The objective of this study was to record the presence of herbicide-resistant weedy radish plants in Argentina. In spring 2008, we found a small population of radish at the end of the flowering stage in an imidazolinone-tolerant canola field treated with imazethapyr. Screening and dose-response tests were conducted to two successive generations. They proved the biotype resistant status, and showed extensive survival (between 50 and 80% of control) to the application of a double dose of four AHAS‑inhibiting herbicides from two different chemical families (imidazolinones and sulfonylureas). Dose-response assays exhibited very high resistance for imazethapyr (LD50 = 2452.5 g a.i. ha-1, GR50 = 2926.9 g a.i. ha-1) and intermediate for metsulfuron (LD50 = 3.0 g a.i. ha-1, GR50 = 43.2 g a.i. ha-1). The acquisition of cross-resistance to different herbicide families would confer an adaptive and invasive advantage in agricultural environments to this biotype. Due to the herbicide rotation conducted in the field, the dispersion of this biotype was restricted. This is the first report of resistance in weedy radish in Argentina.

Low level resistance of goosegrass (Eleusine indica) to glyphosate in Rio Grande do Sul-Brazil

In the last growing seasons, goosegrass (Eleusine spp.) control failures have been observed following application of glyphosate on Roundup Ready® soybean in Rio Grande do Sul (RS) - Brazil, suggesting this species' resistance to the herbicide. Thus, the objectives of this study were to identify the occurrence of goosegrass resistance to the herbicide glyphosate in RS; and to determine the predominant species of the genus Eleusine, as well as the LD50 and GR50 of the suspected resistant biotypes. Two experiments were conducted under greenhouse conditions: one to identify the biotypes resistant to glyphosate, and the other, a dose-response curve experiment, as well as a study of the botanical characteristics of the species. In the first experiment, 39 biotypes were tested, mainly Eleusine indica, collected with suspected resistance to glyphosate. The glyphosate dose was 2,160 g e.a. ha-1, and the control was evaluated at 28 days after treatment. All biotypes were effectively controlled,with the biotypes from the municipality of Boa Vista do Incra showing greater tolerance. Two biotypes suspected of resistance (12.1 and 12.3) and a susceptible biotype in a dose-response experiment were tested at the following doses: 0, 135, 270, 540, 1,080, 1,620, and 2,160 g e.a. ha-1. The results of this experiment showed that biotype 12.1 does not present resistance to glyphosate and biotype 12.3 has a low level resistance since it is effectively controlled by the herbicide at the maximum dose.

Resistance of Raphanus raphanistrum to the herbicide metsulfuron-methyl

The ALS-inhibiting herbicides, especially metsulfuron-methyl, are widely used for weed control, mainly wheat and barley in southern Brazil. Raphanus raphanistrum is a major weed of winter crops. However, in recent years, R.raphanistrum, after being treated with metsulfuron, has shown no symptoms of toxicity, possibly due to herbicide resistance. Aiming to evaluate the existence of R.raphanistrum biotypes resistant to metsulfuron, an experiment was conducted in a greenhouse, in a completely randomized design with four replications. The plots consisted of pots with six plants. The treatments consisted of the interaction of resistant R. raphanistrum (biotype R) and susceptible R. raphanistrum (biotypes S) with ten doses of the herbicide (0.0; 0.6; 1.2; 2.4; 4.8; 9.6; 19.2; 38.4; 76.8 and 153.6 g i.a. ha-1). The application of the test herbicides occurred when the crop was at the stage of 3 to 4 true leaves. The variables analyzed were control and dry matter accumulation. Statistical analysis of dose-response curves was performed by non linear regression. Biotype S was susceptible to the herbicide even at doses below the recommended. Biotype R was insensitive to the herbicide obtaining values of resistance factor (F) higher than 85. The dose-response curve confirmed the existence of R. raphanistrum biotypes with high level of resistance to metsulfuron-methyl.

Multiple resistance of Conyza sumatrensis to Chlorimuron­ethyl and to Glyphosate

Weed resistance to herbicides has been a major issue in Brazil, mainly due to the inefficiency of the herbicides used in no-till areas and to the high cost of these herbicide treatments. Failures in controlling the weed Conyza have been reported in Western and Northern grain crop areas in Paraná (Brazil). This work aimed to evaluate the potential occurrence of C. sumatrensis biotypes resistant to the herbicides chlorimuron-ethyl and glyphosate. Experiments were carried out under greenhouse conditions with four biotypes (Cascavel-2, Toledo-4, Tupãssi-6, and Assis Chateaubriand-7) possibly resistant to, as well as a population considered susceptible to chlorimuron-ethyl and glyphosate. To obtain dose-response curves, eight herbicide doses of chlorimuron-ethyl (0, 2.5, 5, 10, 20, 40, 80 and 160 g ha-1) and glyphosate (0, 90, 180, 360, 720, 1,440, 2,880 and 5,760 g e.a. ha-1) were applied and weed control and shoot biomass evaluations were made. Results provided evidence that two biotypes (Cascavel-2 and Tupãssi-6) were resistant to glyphosate and four biotypes (Cascavel-2, Toledo-4, Tupãssi-6 and Assis Chateaubriand-7) were resistant to chlorimuron­ethyl. Multiple resistance to glyphosate and chlorimuron was confirmed for biotypes Cascavel­2 and Tupãssi 6. This is the first report on multiple resistance in Conyza sumatrensis, worldwide.

Differential susceptibility of biotypes of conyza sumatrensis to Chlorimuron-ethyl herbicide

Horseweed (Conyza spp.) is an annual weed, infesting soybean crops in southern Brazil, with chlorimuron-ethyl being one of the most commonly used herbicides for its control. However, in recent soybean harvests, an unsatisfactory control of this weed using this herbicide was observed, generating suspicion regarding the selection of resistant biotypes. The objective of this study was to evaluate the susceptibility of horseweed biotypes to the herbicide chlorimuron-ethyl. Two experiments were conducted in a greenhouse; in the first one, the biotypes were selected selected, and the second experiment was arranged in a 5 x 5 factorial in a completely randomized design with four replications. The treatments used in the preparation of the dose response curves were doses of the herbicide chlorimuron-ethyl (0.0, 1.56, 3.13, 6.25, 12.5, and 25 g ha-1), applied on the five horseweed biotypes at the 3-4 leaf growth stage. The variables evaluated were visual control percentage and shoot dry weight, compared to the control without herbicide application, and plant acetolactate accumulation. It was concluded that there is a differential susceptibility among the biotypes at doses of less than 20 g ha-1 (dose response curves), which indicates low-level resistance. The practical consequences are the indications of chlorimuron-ethyl application at the maximum doses recomended and that the practice of rotating mechanisms of action must be used in the chemical weed management of these areas.