Semi-lethal Concentration Research Articles

Determination of the toxicity of the freshwater cyanobacterium Woronichinia naegeliana (Unger) Elenkin

Cyanobacterial blooms are undesirable for ecological and health reasons. While Woronichinia naegeliana is a cyanobacterial species that appears frequently in freshwater, information about it is limited. An evaluation of its toxicity was conducted via tests based on the crustaceans Thamnocephalus platyurus and Daphnia pulex. The greatest effect of the aqueous extract obtained from W. naegeliana cells was observed for T. platyurus. The denoted semi-lethal concentration (LC50) after 24 h of exposure was 0.99 mg of dry weight (d.w.) mL−1. A lower toxicity was displayed for D. pulex, although it grew with time. Among the 18 fractions separated from cyanobacterial extract, only one containing the microginin FR3 (MG-FR3) displayed biological activeness against T. platyurus. The remaining products synthesized by W. naegeliana displayed an absence or a low level of toxicity making it impossible to determine the LC50 value. Detailed research revealed that MG-FR3 did not affect the activity of enzymes such as trypsin, chymotrypsin, elastase and thrombin, which indicates another mode of action. The results demonstrated that blooms of W. naegeliana showed toxic activity towards invertebrate zooplankton.

Inactivation of Chironomid Larva in Water with Bacillus thuringiensis subsp. Israelensis

The excessive propagation of Chironomid larva in waterworks is a difficult problem. The inactivation of Bacillus thuringiensis subsp. israelensis (Bti) on different instar Chironomid larva was studied using distilled water as test sample. Furthermore, batch experiments were performed to analyze the influence of pH value, liquid chlorine, and water temperature and ammonia nitrogen on inactivation efficiency of Chironomid larva with Bti. Based on it, removal effects on Chironomid larva in the actual raw water and pre-chlorination coagulation sedimentation water were evaluated. The results showed that Bti possessed good inactivation performance to all larval stages of Chironomid. The 24 h semi-lethal concentration of Bti to the third instar Chironomid larva was 0.423 mg/L. The inactivation rate at pH 9.2 was significantly higher than that at pH 4.4 and 7.1. The liquid chlorine had negative effects on inactivation, indicated by the decreased inactivation rate from 74.22% at liquid chlorine concentration of 0 mg/L to 15.5% at 2.5 mg/L when Bti concentration was 0.5 mg/L. Water temperature ranging from 15℃ to 35℃ did not affect the inactivation efficiency of Bti. At Bti concentration of 0.125 mg/L, the inactivation rate significantly increased from 17.32% at ammonia nitrogen concentration of 0 mg/L to 53.26% at 6 mg/L. The inactivation rate of Chironomid larva was significantly higher in actual raw water than that in pre-chlorination coagulation sedimentation water. It could conclude static immersion with Bti solution in sedimentation tank is an effective method to control Chironomid larva.

Abstract 669: A unique acquired resistance mechanism to agents that target GLI (GANT61) in the Hedgehog (HH) signaling pathway in human colon cancer cells.

Abstract In normal cellular processes, the HH signaling pathway has critical roles in embryonic development, tissue patterning, stem cell function, and differentiation. Canonical HH signaling engages the transmembrane receptor PTCH, the intermediary signaling molecule SMO, and the transcriptional regulators of target genes and the HH signaling response, GLI1 and GLI2. In colon cancers, GLI is aberrantly regulated by oncogenic signaling pathways, driving cell survival and genomic instability involved in oncogenesis, progression and metastasis. Using the small molecule inhibitor of GLI1/GLI2 transcription, GANT61, we have demonstrated that inhibition of HH signaling at the level of GLI induces extensive cell death in human colon carcinoma cell line models in contrast to targeting SMO. Cells with acquired resistance to SMO inhibitors also remain highly sensitive to GANT61, underscoring GLI as a critical target in colon cancer. We have now identified a previously undescribed, novel mechanism by which cancer cells evade cell death following protracted, targeted exposure of GLI by GANT61. The classic pharmacologic approach of stepwise increasing drug selection did not produce a proliferative colon cancer cell line resistant to GANT61, confirming the critical importance of GLI genes to cell survival. In contrast, prolonged exposure (1-3 weeks) of HT29 cells to semi-lethal concentrations of GANT61 (10 μM; HT29-G10), induced GLI1/GLI2 inhibition, reduced proliferation, decreased appearance of γH2AX nuclear foci, and increased methylation of histone H3K9 (H3K9me1-3) within the GLI1 and GLI2 promoters. Methylation of H3K9 lysine residues is correlated with either an inactive or silent state of gene expression, and can exist in mono-, di-, or tri- methylated states. It has been demonstrated by others that methylation at H3K9 in the promoters of p53, p21Waf1 or DHFR genes, can regulate cell proliferation or function at the G1/S transition. In addition to GLI1 and GLI2, H3K9 methylation was also observed in HT29-G10 cells in the promoter regions of FOXM1 and CDC6, G1/S regulatory genes. These are GLI target genes, which regulate the G1/S transition, or PreReplication Complex assembly at initiation of DNA replication, respectively. Data suggest that the GLI2->GLI1->FOXM1->CDC6 promoters are coordinately regulated by H3K9 methylation in a pathway to promote gene silencing, reduce gene transcription, and reduce cell cycle transition at G1/S, constituting a unique mechanism of acquired resistance to drugs that target GLI. It is anticipated that these approaches will provide new insight into critical targets that determine HH-dependent survival, and lead to the translation of agents that target or regulate GLI function. Citation Format: Akwasi Agyeman, Tapati Mazumder, Janet Houghton. A unique acquired resistance mechanism to agents that target GLI (GANT61) in the Hedgehog (HH) signaling pathway in human colon cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 669. doi:10.1158/1538-7445.AM2013-669

A unique acquired resistance mechanism to agents that target GLI (GANT61) in the Hedgehog (HH) signaling pathway

Canonical HH signaling engages the transmembrane receptor PTCH, the intermediary signaling molecule SMO, and the HH signaling response molecules, GLI1 and GLI2. In colon cancers, GLI is aberrantly regulated, driving cell survival and genomic instability in oncogenesis, progression and metastasis. Following inhibition of GLI by a protracted exposure of HT29 to GANT61, we have identified a previously undescribed, novel mechanism by which cancer cells evade cell death. Stepwise selection did not produce a proliferative colon cancer cell line resistant to GANT61, confirming the critical importance of GLI genes to cell survival. In contrast, in HT29 cells, by inhibiting GLI1/GLI2 by prolonged exposure to semi‐lethal concentrations of GANT61 (10 μM; G10), we observed reduced proliferation, decreased appearance of γH2AX nuclear foci, and increased methylation of histone H3K9 (H3K9me1–3) within the promoters of the GLI1 and GLI2 genes. Increased H3K9 methylation was also demonstrated in HT29(G10) cells in the promoter regions of FoxM1 and CDC6, a G1/S and PreReplication Complex assembly regulatory genes, respectively. Data suggest that the GLI2‐>;GLI1‐>;FOXM1‐>;CDC6 promoters are coordinately regulated by H3K9 methylation in a pathway to promote gene silencing, reduce gene transcription, and reduce cell cycle transition at G1/S, constituting a unique mechanism of acquired resistance to drugs that target GLI.This work is supported by NCI awards RO1 CA 32613 (J.A. Houghton) and NCI RO1 108929 (J.A. Houghton)

Effects of Mixed Salt Stress on Germination Percentage and Protection System of Oat Seedling

In order to determine the effect of mixed salt stress on the germination rate of naked oat and protective enzymes of its’ seedling, in this study, using artificial culture box of climate, oat varieties (Yanke 1, Canadian 9, Big oat, Canadian 8, Canadian 6 and Baoluo) were cultivated in mixed salts solution. The results showed, with the increasing of concentration of double salts, the germination ratio and plant height decreased, semi-lethal concentration was between 1.5% and 2.6, utmost concentration was between 2.3 and 3.8%, respectively; the CAT activity of Big oat decreased, but that of other varieties increased first and then decreased, the peak value appeared when concentration of double salts was between 0.4 and 0.6%, respectively. And with the increasing of the concentration of double salt, POD activity and SOD activity of six oat varieties increased first and then decreased and its highest value appeared when concentration of double salts was between 1.2 and 1.8% respectively; the proline contents of all varieties increased, while the MDA contents decreased first and then increased and its minimum value appeared when concentration of double salts was 0.2%. To sum up, under the condition of salt stress, it was likely because of increasing of protective enzyme activity and proline contents to make salt tolerance of oats higher and salt tolerance of different varieties showed significant difference. The salt tolerance order showed that Yanke 1>Canadian 9>Baoluo>Big oat>Canadian 8>Canadian 6.

主要农作物芽期耐盐性比较研究Comparison of Salinity Tolerence among Main Crops at Germination Stage

为了开发利用沿海滩涂资源，为滩涂地区农作物布局提供参考信息，本实验以小麦、高粱、水稻、油菜、青菜、大豆、绿豆和芝麻等8种作物，每种作物随机挑选5个品种(系)为试验材料，采用芽期发芽鉴定法研究了5种盐浓度(0.0%, 0.5%, 1.0%, 1.5%, 2.0%)胁迫下不同作物不同品种之间相对发芽率的变化。以各作物半致死浓度的大小作为作物耐盐性强弱的标准。结果表明，随着盐浓度的升高，相对发芽率明显下降，不同盐浓度处理间相对发芽率的差异均达显著水(P 。不同作物之间差异显著，绿豆耐盐性最高，其半致死盐浓度为1.9%，小麦、油菜次之，为1.2%左右；水稻半致死盐浓度最低(0.84%)。半致死盐浓度下，除大豆外，各作物不同品种之间差异极显著(P ，且遗传力均在68.8%~94.9%之间。因此，可以通过初步耐盐性鉴定，从绿豆、油菜、小麦等耐盐性较好的作物中筛选强耐盐品种在沿海地区推广种植。同时，耐盐种质可以作为作物耐盐性育种亲本以及遗传研究利用。 To compare salinity tolerance among different crops, eight main crops planted in Jiangsu costal areas, including wheat,sorghum, rice, rape oil seed, green vegetables, soybean, mung bean, sesame, with 5 genotypes for each crop, were used as the experimental materials to study the change of the relative germina- tion rate under the condition of the salinity stresses (0.0%, 0.5%, 1.0%, 1.5%, 2.0% NaCl). Semi-lethal NaCl concentration was considered to be the criterion for salinity tolerance screening among different crops. The results showed that the relative germination rates (RGR) were significant decreasing with the increase of NaCl concentration. There were significant differences in RGR among NaCl concentrations and among crops. Of all the test crops, mung bean has the highest semi-lethal salt concentration (1.89%). Wheat and rape oils seed have the second highest semi-lethal salt concentration (1.20%). Rice was the most sensitive crop with only 0.84% of semi-lethal concentration. Furthermore there were a highly significant differences and rela- tively high heritability among different varieties in the same crop at semi-lethal salt concentrations (P Therefore, we can screen a large number of genotypes to explore salinity-tolerant germ- plasm which can be planted in the costal areas of Jiangsu Province. They should also be very useful in genetic improvement for salinity resistance.

Effects of heavy metals on insect immunocompetent cells

The influence of the following heavy metals, copper (Cu), zinc (Zn), cadmium (Cd) and lead (Pb), on haemocytes of the house fly Musca domestica L. was studied under laboratory conditions. House fly larvae were exposed to low or high, semi-lethal concentrations of metals. These particular metals were selected because they are present in polluted environments in Poland. In addition, we studied expression of the stress proteins HSP70 and HSP72 in haemocytes collected from larvae that had been exposed to heavy metal. The obtained results showed changes in haemocytes morphology and phagocytotic plasticity in the experimental flies in comparison to control. The number of prohaemocytes, regarded as stem cells, increased, while granulocytes, responsible for phagocytosis, decreased. However, we have not detected any clear changes in expression of HSP70 or HSP72 in flies treated with low or high concentrations of the heavy metals.

Ameliorative Effects of Brassinosteroid on Excess Manganese-Induced Oxidative Stress in Zea mays L. Leaves

Manganese (Mn) is becoming an important factor limiting crop growth and yields especially on acid soils. The present study was designed to explore the hypothesis that brassinosteroid application can enhance the tolerance of maize ( Zea mays L.) to Mn stress and if so, whether or not the mechanism underlying involves regulation of antioxidative metabolism in leaves. The effects of 24-epibrassinosteroid (EBR) on the growth, photosynthesis, water status, lipid peroxidation, accumulation of reactive oxygen species, and activities or contents of antioxidant defense system in maize plants under Mn stress were investigated by a pot experiment. At supplemented Mn concentrations of 150-750 mg kg −1 soil, the growth of plants was inhibited in a concentration-dependent manner. The semi-lethal concentration was 550 mg Mn kg −1soil. Foliage application with 0.1 mg L −1 EBR significantly reduced the decrease in dry mass, chlorophyll content, photosynthetic rate, leaf water content, and water potential of plants grown in the soil spiked with 550 mg kg −1 Mn. The oxidative stress caused by excess Mn, as reflected by the increase in malondialdehyde (MDA) content and lipoxygenase (LOX, EC 1.13.11.12) activity, accumulation of superoxide radical and H 2O 2, was greatly decreased by EBR treatment. Further investigations revealed that EBR application enhanced the activities of superoxide dismutase (SOD, EC 1.15.1.1), peroxidase (POD, EC1.11.1.7), catalase (EC 1.11.1.6), ascorbate peroxidase (APX, EC 1.11.1.11), dehydroascorbate reductase (DHAR, EC 1.8.5.1), and glutathione reductase (GR, EC 1.6.4.2), and the contents of reduced ascorbate and glutathione, compared with the plants without EBR treatment. It is concluded that the ameliorative effects of EBR on Mn toxicity are due to the up-regulation of antioxidative capacity in maize under Mn stress.

Elemental changes in the brain, muscle, and gut cells of the housefly, Musca domestica, exposed to heavy metals

The toxic effects of heavy metals on organisms are well established. However, their specific action at the cellular level in different tissues is mostly unknown. We have used the housefly, Musca domestica, as a model organism to study the toxicity of four heavy metals: copper (Cu), zinc (Zn), cadmium (Cd), and lead (Pb). These have been fed to larvae at low and high, semi-lethal concentrations, and their accumulation in the head, thorax, and abdomen was subsequently measured in adult flies. In addition, their impact on the cellular concentration of several elements important for cell metabolism-sodium (Na+), magnesium (Mg++), phosphorous (P), sulphur (S), chloride (Cl-) and potassium (K+)-were measured in neural cells, muscle fibers, and midgut epithelial cells. Our study showed that the heavy metals accumulate mainly in the abdomen, in which the concentrations of two of the xenobiotic metals, Cd and Pb, were 213 and 23 times more concentrated, respectively, than in controls. All the heavy metals affected the cellular concentration of light elements in all cell types, but the changes observed were dependent on tissue type and were specific for each heavy metal, and its concentration.

Resveratrol Oxidation inBotrytis cinereaConidia

ABSTRACT Observations using light microscopy showed that approximately 30% of Botrytis cinerea conidia treated with semi-lethal concentrations (i.e., 60 mug/ml) of the grapevine phytoalexin resveratrol possessed intracellular brown coloration. This coloration was never observed in the absence of resveratrol or in conidia treated with resveratrol together with sulfur dioxide (antioxidant compound) or sodium diethyldithiocarbamate (inhibitor of laccase action), suggesting that discoloration resulted from the laccase-mediated oxidation of resveratrol. Further studies using transmission electron microscopy enabled the observation of particular intravacuolar spherical vesicles and of granular material deposits along the tonoplast. These observations are likely to be related to the oxidation of resveratrol by an intracellular laccase-like stilbene oxidase of B. cinerea.